Analysis of Chemical Components of Milk Lab Report

Objective:

The first goal was to gain experience in the separation of natural product. The second

goal was to use basic biochemical qualitative analytical tests. The third goal of the experiment

was to identify the major components of milk.

Introduction:

Milk is an important source of food for many animals and humans. Milk needs to consist

of several different important components to be sufficient enough for certain mammals.

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Proteins, carbohydrates, fat, vitamins, and minerals are the major components of milk. The

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characteristics of the components are important in determining how to separate them from one

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another. Likes dissolve likes; therefore hydrophobic components are dissolved in hydrophobic
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solvents and vice versa for hydrophilic components. “In general, polar liquids are miscible with

other polar liquids but are not miscible with nonpolar liquids,” (Tro 2014: p. 491). Gravity
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filtration and vacuum filtration were used to separate precipitates from a solution.
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Experimental Procedure:

The chemicals used in the experiment included: whole milk, 10% acetic acid, ethyl
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acetate, 1% glucose solution, Biuret’s test solution (0.5% copper sulfate), 10% sodium

hydroxide, lactase, lactose, glucose test solution, 0.1 M ammonium oxalate solution, and 0.5
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ammonium molybdate solution. The materials used in the experiment included: coffee filters,
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funnels, beakers, flasks, test tubes, thermometer, hot plate, watch glass, Pasteur pipettes, rubber

policeman, boiling chip, labeling tape, small mortar and pestle, and 4 glucose urinalysis test
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strips.

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 The beginning of the experiment included heating a large beaker of water. The first

separation called for 100 mL of milk to be heated in a 250 mL in the hot water bath to 40 degrees

Celsius. Once the solution reached 40 degrees Celsius, 11.0 mL of 10% acetic acid solution was

added to the milk and stirred constantly for a minute. By lowering the pH of the milk, curds

formed. Next using a couple coffee filters, the curds were separated from the whey liquid by

gravity filtration. The curds were then transferred to a Buchner funnel with a vacuum filter and

rinsed with 10 mL of distilled water. After the curds were vacuumed for five minutes, the filtrate

from the gravity filtration and the filtrate from the first separation were added together. The

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curds were left to air dry during the rest of the experiment.

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The next step included the second separation. The whey liquid was then heated close to

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boiling to precipitate the whey proteins out. Once the precipitate formed, vacuum filtration was
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used to separate the whey proteins. Both the solid and the liquid were saved, and the solid air-

dried during the rest of the experiment. The qualitative tests for carbohydrates, lactose, calcium,
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and phosphates were performed on the whey liquid.

Fehling’s test for carbohydrates included adding five mL of Fehling’s reagent to one test

tube and adding 5 drops of 1% glucose to the second test tube. After boiling the second test tube
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for five minutes, a brick-red solution indicated a reaction forming copper oxide. The test for

lactose included preparing two test tubes, adding 0.5 mL of distilled water and a spatula tip of
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lactose to both. Lactase was then grinded with a mortar and pestle, and added to the first test
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tube. Lastly, two glucose urinalysis strips were dipped into both test tubes. The test for calcium

included adding one mL of calcium solution to a test tube and then adding one mL of 0.1 M of
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ammonium oxalate. The test for phosphates included adding two mL of 6 M of nitric acid to two

mL of the phosphate solution in a test tube. Next, two mL of 0.5-ammonium molybdate solution

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 was added to the test tube and heated in a hot bath for five minutes. The precipitate of

ammonium phosphomolybdate indicated the presence of phosphate.

The third separation took place in the hood to separate the casein and the butterfat in the

curds. The curds were placed in a 100 mL beaker and 30 mL of ethyl acetate were added to

extract the curd. Once the curds were ground as much as possible, the solution was gravity

filtrated. The rubber policeman was used to save as much casein as possible. The filtrate was

then gathered in an Erlenmeyer flask with a few boiling chips, and heated in a hot bath to

evaporate the ethyl acetate from the solution while stirring constantly. Once the ethyl acetate

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evaporated completely, the flask was placed in an ice water bath for ten minutes. A residue

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formed, and a sample of the residue placed in water to test the solubility.

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The test for protein in the caseins and whey protein was then performed. 2.0 mL of BSA
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protein test solution, five drops of 10% sodium hydroxide, and two drops of 0.5% copper sulfate

were mixed in a small tube. The development of a light violet color in the solution indicates the
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presence of protein. Once the experiment was completed, negative tests were tested on the
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caseins, whey protein, and whey liquid. The Biuret test of proteins was performed on the whey

liquid. The test for carbohydrates was performed on the whey and casein proteins.
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Results:

The first qualitative tests were tested on the whey liquid. The test for phosphate was
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positive. The solution turned yellow and a precipitate formed. Fehling’s test for carbohydrates
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was positive. The solution turned a brick red quickly after being heated. The lactose test was

positive as well. The glucose urinalysis strip turned a brown indicating 30 mmal per liter. The
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negative test used for the whey liquid was the Biuret test for proteins. The test was negative.

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 The Biuret test for proteins was used to test on the whey and casein proteins. The test for

both proteins was positive. The solution containing the whey protein turned a violet, and the

casein protein turned a very deep purple. The negative test used for the proteins was the test for

carbohydrates. The presence of a brick-red solution didn’t form indicating the test was negative.

The butterfat tested in the water was found to be insoluble. The residue of butterfat was a

yellowish white.

Discussion:

In the experiment it was important to understand why certain compounds mixed with

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other solvents and not others. Compounds with like properties the solvent used have a higher

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chance of dissolving. For example, a hydrocarbon does not dissolve in water because a

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hydrocarbon is nonpolar while water is polar. A polar compound doesn’t share electrons equally,
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and a nonpolar compound does share electrons evenly. The separations in this experiment

produced two solids, a liquid, and a residue. The qualitative tests performed proved that the
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components were separated completely.

The main components of milk are protein, carbohydrates, calcium, phosphate, and fat.

There are two types of proteins in milk: caseins and whey. Caseins are water-soluble and can be
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separated by lowering the pH. Whey proteins are soluble even after lowing the pH and are

insoluble when denatured. Denaturation occurs when the proteins are heated causing them to
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unfold into extended chains that causes it to lose its native shape (Tro 2014: p. 82). Butterfat is
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separated by lowering the pH as well, and is precipitated with the caseins. This precipitate is

known as curd. Butterfat contains long hydrocarbon chains that cause it to be hydrophobic, and
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therefore can be dissolved in a nonpolar solvent.

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 The whey liquid leftover after the curd is separated contains the other major components

of milk: whey protein, carbohydrates, vitamins, and minerals. Carbohydrates are sugars, and the

carbohydrates in milk contain lactose, which is a disaccharide Vitamin A, B, and D are the

vitamins present in milk. Vitamin B is water-soluble. Vitamins A and D are fat-soluble. The

minerals phosphate and calcium present in milk are basic elements of bone and tooth enamel.

Conclusion:

Due to the fact that four samples were collected after the separation, experience was

gained in the natural product separation. The four samples were casein protein, butterfat, whey

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protein, and liquid protein. The qualitative tests helped to prove the essential minerals and

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vitamins the milk contains as well.

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References

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 1) Tro, Nivaldo J. Chemistry: A Molecular Approach. Third ed. Pearson Education, 2014.

82,491. Print.

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