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Protein digestion


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Protein digestion

  1. 1. Protein Digestion<br />Presented by: GROUP 1 – DAA<br />Coojacinto<br />De Asis<br />Go<br />Saunar<br />
  2. 2. Proteins are…<br /> biochemical compounds consisting of one or more polypeptides typically folded into a globular or fibrous form, facilitating a biological function. <br />A polypeptide is a single linear polymer chain of amino acids bonded together by peptide bonds between the carboxyl and amino groups of adjacent amino acid residues<br />
  3. 3. Protein digestion <br />The process of digestion is defined as the ‘process by which macromolecules in food are broken down into their component small-molecule subunits’.<br />The macromolecules are the proteins or polypeptides themselves, and the subunits are the amino acids. The bonds holding the subunits together are peptide bonds<br />
  4. 4. Protein Digestion<br />is the degradation of proteins by cellular enzymes enzymes in a process called hydrolysis.<br />Protein digestion takes place in two different phases:<br />In the stomach<br />In the small intestine<br />Both of these phases of digestion are based on several types of enzymes that are called proteinases and proteases.<br />
  5. 5. Proteases- endo- & exo- peptidases; enzymes that degrade proteins by hydrolysis of peptide bonds<br />Proteinases- endo-peptidases; proteases that show specificity for intact proteins<br />
  6. 6. Peptide bond hydrolase<br />Peptidase<br />(=Protease)<br />Endo-acting peptide bond hydrolase<br />Endopeptidase<br />(=Proteinase)<br /> Exo-acting peptide bond hydrolase<br />Exopeptidase<br />
  7. 7. Mouth and Salivary Glands<br /> Chewing and crushing of protein rich foods and mix them with saliva to be swallowed<br />
  8. 8.
  9. 9.
  10. 10. In the Stomach : start of protein Digestion<br />Gastrin<br /> -stimulates Parietal cells to secrete HCL; Chief cells of the gastric glands to secrete pepsinogen<br />Hydrochloric acid<br /> -Denatures protein structure<br /> -Activates pepsinogen(zymogen) to pepsin<br />Pepsin <br /> -hydrolyzes proteins to smaller polypeptides and some free amino acids. <br />
  11. 11. In the intestine<br />. The remainder of protein digestion occurs in the small intestine as the result of the action of enzymes <br /> such as trypsin (secreted by the pancreas) and peptidases (located in the cells that line the small intestine).<br />
  12. 12. In the Small Intestine : enzymes<br />Secretin<br /> - stimulates the pancreas to secrete bicarbonate into the small intestine to neutralize the gastric HCl<br />Cholecystokinin<br />-stimulates secretion of several pancreatic enzyme with activity optima pH 7 to 8.<br />
  13. 13. In the Small Intestine<br />Cont....<br />Trypsin<br /> - activates chymotrypsinogenchymotrypsin<br />procarboxypeptidasescarboxypeptidases<br />proelastaseelastase<br /> -further hydrolyze the peptides that were produced by pepsin in the stomach specifically the peptide bonds next to lysine and arginine<br />Chymotrypsin<br /> -cleaves peptide bonds next to phe, tyr, trp,met, asp, and his<br />
  14. 14. Cont....<br />Carboxypeptidase A & B<br /> -cleave amino acids from the acid (carboxyl) ends of polypeptides<br />Elastase and collagenase<br /> -cleave polypeptides into smaller polypeptides and tripeptides<br />
  15. 15. Cont....<br />Intestinal tripeptidases<br /> -Cleave tripeptides to dipeptides and amino acids<br />Intestinal dipeptidases<br /> -cleave dipeptides to amino acids<br />Intestinal aminopeptidases<br /> -cleave amino acids from the amino ends of small polypeptides(oligopeptides)<br />
  16. 16. Amino acids absorbed<br />Free amino acid  small intestine(villi)Liverblood circulation<br />
  17. 17. Materials:<br />Commercial pepsin<br />Trypsin<br />Concentrated HCl<br />NaOH<br />CuSO4 solution<br />0.5% Na2CO3<br />Hard Boiled Egg (Protein) albumin<br />
  18. 18. Biuret Test<br />a chemical test used to detect the presence of peptide bonds<br />Reagent: Potassium hydroxide (KOH) and hydrated copper(II) sulfate, sodium tartarate<br />Result:<br /><ul><li>(+) test = purple (presence of proteins)
  19. 19. (−) test = blue
  20. 20. blue to pink when combined with short-chain polypeptides (it will not cleave on all peptide chains)</li></li></ul><li>Procedure<br />Action of the Gastric Protease (Pepsin) on Proteins<br />Action of Pancreatic Protease (Trypsin) on Proteins<br />
  21. 21. Action of the Gastric Protease (Pepsin) on Proteins<br />
  22. 22. Action of Pancreatic Protease (Trypsin) on Proteins<br />
  23. 23. B. THE ACTION OF PANCREATIC PROTEASE (TRYPSIN) ON PROTEINS<br />Label 3 tubes, each half full of warm H2O<br />To the first tube add a small amount of grated white of hard boiled egg.<br />To the first tube add a 0.5g of trypsin and 2ml of 0.5% NaCO3.<br />To the second tube add 0.5g trypsin.<br />To the third tube add 2ml of Na2CO3 solution.<br />Keep the tube in a warm H2O bath kept at body temperature for an hour.<br />Filter the contents of each tube and the Biuret test on each of the filtrates.<br />Observe and record the results.<br />
  24. 24. Ideal Results:<br />
  25. 25. POST LAB QUESTIONS<br />
  26. 26. POST LAB QUESTIONS<br />What are the enzymes used in Protein Digestion?<br />Gastric Protease (Pepsin)<br />Pancreatic Protease (Trypsin)<br />Small intestine enzymes (peptidases)<br />
  27. 27. POST LAB QUESTIONS<br />What are the similarities and differences of these enzymes?<br /> These enzyme hydrolyze proteins and their main difference is the location where they are found.<br />
  28. 28. POST LAB QUESTIONS<br />What are the factors that would bring about Protein digestion?<br /><ul><li>the acidity of the food and of the stomach
  29. 29. Temperature
  30. 30. the presence of any digestion inhibitors, such as antacids</li></li></ul><li>POST LAB QUESTIONS<br />Trace the pathway of Protein Digestion<br />Stomach<br />Gastrin stimulates:<br />Parietal cells secrete HCl<br />Chief cells secrete pepsinogen then HCL convert it to pepsin but acts only on certain amino acids<br />Small Intestine<br />released in the lumen<br />Cholecystokinin stimulates pancreatic enzymes such as:<br />TrypsinTrypsinogen (active form)<br />specific for Lys and Arg (carboxyl side)<br />ChymotrypsinChymotrypsinogen (active form)<br />specific for Tyr, Trp, Phe, Leu and Met (carboxyl side) <br />
  31. 31. Protein Digestion Pathway <br /> ( digestion doesn’t happen yet)<br /> Mouth and Salivary Glands<br />Stomach( Gastric Phase)<br />Small Intestine and Pancreas <br />(Pancreatic Phase) <br />Blood<br />
  32. 32. POST LAB QUESTIONS<br />What tests are used to detect the completeness of protein digestion?<br />MILLON’S TEST<br />NITROPRUSSIDE TEST<br />HOPKIN’S COLE TEST<br />XANTHOPROTEIC TEST<br />SAKAGUCHI TEST<br />PAULY TEST<br />
  33. 33. 5.<br />MILLON’S TEST<br /><ul><li>Test for tyrosine (tyr)
  34. 34. Used to detect the presence of soluble proteins
  35. 35. Given by phenols or phenolic substances such as Salicylic acid</li></ul>Principle<br />Mercuric sulphate forms a colored compound with hydroxyl group of Tyr. <br />Result: (+) red ppt<br />
  36. 36. 5.<br />HOPKIN’S COLE TEST<br /><ul><li>specific for tryptophan --- indole group. </li></ul>Principle<br />The indole ring reacts with glyoxylic acid in the presence of a strong acid: H2SO4<br />Reagent: glyoxylic acid<br />Result: violet cyclic product<br />
  37. 37. 5.<br />PAULY’S TEST<br /><ul><li>For tyr, trp, and his</li></ul> Principle<br />Diazotisedsulphanilic acid couples with amino phenol and immidazole to form a colored azocomp’d in cold condition.<br />Result: (+) Deep red color dye<br />
  38. 38. 5.<br />SAKAGUCHI TEST<br /><ul><li>Test for Arg -- guanido group</li></ul>Principle<br />Alkaline solution<br />SakaguchiReagent:α-naphthol and sodium hypochlorite<br />Result: (+) reddish wine color<br />
  39. 39. 5.<br />XANTHOPROTEIC TEST<br /><ul><li>For aromatic groups: tyr, trp, phe(unactivated)</li></ul>Principle<br />Nitration of a Benzene ring with Nitric Acid<br />Reagent: Nitric acid (HNO3) and NaOH<br />Result: (+) yellow (tyr); orange (trp) --alkali<br />
  40. 40. 5.<br />SODIUM NITROPRUSSIDE TEST<br /><ul><li>Bollin’s test
  41. 41. specific for cys – free thiol group (S—H)</li></ul>Principle<br />Cystine whichcontains disulphide linkage (S—S) may be reduced to cysteine<br />reducing agent/s: sodium cyanide, sodium brohydride or sodium bisulphate<br />Result: (+) red complex<br />
  42. 42. What are the factors affecting protein digestion?<br />pH: acidity and alkalinity of environment <br />Regulating activity of inhibitors<br />