Procedure:

Preparation of Solutions:

Molybdate Solution:

Add 13.6 mL of con.sulphuric acid to 35 mL of water and allow the solution to cool. In a separate vessel add 2.5 g of ammonium molybdate to 50 mL of water. Add sulphuric acid solution to it & make up the volume.

Reducing Solution:

Dissolve 11.25 g sodium hydrogen phosphate in 30 mL water. Add 2.25 g sodium sulphate and 0.19 g 1-amino 2-naphthol 4-sulphonic acid and dilute the mixture to 75 mL with water. Stir the mixture thoroughly & filter.

Standard Solutions of Phosphate:

Dissolve 20 mg of potassium dihydrogen phosphate in 100 mL of water. The concentration of phosphate standard solution is 0.2 mg/mL.

• Prepare 9 standard solutions containing 0.5 mL molybdate solution, 0.2 mL reducing solution & x mL of phosphate stock solution + y mL  of distilled water. Where, x+y =9.3 mL and x= 2-0.2n, n = 1,2,…………9.
• Also prepare a blank solution by mixing 0.5 ml of molybdate solution, 0.2 ml of reducing solution and 9.3 mL of water.
• Set the wave length to 635 nm and adjust the colorimeter reading to zero with the blank.
• Place each standard solution of phosphate in turn in the curette and record the absorbance.
• Dilute the soft drink sample 5 times. Mix 1mL of this diluted solution with 0.5 mL molybdate, 0.2 mL reducing solution and 8.3 mL water, so as to get the test solution.
• Measure the absorbance of the sample solutions.
• Plot a graph between absorbance and volume of phosphate in various standard solutions and obtain the calibrated curve.
• From the curve determine the amount of phosphate in the test solution.

The amount of phosphate in the sample of soft drink is 5 x 0.2 x V = 1 x V mg /mL.
5 is the dilution factor.
0.2 mg/mL amount of phosphate in standard solution.
V is the volume of phosphate solution obtained from graph.