22.3 g fresh leaf was air dried to constant mass resulting in 10 g air dry weight. The air dry leaf were pulverized and extracted exhaustively with 80 % methanol. The macerate was concentrated and washed with equal volume of dichloromethane. The aqueous phase was concentrated and lyophilized to yield 3.762 g of “crude” lyophilized powder. The Crude lyophilized powder was dissolved in 20 ml water and partitioned with 20 ml (x2) ethyl acetate.
The lyophilized powders of all 3 fractions were tested in SW872 cells; % viability was determined using MTT and the equivalent Air dry weight and Fresh weight were calculated (triplicates). IC 50 in µg/ml
CRUDE Aqueous fraction Ethyl Acetate Fraction
lyophilized powder: 18.95 ± 0.0 17.27 ± 1.91 14.4 ± 0.83
Air dry Weight: 50.66± 0.0 55.30 ± 6.11 225.75 ± 12.80
Fresh weight: 112.2 ± 0.0 123.9 ± 13.62 502.9 ± 28.71
If expressed in terms of lyophised weight, the Ethyl acetate fraction appears to be more potent than the aqueous fraction. However, if expressed in terms of air dry or fresh weight aqueous fraction is more potent.
What is the best way to expressed IC50 values in order to choose which fraction needs to be further fractionated?
Based on the result displayed here, ethyl acetate fraction of the lyophilised is the most potent fraction. Therefore it should be fractionated further because it appears that its more concentrated with active compounds than other fractions. You should keep working on this fraction.
Dear Nawraj, The results can be expressed both in terms of lyophilized weight or air dry weight, as the case may be. Further, I agree with the answer of Dear Naveed that chance of loss of valuable bioactives is minimum in the lyophilisation process.
