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The Screening of Asiaticosides Microbial Transformation Strain and the Purification, Identification of Transformation Products

Author: DuPeng
Tutor: MaoRenGang
School: Shanghai Normal University
Course: Microbiology
Keywords: bioconversion/biotransformation centella asiatica asiaticosides strain isolation separation purification identification
CLC: R284
Type: Master's thesis
Year: 2010
Downloads: 111
Quote: 3
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Abstract


Centella asiatica saponins revealed important medicinal values, as other saponins their bio-availability is lower. To use microbial fermentation of glycosides of Traditional Chinese Medicine has become aresearch hotspot for improving their bio-availability.In this paper, after screening with esculin, and re-screening, the best microbial strains which can transform the total saponins in the centella asiatica were selected from the soil from different environment. Then the best microbial strain was identified as Penicillium citrinum with morphology, physiology and biochemistry, molecular biology.The methods of TLC and HPLC were discussed for getting a more accurate qualitative and quantitative analysis result. Finally, the best developing solvent of TLC is chloroform: methanol: water=10: 3: 0.2 v/v and the best condition for HPLC was obtained by experiments as: column is Phenomex Luna C18(250mm×4.6mm, 5μm);solvent is methanol(A): water(B)=55: 45 v/v; column temperature is 30℃; the detection wavelength is 205nm; the flow rate is 0.8ml/min.Two aim samples of higher purity were also gained isolated and purified using the methods of Silica Gel Column Chromatography. Two products were identified as 2α,3β,6β,23α-trihydroxy-urs-12-ene-28-O-β-D-glucopranoside (A) and 2α,3β,6β,23α-tetrahydroxy-urs-12-ene-28-oic acid-28-O-β-D-glucopranoside (B) by melting point, optical rotation, high-resolution mass spectrometry, IR, 13CNMR, 1HNMR. The compound B has not been reported yet.Methodological experiments have verified the reliability and accuracy of the HPLC testing analysis. The HPLC method showed a good linear relationship in the range of transformation products were 2.05～10.26μg( r=0.9999) and 2.54～12.68μg( r=0.9998). The average recoveries were 101.1% with RSD 1.3% (n=6) and 101.0% with RSD 0.9% (n=6). It provided a simple and fast way for measurement and analysis of the transformation products from total glucosides of centella asiatica.

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CLC: > Medicine, health > Chinese Medicine > Of Pharmacy > Traditional Chinese medicine chemical
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