As described in Section 1.2 of the General Provisions.

Place 3 to 5 grams of the test sample (5.1) into the dish, cover with the lid and weigh to the nearest 0,1 mg (m₁).

The loss of mass on drying of the sample, expressed as a percentage by mass, is given by:

$\frac{m_1{\mathrm{—\; m}}_2}{m_1{\mathrm{—\; m}}_0}\mathrm{100\; where:}$

×

• m₀

  mass, in g of the dish and its lid after process 5.2;

• m₁

  mass, in g of the dish, its lid and the test portion before drying (process 5.3);

• m₂

  mass, in g of the dish, its lid and the test portion after drying (process 5.4.3 or 5.4.4).

Calculate the loss on drying to the nearest 0,01 %.

The difference in results between two determinations carried out simultaneously or in rapid succession on the same sample, by the same analyst, under the same conditions, shall not exceed 0,1 g of moisture per 100 grams of product.

This repeatability interval should be achieved in 95 % of the times that the method is carried out.

Sulphuric acid, concentrated, S₂O 1,84 g/ml.

Potassium sulphate, anhydrous (K₂SO₄).

Copper (II) sulphate pentahydrate (CuSO₄5H₂O).

Sucrose (C₁₂H₂₂O₁₁).

Boric acid, 40-g/l solution.

Sodium hydroxide, concentrated aqueous solution 30 % (m/m), carbonate free.

Hydrochloric acid, 0,1 mol/1.

Mixed indicator. Mix equal volumes of a 2 g/1 solution of methyl red in at least 95 % (V/V) ethanol and a 1 g/1 solution of methylene blue in at least 95 % (V/V) ethanol.

As described in Section 1.2 of the General Provisions.

If the presence of ammonium caseinate or other ammonium compounds is suspected, carry out the following test. Add to 1 gram of sample in a small conical flask 10 ml of water and 100 mg of magnesium oxide. Rinse down any magnesium oxide adhering to the walls and close the flask with a cork stopper, inserting a piece of moistened red litmus paper between the stopper and the neck of the flask. Mix the contents of the flask carefully and heat the flask in a water bath at 60 to 65 ^o C. If the litmus paper colours blue within 15 minutes ammonia is present, and the method is not applicable (see Section 1).

At the same time as the determination of the nitrogen content of the sample perform a blank determination using 0,5 grams of the sucrose (4.4) instead of the test portion, using the same apparatus, the same quantities of all reagents and the same procedure as described in 6.5. If the titration in the blank determination exceeds 0,5 ml of 0,1 mol/1 acid, the reagents shall be checked and the impure reagent or reagents purified or replaced.

Transfer to the Kjeldahl flask (5.2) 0,3 to 0,4 grams of the test sample (6.1), weighed to the nearest 0,1 mg.

The protein content of the sample, expressed as a percentage by mass, is given by:

$\frac{\left(V_1{\mathrm{—\; V}}_2\right)\mathrm{\times \; T\; \times \; 14\; \times \; 100\; \times \; 6,38}}{\mathrm{m\; \times }\mathrm{1\; 000}}=\frac{\mathrm{8,932}\left(V_1{\mathrm{—\; V}}_2\right)\mathrm{\times \; T}}{m}$

where:

• V₁

  is the volume, in millilitres, of the standard volumetric hydrochloric acid solution (4.7) used in the determination (6.5);

• V₂

  is the volume, in millilitres, of the standard volumetric hydrochloric acid solution (4.7) used in the blank test (6.3);

• T

  is the strength of the standard volumetric hydrochloric acid solution (4.7) in mol/1;

• m

  is the mass, in grams, of the test portion.

Calculate the protein content to the nearest 0,1 %.

The difference between the results of two determinations carried out simultaneously or in rapid succession on the same sample, by the same analyst under the same conditions shall not exceed 0,5 grams of protein per 100 grams of product.

This repeatability interval should be achieved in 95 % of the times that the method is correctly carried out.

Sodium hydroxide solution: 0,1 Mol/1.

Phenolphtalein indicator solution, 10 g/1 in ethanol (95 % V/V) neutralized to the indicator.

As described in Section 1.2 of the General Provisions.

Weigh about 10 grams of the test sample (6.1) to the nearest 10 mg and transfer it to the conical flask (5.2).

Using the 250 ml measuring cylinder (5.6), add 200 ml of freshly boiled and cooled water, previously heated to 60 ^o C. Stopper the flask, mix by swirling and place in the water bath at 60 ^o C (5.8) for 30 minutes. Shake the flask at intervals of about 10 minutes.

Filter, and cool the filtrate to about 20 ^oC. The filtrate must be clear.

Transfer 100 ml of the cooled filtrate into the conical flask (5.5), using the pipette (5.3). Add 0,5 ml of the phenolphtalein indicator solution (4.2), using the pipette (5.4). Titrate with the standard volumetric sodium hydroxide solution (4.1), until the appearance of a faint pink colour, persisting for at least 30 seconds. Determine and record the volume used to the nearest 0,01 ml.

The titratable acidity of the acid casein is given by:

$\frac{\mathrm{20\; \times \; V\; \times \; T}}{m}$

where:

• V

  is the volume, in millilitres, of the standard volumetric sodium hydroxide solution (4.1) used;

• T

  is the strength of the standard volumetric sodium hydroxide solution (4.1) in mol/1;

• m

  is the mass, in grams, of the test portion.

Calculate the titratable acidity to two decimal places.

The difference in results between two determinations carried out simultaneously or in rapid succession on the same sample, by the same analyst, under the same conditions, shall not exceed 0,02 ml of 0,1 mol/1 sodium hydroxide per 1 gram of product.

This repeatability interval should be achieved in 95 % of the times that the method is correctly carried out.

Magnesium acetate tetrahydrate solution, 120 g/1. Dissolve 120 grams of magnesium acetate tetrahydrate [Mg (CH₃ CO₂)₂ 4 H₂ O] in water and make up one litre with water.

Analytical balance

One-mark pipette, 5 ml.

Silica or platinum dishes, about 70 mm diameter and 25 to 50 mm deep.

Drying oven, capable of being controlled at 102 ^oC ± 1 ^oC.

Electrical furnace, capable of being controlled at 825 ^oC ± 25 ^oC.

Boiling water bath

Desiccator containing freshly activated silica gel with a water content indicator or an equivalent desiccant.

As described in Section 1.2 of the General Provisions.

Heat two dishes (A,B) (5.3) in the electrical furnace (5.5), controlled at 825 ^o C ± 25 ^o C, for 30 minutes. Allow the dishes to cool somewhat and then place in the desiccator (5.7) to the temperature of the balance room and weigh to the nearest 0,1 mg.

Weigh, to the nearest 0,1 mg approximately 3 grams of the test sample (6.1), directly into one of the prepared dishes (A).

Using the pipette (5.2), add to the dish (A) exactly 5 ml of the magnesium acetate solution (4.1) so as to wet all of the test portion, and allow to stand for 20 minutes.

To the other prepared dish (B), add with the pipette (5.2) exactly 5 ml of the magnesium acetate solution (4.1).

Evaporate the contents of both dishes (A and B) to dryness on the boiling water bath (5.6).

Place both dishes in the oven (5.4), controlled at 102 ^oC ± 1 ^oC, for 30 minutes.

Heat dish A with its contents on a low flame, a hot plate or under an I/R lamp, until the test portion is completely charred, taking care that it does not burst into flame.

Transfer both dishes (A and B) to the electrical furnace (5.5), controlled at 825 ^oC ± 25 ^oC, and heat for at least one hour until all carbon has disappeared from dish A. Allow both dishes to cool somewhat and then place in the desiccator (5.7) to the temperature of the balance room and weigh to the nearest 0,1 mg.

Repeat the operations of heating for approximately 30 minutes, in the electrical furnace (5.5), cooling and weighing, until the mass remains constant to within 1 mg or begins to increase. Record the minimum mass.

The content of ash, including P₂O₅, in the sample, as a percentage by mass, is given by:

$\frac{\left(m_1{\mathrm{—\; m}}_2\right)—\left(m_3{\mathrm{—\; m}}_4\right)}{m}0\mathrm{\times \; 100}$

where:

• m₀

  is the mass, in grams, of the test portion;

• m₁

  is the mass, in grams, of dish A and residue;

• m₂

  is the mass, in grams, of the prepared dish A;

• m₃

  is the mass, in grams, of dish B and residue;

• m₄

  is the mass, in grams, of the prepared dish B.

Calculate the final result to the nearest 0,01 %.

The difference in results between the determinations carried out simultaneously or in rapid succession on the same sample, by the same analyst, under the same conditions, shall not exceed 0,1 grams per 100 grams of product.

The repeatability interval should be achieved in 95 % of the times that the method is correctly carried out.

Analytical balance

Silica or platinum dish, about 70 mm diameter and 25 to 50 mm deep.

Electrical furnace with air circulation, capable of being controlled at 825 ^oC ± 25 ^oC.

Desiccator, containing freshly activated silica gel with a water content indicator or an equivalent desiccant.

As described in Section 1.2 of the General Provisions.

Heat the dish (4.2) in the electrical furnace (4.3), controlled at 825 ^oC ± 25 ^oC, for 30 minutes. Allow the dish to cool somewhat and then place in the desiccator (4.4) to the temperature of the balance room and weigh to the nearest 0,1 mg.

Weigh, to the nearest 0,1 mg approximately 3 grams of the test sample (5.1) directly into the prepared dish.

Heat the dish with its contents on a low flame, a hot plate or an I/R lamp until the test portion is completely charred, taking care that it does not burst into flame.

Transfer the dish to the electrical furnace (4.3), controlled at 825 ^oC ± 25 ^oC, and heat for at least one hour until all carbon has disappeared from the dish. Allow the dish to cool somewhat and then place in the desiccator (4.4) to the temperature of the balance room and weigh to the nearest 0,1 mg.

Repeat the operations of heating for approximately 30 minutes, in the electrical furnace (4.3), cooling and weighing, until the mass remains constant to within 1 mg or begins to increase. Record the minimum mass.

The ash content of the sample, including P₂O₅, as a percentage per mass, is given by:

$\frac{m_1{\mathrm{—\; m}}_2}{m}0\mathrm{\times \; 100}$

where:

• m₀

  is the mass, in grams, of the test portion;

• m₁

  is the mass, in grams, of the dish and residue;

• m₂

  is the mass, in grams, of the prepared dish.

Calculate the final result to the nearest 0,01 %.

The difference between the results of two determinations carried out simultaneously or in rapid succession on the same sample, by the same analyst, under the same conditions, shall not exceed 0,15 grams of ash per 100 grams of product.

This repeatability interval should be achieved in 95 % of the times that the method is correctly carried out.

Two standard buffer solutions with pH values at 20 ^oC which are known to the second decimal place and will bracket the pH value of the sample under test, for example phtalate buffer solution of pH approximately 4 and a borax buffer solution of pH approximately 9.

Balance, accuracy 0,1 grams.

pH meter, minimum sensitivity 0,05 pH unit, with a suitably calibrated elecrode, e.g. glass electrode and a calomel or other reference electrode.

Thermometer, accuracy 0,5 ^oC.

Conical flask, capacity 100 ml, fitted with a ground glass stopper.

Beaker, capacity 50 ml.

As described in Section 1.2 of the General Provisions.

Record, as the pH of the aqueous solution of caseinate, the value read from the dial of the pH meter to at least two decimal places.

The difference between the results of two determinations carried out simultaneously or in rapid succession on the same sample, by the same analyst, under the same conditions, shall not exceed 0,05 pH unit.

This repeatability interval should be achieved in 95 % of the times that the method is correctly carried out.