Module 3 - Strategic case studies in practice

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Colourant identification

1. Extraction methods

Method A Reagents

Extraction solution: Ethanol or Methanol «Analar"- 3 parts by volume Distilled water -1 part by volume Apparatus Glass beaker - 10 mI Procedure

a) Place one cap or one body or, for a capsule with a low dye content, three caps or bodies in a 10 ml glass beaker. Add 2 to 3 ml of the extraction solution. Leave in the dark for 30 minutes to 1 hour. b) Decant off the supernatant liquid. If much dye remains in the capsule, add a further quantity of the extrac~ion solution. c) Evaporate the supernatant liquid to dryness in a stream of cool air. d) Re-dissolve the extract in the minimum quantity of distilled water. e) If no colour change is observed in the capsule after extraction or the capsule is still strongly coloured, the presence of pigments is indicated. Method B Reagents 0.5% acetic acid solution - 10 ml of dilute acetic acid diluted to 120 ml with water. 50% ammonia solution- Dilute 50 ml of strong ammonia solution to 100 ml with water. · Apparatus Buchner funnel- Hartley 3-section pattern, porcelain (disc diameter 20 mm) Filter paper- Whatman AE 81 (amino ethyl cellulose) paper ( 3 em diameter). Procedure a) Dissolve one of either caps or bodies in 50 ml of water by heating on a water bath of 60°C. b) If pigments are present, transfer to a centrifuge tube, and centrifuge until supernatant liquid is clear. Decant of supernatant liquid. Reheat the solution on a water bath to 60°C. c) Filter the warm solution through Whatman AE81 amino ethyl cellulose paper. The discs of filter are held in a filter holder which is capable of being heated to 60°C. About 6 discs of paper are used. d) Wash the filter paper with two quantities of boiling water. e) Remove the discs. Place in 100 ml of 0,5% acetic acid solution and boil for 5 minutes. 1 f) Remove the discs and drain them. Place in 100 ml water and boil for 5 minutes. Repeat with a further quantity of 100 ml water. g) Dry the discs with sheets of Whatman No 1 filter paper. h) Return discs to holder. Elute the dyes with the minimum quantity of 50% ammania solution. i) Evaporate the solution to dryness in a stream of cool air. Redissolve the extraction in a minimum quantity of water.

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