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Teacher Notes![]() Milk Is a Natural: Biology, Chemistry and NutritionStudent Laboratory KitMaterials Included In Kit
Acetic acid solution, CH3COOH, 1 M, 50 mL
Benedict’s solution, 200 mL Biuret solution, 200 mL Carbohydrate reference solution, dextrose, 50 mL* Ethyl alcohol, 95%, 50 mL Protein reference solution, albumin, 50 mL* Sodium hydroxide solution, NaOH, 1 M, 100 mL Beral-type pipets, graduated, 105 *See Prelab Preparation. Additional Materials Required
Beaker, 400- or 600-mL
Crucible tongs Erlenmeyer flasks, 125-mL, 2 Filter paper, 12.5-cm, 2 pieces Funnel Graduated cylinder, 25- or 50-mL Hot plate Hot vessel gripping device, “hot hands” Paper towels Skim milk, 22 mL Stirring rod Test tubes, medium-size, 7 Test tube holder Test tube rack Thermometer Watch glasses, 3 Prelab PreparationThe carbohydrate reference solution (dextrose) and protein reference solution (albumin) must be freshly prepared. Add 50 mL of distilled water to the bottle. Cap and shake to mix. Safety PrecautionsAcetic acid and sodium hydroxide solutions are corrosive liquids. Ethyl alcohol is a flammable organic solvent and a dangerous fire risk. Benedict’s solution contains copper tartrate and is an alkaline solution. Biuret solution is a highly alkaline solution and is corrosive to eyes and body tissue. Wear chemical splash goggles, chemical-resistant gloves and a chemical-resistant apron. Avoid exposure to eyes and skin. Please consult current Safety Data Sheets for additional safety, handling and disposal information. DisposalPlease consult your current Flinn Scientific Catalog/Reference Manual for general guidelines and specific procedures, and review all federal, state and local regulations that may apply, before proceeding. All solutions from this activity can be disposed of according to Flinn Suggested Disposal Method #26b. Teacher Tips
Correlation to Next Generation Science Standards (NGSS)†Science & Engineering PracticesPlanning and carrying out investigationsAnalyzing and interpreting data Using mathematics and computational thinking Developing and using models Disciplinary Core IdeasMS-PS1.A: Structure and Properties of MatterMS-PS1.B: Chemical Reactions HS-PS1.A: Structure and Properties of Matter HS-PS2.B: Types of Interactions HS-LS1.C: Organization for Matter and Energy Flow in Organisms Crosscutting ConceptsScale, proportion, and quantityStructure and function Systems and system models Performance ExpectationsMS-PS1-2: Analyze and interpret data on the properties of substances before and after the substances interact to determine if a chemical reaction has occurred. Answers to Prelab Questions
Sample Data{11938_Data_Table_1_Protein and carbohydrate content of skim milk}
{11938_Data_Table_2_Qualitative tests for protein and carbohydrate}
Answers to Questions
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Student Pages
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Student Pages![]() Milk Is a Natural: Biology, Chemistry and NutritionIntroductionFrom glossy magazine ads to flashy billboards, the message to drink more milk is all around us. Whole milk is a natural, nutritionally complete food source. It contains all of the essential classes of biological molecules—proteins, carbohydrates and fats—that are important in nutrition. The purpose of this lab is to separate the protein and carbohydrate components of skim milk and verify their identity. Concepts
BackgroundMilk and milk products have been a major food source from earliest recorded history. The principal components of whole milk are proteins (3.5%), carbohydrates (5%) and fat (4%). In addition, milk is also an important source of a variety of essential minerals and vitamins in the diet. In this lab activity, the biochemical nature of proteins and carbohydrates is explored by analyzing the nutritional components of skim milk (milk without the fat component). {11938_Background_Figure_1}
The presence of proteins can be detected using a simple color test based on the reaction of protein molecules with copper ions in basic solution (pH >13). When a protein is allowed to react with CuSO4 in the presence of a strong base, the Cu2+ ions bind to the nitrogen atoms and carbonyl groups in the protein to form a distinctive and stable purple coordination complex. This is called the Biuret test. There are three main proteins in milk: casein, lactalbumin, and lactoglobulin(s). The chief nutritional protein in milk is casein, comprising 80% of the total protein content; it basically serves as a reservoir of amino acids for the body to synthesize its needed proteins. Casein is a phosphoprotein—it contains a large number of phosphate groups attached to the amino acid side chains in the polymer structure. The negatively charged phosphate groups are balanced by positive calcium ions and are thus responsible for the high nutritional calcium content in milk. Because casein is almost completely insoluble in water, it can be easily precipitated from milk by the addition of a small amount of acid at 40 °C. The second group of proteins in milk is made up of smaller proteins that are much more water soluble. There are two main proteins in this group—lactalbumin and lactoglobulin. Lactalbumin is an enzyme that is involved in the synthesis of lactose, the principal carbohydrate component in milk. Lactoglobulin is a generic name for a group of proteins that are responsible for the immunological properties of milk. These proteins have a role far greater than their low quantity suggests, since they constitute the “protective” function of milk. Lactalbumin and lactoglobulin remain in solution when the casein is precipitated from milk upon treatment with acid, but can be forced out of solution by heating milk to higher temperatures (80–90 °C). Carbohydrates Carbohydrates constitute the second class of biochemical and nutritional components in milk. Monosaccharides such as glucose (“blood sugar,” also called dextrose) and fructose are simple carbohydrates and are the fundamental units that make up all other carbohydrate molecules. When two monosaccharide units are joined together, they form disaccharides. Examples of disaccharides include sucrose (cane or table sugar), maltose (malt sugar) and lactose (milk sugar). Polysaccharides are complex carbohydrate polymers composed of many simple sugar molecules joined together. The most familiar polysaccharides are starch and cellulose. The monosaccharide glucose is the principal energy source for metabolism, while the polysaccharide starch serves as a way of storing chemical energy in plants. {11938_Background_Figure_2}
The principal carbohydrate present in milk is lactose, a disaccharide composed of one unit of glucose combined with one unit of galactose. Lactose can be separated from milk by crystallization via prolonged heating to remove most of the water content. The filtrate remaining after the protein and fat components have been removed is heated at 90 °C until the liquid has evaporated. Lactose is isolated as a white powder and is identified by means of Benedict’s test, a general test for “reducing” sugars. Reducing sugars include lactose and maltose (disaccharides) as well as all monosaccharides (such as glucose and galactose). Benedict’s test is based on an oxidation–reduction reaction between the sugar molecule and Cu2+ ion in Benedict’s reagent. The intense blue color of Benedict’s solution fades during a positive test result, which is marked by the formation of an orange-red precipitate consisting of copper(I) oxide (Cu2O). Fats The third class of nutrients in whole milk are fats and oils. Fats and oils are water-insoluble substances found in both plant and animal cells. They can be separated from plant and animal products using nonpolar organic solvents, such as hexane. Fats and oils are members of a diverse class of nonpolar biological molecules called lipids. Other examples of lipids include phospholipids, cholesterol, steroid hormones, and fat-soluble vitamins such as Vitamin A and D. Despite their current political “incorrectness,” fats and oils are essential nutrients that are required for cell membrane structure and function. Fats and oils are also a very efficient way of storing chemical energy and are thus the most important means of long-term energy storage in cells. Whole milk is an emulsion, in which the fat molecules are dispersed in the form of tiny fat globules. The emulsion is stabilized by the presence of protein molecules that adhere to the surface of the fat and prevent it from coalescing and separating out of solution. The fat can be removed from whole milk by extraction with hexane or a similar organic solvent. In skim milk, the fat content has been removed from whole milk by centrifuging it. Fat is less dense than water. When milk is spun very fast in a centrifuge, the fat portion separates out as an upper creamy layer. The upper fat layer is then “skimmed” off—skim milk is the more dense portion that remains behind in a lower layer. Overview of the Milk Experiments This lab activity involves the separation, identification and quantitative analysis of the protein and carbohydrate fractions of skim milk. Skim milk is used rather than whole milk since it lacks fat and thus makes it easier to obtain pure protein and carbohydrate samples. The experimental results will be compared against the information provided on the nutritional label for the amount of protein and carbohydrate in milk. The principal milk protein casein is removed from skim milk by precipitation with mild acid. Dilute acetic acid is added and the solution is heated gently. Casein is a protein that normally contains many negatively charged phosphate groups surrounded by calcium ions in solution. It is this property that keeps the protein “dissolved” or suspended in milk. As the pH of milk is reduced below 5.0, the phosphate groups lose their negative charge and the calcium ions are replaced by bound hydrogen ions from the added acid. The stable protein suspension is broken and casein precipitates from solution. After drying overnight the protein is isolated in the form of a white solid. After removal of the main insoluble protein fraction, the filtrate is heated to a higher temperature (80–90 °C) and the second class of milk proteins (lactalbumin and lactoglobulin) precipitates out of solution. The combined mass of the protein fractions can be used to calculate the percent protein composition in milk. The identity of the protein fractions of milk is confirmed by means of the Biuret test and compared against a reference protein solution (egg albumin). The milk “whey” filtrate remaining after the proteins have been removed is tested with Benedict’s solution to identify the carbohydrates in milk. In an optional exercise, a small portion of the filtrate can be concentrated to dryness by heating for 20–30 minutes. Lactose is isolated as a white solid, and its mass can be used to calculate the percent carbohydrate (sugars) in milk. The steps involved in characterizing the nutritional components of milk are illustrated by means of a flow chart diagram in Figure 3. {11938_Background_Figure_3}
Materials
Acetic acid solution, CH3COOH, 1 M, 2 mL
Benedict’s solution, 12 mL Biuret solution, 12 mL Carbohydrate reference solution, dextrose, 2 mL Ethyl alcohol, 95%, 5 mL Protein reference solution, albumin, 2 mL Sodium hydroxide solution, NaOH, 1 M, 5 mL Beaker, 400- or 600-mL (for hot water bath) Crucible tongs Erlenmeyer flasks, 125-mL, 2 Filter paper, 2 Funnel Graduated cylinder, 25- or 50-mL Hot plate Hot vessel gripping device, such as “hot hands” Paper towels Pipets, Beral-type, graduated, 7 Skim milk, 22 mL Stirring rod Test tubes, medium-size, 7 Test-tube holder Test-tube rack Thermometer Watch glasses, 3 Prelab QuestionsRead the background information and answer the following questions on a separate sheet of paper.
Safety PrecautionsAcetic acid and sodium hydroxide solutions are corrosive liquids. Ethyl alcohol is a flammable organic solvent and a dangerous fire risk. Benedict’s solution contains copper tartrate and is an alkaline solution. Biuret solution is a highly alkaline solution and is corrosive to eyes and body tissue. Avoid exposure of all chemicals to eyes and skin. Wear chemical splash goggles, chemical-resistant gloves and a chemical-resistant apron. Wash hands thoroughly with soap and water before leaving the laboratory. ProcedurePart A. Isolation of Casein
Part D. Qualitative Tests for Protein and Carbohydrate
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