::: MCP 200/300/500 Application Note

Determination of Starch Polarimetric method

Starch Feeding Stuff:::

::: This methods is suitable for determination of starch and high molecular weight starch degradation products in feeding stuff in compliance with the EC Commission Directive 86/174/EEC and Council Directive 96/25/EC. General
Starch is one of the most abundant substances in nature. It is a renewable resource, which is biological decomposable. Starch is a natural product, which is obtained from seeds or roots, like corn, wheat, or potatoes. Starch is a widely spread content, which is found in many products: paper, textile and photo products, glues, colours, concrete, gypsum, adhesives, plastics, food like, soups, sauces, puddings, bakery, confectionary, mayonnaises, crmes, cereal products, as well as in beverages, sweets, feedstuff, but also in cosmetics like lipsticks or facial powders, pharmaceuticals like salves, crmes, pills or in packing materials and many other products.

Safety Precautions
This method does not contain any safety instructions, if any, associated with its use. It is in the responsibility of the user of this method to establish appropriate safety and health practices and determine the applicability of regulatory limitations prior to its use.

Reagents:
25% (w/w) hydrochloric acid, density= 1.126 g/ml 1.128% (w/w) hydrochloric acid. Check the concentration by titration with sodium hydroxide solution ((0.1 mol/l), indicator methyl red (0.1% (w/v)) in 94% (v/v) ethanol. 10 ml=30.94 ml of NaOH 0.1 mol/l Carrez solution I: 21.9 g zinc acetate (Zn (CH3COOH)22H2O 3 g glacial acetic acid make up with water to 100 ml volume Carrez solution II 10.6 g potassium ferrocyanide K4[Fe(CN)6]3H2O in 100 ml water 40% (v/v) ethanol, density 0.948 g/ml at 20 C

Principle
The polarimetric determination of starch content is based on the optically activity of starch. Due to the fact, that starch can not be dissolved with water, hydrochloric acid is used. After dissolution the sample needs to be clarified (precipitation reagent) filtrated, and measured in a polarimeter. An additional blind test has to be performed in order to eliminate the influence of other optically active substances may being present. For the blind test the starch is precipitated, the solution is filtrated and also measured in the polarimeter. From the difference between the two measurements, the starch content can be calculated. Starch has an extremely high specific rotation (>180 , see below). Therefore, also small starch contents can be measured.

Apparatus
MCP 200/300/500 with 200 mm polarimeter tube with Peltier option. Required accuracy 0.01 optical rotation. 250 ml Erlenmeyer flask wit standard ground glass joint and with reflux condenser.

Procedure
Crush the sample finely at least to a particle size that pass trough a 0.5 mm round meshed sieve.

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::: MCP 200/300/500 Application Note

Determination of Starch Polarimetric method

Dairy Products Milk :::

Determination of total optical rotation 2.5 g of crushed sample (weighting accuracy 1 mg) are put into a 100 l volumetric flask. 25 ml of 1.128 % hydrochloric acid are added. The flask is shaked well to achieve an even distribution of the test sample. Now another 25 ml of 1.128% hydrochloric acid is added. The flask is put into a boiling water bath and shaked well and continuous for the first 3 minutes to avoid formation of agglomerates. The amount of water contained in the water bath must be sufficient to keep the water boiling, when the flask is introduced to it. During shaking the flask is not allowed to be taken out of the water bath. Exactly after 15 minutes the flask has to be taken from the water bath, 30 ml of cold water are added and cooled immediately down to 20 C. Now add 5 ml Carrez I solution and shake for one minute. Than 5 ml Carrez solution II are added and again shaked for one minute. Add water up to volume, shake and filtrate. If the filtrate is not clear (in seldom cases) the procedure needs to be repeated by using larger amounts of Carrez solutions, e.g. 10 ml. Measure the optical rotation of the filtrate in the MCP at 20 C using a sample cell of 200 mm optical path length. Determination of optical rotation of substances soluble in ethanol 5 g of crushed sample (weighting accuracy 1 mg) are put into a 100 ml volumetric flask and approx. 80 ml ethanol (40%) are added. Now the flask is left for 1 hour at room temperature. During that time, the flask is shaked vigorously six times to make sure, that the sample is mixed with the ethanol. Afterwards, fill up to volume with ethanol (40%), shake and filtrate. 50 ml of the filtrate (2.5 g sample) are transferred into a 250 ml Erlenmeyer flask and 2.1 ml hydrochloric acid (25%) are added. The flask is shaked well, connected to a reflux condenser and put into a boiling water bath. Exactly after 15 minutes the flask is taken from the water bath, transferred to 100 ml graduated flask, and

rinsed with some cold water. Afterwards, it is cooled down to a temperature of 20 C. Now the sample is clarified with Carrez solution I and II, water is added up to volume. After shaking, the sample is filtrated and its optical rotation is measured at 20 C by using a sample cell of 200 mm optical path length. It is recommended to check, if there is still some starch in the filtrate (Iodine test). Calculation of results The starch content (%) is calculated by following formula:

Starch content % =

2000 ( P P' ) [ ]20 D

P = Total optical rotation in angular degrees. P = Optical rotation in angular degrees of the substances soluble in 40% (V/V) ethanol.

[ ]20 , measD = Specific rotation of pure starch at 20 ured in a polarimeter with the wavelength of 589nm (sodium D-line).
The above formula is only valid for the mentioned masses and dilutions, otherwise corresponding corrections need to be applied. The following specific rotations are conventionally accepted: + 185.9rice starch + 185.4potato starch + 184.6maize starch + 182.7 wheat starch + 181.5barley starch + 181.3oat starch + 184.0 other types of starch and starch mixtures in compound feeding stuff.

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::: MCP 200/300/500 Application Note

Determination of Starch Polarimetric method

Dairy Products Milk :::

Limitations If the following feed material is present in significant amounts in the feeding stuffs, it could result into incorrect results: beet (sugar) products, such as (sugar) beet pulp, (sugar) beet molasses, (sugar) beet pulp molasses, (sugar) beet vinasse, (beet) sugar, Citrus pulp, Linseed, linseed expeller, linseed extracted, Rape seed, rape seed expeller, rape seed extracted, rape seed hulls, Sunflower seed, sunflower seed extracted, sunflower seed, partially decorticated, extracted, Copra expeller, copra extracted, Potato pulp, Dehydrated yeast, Products rich in inulin, Greaves.

Notes
This procedure is also be used (maybe with minor modifications) for the determination of starch content of other samples, e.g. food, corn, potatoes, etc..

Literature
Official Journal of the European Communities. Commission Directive 1999/79/EC (1999). Native starch Determination of starch content Ewers polarimetric method, International Standard, ISO 10520:1997(E). Untersuchung von Lebensmitteln Bestimmung des Strkegehalts in Brot einschlielich Kleingebck aus Brotteigen, Amtliche Sammlung von Untersuchungsverfahren nach $35 LMBG, BVL L 17.00-5 (2003). Untersuchung von Lebensmitteln Bestimmung des Strkegehalts Feinen Backwaren, Amtliche Sammlung von Untersuchungsverfahren nach $35 LMBG, BVL L 18.00-6 (2003). Ewers, E., Zeitschrift fr ffentliche Chemie 14, S. 150-157, (1908). "Feedstuffs Analysis Procedure for Starch" (G-28), Standard Analytical Methods of the Member Companies of the Corn Refiners Association, Inc., 2. Edition, 15. April 1986. Starch in Flour, Polarization method, AOAC Official Method 945.37, First Action 1945 Starch Determination in Cereal Products Using the Modified Evers Polarimetric Method, U.S. Customs Laboratory Methods, USCL Method 11-02

If the sample contains more than 6% of carbonates (calculated in forms of calcium carbonate) they must be removed by treatment with an exactly appropriate amount of diluted sulphuric acid before measurement of total optical rotation. In case of high lactose content (e.g. powdered milk serum or skimmed milk powder), fit a reflux condenser to the flask, after the 80 ml of ethanol are added and put it on a water bath at 50 C for 30 minutes. Leave to cool and proceed as described above.

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