General principles
GENERAL PRINCIPLES TO TAKE INTO ACCOUNT IN THE ASSESSMENT OF OENOLOGICAL PRACTICES AND SUBSTANCES COMPOSINGOENOLOGICAL PRODUCTS
OIV-OENO 602-2022
Oenological practices:
- are safe biological, physical and chemical practices intended for the production and preservation of vitivinicultural products,
- preserve the natural and essential characteristics and composition of wine,
- should not cause consumer confusion or facilitate this.
Wherever possible and reasonable, preference should be given to substances that are extracted from grapes/substances of vitivinicultural origin.
Oenological practices and the use of oenological substances should not mask the effects of the use of defective raw materials, faulty raw materials, or unhygienic practices[1].
Taking this into consideration the OIV establishes a specific description of each oenological practice and any use of oenological substances, including its objectives, purposes and conditions of use. Oenological practices and the use of oenological substances recommended and published by OIV shall only be used for the purposes described and, in the manner, described to provide a benchmark for good manufacturing practice.
Oenological practices and oenological substances are used:
- to prevent or eliminate wine defects (oxidation, bacterial contamination, tartaric precipitation, etc.),
- to improve the process of winemaking (control the fermentations, improve the filterability, improve clarification,…),
- to improve the storage capacity or stability of the vitivinicultural product and maintenance of its organoleptic qualities,
- as additives or processing aids, while the status of the substance depends on their use.
There is a sufficient technological need to justify the use of the oenological substance, or the treatment gives evident advantages, economically or technically, compared to the present practices and consumers are not misled through the use of the oenological treatment agent; the oenological substance provides advantages to consumers and consequently fulfils one or several objectives such as:
- preservation of the nutritive quality of the vitivinicultural products,
- improvement of the storage capacity or stability of the vitivinicultural product and maintenance of its organoleptic qualities.
The OIV establishes a specification for each oenological substance concerning the origin, purity and other necessary indications.
The OIV evaluates and develops new oenological practices or revises those that already exist, also taking into account environmental impact/sustainability according to technological innovations.
The OIV recommends that:
- the quantity of an oenological treatment agent added in the elaboration of a product shall be limited to the lowest possible level necessary to accomplish its desired effect,
- the quantity of the oenological treatment agent that as a result of its use transforms to another product not intended to accomplish any physical or other technical effect in the vitivinicultural product itself, is reduced to the extent reasonably possible.
The OIV recommends maximum numerical use levels for oenological substances with an acceptable daily intake (ADI) set by JECFA or other Food Safety Authorities, also taking account of the opinion of the OIV’s “Food Safety” Group. The OIV also recommends numerical limits for best use of the technological function and/or for quality purposes on the basis of experimental data analysed by its own experts. In the absence of fixed numerical limits from the OIV, the OIV recommends oenological substances shall be used at levels established for GMP, which are the lowest levels possible to provide a specific technological function.
[1] "hygiene", means the measures and conditions necessary to control hazards and to ensure fitness for human consumption of wine, wine-based beverages, table grapes, raisins and other vine-based products taking into account its intended use
Alginic acid
COEI-1-ALGIAC Alginic acid
Sin no. 400
C.A.S. no.: 9005-32-7
- Subject, origin and scope
Alginic acid is a colloidal polysaccharide extracted from various varieties of brown algae in particular from Laminaria. The monomers constituting the -L-glucuronic acid and - D-mannuronic acid are bound in pairs as connections of the type 1 4
-[ α-glu-14- α-glu-14- β-man-1- β-man-1- α-glu |
A clarifying agent, which, after being neutralized before use by potassium chloride, or potassium carbonate or potassium hydrogenocarbonate can be added to the drawn-off liquid, designed to carry out the second fermentation of sparkling wines (foam formation).
Alginic acid is made up on average of 200 basic units of uronic acids.
Their molecular weight ranges between 10 000 and 600 000 U.
- Labeling
The concentration of alginic acid must be indicated on the label, as well as the conditions of safety and conservation.
- Characteristics
Alginic acid exists in powder or filament form, or as amorphous granules of a yellowish white to brown color, insoluble in pure water and the various organic solvents. It can dissolve in water alkalized by sodium carbonate, sodium hydroxide or trisodium phosphate.
- Identifying characteristics
4.1. pH
A suspension of 3% alginic acid in water has a pH ranging between 2 and 3.5.
4.2. Differentiation with other polysaccharides
An alginic acid solution of 5 g/l in sodium hydroxide (dissolve 4.3 g of sodium hydroxide in water and complete to 100 ml) precipitate in gelatinous form by adding a fifth of volume of a 2.5% solution of calcium chloride.
Furthermore, an addition of a half volume of a solution saturated with ammonia sulfate to the solution previously described does not cause any turbidity.
These two tests can be used to differentiate alginic acid from other polysaccharides that may be used in foodstuffs or pharmaceuticals.
4.3. Organoleptic characteristics
Alginic acid must have no taste, or abnormal odor.
- Tests
All the limits described below refer to the dry weight of alginic acid.
5.1. Insoluble in a solution of sodium hydroxide
Dissolve by prolonged magnetic agitation 1 g of alginic acid weighed with precision in 100 ml of a solution of sodium hydroxide (dissolve 4.3 g of sodium hydroxide in water and complete to 100 ml), centrifuge, decant, and wash the residue 5 times with distilled water, with centrifugation and drainage of the washwater each time. Transfer all the residue using distilled water to a Gooch filter that has been tared beforehand (filter made of sintered glass of low porosity), dry for 1 hour at 105°C and weigh again.
The rate of insoluble should not exceed 2% in relation to the dry weight of the alginic acid.
5.2. Loss on desiccation
Determine until constant weight, on a test specimen of 2 g, the loss of weight, at 100-105°C, the alginic acid must be lower than 15 p. 100
5.3. Sulfuric ash
Proceed as indicated in chapter II of the international oenological Codex. The sulfuric ash content should not be higher than 8 p 100 in weight of the alginic acid.
5.4. Preparation of the solution for tests
After weighing the ashes, dissolve them in 2 ml of concentrated hydrochloric acid (R) and 10 ml of water. Heat to activate the dissolution and add water until a volume equal to 25 times the weight of the dry alginic acid is obtained. 1 ml of this solution contains the mineral matter of 0.04 g of dry alginic acid.
5.5. Lead
On the solution prepared for tests (5.4), to carry out the dosage of lead according to the method described in chapter II of the international oenological Codex.
The lead content must be lower than 5 mg/kg.
5.6. Cadmium
On the solution prepared for the tests (5.4), determine the cadmium using the method described in chapter II of the international oenological Codex.
The cadmium content must be lower than 1 mg/kg.
5.7. Mercury
Determine the mercury using the method described in Chapter II of the international oenological Codex.
The mercury content must be lower than 1 mg/kg.
5.8. Arsenic
On the solution prepared for the tests (5.4), determine the arsenic using the method described in Chapter II of the international oenological Codex.
The arsenic content must be lower than 3 mg/kg.
5.9. Bacteriological control
Proceed as indicated in chapter II of the international Oenological Codex for each parameter.
Limit: total viable microorganisms: less than 5 x 103 CFU/g.
5.10. Coliforms
The number of coliforms must be lower than or equal to 1 per g.
5.11. Staphilococca
The number of staphilococca (-haemolytics with positive coagulase) must be lower than or equal to 1 per g.
5.12. Salmonella
The number of salmonella must be lower than 1 per 100 g.
5.13. Yeast
Limit concentration: 5 x 102 CFU per g of preparation.
5.14. Lactic bacteria
Limit concentration: 102 CFU per g of preparation.
5.15. Lactobacillus sp.
Limit concentration: 10 CFU per g of preparation.
5.16. Pediococcus sp.
Limit concentration: absence in a sample of 10 g of preparation.
5.17. Acetic bacteria
Limit concentration: 103 CFU per g of preparation.
5.18. Moulds
Limit concentration: 5 x 102 CFU per g of preparation.
- Storage
Alginic acid must be kept in sealed bags.
Calcium (Alignate)
SIN N°: 402
- Object, origin and scope of application
Calcium alginate is obtained from a 1 % aqueous solution of potassium alginate or alginic acid placed in contact with a 20 % aqueous solution of calcium chloride. Beads of calcium alginate can be produced by dropping droplets of potassium alginate solution into a calcium chloride solution.
Beads of calcium alginate, dry or wet, can contain yeasts or lactic bacteria, dry or wet. They are used for foam forming purposes in the bottle for sparkling wine or to restart alcoholic fermentation in still wines or to start the malolactic fermentation.
These beads can be coated with a double layer of potassium or calcium alginate or with colloidal silica to prevent the precipitation of the yeasts or bacteria incorporated into the beads.
- Labelling
The label should indicate the product’s purity and the safety and storage conditions for calcium alginate, the yeasts or bacteria incorporated into the beads, the expiration date and the lot number.
- Characteristics
Calcium alginate is a translucent gel, which is insoluble in water and wine. It only dissolves in a sodium metaphosphate solution.
An alginic acid precipitate is also produced if 1 ml of sulfuric acid diluted to 10 % (R) is added to 10 ml of an aqueous 1 % (m/v) suspension of calcium alginate.
Anti-foaming agents
COEI-1-ACIGRA Anti-Foaming Agents (Fatty acid mono- and diglycerides)
SIN NO. 471
- Objective, origin and scope of application
The mixture of fatty acid glyceric mono- and diesters (with a small quantity of tri-esters), with fatty oils and acids and alimentary fats are termed mono- and diglycerides. The mixture of mono- and diglycerides used as anti-foaming agents are essentially constituted by oleic acid esters.
The product thus defined can contain small quantities of fatty acids and free glycerol. It is used under appropriate technological conditions and does not leave measurable traces in wine after filtering.
- Labelling
The label must indicate the mono- and diglyceride content of the preparation, the storage and safety conditions, and the final date of use.
- Properties
The product is usually found in the form of an oily liquid with a straw yellow color, a doughy product with an ivory color or a hard waxy solid with a white or off-white color. All of the forms have a pleasant odor and taste. The solid form can be found in flakes, powder or small granules.
The product used as an anti-foaming agent is liquid at normal temperatures, but can become cloudy at low temperatures.
- Solubility
Insoluble in water.
Soluble in ethanol, chloroform and benzene.
- Identifying characteristics
5.1. Hydrolysis of the Sample
Treat 1 g of the sample by reflux using a 0.5 M potassium hydroxide solution for 1 hour. Add 15 ml of water and acidify with hydrochloric acid diluted to 30 pp 100 (v/v) (R) (approximately 4-5 ml). Oily drops or a white/yellowish white precipitate will form. Extract the fatty acids released using 5 ml hexane, separating the solvent. Repeat the extraction with 5 ml of hexane and reunite the two extracts.
Set aside the aqueous phase.
5.2. Detection of the Fatty Acids in the Hexane Extract Using Gas Phase Chromatography
For the purpose of example, use may be made of a semi-polar column, e.g., Carbowax 20M ® measuring 25m x 0.32 mm x 0.25 m phase thickness.
5.3. Detection of Glycerol
Place 5 ml of the aqueous phase in a test tube. Add an excess amount of powdered calcium hydroxide and place the test tube in boiling water for five minutes, stirring from time to time. Cool and filter.
Place one drop of the filtrate in a test tube and add approximately 50 mg of potassium hydrogen sulfate, At the end of the test tube, place a piece of filter paper soaked in the reagent obtained by mixing extemporaneously equal volumes of a sodium nitrosopentacyanoferrate solution (R) and piperidine (F'). Heat using a small flame. A blue coloring of the reactive paper indicates the presence of acrolein.
The color turns red by adding 1M sodium hydroxide solution.
- Tests
6.1. Drying Loss at 100 °C
Weigh exactly a quantity of about 5 g of the product to be analyzed in a glass crystallizing dish with a diameter of 70 mm, which has been preliminarily dried in an oven, cooled in a desiccator and calibrated. Place the crystallizing dish with the fatty material into a 103 °C oven and maintain this temperature for 30 minutes. Remove the crystallizing dish, let cooll in the desiccator, then weigh. Place the sample in the oven again for 30 minutes. Weigh it again after cooling. Drying loss in the oven is completed when weight loss does not exceed 0.05% per half-hour of heating.
Drying loss at 100 °C should be less than 2 pp 100.
6.2. Sulfur Ash
Sulfur ash is quantified as indicated in the Annex using a test sample of 5 g. The sulfur ash should weigh less than 0.2 g/kg.
6.3. Arsenic
Determined as indicated in the Annex using a test sample of 5 g. The arsenic should weigh less than 3 mg/kg.
6.4. Heavy metals
Test for heavy metals either:
- After mineralization at 450 5 °C of the residue left by the drying loss test. Take up the ash using 1 ml of diluted hydrochloric acid (R) and one drop of concentrated nitric acid (R) while heating in a 100 °C water bath to activate dissolution, then decant in a 25 ml volumetric flask, washing the cap with distilled water. Fill up to gauge line.
Draw off a volume of v ml of solution corresponding to 2 g of the sample to be analyzed and proceed with the test for heavy metals as indicated in the Annex.
- or, after liquid mineralization of an sample weighed with precision to about 5 g using concentrated nitric acid (R), Perhydrol and a microwave digester to accelerate the operation.
Decant the liquid obtained in a 25 ml volumetric flask and fill to the line with the wash water. Continue as indicated in the heavy metal tests.
Heavy metal content, expressed in terms of lead, should be less than 10 mg/kg.
6.5. Lead
Using the technique set forth in the Compendium, determine the quantity of lead in one of the two aforementioned preparations (6.4). The lead content should be less than 5 mg/kg.
6.6. Mercury
Using the technique described in the annex, determine the quantity of lead in one of the two aforementioned preparations (6.4). The lead content should be less than 1 mg/kg.
6.7. Cadmium
Using the technique detailed in the annex, determine the quantity of cadmium in one of the two aforementioned preparations (6.4). The lead content should be less than 1 mg/kg.
6.8. Free Fatty Acids
Prepare 125 ml of a mixture of equal volumes of isopropyl alcohol and toluene. Add 2 ml of 1 pp 100 phenolphthalein solution (m/v) in isopropyl alcohol and neutralize using an alkaline solution until a persistent but weak pink coloring appears.
Weigh with precision an amount of approximately 5 g of the sample to be analyzed in a 500 ml conical flask. Add the neutralized solvent mixture and dissolve the test sample, by heating if necessary, while stirring vigorously. Pour the 0.1 M potassium hydroxide solution until a pink color identical to that obtained during the solvent neutralization process is obtained. Let n be the volume in ml poured:
Fatty acid content expressed in g of oleic acid pp 100 (m/m):
- 2.8n / test sample in g
The fatty acid content in terms of oleic acid should be less than 3 pp 100 (m/m).
6.9. Soaps
Weigh precisely about 10 g of the product to be analyzed in a 250 ml conical flask. Add a mixture of 60 ml of acetone and 0.15 ml of 0.5 pp 100 (m/v) bromophenol blue solution in 95% alcohol by volume which has first been neutralized with a 0.1 M hydrochloric acid solution or a 0.1 M sodium hydroxide solution. Gently heat in a 70 °C water bath and titrate with a 0.1 M hydrochloric acid solution until the blue color disappears. Let sit for 20 minutes. Heat until the precipitate redissolves and, if the blue color reappears, continue titration.
1 ml 0.1 M hydrochloric acid solution corresponds to 0.0304 g of sodium oleate (NaC18H33O2).
Soap content expressed in g of sodium oleate pp 100 (m/m):
- 3.04n / test sample in g
Soap content expressed in g of sodium oleate should be less than 6 pp 100 (m/m).
6.10. Monoglycerides
6.10.1. Sample preparation
If the sample is in solid form, melt it by heating it to its melting point at a temperature of less than 10 °C. Liquid samples which are cloudy or have particles in them should also be heated. Mix vigorously.
6.10.2. Method
Weigh precisely a test sample, Q, of approximately 1 g to be analyzed in a 100 ml cylindrical flask. Dissolve using 25 ml of chloroform. Transfer this solution to a decanting glass. Wash the cylindrical flask with 25 ml of chloroform, then with 25 ml of water and add these liquids to the contents of the decanting glass.
Seal the decanting glass hermetically. Stir for 30-60 seconds. Let the two phases separate out (add 1-2 ml of crystallizable acetic acid (R) to break the emulsion). Collect the aqueous phase in a 500 ml conical flask with an emery stopper. Extract the chloroform phase remaining in the decanting glass twice with 25 ml of water. Separate the aqueous phase and place it in the 500 ml conical flask. These aqueous extracts will be used for the free glycerol analysis.
Transfer the chloroform from the decanting glass to a 500 ml conical flask with an emery stopper. Add 50 ml of periodic acetic acid solution (R) while stirring.
In the two other 500 ml conical flasks with emery stoppers to be used as "blanks", place 50 ml of chloroform and 10 ml of water. Add 50 ml of periodic acetic acid solution (R) while stirring to each of the two flasks. Let the three flasks sit at least 30 minutes, but no more than 90 minutes.
While gently stirring, add 20 ml of potassium iodide solution (R) to each of these containers. Let sit at least 1 minute but no more than 5 minutes before volumetric analysis.
Add 100 ml water and titrate with a 0.05 M sodium thiosulfate solution using a magnetic stirrer until the brown color disappears from the aqueous phase. Add 2 ml of starch solution (R) and continue to add the reagent until the iodine disappears from the chloroform layer and the blue color disappears from the aqueous phase.
6.10.3. Calculate the percentage of monoglycerides using the formula:
- B is the average volume in ml of the sodium thiosulfate solution used for analysis of the "blanks" containing chloroform.
- S is the amount of sodium thiosulfate solution in ml used to titrate the sample.
- M is the exact molarity of the sodium thiosulfate solution.
- P is the weight of the sample to be analyzed in the volume of chloroform used for the analysis.
- 17.927 is the molar mass of glycerol monostearate, divided by 20.
The monoglyceride content expressed in terms of glycerol monostearate should be greater than 30 pp 100 (m/m).
6.11. Free glycerol
Add 50 ml of periodic acetic acid solution (R) to the aqueous extracts obtained during the monoglyceride-analysis process. Simultaneously prepare a "blank" by adding to 75 ml of water in a 500 ml conical flask 50 ml of periodic acetic acid solution (R). Continue the determination process as indicated in the method described for monoglycerides.
Calculate the percentage of glycerol using the following formula:
- b is the volume in ml of sodium thiosulfate solution used in the quantitative analysis the "blank" containing 75 ml of water
- S is the volume in ml of sodium thiosulfate solution used in the quantitative analysis of the aqueous extracts
- M is the molarity of the sodium thiosulfate solution.
- Q is the weight of the first sample to to be analyzed (see monoglyceride determination).
- Glycerol content should be less than 7 pp 100 (m/m).
N.B. : Glycerol can also be disclosed and identified by high performance liquid chromatography (HPLC) (5.3).
- Storage
Anti-foaming agents should be kept in completely water-tight containers and away from heat.
Lactic acid
COEI-1-ACILAC L-Lactic acid, D-Lactic acid, D,L-Lactic acid2-hydroxypropanoic acid
N° SIN : 270
C.A.S. number 50-21-5
(L-: 79-33-4; D-: 10326-41-7; DL-: 598-82-3)
chemical formula C3H6O3
Molecular mass: 90.08, density 1.20-1.21.
- Object, origin and field of application
An acid of natural origin obtained by lactic fermentation of sugars or synthetically made; it may contain condensation products such as lactate from lactic acid and dilactide.
It is used for the acidification of musts and wines in the conditions set by the regulation.
- Labelling
The label must mention particularly clearly that it concerns L-lactic or D-Lactic acids obtained by fermentation or D,L-Lactic obtained by a chemical process, the storage conditions and expiration date.
The common commercial products are solutions at 50%-90%.
Solid products containing about 100%-125% of titrable lactic acid also exist. (Note: Lactic acid is hygroscopic and once concentrated by boiling or distillation, it forms condensation products that hydrolyse into lactic acid by dilution and by heating in water).
Purity level: not less than 95.0% and not more than 105.0% of the concentration marked.
- Characteristics
Colourless or slightly yellow and syrupy liquid with a clearly acid flavour to a slightly lactic taste.
- Solubility
- Water at 20°C: very soluble
- Alcohol at 95% vol.: Very soluble
- Ether: very soluble
- Insoluble in chloroform.
- Optical rotation
For L-lactic acid aqueous solution at 2.5 g for 100 ml.
is 2.6° |
For D-lactic acid in aqueous solution at 8 g for 100 ml.
is -2.6° |
- Identity characters
6.1. Characterisation of lactic acid
In a 100 ml conical flask, weigh 10 g of lactic acid, add 5 ml of sulphuric acid 0.5 M, shake, add 25 ml of potassium permanganate at 0.33% and place on a hot plate. Collect the vapour released on a filter paper soaked with a solution at 50% vol/vol of morpholine at 20% and potassium nitrocyanoferrate (II) at 5%.
The filter paper becomes blue.
6.2. Determination of total lactic acid
Titrate the free lactic acid with sodium hydroxide 1 M then hydrolyse the polymerised lactic acid using an excess of sodium hydroxide and then determined by sulphuric acid 0.5 M.
6.3. Colour
Compare the colour with the standards of the alpha scale (colour standards of platinum-cobalt).
6.4. Stereochemical purity
The method is based on the separation by HPLC using a chiral phase of two enantiomers of lactic acid. The product is diluted in water beforehand. Enzymatic determinations can also be performed according to the methods in the Compendium of international methods of analysis of wines and musts.
- Test trials
7.1. Preparation of the test trial solution
For the purity test trials, prepare a solution containing 10% m/v of lactic acid by using the concentration marked.
7.2. Sulphuric ashes
From a 2 g sample of lactic acid, determine the sulphuric ashes as indicated in chapter II of the International Oenological Codex.
The content must be less than or equal to 1 g/kg.
7.3. Chlorides
To 0.5 ml of the test trial solution (7.1), add 14.5 ml of water, 5 ml of diluted nitric acid (R) and 0.5 ml of silver nitrate solution at 5% (R). The solution should satisfy for the test trial, the determination limit of chlorides described in chapter II of the International Oenological Codex.
The chloride content must be less than 1 g/kg expressed in hydrochloric acid.
7.4. Iron
To 10 ml of the test trial solution (7.1), add 1 ml of concentrated hydrochloric acid (R) and 2 ml of the potassium thiocyanate solution at 5% (R). The red colouration obtained must not be darker than the control prepared with 1 ml of the iron salt solution (III) at 0.010 g of iron per litre (R), 9 ml of water and the same quantities of the same reagents.
The iron content must be less than 10 mg/kg.
Iron can also be determined by atomic absorption spectrometry according to the method described in chapter II of the International Oenological Codex.
7.5. Lead
Using the test trial solution (7.1), apply the method described in chapter II of the International Oenological Codex.
Lead content should be less than 2 mg/kg.
7.6. Mercury
Using the test trial solution (7.1), determine the mercury according to the method described in chapter II of the International Oenological Codex.
Mercury content should be less than 1 mg/kg.
7.7. Cadmium
Using the test trial solution (7.1), determine the cadmium according to the method described in chapter II of the International Oenological Codex.
Cadmium content should be less than 1mg/kg.
7.8. Arsenic
Using the test trial solution (7.1), determine the arsenic according to the method described in chapter II of the International Oenological Codex.
Arsenic content should be less than 3 mg/kg.
7.9. Sulphates
To 1 ml of the test trial solution (7.1), add 18 ml of water, 1 ml of diluted hydrochloric acid at 10% (R) and 2 ml of barium chloride solution at 10% (R). The solution should satisfy for the test trial, the determination limit of sulphates described in chapter II of the International Oenological Codex.
Sulphate content should be less than 1 g/kg, expressed in sulphuric acid.
7.10. Cyanides
In a 40 ml volumetric flask containing 25 ml of distilled water and 2.5 ml of buffer solution at pH 7.5 (R), introduce 0.4 ml of the test trial solution (7.1), add 0.3 ml of chloramine T solution at 0.1% (R). Wait 90 seconds and add 6 ml of pyridine-pyrazolone reagent (R). Complete to 40 ml with distilled water and mix. The colouration obtained must not be darker than that obtained by treating the same way 4 ml of a freshly prepared potassium cyanide solution titrating 1 mg of hydrocyanic acid per litre (R).
Free cyanide content expressed in hydrocyanic acid should be less than 1 mg/kg.
7.11. Citric acid
To 5 ml of the test trial solution (7.1), add 5 ml of water, 2 ml of mercury sulphate solution (II) (R), bring to the boil and add a few drops to the potassium permanganate solution at 2% (R). No white precipitate should form.
7.12. Citric, oxalic, tartaric and phosphoric acids
Dilute 1 ml of the test trial solution (7.1) in 10 ml of water, add 40 ml of the calcium hydroxide solution (R) bring to the boil for 2 minutes. No turbidity should form.
7.13. Sugars
Add 2 ml of the test trial solution (7.1) to 10 ml of cupro-alkaline reagent (R). No red precipitate should form.
- Storage
Lactic acid should be stored in hermetically sealed containers away from heat and light.
Malic acid
COEI-1-ACIMAL L-Malic acid , DL-Malic acid
2-hydroxybutanedioic acid
N° SIN: 296
C.A.S. number 617-48-1
Chemical formula C4H6O5
Molecular mass: 134.09
- Object, origin and field of application
An acid of natural origin contained in most fruit (L-malic acid) or synthetically made: DL-malic.
It is used for the acidification of musts and wines in the conditions set by the regulation.
- Labelling
The label must mention particularly clearly that it is L-malic or D,L-malic acid, the storage conditions and date of expiry.
Malic acid content should be at least 99%.
- Characteristics
White or off-white crystalline powder or granules with a clearly acid flavour.
Melting point of D,L-malic: 127°C-132°C
Melting point of L-malic: 100°C.
- Solubility
- Water at 20°C: 55.8 g/100
- Alcohol at 95% vol.: 45.5 g/100.
- Ether: 0.84 g/ 100
- Optical rotation
For the L-Malic acid in aqueous solution at 8.5 g for 100 ml.
is -2.3° |
- Identity characters
6.1. Characterisation of malic acid
Malic acid can be determined by an enzymatic process according to the methods in the Compendium of international methods of analysis of wines and musts (specifically L-malic and D-malic acids. Malic acid can also be determined by HPLC according to the method in the Compendium of international methods of analysis of wines and musts.
- Test trials
7.1. Preparation of the test trial solution
For purity trials, prepare a solution containing 10% m/v of malic acid.
7.2. Sulphuric cinders
From a 2 g sample of malic acid, determine the sulphuric cinders as indicated in chapter II of the International Oenological Codex.
Content must be less than or equal to 1 g/kg.
7.3. Chlorides
To 0.5 ml of the test trial solution (7.1), add 14.5 ml of water, 5 ml of diluted nitric acid (R) and 0.5 ml of silver nitrate solution at 5% (R). The solution should satisfy for the test trial, the determination limit of chlorides described in chapter II of the International Oenological Codex.
Content must be less than 1 g/kg expressed in hydrochloric acid.
7.4. Iron
To 10 ml of the test trial solution (7.1), add 1 ml of concentrated hydrochloric acid (R) and 2 ml of potassium thiocyanate solution at 5% (R). The red colouration obtained should not be darker than that of the control prepared with 1 ml of an iron salt solution (III) at 0.010 g of iron per litre (R), 9 ml of water and the same quantities of the same reagents.
Content must be less than 10 mg/kg.
Iron can also be determined by atomic absorption spectrometry according to the method described in chapter II of the International Oenological Codex.
7.5. Lead
Using the test trial solution (7.1), apply the method described in chapter II of the International Oenological Codex.
Lead content must be less than 5 mg/kg.
7.6. Mercury
Using the test trial solution (7.1), determine the mercury according to the method described in chapter II of the International Oenological Codex.
Mercury content must be less than 1 mg/kg.
7.7. Cadmium
Using the test trial solution (7.1), determine the cadmium according to the method described in chapter II of the International Oenological Codex.
Cadmium content must be less than 1 mg/kg.
7.8. Arsenic
Using the test trial solution (7.1), determine the arsenic according to the method described in chapter II of the International Oenological Codex.
Arsenic content must be less than 3 mg/kg.
7.9. Sulphates
To 1 ml of the test trial solution (7.1), add 18 ml of water, 1 ml of diluted hydrochloric acid at 10% (R) and 2 ml of the barium chloride solution at 10% (R). The solution should satisfy for the test trial, the determination limit of sulphates described in chapter II of the International Oenological Codex.
Sulphates content must be less than 1 g/kg, expressed in sulphuric acid.
7.10. Cyanides
In a 40 ml volumetric flask containing 25 ml of distilled water and 2.5 ml of buffer solution at pH 7.5 (R), introduce 0.4 ml of the test trial solution (7.1), add 0.3 ml of chloramine T solution at 0.1% (R). Wait 90 seconds and add 6 ml of pyridine-pyrazolone reagent (R). Complete to 40 ml with distilled water and mix. The colouration obtained must not be darker than that obtained by treating the same way 4 ml of a freshly prepared potassium cyanide solution titrating 1 mg of hydrocyanic acid per litre (R).
Free cyanide content expressed in hydrocyanic acid must be less than 1 mg/kg.
7.11. Sugars
Add 2 ml of the test trial solution (7.1) to 10 ml of cupro-alkaline reagent (R). No red precipitate should form.
7.12. Fumaric and maleic acids
Limit in fumaric acid: 1% in weight.
Limit in maleic acid: 0.05% in weight. These acids are determined by HPLC according to the method described in the Method of Analysis of Wines and Musts in the same way as malic and tartaric acids.
- Storage
Malic acid should be stored in hermetically sealed containers away from heat and light.
Rectified alcohol of agricultural origin
COEI-1-ALCAGR Rectified alcohol of agricultural origin
- Objective, Origin and Scope of Application
Rectified, or "neutral," alcohol obtained by distilling and rectifying alcohol from wine, wine sediments or alcoholic fermentation products derived from from grape or raisin marcs, and all other plant-based substances of agricultural origin.
Rectified alcohol of agricultural origin forms an ingredient of some spirits and special wines.
- Composition
At a temperature of 20 °C, 100 parts of this alcohol contain at least 96 parts ethanol.
Note: The tests and controls described below in italics are not mandatory and are performed only upon request.
- Properties
Colorless, clear, volatile liquid with a penetrating odor and fiery flavor. It is flammable and burns without smoke and with a blue flame.
It should be distilled completely at between 78 and 79 °C.
3.1. Solubility
Neutral alcohol is miscible in water in all proportions with a notable release of heat and contraction of volume. It is also mixable with in acetone, chloroform, ethyl ether, glycerol, and an equal volume of castor oil.
3.2. Characterization Procedure
- Slowly heat a mixture of 1 ml neutral alcohol, twenty drops of concentrated sulfuric acid (R) and 10 g of sodium acetate (R) in a test tube. A strong, characteristic odor of ethyl acetate will be released.
- Mix several drops of alcohol and 1 ml of concentrated sulfuric acid (R), then add several drops of 10 pp 100 potassium dichromate solution. The liquid will become green and emit the odor of ethanal.
- Dilute 0.5 ml of alcohol with 4.5 ml of water. Add 1 ml of 1M sodium hydroxide solution, then slowly add 2 ml of iodized potassium iodide (R). An odor of iodoform will be produced, following by the formation of a yellow precipitate.
- Analysis of Agricultural Origin
This analysis is carried out by measuring the ethanol 14C/12C ratio (scintillation) in accordance with the method described in the Spirits Compendium.
- Tests
4.1. Appearance
Take two identical test tubes made of alkali-lime glass about 250 mm high, fill one with alcohol, the other with water, which will serve as a control. Examine the liquids along the cylinder's axis. The alcohol should not exhibit any noticeable coloration.
In one test tube about 250 mm high and 25 mm in diameter, pour 40 ml of alcohol, then dilute it with 80 ml of water. The mixture should not cloud nor present any odor or foreign taste.
4.2. Foreign Odoriferous Substances
Let 10 ml of alcohol evaporate spontaneously on a strip of white filter paper. No foreign odor should be perceived during or after evaporation.
4.3. Dry Extract or Non-Volatile Residue
In a 25 ml calibrated dish, heat to 100 °C in a water bath, then slowly evaporate 100 ml of alcohol. Weigh. The dry extract should be less than 1.5 g/hl 100% ethanol by volume.
4.4. Heavy Metals
Take up, using 10 ml dilute chlorhydric acid (R), any residue left from the evaporation of 100 ml alcohol during the dry extract analysis. After heating for several minutes in a 100 °C water bath to stimulate dissolution of this residue, decant the acid solution in a 25 ml volumetric flask after the calibrated dish has been washed three times with 5 ml of water and the volume raised to 25 ml. Take a 5 ml sample of this solution in a test tube. Add 2 ml of a pH 3.5 (R) buffer solution, 7.5 ml of water and 1.2 ml of thioacetamide reagent (R). The solution should not produce any white or black precipitates nor any brown or other coloration. At the very least, any coloration produced should be no more intense than that obtained using the general method. (Heavy metal conent expressed in lead, after 50% concentration of the alcohol, should be 0.5 mg/l).
4.5. Lead
Using the method set forth in the Compendium, perform the lead analysis lead in the solution obtained in the previous paragraph. (Lead content should be less than 0.5 mg/l).
4.6. Mercury
Using the method described in the annex, carry out the mercury analysis in the solution obtained in Paragraph
(Mercury content should be less than 0.2 mg/l).
4.7. Arsenic
Using the method described in the annex, carry ou the arsenic analysis in the solution obtained in Paragraph 4.4. (Arsenic content should be less than 0.5 mg/kg after 50% alcohol concentration).
4.8. Ketones, propan-2-ol and 2-methylpropan-1-ol
Add 3 ml of water and 10 ml of mercury sulfate (II) solution (R) to 1 ml of alcohol, then heat in a 100 °C water bath. No precipitate should form in the first three minutes.
4.9. Permanganate Decolorization Time (Barbet Test)
Pour 50 ml of the alcohol sample into a flask. Add 2 ml of freshly prepared potassium permanganate solution to 0.20 g/l (R). Place the container in a 15 °C water bath and start a stopwatch. Avoid directly exposing the sample to natural or artificial light during the test.
Simultaneously, place 50 ml of the comparison solution in the 15 °C water bath. This solution is obtained by mixing 3 ml of 5 pp 100 cobalt chloride solution (R), 4.2 ml of 4 pp 100 uranyl nitrate solution (R) and filling to 50 ml with distilled water. Compare the test color to the standard. Stop the timer when the colors are identical. Note the amount of time elapsed. The decolorization time of the permanganate should be at least 20 minutes.
4.10. Sulfured Derivatives
Add approximately 1 ml of mercury, then 20 ml of alcohol to a test tube. Agitate for 1-2 minutes. The surface of the mercury should remain brilliant with no black clouding.
4.11. Methanol
4.11.1. Colorimetric Analysis
Standard solution: weigh 5 g of methanol in a 50 ml volumetric flask, then top off to the line with ethanol (free of methanol).
In a 1-liter volumetric flask, place 1 g of the preceding solution (i.e., 1.25 ml) containing 125 mg of methanol, 250 ml of pure alcohol (methanol free). Top off with water to 1000 ml.
Test technique: place 1250/A ml of alcohol in a volumetric flask (A is the alcoholmetric titer of the alcohol to be tested.) and fill to the gauge line with water. Place 1 ml of alcohol, diluted to 25 pp 100 in a test tube. Add four drops of 50 pp 100 (m/m) phosphoric acid (R), four drops of 5 pp 100 (m/m) potassium permanganate solution (R), then stir and let sit 10 minutes. Decolorize the permanganate with several (typically 8) drops of 2 pp 100 (m/v) of potassium anhydrous sulfite (metabisulfite) (R), avoiding any excess. Add 5 ml of chromotropic sulfuric acid solution (R). Place in a 70 °C water bath for 20 minutes. No violet color should appear, or in the event it does appear, it should not be more intense than that of a control prepared using the same technique and the same reagents, with 1 ml of the aforementioned standard solution (maximum methanol content is 50 g/hl at 100% vol.).
4.11.2. Gas phase chromatography Analysis
- Equipment (example):
- Gas phase chromatograph with a flame ionization detector
- Semi-polar capillary columns, for example Carbowax 20 M .
Test technique:
Prepare a water-alcohol solution using 1 g per liter of the internal standard (4-methylpentane-2-ol) in 50 pp 100 alcohol by volume.
Prepare the solution to be analyzed by adding 5 ml of this solution to 50 ml of alcohol reduced to 50 pp 100 by volume.
Prepare a reference solution of methanol at 100 mg per liter of alcohol at 50 pp 100 by volume. Add 5 ml of the internal standard solution to 50 ml of this solution.
Inject 2 microliters of the solution to be analyzed added to the internal standard solution, into the chromatograph.
The oven temperature should be 90 °C and the supporting gas flow rate should be 25 ml per minute. These settings are given as an example.
- S: surface of the methanol peak of the reference solution
- : surface of the methanol peak of the solution to be analyzed
- i: surface of the internal standard solution peak in the solution to be analyzed
- I: surface of the internal standard solution peak in the reference solution
The methanol content, expressed in milligrams per liter of alcohol at 50 pp 100 by volume, is given by the formula:
The content in grams per hectoliter of pure alcohol is 0.20C (maximum content in methanol 50 g/hl of ethanol at 100% by volume).
4.12. Ammonium Hydroxide and Nitrogenous Bases
Pour 50 ml of the alcohol to be examined into a 200 ml flask. Add 40 ml of water and two drops of phosphoric acid (ρ 20 = 1.58). Distill and collect the 80 ml that are returned. Add 2 ml of 10 pp 100 sodium hydroxide ® to the cooled residue. Distill again and collect approximately 7 ml of distillate in a test tube to which had previously been added 2 ml of water and one drop of methyl red solution ®. The distillate should be drawn to the bottom of the tube using a slender tube. Titrate using a solution of 0.01 M hydrochloric acid until the indicator turns to red. Let n be the number of milliliters of 0.01 M hydrochloric acid solution used.
1 ml of 0.01 M hydrochloric acid solution corresponds to 0.00014 g of nitrogen (ammoniacal or volatile nitrogen bases).
The quantity of ammoniacal nitrogen or nitrogenous bases expressed in milligrams of nitrogen per liter of ethanol is:
280n/A
Where A is the alcohometric titer by volume of the alcohol studied.
Neutral alcohol should not contain more than 1 mg of nitrogen (ammoniacal or of volatile nitrogenous bases) per liter of ethanol.
(Maximum ammonium hydroxide and nitrogenous base content is expressed in terms of nitrogen is 0.1 g/hl of ethanol at 100% by volume).
4.13. Acidity
Place 100 ml strengthened of 50 pp 100 by volume alcohol in a 250 ml conical flask. Add one drop of phenol red solution (R) and add 0.01 M sodium hydroxide, one drop at a time, until red, where n is the number of milliliters used.
1 ml of 0.01 M sodium hydroxide corresponds to 0.0006 g of acetic acid.
Acidity expressed in milligrams of acetic acid per liter of ethanol is equal to 12n.
This acidity should be less than 15 mg/l of ethanol (or 1.5 g/hl) at the time the alcohol is delivered.
(Maximum acidity expressed in terms of acetic acid is 1.5 g/hl of ethanol at 100% by volume).
Note: Indicator movement should be stable and clear cut during quantitative analysis of the acidity. If it is not, and especially if the acidity exceeds 15 mg/l, a new test should be conducted after the sample is degassed using the following technique.
100 ml of alcohol at 50 pp 100 by volume is placed in a 250 ml flask whose stopper has two tubes through it.
One tube permits the flask to be kept under a vacuum using a glass filter pump. Pressure is kept between 55 and 65 cm of mercury.
During the procedure, the other tube allows air bubbling from which carbon dioxide is removed by using a sode wash bottle. To accomplish this, the tube has a capillary portion which is submerged in the alcohol. The rate of air flow through the wash bottle is approximately 1 ml per second.
The procedure should last between 3 and five minutes. Titration is accomplished in the same flask.
4.14. Esters
Add 10 ml of 0.1 M sodium hydroxide solution measured with precision to the solution prepared to analyze acidity as detailed under 4.13 (or 100 ml of alcohol at 50% by volume). Cork the flask and stir while maintaining a temperature equal to or slightly higher than 20 °C. After 24 hours of contact, titrate the excess sodium hydroxide using a 0.1 M solution of hydrochloric acid, where n is the number of milliliters used.
To determine the quantity of 0.1 M hydrochloric acid solution which will neutralize 10 ml of 0.1 M sodium hydroxide solution in the presence of the same quantity of alcohol and of the same indicator movement obtained by decreased pH intervals, perform the following test: place 100 ml of degasified 50 pp 100 alcohol in a 250 ml conical flask. Add one drop of phenol red solution (R) and n milliliters of 0.1 M sodium hydroxide. Which cuase the indicator to turn to red. Add 10 ml of the 0.1 M sodium hydroxide solution, and, immediately thereafter, add 0.1 M hydrochloric acid solution to obtain the same movement of the indicator, that is, nnof the volume used.
1 ml of 0.1 M sodium hydroxide solution corresponds to 0.0088 g of ethyl acetate. The ester concentration, expressed in milligrams of ethyl acetate contained in 1 liter of ethanol is:
This content level should not exceed 13 mg for 1 liter of ethanol (or 1.3 g/hl) at the time the alcohol is delivered.
(Maximum ester content expressed in terms of ethyl acetate is 1.3 g/hl of ethanol at 100% of volume).
4.15. Aldehydes
Standard solution: Place 268.3 mg of pure acetal (boiling point: 102°C) in a 100 ml volumetric flask. Top off to the line with 50 pp 100 alcohol by volume, free of aldehydes.
Dilute this solution to 1/10 in 50 pp 100 alcohol by volume, is free of aldehydes. The solution obtained contains 100 mg of ethanal per liter of 50 pp 100 alcohol by volume, or 20 g in 100 liters of ethanol.
Test procedure: Place 10 ml of alcohol reduced to 50 pp 100 by volume in a test tube. In a second test tube, place 5 ml of the solution containing 100 mg of ethanal per liter of alcohol at 50 pp 100 and 5 ml of alcohol at 50 pp 100 by volume which is free of aldehydes. Add to the two tubes 4 ml aniline red chlorhydrate solution decolorized by sulfuric acid (R), stir, and compare the colorations obtained after 20 minutes.
The alcohol to be tested should have a color approximately equal to that of the standard solution.
(Maximum aldehyde content expressed in ethanal is 0.5 g/hl at 100% of volume).
Note concerning 50 pp 100 alcohol by volume without aldehydes: Place 100 ml of alcohol diluted to 50 pp 100 by volume in a 250 ml flask with 2 g of metaphenylene diamine (R) and two pieces of pumice stone. Connect the flask to a reflux condenser and maintain a gentle boil for one hour. After cooling, connect the flask to the distilling apparatus and slowly distill without overheating the walls. Collect 75 ml of distillate in a 100 ml volumetric flask. Fill to the line with distilled water.
4.16. Superior Alcohols
Propan-1-ol, 2-methylpropan-1-ol, 2- and 3-methylbutan-1-ol.
Quantitative analysis by gas phase chromatography (see methanol).
Maximum content for the sum of each of the alcohols: 0.5 g/hl of ethanol at 100% of volume.
4.17. Furfural
Place 10 ml of alcohol reduced to 50% by volume in a test tube with an emery stopper. Add 0.5 ml of aniline (R) and 2 ml of crystallizable acetic acid (R). Stir. No salmon pink coloration should be perceptible after 20 minutes.
- Storage
The alcohol should be stored in inert containers which will not give off metals, ions or plastic constituents.
The containers, as well as storage methods,must be in compliance with safety standards.
Rectified alcohol of viti-vinicultural origin
COEI-1-ALCVIT Rectified alcohol of viti-vinicultural origin
- Objective, Origin and Scope of Application
Alcohol obtained exclusively by distillation and rectification from wine, grape marcs, wine sediments, or fermented raisins.
Rectified alcohol of viti-vinicultural origin constitues a constituent of some spirits and special wines.
- Composition
At a temperature of 20 °C, 100 parts of this alcohol contain at least 96 parts ethanol.
Note: The tests and controls described below in italics are not mandatory and are performed only upon request.
- Properties
Colorless, clear, volatile liquid with a penetrating odor and fiery taste. It is flammable and burns without smoke and with a blue flame.
It should be distilled completely at between 78 and 79 °C.
3.1. Solubility
Neutral alcohol is miscible in water in all proportions with a notable release of heat and contraction of volume. It is also mixable with in acetone, chloroform, ethyl ether, glycerol, and an equal volume of castor oil.
3.2. Characterization Procedure
- Slowly heat a mixture of 1 ml neutral alcohol, twenty drops of concentrated sulfuric acid (R) and 10 g of sodium acetate (R) in a test tube. A strong, characteristic odor of ethyl acetate will be released.
- Mix several drops of alcohol and 1 ml of concentrated sulfuric acid (R), then add several drops of 10 pp 100 potassium dichromate solution. The liquid will become green and emit the odor of ethanal.
- Dilute 0.5 ml of alcohol with 4.5 ml of water. Add 1 ml of 1M sodium hydroxide solution, then slowly add 2 ml of iodized potassium iodide (R). An odor of iodoform will be produced, following by the formation of a yellow precipitate.
3.3. Determination of Viti-vinicultural Origin
This analysis is carried out by measuring the ethanol 14C/12C ratio (scintillation) in accordance with the method described in the Spirits Compendium.
3.4. If necessary, the viti-vinicultural source of the alcohol can be determined using isotopic methods detailed in the Compendium of Wine and Must Analysis Methods.
- Tests
Test are identical to those for rectified alcohol of agricultural origin , but with the following content limits:
4.1. Methanol
Maximum content 50 g/hl of ethanol at 100% by volume.
4.2. Acidity
Maximum acetic acid content 1.5 g/hl of ethanol at 100% by volume.
4.3. Esters
Maximum content of ethyl acetate 1.3 g/hl of ethanol at 100% by volume (or 5 g/hl).
4.4. Aldehydes
Maximum ethanal content 0.5 g/hl of ethanol at 100% by volume.
4.5. Superior Alcohols
Maximum content 0.5 g/hl of ethanol at 100% by volume.
4.6. Preparing the solution for tests
Using 10 ml of dilute hydrochloric acid (R), take up the residue left by evaporating 100 ml of alcohol during the dry extract analysis. After heating for several minutes in a 100 °C water bath to stimulate dissolution of this residue, decant the acid solution in a 25 ml volumetric flask, and wash the dish three times with 5 ml of water and filled to 25 ml.
4.7. Heavy metals
Place 5 ml of the prepared solution in a test tube in accordance with paragraph 4.6. Add 2 ml of pH 3.5 (R) buffer solution, 7.5 ml of water and 1.2 ml of thioacetamide reagent (R). The solution should not yield any white or black precipitate nor any brown or coloring. At the very least, any coloring produced should be no more intense than that obtained using the general method (heavy metals content expressed in terms of lead, after 50% concentration of the alcohol, should be 0.5 mg/l).
4.8. Lead
Using the method set forth in the Compendium, conduct the quantitative lead analysis on the solution prepared for testing (under paragraph 4.6) (lead content should be less than 0.5 mg/l).
4.9. Mercury
Carry out the quantitative mercury analysis on the solution prepared for testing (under Paragraph 4.6), implementing the technique described in the annex (mercury content should be less than 0.2 mg/l).
4.10. Arsenic
Conduct the quantitative arsenic analysis on the solution prepared for testing (Paragraph 4.6), using the method described in the annex (Arsenic content should be less than 0.5 mg/kg).
- Storage
Alcohol should be stored in inert containers which will not give off metals, ions or plastics constituents.
The containers as well as the storage methods must comply with safety standards.
Ammonium chloride
COEI-1-AMMCHL Ammonium chloride
Ammonia Hydrochloride
Ammonii Chloridum
NCl=53.50
SIN NO.: 510
- Objective, Origin and Domain of Application
This product is used as a fermentation activator and is reserved for fermentation operations. It makes available ammonium ions which can be directly assimilated by the yeast.
Statutory limits regulate the amount of ammonium added.
- Labeling
The concentration of this product should be indicated on the label, including cases in which it is mixed. In addition, safety and storage conditions should be stipulated.
- Centesimal Composition
Cl |
66.22 |
NH3 |
31.78 |
- Properties
Colorless, odorless crystals with a fresh, salty and piquant taste. It sublimes without decomposing and is stable in air.
- Solubility
Water at 20 °C |
350.8 g/l |
Water at 100 °C |
758 g/l |
Alcohol, 95% by vol. |
13.3 g/l |
- Identifying Characteristics
Aqueous solutions of ammonium chloride produce reactions of ammonium and those of chloride.
- Testing
7.1. Sulfur Ash
When quantitifed as indicated in the Annex, the sulfur ash content of the ammonium chloride should not be greater than 0.2 per 100.
7.2. Preparing the solution for tests
Prepare an aqueous solution from NH4Cl crystals at 10 per 100 (m/v).
7.3. Sulfates
To 1 ml of solution prepared for tests under paragraph 7.2, add 2 ml of hydrochloric acid diluted to 10 pp 100 (m/v) (R), 17 ml of water and 2 ml of barium chloride solution (R). The mixture should be clear, or else the opalescence observed after 15 minutes should be less than that of the control solution prepared as indicated in the Annex. (Sulfate content expressed in terms of sulfuric acid should be less than 1 g/kg).
7.4. Nitrates
Mix 5 ml of concentrated sulfuric acid (R) and 0.5 ml of an extemporaneously prepared iron (II) sulfate solution at 5 pp 100 in a test tube. Without mixing, pour 5 ml of the solution prepared under paragraph 7.2. No coloration should be observed at the surface line separating the two solutions.
7.5. Phosphates
To 0.5 ml of the solution prepared for testing under Paragraph 7.2, add 5 ml of water and 10 ml of nitro-vanadomolybdic (R) reagent. Leave in contact for 15 minutes at 20 °C. If a yellow coloration appears, it should be less intense than that obtained by adding 0.5 ml of a solution of 0.05 g of phosphorous per liter (R), 5 ml of water and 10 ml of nitro-vanadomolybdic (R) reagent. (Phosphate content expressed in terms of phosphorous less than 500 mg/kg).
7.6. Iron
To 5 ml of solution prepared under paragraph 7.2, add 1 ml of concentrated hydrochloric acid (R), one drop of 2 pp 100 potassium permanganate and 2 ml of 5 pp 100 potassium thiocyanate (R).
If a red coloration appears, it should be less intense than that of a control prepared with 2.5 ml of an iron (III) solution containing 0.01 g of iron per liter (R), 2.5 ml of water and the same quantities of the same reagents. (Iron content should be less than 50 mg/kg).
The iron may also be quantitatively analyzed using atomic absorption spectometry, in accordance with the method detailed in the Compendium.
7.7. Arsenic
Using the method indicated in the annex, test for arsenic in the test solution prepared in accordance with Paragraph 7.2. (Arsenic content should be less than 3 mg/kg.)
7.8. Lead
Using the method described in the Compendium, quantify the lead in the solution obtained under Paragraph 7.2. (Lead content should be less than 2 mg/kg.)
7.9. Mercury
Using the method described in the annex, test for mercury in the solution prepared for testing under §7.2. (Mercury content should be less than 1 mg/kg.
7.10. Quantitative Ammonia Analysis
Dilute the solution prepared for testing under paragraph 7.2 to one-tenth strength, then place 10 ml of this dilute solution (i.e., 0.1 g of ammonium chloride) in a steam distillation device. Add 10 ml of 30% sodium hydroxide (R) and distill 100 ml. Quantify the distilled ammonia using 0.1 M hydrochloric acid. Let n be the number of milliliters used:
100 g of ammonium chloride contains 1.7 n g of ammonia (NH3).
(Ammonia content greater than 31.5 pp 100).
7.11. Quantitative Hydrochloric Acid Analysis
Take a 10 ml sample of the solution prepared for testing under paragraph 7.2, which has been diluted to one-tenth strength. Place the sample in a cylindrical flask. Add 20 ml of 0.1 M silver nitrate solution, 1 ml of concentrated nitric acid (R), 5 ml of iron (III) sulfate solution and 10 pp 100 of ammonium (R). Titrate the excess silver nitrate with a 0.1 M potassium thiocyanate solution. Let n be the number of milliliters used:
100 g of ammonium chloride contains 3.65 (20-n) g of hydrochloric acid (HCl). (Hydrochloric acid content greater than 67.5 pp 100).
- Storage
Ammonium chloride must be stored in water-tight containers away from heat.
Ammonium hydrogen sulfite
COEI-1-AMMHYD Ammonium hydrogen sulfite
Ammonium bisulfite
NH4HSO3 = 99.07
- Objective, origin and scope of application
This product falls in the category of preservatives and is used exclusively for fermentation operations. It makes available sulfur dioxide and ammonium ions, which can be directly assimilated by the yeast. There are regulatory restrictions on the amount of ammonium that can be added and on sulfur dioxide content.
- Labeling
The concentration of this product, as well as the safety and storage conditions, should be indicated on the label.
- Centesimal composition
NH3 |
17.16 |
SO2 |
64.67 |
- Properties
Ammonium hydrogen sulfite always takes an aqueous solution form. This solution emits a piquant sulfur dioxide odor.
- Solubility
Water at 60 °C |
847 g/l |
Alcohol, 95% by vol. |
Slightly soluble |
- Identifying characteristics
Aqueous solutions of ammonium hydrogen sulfite produce reactions of ammonium (release of ammonia in the presence of sodium hydroxide when heated) and sulfur dioxide (filter paper soaked in potassium iodate and starch turns blue).
- Tests
7.1. Sulfur Ash
As quantified as indicated in the Annex, the proportion of ammonium hydrogen sulfite ash should not be greater than 0.2 per 100.
7.2. Preparing the Solution for Tests
Prepare a 10 pp 100 (m/v) solution.
7.3. Iron
To 5 ml of the solution prepared for testing under paragraph 2, add 1 ml of concentrated hydrochloric acid (R), one drop of 2 pp 100 potassium permanganate (R) and 2 ml of 5 pp 100 potassium thiocyanate (R).
If a red colorating appears, it should be less intense than that of a control prepared with 2.5 ml of an iron (III) solution of 0.01 g of iron per liter (R), 2.5 ml of water and the same quantities of the same reagents.
(Iron content should be less than 50 mg/kg).
The iron may also be quantified by means of atomic absorption spectrometry, using the technique described in the Compendium.
7.4. Lead
Use the method detailed in the Compendium on the solution in a concentration of 10 pp 100 prepared for testing (under 7.2) and diluted to one one-twentieth.
7.5. Mercury
Test for mercury in the solution prepared for testing (under 7.2) using the technique detailed in the annex. (Mercury content should be less than 1 mg/kg.)
7.6. Arsenic
Using the method indicated in the Annex, test for arsenic in 2 ml of the test solution prepared for testing in accordance with paragraph 7.2. (Arsenic content should be less than 3 mg/kg).
7.7. Quantitative Ammonia Analysis
Dilute the solution prepared for testing under paragraph 7.2 to one-tenth strength, then place 10 ml of this dilute solution (0.10 g of ammonium hydrogen sulfite) in a steam distillation device (described in the annex). Add 10 ml of 30 pp 100 sodium hydroxide (R) and distill 100 ml. Quantify the distilled ammonia using 0.1 M hydrochloric acid. Let n be the number of milliliters used:
100 g of ammonium hydrogen sulfite contain 1.7 n g of ammonia (NH3). Ammonia content should be greater than 16.5 pp 100 (m/m).
7.8. Quantitative Sulfur Dioxide Analysis
In a 200 ml conical flask, place 50 ml of cold water, then 5 ml of the freshly prepared ammonium hydrogen sulfite solution. Titrate with 0.05 M iodine in the presence of starch. Let n be the volume of iodine used.
SO2 content per 100 g: 6.4n
Ammonium hydrogen sulfite should contain at least 62 pp 100 SO2.
- Storage
Ammonium hydrogen sulfite solutions should be stored in hermetically sealed containers away from heat and cold.
Diammonium hydrogen phosphate
COEI-1-PHODIA Diammonium hydrogen phosphate
Ammonium hydrogen phosphate
Ammonii phosphas
SIN N°.342
- Objective, Origin and Scope of Application
This product is used as a fermentation activator and is reserved for fermentation operations. It makes available ammonium ions, which can be directly assimilated by the yeast. Excess phosphates can lead to iron breakdown.
Statutory provisions limit the amount of ammonium that can be added.
- Labelling
The concentration of this product should be indicated on the label, including cases of mixtures. In addition, safety and storage conditions should also be stipulated.
- Centesimal Composition
H3PO4 |
74.21 |
P2O5 |
53.75 |
NH3 |
25.79 |
- Properties
Colorless, monoclinic crystals. This salt slowly loses small quantities of ammonia in air.
- Solubility
Water at 20 °C |
689 g/l |
Water at 100 °C |
1060 g/l |
Alcohol, 95% by vol. |
insoluble |
- Identifying Characteristics
6.1. Prepare a 1 pp 100 (m/v) solution in water. The solution has a pH of approximately 8, and a slight pink color is produced with several drops of phenolphthalein (R). At 25 °C, the pH of this solution should be between 7.8 and 8.4.
6.2. This solution produces a yellow precipitate with a nitro-molybdic reagent (R).
6.3. When heated with several drops of 30% sodium hydroxide solution (R), this solution releases ammonia.
- Tests
7.1. Sulfur Ash
Quantified as indicated in the Annex, the proportion of diammonium phosphate ash should not be greater than 5 g/kg.
7.2. Preparing the solution for tests
Prepare a 10 pp 100 (m/v) solution.
7.3. Chlorides
To 0.5 ml of the solution prepared for testing under Paragraph 7.2, add 14.5 ml of water, 5 ml of nitric acid diluted to 10 pp 100 (R) and 0.5 ml of 5 pp 100 silver nitrate solution (R). After 15 minutes at rest in the dark, there should be no clouding, or any clouding visible should be less intense than that observed in the control prepared as detailed in the annex. (Hydrochloric acid content is less than 1 g/kg).
7.4. Sulfates
To 1 ml of solution prepared for tests under paragraph 7.2, add 2 ml of dilute hydrochloric acid (R), 17 ml of water and 2 ml of barium chloride solution (R). The mixture must not form any precipitate or any opalescence; or else, any opalescence that does occur should be less intense than that observed in the control prepared as indicated in the Annex. (Sulfuric acid content should be less than 1 g/kg).
7.5. Oxalic acid
To 5 ml of solution prepared for tests under paragraph 7.2, add 20 drops of acetic acid (R) and 5 ml of solution saturated with calcium sulfate (R). The solution should remain clear.
7.6. Iron
To the 5 ml of solution prepared under paragraph 2, add 1 ml of concentrated hydrochloric acid (R) and 1 ml of 5 pp 100 potassium thiocyanate solution (R).
Coloring should be less intense than that of a control prepared with 2.5 ml of an iron solution in a concentration of 10 mg of iron per liter (R), 2.5 ml of water and the same quantities of the same reagents. (Iron content should be less than 50 mg/kg.)
Iron may also be analytically quantified by atomic absorption spectometry, according to the method specified in the Compendium.
7.7. Lead
By implementing the method detailed in the Compendium, carry out quantitative analysis of the solution prepared for testing according to Paragraph 7.2. (Lead content should be less than 5 mg/kg).
7.8. Mercury
Test for mercury in the solution prepared for testing (Par. 7.2), in accordance with the method detailed in the Compendium. (Mercury content should be less than 1 mg/kg.)
7.9. Arsenic
Using the method indicated in the Annex, test for arsenic in 2 ml of the test solution prepared in accordance with paragraph 7.2. (Arsenic content should be less than 3 mg/kg.)
7.10. Quantitative Ammonia Analysis
Dilute the solution prepared under Paragraph 7.2 to one-tenth strength, then place 10 ml of this dilute solution (0.10 g of ammonium phosphate) in a steam distillation device (described in the Annex). Add 10 ml of water, 10 ml of 30% sodium hydroxide (R) and distill 10 ml. Analytically quantify the distilled ammonia using 0.1 M hydrochloric acid. Let n be the number of milliliters used:
100 g of ammonium phosphate contains 1.7 n g of ammonia (NH3). (Minimum content is 25 pp 100).
7.11. Quantitative Analysis of Phosphoric Acid
Place 25 ml of the solution prepared under paragraph 7.2 in a conical flask. Add 5 drops of phenolphthalein (R). The solution should have a pale pink color. If not, add just enough 0.1 M sodium hydroxide solution to cause incipient movement of the indicator. Add 10 drops of bromocresol green (R) and use a burette to pour 0.5 M sulfuric acid until the indicator turns green.
Let n be the volume in ml used:
One liter of 0.5 M solution corresponds to 71 g of phosphoric anhydride or 98 g of phosphoric acid.
Proportion of ammonium phosphate in g per 100 g:
in phosphoric anhydride |
2.84 n |
in phosphoric acid |
3.92 n |
The proportion of phosphoric anhydride must range between 51.6 and 55 pp 100, or between 71.5 and 76 pp 100 of phosphoric acid.
- Storage
Ammonium phosphate must be stored away from moisture and heat, and in hermetically sealed containers.
Ammonium sulfate
COEI-1-AMMSUL Ammonium sulfate
Ammonium sulfuricum
(NH4)2SO4 = 132.10
SIN NO. 517
- Objective, Origin and Scope of Application
This product is used as a fermentation activator and is reserved for fermentation operations. It adds ammonium ions that can be directly assimilated by the yeast. The sulfates added are completely soluble in wine.
Statutory restrictions govern the addition of ammonium.
- Labelling
The concentration of this product should be indicated on the label, including mixtures. In addition, safety and storage conditions should be noted.
- Centesimal Composition
H2SO4 |
74.22 |
NH3 |
25.78 |
SO3 |
60.59 |
N |
21.20 |
- Properties
Transparent, anhydrous crystals with a bitter, pungent taste, which are similar to potassium sulfate crystals, with which this salt is isomorphous.
- Solubility
Water at 20 °C |
509 g/l |
Water at 100 °C |
1040 g/l |
Alcohol, 90% by vol. |
Insoluble |
Acetone |
Insoluble |
- Identifying Characteristics
Solutions of this salt in water in a concentration of 1 pp 100 (m/v) has a pH of approximately 5.5. This solution allows reactions of ammonium and those involving sulfates.
- Tests
7.1. Sulfur Ash
The concentration of sulfur ash of ammonium sulfate prepared as explained in the annex in a test sample of 1 g must not exceed 5 g/kg.
7.2. Preparing the Solution for Tests
Prepare a 10 pp 100 (m/v) solution.
7.3. Chlorides
To 0.5 ml of the solution prepared for testing under paragraph 7.2, add 14.5 ml of water, 5 ml of nitric acid (R) diluted to a concentration of 10 pp 100 and 0.5 ml of 5 pp 100 silver nitrate solution (R). After 15 minutes at rest in the dark, there should be no clouding; or else, any clouding visible should be less intense than that observed in the control prepared as indicated in the annex. (Hydrochloric acid content must be less than 1 g/kg).
7.4. Phosphates
To 0.5 ml of the solution prepared for tests under paragraph 7.2, add 5 ml of water and 10 ml of nitro-vanadomolybdic reagent (R). Leave in contact for 15 minutes at 20 °C. If a yellow coloring appears, it should be less intense than that obtained by adding, to 0.5 g of a solution containing 0.05 g phosphorous per liter, 5 ml of water and 10 ml of nitro-vanadomolybdic reagent. (Phosphate content expressed in terms of phosphorous should be less than 500 mg/kg).
7.5. Nitrates
Mix 5 ml of concentrated sulfuric acid (R) and 0.5 ml of an previously prepared iron (II) sulfate solution in a concentration of 5 pp 100 (m/v) in a test tube. Without mixing, pour 5 ml of the solution obtained by dissolving 2 g of ammonium sulfate in 10 ml of water. No coloring should be observed at the surface separating the two solutions
7.6. Iron
To 5 ml of the solution prepared for testing under paragraph 7.2, add 1 ml of concentrated hydrochloric acid (R), one drop of 2 pp 100 potassium permanganate (R) and 2 ml of 5 pp 100 potassium thiocyanate solution (R).
If a red coloring appears, it should be less intense than that of a control prepared with 2.5 ml of an iron (III) solution in a concentration of 0.01 g of iron per liter (R), 2.5 ml of water and the same quantities of the same reagents. (Iron content should be less than 50 mg/kg).
The proportion of iron may also be quantified by atomic absorption spectometry, using the technique detailed in the Compendium.
7.7. Lead
Use the quantitative analysis technique detailed in the Compendium on the solution prepared for testing under paragraph 7.2.
(Lead content should be less than 5 mg/kg.)
7.8. Mercury
Test for mercury concentration in the solution prepared for testing (7.2), using the method explained in the annex. (Mercury content should be less than 1 mg/kg.)
7.9. Arsenic
Using the method indicated in the Annex, test for arsenic concentration in the test solution prepared in accordance with paragraph 2. (Arsenic content should be less than 3 mg/kg.)
7.10. Quantitative Analysis of Ammonia
Dilute the test solution prepared under paragraph 7.2 to one-tenth strength, then place 10 ml of this dilute solution (0.10 g of ammonium sulfate) in a steam distillation device (described in the Annex). Add 20 ml of 30% sodium hydroxide (R) and distill 100 ml. Quantitatively analyze the distilled ammonia using 0.1 M hydrochloric acid. Let n be the number of milliliters used:
100 g of ammonium sulfate contains 1.7 n g of ammonia (NH3).
(Ammonia concentration greater than 25 pp 100.)
7.11. Quantitative Analysis of Sulfuric Acid
Dilute the test solution prepared for testing under paragraph 7.2 to one-tenth strength, then take 25 ml of this solution and add 75 ml of water and 1 ml concentrated hydrochloric acid (R). Bring to a boil while slowly adding a small excess of barium chloride solution (R). Let the precipitate form for 30 minutes in a 100 °C water bath. Collect the precipitate, then wash, calcine in an oven at 600 °C and weigh. Let p be the weight of the barium sulfate precipitate:
100 g of ammonium sulfate contains 16.80 p g of sulfuric acid (H2SO4). (Sulfuric acid content greater than 73.5 pp 100.)
- Storage
Ammonium sulfate should be stored in a dry place in hermetically sealed containers, away from heat.
Silver (chloride)
COEI-1-CHLARG Silver[1] chloride
N° CAS : 7783-90-6
- Object, origin and scope of application
This monograph relates to silver chloride used for adsorption into an inert carrier material with a view to its use in wine.
Silver chloride is used for the treatment of wines to remove fermentation and storage-related abnormal odours (odours caused by reduction reactions, characterised by the presence of hydrogen sulphide and thiols).
Silver sulphide formed during the treatment remains adsorbed by the inert carrier material and together they can be separated by filtration.
The inert carrier materials, such as, for instance, kieselguhr (diatomaceous earth), bentonite, kaolin, etc. should comply with the prescriptions of the International Oenological Codex.
- Labelling
The product concentration, batch number, use-by-date, safety warnings and storage conditions should be indicated on the label.
- Appearance
Silver chloride, in its pure state, is a white solid matter.
- Composition (test trials)
The silver chloride used should have a minimum purity of 99%. Determination of the silver content is conducted according to the atomic absorption spectrophotometry (AAS) method (7.8).
The silver chloride content in the inert carrier material should be higher than or equal to 2%.
- Identification of silver chloride
On exposure to light, silver chloride undergoes photolytic decomposition (with darkening).
Silver chloride is partially soluble in a 3% ammoniacal solution (bromide and iodide do not go into solution in the cold) and subsequent addition of potassium iodide solution results in the precipitation of yellow silver iodide (higher sensitivity to light than AgCl). Alternatively, a diluted solution of red potassium hexacyanoferrate(III) can be added instead of iodide. A brown precipitate (Ag3[Fe(CN)6]) is formed.
- Solubility of silver chloride
In water at 25 °C: 0.00188 g/L.
Insoluble in alcohol and nitric acid.
Soluble in sulphuric acid, hydrochloric acid, thiosulphate and ammonium solutions upon complex formation.
- Tests
7.1. Preparation of test solution
Place 0.5 g of sodium chloride and 20 mL of 0.1 mol/L sodium thiosulphate solution in a 50 mL beaker. Mix for 30 minutes. Afterwards, allow to rest/sediment for 5 minutes. Filter the supernatant using a single-use syringe with a filter, pore size 0.45 µm. Transfer 0.5 mL filtrate to a 100 mL volumetric flask and fill up to the calibration mark with distilled water.
7.2. Appearance of test solution
The solution must be colourless, possibly cloudy. The filtrate is colourless.
7.3. Iron
Determine the content according to the atomic absorption spectrophotometry (AAS) method described in Chapter II of the International Oenological Codex; content below 5 mg/kg.
7.4. Nickel
Determine the content according to the atomic absorption spectrophotometry (AAS) method described in Chapter II of the International Oenological Codex; content below 5 mg/kg.
7.5. Lead
Determine the content according to the atomic absorption spectrophotometry (AAS) method described in Chapter II of the International Oenological Codex; content below 5 mg/kg.
7.6. Mercury
Determine the content according to the atomic absorption spectrophotometry (AAS) method described in Chapter II of the International Oenological Codex; content below 1 mg/kg.
7.7. Arsenic
Determine the content according to the atomic absorption spectrophotometry (AAS) method described in Chapter II of the International Oenological Codex; content below 3 mg/kg.
7.8. Silver
Determination by atomic absorption spectrophotometry (AAS), described in the Compendium of International Methods of analysis of wines an musts, after preparation of a test solution (7.1). Calibration with 1 mg/L, 2.5 mg/L and 5 mg/L Ag-reference solutions.
- Storage
Silver chloride must be stored in a dry place, protected from light in hermetically sealed packaging.
[1] Silver chloride used for the treatment of wine should be adsorbed into an inert carrier material
Argon
COEI-1-ARGON
Argon Ar = 40.0
N° SIN: 938
N°CAS = 7440-37-1
- Object, origin and field of application
Neutral gas, used for operations of inerting or degassing, it is used in a mixture of nitrogen and/or of carbon dioxide.
- Labelling
The label must mention the nature of the gas and refer to its composition and purity. The safety conditions must also be indicated on the packages.
- Characteristics
Colourless and odourless gas without flavour. Non flammable, it does not support combustion.
The weight of a litre of argon under the pressure of 760 mm of mercury is 1.784 g at 0°C. A volume of water dissolves 0.0336 volume of argon at 20°C.
- Test trials
The global purity of the argon used in oenology must not be less than 99% of argon in volume.
Before any measurement it is advisable to allow any gas to escape for a few minutes in order to purge the piping.
4.1. Chromatographic dosage
The search and determination of gases: Nitrogen, carbon monoxide (less than 10 µl/l), oxygen (10 ml/l), hydrogen, carbon dioxide (less than 300 µl/l), etc., are quickly obtained by chromatography in gaseous phase according to the method in chapter II of the International Oenological Codex.
The total surface are of hydrogen chromatographic peaks, of oxygen and nitrogen must not exceed 1% of gas surfaces to be examined.
The following chemical methods can also be used for oxygen.
4.2. Oxygen dosage by chemical method
Preparation of the flask for searching oxygen:
- Introduce in a 24 ml flask about two fragments of copper turnings of 2 cm, 16 ml of ammoniac solution of copper sulphate (R), then 2 ml of hydrazine dihydrochloride solution (R).
- Seal the flask with a rubber stopper that is easy to pierce with a needle for hypodermic injections. Crimp the neck with a metallic capsule. Then cover the capsule with wax in order to ensure perfect water tightness. Shake the flask and allow to stand away from light until complete discolouration is obtained after about eight days.
Conduct of the test trial:
- Pierce the flask’s stopper to search for oxygen with a needle of 8/10 millimetre for hypodermic injection (take care so as not to plunge it into the liquid) that then will be used for evacuating the gas after bubbling. Then introduce a second needle of the same diameter releasing the gas and plunging it into the liquid. After a minute of bubbling, a noticeable colouration should not be observed. In the presence of oxygen, the liquid quickly becomes blue and the colour darkens with time.
- Packaging
The argon is supplied in highly resistant steel cylinders painted in white with needle valves. The resistance of these cylinders must be checked periodically.
Ascorbic acid
COEI-1-ASCACI Ascorbic acid
2,3-didehydro-L-threohexano-4-lactone
Acidum ascorbicum
L-Ascorbic Acid
C6H8O6 = 176.1
SIN NO. 300
- Objective, Origin and Scope of Application
Ascorbic acid is the enolic form of 3-oxo-L-gulofuranolactone (2,3-didehydro-L-threohexano-4-lactone).
This product falls into the category of antioxidants and is used as a reducing agent used to prevent oxidation.
Its use is subject to statutory regulations regarding limits.
- Labelling
The concentration of this product should be indicated on the label, including cases in which it is used in mixtures, as should the safety and storage conditions.
- Properties
Odorless white or very pale yellow crystalline powder with an acidic flavor. Aqueous solutions rapidly decay in air and light and have a maximum stability at pH 5.4. Melting point in a capillary tube: approximately 190 °C with decomposition.
Ascorbic acid in aqueous solution has a pH of less than or equal to 3.
- Solubility
Water at 20 °C |
290 g/l |
Alcohol, 95% by vol. |
320 g/l |
Methanol |
125 g/l |
Acetone |
soluble |
Benzene, chloroform, ethyl ether, petroleum ether |
insoluble |
- Rotatory Power
In a 10 pp 100 (m/v) aqueous solution, ascorbic acid has a specific rotatory power
is between +20,5° et +21,5° |
- Absorption in Ultraviolet Light
Ascorbic acid in alcohol solutions in a concentration of 10 mg/l exihibits an absorption spectrum with a maximum of approximately 244 nm.
The solution has a specific extinction of:
approximately 560 |
- Identifying Characteristics
7.1. Preparation of the Solution for Testing
Dissolve 5 g ascorbic acid in water and fill to 100 ml using the same solvent.
7.2. Add 0.5 g monosodium carbonate to 2 ml of the solution prepared for testing (Par. 7.1).
7.3. Add several drops nitric acid diluted to 10 pp 100 (R) and several drops silver nitrate in a concentration of 1 pp 100 (R) to 1 ml of the solution prepared for tests (Par. 7.1). A gray precipitate will form.
7.4. To 1 ml of the solution prepared for testing (Par. 7.1) add one drop of recently prepared sodium nitrohexacyanoferrate (III) Na2[Fe(CN)5NO], 2H2O (sodium pentacyanonitrosylferrate) in a concentration of 5 pp 100 (m.v), and 2 ml of 10 pp 100 diluted sodium hydroxide solution (R). Then, add 0.6-0.7 ml of concentrated hydrochloric acid (R) and stir. The yellow color will turn to blue.
7.5. Add drop by drop 2 ml of 2,6-dichlorophenolindophenol solution (R) to the solution prepared for testing (Par. 7.1). It will instantly become decolored.
- Tests
8.1. Sulfur Ash
As determined in 1.0 g ascorbic acid, the proportion of sulfur asheshould not be greater than 1 g/kg.
8.2. Appearance of the Solution
The solution prepared for tests under paragraph 7.1 should be clear and colorless.
8.3. Determining pH
The pH of the solution prepared for tests under paragraph 7.1 should be between 2.4 and 2.8.
8.4. Heavy metals
10 ml of the solution prepared for tests under paragraph 7.1 should meet the heavy metal limit requirements described in the annex. (The heavy metal concentration expressed in terms of lead should be less than 10 mg/kg).
8.5. Lead
Use the technique detailed in the Compendium to analyze the solution prepared for tests (Par. 7.1). (Lead concentration should be less than 2 mg/kg).
8.6. Mercury
Use the technique described in the annex to analyze the solution prepared for tests (Par. 7.1). (Mercury concentration should be less than 1 mg/kg.)
8.7. Arsenic
Using the method indicated in the Annex, test for arsenic in the test solution prepared in accordance with paragraph 7.1. (Arsenic concentration should be less than 3 mg/kg).
8.8. Iron
Implement the atomic absorption technique described in the Compendium to analyze the solution prepared for tests (Par. 7.1). (Iron concentration should be less than 5 mg/kg.)
8.9. Copper
Implement the atomic absorption technique described in the Compendium to analyze the solution prepared for tests (Par. 7.1). (Copper concentration should be less than 2 mg/kg.)
8.10. Moisture
Dehydration loss after drying in a desiccator under a vacuum and in the presence of sulfuric acid for 24 hours must be less than 0.4%.
8.11. Quantitative Analysis
In 80 ml of recently boiled and cooled water to which 10 ml of sulfuric acid diluted to 10 pp 100 (R) has been added, dissolve a test sample weighed precisely at about 0.20 g. Add 1 ml of starch (R) and titrate using 0.05 M iodine until a persistent blue coloration appears.
1 ml of 0.05 M iodine corresponds to 8.81 mg ascorbic acid.
The product should contain at least 99 pp 100 ascorbic acid.
- Storage
Ascorbic acid should be stored in tightly sealed non-metal containers in a dark place. Aqueous solutions decay rapidly in air and light.
Isoascorbic acid
Isoascorbic acid, or D-ascorbic acid or erythorbic acid has the same antioxidant power as ascorbic acid and can be used for the same oenological purpose.
This acid exhibits the same appearance and the same solubility properties as ascorbic acid.
It is, optically, the reverse of ascorbic acid and has, under the same conditions, a specific rotatory power of:
is between -20° et -21,5° |
With the exception of rotatory power, this acid should exhibit the same properties as ascorbic acid, respond in the same way to the identifying reactions, pass the same tests and responds to the same quantitative analysis.
Note 1: The vitamin C efficacity of isoascorbic acid is approximately 1/20 of that of ascorbic acid.
Note 2: There is a preliminary draft resolution calling for registration of this product in the International Code of Oenological Practices.
Nitrogen
COEI-1-AZOTE Nitrogen
Nitrogenum
N = 14.007
SIN NO. 941
- Objective, Origin and Scope of Application
Neutral gas used to render inert or to degas. It can be used pure or mixed with carbon dioxide.
- Labelling
The label should mention the nature of this gas and reference its composition and purity, as well as its safety and storage conditions.
- Properties
Colorless, odorless, flavorless gas. It is not flammable and does not maintain combustion.
The weight of a liter of nitrogen under normal conditions is 1.250 g.
Under a pressure of 760 mm of mercury at 20 °C, a volume of water dissolves a 0.01507 volume of nitrogen, while a volume of alcohol dissolves a 0.1224 volume of nitrogen.
- Tests
The purity of nitrogen used for oenological purposes should be 99 parts per 100 by volume.
Before undertaking any measurement, the gas should be allowed to escape for several moments in order to clean out the lines.
Gas detection and quantitative analysis: oxygen, carbon monoxide, argon, carbon dioxide, etc. are most rapidly detected using gas phase chromatography. (See this method in the Annex.)
The following chemical methods can also be used.
4.1. Phosphorous-containing Hydrogen, Arsenical Hydrogen and Reducing Substances
Let 1 liter of nitrogen to flow into a mixture of 10 ml of ammoniacal silver nitrate (R) and 15 ml of water.
Regulate the flow of gas so that the gas flows into the solution in approximately 15 minutes.
There should be no clouding or brown coloration when compared with an identical control solution through which no gas will flow.
4.2. Oxygen
Prepare a flask to test for oxygen as follows:
Place 2 turned pieces of copper of approximately 2 cm2, 16 ml of ammoniacal copper sulfate solution (R) and 2 ml of hydrazine dichlorhydrate in a 24 ml flask.
Stop the flask with a rubber stopper which can easily be pierced with a hypodermic needle. Seal the collar with a metal cap, then cover the cap with wax to ensure a perfectly airtight seal. Shake the flask, then let it sit in the dark until the color disppears completely, after approximately eight days.
Conducting the test:
Pierce the flask stopper with a 8/10 mm hypodermic needle (take care not to dip the needle into the liquid). This will allow gas to escape after bubbling. Next, insert a second hypodermic needle of the same diameter and plunge it into the liquid. After a minute of bubbling, there should be no significant coloring. In the presence of oxygen, the liquid will rapidly turn blue and the color becomes more intense over time.
The nitrogen must incorporate less than 10 ml/l oxygen.
- Packing and Storage
Nitrogen is delivered in high-strength steel canisters which are painted black and equipped with a needle valve tap. The strength of the canisters should be checked periodically.
Lactic acid bacteria
COEI-1-BALACT Lactic acid bacteria
- Object, origin and field of application
Lactic acid bacteria are used in oenology to perform malolactic fermentation. The lactic acid bacteria must belong to the Oenococcus, Leuconostoc, Lactobacillus and Pediococcus genus and must be isolated from grapes, musts, wine or have been derived from these bacteria.
The use of genetically modified bacteria will be governed by the currently applicable legislation.
The strains of lactic acid bacteria must be kept under conditions which most favour their genetic stability.
Lactic acid bacteria used in oenology must transform the malic acid in must and wine into lactic acid and carbon dioxide. This should produce biogenic amines in the smallest possible quantities, and must not produce an off taste.
- Labelling
The following information must be indicated on the label:
- The genus name and specie(s) in addition to the reference(s) of the strain(s) in the case that there is a registration body.
- Selecting body
- Operating instructions method and possible reactivation additives recommended by the manufacturer.
- The minimum number of viable cells per gram of preparation that is guaranteed by the manufacturer,
- The manufacturing batch number, in addition to the expiration date and storage conditions with a storage temperature recommended by the manufacturer.
- Where relevant, the indication that lactic acid bacteria were obtained by genetic modifications and their modified character(s).
- The additives.
- Characteristics
Lactic acid bacteria are marketed in liquid, frozen or powder form obtained by lyophilisation or drying, in pure culture or in association with pure cultures.
- Test trials
4.1. Humidity for lyophilisated or dried bacteria
Measured by the weight loss of 5 g of the product, dried at 105 °C until constant weight (about 3 hours).
Maximum content should not exceed 8 %.
4.2. Lead
Proceed with the determination according to the method in chapter II of the International Oenological Codex.
Content should be less than 2 mg/kg of dry matter.
4.3. Mercury
Proceed with the determination according to the method in chapter II of the International Oenological Codex.
Content should be less than 1 mg/kg of dry matter.
4.4. Arsenic
Proceed with the determination according to the method in chapter II of the International Oenological Codex.
Content should be less than 3 mg/kg of dry matter.
4.5. Cadmium
Proceed with the determination according to the method in chapter II of the International Oenological Codex.
Content should be less than 1 mg/kg of dry matter.
4.6. Viable lactic acid bacteria[1]
Proceed with counting according to the method in chapter II of the International Oenological Codex.
The number should be more or equal to 108 CFU/ml for frozen or liquid bacteria.
The number should be more or equal to 1011 CFU/g for lyophilisated or dried bacteria.
4.7. Mould
Proceed with counting according to the method in chapter II of the International Oenological Codex.
The number should be less than 103 CFU/g.
4.8. Contaminant acetic acid bacteria
Proceed with counting according to the methods in chapter II of the International Oenological Codex.
The number of acetic bacteria should be less than 103 CFU/g for frozen or liquid lactic acid bacteria or 104 CFU/g for lyophilisated or dried lactic acid bacteria.
The sum of Acetobacter + Gluconobacter should be less than 103 CFU/ml for frozen or liquid lactic acid bacteria or 104 CFU/g for lyophilisated or dried lactic acid bacteria.
4.9. Yeasts contaminants
Proceed with counting according to the methods in chapter II of the International Oenological Codex.
The number of viable cells of total contaminant yeasts must be less than 103 CFU/g for lyophilisated or dried lactic acid bacteria or 102 CFU/ml for frozen or liquid lactic acid bacteria.
4.10. Salmonella
Proceed with counting according to the method in chapter II of the International Oenological Codex.
Absence should be checked on a 25 g sample.
4.11. Pseudomonas aeruginosa[2]
4.12. Escherichia coli
Proceed with counting according to the method in chapter II of the International Oenological Codex using a selective differential medium for Escherichia coli. MET in the annex. A lactic acid bacteria stock suspension is carried out in a trypone salt solution using 1 g of lactic acid bacteria for 10 ml of solution (total volume). 2 ml of stock solution is transferred to each dish using 5 different dishes. Absence should be checked on 1 g sample.
4.13. Staphylococci
Proceed with counting according to the method in chapter II of the International Oenological Codex. The presence of staphylococci is evaluated by an enrichment culture in a liquid Giolitti and Cantoni medium followed by a confirmation on a solid Baird Parker medium in the annex.
A lactic acid bacteria stock suspension is carried out in a salt tryptone solution using 1 g of lactic acid bacteria for 10 ml of solution (total volume). 10 ml of stock suspension is used to inoculate a Giolitti and Cantoni medium to Tween 80 double concentration. Cultures are incubated 48 hours at 37 °C.
In the case that the Giolitti and Cantoni medium gives positive results, the presence of Staphylococci is confirmed by isolation on a solid Barid Parker medium. A positive culture medium loop is used to inoculate solid BP mediums to obtain isolated colonies.
Absence should be checked on 1 g sample.
4.14. Coliforms
Proceed with counting according to the method in chapter II of the International Oenological Codex using a selective differential medium for coliforms, desoxycholate gelose in the annex. A lactic acid bacteria stock suspension is carried out in a salt tryptone solution using 1 g of lactic acid bacteria for 10 ml of solution (total volume). 2 ml of stock solution are transferred is each dish using 5 different dishes.
The number of coliforms should be less than 102 CFU/g.
- Additives
They must be in conformity with regulations in force.
- Storage conditions
Always refer to manufacturer’s recommendations.
Appendix: Preparation of a leaven “pied de cuve malo” to innoculate 100hL of wine or any volume from the values in brackets in %, the quantities of powder are expressed in g/L |
[1] Except for specific bacteria intended for acidic wines (pH up to 2.85) that should be used with a pre-multiplication process (see Annex) in the must or wine, where the population cannot be less than 109 CFU/g.
[2] Point to be studied at a later date by the expert group “Microbiology”.
Bentonites
COEI-1-BENTON Bentonites
Bentonita
N° SIN: 558
- Object, origin and field of application
Bentonites are hydrous aluminium silicates belonging to the montmorillonite group. The brute formula is:
Si4 (Al (2-x) Rx) (O10, H2O)(Cex, nH2O) or Si4(Al(2-x)Rx)(H2O)n where: |
R = Mg, Fe, M, Zn, Ni |
Ce (exchangeable cations) = Ca, Na, Mg. |
Bentonites are used for clarification operations or protein stabilisation in musts and wine. Bentonites fix to certain unstable proteins which allows them to be eliminated.
Bentonites are capable of fixing coloured matter.
- Labelling
The nature of the bentonite (natural sodium, calcium, and activated calcium), batch number and the optimal expiration date for activated bentonites will be indicated on the label. The mention of risks and safety concerning the presence of crystalline silica should also be indicated.
2.1. Natural Bentonites:
Depending on the nature of the of exchangeable cations present, there are 2 naturally occurring types of bentonite:
- Sodium bentonite, it swells and absorbs readily where sodium is the major exchangeable cation.
- Calcium bentonite, where calcium is the major exchangeable cation, it is lower swelling and lower absorbent than sodium bentonites.
These two types of betonites are simply grinded before their commercialisation after possibly being dried at 80°C to 90°C.
2.2. Activated bentonites:
In order to improve the adsorption properties of calcium betonites, they are most often activated by sodium carbonate, then dried and grinded. This results in activated calcium bentonites with properties equal or superior to sodium bentonites.
The properties of these bentonites thus activated or permuted are less stable in time (3 to 18 months) and depend on the activation of magnesium, calcium, and sodium levels.
These different types of bentonites are in the form of powder, spherical or cylindrical granules. Colour can vary from white for the purest products to grey, beige or green for others.
- Test trials
3.1. Odour
Bentonite should not have any undesirable odour (e.g. no mould) and should not change the taste of wine.
3.2. pH level
Shake 5g of bentonite with 100 ml of distilled water for 5 minutes. Allow to stand for 1 hour. Measure the pH level of the supernatant liquid. Natural calcium bentonites have a neutral pH (level around 6.5 to 8.5) whereas activated calcium bentonites have a much more alkaline pH (level around 8.5 to 10.0). Natural sodium bentonites have a wider range of pH (level around 4.7 to 10.0).
3.3. Loss during desiccation
The desiccation of 5 g of bentonite at 105°C during 4 hours causes a weight loss of 5% to 15% of the initial weight (often around 10%).
3.4. Preparation of the test trial solution
Weigh p g of bentonite containing 10 g of anhydrous bentonite.
In a 500 ml flask with a large opening which can be hermetically sealed, add 100 ml of tartaric acid solution to 5 g per litre until the solution has a pH level of 3 (R). Sprinkle the bentonite trial sample in the constantly shaken solution (for example using a magnetic stirrer) and a funnel. After this addition, shake vigorously for 5 minutes. Allow to stand for 24 to 48 hours. Decant, centrifuge or filter if necessary to obtain at least 100 ml of clear liquid.
All the following set limits for bentonite are for the weight of dried bentonite.
3.5. Montmorillonite content
Minimum rate:
- Manufacturer indicates that the content should not be under 80% by x-ray diffraction analysis.
3.6. Different forms of free silica content
- Crystal silica content must be less than 3% (quartz N° CAS 14080-60-7, cristobalite N° CAS 14464-46-1).
- Particle holdings under 10 microns must be less than 10%.
- Respirable crystal silica content must be under 0.3%.
These standards must be written on the security form supplied by the manufacturer.
3.7. Lead
In the test trial solution (3.4) determine the lead content using the method described in Chapter II.
Lead content must be less than 5 mg/kg.
3.8. Mercury
Determine the mercury content according to the method described in Chapter II with the test trial solution (3.4).
Mercury content should be less than 1 mg/kg.
3.9. Arsenic
Determine the arsenic content of 5 ml of test trial solution (3.4) according to the method in Chapter II.
Soluble arsenic content should be less than 2 mg/kg.
3.10. Iron
Add 12.5 ml of water, 1 ml concentrated hydrochloric acid (R) and 2 ml of potassium thiocyanate at 5% (R) to 5 ml of the test trial solution (3.4). The red coloration should be lighter than what is obtained when using 2.5 ml citric acid at 5% at pH 3 (R), 1 ml concentrated hydrochloric acid (R), 15 ml of iron salt solution (III) at 0.010 g of iron per litre (R) and 2 ml of potassium thiocyanate solution at 5% (R).
Iron content should be less than 600 mg/kg).
Iron can also be determined by atomic absorption spectrometry according to the method in Chapter II.
3.11. Aluminium
On the test trial solution (3.4), find extractable aluminium according to the method described in Chapter II.
Extractable aluminium content should be less than 2.5 g/kg.
3.12. Calcium and magnesium
On the test trial solution (3.4), determine calcium and magnesium using the methods outlined in the Compendium of International Methods of Analysis of Wine and Musts.
Calcium and soluble magnesium combined should be less than 100 meq for 100 g.
3.13. Sodium
On the test trial solution (3.4), determine sodium using the method outlined in the Compendium of International Methods of Analysis of Wine and Musts.
Soluble sodium content should be less than 10 g/kg for natural bentonites and less than or equal to 35 g/kg for activated bentonites.
3.14. Presence of large particles
- Put 1 litre of water in a 1.5 litre long stem glass.
- Slowly add while shaking the liquid, a quantity of bentonite corresponding to 50 g of dried bentonite.
- Shake vigorously 2 to 3 minutes and allow to stand for 24 hours.
- Shake 2 to 3 minutes and allow to stand for 2 minutes.
- Using a siphon, take off 9/10 of the cloudy liquid exceeding 100 ml and leave the deposits at the bottom of the glass.
- Add 900 ml of water.
- Shake 1 minute.
- Allow to stand for 2 minutes and repeat to obtain 5 washings.
- Remove the deposit and put in a capsule.
- Dry and weigh.
- The residue must be less than 8 g for 100 g.
3.15. De-acidification tests trials
- Weigh (p) of bentonite containing 0.2 g of dried bentonite.
- Put this in a 125 ml flask containing 50 ml of citric acid 0.033 M solution (R).
- Shake vigorously for 5 minutes and allow to stand for 30 minutes.
- Either filter or centrifuge.
- Take 10 ml of filtrate and titrate with an acid solution of 0.1 M of sodium hydroxide with a drop of phenolphthalein solution (R), that is n ml the volume poured to obtain a colour change in the indicator: 250 (10 – n) is the number of milliequivalent of acids fixed or neutralised for 100 g of bentonite.
- The maximum limit is 2.5 eq/kg.
3.16. Rate of swelling
Swelling indicator: specific test is necessary.
2 g of bentonite is strewn over 100 ml of demineralised water and 100 ml of wine in a graduated test tube cylinder. After 24 hours, weigh the volume of bentonite. This will be expressed in ml/g of dried product.
3.17. Protein adsorption test trial (for bentonite to go through deproteinisation)
3.17.1. Preparation of test trial solution:
Mix 5 g of egg white with a sufficient amount of citric acid solution of 5 g per litre (pH=3) to make 1 litre. Filter. Determine total nitrogen on 100 ml of this solution by using the procedure described in Chapter II. This solution contains approximately 90 mg of total nitrogen for 575 mg of proteins per litre.
3.17.2. For each test trial using 100 ml of this solution, mix increasingly larger doses of bentonites prepared in a 5% suspension in order to process doses of 0.1 to 0.8 g/l. Shake vigorously and maintain at 15°C–20 °C for 6 hours. Centrifuge and proceed with determinations of nitrogen or residual proteins.
A de-proteinising bentonite should eliminate at least 50% of the proteins in a synthetic solution with a 0.4 g/l dose.
3.18. Determining the specific adsorption surface (or the adsorption indicator for methylene blue)
See method described in annex.
The accepted limit should be 300 mg/100g.
- Storage
Bentonites must be stored in a ventilated area in watertight containers away from volatile objects that they could adsorb.
Annex: Determination of the specific surface of adsorption of bentonite
- General information
1.1. Aim of the test trial
This test trial enables to measure the capacity of bentonite to adsorb methylene blue.
Clays, organic matters, and iron hydroxide preferentially adsorb methylene blue. This capacity takes into account the activity on the surface of these elements. We call, “blue value” of bentonites, the quantity expressed in grams of methylene blue adsorbed per 100 g of bentonites.
1.2. Principle of the test trial
Elemental doses of a methylene blue solution are injected successively into an aqueous solution containing the trial sample. The adsorption of blue is checked after each addition by making a spot on a paper filter (spot test, see paragraph 5).
For a simple conformity check, the specified quantity of blue is injected once.
- Equipment and reagent
2.1. A 25 ml burette graduated 1/10 ml.
2.2. Paper filter: quantitative and without ashes (< 0.010); weight: 95 g/m2; thickness: 0.20 mm; filtration speed 75; retention: 8 micrometers.
2.3. A glass rod: 300 mm length; 8 mm diameter.
2.4. A magnetic stirrer and magnetic stirring bar.
2.5. Methylene blue of medicinal quality at 10g/l 0.1 g/l.
The maximum duration for using the solution is one month. The solution must be stored away from light.
2.6. Demineralised or distilled water.
- Preparation of test trial samples
Add 10 g of bentonite in 200 ml of distilled water, allow to swell for 2 hours, then homogenise by shaking.
- Carrying out test trial
4.1. Definition of spot test
After each addition of blue (see paragraph 5.2), this test involves taking a drop of suspension that is placed on a paper filter using a glass rod. The spot that is formed is composed of a central deposit of matter, blue in colour surrounded by a humid colourless area.
The drop must be such that the diameter of the deposit is between 8 and 12 mm.
The test is positive if a persistent light blue ring appears around the middle deposit in the humid zone. The test is negative if the ring is colourless.
4.2. Determination
Using a burette, pour 2 ml of blue solution in a container with 200 ml of suspension of bentonite maintained in agitation. After 2 minutes, add 1 ml of blue solution. This addition is followed by the spot test on filter paper.
Allow the asorption of blue to occur which is not instantaneous. Meanwhile tests should be conducted minute by minute.
If the light blue ring disappears at the fifth spot, proceed with elemental additions of 0.2 ml of blue and then 0.1 ml.
Each addition is followed by tests conducted minute by minute.
Renew these operations until the test remains positive for 5 consecutive minutes: the determination is considered as ended.
That is V ml poured.
- Expression of results
5.1. Blue value
The blue value expressed in grams of blue for 100 g of bentonite is shown in the following formula:
- V is the value of blue methylene poured in ml.
5.2. Conformity check compared to a given specification
The specification is expressed in blue value for 100 g of bentonite, or s of this value.
The volume of blue solution to be added in one time to the preparation (3) is:
The spot test is done after eight minutes of shaking. If it is negative, the bentonite complies with the specification.
Beta-glucanase
COEI-1-BGLUCA Beta-Glucanases from Trichoderma Sp.
(E.C. 3-2-1-58)
(C.A.S. No. 9073-49-8)
Glucan 1,3-beta-glucosidase
(exo-1,3-beta-glucosidase; beta-1,3-glucan exo-hydrolase; exo-1,3-beta-glucanase; endo-1,3-beta-glucanase)
and glucan 1,6-beta-glucosidase
General specifications
The specifications must comply with general specifications for enzymatic preparations that appear in the International Oenological Codex.
- Object, origin and field of application
The degradation of beta-glucans present in wines, in particular those from grapes affected by Botrytis cinerea or yeast glucans. These molecules of a very high molecular weight hydrolyse the beta-1,3 and beta-1,6 bonds of 1,3 (1,6)-beta-D-glucans with glucose production.
Secondary activities: hemicellulases, cellulases.
The beta-1,3-D-glucanases are produced from Trichoderma harzianum .and/or Trichoderma ressei
The preparation of the enzyme is without any harmful consequences as is production and purification.
Beta-glucanases do not contain any substances, micro-organisms nor collateral enzymatic activities that can:
- be harmful to health,
- be harmful to the quality of the products treated,
- lead to the formation of undesirable products or flavour problems.
There are regulatory limits for the use of beta-glucanases in wine.
- Labelling
The concentration of the product must be indicated on the label, as well as the safety conditions, storage conditions and the expiry date.
- Characteristics
In general, it is greyish to light brown amorphous powder or light brown to dark brown liquids or granules.
- Solubility
Soluble in water and practically insoluble in ethanol.
- Enzymatic activity
Activity is the quantity of enzyme necessary for liberating in standardised conditions (see activity measured according to a method to be described), a quantity of reducing sugars corresponding to 1 µmole of glucose per minute.
Remark: the enzyme produced according to paragraph 6 simultaneously has beta-1,3-glucanase and beta-1,6-glucanase activities which gives it the sought oenological properties.
- Source of enzyme and production means
The beta (-1,3-1,6) glucanases are produced by submerged culture of a selected non pathogenic, non toxic strain of Trichoderma harzianum and/or Resei that is not genetically modified, in pure culture.
- Diluents, preservatives and additives
The preparation of beta-glucanase is generally in the form of granules. These products are prepared with food diluents or food additives such as maltodextrin, sodium citrate, citric acid, starch or glucose.
- Test trials
8.1. Loss at desiccation: Less than 10%. (does not apply to liquid preparations)
8.2. Ashes/Sulphuric ashes
Determine the sulphuric cinders according to the method in Chapter II of the International Oenological Codex.
The rate of sulphuric ashes of beta-glucanases should not be more than 2% of dry matter.
8.3. Preparation of the test solution
Dissolve 5 g of beta-glucanases in 100 ml of water.
8.4. Heavy metals
Add 2 ml of buffer solution pH 3.5 (R) and 1.2 ml of thioacetamide reagent (R) to 10 ml of the test trial solution (8.3). No precipitate should form. If a brown colouration appears, it should be
lighter than the control prepared as indicated in Chapter II of the International Oenological Codex.
The heavy metal content expressed in lead should be less than 30 mg/kg.
8.5. Arsenic
In 2 ml of test trial solution (8.3), search by the method indicated in
- Chapter II of the International Oenological Codex.
Arsenic content should be less than 3 mg/kg.
8.6. Lead
Using the test trial solution (8.3) determine the lead according to the method described.
Lead content should be less than 5 mg/kg.
8.7. Mercury
Using the test trial solution (8.3) determine the mercury according to the method described in Chapter II of the International Oenological Codex.
Mercury content should be less than 0.5 mg/kg.
8.8. Cadmium
Using the test trial solution (8.3) determine the cadmium according to the method described in chapter II of the International Oenological Codex.
Cadmium content should be less than 0.5 mg/kg.
8.9. Biological contaminants
Total microorganisms |
less than 5 104 CFU/g of preparation |
Total bacteria |
less than 103 CFU/g of preparation |
Total coliforms |
less than 30 CFU/g of preparation |
Escherichia coli |
absence checked on a 25 g sample |
St. aureus[1] |
absence checked on a 1 g sample |
Salmonella |
absence checked on a 25 g sample |
Sulfitoreducing anaerobia |
less than 30 CFU/g of preparation |
Yeasts |
maximum content 102 CFU/g of preparation |
Total lactic bacteria |
absence checked on a 10 g sample |
Acetic bacteria |
maximum content 102 CFU/ g of preparation |
Moulds |
maximum content 102 CFU/g of preparation |
Antibiotic activity[2] |
not detectable |
Mycotoxins[3] |
not detectable |
- Storage
In a solid form, the preparation can be stored for several years and in a liquid form for several months at a low temperature (+5°C).
[1] Method to be defined by the Sub-Commission of Methods of Analysis
[2] Method to be defined by the Sub-Commission of Methods of Analysis
[3] Method to be defined by the Sub-Commission of Methods of Analysis
Pieces of oak wood
COEI-1-WOOPIE Pieces of oak wood
- Object, origin and field of application
Pieces of oak wood used for winemaking and for passing on certain constituents to the wine in conditions set by regulations.
The pieces of oak wood must come exclusively from the Quercus genus.
They can possibly be left in their natural state or they can be heated to a low, medium or high temperature but they must not be charred including on the surface, nor be carbonaceous, nor friable when touched.
No compound should be added to them for the purpose of increasing their natural aromatising capacity or their extractible phenolic compounds.
Likewise, they must not undergo any chemical, enzymatic or physical treatment other than heating.
- Labelling
The label must mention the varietal origin of the oak and the intensity of any heating, the storage conditions and safety precautions.
- Dimensions
The dimensions of these particles must be such that at least 95% in weight be retained by the screen of 2 mm mesh (9 mesh).
- Purity
The pieces of oak wood must not release any substances in concentrations which may be harmful to health,
- Storage conditions
The pieces of oak wood must be stored in sufficiently dry and odourless conditions free from substances liable to contaminate them.
- Introduction in wine
Where bags or other containers are used as the means of introducing pieces of oak wood or related support system into wine, they must be made from materials that are approved for food contact in the country of use, and which do not release any substances into the wine in concentrations which may be harmful to health, or jeopardise to the quality of the final product.
Annex A: Determination of the size of pieces of oak wood by screening
- Introduction
The use of pieces of oak wood, commonly called chips, to treat wine is authorised provided they comply with the specifications of the Oenological Codex (resolution OENO 3/2005). In particular, the pieces of oak wood used must meet a size requirement, and it is specified that "The dimensions of these particles must be such that at least 95% in weight be retained by the screen of 2 mm mesh (9 mesh)". The following operating procedure provides a method of sampling and then screening that can be used to verify this requirement.
- Field of application
The method applies to oak wood test samples of more than 0.5 kg.
- Principle
After dividing up the initial test sample, a known quantity of pieces of oak wood (approximately 200g) is placed on a vibrating screen. By weighing the pieces of oak wood remaining on the screen after shaking, it is possible to determine the percentage by weight of particles retained by the screen.
- Equipment
- Standard laboratory equipment.
- Screen of 2 mm mesh (9 mesh), 30 cm in diameter, mounted on a vibrating plate provided with a recovery tray.
- Weighing machine capable of weighing to within 0.1 g.
- Slotted test specimen divider (see figure below as an example).
Slotted test sample divider (EN 1482-1: 2007 Scheme proposed as an example |
- Division of test sample
When the size of the test sample has to be reduced to obtain “sub-samples” of 200 g which retain a homogeneous nature representative of the initial test sample, a slotted test sample divider can be used which allows random separation of the test sample into 2 parts.
The test sample is poured entirely into the divider in order to separate it into two statistically equivalent parts. Half is put aside, while the other half is again split by means of the chip spreader. This operation is repeated as often as necessary, half being eliminated at each stage with the aim of obtaining 2 “sub-samples” of about 200 g each.
- Operating procedure
- Weigh the empty screen (WES).
- Weigh the empty recovery tray (WET).
- Tare the screen + recovery tray unit and place on it about 200 g of pieces of oak wood weighed to within 0.1 g. Let WOAK be the weight of the pieces of oak wood to be screened.
- Place the unit on the vibrating plate and close the cover with the clamping loops.
- Start up the device and allow it to vibrate for 15 minutes.
- Weigh the screen containing the remaining particles that have not passed through the 2mm meshes (WPS).
- Weigh the recovery tray containing the particles that have passed through the screen (WPT).
A second test is performed in these conditions on the second sub-sample of pieces of oak wood coming from the same initial test sample.
Comment: Weighing of the recovery tray before and after screening (WRT and WPT) serves to verify that there has been no loss of test sample during the operation.
One should have:
- Calculation
The percentage (by weight) of particles retained by the screen of 2mm mesh is given by the following formula:
This calculation is performed for each of the 2 sub-samples coming from the initial test sample; the percentage of particles retained corresponds to the mean of the 2 results.
- Bibliography
- Resolution OENO 3/2005 PIECES OF OAK WOOD
- EN1482-1 - Fertilizers and liming materials. Sampling and sample preparation. Part 1: Sampling.
Wood for wine containers
COEI-1-WOOCON Wood for wine containers
- Subject, origin and scope
The wood of containers used during the making, storage or transport of wines.
The pieces of wood must exclusively originate from species recognized as being suitable to store wine (oak, chestnut)
They can possibly be left in their natural state or they can be heated to a low, medium or high temperature, but they must not be charred, including on the surface, nor be carbonaceous, nor friable when touched.
No compound should be added to them for the purpose of increasing their natural aromatizing capacity or their extractable phenolic compounds.
They must not undergo any chemical, enzymatic or physical treatment other than heating when used for new containers.
If they have undergone chemical or physical treatment, in particular to clean containers having already been used, it is recommended to ensure the perfect harmlessness of any such treatment for materials in contact with foodstuffs, and in particular to ensure that sufficient rinsing has eliminated any trace of certain products that are not authorized in wine.
- Container marking and/or accompanying document
Container markings or the accompanying document must indicate the origin of the botanical species of wood, the intensity of any heating and the safety instructions.
- Purity
Wooden containers must not release substances in concentrations which may be harmful to health.
- Storage
Wooden containers must be washed before first use and then stored under suitable conditions to prevent any development of undesirable micro-organisms when the containers are empty.
Calcium carbonate
COEI-1-CALCAR Calcium carbonate
Calcii carbonas
CaCO3 = 100.1
SIN NO. 170
- Objective, origin and scope of application
This product is used for deacidification. The transport of calcium ions causes salification of free tartaric acid. The use of calcium carbonate is also authorized when using the so-called "double salt" method of deacidification. It may then contain small quantities of calcium tartromalate (double salt) and/or calcium tartrate There are regulations governing the use of this product..
- Labelling
The label should indicate the proportion of pure ccalcium carbonate and the safety and storage requirements.
- Centesimal composition
Carbon dioxide |
43.97 |
Calcium |
40.04 |
- Properties
Calcium carbonate exists as a white powder with the reaction properties of carbonates. In solution in a concentration of 5 pp 100 (m/v) in dilute acetic acid (R), it yields calcium reactions.
- Solubility
Insoluble in water
Insoluble in alcohol at 95% by vol.
Soluble with effervescence in dilute acetic acid, hydrochloric acid and nitric acid solutions
- Tests
6.1. Desiccation loss
Weigh 2 g calcium carbonate in a dish. Place in an oven at 200 °C for 4 hours. Weight loss should not exceed 2 pp 100.
6.2. Substances Soluble in Water
Mix 2 g of ground calcium carbonate with 20 ml of boiled water. Filter. Collect 10 ml. The solution should be neutral. Dry evaporate. The residue should not be greater than 1 pp 100l.
6.3. Ammoniacal Ions
Place 2 g of calcium carbonate, 25 ml of distilled water and 5 ml of 30 pp 100 sodium hydroxide solution (R) in the flask of a distillation device.
Distill and collect 20 ml distillate in 40 ml 4 pp 100 boric acid (R) in the presence of methyl red (R). Two drops of 0.1 M hydrochloric acid solution should be sufficient to cause the indicator to turn color.
6.4. Barium
Dissolve 0.50 g of calcium carbonate in 10 ml of nitric acid diluted to 10 pp 100 (R). Add 10 ml of saturated calcium sulfate solution (R). The mixture should remain llear.
6.5. Preparing the Solution for Tests
Dissolve 10 g of calcium carbonate in 100 ml of 10 pp 100 dilute acetic acid (m/v) (take care as there will be effervescence due to the release of carbon dioxide).
6.6. Magnesium
Use the method described in the Compendium on the solution prepared for testing under paragraph 6.5. (Content should be less than 1 pp 100 by weight).
6.7. Iron
Use the atomic absorption spectrometry method described in the Compendium on the solution prepared under paragraph 6.5. (Iron content should be less than 300 mg/kg).
6.8. Lead
Using the technique described in the annex to quantitatively analyze the lead in the solution prepared for testing (Par. 6.5). (Lead content should be less than 2 mg/kg).
6.9. Mercury
Implement the technique described in the annex to quantitatively analyze the mercury in the solution prepared for testing (Par. 6.5). (Mercury content should be less than 1 mg/kg).
6.10. Arsenic
Using the method described in the annex, test for arsenic in the solution prepared for testing (Par. 6.5). (Arsenic content should be less than 3 mg/kg).
6.11. Sodium
In accordance with the method described in the Compendium, quantitatively determine sodium content by flame photometry in the solution prepared for testing (Par. 6.5). (Sodium content should be less than 500 mg/kg).
6.12. Quantitative Analysis
Dissolve a precisely weighed sample p of about 2 g in 50 ml of a 1 M hydrochloric acid solution. Bring to a boil. Allow to cool and and titrate the excess hydrochloric acid solution using 1 M sodium hydroxide solution and methyl red (R). Let n be the amount in ml of 1 M sodium hydroxide solution used:
1 ml of 1 M hydrochloric acid corresponds to 0.05005 g calcium carbonate. Parts per 100 of calcium carbonate in the product tested:
The wine-making product must contain a minimum of 98 pp 100 calcium carbonate.
- Storage
Calcium carbonate should be stored in a dry place in hermetically sealed containers away from volatile elements it could adsorb.
Potassium carbonate
COEI-1-POTCAR Potassium carbonate
Potassium carbonate anhydrous (K2CO3, CAS No. 584-08-7)
Potassium carbonate hydrate (2K2CO3 · 3H2O, CAS No.: 6381-79-9)
- Objective, origin and scope of application
The addition of potassium carbonate can be used to deacidify musts and wines.
- Labelling
The label should indicate the product's purity, lot code, date of manufacture, storage conditions and expiration date.
- Characteristics
Anhydrous potassium carbonate (K2CO3) is the potassium salt of carbonic acid and occurs as a white, odourless, hygroscopic powder. The hydrate form (2K2CO3 · 3H2O) occurs as small, white, translucent crystals or granules.
- Identifying characteristics
- Solubility: Very soluble in water, insoluble in ethanol (95% by vol).
- Carbonate: Potassium carbonate is soluble with effervescence in dilute acetic acid or hydrochloric acid solutions, evolving a colourless gas (CO2) that, when passed into calcium hydroxide solution, produces a white precipitate immediately.
- Potassium: The presence of potassium imparts a violet colour to a non-luminous flame if not masked by the presence of small quantities of sodium.
- Tests
The limits are determined according to the values observed during production in line with the good manufacturing practices.
5.1. Desiccation Loss
Through the desiccation of 3 g of potassium carbonate for 4 hours at 180°C, for the anhydrous form, the loss of weight must be lower than 1%, for the hydrate form, the loss of weight must be between 10,0% and 16,5%.
5.2. Preparing the Solution for Tests
Dissolve 1 g of potassium carbonate in 20 mL water.
5.3. Substances Insoluble in Water
Filter the solution prepared for testing under Paragraph 5.2. on a membrane of cellulose ester with a diameter of the pore lower or equal to 0,5 μm, no residue can be detected.
5.4. Iron
Using the atomic absorption spectrometry technique detailed in chapter II of the International Oenological Codex, determine the iron content in the test solution (5.2); the content should be less than 10 mg/kg.
5.5. Lead
Using the technique set forth in chapter II of the International Oenological Codex, determine the lead content in the test solution (5.2); the content should be less than 5 mg/kg.
5.6. Mercury
Using the technique described in chapter II of the International Oenological Codex, determine the mercury content in the test solution (5.2); the content should be less than 1 mg/kg.
5.7. Arsenic
Using the technique described in chapter II of the International Oenological Codex, determine the arsenic content in the test solution (5.2); the content should be less than 3 mg/kg.
5.8. Sodium
Determine the sodium content in the test solution (5.2) using flame photometry described in chapter II of the International Oenological Codex; the content should be less than 1%.
5.9. Cadmium
Using the technique described in chapter II of the International Oenological Codex, determine the cadmium content in the test solution (5.2); the content should be less than 1 mg/kg.
5.10. Potassium Carbonate Content
Sample: 1 g previously dried.
Analysis: Transfer sample to a beaker and dissolve it in 50 mL water. Add 2 drops of methyl red TS and, while constantly stirring, slowly titrate with 1 N hydrochloric acid until the solution becomes faintly pink. Heat the solution to boiling, cool, and continue titration until the faint pink colour no longer fades after boiling. The product intended for wine-making should contain a minimum of 98% potassium carbonate.
- Storage
Potassium carbonate should be stored in airtight containers.
Calcium phytate
COEI-1-CALPHY Calcium phytate
Calcium inositol hexaphosphate
Calcii phytas
C6H6Ca6O24P6,3H20= 942.11
- Objective, Origin and Scope of Application
Calcium phytate is the salt of the inositol hexaphosphoric ester, or inositohexaphosphoric or phytic acid.
In its calcium and magnesium double salt forms, phytic acid composes phytin, a reserve form of phosphorous in plants.
Since it is an iron (III) complexing agent approved for removal of excess iron in wines, its use must be strictly monitored.
Any excess phytate with respect to the iron (III) content causes deposits to build up when the slightest oxidation occurs.
- Labelling
The label should indicate product concentration even when used in mixtures, as well as its safety and storage conditions.
- Properties
White powder with an acidulous taste, which is minimally soluble in water, soluble in dilute strong acids, and difficult to dissolve in wine, in which solubility is incomplete.
Aqueous calcium phytate solution possess an acidic nature, which is disclosed by movement of the indicator to litmus. It yields calcium reactions.
- Tests
4.1. Desiccation Loss
Dry a 1 g sample of calcium phytate in an oven at 105 °C until a constant weight is obtained. Weight loss should be less than 12 pp 100.
Limits indicated below are for dry product.
4.2. Ash
Incinerate a 0.250 g test sample of calcium phytate at 550 °C. The residue should not be less than 65 pp 100 nor greater than 72 pp 100 of the dry product contained in the test sample.
4.3. Insoluble Substances
Prepare a first solution contaning 1 g of calcium phytate, 7 ml of 1M hydrochloric acid solution, and 93 ml of distilled water. Separately, prepare a solution of 1 g of calcium phytate with 50 ml of distilled water and 1.5 ml pure phosphoric acid (R). Filter each of the solutions separately and collect the deposit. Wash and dry the deposit at 100 °C. Each residue should be less than 1 part per 100 (10g/kg) of dried product at 105 °C.
4.4. Starch
Add several drops of iodinated water (R) to the residues obtained under Paragraph 4.3; no blue coloration should develop.
4.5. Sugars
- Stir 3 g of calcium phytate with 15 ml of distilled water.
- Filter.
- The filtrate should not reduce the cupro-alkaline reagent (R) before or after the sucrose inversion.
4.6. Albumin
- Dissolve 1 g of the product in a mixture of 1 ml of concentrated hydrochloric acid (R) and 3 ml of distilled water.
- Add 3 ml of 30% sodium hydroxide solution (R).
- Filter.
- When one drop of 4 pp 100 (m/v) copper (II) sulfate solution is added to the filtrate, no violet color should appear.
4.7. Preparing the Solution for Tests
Macerate a quantity of calcium phytate containing 5 g dry product with 100 ml of 10 g per liter citric acid (R) for 24 hours while agitating from time to time. Filter.
4.8. Iron
- Add 1 ml of concentrated hydrochloric acid (R) and 2 ml of 5 pp 100 potassium thiocyanate to 10 ml of test solution prepared under paragraph 4.7.
- The resulting colorating should be less intense than that produced by a control tube prepared with 2.5 ml solution in a concentration of of 0.010 g of iron per liter (R), 7.5 ml of distilled water, 1 ml of concentrated hydrochloric acid (R) and 2 ml of 5 pp 100 thiocyanate (R). (Iron content should be less than 50 mg/kg).
4.9. Lead
Using the method described in the Compendium, quantify lead analytically in the test solution prepared according to Par. 4.7. (Lead content should be less than 5 mg/kg).
4.10. Mercury
Using the method described in the annex, quantify mercury analytically in the test solution prepared according to Par. 4.7. (Mercury content should be less than 1 mg/kg).
4.11. Arsenic
Using the method described in the annex, quantify arsenic analytically in the test solution prepared according to Par. 4.7. (Arsenic content should be less than 3 mg/kg).
4.12. Mineral phosphates
- Place 0.50 g calcium phytate in a 200 ml volumetric flask.
- Add 100 ml of distilled water and 5 ml of concentrated nitric acid (R).
- Agitate for 15 minutes at 20 °C and top off to 200 ml with distilled water.
- To 10 ml of this solution, add 10 ml of nitro-vanadomolybdic reagent (R).
- Leave in contact for 15 minutes at 20 °C. The resulting color should be less intense than that produced by adding 5 ml distilled water and 10 ml nitro-vanadomolybdic reagent (R) to 5 ml of a monopotassic phosphate solution containing 0.05 g phosphorous per liter (R).
- (Mineral phosphate content, expressed in terms of phosphorous, should be less than 1 pp 100).
4.13. Glycerophosphates
- Heat 0.50 g of calcium phytate in the presence of monopotassic sulfate.
- No acrolein fumes (odor of burnt horn) should be released.
4.14. Total Phosphorous Determination
- Weigh precisely a 0.25 g sample of calcium phytate which has already been dried at 105 °C.
- Place it in a flask which is ground and polished so it can be fitted with a tube 8 mm in diameter and 1 m long which will serve as a reflux condenser.
- Add 5 ml of concentrated sulfuric acid (R) and 0.5 ml concentrated nitric acid (R).
- Bring to boiling under reflux for approximately 15 minutes.
- After cooling, decant the contents of the flask diluted with water in a 1 liter volumetric flask.
- Wash the condenser and flask with water by pouring these liquids in the volumetric flask, and fill to gauge line after bringing the temperature to 20 °C. Agitate.
- Add 10 ml of nitro-vanadomolybdic reagent (R) to 10 ml of this solution. Agitate in a 20 °C water bath and let sit in the water bath for 15 minutes. The intensity of the resulting color should be equal to or greater than that of a control prepared under the same conditions using 8 ml of monopotassic phosphate solution in a concentration of 0.05 g of
phosphorous per liter (R), 2 ml of water and 10 ml of nitro-vanadomolybdic reagent (R).
Total phosphorous analysis can also be determined using a spectrophotometer with a wavelength of 425 nm whose calibration curve was obtained based on 4-6-8-10 ml of solution in a concentration of 0.05 mg phosphorous per liter (R).
Calcium phytate should contain at least 15 parts of phosphorous per 100 , as compared with a product dried at 105 °C.
- Storage
Calcium phytate should be stored in a dry place in hermetically sealed containers.
Calcium tartrate
COEI-1-CALTAR Calcium tartrate
Dextrorotatory Calcium Tartrate
Calcium tartaricum
(OOC-CHOH-CHOH-COO) Ca, 4H2O
Tetrahydric L(+)-2,3- calcium dihydoxybutanedioate
(OOC-CHOH-CHOH-COO) Ca, H2O
C4H12CaO10 = 260.13
SIN No. 354
- Objective, Origin and Scope of Application
A natural wine salt primarily originating from wine residues. It is therefore typically found in L(+) form. It usually crystallizes in tetrahydrated form.
This product promotes triggering of the precipitation of the natural calcium tartrate in wine by means of a seeding technique.
- Labelling
The label should indicate product concentration, even when used in mixtures, as well as its safety and storage conditions.
- Centesimal Composition
Tartaric acid |
57.7 |
Calcium |
15.4 |
Water |
27.9 |
- Properties
Fine, crystalline powder with a white or off-white color. Tasteless. Melting point is 270 °C.
- Solubility
Water at 20 °C |
0.525 g/l |
Alcohol, 95% by vol. |
0.15 g/l |
Ethyl ether |
0.01 g/l |
- Tests
6.1. Rotatory Power
Dissolve 1 g of the substance in 1l of 1 M hydrochloric acid. After it has completely dissolved, it gives:
Rotatory power is sensitive to slight variations in pH.
6.2. pH in Saturate Solution
- Add 1 g of the product to 100 ml of distilled water.
- After shaking for one hour and allowing the precipitate to resettle (15 minutes), an increase in pH of between 1.5 and 2.5 pH units should be observed.
6.3. Desiccation Loss
- Desiccation loss is determined up to constant weight in precisely-weighed sample of about 1 g.
- At a temperature of between 100 and 105 °C, weight loss should be less than or equal to 2.5 pp 100.
6.4. Preparing the Solution for Tests
Dissolve a sample precisely weight to about 1 g in 100 ml of 1 M hydrochloric acid.
6.5. Sulfates
- Take 10 ml of the test solution (Par. 6.4) and add to it 1 ml of 10 pp 100 barium chloride solution (R).
- After homogenization, let sit after 15 minutes.
- No clouding should occur.
- If clouding does occur, it should be less intense than that in a control prepared using the method indicated in the Annex. (Sulfate content, expressed in terms of sulfuric acid, should be less than 1 g/kg).
6.6. Heavy Metals
Add 0.5 ml of concentrated ammonium hydroxide (R), 2 ml of pH 3.5 buffer solution (R) and 1.2 ml of thioacetamide reagent (R) to 10 ml of the test solution prepared under paragraph 6.4. (Heavy metal content, expressed in terms of lead, should be less than 10 mg/kg).
6.7. Lead
Using the method described in the Compendium, quantify lead analytically in the test solution prepared according to Par. 6.4. (Lead content should be less than 5 mg/kg).
6.8. Mercury
Using the method described in the Annex, quantify mercury analytically in the test solution prepared according to Par. 6.4. (Mercury content should be less than 1 mg/kg).
6.9. Arsenic
Using the method described in the Annex, quantify arsenic analytically in the test solution prepared according to Par. 6.4.
(Arsenic content should be less than 3 mg/kg).
6.10. Basic Residue Determination
- Dissolve a sample, p, of tetrahydric calcium tartrate weighed precisely at about 0.5 g in 25 ml of 1 M hydrochloric acid solution (R).
- Bring to boiling under reflux and allow to cool.
- Titrate the excess acid using 1 M sodium hydroxide solution (R) and in the presence of methyl red (R). Let n be the quantity in millimeters of the 1 M sodium hydroxide solution used. 1 ml of 1 M hydrochloric acid corresponds to 0.05005 g of calcium carbonate.
- The content in parts per 100 of calcium carbonate is:
The products used in winemaking should contain a maximum of 3 pp 100 basic residues expressed in terms of calcium carbonate.
- Storage
Calcium tartrate should be stored away from moisture in hermetically-sealed containers.
Calcium sulfate
COEI-1-CALSUL Calcium sulfate
Ca x 2 O (Dihydrate)
CAS NUMBER 10101-41-4
- Objective and scope of application
This product is used for must acidification in the production of liqueur wines. Calcium sulfate added reacts with tartrate ions of the must producing insoluble calcium tartrate and releasing ion sulfate in the must. These facts originate modifications in ions equilibria that liberates proton ions and reduces the pH without increasing the titratable acidity.
- Labelling
The label should indicate the nature of calcium sulfate, batch number and the storage and safety requirements.
- Stoichiometric composition
CaSO4 |
79.1 % |
H2O |
20.9 % |
- Properties
Calcium sulfate dihydrate exists as a white amorphous powder. Not to be confused with the anhydrous form which is very hygroscopic and sets in contact with must.
- Solubility
Slightly soluble in water and soluble in hydrochloric, sulphuric and nitric acid solutions.
- Tests
6.1. Desiccation losses
Free water: Weigh 50 g of calcium sulphate in a dish. Place it in an oven at 40 ºC until constant weight. Weight loss should not exceed 2 %.
Free and bonded water: Place another sample in an oven at 200 º C during 4 h. Total weight loss should not exceed 23 %.
6.2. Preparing the Solution for Tests
Weigh 10 g of calcium sulphate. In a 500 ml erlenmeyer flask which can be hermetically sealed, add 200 ml of tartaric acid solution at 5 g/L per litre and bring to pH 3 with HCl 0.1 N. Put this in a magnetic mixer, sprinkle gently the calcium sulphate and mix for 1 hour at a temperature of 20 2°C. Allow to settle and filter by eliminating the first 50 ml of filtrate. Collect at least 100 ml of clear liquid.
6.3. Lead
Using the technique described in the Compendium, analyse quantitatively the lead in the solution prepared for testing (Par. 6.2). Lead content in calcium sulfate should be less than 2 mg/kg.
6.4. Mercury
Using the technique described in the Compendium, analyse quantitatively the mercury in the solution prepared for testing (Par. 6.2). Mercury content in calcium sulfate should be less than 1 mg/kg.
6.5. Arsenic
Using the technique described in the Compendium, analyse quantitatively the arsenic in the solution prepared for testing (Par. 6.2). Arsenic content in calcium sulfate should be less than 3 mg/kg.
6.6. Iron
Using the technique described in the Compendium, analyse quantitatively the iron in the solution prepared for testing (Par. 6.2). Iron content should be less than 200 mg/kg.
6.7. Quantitative Analysis
- Any method of analysis included in the Compendium could be used. In the case of using the gravimetric method OIV-MA-AS321-05A, use the following procedure.
- Weigh 250 milligrams of the sample dried at 40 ºC with a precision of 1 mg and dissolve it in 10 mL HCl 1M.
- Take 5 mL of this solution and add 0,5 mL of HCl 2 M and 1,5 mL of a solution of BaCl2 400 g/L. Stir with a glass stirrer; rinse the stirrer with a little distilled water and leave to stand for 5 min.
- Centrifuge for 5 min at 3.000 rpm, then carefully decant the supernatant liquid.
- Wash the barium sulfate precipitate as follows: add 10 mL hydrochloric acid 2 M, place the precipitate in suspension and centrifuge for 5 min at 3.000 rpm, then carefully decant the supernatant liquid.
- Repeat the washing procedure twice as before using 15 mL distilled water each time.
- Quantitatively transfer the precipitate, with distilled water, into a tared platinum capsule and place over a water bath at 100°C until fully evaporated.
- The dried precipitate is calcined several times briefly over a flame until a white residue is obtained. Leave to cool in a desiccator and weigh:
- Calculations
Content of calcium sulfate dihydrate in the product (%)= |
- Where p is the measured weight of BaSO4 in mg.
If other method of analysis of sulphates included in the Compendium is used to analyse the initial solution of calcium sulfate prepared for quantitative analysis:
Content of calcium sulfate dihydrate in the product (%)= |
- Where c is the concentration of sulfates in mg/L of K2SO4
The wine-making product must contain a minimum of 90 pp 100 calcium sulfate.
- Storage
Calcium sulfate should be stored in a dry place in hermetically sealed containers away from volatile elements it could adsorb.
Caramel
COEI-1-CARAME Caramel
N° SIN : 150
- Object, origin and field of application
Caramel can be found in liquid form or solid form ranging in colour from dark brown to black. Colouring wine in the stricto sensu is not allowed but caramel is used as a colouring agent in certain liquor wines, spirit beverages of vitivinicultural origin and wine-based beverages.
- Definitions
Caramel (or ordinary caramel) (Class I) (SIN: 150a)
- Caramel (or ordinary caramel) is prepared by controlled heating of carbohydrates made up of glucose and fructose monomers and/or their respective polymers (for example, glucose syrup, saccharose and/or inverted sugars syrups). To favour caramelisation, acids, bases and salts excluding ammonium compounds can be used.
Caustic sulphite caramel (Class II) (SIN:150b)
- Caustic sulphite caramel is prepared by controlled heating of carbohydrates as defined for ordinary caramel, with or without acids or bases, in the presence of sulphite compounds (sulphuric acid, potassium sulphite, potassium hydrogen sulphite, sodium sulphite and sodium hydrogen sulphite). No ammonium compounds are used.
Ammonia caramel (Class III) (SIN:150c)
- Ammonia caramel is prepared by controlled heating of carbohydrates as defined for ordinary caramel, with or without acids or bases, in the presence of ammonium compounds (ammonium hydroxide, ammonium carbonate, ammonium hydrogen carbonate, and ammonium phosphate). No sulphite compounds are used.
Ammonium sulphite caramel (Class IV) (SIN: 150d)
- Ammonia sulphite caramel is prepared by controlled heating of carbohydrates as defined for ordinary caramel, with or without acids or bases, in the presence of sulphite and ammonium compounds (sulphuric acid, potassium sulphite, potassium hydrogen sulphite, sodium sulphite, sodium hydrogen sulphite, ammonium hydroxide, ammonium carbonate, ammonium hydrogen carbonate, ammonium phosphate, ammonium sulphate, ammonium sulphite and ammonium hydrogen sulphite.
- Labelling
The concentration of the product and whether it was mixed, must be indicated on the label in addition to the storage conditions.
- Test trials
4.1. Intensity of the colouring
The intensity of the colouring is defined as the absorbance of a liquid solution of 0.1% (m/v) concentrated caramel measured in a 1 cm space of optical pathway with light waves of 610 nm.
4.2. Total Nitrogen
Apply the method described in Chapter II of the International Oenological Codex to 2 g of exactly measured caramel.
4.3. Preparation of the solution for the test trials
- Place 2 g of caramel in a capsule; put in heat chamber at 105°C for 4 hours than incinerate carefully without going beyond 550°C.
- Take the cinders and put in 10 ml of 10% hydrochloric acid (R). Heat a little and transfer to a graduated 50 ml flask and rinse the capsule with water and fill up to the indicator.
4.4. Heavy metals
- Take 10 ml of the solution prepared for the trial tests as in point 4.3, and add 2 ml of 3.5 pH buffer solution (R) and 1.2 ml of thioacetamide reagent (R).
- If the solution turns brown, it must be less brown than the control sample, as indicated in Chapter II of the International Oenological Codex.
4.5. Lead
Using the solution for test trials as prepared in the point 4.3, measure out the lead as indicated in Chapter II of the International Oenological Codex. Please refer to point 5 for maximum contents.
4.6. Mercury
Measure out the mercury using the method described in Chapter II of the International Oenological Codex.
Please refer to point 5 for maximum contents.
4.7. Cadmium
- Test solution prepared according to point 4.3;
- Measure out the cadmium using the method described in Chapter II of the International Oenological Codex.
Please refer to point 5 for maximum contents.
4.8. Arsenic
- Test solution prepared according to point 4.3;
- Measure out the arsenic using the method described in Chapter II of the International Oenological Codex.
Please refer to point 5 for maximum contents.
4.9. Colouring matter retained on DEAE cellulose
See method as described by JECFA published in the Compendium of food additive specifications, FAO Food and Nutrition Paper 52 Add. 8.
4.10. Colouring matter retained on phosphorylcellulose
See method as described by JECFA published in the Compendium of food additive specifications, FAO Food and Nutrition Paper 52 Add. 8.
4.11. 4‑Methylimidazole
See method as described by JECFA published in the Compendium of food additive specifications, FAO Food and Nutrition Paper 52 Add. 8.
4.12. 2‑Acetyl‑4‑tetrahydroxybutylimidazole
See method as described by JECFA published in the Compendium of food additive specifications, FAO Food and Nutrition Paper 52 Add. 8.
4.13. Total sulphur
See method as described by JECFA published in the Compendium of food additive specifications, FAO Food and Nutrition Paper 52 Add. 8.
4.14. Sulphur dioxide
The method used can be found in the O.I.V. Compendium of International Methods of Analysis of Wine and Musts.
- Particular specifications
5.1. Ordinary caramel
Colouring matter retained on DEAE cellulose |
Not more than 50% |
Colouring matter retained on phosphorylcellulose |
Not more than 50% |
Colour intensity |
0.01 – 0.12 |
Total nitrogen |
Not more than 0.1% |
Total sulphur |
Not more than 0.3% |
Arsenic |
Not more than 1 mg/kg |
Lead |
Not more than 2 mg/kg |
Mercury |
Not more than 1 mg/kg |
Cadmium |
Not more than 1 mg/kg |
Heavy metals (expressed in Pb) |
Not more than 25 mg/kg |
5.2. Caustic sulphite caramel
Colouring matter retained on DEAE cellulose |
Not more than 50% |
Colour intensity |
0.06 – 0.10 |
Total Nitrogen |
Not more than 0.2%[1] |
Total sulphur dioxide |
Not more than 0.2% [2] |
Total sulphur |
1.3 – 2.5%[3] |
Sulphur retained on DEAE cellulose |
Over 40% |
Percentage of optical colour density retained on DEAE cellulose |
19-34 |
OD 280/560 ratio |
Over 50 |
Arsenic |
Not more than 1 mg/kg |
Lead |
Not more than 2 mg/kg |
Mercury |
Not more than 1 mg/kg |
Heavy metals (expressed in lead) |
Not more than 25 mg/kg |
5.3. Ammonia caramel
Colouring matter retained on DEAE cellulose |
Not more than 50% |
Colour matter retained on phosphorylcellulose |
Not more than 50% |
Colour intensity |
0.08 – 0.36 |
Ammoniac nitrogen |
Not more than 0.4%[4] |
4‑Methylimidazole |
Not more than 250 mg/kg[5] |
2‑Acetyl‑4‑tetrahydroxybutylimidazole |
Not more than 10 mg/kg[6] |
Total sulphur |
Not more than 0.3%[7] |
Total nitrogen |
1.3 – 6.8%[8] |
Percentage of optical colour density |
13-35 |
Arsenic |
Not more than 1 mg/kg |
Lead |
Not more than 2 mg/kg |
Mercury |
Not more than 1 mg/kg |
Cadmium |
Not more than 1 mg/kg |
Heavy metals (expressed in lead) |
Not more than 25 mg/kg |
5.4. Ammonium sulphite caramel
Colouring matter retained on DEAE cellulose |
Not more than 50% |
Colour intensity |
0.10 – 0.60 |
Ammoniac nitrogen |
Not more than 2.6%[9] |
Sulphur dioxide |
Not more than 0.5%[10] |
4‑Methylimidazole |
Not more than 250 mg/kg[11] |
Total nitrogen |
0.5 – 7.5%[12] |
Total sulphur |
1.4 – 10%[13] |
Nitrogen/sulphur precipitation by alcohol ratio |
0.7 – 2.7 |
OD precipitation by alcohol ratio[14] |
8-14 |
OD 280/560 ratio[15] |
Not more than 50 |
Arsenic |
Not more than 1 mg/kg |
Lead |
Not more than 2 mg/kg |
Mercury |
Not more than 1 mg/kg |
Cadmium |
Not more than 1 mg/kg |
Heavy metals (expressed in lead) |
Not more than 25 mg/kg |
- Storage conditions
Caramel must be stored in a closed container.
- References
- Directive 95/45/CE Journal officiel des Communautés européennes, L 226, 22 September 1995.
- Compendium of food additive specifications, Addendum 8, FAO Food and Nutrition Paper 52 Add.8.
- Joint FAO/WHO Expert Committee on Food Additives (JECFA) ISBN 92-5-104508-9.
[1] Expressed by the intensity of equivalent colouring; or compared to a product with a colour intensity of 0.1 unit of absorption.
[2] Expressed by the intensity of equivalent colouring; or compared to a product with a colour intensity of 0.1 unit of absorption.
[3] Expressed by the intensity of equivalent colouring; or compared to a product with a colour intensity of 0.1 unit of absorption.
[4] Expressed by the intensity of equivalent colouring; or compared to a product with a colour intensity of 0.1 unit of absorption.
[5] Expressed by the intensity of equivalent colouring; or compared to a product with a colour intensity of 0.1 unit of absorption
[6] Expressed by the intensity of equivalent colouring; or compared to a product with a colour intensity of 0.1 unit of absorption
[7] Expressed by the intensity of equivalent colouring; or compared to a product with a colour intensity of 0.1 unit of absorption
[8] Expressed by the intensity of equivalent colouring; or compared to a product with a colour intensity of 0.1 unit of absorption
[9] Expressed by the intensity of equivalent colouring; or compared to a product with a colour intensity of 0.1 unit of absorption.
[10] Expressed by the intensity of equivalent colouring; or compared to a product with a colour intensity of 0.1 unit of absorption.
[11] Expressed by the intensity of equivalent colouring; or compared to a product with a colour intensity of 0.1 unit of absorption.
[12] Expressed by the intensity of equivalent colouring; or compared to a product with a colour intensity of 0.1 unit of absorption.
[13] Expressed by the intensity of equivalent colouring; or compared to a product with a colour intensity of 0.1 unit of absorption.
[14] The optical densities of precipitation by alcohol is defined as the optical density of precipitation at 280 nm divided by the optical density at 560 nm (in a 1 cm space).
[15] The optical densities of precipitation by alcohol is defined as the optical density of precipitation at 280 nm divided by the optical density at 560 nm (in a 1 cm space).
Caseins
COEI-1-CASEIN Caseins (Lactic Casein or Caseina acids)
- Object, origin and field of application
Casein, a heteroprotein containing phosphorous, is found in milk in the state of calcium salt.
It is obtained by coagulating skim milk.
It is the fining agent indicated for the treatment of oxidations in wine. It can only be used in alkaline water with potassium carbonate or potassium hydrogenocarbonate.
Casein adsorbs polyphenols, in particular oxidised polyphenols.
- Labelling
The concentration of casein used for the preparation must be indicated on the label including in the case of a mixture, as well as the storage conditions.
- Characteristics
Casein is a yellowish white coloured powder. It is amorphous, odourless and insoluble in pure water and various organic solvents. It can have a slight lactic odour. In alkaline water or in saline solutions with alkaline reactions, it swells and produces a colloidal solution: 100 ml of alkaline water for 1 g of potassium hydroxide or sodium hydroxide, dissolve 10 g of casein in a water bath at 100°C. The solution diluted 20 times its volume in water is cloudy; it should be free of lumps.
The so-called soluble caseins are mixed with pure powder and/or potassium carbonate (maximum 20%), or potassium hydrogenocarbonate).
Caseins used in oenology are fit for human consumption.
- Identifying characteristics
4.1. Casein doesn’t precipitate by heating its alkaline solution. This solution precipitates by acidification once the pH is less than 5.
4.2. Casein ashes contain phosphates characterised by the nitromolybdic reagent (R).
- Test trials
Casein should have no flavour, nor abnormal odour (rotten, mouldy, putrid, etc.)
5.1. Acidity
5.1.1. Principle
Determining free acidity in casein by an acidobasic determination of an aqueous extract of the product.
5.1.2. Reagents
- Sodium hydroxide 0.1 M
- Phenolphthalein, solution at 10 g/l in ethanol
5.1.3. Procedure
Preliminary test:
- Homogenise the product by shaking vigorously;
- Put 50 g of the product on a strainer (metal mesh strainer 200 mm in diameter, nominal size of 500 µm for the opening with a receptacle (Standard ISO 3310/1);
- If 50 g of the product passes through completely, use the product as it is;
- If the 50 g of the product do not pass through, grind the product until 50 g do pass through.
During all these operations, avoid changing the water content of the product.
Preparation for the test trial solution:
- Take approximately 10 g to the nearest 10 mg of the 50 g passed through the strainer, or m of this mass.
- Put the mass m in a 250 ml conical flask.
- Pour 200 ml of recently boiled distilled water brought to 60°C into the flask.
- Shake the closed flask.
- Allow to stand for 30 minutes in a water bath at 60 °C while shaking the flask every 10 minutes.
- Filter.
- The filtrate at 20°C must be clear.
Carrying out the test:
- Take 100 ml of filtrate.
- Place the test sample in a 250 ml conical flask.
- Add 0.5 ml of phenolphthalein solution to the flask.
- Titrate using 0.1 M sodium hydroxide solution.
- Let V represent the volume used.
5.1.4. Calculation
Free acidity in casein expressed in meq/l is equal to:
- V is the volume in ml of sodium hydroxide used.
- T is the exact mole fraction of the sodium hydroxide solution.
- m is the mass density in g of the test trial sample.
Acidity expressed as lactic acid should be less than 1.6 g/l.
5.2. pH
- Shake 10 g of casein in 100 ml of water for a few minutes.
- Decant; the pH of the solution should be less than or equal to 5 for pure casein.
5.3. Loss by dessication
- Determine the weight loss of 2 g of the test trial sample by drying to constant weight at 100°C-105°C.
- Weight loss of casein must be less than 12%.
All the limits set below apply to dried products.
5.4. Ashes
- Incinerate the residue left in the weight loss determination by dessication, without exceeding 600 °C.
- The rate of the ashes should be less than 3% for casein acid and less than 23% for the casein acid and potassium carbonate or potassium hydrogenocarbonate mixture.
5.5. Preparation of test trial solution
- After determining the weight of the ashes, dissolve them in 2 ml of concentrated hydrochloric acid (R) and 10 ml of water.
- Heat to dissolve and add water until reaching a volume equal to 25 times the weight of dried casein. 1 ml of this solution contains 0.04 g of dried casein mineral matters.
5.6. Iron
Take 10 ml of the test trial solution (5.5), and add 1 ml of concentrated hydrochloric acid (R), 3 drops of hydrogen peroxide solution at 3 volumes(R) and 2 ml of potassium thiocyanate solution at 5% (R).
If a red colouration appears, it must be lighter than the control prepared with 8 ml of iron solution (III) at 0.01 g of iron per litre (R), 2 ml of water and the same volumes of concentrated hydrochloric acid (R) and potassium thiocyanate solution at 5% (R).
Iron content should be less than 200 mg/kg.
This determination can also be carried out by atomic absorption spectrophotometry.
5.7. Lead
On the test trial solution (5.5), determine the lead according to the method described in Chapter II of the International Oenological Codex.
Lead content should be less than 5 mg/kg.
5.8. Cadmium
On the test trial solution (5.5), determine the cadmium according to the method described in Chapter II of the International Oenological Codex.
Cadmium content should be less than 1 mg/kg.
5.9. Mercury
Determine the mercury according to the method described in Chapter II of the International Oenological Codex.
Mercury content should be less than 1 mg/kg.
5.10. Arsenic
On the test trial solution (5.5), determine the arsenic according to the method described in Chapter II of the International Oenological Codex.
Arsenic content should be less than 3 mg/kg.
5.11. Total nitrogen
Introduce approximately 0.20 g of casein precisely weighed in a mineralisation flask with 15 ml of concentrated sulphuric acid (R) and 2 g of mineralisation catalyst (R) and continue the operation according to the method in chapter II of the International Oenological Codex.
Total nitrogen content must be more than 13%.
5.12. Proteins
Protein content should not be less than 82% of weight (total nitrogen 6.38).
5.13. Fat content
Determine the fat content using the gravimetric Schmid-Bondzynski-Ratslaff method (standard ISO 5543).
Fat content should be less than 2%.
5.14. Bacteriological monitoring
Proceed as indicated in chapter II of the International Oenological Codex.
Limit: total viable microorganisms: less than 3 x 104 CFU/g.
5.15. Coliforms
Proceed with counting according to the method in chapter II of the International Oenological Codex.
Absence must be checked on a sample of 25 g.
5.16. Staphylococci
Proceed with counting according to the method in chapter II of the International Oenological Codex.
The number of staphylococci (ß-hemolytiques positive coagulase) must be less than or equal to 1 per g.
5.17. Escherichia Coli
Proceed with counting according to the method in chapter II of the International Oenological Codex.
Absence must be checked on a sample of 1 g.
5.18. Salmonella
Proceed with counting according to the method in chapter II of the International Oenological Codex.
The number of salmonella should be less than 1 per 100 g.
5.19. Yeasts
Proceed with counting according to the method in chapter II of the International Oenological Codex.
Content limit: 103 CFU/g of preparation.
5.20. Lactic bacteria
Proceed with counting according to the method in chapter II of the International Oenological Codex.
Content limit: 102 CFU/g of preparation.
5.21. Lactobacillus sp.
Content limit: 10 CFU/g of preparation.
5.22. Pediococcus sp.[2]
Content limit: absence in a 10 g preparation sample.
5.23. Acetic bacteria
Proceed with counting according to the method in chapter II of the International Oenological Codex.
Content limit: 103 CFU/g of preparation
5.24. Mould
Proceed with counting according to the method in chapter II of the International Oenological Codex.
Content limit: 103 CFU/g of preparation
- Storage
Casein must be stored in watertight bags between 5°C and 20°C with relative humidity less than 65%. Its shelf life is 24 months.
- References
- Standard ISO 5543.
Method to be defined later on
[2] Method to be defined later on
Cellulose
COEI-1-CELLUL Cellulose
(
INS N°: 460
- Object, origin and field of application
Cellulose is obtained from mechanical processing and purification from an alpha-cellulose, which comes directly from vegetable fibres. Its molecular weight is 1.5. Dalton. Cellulose fibre is used for its absorbency traits, mainly for the filtration of wine.
- Labelling
The concentration of the product and whether it was mixed, must be indicated on the label in addition to the change.
- Characteristics
Cellulose is a white odourless, flavourless, fibre. It is insoluble in water.
- Test trials
4.1. pH
- Mix 5g of cellulose in 40 ml of water free of carbon dioxide, for 20 minutes.
- Centrifuge.
- The pH of the supernatant will be between 5.0 and 7.5.
4.2. Humidity and volatile matter
- Put 5 g of cellulose in an incubator at 105°C for 3 hours.
- Mass loss must not exceed 8%.
All of the maximum limits set below refer to the dried product.
4.3. Starch
- Add 90 ml of water (R) to 10 g of microcrystalline cellulose and boil for 5 minutes.
- Filter when hot. Cool and add 0.1 ml of 0.05 Miodine to the filtrate.
- A blue colour should not appear.
4.4. Ashes
- Incinerate at 600 25°C the residue obtained according to point 4.2, for 4 hours.
- The weight of the ashes should not exceed 2%.
4.5. Preparation of the test solution
- After weighing, dissolve ashes in 2 ml of concentrated hydrochloric acid (R) and 10 ml of water (R).
- Heat in order to dissolve and fill the water up to 50 ml. (R).
4.6. Iron
Determine iron using an atomic absorption spectrophotometer (following the method described in Chapter II on the test solution (4.5).
Iron content must be less than 100 mg/kg.
4.7. Lead
Measure out lead following the method described in Chapter II on the test solution (4.5). Lead content must be less than 5 mg/kg.
4.8. Mercury
Measure out mercury following the method described in Chapter II on the test solution (4.5).
Mercury content must be less than 1 mg/kg.
4.9. Cadmium
Measure out cadmium as described in Chapter II on the test solution (4.5).
Cadmium content must be less than 1 mg/kg.
4.10. Arsenic
Measure out arsenic following the method described in Chapter II on the test solution (4.5).
Arsenic content must be less than 2 mg/kg.
4.11. Calcium
Determine calcium using an atomic absorption spectrophotometer (see method described in Chapter II on the test solution (4.5). Calcium content must be less than 500 mg/kg.
4.12. Water soluble substances
Evaporate the aliquot part of the supernatant obtained when measuring the pH level at point 4.1, in an incubator at 105°C for 3 hours. The soluble substance content should not exceed 0.25%.
- Storing conditions
Cellulose should be kept in a well-ventilated place in sealed packages away from volatile substances susceptible of being adsorbed.
Cellulose powdered
POWDERED CELLULOSE
(C12H20O10)n
INS No.: 460 (ii)
CAS No.: 9004-34-6
OIV-OENO 681-2022
1. Object, origin and scope of application
Powdered cellulose is cellulose of food-grade quality. Cellulose is a linear glucose homopolymer composed of glucopyranose units linked by β-1,4-glycosidic bonds; its degree of polymerisation (DP) is dependent on the origin of the cellulosic material.
Powdered cellulose is a non-modified, purified cellulose, obtained by the mechanical disintegration of alpha-cellulose procured in pulp form from fibrous plant material.
Powdered cellulose plays a “support” role in clarified fermentation media; it allows for improved “degassing” of carbon dioxide at the start of alcoholic fermentation and thus shortens the latency phase. It increases the fermentability of must.
2. Labelling
The following should be mentioned on the label:
- the identification of the cellulose and its use in food,
- the concentration of the product, including in the case of a mixture,
- safety and storage conditions
- the batch no.,
- the expiry date.
3. Characteristics
Powdered cellulose comes in the form of flakes or very fine fibres, and is whitish in colour, odourless and flavourless.
4. Test limits and methods
4.1 Identification
Place approx. 10 mg powdered cellulose on a watch glass and disperse into 2 mL iodinated zinc chloride solution (R). The solution will turn a blue-purple colour.
4.2 Solubility
Powdered cellulose is insoluble in water, ethanol, ether and diluted acids, and slightly soluble in sodium hydroxide solution.
4.3 Purity
The powdered cellulose content should not be less than 92%.
4.4 Size of fine particles
The size of fine particles should not be less than 5 µm; the number of particles smaller than 5µm should not exceed 10%.
4.5 pH
Mix approx. 10 g dry cellulose in 90 mL water free of carbon dioxide for 60 min. Centrifuge. The pH of the supernatant liquid should be between 5.0 and 7.5.
4.6 Water-soluble substances
Mix around 6 g of sample, dried beforehand, with 90 mL water that has recently been boiled then cooled. Leave to rest for 10 min. Filter using a membrane with porosity of 3 µm, throw away the first 10 mL of filtrate and pass the filtrate through the same filter a second time if necessary to obtain a clear filtrate. Evaporate a 15-mL portion of filtrate to dryness in a tared evaporating dish over a water bath, and dry to 105 °C for 1 h. Weigh the dish containing the dry residue. Less than 15 mg of residue should be obtained.
4.7 Detection of starch
Add 90 mL demineralised water (R) to 10 g powdered cellulose and boil for 5 min. Filter while hot with a 25-µm membrane filter. Cool and add 0.1 mL 0.05 M iodine solution to the filtrate. No blue colouration should appear. With very fine grains, a light blue colour may be observed after the addition of the iodine solution but will disappear after 30 minutes.
4.8 Loss on drying
Place 1 g powdered cellulose in a tared dish in an oven at 100-105 °C for 3 hours. The loss on drying should not exceed 7.0%.
4.9 Ashes
Incinerate the residue obtained in point 4.8 at 800 ± 25 °C for 4 hours. The weight of the ashes should not be greater than 0.3%.
All of the limits set below relate to dry products.
5. Preparation of the test solution
After weighing, dissolve the ashes in 2 mL concentrated hydrochloric acid (R) and 10 mL water (R). Heat to activate dissolution and make up to 50 mL with water.
5.1 Iron
Determine the iron content of the solution prepared for testing purposes (5) by atomic absorption spectrophotometry according to the method described in Chapter II. The iron content should be less than or equal to 10 mg/kg.
5.2 Lead
Determine the lead content of the solution prepared for testing purposes (5) according to the method described in Chapter II. The lead content should be less than 2 mg/kg.
5.3 Mercury
Determine the mercury content of the solution prepared for testing purposes (5) according to the method described in Chapter II. The mercury content should be less than 1 mg/kg.
5.4 Cadmium
Determine the cadmium content of the solution prepared for testing purposes (5) according to the method described in Chapter II. The cadmium content should be less than 1 mg/kg.
5.5 Arsenic
Determine the arsenic content of the solution prepared for testing purposes (5) according to the method described in Chapter II. The arsenic content should be less than 1 mg/kg.
6. Storage
Powdered cellulose should be stored in a well-ventilated place in airtight packaging, away from any volatile substances that it could adsorb.
Mycrocristalline cellulose
COEI-1-CELMIC Microcristlaline cellulose
INS N°: 460
- Object, origin and field of application
Microcrystalline cellulose is purified cellulose and is partially depolymerised. It comes from the treatment of alpha-cellulose mineral acids from plant fibres. Its molecular weight is approximately 36 000
Microcrystalline cellulose plays an important role in “supporting” very clarified fermentation as it increases the fermentability of the juices.
- Labelling
The concentration of the product must be mentioned on the label and if there is mixing as well as the method of preservation.
- Characteristics
Cellulose is found in microcrystalline powder form. white. odourless and tasteless. It is almost insoluble in water. acetone. ethanol. toluene. diluted acids and in 50 g/l sodium hydroxide solutions.
- Identification
4.1. In a watch glass. put approximately 10 mg of microcrystalline cellulose and add 2 ml of zinc chloride iodated solution (R). The solution turns bluish purple.
4.2. Degree of polymerisation
- Put 1.300 g of microcrystalline cellulose in a 125 ml conical flask. Add 25 ml of water (R) and 25 ml of 1M cupriethylenediamine hydroxide.
- Immediately pass a nitrogen current. Close the flask and mix until completely dissolved.
- For 7 ml of the solution into an appropriate glass capillary viscosimetric tube.
Time how long it takes between two lines on the viscosimeter and express the time measured in (t1). Calculate the kinematic viscosity V1 of the solution using the following formula:
In which k1 is the viscosimeter constant.
Take out an appropriate volume of 1M cupriethylenediamine hydroxide and dilute with the same volume of water. (R). Using an appropriate capillary viscosimeter. determine the time flow of this solution.
Calculate the kinematics viscosity V2 of the solvent using the following formula.
In which k2 is the viscosimeter constant.
Determine the relative viscosity ηrel of the microcrystalline cellulose sample. using the following formula:
Determine the intrinsic viscosity []c by extrapolation. using the intrinsic viscosity table in Annex.
Calculate the degree of polymerisation P. using the formula:
In which m is the mass. in grams of the trial and b is the value obtained in the test trial " loss through drying " in %.
The degree of polymerisation is not over 350.
4.3. pH
- Shake for 20 minutes about 5 g of cellulose in 40 ml of water free of carbon dioxide.
- Centrifuge.
- The pH of the supernatant liquid must be between 5.0 and 7.5.
4.4. Soluble substances in ether
- Prepare a column of 10.0 g of microcrystalline cellulose in a glass tube with an inside diameter of approximately 20 mm.
- Put 50 ml of ether free of peroxides (R). through the column and evaporate the eluate until bone dry.
- The residue should not be above 5.0 mg (0.05%).
4.5. Soluble substances in water
- Mix 5.0 g of microcrystalline cellulose with 80 ml of water (R) for 10 mn.
- Filter in a vacuum and collect the filtrate in a weighed vase.
- Evaporate over a bath of 100° C water until bone dry and dry at 100-105°C for 1 hour.
- The residue is not above 12.5 mg (0.25%).
4.6. Starch
- Add 90 ml of water (R) to 10 g microcrystalline cellulose. and boil for 5 mn.
- Filter when hot.
- Let cool and add 0.1 ml iodine 0.05 M to filtrate.
- There is no blue colouring.
4.7. Loss through drying
- Put 1 g of cellulose in a mass capsule for 3 hours in an incubator at 100-105°C.
- Loss through drying should not be more than 6.0%.
All limits set below refer to the dried product.
4.8. Ashes
- Incinerate at 600 ± 25°C the residue obtained in point 4.7. for 4 hours.
- The mass of the ashes should not be more than 0.1%.
4.9. Preparation of test solution
- After weighing. dissolve the ashes in 2 ml of concentrated hydrochloric acid (R) and 10 ml of water (R).
- Heat to activate the dissolution and fill up to 50 ml with water.
4.10. Iron
Determine iron with an atomic absorption spectrophotometer following the method described in Chapter II into the test solution (4.9).
Iron content must be less than less or equal to 10 mg/kg.
4.11. Lead
Determine the lead according to the method described in Chapter II. into the test solution (4.9).
Lead content must be less than 5 mg/kg.
4.12. Mercury
Determine the mercury according to the method described in Chapter II
Mercury content must be less than 1 mg/kg.
4.13. Cadmium
Determine the cadmium according to the method described in Chapter II. into the test solution (4.9).
Cadmium content must be less than 1 mg/kg.
4.14. Arsenic
Determine the arsenic according to the method described in Chapter II.
Arsenic content must be less than 1 mg/kg.
4.15. Calcium
Determine the calcium with an atomic absorption spectrophotometer. following the method described in Chapter II. into the test solution (4.9).
Calcium content must be less than 500 mg/kg.
5. Storing conditions
Cellulose must be stored in a well-ventilated place in sealed packages away from volatile substances which it might adsorb.
Table of instrinsic viscosity |
||||||||||
Intrinsic viscosity. [η]c. according to value of relative viscosity.. ηrel |
||||||||||
[η]c |
||||||||||
rel |
0.00 |
0.01 |
0.02 |
0.03 |
0.04 |
0.05 |
0.06 |
0.07 |
0.08 |
0.09 |
1.1 |
0.098 |
0.106 |
0.115 |
0.125 |
0.134 |
0.143 |
0.152 |
0.161 |
0.170 |
0.180 |
1.2 |
0.189 |
0.198 |
0.207 |
0.216 |
0.225 |
0.233 |
0.242 |
0.250 |
0.259 |
0.268 |
1.3 |
0.276 |
0.285 |
0.293 |
0.302 |
0.310 |
0.318 |
0.326 |
0.334 |
0.342 |
0.350 |
1.4 |
0.358 |
0.367 |
0.375 |
0.383 |
0.391 |
0.399 |
0.407 |
0.414 |
0.422 |
0.430 |
1.5 |
0.437 |
0.445 |
0.453 |
0.460 |
0.468 |
0.476 |
0.484 |
0.491 |
0.499 |
0.507 |
1.6 |
0.515 |
0.522 |
0.529 |
0.536 |
0.544 |
0.551 |
0.558 |
0.566 |
0.573 |
0.580 |
1.7 |
0.587 |
0.595 |
0.602 |
0.608 |
0.615 |
0.622 |
0.629 |
0.636 |
0.642 |
0.649 |
1.8 |
0.656 |
0.663 |
0.670 |
0.677 |
0.683 |
0.690 |
0.697 |
0.704 |
0.710 |
0.717 |
1.9 |
0.723 |
0.730 |
0.736 |
0.743 |
0.749 |
0.756 |
0.762 |
0.769 |
0.775 |
0.782 |
2.0 |
0.788 |
0.795 |
0.802 |
0.809 |
0.815 |
0.821 |
0.827 |
0.833 |
0.840 |
0.846 |
2.1 |
0.852 |
0.858 |
0.864 |
0.870 |
0.876 |
0.882 |
0.888 |
0.894 |
0.900 |
0.906 |
2.2 |
0.912 |
0.918 |
0.924 |
0.929 |
0.935 |
0.94 1 |
0.948 |
0.953 |
0.959 |
0.965 |
2.3 |
0.971 |
0.976 |
0.983 |
0.988 |
0.994 |
1.000 |
1.006 |
1.011 |
1.017 |
1.022 |
2.4 |
1.028 |
1.033 |
1.039 |
1.044 |
1.050 |
1.056 |
1.061 |
1.067 |
1.072 |
1.078 |
2.5 |
1.083 |
1.089 |
1.094 |
1.100 |
1.105 |
1.111 |
1.116 |
1.121 |
1.126 |
1.131 |
2.6 |
1.137 |
1.142 |
1.147 |
1.153 |
1.158 |
1.163 |
1.169 |
1.174 |
1.179 |
1.184 |
2.7 |
1.190 |
1.195 |
1.200 |
1.205 |
1.210 |
1.215 |
1.220 |
1.225 |
1.230 |
1.235 |
2.8 |
1.240 |
1.245 |
1.250 |
1.255 |
1.260 |
1.265 |
1.270 |
1.275 |
1.280 |
1.285 |
2.9 |
1.290 |
1.295 |
1.300 |
1.305 |
1.310 |
1.314 |
1.319 |
1.324 |
1.329 |
1.333 |
3.0 |
1.338 |
1.343 |
1.348 |
1.352 |
1.357 |
1.362 |
1.367 |
1.371 |
1.376 |
1.381 |
3.1 |
1.386 |
1.390 |
1.395 |
1.400 |
1.405 |
1.409 |
1.414 |
1.418 |
1.423 |
1.427 |
3.2 |
1.432 |
1.436 |
1.441 |
1.446 |
1.450 |
1.455 |
1.459 |
1.464 |
1.468 |
1.473 |
3.3 |
1.477 |
1.482 |
1.486 |
1.491 |
1.496 |
1.500 |
1.504 |
1.508 |
1.513 |
1.517 |
3.4 |
1.521 |
1.525 |
1.529 |
1.533 |
1.537 |
1.542 |
1.546 |
1.550 |
1.554 |
1.558 |
3.5 |
1.562 |
1.566 |
1.570 |
1.575 |
1.579 |
1.583 |
1.587 |
1.591 |
1.595 |
1.600 |
3.6 |
1;604 |
1.608 |
1.612 |
1.617 |
1.621 |
1.625 |
1.629 |
1.633 |
1.637 |
1.642 |
3.7 |
1.646 |
1.650 |
1.654 |
1.658 |
1.662 |
1.666 |
1.671 |
1.675 |
1.679 |
1.683 |
3.8 |
1.687 |
1.691 |
1.695 |
1.700 |
1.704 |
1.708 |
1.712 |
1.715 |
1.719 |
1.723 |
3.9 |
1.727 |
1.731 |
1.735 |
1.739 |
1.742 |
1.746 |
1.750 |
1.754 |
1.758 |
1.762 |
4.0 |
1.765 |
1.769 |
1.773 |
1.777 |
1.781 |
1.785 |
1.789 |
1.792 |
1.796 |
1.800 |
4.1 |
1.804 |
1.808 |
1.811 |
1.815 |
1.819 |
1.822 |
1.826 |
1.830 |
1.833 |
1.837 |
4.2 |
1.841 |
1.845 |
1.848 |
1.852 |
1.856 |
1.859 |
1.863 |
1.867 |
1.870 |
1.874 |
4.3 |
1.878 |
1.882 |
1.885 |
1.889 |
1.893 |
1.896 |
1.900 |
1.904 |
1.907 |
1.911 |
4.4 |
1.914 |
1.918 |
1.921 |
1.925 |
1.929 |
1.932 |
1.936 |
1.939 |
1.943 |
1.946 |
4.5 |
1.950 |
1.954 |
1.957 |
1.961 |
1.964 |
1.968 |
1.971 |
1.975 |
1.979 |
1.982 |
4.6 |
1.986 |
1.989 |
1.993 |
1.996 |
2.000 |
2.003 |
2.007 |
2.010 |
2.013 |
2.017 |
4.7 |
2.020 |
2.023 |
2.027 |
2.030 |
2.033 |
2.037 |
2.040 |
2.043 |
2.047 |
2.050 |
4.8 |
2.053 |
2.057 |
2.060 |
2.063 |
2.067 |
2.070 |
2.073 |
2.077 |
2.080 |
2.083 |
4.9 |
2.087 |
2.090 |
2.093 |
2.097 |
2.100 |
2.103 |
2.107 |
2.110 |
2.113 |
2.116 |
5.0 |
2.119 |
2.122 |
2.125 |
2.129 |
2.132 |
2.135 |
2.139 |
2.142 |
2.145 |
2.148 |
5.1 |
2.151 |
2.154 |
2.158 |
2.160 |
2.164 |
2.167 |
2.170 |
2.173 |
2.176 |
2.180 |
5.2 |
2.183 |
2.186 |
2.190 |
2.192 |
2.195 |
2.197 |
2.200 |
2.203 |
2.206 |
2.209 |
5.3 |
2.212 |
2.215 |
2.218 |
2.221 |
2.224 |
2.227 |
2.230 |
2.233 |
2.236 |
2.240 |
5.4 |
2.243 |
2.246 |
2.249 |
2.252 |
2.255 |
2.258 |
2.261 |
2.264 |
2.267 |
2.270 |
5.5 |
2.273 |
2.276 |
2.279 |
2.282 |
2.285 |
2.288 |
2.291 |
2.294 |
2.297 |
2.300 |
5.6 |
2.303 |
2.306 |
2.309 |
2.312 |
2.315 |
2.318 |
2.320 |
2.324 |
2.326 |
2.329 |
5.7 |
2.332 |
2.335 |
2.338 |
2.341 |
2.344 |
2.347 |
2.350 |
2.353 |
2.355 |
2.358 |
5.8 |
2.361 |
2.364 |
2.367 |
2.370 |
2.373 |
2.376 |
2.379 |
2.382 |
2.384 |
2.387 |
5.9 |
2.390 |
2.393 |
2.396 |
2.400 |
2.403 |
2.405 |
2.408 |
2.411 |
2.414 |
2.417 |
6.0 |
2.419 |
2.422 |
2.425 |
2.428 |
2.431 |
2.433 |
2.436 |
2.439 |
2.442 |
2.444 |
6.1 |
2.447 |
2.450 |
2.453 |
2.456 |
2.458 |
2.461 |
2.464 |
2.467 |
2.470 |
2.472 |
6.2 |
2.475 |
2.478 |
2.481 |
2.483 |
2.486 |
2.489 |
2.492 |
2.494 |
2.497 |
2.500 |
6.3 |
2.503 |
2.505 |
2.508 |
2.511 |
2.513 |
2.516 |
2.518 |
2.52 1 |
2.524 |
2.526 |
6.4 |
2.529 |
2.532 |
2.534 |
2.537 |
2.540 |
2.542 |
2.545 |
2.547 |
2.550 |
2.553 |
6.5 |
2.555 |
2.558 |
2.561 |
2.563 |
2.566 |
2.568 |
2.571 |
2.574 |
2.576 |
2.579 |
6.6 |
2.581 |
2.584 |
2.587 |
2.590 |
2.592 |
2.595 |
2.597 |
2.600 |
2.603 |
2.605 |
6.7 |
2.608 |
2.610 |
2.613 |
2.615 |
2.618 |
2.620 |
2.623 |
2.625 |
2.627 |
2.630 |
6.8 |
2.633 |
2.635 |
2.637 |
2.640 |
2.643 |
2.645 |
2.648 |
2.650 |
2.653 |
2.655 |
6.9 |
2.658 |
2.660 |
2.663 |
2.665 |
2.668 |
2.670 |
2.673 |
2.675 |
2.678 |
2.680 |
7.0 |
2.683 |
2.685 |
2.687 |
2.690 |
2.693 |
2.695 |
2.698 |
2.700 |
2.702 |
2.705 |
7.1 |
2.707 |
2.710 |
2.712 |
2.714 |
2.717 |
2.719 |
2.721 |
2.724 |
2.726 |
2.729 |
7.2 |
2.731 |
2.733 |
2.736 |
2.738 |
2.740 |
2.743 |
2.745 |
2.748 |
2.750 |
2.752 |
7.3 |
2.755 |
2.757 |
2.760 |
2.762 |
2.764 |
2.767 |
2.769 |
2.771 |
2.774 |
2.776 |
7.4 |
2.779 |
2.781 |
2.783 |
2.786 |
2.788 |
2.790 |
2.793 |
2.795 |
2.798 |
2.800 |
7.5 |
2.802 |
2.805 |
2.807 |
2.809 |
2.812 |
2.814 |
2.816 |
2.819 |
2.821 |
2.823 |
7.6 |
2.826 |
2.828 |
2.830 |
2.833 |
2.835 |
2.837 |
2.840 |
2.842 |
2.844 |
2.847 |
7.7 |
2.849 |
2.851 |
2.854 |
2.856 |
2.858 |
2.860 |
2.863 |
2.865 |
2.868 |
2.870 |
7.8 |
2.873 |
2.875 |
2.877 |
2.879 |
2.881 |
2.884 |
2.887 |
2.889 |
2.891 |
2.893 |
7.9 |
2.895 |
2.898 |
2.900 |
2.902 |
2.905 |
2.907 |
2.909 |
2.911 |
2.913 |
2.915 |
8.0 |
2.918 |
2.920 |
2.922 |
2.924 |
2.926 |
2.928 |
2.931 |
2.933 |
2.935 |
2.937 |
8.1 |
2.939 |
2.942 |
2.944 |
2.946 |
2.948 |
2.950 |
2.952 |
2.955 |
2.957 |
2.959 |
8.2 |
2.961 |
2.963 |
2.966 |
2.968 |
2.970 |
2.972 |
2.974 |
2.976 |
2.979 |
2.981 |
8.3 |
2.983 |
2.985 |
2.987 |
2.990 |
2.992 |
2.994 |
2.996 |
2.998 |
3.000 |
3.002 |
8.4 |
3.004 |
3.006 |
3.008 |
3.010 |
3.012 |
3.015 |
3.017 |
3.019 |
3.021 |
3.023 |
8.5 |
3.025 |
3.027 |
3.029 |
3.031 |
3.033 |
3.035 |
3.037 |
3.040 |
3.042 |
3.044 |
8.6 |
3.046 |
3.048 |
3.050 |
3.052 |
3.054 |
3.056 |
3.058 |
3.060 |
3.062 |
3.064 |
8.7 |
3.067 |
3.069 |
3.071 |
3.073 |
3.075 |
3.077 |
3.079 |
3.081 |
3.083 |
3.085 |
8.8 |
3.087 |
3.089 |
3.092 |
3.094 |
3.096 |
3.098 |
3.100 |
3.102 |
3.104 |
3.106 |
8.9 |
3.108 |
3.110 |
3.112 |
3.114 |
3.116 |
3.118 |
3.120 |
3.122 |
3.124 |
3.126 |
9.0 |
3.128 |
3.130 |
3.132 |
3.134 |
3.136 |
3.138 |
3.140 |
3.142 |
3.144 |
3.146 |
9.1 |
3.148 |
3.150 |
3.152 |
3.154 |
3.156 |
3.158 |
3.160 |
3.162 |
3.164 |
3.166 |
9.2 |
3.168 |
3.170 |
3.172 |
3.174 |
3.176 |
3.178 |
3.180 |
3.182 |
3.184 |
3.186 |
9.3 |
3.188 |
3.190 |
3.192 |
3.194 |
3.196 |
3.198 |
3.200 |
3.202 |
3.204 |
3.206 |
9.4 |
3.208 |
3.210 |
3.212 |
3.214 |
3.215 |
3.217 |
3.219 |
3.221 |
3.223 |
3.225 |
9.5 |
3.227 |
3.229 |
3.231 |
3.233 |
3.235 |
3.237 |
3.239 |
3.241 |
3.242 |
3.244 |
9.6 |
3.246 |
3.248 |
3.250 |
3.252 |
3.254 |
3.256 |
3.258 |
3.260 |
3.262 |
3.264 |
9.7 |
3.266 |
3.268 |
3.269 |
3.271 |
3.273 |
3.275 |
3.277 |
3.279 |
3.281 |
3.283 |
9.8 |
3.285 |
3.287 |
3.289 |
3.291 |
3.293 |
3.295 |
3.297 |
3.298 |
3.300 |
3.302 |
9.9 |
3.304 |
3.305 |
3.307 |
3.309 |
3.311 |
3.313 |
3.316 |
3.318 |
3.320 |
3.321 |
10 |
3.32 |
3.34 |
3.36 |
3.37 |
3.39 |
3.41 |
3.43 |
3.45 |
3.46 |
3.48 |
11 |
3.50 |
3.52 |
3.53 |
3.55 |
3.56 |
3.58 |
3.60 |
3.61 |
3.63 |
3.64 |
12 |
3.66 |
3.68 |
3.69 |
3.71 |
3.72 |
3.74 |
3.76 |
3.77 |
3.79 |
3.80 |
13 |
3.80 |
3.83 |
3.85 |
3.86 |
3.88 |
3.89 |
3.90 |
3.92 |
3.93 |
3.95 |
14 |
3.96 |
3.97 |
3.99 |
4.00 |
4.02 |
4.03 |
4.04 |
4.06 |
4.07 |
4.09 |
15 |
4.10 |
4.11 |
4.13 |
4.14 |
4.15 |
4.17 |
4.18 |
4.19 |
4.20 |
4.22 |
16 |
4.23 |
4.24 |
4.25 |
4.27 |
4.28 |
4.29 |
4.30 |
4.31 |
4.33 |
4.34 |
17 |
4.35 |
4.36 |
4.37 |
4.38 |
4.39 |
4.41 |
4.42 |
4.43 |
4.44 |
4.45 |
18 |
4.46 |
4.47 |
4.48 |
4.49 |
4.50 |
4.52 |
4.53 |
4.54 |
4.55 |
4.56 |
19 |
4.57 |
4.58 |
4.59 |
4.60 |
4.61 |
4.62 |
4.63 |
4.64 |
4.65 |
4.66 |
Carbon (oenological)
COEI-1-CHARBO Oenological carbon
INS N° :153
- Object, origin and field of application
Oenological carbon are of plant (generally wood) origin. In order to increase their adsorption properties, the carbon is subjected to activation either at a high temperature or a lower temperature in the presence of an acid, (generally phosphoric acid). Oenological carbon must not be activated with a metal-based catalyser like zinc chloride.
It is in the form of very fine and light black powder, or in granulated form.
There are humid preparations which can reduce dust accumulation. In this case, weight loss as described in 3.1 can reach 60%.
Carbon can be agglomerated with bentonite.
Carbon for oenological purposes is used to correct alterations due to fungus in addition to the colour of white musts from purple, spotted or oxidised grapes. The carbon can eliminate anthocyanins and oxidised or non-oxidised polyphenols in addition to polysaccharides. The carbon are used to correct the organoleptic characteristics of musts made from grapes altered by fungus.
Oenological carbon can also be used to reduce the presence of Ochratoxin A in musts, the musts during fermentation and in white wine.
Decolourising carbon has a relatively weak deodorising effect.
Absorption by carbon is not very selective and depends on its structure, porosity and specific surface area.
The limit concerning the use of carbons should be compliant with the prescriptions of the OIV International Code of Oenological Practices (expressed by weight of dry carbon).
- Labelling
The label should indicate the storage conditions, the expiration date for humid solutions, and a mention of whether there are existing regulations regarding the usage of the product and specify if it concerns decolourising or deodorising carbon.
- Test trials
3.1. Loss with dessication
- Put 5g of carbon in a silica capsule and heat to 100°C in an incubator.
- After 3 hours of dessicating, weight loss should not be more than 20%.
All limits set for carbon refer to dry carbon weight.
3.2. Ashes
- Incinerate the previously obtained dry residue at 500°C-600°C.
- These ashes should not be more than 10%.
- Carbon agglomerated with bentonite should have ashes more than 10% and less than 30%.
3.3. Soluble matter in acids
- Boil 5 g of dried carbon with 20 ml of concentrated hydrochloric acid (R) and 100 ml of water.
- Once cooled, filter using a fine filter or membrane.
Evaporate the filtrate and dry at 100°C–105°C. The soluble matter content in acids should not be more than 5%.
3.4. Chlorides
- Shake 0.067g of dried carbon and 20 ml of distilled water.
- Filter.
- Add 5 ml of diluted nitric acid (R) to 5 ml of filtrate.
- Fill up to 20 ml and add 0.5 ml of silver nitrate solution at 5% (R).
- Compare any opalescence or cloudiness to a prepared control sample as indicated in the annex. Other methods such as ionic chromatography can be used.
- Chloride content should not be more than 3g/kg.
3.5. Cyanides
- Put a quantity of carbon containing 1 g of dried carbon with 10 ml of diluted sulphuric acid (R) in a 100 ml conical flask.
- Adapt to the conical flask a pressure relief tube plunged into approximately 2 ml of saturated borax solution (R) in a test tube.
- Distil and gather 2 to 3 ml of distillate.
- Add 5 drops of potassium anhydrosulphite solution at 2% (R) and leave for 5 minutes.
- Add 1 ml of iron sulphate solution (II) at 5% (R) and leave for 15 minutes.
- Then add 2 drops of phenolphthalein (R). Use a saturated borax solution (R) to make the solution a little more alkaline.
- Leave for 5 minutes.
- Add 2 drops of iron sulphate (III) and ammonia solution at 10% (R) and 1 ml of concentrated hydrochloric acid (R).
- No colouration nor blue precipitate should form.
3.6. Polycyclic aromatic hydrocarbons
Polycyclic aromatic hydrocarbons including benzoa]pyrene are extracted by hexane; the solvent is evaporated and the residue is taken by the methanoltetrahydrofuran mixture for HPLC analysis following the method described in chapter II.
Note: It is also possible to determine benzoa]pyrene by gas chromatography by using an apolar capillary column with detection by mass spectrometry following the method described in chapter II of the International Oenological Codex.
Benzoa]pyrene content should not be more than 10 μg/kg.
3.7. Sulphides
- Put a quantity of carbon containing 1 g of dried carbon with 10 ml of diluted hydrochloric acid and 10 ml of water in a 50 ml flask.
- Distil and collect 5 ml of distillate in a test tube containing 5 ml of 1 M sodium hydroxide solution.
- 0.5 ml of lead nitrate solution at 1 g per litre (R) is added to 1 ml of test trial solution.
- There should be no brown colouring or black precipitate.
- Sulphide content expressed in sulphur should not be more than 20 mg/kg.
3.8. Preparation of test trial solution
- Put a quantity of carbon corresponding to 2.5 g of dried carbon with 50 ml of a citric acid solution at 5 g a litre with a pH of 3 (R), in a conical flask with a wide opening that can be sealed.
- Shake vigorously for 5 minutes and allow to stand at least 12 hours.
- Filter through a fine filter or a membrane in order to obtain a clear solution.
3.9. Iron
- Add 5 ml of water, 1 ml of concentrated hydrochloric acid, 2 ml of 5% potassium thiocyanate solution (R) to 5 ml of test trial solution as prepared in point 3.8.
- The colouration obtained should be lighter than the control sample prepared with 10 ml of iron salt solution (II) at 0.010 g of iron per litre (R), and 1 ml of concentrated hydrochloric acid (R), 2 ml of 5% potassium thiocyanate solution (R). Atomic absorption spectrophotometry can also be used.
Iron content should not be more than 200 mg/kg.
3.10. Lead
Determine the lead according to the method described in chapter II on the test trial solution prepared according to point 3.8.
Lead content should not be more than 2 mg/kg.
3.11. Mercury
Determine the mercury according to the method described in chapter II on the test trial solution prepared according to point 3.8.
Mercury content should not be more than 1 mg/kg.
3.12. Arsenic
Determine the arsenic according to the method described in chapter II on the test trial solution prepared according to point 3.8.
Arsenic content should not be more than 3 mg/kg.
3.13. Calcium
Determine the calcium according to the method described in chapter II on the test trial solution prepared according to point 3.8.
Calcium content should not be more than 10 g/kg.
3.14. Cadmium
Determine cadmium according to the method described in chapter II on the test trial solution prepared according to point 3.8.
Cadmium content should not be more than 1 mg/kg.
3.15. Zinc
Determine the zinc according to the method described in chapter II on the test trial solution prepared according to point 3.8.
Zinc content should not be more than 25 mg/kg.
3.16. Specific surface area
The specific surface area of a decolourising carbon must be between 600 and 2000 m2/g.
Methylene blue decolourisation is the method used. (Methylene blue indicator).
3.17. Methylene blue indicator
- Prepare 4 conical flasks and place 0.1 g of carbon.
- Add 10, 15, 17 and 20 ml of methylene blue solution at 1.2 g/l (absorbance at 620 nm is between 0.830 and 0.850).
- After shaking for 5 minutes, filter through a slow filter and note the volume of the solution in the conical flask which underwent decolourisation.
Depending on the results, repeat this experiment with different volumes of solution.
Put the solution in a spectrophotometer at 664 nm with the absorbance value of 0.08 with an optical path of 1 cm.
The volume of the methylene blue test solution in ml just discoloured, represents the methylene blue indicator.
- Phenol index
4.1. Introduction
When activated carbon is applied in the treatment of wine, the phenol index can be used to define a limit value over which the carbon is considered as a decolouriser and under which it is regarded as a deodoriser. The phenol index selected is the AWWA B600-90 index
4.2. Principle:
AWWA phenol index: this index, expressed in g of carbon scaled to the dry weight per l of solution represents the carbon powder concentration required to decrease the phenol concentration of a solution from 200 mg/l to 20 mg/l.
4.3. Description of the AWWA method:
This index is determined using an adsorption isotherm based on at least 4 different weights of carbon put in contact with a phenol solution.
This isotherm represents the weight of phenol adsorbed in mg/l/g carbon, in relation to the residual phenol concentration in the solution, expressed in mg/l.
4.4. Reagents
4.4.1. Pure disodic hydrogenophosphate Na2HPO4 for analysis
4.4.2. Distilled water
4.4.3. Pure phosphoric Acid (H3PO4)
4.4.4. Pure phenol
4.4.5. Buffer solution A of disodic hydrogenophosphate with a pH of 6.5 at 104 g/l
In a 1-litre graduated flask, dissolve 104 g of disodic hydrogenophosphate (4.4.1) in 300 ml of hot water (4.4.2), add 14 ml of phosphoric acid (4.4.3) and make up to one litre. Homogenise. Check that the pH is 6.5 0.1
4.4.6. Buffer solution B of disodic hydrogenophosphate with a pH of 6.5 at 10.4 g/l
In a 1-litre graduated flask, place 100 ml of buffer solution A at 104 g/l (4.4.5) and make up with water (4.4.2). Homogenise.
4.4.7. Phenol solution with 1 g/l
In a 100-ml graduated flask, place 100 mg of phenol (4.4.4) and make up to 100 ml with water (4.4.2). Obtain complete dissolution by stirring.
4.4.8. Calibration solutions of phenol with 20, 40, 60, 80, 100, and 120 mg/l
In a series of 100-ml graduated flasks, respectively place 2 ml, 4 ml, 6 ml, 8 ml, 10 ml, and 12 ml of the phenol solution with 1 g/l (4.4.7). Make up to 100 ml using buffer solution B (4.4.6).
4.4.9. Phenol solutions with 200 mg/l
In a 1-litre flask, place 200 ml of the phenol solution at 1 g/l (4.4.7), add 100 ml of buffer solution A (4.4.5), make up to 1 l with water (4.4.2). Homogenise.
4.4.10. Measuring the phenol index of oenological carbon powder
Note: The water content of the carbon must be known in order to scale the index to the dry carbon weight.
4.5. Apparatus
4.5.1. Laboratory glassware i.e.: graduated precision pipettes to measure small volumes, 100-ml and 1-l graduated flasks, funnels, and 300-ml conical bottles
4.5.2. Filter paper
4.5.3. Laboratory balance, precision to within 0.10 mg
4.5.4. Spectrometer capable of operating in the ultraviolet spectrum and housing quartz tanks with an optical thickness of 1 cm.
4.5.5. Laboratory shaker (it is not recommended to use a magnetic bar)
4.6. Procedure
4.6.1. Phenol calibration curve.
Measure the absorbance at 270 nm in tanks with an optical thickness of 1 cm (4.5.4) of each phenol solution with 20, 40, 60, 80, 100, and 120 mg/l (4.4.8). Calculate the straight regression line of the absorbance in relation to the phenol concentration.
Note:The blank is based on buffer solution B (4.4.6).
4.6.2. Determine the residual phenol for each carbon (4.4.10)
In a series of 300-ml conical flasks, place 200 ml of phenol solution at 200 mg/l (4.4.9), then respectively 0.4, 0.5, 0.6 and 0.7 g of carbon; close the bottle.
For these 4 preparations, stir for 30 minutes (4.5.5) so that the carbon remains in suspension.
Filter on paper (4.5.2) the 4 samples containing the carbon and a blank (phenol solution with 200 mg/l (4.4.8) without carbon).
Measure the absorbance at 270 nm in tanks with an optical thickness of 1 cm (4.5.4) of each one of the filtered solutions.
Note 1 The blank is based on buffer solution B (4.4.6).
Note 2 At least one of the quantities of carbon must adsorb 90% of the phenol in the solution; if not, widen the carbon weight range.
4.7. Calculations
4.7.1. Determine the percentage of residual phenol in each filtrate for each activated carbon:
residual % = milligram per litre of residual phenol filtrate * 100/200 (milligram per litre of phenol in the test solution). i.e. a = % residual phenol |
4.7.2. Determine the percentage of X (adsorbed phenol):
% of X = 100 -% residual in the filtrate. i.e. X = 100 – a |
4.7.3. The quantities of activated carbon for 200 ml of phenol solution are multiplied by 5 to obtain the quantities of activated carbon, i.e. M in grams per litre.
4.7.4. Calculate the percentage of the value of X/M for each activated carbon.
4.7.5. Plot the isotherm: percentage of residual normality of the filtrate on the X-axis (a) and the percentage of X/M on the Y-axis using 2x2 logarithmic paper; establish the straight regression line and determine the regression equation. It is also possible to calculate the regression using the logarithm for the values of a and X/M.
4.7.6. Determine X/M at 10%; i.e. C (when the residual phenol concentration of the filtrate is 10%).
4.7.7. Phenol index in grams per litre = 90/C * (100 -% of humidity/100); i.e. P
This formula refers to activated carbon without humidity.
4.7.8. Limit values
A carbon is regarded as a deodouriser if its phenol index is lower than 3.5
4.7.9. Examples
Standard cuve |
|
Abs at 270 nm |
Phenol mg/l |
0.303 |
20 |
0.603 |
40 |
0.8777 |
60 |
1.3443 |
100 |
1.53 |
120 |
Calibration straight line for phenol titration |
A1 |
g of carbon |
% humidity |
Dry weight |
Abs |
C phenol |
0,25 |
1.31 |
0.2467 |
0.309 |
23 |
|
0,6255 |
1.31 |
0.6173 |
0.065 |
5 |
|
0,7136 |
1.31 |
0.7043 |
0.0454 |
3 |
|
0,7829 |
1.31 |
0.7726 |
0.0367 |
3 |
A1 Calculations |
|||||||
a |
X |
M |
a |
X/M |
Log a |
Log X/M |
|
11.53 |
88.47 |
1.23 |
11.53 |
71.72 |
1.061829 |
1.855636 |
|
2.42 |
97.58 |
3.09 |
2.42 |
31.61 |
0.384605 |
1.499871 |
|
1.69 |
98.31 |
3.52 |
1.69 |
27.92 |
0.228747 |
1.445885 |
|
1.37 |
98.63 |
3.86 |
1.37 |
25.53 |
0.136357 |
1.407065 |
|
c |
P |
||||||
10.00 |
66.16 |
1 |
1.8206 |
1.3 |
Adsorption isotherm of carbon A1 |
A2 |
g of carbon |
% Humidity |
Dry weight |
Abs |
C phenol |
0.4054 |
1.60 |
0.3989 |
0.9969 |
74 |
|
0.6012 |
1.60 |
0.5916 |
0.679 |
51 |
|
0.7914 |
1.60 |
0.7787 |
0.4972 |
37 |
|
0.9032 |
1.60 |
0.8887 |
0.4126 |
31 |
|
1.4040 |
1.60 |
1.3815 |
0.2654 |
20 |
A2 Calculations |
|||||||
a |
X |
M |
a |
X/M |
Log a |
Log X/M |
|
37.18 |
62.82 |
1.99 |
37.18 |
31.49 |
1.570343 |
1.498241 |
|
25.33 |
74.67 |
2.96 |
25.33 |
25.25 |
1.403561 |
1.402199 |
|
18.54 |
81.46 |
3.89 |
18.54 |
20.92 |
1.268222 |
1.320569 |
|
15.39 |
84.61 |
4.44 |
15.39 |
19.04 |
1.187221 |
1.279687 |
|
9.90 |
90.10 |
6.91 |
9.90 |
13.04 |
0.995635 |
1.115409 |
|
c |
P |
||||||
10 |
13.70 |
1 |
1.1367 |
6.5 |
Adsorption isotherm of carbon A2 |
4.7.10. Collaborative analysis: AWWA phenol indices in g/l
Lab 1 |
Lab 2 |
Lab 3 |
Lab 4 |
|
A1 |
1.7 |
1.53 |
1.8 |
1.3 |
A2 |
5.1 |
4.56 |
6.2 |
6.5 |
A3 |
1.3 |
1.29 |
1.8 |
1.4 |
A4 |
5.8 |
4.95 |
10.0 |
7 |
B1 |
11.4 |
7.18 |
10.6 |
7.6 |
B2 |
1.8 |
1.47 |
2.3 |
1.4 |
B3 |
49.4 |
21.97 |
18.0 |
17.5 |
B4 |
2.9 |
2.80 |
3.6 |
2.6 |
C1 |
1.9 |
1.69 |
2.3 |
1.8 |
C2 |
1.7 |
1.56 |
2.0 |
1.5 |
C3 |
5.4 |
4.71 |
6.2 |
4.9 |
C4 |
5.4 |
4.55 |
6.0 |
4.7 |
Reproducibility: 2.88 for the 5.86 general average SR = 1,03
- Determination of the decolourisation capacity of carbon
5.1. Principle
Measuring the decolourisation of an oenocyanin solution with a precise amount of carbon under defined conditions.
5.2. Apparatus:
Equipment:
5.2.1. Precision balance in mg
5.2.2. 5.2.2 Magnetic stirrer
5.2.3. Absorption spectrophotometer for OD to 420, 520 and 620 nm measures Glassware:
5.2.4. 250 ml cylindrical flask
5.2.5. 250 ml conical flask
5.2.6. 200 ml volumetric flask
5.2.7. Chamber with a 1 mm optical path for an absorption spectrophotometer.
5.3. Reagents
5.3.1. Very pure demineralised water
5.3.2. Crystallised acetic acid
5.3.3. Tartaric acid
5.3.4. Crystallised sodium acetate
5.3.5. 96% volume ethanol
5.3.6. Oenocyanin powder
5.4. Preparation of oenocyanin solution
5.4.1. Pour approximately 150 ml of demineralised water (5.3.1) in a 250 ml cylindrical flask (5.2.4).
5.4.2. Shake (5.2.2).
5.4.3. Weigh 0.900 g 0.001 g of oenocynanin (5.3.6) and dissolve by adding small amounts while stirring in a vortex mixer.
5.4.4. Weigh 1.400 g 0.01 g of tartaric acid (5.3.3) and pour into the cylindrical flask (5.2.4).
5.4.5. Pour 0.8 ml of crystallised acetic acid (5.3.2) and 1.4 g of crystallised sodium acetate (5.3.4).
5.4.6. Shake continuously until completely dissolved (5.2.2)
5.4.7. Transfer to a 200 ml volumetric flask (5.2.6).
5.4.8. Adjust to 200 ml with the rinsing water from the cylindrical flask of the 5.4 preparation.
5.4.9. Transfer again into a 250 ml cylindrical flask (5.2.4).
5.4.10. Shake (5.2.2).
5.4.11. Centrifuge 150 ml of the solution for 10 minutes at 10,000 g place the supernatant in a cuvette with 1 mm optical path
5.4.12. Measure the colour intensity of the spectrometer (5.2.3)
CI1=OD 420+ OD 520+ OD 620 |
CI1= |
5.5. Decolourisation by carbon
5.5.1. Weigh 100 mg of dried carbon.
(Measure the humidity in order to define the exact dose of humid carbon to be used).
5.5.2. Put the carbon in 100 ml of oenocyanin solution with colour intensity
CI1= 4 |
5.5.3. Shake for 30 minutes (5.2.2).
5.5.4. Allow to stand for 10 minutes and centrifuge 10 ml of this mixture for 10 minutes at 10,000 g.
5.5.5. Measure the colour intensity with a spectrometer (5.2.3) under 1 mm of optical path:
CI2=OD 420+ OD 520+ OD 620 |
5.6. Calculation of decolourisation capacity
The decolourisation capacity (DC):
DC=100 (CI1-CI2)/CI1 |
Carbon is considered as a ‘decolourising agent’ when DC is more than or equal to 40.
- Storage
Carbon cannot be stored in open bags because of its adsorption capacities. Oenological carbon must be stored in sealed packages away from volatile substances that it could adsorb.
Chitin-glucan
COEI-1-CHITGL Chitin-glucan
[]m - []n
CAS number Chitin: 1398-61-4
CAS number β-glucan: 9041-22-9
- Purpose, origin and scope
Chitin-glucan is of fungus origin and is a natural polymer, the main component of the cellular walls of Aspergillus niger. It is initially extracted and purified from the mycelium of Aspergillus niger. This fungal resource is a by-product of the citric acid produced for the food and pharmaceutical markets.
Chitin-glucan is composed of polysaccharides chitin (repeat units N-acetyl-D-glucosamine) and 1,3-ß-glucan (repeat unit D-glucose). The two polymers are covalently connected and form a three-dimensional network. The chitin/glucan ratio ranges from 25:75 to 60:40 (m/m).
It is used as a fining agent of musts during racking in order to reduce the colloid content and cloudiness.
It is also used for stabilising wines prior to bottling after alcoholic fermentation. This polymer has a stabilising capacity with respect to ferric breakages. It also helps eliminate undesirable compounds such as heavy metals (lead, cadmium), mycotoxins, etc.
- Synonyms
Poly(N-acetyl-D-glucosamine)-poly(D-glucose) and 1,3- β-glucan
- Labelling
The following information must be stated on the packaging label: fungal origin, product for oenological use, use and conservation conditions and use-by date.
- Characters
4.1. Aspect
Chitin-glucan comes in the form of a white, odourless and flavourless powder. Chitin-glucan is almost completely insoluble in aqueous or organic medium.
4.2. Purity and soluble residues
The purity of the product must be equal to or higher than 95 %.
Dissolve 5 g of chitin-glucan in 100 ml of bidistilled water and agitate for 2 minutes. Filter after cooling on a fine mesh filter or membrane.
Evaporate the filtrate and dry at 100-105 °C. The content of solubles should not be higher than 5 %.
- Tests
5.1. Identification and chitin-glucan ratio
5.1.1. Determination of the chitin-glucan ratio
The chitin/glucan ratio is determined using the 13C NMR spectrum in solid phase, by comparison with the spectrum of a pure chitin reference sample.
This method is detailed in appendix I.
5.2. Loss during desiccation
In a glass cup, previously dried for 1 hour in an oven at 100-105 °C and cooled in a desiccator, place 10 g of the analyte. Allow to desiccate in the drying oven at 100-105 °C to constant mass. Weigh the dry residue amount after cooling in the desiccator.
The weight loss must be lower than 10 %.
Note: all the limits stated below are reported in dry weight except for the microbiological analyses
5.3. Ashes
Incinerate without exceeding 600°C the residue left from the determination of the loss during desiccation as described in 5.2. Allow to calcine for 6 hours. Allow the crucible to cool in a desiccator and weigh.
The total ash content should not be higher than 3 %.
5.4. Preparation of the test solution
Before determining the metals, the sample is dissolved by acid digestion (HNO3, H2O2 and HCl). Mineralisation is performed in a closed microwave system. The sample undergoes neither crushing nor drying before mineralisation.
The reagents used for the mineralisation of chitin-glucan are as follows: HNO3 (65 %) (Suprapur), HCl (37 %) (Suprapur), H2O2 (35 %). The 0.5 to 2 g sample of chitin-glucan is placed in a flask to which are added 25 ml of HNO3, 2 ml of HCl and 3 ml of H2O2. This is submitted to microwave digestion (Power of 60 % for 1 min, 30 % for 10 min, 15 % for 3 min, and 40 % for 15 min). The solution is diluted in a volumetric flask with bidistilled water to a final volume of 25.0 ml.
The metal contents can then be determined.
5.5. Lead
Lead is determined by atomic absorption spectrophotometry, using the method described in appendix II.
The lead content must be lower than 1 mg/kg.
It is also possible to achieve lead determination by atomic absorption, using the method described in chapter II of the International Oenological Codex.
5.6. Mercury
Mercury is determined by atomic absorption spectrophotometry, using the method described in appendix II.
The mercury content must be lower than 0.1 mg/kg.
It is also possible to achieve mercury determination by atomic absorption, using the method described in chapter II of the International Oenological Codex.
5.7. Arsenic
Arsenic is determined by atomic absorption spectrophotometry, using the method described in appendix II.
The arsenic content must be lower than 1 mg/kg.
It is also possible to achieve arsenic determination by atomic absorption, using the method described in chapter II of the International Oenological Codex.
5.8. Cadmium
Cadmium is determined by atomic absorption spectrophotometry, using the method described in appendix II.
The cadmium content must be lower than 1 mg/kg.
It is also possible to achieve cadmium determination by atomic absorption, using the method described in chapter II of the International Oenological Codex.
5.9. Chromium
Chromium is determined by atomic absorption spectrophotometry, using the method described in appendix II.
The chromium content must be lower than 10 mg/kg.
It is also possible to achieve chromium determination by atomic absorption, using the method described in chapter II of the International Oenological Codex.
5.10. Zinc
Zinc is determined by atomic absorption spectrophotometry, using the method described in appendix II.
The zinc content must be lower than 50 mg/kg.
It is also possible to achieve zinc determination by atomic absorption, using the method described in chapter II of the International Oenological Codex.
5.11. Iron
Iron is determined by atomic absorption spectrophotometry, using the method described in appendix II.
The iron content must be lower than 100 mg/kg.
It is also possible to achieve iron determination by atomic absorption, using the method described in chapter II of the International Oenological Codex.
5.12. Copper
Copper is determined by atomic absorption spectrophotometry, using the method described in appendix II.
The copper content must be lower than 30 mg/kg.
It is also possible to achieve copper determination by atomic absorption, using the method described in chapter II of the International Oenological Codex.
5.13. Microbiological control
5.13.1. Total bacteria count
The total bacteria count is performed according to the horizontal method by means of the colony count technique at 30 °C on the PCA medium described in appendix III.
Less than 1000 CFU/g of preparation.
It is also possible to carry out the enumeration as described in chapter II of the International Oenological Codex.
5.13.2. Enterobacteria
The enumeration of Enterobacteria is carried out according to the horizontal method by means of the colony count technique at 30 °C described in appendix IV.
Less than 10 CFU/g of preparation.
5.13.3. Salmonella
Carry out the enumeration as described in chapter II of the International Oenological Codex.
Absence checked on a 25 g sample.
5.13.4. Coliform bacteria
Carry out the enumeration as described in chapter II of the International Oenological Codex.
Less than 100 CFU/g of preparation.
5.13.5. Yeasts
The enumeration of yeasts is carried out according to the horizontal method by means of the colony count technique at 25 °C on the YGC medium described in appendix V.
Less than 100 CFU/g of preparation.
It is also possible to carry out the enumeration as described in chapter II of the International Oenological Codex.
5.13.6. Moulds
The enumeration of moulds is carried out according to the horizontal method by means of the colony count technique at 25 °C on the YGC medium described in appendix VI.
Less than 100 CFU/g of preparation.
It is also possible to carry out the enumeration as described in chapter II of the International Oenological Codex.
- Ochratoxin A testing
Prepare an aqueous solution (distilled water) of chitin-glucan at 1 % and agitate for 1 hour, then carry out determination using the method described in the Compendium of International Methods of Analysis of Wine and Musts.
Less than 5 µg/kg.
- Storage
Keep container closed and store in a cool and dry place.
Appendix I: Determination of the chitin/glucan ratio
- Principle
This method consists in determining the chitin/glucan ratio using the 13C RMN spectrum in solid phase.
- Reagents and materials
- Chitin glucan sample
- Osmosis purified water
- Hydrochloric acid 1 M
- Pure ethanol
- Pure chloroform
- Pure methanol
- Acetone
- Standard laboratory material, pipettes, cylindrical glass vases, porosity filters 30 μm…
- Rotary shaker
- Laboratory centifuge
- Conductimeter
- Nuclear Magnetic resonance apparatus
- Sample preparation
Before the determination, samples are prepared according to a precise protocol as described below:
3.1. Washing with HCl 1 M (2.3)
This step consists in mixing 2 g of chitin-glucan (2.1) and 40 ml of HCl 1 M in a tube flask.
This mixture is agitated for 30 min at 320 rpm then centrifuged at 4000 rpm for 10 min. The supernatant is eliminated.
This step is repeated once.
3.2. Washing with osmosis purified water
This step consists in mixing the sediment from the previous step with 40 ml of osmosis purified water (2.2).
This mixture is centrifuged for 10 min at 4000 rpm. The supernatant is eliminated.
This step is repeated until the supernatant conductivity is lower than 100 μS/cm.
3.3. Washing with ethanol
This step consists in mixing the sediment from the previous step with 40 ml of ethanol (2.4).
This mixture is centrifuged for 10 min at 4000 rpm. The supernatant is eliminated.
This step is repeated once.
3.4. Washing with chloroform/methanol
This step consists in mixing the sediment from the previous step with 40 ml of a of 50/50, v/v of chloroform (2.5) and methanol (2.6) mixture.
This mixture is agitated for 30 min at 320 rpm then centrifuged at 4000 rpm for 10 min. The supernatant is eliminated.
This step is repeated once.
3.5. Washing with acetone and drying
This step consists in mixing the sediment from the previous step with 40 ml of acetone (2.7).
This mixture is agitated for 30 min at 320 rpm then centrifuged at 4000 rpm for 10 min.
After centrifugation, pour the supernatant on a 30 μm filter, rinse the tube flask with acetone (2.7) and pour everything on the filter.
Place the material located on the filter in a crystallising dish and allow to dry.
After drying, the product is ready to be analysed by NMR.
- Procedure
The prepared samples are then analysed on the Brücker Avance DSX 400WB nuclear magnetic resonance instrument (or the equivalent).
The analysis conditions are as follows:
- Magnetic field: 9.04 Tesla
- Larmor frequency: 83 kHz
- Time interval between 2 magnetic pulses: 5s
- Time period during which the magnetic pulse is applied: 5,5ms
- Number of magnetic pulse sequences: 3000
- Expression of the results
5.1. The beta-glucan content is determined from the area of the four resonance bands.
5.2. The results are expressed in mol %.
Appendix II : Metal determination by atomic emission spectroscopy
- Principle
This method consists in measuring atomic emission by an optical spectroscopy technique.
- Sample preparation
Before the determination of metals, the sample is dissolved by acid digestion (HNO3, H2O2 and HCl). Mineralisation takes place in closed microwave system. The sample undergoes neither crushing nor drying before mineralisation.
The reagents used for the mineralisation of chitosan are as follows: HNO3 (65 %) (Suprapur), HCl (37 %) (Suprapur), H2O2 (35 %). The 0.5 to 2 g sample of chitin-glucan is placed in a flask to which are added 25 ml of HNO3, 2 ml of HCl and 3 ml of H2O2. The whole is then submitted to microwave digestion (Power of 60 % for 1 min, 30 % for 10 min, 15 % for 3 min, and 40 % for 15 min). The solution is then diluted in a volumetric flask with bidistilled water to a final volume of 25.0 ml.
The metal contents can then be determined.
- Procedure
The dissolved samples are nebulised and the resulting aerosol is transported in a plasma torch induced by a high frequency electric field. The emission spectra are dispersed by a grating spectrometer and the line intensity is evaluated by a detector (photomultiplier). The detector signals are processed and controlled by a computer system. A background noise correction is applied to compensate for the background noise variations.
- Expression of the results
The metal concentrations in the oenological products are expressed in mg/kg
Appendix III : Total bacteria count by counting the colonies obtained at 30 °C
PCA medium
Composition:
Peptone |
5.0 g |
Yeast extract |
2.5 g |
Glucose |
1.0 g |
Agar-agar |
15 g |
Adjusted to |
pH 7.0 |
Water |
complete to 1000 ml |
The medium is sterilised before use in an autoclave at 120 °C for 20 min.
The Petri dishes are inoculated by pour plate method and spiral plating method.
After inoculation, they are incubated at 30 °C in aerobiosis for 48 to 72 hours.
Count the CFU number.
Appendix IV : Enumeration of Enterobacteria is carried out according to the horizontal method by means of the colony count technique
at 30 °C
VRBG medium
Composition:
Peptone |
7 g |
Yeast extract |
3 g |
Glucose |
10 g |
Sodium Chloride |
5 g |
Crystal Violet |
0.002 g |
Neutral Red |
0.03 g |
Agar-agar |
13 g |
Bile salts |
1.5 g |
Adjusted to |
pH 7.4 |
Water |
complete to 1000 ml |
The medium is sterilised before use in an autoclave at 120 °C for 20 min.
The Petri dishes are inoculated by pour plate method and spiral plating method.
After inoculation, they are incubated at 30 °C in aerobiosis for 18 to 24 hours.
Count the CFU number.
Appendix V : Enumeration of yeasts by counting
YGC medium
Composition:
Yeast extract |
5.0 g |
D-glucose |
20 g |
Agar-agar |
14.9 g |
Choramphenicol |
0.1 g |
Adjusted to |
pH 6.6 |
Water |
complete to 1000 ml |
The medium is sterilised before use in an autoclave at 120 °C for 20 min.
The Petri dishes are inoculated by pour plate method and spiral plating method.
After inoculation, they are incubated at 25 °C in aerobiosis for 3 to 5 days without being turned over.
Count the number of yeasts.
Appendix VI :Enumeration of the moulds by counting
YGC medium
Composition:
Yeast extract |
5.0 g |
D-glucose |
20 g |
Agar-agar |
14.9 g |
Choramphenicol |
0.1 g |
Adjusted to |
pH 6.6 |
Water |
complete to 1000 ml |
The medium is sterilised before use in an autoclave at 120 °C for 20 min.
The Petri dishes are inoculated by pour plate method and spiral plating method.
After inoculation, they are incubated at 25 °C in aerobiosis for 3 to 5 days without being turned over.
Count the number of moulds.
Chitosan
COEI-1-CHITOS Chitosan
[]n
CAS number Chitosan : [9012-76-4]
Chitosan |
- Purpose, origin and applicability
Chitosan, a natural polysaccharide prepared of fungal origin, is initially extracted and purified from reliable and abundant food or biotechnological fungal sources such as Agaricus bisporus or Aspergillus niger.
Chitosan is obtained by hydrolysis of a chitin-rich extract. Chitin is a polysaccharide composed of several N-acetyl-D-glucosamine units interconnected by β (1.4) type linkages.
Chitosan is composed of glucosamine sugar units (deacetylated units) and N-acetyl-D-glucosamine units (acetylated units) interconnected by β (1.4) type linkages.
It is used as a fining agent in the treatment of musts for flotation clarification to reduce cloudiness and the content of unstable colloids.
It is also used for stabilising wines. This polymer actually helps eliminate undesirable micro-organisms such as Brettanomyces.
- Synonyms
Poly(N-acetyl-D-glucosamine)-poly(D-glucose).
- Labelling
The following information must be stated on the packaging label: exclusively fungal origin, product for oenological use, use and conservation conditions and use-by date.
- Characters
4.1. Aspect and solubility
Chitosan comes in the form of a white, odourless and flavourless powder. Chitin-glucan is almost completely insoluble in aqueous or organic medium.
4.2. Purity and soluble residues
The purity of the product must be equal to or higher than 95 %.
Dissolve 5 g of chitin-glucan in 100 ml of bidistilled water and agitate for 2 minutes. Filter after cooling on a fine mesh filter or membrane.
Evaporate the filtrate and dry at 100-105 °C. The content of solubles should not be higher than 5 %.
- Tests
5.1. Determination of the acetylation degree and chitosan origin
5.1.1. Determination of the acetylation degree
The acetylation degree is determined by potentiometric titration, using the method described in Appendix I.
5.1.2. Determination of the source
Chitosan, as a natural polymer, is extracted and purified from fungal sources; it is obtained by hydrolysis of a chitin-rich extract. This chitosan is considered identical to chitosan from shellfish in terms of structures and properties.
An identification of the origin of chitosan is made based on 3 characteristics: content of residual glucans (refer to method in annex II), viscosity of chitosan in solution 1 % and settled density (following settlement).
Only fungal origin chitosan has both contents of residual glucan > at 2 %, a settled density ≥ at 0,7 g/cm3 and viscosity in solution 1 % in acetic acid 1 % < at 15 cPs
5.2. Loss during desiccation
In a glass cup, previously dried for 1 hour in an oven at 100-105 °C and cooled in a desiccator, place 10 g of the analyte. Allow to desiccate in
the drying oven at 100-105 °C to constant mass. Weigh the dry residue amount after cooling in the desiccator.
The weight loss must be lower than 10 %.
Note: all the limits stated below are reported in dry weight except for the microbiological analyses
5.3. Ashes
Incinerate without exceeding 600 °C the residue left from the determination of the loss during desiccation as described in 5.2. Allow to calcine for 6 hours. Allow the crucible to cool in a desiccator and weigh.
The total ash content should not be higher than 3 %.
5.4. Preparation of the test solution
Before determining the metals, the sample is dissolved by acid digestion (HNO3, H2O2 and HCl). Mineralisation is performed in a closed microwave system. The sample undergoes neither crushing nor drying before mineralisation.
The reagents used for the mineralisation of chitosan are as follows: HNO3 (65 %) (Suprapur), HCl (37 %) (Suprapur), H2O2 (35 %). The 0.5 to 2 g sample of chitosan is placed in a flask to which are added 25 ml of HNO3, 2 ml of HCl and 3 ml of H2O2. This is submitted to microwave digestion with a maximum power of 1200 watts; Power of 60 % for 1 min, 30 % for 10 min, 15 % for 3 min, and 40 % for 15 min). The solution is diluted in a volumetric flask with bidistilled water to a final volume of 25.0 ml.
The metal contents can then be determined.
5.5. Lead
Lead is determined by atomic absorption spectrophotometry, using the method described in appendix II.
The lead content must be lower than 1 mg/kg.
It is also possible to achieve lead determination by atomic absorption, using the method described in chapter II of the International Oenological Codex.
5.6. Mercury
Mercury is determined by atomic absorption spectrophotometry, using the method described in appendix II.
The mercury content must be lower than 0.1 mg/kg.
It is also possible to achieve mercury determination by atomic absorption, using the method described in chapter II of the International Oenological Codex.
5.7. Arsenic
Arsenic is determined by atomic absorption spectrophotometry, using the method described in appendix II.
The arsenic content must be lower than 1 mg/kg.
It is also possible to achieve arsenic determination by atomic absorption, using the method described in chapter II of the International Oenological Codex.
5.8. Cadmium
Cadmium is determined by atomic absorption spectrophotometry, using the method described in appendix II.
The cadmium content must be lower than 1 mg/kg.
It is also possible to achieve cadmium determination by atomic absorption, using the method described in chapter II of the International Oenological Codex.
5.9. Chromium
Chromium is determined by atomic absorption spectrophotometry, using the method described in appendix II.
The chromium content must be lower than 10 mg/kg.
It is also possible to achieve chromium determination by atomic absorption, using the method described in chapter II of the International Oenological Codex.
5.10. Zinc
Zinc is determined by atomic absorption spectrophotometry, using the method described in appendix II.
The zinc content must be lower than 50 mg/kg.
It is also possible to achieve zinc determination by atomic absorption, using the method described in chapter II of the International Oenological Codex.
5.11. Iron
Iron is determined by atomic absorption spectrophotometry, using the method described in appendix II.
The iron content must be lower than 100 mg/kg.
It is also possible to achieve iron determination by atomic absorption, using the method described in chapter II of the International Oenological Codex.
5.12. Copper
Copper is determined by atomic absorption spectrophotometry, using the method described in appendix II.
The copper content must be lower than 30 mg/kg.
It is also possible to achieve copper determination by atomic absorption, using the method described in chapter II of the International Oenological Codex.
5.13. Microbiological control
5.13.1. Total bacteria count
The total bacteria count is performed according to the horizontal method by means of the colony count technique at 30 °C on the PCA medium described in appendix III.
Less than 1000 CFU/g of preparation.
It is also possible to carry out the enumeration as described in chapter II of the International Oenological Codex.
5.13.2. Enterobacteria
The enumeration of Enterobacteria is carried out according to the horizontal method by means of the colony count technique at 30 °C described in appendix IV.
Less than 10 CFU/g of preparation.
5.13.3. Salmonella
Carry out the enumeration as described in chapter II of the International Oenological Codex.
Absence checked on a 25 g sample.
5.13.4. Coliform bacteria
Carry out the enumeration as described in chapter II of the International Oenological Codex.
Less than 100 CFU/g of preparation.
5.13.5. Yeasts
The enumeration of yeasts is carried out according to the horizontal method by means of the colony count technique at 25 °C on the YGC medium described in appendix VI.
Less than 100 CFU/g of preparation.
It is also possible to carry out the enumeration as described in chapter II of the International Oenological Codex.
5.13.6. Moulds
The enumeration of moulds is carried out according to the horizontal method by means of the colony count technique at 25 °C on the YGC medium described in appendix VII.
Less than 100 CFU/g of preparation.
It is also possible to carry out the enumeration as described in chapter II of the International Oenological Codex.
- Ochratoxin A testing
Prepare an aqueous solution (distilled water) of chitosan at 1 % and agitate for 1 hour, then carry out determination using the method described in the Compendium of International Methods of Analysis of Wine and Musts.
Less than 5 μg/kg.
6.1. Storage
Keep container closed and store in a cool and dry place.
Appendix I: Determination of the acetylation degree
- Principle
This method consists in determining the acetylation degree of chitosan by titration of the amino groups. The acetylation degree is the ratio of the number of N-acetyl-glucosamine units to the number of total monomers.
This method is based on the method described by Rinaudo et al., (1999).
The titration of a chitosan solution by means of NaOH at 0.1 M must be performed in order to identify two pH jumps from 0 to 14.
Chitosan is dissolved in 0.1M HCl, the amino groups (on the deacetylated glucosamine units (G)) are positively charged (HCl in excess)).
The chitosan solution (of known quantity) is titrated by NaOH of known concentration. In the first part of the reaction, the excess quantity of HCl is determined:
1.1. HCl (excess)+NaOH+ NH3+Cl- --> NaCl + H2O + NH3+Cl-
After the first pH jump, the quantity of charged amino groups is determined:
1.2. HCl +H2O + NH3+Cl-+ NaOH --> NH2 + 2H2O + 2NaCl
1.3. After the second pH jump, the excess quantity of NaOH is measured.
The determination of the NaOH volume between the two jumps makes it possible to identify the quantity of charged amines.
- Reagents and materials
2.1. Commercial preparation of chitosan
2.2. Distilled or deionised water
2.3. Chlorhydric acid 0,3 M
2.4. Sodium Hydroxide 0,1M
2.5. Glass cylindrical flasks, pipettes, burettes…
2.6. Magnetic mixer and stir bar
2.7. pH-meter with temperature sensor.
- Samples preparation
Before determination, the samples are prepared according to the protocol described hereafter:
100 mg of chitosan are placed into a cylindrical flask to which 3 ml of 0.3 M HCl and 40 ml of water are added. Agitate for 12 hours.
- Procedure
First introduce the pH electrode of the pH-meter as well as the temperature sensor into the cylindrical flask. Check that the pH value is lower than 3.
To bring to pH = 1, add a V1 volume (ml) of HCl 0.3 M and agitate.
Then to bring to pH = 7 with a V2 volume (ml) of 0.1 M NaOH
These operations can be carried out using an automatic titrator.
- Expression of results
The acetylation degree of chitosan is expressed in %. This formula is the ratio of the mass of acetylated glucosamine (aG) units in g actually present in the sample, to the mass in g that would be present if all the groups were acetylated, where:
- Mcs: dry weight of chitosan in g
For a 1 g sample
With G = Glucosamine part; a = acetylated part
aG weight actually present (in g) =
aG weight if all the deacetylated groups were acetylated (in g) =
The acetylation degree will be equal to DA, where:
Bibliography
- Rinaudo, M., G. Pavlov and J. Desbrieres. 1999. Influenced of acetic acid concentration on the solubilization of chitosan. Polym. 40, 7029-7032.
Appendix II: Determination of the residual glucan content
- Principle
This method consists in determining the content of residual glucans in chitosan by means of spectrophotometry.
This method is based on a colorimetric reaction with a response depending on the degradation of the starch hydrolysates by hot concentrated sulphuric acid.
This degradation gives a brown yellow compound with a colour intensity proportional to the content of residual glucans.
- Reagents and materials
2.1. Glucan 97% (Société Mégazyme)
2.2. Commercial preparation of chitosan
2.3. Distilled or deionised water
2.4. Ethanol
2.5. Acetic acid 1%
2.6. Solution of phenol 5%
2.7. Glacial acetic acid 100%
2.8. Glass cylindrical flasks, pipettes, volumetric flasks,…
2.9. Magnetic mixer and stir bar
2.10. Chronometer
- Preparation of the standard range
A stock solution of glucan (glucan with a purity of 97 % is provided by the company Megazyme) is prepared according to the precise protocol described hereafter:
500 mg of glucan are introduced into a volumetric flask of 100 ml into which 6 ml of ethanol and 80 ml of distilled water are added.
Agitate and boil out to allow glucan dissolution
Allow to cool, adjust to the filling mark with water
Agitate for 30 minutes.
Pour 1 ml of this solution into a 50 ml volumetric flask and adjust to the filling mark with 1 % acetic acid.
The solution is ready to use to produce the standard range according to the protocol hereafter.
Stock solution V (ml) |
Water V (ml) |
Glucan M (μg) |
0 |
1 |
0 |
0.1 |
0.9 |
10 |
0.3 |
0.7 |
30 |
0.5 |
0.5 |
50 |
0.7 |
0.3 |
70 |
- Samples preparation
Before determination, the samples are prepared according to the protocol described hereafter:
100 mg of chitosan are placed into a 50 ml volumetric flask to which 25 ml of 1 % acetic acid are added.
Agitate for 12 hours then adjust to the filling mark.
- Procedure
In a test tube, add 1 ml of the analyte solution, 1 ml of phenol at 5 % and 5 ml of concentrated sulphuric acid.
Agitate this mixture using a vortex for 10 s, then allow to cool for 1 hour.
The absorbance A is measured at 490 nm.
- Expression of the results
Determine the glucan content in µg/g from the calibration curve (0-70 μg). This content is expressed in μg/g of chitosan.
Appendix III: Metal determination by atomic emission spectroscopy
- Principle
This method consists in measuring atomic emission by an optical spectroscopy technique.
- Sample preparation
Before the determination of metals, the sample is dissolved by acid digestion (HNO3, H2O2 and HCl). Mineralisation takes place in closed microwave system. The sample undergoes neither crushing nor drying before mineralisation.
The reagents used for the mineralisation of chitosan are as follows: HNO3 (65 %) (Suprapur), HCl (37 %) (Suprapur), H2O2 (35 %). The 0.5 to 2 g sample of chitosan is placed in a flask to which are added 25 ml of HNO3, 2 ml of HCl and 3 ml of H2O2. The whole is then submitted to microwave digestion (Power of 60 % for 1 min, 30 % for 10 min, 15 % for 3 min, and 40 % for 15 min). The solution is then diluted in a volumetric flask with bidistilled water to a final volume of 25.0 ml.
The metal contents can then be determined.
- Procedure
The dissolved samples are nebulised and the resulting aerosol is transported in a plasma torch induced by a high frequency electric field. The emission spectra are dispersed by a grating spectrometer and the line intensity is evaluated by a detector (photomultiplier). The detector signals are processed and controlled by a computer system. A background noise correction is applied to compensate for the background noise variations.
- Expression of the results
The metal concentrations in chitosan are expressed in mg/kg.
Appendix IV: Total bacteria count by counting the colonies obtained at 30 °C
PCA medium
Composition:
Peptone |
5.0 g |
Yeast extract |
2.5 g |
Glucose |
1.0 g |
Agar-agar |
15 g |
Adjusted to |
pH 7.0 |
Water |
complete to 1000 ml |
The medium is sterilised before use in an autoclave at 120 °C for 20 min.
The Petri dishes are inoculated by pour plate method and spiral plating method.
After inoculation, they are incubated at 30 °C in aerobiosis for 48 to 72 hours.
Count the CFU number.
Appendix V: Enumeration of Enterobacteria is carried out according to the horizontal method by means of the colony count technique at 30 °C
VRBG medium
Composition:
Peptone |
7 g |
Yeast extract |
3 g |
Glucose |
10 g |
Sodium Chloride |
5 g |
Crystal Violet |
0.002 g |
Neutral Red |
0.03 g |
Agar-agar |
13 g |
Bile salts |
1.5 g |
Adjusted to |
pH 7.4 |
Water |
complete to 1000 ml |
The medium is sterilised before use in an autoclave at 120 °C for 20 min.
The Petri dishes are inoculated by pour plate method and spiral plating method.
After inoculation, they are incubated at 30 °C in aerobiosis for 18 to 24 hours.
Count the CFU number.
Appendix VI : Enumeration of yeasts by counting
YGC medium
Composition:
Yeast extract |
5.0 g |
D-glucose |
20 g |
Agar-agar |
14.9 g |
Choramphenicol |
0.1 g |
Adjusted to |
pH 6.6 |
Water |
complete to 1000 ml |
The medium is sterilised before use in an autoclave at 120 °C for 20 min.
The Petri dishes are inoculated by pour plate method and spiral plating method.
After inoculation, they are incubated at 25 °C in aerobiosis for 3 to 5 days without being turned over.
Count the number of yeasts.
Appendix VII : Enumeration of the moulds by counting
YGC medium
Composition:
Yeast extract |
5.0 g |
D-glucose |
20 g |
Agar-agar |
14.9 g |
Choramphenicol |
0.1 g |
Adjusted to |
pH 6.6 |
Water |
complete to 1000 ml |
The medium is sterilised before use in an autoclave at 120 °C for 20 min.
The Petri dishes are inoculated by pour plate method and spiral plating method.
After inoculation, they are incubated at 25 °C in aerobiosis for 3 to 5 days without being turned over.
Count the number of moulds.
Citric acid, monohydrate
COEI-1-CITACI Citric acid, monohydrate
Monohydrated 3-Carboxy-3-hydroxypentanedioic acid
Acidum citricum
SIN NO. 330
- Objective, Origin and Scope of Application
Citric acid can be used to chemically acidify wines or as a stabilizing agent to limit, in particular, the risks of iron breakdown, or again, for prewashing filter plates. Its maximum proportions in wine may be subject to statutory limits.
- Labelling
The label should indicate product concentration, even when included in mixtures, as well as its safety and storage conditions.
- Properties
Citric acid is found in the form of colorless, translucent crystals which are rather friable and slightly efflorescent, or in crystalline powder form.
- Solubility
Water at 20 °C |
very soluble |
Alcohol, 95% by vol. |
very soluble |
Glycerol |
very soluble |
Ethyl ether |
31.5 g/l |
Aqueous citric acid is inert in polarized light.
- Identifying Characteristics
5.1. Verify total solubility in water. A 1 pp 100 solution (m/v) shows an acid reaction to methyl orange (R).
5.2. Place 2 ml of an aqueous 1 g/l citric acid solution and 0.5 ml of mercury (II) sulfate solution (R) in a test tube. Bring to a boil and add several drops of 2 pp 100 potassium permanganate solution (R). A white precipitate should form.
5.3. Add 1 drop of bromine water (R), 3 drops of concentrated sulfuric acid (R) and 1 drop of saturated potassium permanganate solution to 0.1 ml of 10 pp 100 (m/v) aqueous citric acid solution.Bring to a boil.
Add 2 ml of concentrated sulfuric acid (R). Heat again until completely dissolved. Let cool, then add 0.1 ml of beta-naphthol (R). A green coloring should appear. A pink coloring is obtained under the same conditions if sulforesorcin reagent (R) is used under the same conditions.
5.4. Place 5 ml of chloroform or dichloromethane in a test tube. Add 100-200 mg of citric acid. Shake. The crystals or crystalline powder should collect together at the surface of the liquid. Under these same conditions, tartaric acid collects at the very bottom of the tube.
- Tests
6.1. Foreign Substances
Citric acid should be soluble without residue in its weight of water and in twice its weight of 95% alcohol (by volume).
6.2. Sulfur Ash
After calcination at 600 °C 25 °C, the concentration of sulfur ash should not be greater than 0.5 g/kg.
6.3. Tartaric Acid Determination
Add 2 drops of sulforesorcinic reagent (R) and 2 drops of 10 pp 100 (m/v) citric acid solution to 2 ml of concentrated sulfuric acid (R). Heat to 150 °C. The solution should not develop a violet coloring.
6.4. Preparing the Solution for Tests
Prepare a 10 parts per 100 (m/v) solution.
6.5. Chlorides
Add 14.5 ml of water, 5 ml of nitric acid diluted to 10 pp 100 (R) and 0.5 ml of 5 pp 100 silver nitrate solution (R) to 0.5 ml of the solution prepared for tests under paragraph 6.4. After sitting for 15 minutes in the dark, there should be no clouding. If clouding does occur, it should be less intense than that observed in a control prepared as indicated in the Annex. (Chloride content expressed in terms of hydrochloric acid should be less than 1 g/kg).
6.6. Sulfates
Add 18 ml of water, 1 ml of diluted hydrochloric acid (R) and 2 ml of 10 pp 100 barium chloride solution diluted to 10 pp 100 (R) to 1 ml of the solution prepared for tests under paragraph 6.4. After 15 minutes, there should be no clouding. If tclouding does occur, it should be less intense than that observed in a control prepared by replacing the test solution with 1 ml of 0.1 g/l sulfuric acid solution. (Sulfate content expressed in terms of sulfuric acid should be less than 1 g/kg).
6.7. Oxalic Acid and Barium
Neutralize 5 ml of the solution prepared for tests under paragraph 6.4 by adding concentrated ammonium hydroxide (R). Add 2 drops of acetic acid (R) and 5 ml of saturated calcium sulfate solution (R). There should be no clouding.(Oxalate content expressed in terms of oxalic acid should be less than 0.1g/kg).
6.8. Iron
Add 1 ml of concentrated hydrochloric acid (R) and 2 ml of 5 pp 100 potassium thiocyanate solution (R) to 10 ml of the solution prepared for tests under paragraph 6.4. The resulting red coloration should be less intense than that observed in a control using 1 ml of iron (III) salt solution in a concentration of 0.010 g of iron per liter, 9 ml of water and the same quantities of the same reagents. (Iron content should be less than 10 mg/kg).
Iron may also be analytically quantified by atomic absorption spectometry in accordance with the technique detailed in the Compendium.
6.9. Cadmium
Using the method described in the Annex, quantify cadmium analytically in the test solution prepared according to Par. 6.4. (Cadmium content should be less than 1 mg/kg).
6.10. Lead
Using the method described in the Compendium, determine lead content analytically in the test solution prepared according to Par. 6.4. (Lead content should be less than 1 mg/kg).
6.11. Mercury
Using the method described in the Annex, determine the mercury content analytically in the test solution prepared according to Par. 6.4. (Merucry content should be less than 1 mg/kg).
6.12. Arsenic
Using the method described in the Annex, determine the arsenic content analytically in the test solution prepared according to Par. 6.4. (Arsenic content should be less than 1 mg/kg).
- Storage
Citric acid should be stored in a dry place in air-tight bags.
Carboxymethylcellulose (cellulose gum, CMC)
COEI-1-CMC Carboxymethylcellulose (cellulose gum) (CMC)
INS no. 466
CAS [9004-32-4]
- where R = H or CH2COONa
- Subject, origin and scope
Carboxymethylcellulose (cellulose gum) for oenological use is prepared exclusively from wood by treatment with alkali and monochloroacetic acid or its sodium salt. Carboxymethylcellulose inhibits tartaric precipitation through a "protective colloid" effect. A limited dose is used.
- Synonyms
Cellulose gum, CMC, Sodium CMC, Sodium salt of a carboxymethyl ether of cellulose, NaCMC
- Labelling
Labelling must mention that the carboxymethylcellulose is for use in food, as well as safety and preservation conditions.
- Characteristics
4.1. Description
Granular or fibrous powder, blank or slightly yellowish or greyish, slightly hygroscopic, odourless and tasteless. This may be proposed in
the form of a concentrate for solution in wine prior to use. Solutions must contain at least 3,5 % carboxymethylcellulose.
4.2. Chemical formula
The polymers contain anhydroglucose units substituted with the following general formula: [C6H7O2(OH)x(OCH2COONa)y]n where
- N is the degree of polymerisation
- x = from 1.50 to 2.80
- y = from 0.2 to 1.50
- x + y = 3.0
- (y = degree of substitution)
Note: Only the carboxymethylcellulose possessing a degree of substitution between 0.6 and 1.0 are completely soluble.
4.3. Degree of substitution
Evaluate the degree of substitution using the method described below. The degree of substitution must lie between 0.60 and 0.95.
4.4. Molecular weight
Ranges from 17,000 to 300,000 (degree of polymerisation from 80 to 1,500). The molecular weight can be evaluated through measurement of viscosity.
The viscosity of a 1 % solution must lie between 10 and 15 , or between 20 and 45 for a 2 % solution, or between 200 and 500 for a 4 % solution.
4.5. Composition
Measure the carboxymethylcellulose composition using the method described below. The carboxymethylcellulose content must be at least 99.5 % of the anhydrous substance.
- Tests
5.1. Solubility
Forms viscous colloidal solution with water. Insoluble in ethanol.
5.2. Foam test
Vigorously shake a 0.1 % solution of the sample. No layer of foam appears (this test distinguishes sodium carboxymethylcellulose from other cellulose ethers and from alginates and natural gums).
5.3. Precipitate Formation
To 5 mL of an 0.5% solution of the sample add 5 mL of a 5 % solution of copper sulfate or of aluminium sulfate. No precipitate appears. (This test permists the distinction of sodium carboxymethyl cellulose ethers from other cellulose ethers, and from gelatine, carob bean gum and tragacanth gum)
5.4. Colour reaction
Add 0.5 g of powdered carboxymethylcellulose sodium to 50 mL of water, while stirring to produce a uniform dispersion. Continue the stirring until a clear solution is produced. To 1 mL of the solution, diluted with an equal volume of water, in a small test tube, add 5 drops of 1naphthol. Incline the test tube, and carefully introduce down the side of the tube 2 mL of sulfuric acid so that it forms a lower layer. A red-purple colour develops at the interface.
5.5. Moisture - Loss on drying
Measure the loss on drying using the method described below. Not more than 12 % after drying.
5.6. pH of a 1 % solution
No less than 6 and no more than 8.5 pH units.
5.7. Arsenic
Quantifythe arsenic using the method described in chapter II. The arsenic content must be lower than 3 mg/kg
5.8. Lead
Quantify the lead using the method described in chapter II. The lead content must be lower than 2 mg/kg
5.9. Mercury
Quantify the mercury using the method described in chapter II. The mercury content must be lower than 1 mg/kg
5.10. Cadmium
Quantify the cadmium using the method described in chapter II. The cadmium content must be lower than 1 mg/kg
5.11. Free Glycolate
Quantify the glycolate using the method described below. The carboxymethylcellulose should not contain more than 0.4 % (calculated in sodium glycolate percentage of the anhydrous substance).
5.12. Sodium
Quantify the sodium using the method described in chapter II. The sodium content must be lower than 12.4 % of the anhydrous substance
5.13. Sodium chloride
Quantify the sodium chloride using the method described below. The carboxymethylcellulose must not contain more than 0.5 % of the anhydrous substance.
Annexes
- Loss on drying
1.1. Objective
This test determines the volatile part of carboxymethylcellulose. The result of this test is used to calculate the total solids of the sample and by extension, all the volatile substances at the test temperature are regarded as moisture.
1.2. Interest and use The measurement of water content (by taking account of the purity) is used to measure the quantity of carboxymethylcellulose in commercial products.
1.3. Equipment
1.3.1. Drying oven at 105 °C 3 °C;
1.3.2. Weighing bottle 50 mm in internal diameter and 30 mm in height or equivalent;
1.3.3. Precision balance
1.4. Test
1.4.1. Weigh between 3 and 5 g of sample to ± 1 mg, in a weighing bottle which has already been tared.
1.4.2. Place the weighing bottle without its lid in the drying oven for four hours. Let cool in a desiccator, replace the lid and weigh.
1.4.3. Continue the process until constant weight
1.5. Calculation
1.5.1. Calculate the percentage of the water content M according to the formula:
Where
- A = loss of weight by drying (in g);
- B = initial mass of sample.
- Sodium Glycolate
2.1. Objective
This test covers the determination of sodium glycolate contained in the purified carboxymethylcellose containing not more than 2 % sodium glycolate.
2.2. Summary of the test method
Carboxymethyl cellulose dissolved in acetic acid (50 %), precipitated with acetone in the presence of sodium chloride and the insoluble is eliminated by filtration. The filtrate containing the glycolate sodium (in the form of glycolic acid) is treated to remove the acetone and reacts with 2,7-dihydroxynaphthalene. The resulting colour is measured at 540 nm with a calibrated spectrophotometer using solutions of known concentrations.
2.3. Interest and use
This test method (along with moisture and sodium chloride) is must been used when measuring the quantity of polymer in the substance. It must be used to check the purity of carboxymethylcellulose required by public health regulations.
2.4. Equipment
2.4.1. Spectrophotometer capable of carrying out analysis at 540 nm;
2.4.2. Spectrophotometer cells, 1 cm of optical path
2.4.3. Aluminium paper in squares approximately 50 × 50 mm;
2.4.4. Precision balance
2.5. Reagents
2.5.1. Acetic acid, glacial (purity ≥ 99 %);
2.5.2. Acetone (purity ≥ 99 %);
2.5.3. 2,7-dihydroxynaphtalene solution (0.100 g/L): Dissolve 100 mg 1 mg of 2,7dihydroxynaphthalene (naphthalenediol) in 1 L of sulphuric acid. Before using, allow the solution to stand until the initial yellow colo disappears. If the solution is dark, eliminate it and prepare a new one with a different supply of sulphuric acid. This solution remains stable for one month when stored in a dark bottle;
2.5.4. Standard glycolic acid solution at 1 mg/mL: dry several grams of glycolic acid in a desiccator for at least sixteen hours at room temperature. Weigh 100 mg ± 1 mg, pour into a 100 mL graduated flask, dissolve with water, adjust with water to the filling mark. Do not keep solution longer than 30 days;
2.5.5. Sodium chloride (NaCl, purity ≥ 99 %);
Sulphuric acid concentrate (H2SO4 purity ≥ 98 %, ρ≥ 1.84).
2.6. Preparation of the calibration curve
2.6.1. In a series of five graduated 100 mL volumetric flasks, pour 0, 1, 2, 3 and 4 mL of the glycolic acid reference solution (to 1 mg / mL). Into each flask, add 5 mL of water, then 5 mL of glacial acetic acid, make up with acetone to the filling mark and mix. These flasks contain respectively, 0, 1, 2, 3 and 4 mg of glycolic acid.
2.6.2. Pipet 2 mL of each of these solutions and transfer them into five 25 mL graduated flasks. Evaporate the acetone by heating the open graduated flasks, laid out vertically, in a water bath for exactly 20 min. Remove from the water bath and let cool at room temperature.
2.6.3. Add 5 mL of 0.100 g/L 2,7-dihydroxynaphtalene solution, mix thoroughly, then add an additional 15 mL of 2,7-dihydroxynaphtalene solution and mix. Cover the mouth of the flasks with a small piece of aluminium foil, place the flasks upright in the water bath for 20 min. Remove from the water bath, let cool at room temperature and add sulphuric acid to the filling mark.
2.6.4. Measure the absorbance of each sample at 540 nm against the blank using 1 cm optical depth cells. Plot the absorbance curve according to the corresponding quantity of glycolic acid (in mg) in each flask.
2.7. Test method
2.7.1. Weigh 0.500 g ± 0.001 g of sample and transfer into a 100 mL beaker. Moisten the sample entirely with 5 mL of acetic acid, followed by 5 mL of water, stir with a glass rod until dissolution is complete (usually requires approximately 15 minutes). Slowly add 50 mL of acetone while stirring, then approximately 1 g of sodium sulphate. Continue to stir for several minutes to ensure complete completely precipitation the carboxymethylcellulose.
2.7.2. Filter using a paper filter previously soaked with small amount of acetone, and collect the filtrate in a 100 mL graduated flask. Use 30 mL of acetone to facilitate transfer of solid matter and to wash the filter cake. Make up to the filling mark with acetone and mix.
2.7.3. In another 100 mL graduated flask, prepare a blank with 5 m