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    Home > Chemicals Industry > Chemical Technology > Determination of protein in soy milk beverage (2)

    Determination of protein in soy milk beverage (2)

    • Last Update: 2021-07-18
    • Source: Internet
    • Author: User
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    Safety reminder

    Because the temperature of the digestion process can reach 400℃, care should be taken to avoid scalds during operation; the digestion process will produce toxic gases such as sulfur dioxide , so it must be carried out in a fume hood, and the laboratory must be well ventilated to avoid poisoning; after digestion, Do not put the nitrogen bottle directly on the test bench.
    First, the high temperature will burn the test bench; second, the nitrogen bottle will burst when it is cold.
    Put the nitrogen bottle on the wooden bracket until it cools to room temperature


    Operation steps

    (1) Digestion Accurately weigh 10-25g of soy milk beverage (about 30-40mg nitrogen), carefully transfer it into a dry and clean 100mL or 250mL Kjeldahl flask, add 0.
    2g of
    copper sulfate , 6g potassium sulfate and concentrated sulfuric acid 20mL, shake gently, install the digestion device according to Figure 3-6, and diagonally support it on the asbestos net with small holes at an angle of 45°
    Use an electric furnace to heat it with a small fire.
    After all the contents are carbonized and the foam stops producing, increase the firepower to keep the liquid in the bottle slightly boiling until the liquid turns blue-green and clear and transparent, and then continue heating for 0.

    Remove and cool, carefully add 20mL of distilled water, after leaving to cool, transfer to a 100mL volumetric flask, rinse the ammonia flask three times with a small amount of water, and transfer the lotion to the volumetric flask to a constant volume


    (2) Distillation Connect the Kjeldahl bottle according to the distillation device in Figure 3-7, plug the mouth of the bottle tightly, and insert the lower end of the condenser into the liquid surface of the absorption bottle (the bottle is pre-filled with 10 mL of 20g/L boric acid solution and mixed indicator 2 ~3 drops)
    Accurately draw 2.
    0 mL of sample treatment solution from the small glass cup into the reaction chamber, wash the small glass cup with 10 mL of water and make it flow into the reaction chamber, and then close the rod-shaped glass stopper

    Pour 10 mL of 400g/L
    sodium hydroxide solution into the small glass cup, lift the glass stopper to slowly flow into the reaction chamber, immediately cover the glass stopper tightly, and add water to the small glass cup to prevent air leakage
    Clamp the screw clamp and start the distillation


    Figure 3-7 Diagram of Nitrogen Distillation Unit

    1- Electric furnace 2- Steam generator (2L flask) 3- Screw clamp

    4-Small glass cup and rod-shaped glass stopper 5-Reaction chamber 6-Reaction chamber

    7-Rubber tube and screw clamp 8-condensation tube 9-distillate receiving bottle

    After distilling for 10 minutes, move the distillate receiving bottle, and the liquid level will leave the lower end of the condenser, and distill for another 1 minute
    Then rinse the outside of the lower end of the condenser tube with a small amount of water, and remove the distillate receiving bottle


    (3) Titration: Titrate to the end point with a standard titration solution of sulfuric acid or hydrochloric acid
    Among them, 2 parts of methyl red ethanol solution and 1 part of methylene blue ethanol solution indicator, the color changes from purple-red to gray, pH 5.
    4; 1 part of methyl red ethanol solution and 5 parts of bromocresol green ethanol solution indicator, the color is changed from The wine red turns to green, and the pH is 5.

    At the same time, make a reagent blank


    Result calculation

    Where x——the protein content in the sample, g/100g

    c——Concentration of HCI standard solution, mol/L

    V 1 ——Volume of hydrochloric acid standard solution consumed when titrating sample absorption solution, mL

    V 2 ——Volume of hydrochloric acid standard solution consumed when titrating blank absorption solution, mL

    V 3 ——The volume of the aspirated liquid, mL

    m——sample mass, g

    The millimolar mass of 1/2N
    2 , g/mmol

    F——The coefficient of nitrogen conversion to protein
    General food is 6.
    25; pure milk and pure dairy products are 6.
    38; flour is 5.
    70; corn and sorghum are 6.
    24; peanuts are 5.
    46; rice is 5.
    95; soybeans and their crude processed products are 5.
    71; soy protein products are 6.
    25; meat and meat The product is 6.
    25; barley, millet, oats, rye is 5.
    83; sesame, sunflower is 5.
    30; compound formula food is 6.
    25, expressed by the arithmetic mean of two independent determination results obtained under repeatable conditions, when the protein content is ≥1g/100g , The result retains three significant digits; when the protein content is less than 1g/100g, the result retains two significant digits


    The absolute difference between two independent determination results obtained under repeatability conditions shall not exceed 10% of the arithmetic mean


    (1) This method does not apply to the determination of food containing inorganic nitrogen-containing substances and organic non-protein nitrogen-containing substances

    (2) The reagent solution used in this method should be prepared with ammonia-free distilled water

    (3) Precautions during digestion

    ①Do not use strong fire for digestion.
    Keep boiling gently.
    Pay attention to turning the Kjeldahl flask constantly to wash off the solid residue attached to the wall of the flask with the condensed acid and promote its complete digestion


    ② If the sample contains a lot of fat or sugar, a large amount of foam is likely to be produced during the digestion process.
    To prevent the foam from overflowing out of the bottle, apply a small fire to heat it and shake it constantly; or add a small amount of octanol or liquid paraffin or silicone oil to dissipate.
    Foaming agent, and also pay attention to control the intensity of the heat source


    ③When the sample digestion solution is not easy to be clear and transparent, cool the Kjeldahl flask, add 2~3mL of 30% hydrogen peroxide , and continue heating and digestion

    ④Generally, after digestion until it becomes transparent, continue digestion for 30 minutes, but for samples containing nitrogen compounds that are particularly difficult to ammoniated, such as lysine, histidine, tryptophan, tyrosine or proline, etc.
    , Need to extend the digestion time appropriately

    If the organic matter is completely decomposed, the digestive juice will be blue or light green, but when the iron content is high, it will be darker green


    ⑤In the digestion reaction, in order to accelerate the decomposition of protein and shorten the digestion time, substances such as potassium sulfate and copper sulfate are often added
    potassium sulfate added functions to raise the boiling point of the solution, to accelerate the role of organic matter decomposition
    Copper sulfate acts as a catalyst and can indicate the completion of the digestion reaction

    There are many types of catalysts that can be used in the Kjeldahl method.
    In addition to copper sulfate, there are also mercury oxide, mercury, selenium powder, etc.
    However, considering the effect, price and environmental pollution and other factors, the most widely used is
    copper sulfate
    A small amount
    of hydrogen peroxide , potassium hypochlorite, etc.
    often added as an oxidant to accelerate the oxidative decomposition of organic matter


    (4) Precautions during distillation

    ①The distillation device must not leak gas

    ② If the amount of alkali added before distillation is insufficient, the digestive solution will be blue and no copper hydroxide will be precipitated.
    At this time, the
    amount of
    sodium hydroxide should be increased

    ③After the distillation is completed, the lower end of the condenser should be lifted from the liquid surface to clean the nozzle, and then steam for 1 min before turning off the heat source, otherwise the absorption liquid may be sucked back

    ④The temperature of the boric acid absorption solution should not exceed 40°C, otherwise the absorption of ammonia will weaken and cause loss.
    At this time, it can be used in a cold water bath


    ⑤When the sampling amount is large, if the dry sample exceeds 5g, the amount of sulfuric acid can be increased according to the proportion of 5mL per 1g sample

    ⑥The mixed indicator is green in alkaline solution, gray in neutral solution, and red in acid solution

    Related Links: Determination of Protein in Soy Milk Beverage (1)



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