Sample Preparation: Grounded samples of 10 g were extracted using 70% of ethanol (100 mL 1:10 w/v)) twice at room temperature for 24h and 6 h respectively. The mixture obtained were subsequently filtered with Whatman No. 5 filter paper. Filtrate obtained …show more content…
Defatted and dried sample (2.0 g) was ground separately and refluxed with 20.0 ml of 80% ethyl alcohol for 6.0 h. The extracts was filtered and made up to 25 ml. The samples (0.2 and 0.4 ml) and standard glucose (0.2-1.0 ml from 0.1mg/ml D-glucose stock) were taken in a series of test tubes and the volume was made up to 1.0 ml. A reagent blank was prepared by taking 1.0 ml water alone. Anthrone reagent (0.5g anthrone and 10g thiourea, in 1L 66% H2SO4) was added to all test tubes to a final volume of 5.0 ml and heated in a 90°C water bath for 15 min. The blue color developed was read at 620 nm in a UV-Vis spectrophotometer against the reagent blank. The amounts of soluble carbohydrates in samples were calculated from the glucose standard …show more content…
Samples were grinded to powder and passed through 0.5mm sieve. Sample were defatted and crude protein was estimated (Awan & Rahman, 2006) to determine the sample quantity taken for amino acid analysis. Diluted hydrochloric acid was used for Free amino extraction. Nitrogenous macromolecules that were co-extracted were precipitated using sulfosalicylic acid and removed by filtration. A mixture of performic acid and phenol was used for Oxidation at 0°C. Hydrolysates of the sample were then adjusted to pH 2.2 using 7.5M sodium hydroxide sol. Amino acid analyzer based on ion exchange chromatography was used to separate amino acids. Amino acids were determined by Post column derivatization of amino acids with ninhydrin and photometric detection was taken at 570 nm and 440 nm for