Carbohydrate Isolation and Characterization Activity 5 PDF

ACTIVITY # 5 Isolation and Characterization of Carbohydrates Isolation, Hydrolysis and General Tests for Polysaccharides JRDIMAGUIBA  Materials for isolation of carbohydrates CHICKEN LIVER- glycogen Extraction...

ACTIVITY # 5 Isolation and Characterization of Carbohydrates Isolation, Hydrolysis and General Tests for Polysaccharides JRDIMAGUIBA  Materials for isolation of carbohydrates CHICKEN LIVER- glycogen Extraction Precipitation by ethanol -mincing and blending General Tests for CHO -0.1% acetic acid for ppt’n Acid hydrolysis Qualitative Tests  Materials for isolation of carbohydrates POTATO-starch Extraction General Tests for CHO -chopping and grinding -settling of starch Acid hydrolysis Qualitative Tests GENERAL TESTS FOR CARBOHYDRATES  Molisch Test Reagent: Molisch rgt (5% α-napthol in 95% ethanol) Positive result: purple ring at the junction of 2 solutions  Iodine Test Reagent: KI or I2 Positive result: blue solution QUALITATIVE TESTS FOR CARBOHYDRATES Bial’s Test Mucic acid test for galactose Mucic acid Test Galactose crystals in Mucic acid test Lactose crystals in Mucic acid test  Phenylhydrazone/ Osazone Test a chemical test used to detect reducing sugars This test even allows the differentiation of different reducing sugars on the basis of the time of appearance of the complex. Principle of Phenylhydrazone/ Osazone Test consists of phenylhydrazine in acetate buffer carbohydrates with free or potentially free carbonyl groups react with phenylhydrazine to form osazone The condensation-oxidation-condensation reaction between three molecules of phenylhydrazine and carbon one and two of aldoses and ketoses yields 1, 2-diphenyhydrazone, which is known as osazone. Principle of Phenylhydrazone/ Osazone Test Osazone appears as yellow-colored crystals of characteristic shape, solubility, melting point, and time of formation. Osazone is different for different sugars. Since both carbons 1 and 2 are involved in the reaction, C-2 epimers produce the same osazone. Similarly, ketoses with a configuration identical to aldoses below C-2 give the same osazones, e.g. glucose and fructose. Osazone formation using phenylhydrazine https://microbenotes.com/osazone-test/ A typical reaction showing the formation of an osazone. D-glucose reacts with phenylhydrazine to give glucosazone.  Chart for the identification of reducing sugar through osazone/phenylhydrazone test Uses of Phenylhydrazone/ Osazone Test used to distinguish lactose from maltose during the identification of unknown sugars a simple, cheap, and relatively less time-consuming test for the identification and differentiation of different sugars encountered in clinical practice can also be used for locating sugars in plant tissues. Limitations of Phenylhydrazone/ Osazone Test gives a positive result for sucrose when if boiled for 30 minutes or more even though sucrose is a non-reducing sugar not effective if the sample contains a mixture of different sugars Similarly, large quantities of sugars are required for a positive result. Thank you!