Emodin-8-o-beta-d-glucosidase Preparation Methods To Explore

Emodin-8-O-β-D-glucopyranoside exists in herbs such as Radix et RhizomaRhei, Polygonum multiflorum Thunb., Polygonum cuspidatum Sieb.et Zucc. etc. Atpresent, studies have shown that its true right brain acetylcholinesterase activitywas reversible inhibition. Emodin-8-O-β-D-glucopyranoside can not only enhancethe content of central acetylcholine, but also be of a protective effect to injuryhippocampal neurons, therefore, emodin-8-O-β-D-glucopyranoside is a truereversible inhibitor of acetylcholinesterase, meanwhile is hippocampal neuronsinjury agent, so can improve a number of areas of Alzheimer's disease, such as theincidence and development. Emodin-8-O-β-D-glucopyranoside significantlyimproved disfunction by scopolamine lead to learning and memory in mice, andsignificantly improve learning and memory in normal mice.Compared with thepositive drug Huperzine A, there was no significant difference between them.Therefore, emodin-8-O-β-D-glucopyranoside has a good development prospectsfor senile dementia and mental raise.Herbs contain a certain amount of emodin-8-O-β-D-glucopyranoside,whichcontain emodin (Polygonum cuspidatum, Polygonum multiflorum, Radix et Rhizo-ma Rhei, Cassia tora L. or Cassia obtusifolia L., etc.), but the content of emodin-8-O-β-D-glucopyranoside is low. At present, biotransformation has become auseful tool of natural product structure modification and innovation.It is achemical reaction which modifies structures of substrate using enzymes of thebiological metabolism process. Biotransformation has advantages of highselectivity, mild conditions, unpollution, low cost, few side products. Therefore,the issue will make some research on preparations of emodin-8-O-β-D-glucopy-ranoside by biotechnology.It is of significance to research and develop drugscombating Alzheimer's for the gradual aging society.This paper is divided into four parts,including summary of related literatures,research on Preparation of emodin-8-O-β-D-glucopyranoside, content determ-ination of emodin-8-O-β-D-glucopyranoside from Polygonum multiflorum Thunb."by HPLC and summary and discussion.The first part: summary of related literaturesFirstly studies on chemical constitutes and pharmacological activities of Poly- gonum multiflorum and PoIygonum cuspidatum Sieb.et Zucc.were reviewed, andthen application and development of biotransformation and extraction by enzymein pharmacy were reduced and concluded.The second part: research on Preparation of emodin-8-O-β-D-glucopyranosi-deChapter one: preparation of emodin-8-O-β-D-glucopyranoside by routine me-thodemodin-8-O-β-D-glucopyranoside, physcion-8-O-β-D-glucopyranoside, emo-din and polydation were isolated and purified from Polygonum multiflorum andPolyg onum cuspidatum Sieb.et Zucc.by solvent extraction colomn-chromatogra-phy.Chapter two: preparation of emodin-8-O-β-D-glucopyranoside by biotransfor-mation1 Conversion emodin into emodin-8-O-β-D-glucopyranoside by plant cellculturesemodin-8-O-β-D-glucopyranoside was produced through biotransformation ofemodin by plant cell cultures for the first time, and the best conditions were select-ed for biotransformation.The method of content determination of emodin-8-O-β-D-glucopyranoside inCatharanthus roseus cell suspension cultures was established for the first time.Method: column: DiamonsiL^TM (Diamond) C₁₈ column (250mm×4.6mm,5μm);mobile phase: methanol-water-formic acid (70: 29.6:0.4) V/V; the rate of flow:1 mL·min^-1. detection wavelength: 280nm; temperature of column: room temper-ature. The regression equation: Y=3E+06X-54.143, the correlation coefficient r=0.9998, the linear range of emodin-8-O-β-D-glucopyranoside was: 0.015μg～0.075μg.2 Extraction process of combine anthraquinone from herbs by biologicalenzyme tecnology.the optimum extraction process of combine anthraquinone from Radix etRhizoma Rhei by cellulase was screened by orthogonal experimental for the firsttime, which was appraised by weighted evaluation method with two indicators ofaverage content determination of combine anthraquinone and the rate of paste, Thescore (X) of average content determination of combine anthraquinone (%) is forthe right to 60% and the score (Y) of paste rate is for the right to 40%,the overallscore is that Z=60%X+40%Y. So the optimum extraction process is that 40℃,hydrolysis for 1h, and pH=7.3 Screening of derivation of the substrate 6-acetylemodin by microorga-nism 6-acetylemodin is the substrate for preparation of emodin-8-O-β-D-glu- copyranoside by microorganism. In this experiment the method of screening singlefactor was employed, with reference to the findings to IR spectrometry and resultsof TLC, the best experimental conditions of 6-acetylemodin was screened:Preparing emodin in home flask, dring for 1 h at 50℃, adding acetic anhydride byemodin-reagent (1:3), keeping in the boiling water bath for 30 minutes then addingcold water 100mL quickly, vibrating, refrigerators placed 24h, vacuum filtering,watering to no sour, 50℃dring 50min.The third part: Content determination of emodin-8-O-β-D-glucopy ranosidefrom Polygonum multiflorum Thunb.Method of HPLC determination of emodin-8-O-β-D-glucopyranoside fromPolygonum multiflorum Thunb. was established for the first time: column: Diam-onsil^TM (Diamond) C₁₈ column (250mm×4.6mm,5μm); mobile phase: methanol-water-formic acid (70:29.6:0.4) V/V; the rate of flow: 1mL·min^-1;detection wa-velength: 280nm; the temperature of column: room temperature. The regressionequation: Y=3E+06X+2187.5,with a correlation coefficient of r=0.9999, emodin-8-O-β-D-glucopyranoside linear in the range of 0.060μg～0.300μg. results of det-ermination show that the method is simple and accurate, with good repeat-abilityand recovery. So the method can be used in the content determination ofemodin-8-O-β-D-glucopyranoside from Polygonum multiflorum Thunb.The conte-nt determination of emodin-8-O-β-D-glucopyranoside is 0.086%.The fourth part: conclusion and discussion...