*2.4.6. Determination of vitamin-C*

**Preparation of standard solution:** 5 mg of ascorbic acid was taken and dilute in 5 ml of distilled water to give concentration of 1 mg/1 ml.

**Procedure:** (1) Take 25 ml of test sample and add 25 ml of 20% Meta phosphoric acid. (2) Dilute it to 100 ml with distilled water. (3) Take 10 ml of aliquot from the above solution and add 2.5 ml of acetone. (4) Titrate it with 0.05% dye solution till a pink color persist for 15 s (V1). (5) For the standard readings take 0.05 g of vitamin C (ascorbic acid) (A). (6) Add 60 ml of 20% Metaphosphoric acid and dilute it to 50 ml with distilled water. (7) Titrate the known volume of the above solution with 0.05% dye solution till a pink color persist for 15 s (V2).

Calculation:

powder was placed in a porous thimble and kept in the middle compartment. For the procedure, the solvent in the lower container is heated to its boiling temperature (different solvents have different boiling temperature to maintain), and reflux condenser vapor passes through the side arm up. The thimble containing the material to be extracted using the vapor liquefies and drips. Here, central compartment extract gradually collects from warm water percolates through the material and the wall of the thimble. The entire liquid in the central compartment flows through the side tube and back into the lower solvent container of the height of the extract reaches to the top of the siphon. The extract removed in a petri dish and left aside to evaporate. After evaporation of solvent, the leftover extract was filled in the eppendorf tube.

Stock solution is prepared by dissolving 30 mg of methanolic, hexanoic and distilled water extract in to 30 ml of methanol, hexane and distilled water respectively to give the concentra-

(1) Ten grams of fruit sample were taken in a Petri dish and kept in a hot air oven at 90–100°C for 5–6 h. (2) Weight of the fruit was measured after it was completely dry. (3) Loss in weight

(1) A mixture of 50 ml of methanol and diethyl ether were taken in 1:1 concentration. (2) From 30 ml of stock solution, 1 ml of test sample was added in the mixture and few drops of phenolphthalein as an indicator. (3) The solution was titrated with 0.1 N NaOH. (4) The change in

The biuret test is a chemical test used to detect the presence of peptide bonds. In the presence of peptides (–CO-NH- groups), a copper (II) ion forms violate coordination complexes in an alkaline solution. Copper (II) reduces to copper (I). The intensity of the color complex is

**Reagents:** (1) Biuret reagent: Dissolve 3 g of CuSO4.5H2O and 9 g of sodium potassium tartrate in 500 ml of 0.2 N NaOH solutions. To this solution, add 5 g of potassium iodide and make about 1 L with 0.2 N NaOH. (2) Standard protein solution: 5 mg of bovine serum albu-

This process is then repeated with the other solvents.

126 Phytochemicals - Source of Antioxidants and Role in Disease Prevention

**2.4. Analysis of nutritional value**

tion of 1 mg/1 ml [13].

*2.4.2. Estimation of moisture*

*2.4.3. Estimation of fat content*

*2.4.4. Estimation of crude protein*

min/ml water. Prepare fresh.

measured by colorimetrically at 520 nm.

Calculation

*2.4.1. Sample preparation and nutritive analysis*

was regarded as a measure of moisture content.

color was observed from light yellow to a brownish coffee color.

Acid value (mg/L) <sup>=</sup> (*Test* <sup>−</sup> *Blank*)*X*0.1*X*254 \_\_\_\_\_\_\_\_\_\_\_\_\_\_\_\_\_\_\_\_ *Sampled used* (*ml*)

Calculation:

$$\text{Amount of } \text{acorbic acid (mg/100ml)} = \frac{\text{(A mg)} \times \text{(U2)} \times \text{250 X100}}{\text{(V1 mol/X (B)X (wt of Sample)}}$$

A mg = Std. vitamin C taken; V1 ml = Vol. of dye taken to titrate the sample; V2 ml = Vol. of dye taken to titrate std. vitamin C; B = Total vol. of std. solution taken which is to be titrated against 0.05% dye; 250 = Total vol. of std. solution made after dilution.
