ANNEX IISupplementary provisions
ANNEX II Determination of starches or dextrins or other modified starches content in goods of CN codes 3505 20 10 to 3505 20 90 and of amylaceous substances content in goods of CN codes 3809 10 10 to 3809 10 90 I. PRINCIPLE Starch is converted by acid hydrolysis into reducing sugars which are determined by volume using Fehling’s solution. II. APPARATUS AND REAGENTS 1. 250 ml flask; 2. 200 ml graduated flask; 3. 25 ml graduated burette; 4. Hydrochloric acid at 1,19 density; 5. Potassium hydroxide solution; 6. Decolourising charcoal; 7. Fehling’s solution; 8. Methylene blue solution (1 %). III. METHOD Into a 250 ml flask place a sample containing about 1 g of starch. Add 100 ml of distilled water and 2 ml of hydrochloric acid. Bring to the boil and reflux for three hours. Transfer the contents of the flask and rinsings into a 200 ml graduated flask. Cool and nearly neutralise with potassium hydroxide solution. Add distilled water to 200 ml and filter through a little decolourising charcoal. Then pour the solution into a graduated burette and reduce 10 ml of Fehling’s solution by the following method: Into a flat-bottomed flask of about 250 ml pour 10 ml of Fehling’s solution (5 ml of solution A and 5 ml of solution B). Shake until clear and add 40 ml of distilled water and a small quantity of quartz or pumice. Place the flask on a square asbestos plate with a round hole of about 6 cm diameter in the centre, the asbestos in turn resting on a piece of wire gauze. Heat the flask at such a rate that the liquid begins boiling after about two minutes. From the burette, add to the boiling liquid successive quantities of the sugar solution until the blue colour of the Fehling’s solution becomes hardly discernible; then add 2 or 3 drops of methylene blue solution as indicator, and complete the titration by adding further quantities of the sugar solution, drop by drop, until the blue colour of the indicator disappears. For greater accuracy repeat the titration under the same conditions, but adding without a break almost all the sugar solution required to reduce the Fehling’s solution. In this second titration, the reduction of the Fehling’s solution should occur within three minutes. Keep boiling for exactly two further minutes, adding the reagent within one minute drop by drop to the boiling solution until the blue colour disappears. The percentage by weight of starch in the sample is determined by means of the following formula: starch % = ((T × 200 × 100)/(n × p)) × 0,95 where: T : is the quantity in grams of anhydrous dextrose corresponding to 10 ml of Fehling’s solution (5 ml of solution A and 5 ml of solution B). This titer corresponds to 0,04945 g of anhydrous dextrose when solution A contains 17,636 g of copper per litre; n : is the number of ml of the sugar solution used for titration; p : is the weight of the sample amount; 0,95 : is the rate of conversion of anhydrous dextrose into starch. IV. PREPARATION OF FEHLING’S SOLUTIONS Solution A : In a graduated flask dissolve 69,278 g of pure crystallised copper sulphate — Analytical Reagent (CuSO 4 5H 2 O) — free from iron in distilled water and bring the volume to 1 litre with distilled water. The correct strength of this solution must be verified by a quantitative determination of the copper. Solution B : In a graduated flask dissolve 100 g of sodium hydroxide and 346 g of double sodium potassium tartrate (Rochelle salt) in distilled water and bring the volume to 1 litre with distilled water. The two solutions A and B must be mixed in equal quantities immediately before use. 10 ml of Fehling’s solution (5 ml of solution A and 5 ml of solution B) is completely reduced, under the conditions described at section III, by 0,04945 g of anhydrous dextrose.