Microbial Diversity Analysis And Pharmacodynamics Study Of Liushenq

Traditional Chinese medicine fermentation processing has a long history of the application in our country.Medicated leaven,also know as Liu Shenqu,is made from flour,wheat bran,red bean,bitter almond,Artemisia annua,Xanthium sibirieum,Polygonum hydropiper fermentation by a certain percentage.Its curative effect is promoting digestion and relieving stasis,strengthening the spleen and stomach.As its curative effect is very significant,it is one of the most widely used traditional fermented medicine.Currently,the traditional way of medicated leaven production is natural fermentation.As it’s process of ferment restricted by season and relying on subjective experience,leading to poor sanitation and unstable quality.Some miscellaneous bacterium will produce toxic metabolites,such as aflatoxin,which affects the application of medicated leaven.Now,most of the research selects strains for fermentation witch is based on traditional culture purification technology.It can’t parse microbes contained in medicated leaven fully.At the same time,enzyme activity(amylase and protease)or medicinal active ingredients are mostly known as indexes for quality evaluation,this methods is inaccurate.So the important premises of medicated leaven are understanding microbial components and their differences fully and searching for a pharmacodynamics index that can evaluate the quality of medicated leaven and ascertain the optimal fermentation strain.The main research contents of this study are as follows:(1)Use high-throughput sequencing technology to evaluate microbial diversity of medicated leaven.The results show that:Microbiological compositions of medicated leaven from various producing areas are different,and dominant microbial communities are existed,amylase activity and pH known as indexes for evaluating medicated leaven are unilateral.Dominant microorganism from Anhui、Sichuan、Shandong medicated leaven are Lactobacillus and Candida,but their abundance are different.Dominant microorganism from Fujian、Hebei、Shaanxi、ZheJiang medicated leaven respectively are Weissella and Candida;Pantoea、Pseudomonas and Alternaria;Lactobacillus and Aspergillus;Pantoea、Pseudomonas and Aspergillus.(2)Use HPLC to compare medicated leaven fingerprint from various producing areas,and detect the content of aflatoxin B1.The results show that:Compared with water decoction,the fingerprint of methanol extract contains more common peaks and its similar degree are also higher(compared with the reference fingerprint).The raw materials of lab-fermented medicated leaven are identical,but fermentative strains are different,the similar degree are low,it shows fermentative strains could make the same chemical composition translate different chemical compositions.Combine with the results of microbial diversity analysis,we speculate the low abundant microflora play a very important role in the process of fermentation.The content of aflatoxin B1 from different medicated leaven are not overweight.(3)Investigate the effects of medicated leaven on activity and ODC mRNA expression of IEC-6 cell.The results show that:The effect that methanol extract promots IEC-6 proliferation is far superior to water decoction,and there is a dose effect relationship.Combine with the results of,we find there are more effective components in methanol extract and the fingerprint of methanol extract could show chemistry changes preferably.Methanol extract of medicated leaven from Sichuan、Hebei、Shaanxi and synergistic fermentation medicated leaven could raise ODC gene expression observably,and IEC-6 cell survival rate is positively related to ODC gene expression,this result further prove IEC-6 proliferation can be used as a pharmacodynamics index that can evaluate the quality of medicated leaven and ascertain the optimal fermentation strain.(4)Culture separately and purify dominant microorganism of leaven production from Sichuan、Shaanxi and use these microorganism for pure breed fermentation,screen of strains of fermentation with the activity of IEC-6 cell.The results show that:Find 6 types microorganism,are Lactobacillus pontis、Aspergillus niger、pediococcus acidilactici、Pediococcus pentosaceus、Cryptococcus neoformans、Lichtheimia ramose.Forthe effect that promots IEC-6 proliferation,fungus are slightly superior to bacteria,the best fermentation strain in turn:Aspergillus niger≈ Lichtheimia ramosa>Pediococcus acidilactici>Lactobacillus pontis>Pediococcus pentosaceus...