3. OBJECTIVES To measure ascorbic acid level in food products Identify the sources of ascorbic acid Master of extraction and determination of ascorbic acid (vitamin c) in food products Explain the clinical significance of Vitamin C

4. Outline of Today’s lecture 1. Introduction 2. Sources of Vitamin C Functions of Vitamin C 3. Functions of Vitamin C 4. Spectrophotometric determination

5. What is Ascorbic Acid? Ascorbic acid is a naturally occurring organic compound with antioxidant properties. It is a white solid, but impure samples can appear yellowish. It dissolves well in water to give mildly acidic solutions. Ascorbic acid is one form of vitamin C.organic compoundantioxidantvitamin C

6. 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.

7. Sources of Vitamin C It can be obtained from citrus fruits, tomatoes, potatoes, and fresh vegetables, particularly from red and green peppers. It is now made synthetically. All animals except man, other primates, and guinea pigs, are capable of synthesizing their own ascorbic acid; man must obtain it from food.

11. 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

13. Symptoms of a Vitamin C Deficiency Fatigue – Early on, someone with a vitamin C deficiency will tend to get tired easily and experience reduced energy. Mood Changes – Individuals with a vitamin C deficiency may become irritable or short tempered. Weight Loss – As with other “wasting conditions” someone with a vitamin C deficiency may experience sudden weight loss. Joint and Muscle Aches – Chronic pain in the limbs or joints can be a symptom of a vitamin deficiency.

14. Continue…. Dental Conditions – Just as a healthy daily dose of vitamin C contributes to healthy teeth and gums, a deficiency can cause deterioration of the gums. Dry Hair and Skin – A change in hair and skin conditions can also signal that the body is not getting enough of vitamin C and other essential vitamins and minerals. Infections – When an individual does not have enough vitamin C over time, this can have a negative impact on general healing of wounds and the fighting of infections.

15. Deficiency of Ascorbic acid

17. “ Scurvy “

18. Two of the chemical properties of ascorbic acid are its acidity, due to the enolic hydrogen at C-3 (see Reaction 1) and its oxidation to dehydroascorbic acid (see Reaction 2).

20. The reducing power of ascorbic acid is the basis for most of the quantitative analytical procedures for estimation of this compound, such as titration of the dye, 2,6- dichlorophenolindophenol (DCPIP) (see Reaction 3).

22. Procedure A- Preparation of samples: 1-Citrus fruit juice and bottled juices. The juice from fresh fruit can be obtained with a lemon squeezer followed by filtration through course filter paper. These steps will remove fibrous materials that might plug the tip of the pipettes. Pipette 0.5 ml of fruit juice into a test tube and dilute with 4.5 ml of 1% phosphate-citrus solution. Cover the tube with parafilm and invert several times to mix. Remember to account for these dilution factors when calculating your vitamin C concentration in the sample.

23. 2-Vegetables. Homogenize 1 gm (weight to 0.01gm) of vegetable with 5-10 ml of 1% phosphate – citrate using a mortar and pestle. Pour off the liquid into a filter funnel. Extract the solids a second time with 10 ml of 1% phosphate-citrate and filter. Combine the filtrate into a volumetric flask. Add 1% phosphate- citrate to the filtrate until the volume is 25 ml. Remember to account for these dilution factors when calculating your vitamin C concentration

24. Preparation of Cuvettes Sample (Reagent+Sample Standard (Reagent+ Vit.C)) Blank (only Reagent)

25. S#2S#1Std. Reagents (ml) --0.05 Vitamin C Standard -0.05- Sample # 1 0.05-- Sample # 2 0.450.450.45 Phosphate – Citrate buffer 1.01.01.0DCPIP

26. Mix all the tubes; zero the spectrophotometer at 518 nm with the blank (only distilled water). Read all the standard and samples at 518 nm. Record your results for further calculations.

27. Concentration of vitamin C (mg/ml)= Absorbance of Sample X Conc. of Standard X 10 Absorbance of Standard