376 BCH Miss Tahani Al-shehri 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 Dehydroscorbic acid Ascorbic 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 Measure the volume of the urine. Pipette 5 ml of urine into a 100 ml volumetric flask and make up to the mark with 5% acetic acid. Pipette 5 ml of DCPIP dye solution into a conical flask. Titrate with standard ascorbic acid solution (0.02 mg/ ml) until the solution just becomes colourless.

METHOD cont.. Calculate how many milligrams of ascorbic acid are equivalent to 5 ml of the dye solution. Pipette 5 ml of dye into a conical flask. 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 Repeat the titrate and take the average

METHOD cont.. Calculate how much ascorbic acid present in sample urine diluted 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 =…………………………………………………………