Effects Of Processing Methods On The Components Of Polygonatum Polysaccharide And The Related Pharmacological Effects Of "Invigorating Qi And Nourishing Yin"

Objective:Polygonatum sibiricum(PS)is a kind of Medicine-Food traditional Chinese medicine.It has been widely used in clinic,In history,there were many processing methods of PS,such as single steaming,double steaming,nine cycles of steaming and solarization.By the Qing Dynasty,"nine cycles of steaming and solarization" was the mainstream method for the processing of PS.Due to the complexity and time-consuming of the nine cycles of steaming and solarization process,in order to reduce costs and save time,the single-steaming or two-steaming process is mostly used in the commercial PS extract.Modern research shows that the steaming time and times of PS have great influence on the chemical composition of PS.After about 25h of steaming(fourfold processed PS)some of the chemical components reach the maximum value and then decrease.However,it is not clear what effect the change of chemical composition will have on the pharmacodynamics.Therefore,this experiment simulates the ancient method of "nine cycles of steaming and solarization" to produce fourfold processed PS and ninefold processed PS.At the same time,we select the commercial PS(in line with the processing specifications of Zhejiang Province)as the object of this study to investigate the differences in chemical composition and pharmacodynamics of different processed products of PS.Finally,in vitro experiments were performed to verify the efficacy of the relatively rare fourfold processed PS.To provide reference for clinical use to select appropriate processed products.Methods:1.Preparation and chemical content determination of different processed products of PS:Simulated the ancient method of processing(nine cycles of steaming and solarization),using modem equipment,to obtain fourfold and ninefold processed PS.Observe the appearance,color,smell and taste of four different kinds of processed products of PS(raw PS,commercial PS,fourfold processed PS,ninefold processed PS).The water content is determined by the drying method,the alcohol soluble extract is determined by hot dip method,the polysaccharide content and the reducing sugar content is determined by the anthrone-sulfuric acid method.2.Effect of different processed products of PS on the deficiency of both qi and yin model rats:The rat model of qi and yin deficiency was established by continuous modeling for 2 weeks using 3 factors(drugs with hot nature,food restriction,weight-bearing swimming).Rats with successful modeling were randomly divided into model group,positive groupP the commercial PS grouP,fourfold processed PS group and ninefold processed PS group.Each group was given 10g/kg PS water extractor of different processed products or positive drug(sheng mai yin)for 6 weeks.Observe the general behavioral indicators of deficiency of both qi and yin rats during the administration,such as body weight,tail diameter,surface temperature,grip strength,pain threshold and thermal imaging.After the last administration,detection Hb by tail tip blood.Collection blood samples from orbital to detect ALT,AST,GLU,TG,Cr;and detect IgA,IgM,cAMP,cGMP by Elisa.After 6 weeks of administration,stop the administration and modeling,and the rats were recovered without intervention for 4 weeks,the weight changes were observed,and the effects of different processed products on the Qi-Yin deficiency model rats were compared.3.Study on the protection of premature senescence of human embryo lung diploid fibroblast(2BS)cells by polygonatum polysaccharide from fourfold processed PS:Adopt fourfold processed PS from the first part of the experiment,extract its polygonatum polysaccharide(FPP)for in vitro cell experiments by ultrasonic extraction.Treate the 30-generation(PD)2BS cells with different concentrations of FPP(0.01,0.1,1 mg/ml),after 2h,treate some of them with H2O2 for 3 days,and culture others for 72 hours.After the end of the culture,measure the cell viability by MTT assay,and determine the staining rate of?-galactosidase(SA-?-gal)in 2BS cells.Results:1.Preparation and chemical content determination of different processed products of PS:?The appearance,color,cross-section,odor and taste of PS processed by different methods are different.With the increase of steaming times,the feeling of hemp tongue disappears,the smell is sweet first,then bitter slightly,and the color of cross-section and surface is gradually deepened,turning to lacquer black.?The water content of the same processed products is slightly different.The water content of raw PS,commercial PS,fourfold PS and ninefold PS were 12.29%,14.00%,12.45%and 11.77%.?The content of alcohol-soluble extracts in different processed products is different.With the increase of steaming times,the content of alcohol-soluble extracts first increased and then decreased.The content of extract of raw PS,commercial PS,fourfold PS and ninefold PS were 22.14%?46.78%?64.30%?53.63%.?The content of total polygonatum polysaccharide in raw PS and steamed PS has significant difference.The content of total polysaccharide of raw PS,commercial PS,fourfold PS and ninefold PS were 7.31%,1.99%,1.51%and 1.52%.After steaming,the content of polygonatum polysaccharide decreased significantly.With the increase of steaming times,the content of polygonatum polysaccharide gradually stabilized.?The content of reducing sugar in different processed products of PS is quite different.The content of reducing sugar of raw PS,commercial PS,fourfold PS and ninefold PS were 13.00%?14.43%?51.69%?37.00%.With the increase of steaming times,the change trend of reducing sugar content is similar to that of extract content.The content of reducing sugar in fourfold processed PS is the highest.2.Pharmacodynamical research of different processed products of PS on Qi-Yin deficiency model rats:Modeling for 2 weeks,the average body weight of the model group was 174.90g,the normal group was 355.22g,only 1/2 of the normal group,showing the symptoms of emaciation in deficiency of both qi and Yin;the swimming time of the rats decreased significantly,the average swimming time decreased from 22.14min to 5.47min,showing the fatigue in deficiency of both qi and Yin Symptoms of physical or mental distress.At the same time,the GLU level of the model group was significantly reduced,the surface temperature was significantly increased,and the symptoms of deficiency of Qi and Yin such as impatience,sensitivity,irritability,and hair become no gloss,which indicated that the model was successful.After 6 weeks of administration,compared with the model group,the effects of different processed products of PS on the model rats of deficiency of Qi and Yin:?The commercial PS can significantly improve the general physical signs(pain threshold,surface temperature,grip strength,thermal imaging),blood biochemical level(AST,GLU,TG),protein level(Mb,Hb),immune system(IgA,IgM)and cyclic nucleotide system of rats with deficiency of Qi and Yin.?The fourfold PS can significantly improve the general physical signs(body weight,tail diameter,pain threshold,surface temperature,grip strength,thermal imaging),blood biochemical level(ALT,AST,GLU,TG),protein level(Mb,Hb),immune system(IgA,IgM),cyclic nucleotide system of rats with deficiency of both qi and Yin,and has a better recovery effect on the body weight of rats after stopping the drug.?The ninefold PS can significantly improve the general physical signs(body weight,pain threshold,surface temperature,grip strength),blood biochemical level(ALT,AST,GLU),MB and IgA of rats with deficiency of Qi and Yin.? Among the indexes,the comparison of different processed products shows that the commercial PS has certain advantages in improving hemoglobin and immunoglobulin;the fourfold PS have certain effects in various indexes,especially in regulating blood lipid metabolism and recover body weight;the nine preparations have better effects in improving liver function and immunoglobulin IgA.3.Study on the anti-H2O2-induced aging of human embryonic lung diploid fibroblast 2BS cells by polygonatum polysaccharide from PS:?polygonatum polysaccharide from fourfold processed PS(0.01?0.1mg/ml)can delay the natural aging of 2BS cells and promote cell proliferation.High dose shows slight inhibition.?The medium dose of polygonatum polysaccharide from fourfold processed PS can resist the senescence of 2BS cells caused by sublethal dose of H2O2 treatment and reduce apoptosis.Low dose have similar effects,but no significant differences.High doses show the opposite effect.?The medium dose of polygonatum polysaccharide from fourfold processed PS can significantly reduce the positive rate of SA-?-gal staining in 2BS cells treated with sublethal dose of H2O2.Conclusion:1.The appearance,color,odor,taste,extract content,total polysaccharide content and reducing sugar content of different processed products of PS(raw PS,commercial PS,fourfold processed PS and ninefold processed PS)are quite different.The total polysaccharide content of raw PS is the highest,the extract and reducing sugar content of fourfold processed PS is the highest.2.Different processed products of PS can improve the general signs(body weight,grip,diameter of tail root,surface temperature,thermal imaging,pain threshold),oxygen supply capacity(decrease Mb,AST,increase Hb),liver function(decrease AST,ALT)and glycolipid metabolism(decrease TG,increase GLU),increase serum IgA,IgM,regulate cyclic nucleotide system(cAMP/cGMP)and help rats recover their body weight.Commercial PS has certain advantages in improving hemoglobin and immunoglobulin;the fourfold processed PS has certain curative effect in all indexes,especially in regulating blood lipid metabolism and recover body weight;the ninefold processed PS has outstanding effect in improving liver function and IgA.Different processed products have their own advantages and disadvantages.Appropriate selection should be made during clinical use.3.The polygonatum polysaccharide from fourfold processed PS can resist the premature senescence of cells caused by sublethal dose of H2O2,protect 2BS cells,delay the natural senescence of 2BS cells and promote cell proliferation.