LAS 3 - Proteins Part 1 PDF

PROTEINS ISOLATION OF PROTEIN Isolation of Gluten - Crude gluten - is a mixture of proteins extracted from wheat flour. It's primarily composed of two proteins: glutenin and gliadin. These proteins are responsible for the elastic proper...

PROTEINS ISOLATION OF PROTEIN Isolation of Gluten - Crude gluten - is a mixture of proteins extracted from wheat flour. It's primarily composed of two proteins: glutenin and gliadin. These proteins are responsible for the elastic properties of dough, allowing it to be stretched and molded. Isolation of Bean Protein - 1M Acetic Acid - The one who precipitates the bean protein from the mixture. Isolation of Myoglobin from Muscle Isolation of Albumin from Egg White POST LABORATORY QUESTIONS Isolation of Gluten 1. State the relationship of iodine test and complete removal of starch - The iodine test is used to determine the presence of starch. The starch reacts with iodine to produce a blue-black color. Therefore, when the solution is no longer blue-black when tested on the wash water of the dough, it signifies that all starch has been completely washed out. 2. How does the amount of the insoluble material left compare with the amount of the original sample used? - The amount of insoluble material left is significantly less than the original amount of wheat flour. This is because the majority of the wheat flour is composed of starch and starch is soluble in water and could be washed away during the process. The gluten which is insoluble in water remains as a residue. 3. What composes crude gluten? - Crude gluten is primarily composed of two proteins: glutenin and gliadin. These proteins are responsible for the elastic properties of dough, allowing it to be stretched and molded. Isolation of Bean Protein 1. State the purpose of soaking the beans in water - Soaking the beans in water hydrates the beans, softening it making it easier to grind. This process also helps activate enzymes in the beans which can break down complex carbohydrates making protein extraction more efficient. 2. What are the predominant proteins present in beans? - The predominant proteins in beans are legumins and globulins. These are storage proteins that provide a rich source of amino acids. 3. How are these bean proteins precipitated by an acid? - Bean proteins are precipitated by acid due to the isoelectric point (pI) of the proteins. The pI is the pH at which a protein has no net electrical charge. When the pH of the solution is adjusted to the pI of the protein, the solubility of the  protein decreases, leading to precipitation. Acetic acid, being an acid, lowers the pH of the solution, causing the bean proteins to precipitate out. - The addition of acetic acid causes the bean protein to gain positive charges from being an isoelectric point (no charges), this gain of positive charges causes the bean protein to repel the charges of the hydrogen ion of the water thus precipitating out. Classify the following proteins according to their biological functions and describe: Proteins Class of biological function Description of the function Albumin Transport protein Maintains osmotic pressure in blood, transports fatty acids, hormones, drugs, and other substances. Gluten Structural protein Provides elasticity and viscosity to dough, forming the structure of bread and other baked goods. Myoglobin Storage protein Stores oxygen in muscle tissue for use during cellular respiration. CHARACTERIZATION OF PROTEINS (SUMMARY) Concentrations of isolated protein from the experiment: Gluten - 0.5g each test tubes Bean Protein - Precipitated bean protein is diluted using 15 ml distilled H2O (3ml each test tubes) Albumin - 3 ml each test tubes Test Test for presence Color Biuret Test General test for proteins, at Purple/Violet least 2 peptide bonds Ninhydrin Test Amino acid Blue/Violet Millon’s Test Tyrosine & phenol solution Brick Red PPT Hopkins-Cole Test Tryptophan Purple Ring Lead Acetate test Sulfur containing amino acids Black to Brown  (Methionine, cysteine, homocysteine, and taurine) Xanthoproteic Test Aromatic amino acids (e.g. Yellow tyrosine, tryptophan and phenylalanine) Sakaguchi Test Arginine Red CHARACTERIZATION OF PROTEINS (PRINCIPLE) General Test Biuret Test - The Biuret test detects the presence of peptide bonds, which are indicative of proteins. When a protein solution is mixed with Biuret reagent (a solution of copper sulfate), the peptide bonds in the protein complex with copper ions (Cu²⁺) under alkaline conditions. This complex forms a violet or purple color, indicating the presence of proteins. The intensity of the color is proportional to the number of peptide bonds, which correlates with protein concentration. Ninhydrin Test - The Ninhydrin test is used to detect free amino acids. When Ninhydrin reacts with the free amino group (-NH₂) of amino acids, it undergoes a decarboxylation reaction, resulting in the formation of a blue or purple-colored compound known as Ruhemann's purple. This reaction is sensitive and can detect even small amounts of amino acids. Ninhydrin also reacts with peptides and proteins but with a different color intensity Specific Test Millon’s Test - Millon’s test is specific for detecting phenolic compounds, particularly the amino acid tyrosine. In this test, Millon's reagent (mercuric nitrate in nitric acid) reacts with the phenolic group of tyrosine when heated. This reaction produces a red-colored complex, indicating the presence of tyrosine in the sample. This test is useful for distinguishing proteins that contain tyrosine from those that do not. Hopkins-Cole Test - The Hopkins-Cole test is specific for the detection of the amino acid tryptophan. In this test, the protein sample is mixed with glyoxylic acid and concentrated sulfuric acid. The indole ring of tryptophan reacts with glyoxylic acid under acidic conditions, forming a violet ring at the interface of the two liquids. This test is used to confirm the presence of tryptophan in proteins. Lead Acetate Test - The Lead Acetate test is used to detect sulfur-containing amino acids, such as cysteine. When a protein containing cysteine is treated with lead acetate, the sulfur in cysteine forms a black precipitate of lead sulfide (PbS). This reaction indicates the presence of cysteine or other sulfur-containing amino acids in the protein. Xanthoproteic Test - The Xanthoproteic test detects aromatic amino acids, particularly tyrosine, tryptophan, and phenylalanine. When proteins are treated with concentrated nitric acid, the aromatic rings of these amino acids undergo nitration, resulting in a yellow-colored complex. Upon addition of an alkali, the color intensifies to orange, confirming the presence of aromatic amino acids. Sakaguchi Test - The Sakaguchi test specifically detects the amino acid arginine. In this test, the guanidine group of arginine reacts with α-naphthol and sodium hypochlorite under alkaline conditions to form a red-colored complex. The presence of a red color indicates the presence of arginine in the protein sample.