E2 vit.c exp


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E2 vit.c exp

  1. 1. Introduction : Vitamin C (L-Ascorbic acid) is an important vitamin. Lack of ascorbic acid in diet causes scurvy, a disease characterized by weakness, small hemorrhages throughout the body that causes gums and skin to bleed, and loosening of the teeth. Scurvy was a serious problem for English sailors in the 1600s and 1700s. James Lind, a British doctor. His test was conducted at sea in May 1747 and consisted of two groups of men. One group was provided with lemon juice in addition to their normal rations, the other was not. This test was considered to be the first example of a controlled experiment comparing results on two populations of a factor applied to one group only with all other factors the same. The results of the tests conclusively showed that lemons prevented the disease. Around 1930 two scientists working independently isolated and published their findings on vitamin C. The men found that vitamin C prevented and treated scurvy. The term ascorbic acid was adopted to describe its ability to prevent scurvy.Since this discovery, the Royal Navy make sure that all sailors had lemon juice to drink when they were at sea for longer than one month. Figure 1: Structure of vitamin C (ascorbic acid). Ascorbic acid (C6H8O6) is a water-soluble vitamin, whose structure is shown in Figure 1. Vitamin C is easily oxidized, and the majority of its functions in vivo rely on this property. It plays a key role in the body’s synthesis of collagen and norepinephrine by keeping the enzymes responsible for these processes in their active reduced form. Vitamin C may also play a role in detoxifying by-products of respiration. The human body cannot produce ascorbic acid, and so it must be obtained entirely through one’s diet. A very small daily intake of vitamin C (10-15 mg/day for an adult) is required to avoid deficiency and stave off scurvy. The body requires extra vitamin C when fending infections. Since ascorbic acid is a water-soluble vitamin, risk of getting an over dose is minimal and any unused vitamin C will be excreted. The minimum daily requirement is 30 mg, and the recommended daily allowance is 60-70 mg per person. Fruits, vegetables, and organ meats (e.g., liver and kidney) are generally the best sources of ascorbic acid; muscle meats and most seeds do not contain significant amounts of ascorbic acid. The amount of ascorbic acid in plants varies greatly, depending on such factors as the variety, weather, and maturity. But the most significant determinant of vitamin C content in foods is how the food is stored and prepared. Since vitamin C is easily oxidized, storage and the cooking in air leads to the eventual oxidation of vitamin C by oxygen in the atmosphere. In addition, ascorbic acid’s water-solubility means that a significant amount of vitamin C present in a food can be lost by boiling it and then discarding the cooking water.
  2. 2. Title : Estimation of Vitamin C from Supplied Sample Theory: Ascorbic acid as the name implies possesses the usual acidic property, (donation of H+ ion). The acidity of vitamin C is not due to the carboxylic grouptied up in lactone from but to the ionization of enol group. The method of estimation of vitamin C is based on the stoichiometric reduction of dye 2,6-dichlorophenolindophenol to colourless compound by ascorbic acid. The titration is conducted in presence of metaphosphoric acid mixture in order to inhibit the ascorbic acid oxidation catalyzed by certain metallic ions (such as copper & silver ions present in distilled water). In aqueous system, the vitamin is easily oxidized, the instability increases with increase in pH. Metaphosphoric acid stabilizes the solution by lowering the pH. As a result of titration vitamin C gets oxidized to dehydro ascorbic acid structurally as follows. Vit. C + 2,6-dichlorophenolindophenol ↔ L-dehydroascorbic acid + 2,6-dichlorophenolindophenol (Oxidized form. Blue) (Colourless) Objectives : (1) To examine the percentage of vitamin C content in supplied sample. (2) To calculate the content of vitamin C of supplied sample based on content of vitamin C from ascorbic acid powder as reference. Materials: Slice of guava , 2,6-dichlorophenolindophenol , 3% metaphosphoric acid ,standard Vit.C solution. Apparatus : Burette , conical flasks , retort stand , beakers , mortar. Figure 2: Burette with retort stand Figure 3: Mortar Figure 4: Centrifuge Procedure : (1) 5.0 grams of guava is weighed. After that, it is grinded into guava juice with 20ml 3%metaphosporic acid. Guava juice is poured into two small tube (10ml each) and send to centrifuge. (2) Standard vitamin C solution is prepared by taking 10 mg of vitamin C mixed with 100 ml of 3% metaphosphoric acid in 100 ml volumetric flask.
  3. 3. (3) 10 ml of standard vitamin C solution is taken in a conical flask and it is titrated with prepared dye from a burette. The titration is terminated by the appearance of a permanent light pink colour in the titration medium. The operation is repeated for two times and the burette reading is recorded each time. (4) The supplied sample is transferred in a 50 ml volumetric flask make up to the mark by 3% metaphosphoric acid. 10 ml of this supplied solution is taken into a conical flask and it is titrated with the dye. This operation is repeated for two times and the reading is taken each times. Results Percentage error of burette: ±0.01 ml Table1: Titration volume for standard Vit. C No. of Observation Volume of standard Vit.C(ml) Amount of Vit. C (mg) Initial Burette Reading (ml) Final Burette Reading (ml) Average Reading (ml) 1 10 1 0.00 26.00 25.75 2 10 1 0.00 25.50 Table2: Titration volume for 5.0g of Guava No. of observation Volume of Guava Juice(ml) Initial Burette Reading (ml) Final Burette Reading (ml) Average Reading (ml) 1 10 0.00 13.40 12.90 2 10 0.00 12.40 Content of Vitamin C in Guava x=dye used to titrate ascorbic acid y=dye used to titrate guava juice 10mg of standard vitamin C 100ml of solution 1mg of standard vitamin C 10ml of solution mg of vitamin C × 5ml = x 5ml = 0.5010 mg 0.5010 mg x 20 = 50.10 mg = 0.0501 g *In every 100g of guava, there is 0.0501g of vitamin C in the content. Discussion Vitamin C (ascorbic acid) gets oxidized to its dehydro form by air specially at alkaline pH. However, it is suitable in an acidic solution. Therefore, vitamin C is extracted in metaphosphoric acid or in a mixture of metaphosphoric and dilute acetic acid. Its estimation in the extract is
  4. 4. carried out by titrating it against 2.6-dichlorophenol indophenols solution. Oxidized form of this dye is blue in colour in an alkaline medium and red in an acidic medium. Reduced form of the dye, on the other hand, is colourless and is termed as its leuco form. The redox reaction occurring during the titration process below: From the results of experiment, I had found that there is varies between the result of vit. C content come from the guava juice with the standard vitamin C solution. This variation may due to several factors as below: 1) Production factors and climate conditions: High nitrogen fertilizer rates can lower vitamin C levels in citrus fruits. Proper potassium levels are also needed for good vitamin C levels. Additionally, climate, especially temperature -- total available heat -- affect vitamin C levels. Areas with cool nights produce citrus fruits with higher vitamin C levels. Hot tropical areas produce fruit with lower levels of vitamin C. Environmental conditions that increase the acidity of citrus fruits also increase vitamin C levels. 2) Maturity state and position on the tree: Vitamin C decreases during the ripening process. Immature fruit has the highest levels. The position on the tree also affects vitamin C levels. Since sunlight exposure enhances vitamin C levels, fruit positioned on the outside of the tree and on the south side have higher levels. Shaded inside fruit has the lowest. 3) Handling and storage: Oxygen is the most destructive ingredient in juice causing degradation of vitamin C. However, one of the major sugars found in guava juice, fructose, can also cause vitamin C breakdown. The higher the fructose content, the greater the loss of vitamin C. Conversely, higher acid levels of citric and malic acids stabilize vitamin C. Conclusion Throughout this experiment, I had found that 100g of guava contain 0.0501g of Vitamin C. Reference 1) http://en.wikipedia.org/wiki/Vitamin_C 2) http://www.ultimatecitrus.com/vitaminc.html 3) Biochemistry Fourth Edition, Reginald H.Garret /Charles M.Grisham 4) Experimental Biochemistry A Student Companion