ESTIMATION OF COLCHICINE:In the first method for extraction (Method 1), 0.5 g of powdered plant material was extracted twice with 25 ml of petroleum ether with frequent shaking for 1 h, followed each time by filtration. The solid residues were air dried and then extracted with 10 ml of dichloromethane at room temperature for 30 min with frequent shaking. Then 10% solution of ammonia (0.5 ml) was added to the mixture with vigorous shaking for 10 min; the mixture was left undisturbed for 30 min and then filtered. The residue was washed twice with 10 ml of dichloromethane and then combined with the filtrate. The organic phase was evaporated to dryness and then dissolved in 1 ml of 70% ethanol to yield the test sample.
In the second method (Method 2), 12 g of plant material was extracted for 6 h in a Soxhlet extractor with methanol. The extract was diluted with distilled water and then partitioned against petroleum ether, and finally the aqueous phase was extracted with chloroform. The chloroform extract was then evaporated and the residue redissolved in methanol and filtered through 0.45 µm filter. The filtrate thus obtained constituted the test sample.
In the third method (Method 3), 20 g of freeze dried material was extracted using 200 ml methanol in a cold room (10°) overnight and the homogenate was centrifuged at 1252 × g for 5 min. The methanolic extract was evaporated to dryness and then the residues redissolved in 50 ml water. The aqueous extract was then centrifuged at 7826 × g for 5 min. The supernatant was first partitioned twice against petroleum ether and then discarded, and then once in diethyl ether, discarding the supernatant each time. The residue was washed five times with equal volumes of chloroform, which was retained and evaporated to dryness. The chloroform residue was redissolved in 95% HPLC grade methanol and then filtered through a 0.45 µm millipore filter to yield the test sample.
Identification of colchicine was done by...
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