Ascorbic Acid Saturation Test Determination of Ascorbic Acid

Ascorbic Acid Saturation Test & Determination of Ascorbic Acid Content of Some Fruit Juices. 376 BCH Miss Tahani Al-shehri

OBJECTIVES • To measure the concentration of ascorbic acid urine sample after administration of a dose of ascorbic acid.

Introduction • Vitamin C is a water-soluble vitamin that is necessary for normal growth and development. • Water-soluble vitamins dissolve in water. The body cannot store them. Leftover amounts of the vitamin leave the body through the urine. That means you need a continuous supply of such vitamins in your diet.

Introduction Ascorbic acid Dehydroscorbic acid

Functions of Vitamin C • Vitamin C is required for the growth and repair of tissues in all parts of your body. • It is necessary to form collagen • Vitamin C is one of many antioxidants in the body * The body does not manufacture vitamin C on its own ( Why ? )

Deficiency of Ascorbic acid • “ Scurvy “

ascorbic acid saturation test • Administered ascorbic acid is taken up by the tissues until they are saturated, after which there is a sharp rise in the urinary ascorbic acid excretion.

Principle • In a protein-free filtrate of plasma or urine, ascorbic acid has the property of decolorizing a blue dye, dichloro-phenolindophenol DCPIP and the amount that is decolorized is a measure of the amount of the vitamin C present.

Principle

Notes • Estimation of urinary ascorbic acid when the specimen arrives in the laboratory must carried out immediately. • Ascorbic acid is unstable in urine and a special bottle containing 20 ml of glacial acetic acid must be used for the urine collection. • Glacial acetic acid is corrosive and any spilt on the skin should immediately be washed off with a large volume of water.

METHOD 1. Measure the volume of the urine. 2. Pipette 5 ml of urine into a 100 ml volumetric flask and make up to the mark with 5% acetic acid. 3. Pipette 5 ml of DCPIP dye solution into a conical flask. 4. Titrate with standard ascorbic acid solution (0. 02 mg/ ml) until the solution just becomes colourless.

METHOD cont. . 5. Calculate how many milligrams of ascorbic acid are equivalent to 5 ml of the dye solution. 6. Pipette 5 ml of dye into a conical flask. 7. Titrate with the dilute urine sample. The titration must be done very rapidly, otherwise slow-reacting reducing substances present in the urine will contribute to the reduction of the dye 8. Repeat the titrate and take the average

METHOD cont. . 9. Calculate how much ascorbic acid present in sample urine diluted 10. Calculate milligrams of ascorbic acid in total urine collected.

Results • Volume of urine =………………. • Mg of ascorbic acid equivalent to 5 ml of the dye solution =…………………… • The conc. Of ascorbic acid present in sample urine diluted =………………… • Mg of ascorbic acid in total urine collected =……………………………