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Mothed of Analysis of Ginkgo Biloba Extract
Total flavonol glycosides, determine according to HPLC
Chromatography conditions and system suitability: using octadecylsilane bonded silica gel (C18)as filling agent; methanol : 0.4% phosphoric acid solution ( 50:50) as mobile phase. Detection wavelength is 360nm and the number of the theoretical plates of the column is not less than 2500.
Reference solution: accurately weigh reference quercetin, kaempferol and isorhamnetin previously dried overnight over phosphorous pentoxide, add methanol to produce three solutions containing 0.03mg/ml. 0.03mg/ml, 0.02mg/ml respectively as the reference solutions.
Test solution: accurately weigh 35mg of the test material, added the mixed solution of methanol:25% hydrochloric acid (4:1), heated under reflux in water bath for 2 hours, quickly cooling to room temperature and transfered to a 50ml volumetric flask, diluted with methanol to the scale and shake well. Filter with microporous membrane ( 0.5um), take the filtrate as testing solution
Procedure: accurately inject 10ul each of the above three reference solutions and the test solution into the column separately, and determine the contents of quercetin, kaempferol and isorhamnetin respectively. Calculated the content of total amount of flavonol glycosides using the following equation.
The content of total amount of flavonol glycolsides=( the content of quercetin + the content of kaempferol + the content of isorhamnetin) X 2.51
Terpenoid lactones: carry out the test according to HPLC method ( appendix VI D)
Chromatography conditions and system suitability: Using octadecylsilane bonded silica gel(C18)as filling agent and methanol:water(35:65) as mobile phase, detected by refrative index detector. The number of theoretic
al plate should not be less than 2500 calculated on the basis of terpenoic lactone peak and the separation degree between the peak of bilobalide and the C peak of ginkgolide should be greater than 1.5.
Reference solution: weigh accurately proper quantities of bilobalide, ginkgolide A, ginkgolide B and ginkgolide C, add methanol to produce per ml containing 2mg, 1mg, 1mg of mixed reference solution .
Test solution: weigh accurately about 150mg of the test materials, added 10ml water and heated in water bath to dissolve, added 2 drops of 2% hydrochloric acid and extracted 4 times under shaking with ethyl acetate( 15ml,10ml,10ml,10ml), combined the phase of ethyl acetate and washing with 20ml of 5% sodium acetate solution. Separately take sodium acetate solution and wash again with 10ml ethyl acetate. Combine ethyl acetate extract solution with wash solution and wash 2 times with water each 20ml. Separately take washed solution and wash with 10ml ethyl acetate. Combine ethyl acetate solutions, recover ethyl acetate to dryness. Dissolve the residue with acetone and transfer to a 5ml volumetric flask and add acetone to scale and shake well.
Procedure: Pipet accurately 10ul,15ul of reference solution and 10ul of test solution respectively,
Inject into column and determine. Calculate the content of bilobalide, ginkgolideA, ginkgolide B and ginkgolide C respectively using the external standard two-point log equation method.
It contains terponid lactones not less than 6.0% in form of the sum of bilobalide( C15H18O8), ginkgolide A( C20H24O9), ginkgolide B( C20H24O10), ginkgolide C( C20H24O11), calculated on the dried basis.
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