Processing of Horticultural Crops (1+2)

Exercise

Practical 10 - Ascorbic Acid

Aim: To estimate ascorbic acid in fresh and processed fruit and vegetable products.

Theory and Principle: Fruit, vegetables and their products are important sources of ascorbic acid. The ascorbic acid is present in sufficient quantity in aonla, guava, grapefruit, lemon, pineapple, strawberry fruits etc. Barbados cherry is the chief source of vitamin C. The products manufactured from these fruits are also considered as rich in ascorbic acid and the contents available in the commodities can be detected by using 2, 6 dichlorophenol - indophenol visual titration method.

2, 6 dichlorophenol - indophenol visual titration method: The method is based on reduction of 2, 6 dichlorophenol – indophenols dye. The dye, which is blue in alkaline solution and red in acidic solution, is reduced by ascorbic acid to a colourless form. The reduction is quantitative and specific for ascorbic acid in solutions in the pH range of 1.0 - 3.5. In estimation of ascorbic acid, the prepared sample is titrated against standard 2, 6 dichlorophenol – indophenols dye to a pink end point. The titre is then used to calculate the ascorbic acid in the sample.

Apparatus, reagents and glassware required

1. 3% metaphosphoric acid (HPO₃): Dissolve 3g sticks of HPO₃ in distilled water (100 ml). prepare 1 litre solution of 3% HPO₃ as it also required in sample preparation, 0.1% oxalic acid can also be used in place of metaphosphoric acid.
2. Ascorbic acid solution: Weigh 100 mg L-ascorbic acid and dissolve in 3% HPO₃ and make volume up to 100 ml with HPO₃. Dilute 10 ml to 100 ml with 3% HPO₃ (1ml=0.1mg ascorbic acid)
3. Sodium bicarbonate
4. Dye solution: Dissolve 50 mg of the Sodium salt of 2, 6 dichlorophenol indophenol dye in 150 ml hot glass distilled water containing 42 mg of Sodium bicarbonate. Cool and dilute with distilled water to 200 ml. Store in refrigerator and standardize.
5. Beakers - 100, 250ml
6. Volumetric flasks - 100, 250ml
7. Measuring cylinder - 250ml
8. Pipette - 10ml

Procedure for estimation
Standardization of dye: To 5 ml of Standard ascorbic acid (1ml=0.1mg) and 5 ml HPO_3. Titrate this solution with the dye solution to a pink colour which should persist for 15 seconds. Determine the dye factor i.e. ascorbic acid per ml of the dye.


Sample preparation:

1. Fresh Fruits/vegetables: Crush/grind fruit or vegetable parts (known weight) along with 3% HPO₃, make to volume (100 ml) with HPO₃. Filter or centrifuge.
2. Fruit Juices: To 10-20 ml juice add 3% HPO₃ and make volume to 100 ml. Filter or centrifuge.
3. Solid and semi solid food: Take 10 g sample, blend with 3% HPO₃ and make up to 100 ml with HPO₃. Filter or centrifuge.

Procedure for titration: Take (2-10ml) the HPO₃ extract of the sample and titrate with the standard dye to a pink end point persisting for at least 15 seconds. The titre of the sample should be such that titre should not exceed 3-5 ml.

Samples containing sulphur dioxide: Sulphur dioxide when present in fruit products like squash, jam, drinks etc reduces the indophenol dye and this interferes in the ascorbic acid estimation. To eliminate the SO₂ interference use formaldehyde condensation method as under:

To 10ml filtrate in a test tube, add 1ml of 40% formaldehyde and 0.1ml of HCl, keep for 10 minutes and titrate as earlier. Record titre and calculate ascorbic acid.

Problem: 10g of guava squash sample was made up 250ml using 3% metaphophoric acid and filtered. 10ml of the filtrate was titrated against standard 2,6 dichlorophenol dye having dye factor of 0.1mg ascorbic acid/ml dye. Calculate ascorbic acid in the guava squash when titre value was 5 ml.

Solution:

• Given: Wt of sample = 10g
• Volume made up = 250 ml
• ml of sample taken for estimation = 10ml
• Dye factor = 0.1
• Titre value = 5ml

5 × 0.1 × 250 × 100

= --------------------------------

10 × 10

mg ascorbic acid/100g = 125mg/100g


Inference: The given sample of guava squash contained 125mg/100g ascorbic acid.


Observations

Sample No.	Weight of sample taken	Volume made up	Aliquot used for estimation	Volume (ml) of standard dye used
1.
2.
3.
Mean