Post-Lab Qualitative Tests for Carbohydrates and Lipids PDF

POST LAB: Activity No. 1- CARBOHYDRATES IDENTIFICATION Carbohydrates  These are polyhydroxy aldehydes or polyhydroxy ketones or compounds which yield them on hydrolysis.  Polyhydroxy signifies that the aldehydes/ketone...

POST LAB: Activity No. 1- CARBOHYDRATES IDENTIFICATION Carbohydrates  These are polyhydroxy aldehydes or polyhydroxy ketones or compounds which yield them on hydrolysis.  Polyhydroxy signifies that the aldehydes/ketone group has several hydroxy ( -CH ) group in addition to their several -OH group. CARBOHYDRATES 1. Molish Test - The formation of violet color is due to the production of – violet ring furfuraldehyde or related compounds of the action of the acids in carbohydrates. The color is due to the action of these aldehyde on alpha- naphtol. -CHO + α-naphtol H2SO4 violet ring is formed at the (dehydrating agent) conjunction of 2 liquids a. Glucose (hexealdose) b. maltose + α - naphtol H2SO4 all positive c. starch REDUCING 2. Benedict's Test - also called Benedict's solution or Benedict's test) is a SUGAR reagent named after an American chemist, Stanley Rossiter Benedict. It is – brick red used as a test for the presence of reducing sugars such as glucose, fructose and maltose, or more generally for the presence of aldehydes (except aromatic ones). It is often used in place of Fehling's solution. Benedict's reagent contains blue copper(II) sulfate (CuSO4) which is reduced to red copper(I) oxide (Cu2O) by aldehydes, also oxidizing them to carboxylic acids. The copper(I) oxide is insoluble in water and so precipitates. + (positive) aldehyde - (negative) ketone R - CHO + CuSO4 + NaCO3 sodium nitrate Benedict's reagent C6H5O7Na3.2H20 brick red precipitate Or (Cu2O) sodium nitrate C6H5O7NCl3.2H2 a. glucose + b. fructose (ketohexose - c. galactose (aldohexose) + d. sucrose + CuSO4 + NaCO3 - e. maltose + f. lactose + g. starch -  MONO 3. Barfoed's Test - is a chemical test used for detecting the presence of FROM monosaccharides. It is based on the reduction of copper(II) acetate to copper(I) DISACC oxide (Cu2O), which forms a brick-red precipitate. (Disaccharides may also react, – brick red but the reaction is much slower.) A number of other substances, including sodium chloride, may interfere a. glucose + b. fructose + c. maltose + Cu(C2H3O2)2 + HAc - d. lactose - Note: Maltose and lactose will give a red precipitate if it will be heat continuously. REDUCING 4. Fehling's test - This contains copper (Cu+) complex ion where in a red SUGAR precipitate of copper(I) oxide [Cu2O] is formed. This is actually the test for – brick red glucose (sugar) in urine, since glucose is an aldehyde. Somewhat similar with Benedict's Test + (positive) aldehyde - (negative) ketone  Rochelle salt, colorless to blue-white orthorhombic crystalline salt with a saline, cooling taste. It is also called Seignette salt after Pierre Seignette, an apothecary of La Rochelle, France, who was the first to make it (c.1675). Chemically, it is potassium sodium tartrate, KNa (C4H4O6)·4H2O. R - CHO + CuSO4 + NaoH Rochelle salt RCOONa + CU2O (brick (sodium potassium tartrate) red precipitate) + Na2SO4 + H2O a. Glucose + b. fructose + CuSO4 + NaoH Rochelle salt + c. sucrose - d. maltose + e. lactose + f. starch solution - g. acidified glycogen + REDUCING 5. Nylander Test SUGAR – black Bi(OH)2NO3 + KOH reducing sugar Bi(OH) + KNO 3 (Bismuth subnitrate) ∆ Bi(OH)3 reducing sugar 2Bi + 3O + H2O ∆ Metallic bismuth (black precipitate) a. glucose + b. sucrose - c. maltose + d. lactose + Bi(OH)3 + e. galactose +  f. starch - g. fructose + KETOSE 6. Seliwanoff’s test - is utilized for the differentiation of aldose and ketose FROM sugars. A ketose is said to be present if the mixture turns red. For a general ALDOSE preview, one way to differentiate ketoses from aldoses is to look for the – ketose: deep cherry red functional group attached to it. When a ketone is attached to it, it is a ketose; – aldose: faint pink however, if an aldehyde is attached to it, it is an aldose. Ketoses are more rapidly dehydrated than aldoses when heated. The dehydrated ketose then reacts with the resorcinol to produce a deep cherry red color. Aldoses may react slightly to produce a faint pink color.  + (positive) ketone - (negative) aldose ketone + resorcinol-HCl hydroxymethyl furfural condenses red color compound a. glucose - b. fructose + resorcinol-HCl + c. sucrose + REDUCING 8. Picric acid Test - It is another test for detection of reducing sugars. The SUGAR reducing sugars react with picric acid to form a red colored picramic acid. – mahogany red Appearance of mahogany red (dark brown-red) color would indicate the presence of reducing sugars in the sample solution. POST LAB: Activity No. 2- TESTS FOR INDIVIDUAL CARBOHYDRATES A. Osazone Formation HEAT: REDUCING SUGAR + PHENYLHYDRAZINE = YELLOW PPT  If a solution of reducing sugar is heated with phenylhydrazine, a yellow precipitate is obtained. The precipitated compound is called osazones.  The formation of these distinctive crystalline derivatives is useful for comparing the structures of sugars.  Osazones of monosaccharides (glucose & fructose) are formed on hot after about 15 minutes, and have the same crystal shape needles under the microscope.  Osazones of reducing disaccharides (maltose & lactose) are formed after a longer time (up to 30 minutes) and crystal appears slowly after cooling and can be distinguished under the microscope as follows: Lactose gives crystals in the form of a tuft of needles. Maltose gives crystals in the form of broad needles. Glucose + Phenylhydrazine-NaAc Phenylglucosazone Fructose + Phenylhydrazine-NaAc Phenylfructosazone Lactose + Phenylhydrazine-NaAc Phenyllactosazone Maltose + Phenylhydrazine-NaAc Phenylmaltosazone B. Hydrolysis of Starch and Glycogen  Iodine Test - Organic analysis relies on certain chemical reaction particularly functional group such as amine, alcohol, aldehyde, alkene, ester, carboxylic acid and ether.  Iodine Solution - iodine dissolved in an aqueous solution of potassium iodide (IKI)  reacts with starch producing a deep blue black color. Starch Iodine Sol’n dextrins maltose D-glucose Starch (deep blue) dextrins (amylodextrins) faint blue Erythrodextrin (red) achrodextrin (faint red/pink) Maltose (colorless) glucose (colorless)  Activity No. 3 THE FERMENTATION OF SUGARS USING YEAST Sugar fermentation results in the production of ethanol and carbon dioxide. In the case of sucrose, the fermentation reaction is: C12H22O11(aq)+H2O Yeast 4C2H5OH(aq)+4CO2(g) Lactose is also C12H22O11 but the atoms are arranged differently. Before the disaccharides sucrose and lactose can undergo fermentation, they have to be broken down into monosaccharides by the hydrolysis reaction shown below: C12H22O11+H2O⟶2C6H12O6 The hydrolysis of sucrose results in the formation of glucose and fructose, while lactose produces glucose and galactose. sucrose + water ⟶ glucose + fructose lactose + water ⟶glucose + galactose The enzymes sucrase and lactase are capable of catalyzing the hydrolysis of sucrose and lactose, respectively. The monosaccharides glucose, fructose and galactose all have the molecular formula C6H12O6 and ferment as follows: C6H12O6(aq) Yeast 2C2H5OH(aq)+2CO2(g)  Activity No. 4 Properties of Lipids A. Solubility Test 1. oil + CHCl3 chloroform = Miscible 2. oil + EtOH = Immiscible etoh tubig — di natutunaw 3. oil + H2O = Immiscible Principle: Follows Like dissolves Like Principle. B. The Grease-spot Test for Lipids 1. Water = absorbed by the paper 2. oil = formed translucent C. Acrolein Test – Test for Glycerol - indicates the presence of glycerol, components of fats and oil - on continued heating with KHSO4 (Catalyst) glycerol is dehydrated forming acrolein or acrylic aldehyde (odor of burned copra). - the reaction is Fat Fatty acid + glycerol H | H H – C – OH H–C=O I heated I H – C – OH ---------- > H- C + 2 H2O I KHSO4 II H – C – OH H–C | | H H Acrolein/ Acraldehyde D. Test for Sterol Sterol – a hydroxyl derivatives of steroids ( cyclopentanoperhydrophenanthrene ) 1. Lieberman-Burchard Test - The sterol is treated with chloroform, CHCl3, acetic anhydride, and concentrated H2SO4 to produce an emerald green color. CHCl3 + H2SO4 + Butter + acetic anhydride Sterol (emerald green color) 2. Salkowski Test - A solution of the sterol in chloroform, CHCl3, is shaken with concentrated H2SO4. - A positive test is the formation of cherry red color that appears in both layers. CHCl3 + H2SO4 + Butter Sterol (cherry red color)  Activity No. 5 Saponification Saponification - alkaline hydrolysis of fats or oils - when fat is hydrolyzed by means of strong alkali’s such as NaOH or KOH, soap and glycerol are formed. - the reaction is Fat Salts of Fatty acid + glycerol (Soap)