| Purpose:The processing technology of traditional Chinese medicine is the summary of clinical practice of medical scientists of past dynasties.Clinical practice has proved that processing can reduce the toxic and side effects of traditional Chinese medicine and play a vital role in clinical application.Belamcanda chinensis(L.)DC the dry rhizome of was first published in the"Shennong materia medica classic".It has the effects of clearing and detoxifying,eliminating phlegm and promoting swallowing.Belamcanda chinecsis has a small poison.Taking it for a long time makes people weak,and taking it more often makes people diarrhea.In order to further improve the safety and effectiveness of clinical medication and reduce the toxic and side effects of Belamcanda chinecsis,the research group conducted a preliminary study on the preparation of Belamcanda chinecsis from rice swill.It was found that the processing and drying of rice swill could significantly reduce the levels of ALT and AST in liver function of model animals and promote the secretion of amylase in rats;At the same time,due to the existence of rice swill,the inherent enzyme in Belamcanda chinecsis medicinal materials and the enzyme in rice swill can convert the flavonoid glycosides in Belamcanda chinecsis medicinal materials into glycosides to maximize the drug efficacy.Therefore,the processing of Belamcanda chinecsis medicinal materials with rice swill has a certain detoxification and synergistic effect.In order to explore the influence of rice swill on Belamcanda chinecsis medicinal materials and further clarify its processing principle,this study combines the traditional Chinese medicine theory of"bitter cold hurts the spleen"with modern pharmacology,and studies the detoxification and synergistic effect of rice swill processing Belamcanda chinecsis from the methods of intestinal,florapharmacodynamics,metabolomics,analytical and chemistry,so as to ensure the safety and effectiveness of clinical medication,to provide a scientific research basis for further replacing Belamcanda chinecsis raw products in clinical application.Material and method:1 The changes of pharmacodynamics and toxicity indexes of Belamcanda chinecsis raw products and rice swill before and after processing were determined by observing the general state of normal rats,measuring gastrointestinal hormones and digestive enzymes,detecting biochemical indexes of liver and kidney function and measuring intestinal flora.Based on the theory of"bitter cold"and"bitter cold hurts the spleen",the detoxification effects of raw Belamcanda chinecsis and rice swill before and after processing were studied.Sixty normal SD rats were randomly divided into 6 groups according to body weight:blank group,Belamcanda chinecsis raw product group,Belamcanda chinecsis processed product group,raw product aglycone group,processed product aglycone group and raw product glycoside group.The blank group,sham operation group and model group were perfused with the same amount of distilled water every day;The administration group was given each extract(equivalent to 20 g/kg)every day for 21 consecutive days.During the experiment,the mental state,food intake,fecal color and state of rats were observed every day;After the experiment,blood was taken from abdominal aorta and serum was separated for subsequent determination of gastrin,amylase and liver and kidney function;The spleen and liver of rats were taken for the subsequent calculation of spleen index and the pathological changes of liver and spleen were observed;1cm cecal tissue was put into sterile EP tube for subsequent 16S high increment sequencing experiment to determine the changes of intestinal flora.The results were analyzed to explore the detoxification mechanism of rice swill,so as to provide scientific research support for the safety of Belamcanda chinecsis in clinical application.2 The rat model of acute lung injury induced by LPS was used to compare the effects of lung pathological changes and lung dry/wet weight in rats,and determined the changes of pharmacodynamic indexes before and after drying processing.Non targeted"metabolomics"technology was used to compare the changes of serum metabolic components of Belamcanda chinecsis raw products and rice swill processed products in rats with ALI.Ninety normal SD rats were randomly divided into 9groups according to their body weight,namely blank control group,sham operation group,model control group,Suhuang Zhike capsule group,dexamethasone acetate group,raw Belamcanda chinecsis high and low dose groups,and rice swill Belamcanda chinecsis high and low dose groups.Rats in the non experimental group were given the same volume of distilled water.The rats in the experimental group were given dried raw extract(low and high)dose and processed extract(low and high)dose respectively.The positive group of traditional Chinese medicine was given Suhuang Zhike capsule.Western medicine positive group was given dexamethasone acetate tablets.The administration volume was 10 ml/kg,which was administered continuously for 5 days.Sixty mins after the last administration,the rats except the blank group were anesthetized by intraperitoneal injection of 6%chloral hydrate,fixed in the supine position on the operating board,disinfected the neck,cut the neck skin,exposed the trachea,drip LPS(4 mg/kg)into the airway,equal doses of normal saline were dropped in the sham group.Sew up after operation,continue the administration until the 7th day(48 h after modeling),and take materials.The lung tissue was taken from the right lung and fixed with 6%formaldehyde for subsequent observation of pathological changes and calculation of pathological score;The left lung was weighed and dried to calculate the dry wet weight ratio of rats.Blood was taken from abdominal aorta and centrifuged at low temperature to obtain serum for subsequent serum metabolomics determination.The results were analyzed to explore the synergistic principle of Belamcanda chinecsis processed with rice swill,so as to provide support for the effectiveness of clinical medication.3 Previous studies have proved that rice swill can reduce toxicity and increase efficiency after processing,but whether it can replace raw Belamcanda chinecsis in clinic is unclear.In this study,UPLC-Q-TOF-MS~Etechnology was used to compare the components of 16 different batches and different producing areas of Belamcanda chinecsis raw products and rice swill processed products.The feasibility of replacing raw Belamcanda was discussed from the difference of chemical composition.The selection of Belamcanda samples is representative.We selected Belamcanda from different regions and different ripening times to comprehensively evaluate the effective components of Belamcanda,so as to reduce the result deviation caused by regional limitations and different ripening times..The multivariate analysis methods of PCA,PLS-DA and OPLS-DA were used to analyze the chemical components of 16batches of dried raw products from different producing areas.Finally,the VIP value and loading diagram were used to determine the different components of chemical components before and after dried processing,so as to find the changes of chemical components after dried processing.Results:1 Comparison of toxic and side effects between Shegan raw products and rice swill processed productsIn the general state observation results of normal rats,we found that from the14th day after administration,the rats in the raw product group began to show signs such as fatigue,eating less,liking to pile up,irregular stool,yellowing hair color,etc.,and the rats in the raw product group and the raw glycoside group began to show slow weight growth at the 21st day,and the rats in the raw product group and the raw glycoside group had the least food intake.The results of spleen index showed that the raw group was lower than the blank group,while the processed group was higher than the blank group.The spleen index of the processed group was significantly higher than that of the raw group(P<0.05).There were no significant pathological changes in spleen and liver parenchyma in each group.The determination results of gastrin and amylase in serum showed that the secretion of gastrin and amylase in processed products group increased significantly,the secretion of gastrin and amylase in raw aglycones group and glycosides group decreased,and the decrease was more obvious in glycosides group(P<0.05),which was consistent with the results of our previous experiments.The results of liver and kidney function test showed that compared with the blank group,ALP and TBIL in the raw product group were significantly higher(P<0.05),and there was no significant difference between the processed product group and the blank group(P>0.05).ALT and AST in the biochemical indexes of liver function in the raw product group showed an upward trend,but there was no significant difference(P>0.05).There was no significant difference in the effects of other indexes of liver function between the raw product group and the blank group.BUN and CREA in each treatment group were lower than those in the blank group,but there was no significant difference(P>0.05).The results of intestinal flora study showed that the number of ASV/OTU in the five treatment groups was lower than that in the blank group,and the reduction of raw glycosides in Belamcanda chinecsis was the most obvious.In terms of species richness and diversity,there was no significant difference among the groups.Among the specific differential bacterial groups obtained by LEfse analysis,the differential bacteria between Belamcanda chinecsis raw product group and processed product group are uncultured Clostridium,Eubacterium,Eubacilli and Trichospirillaceae.Among them,the abundance of Eubacteriaceae and Eubacteriaceae increased significantly in the dried processed products group,while the abundance of trichospiroidaceae decreased significantly(P<0.05).The difference between the aglycones of raw and processed products of Belamcanda is Clostridium＿ⅩⅤⅢ.Uncultured Firmicutes and Eubacteria.The difference bacteria among the raw product group,raw product aglycone group and raw product glycoside group are Enterococcaceae,Enterococcus,Ruminococcus.For KEGG prediction pathway,there are two different prediction pathways between raw product group and processed product group:C5 branch dicarboxylic acid metabolism and RNA degradation;There are Nitrotoluene degradation,Porphyrin and Chlorophyll metabolism pathways between raw aglycones and processed aglycones;There are metabolic pathways such as Amino acid metabolism and Antibiotic resistance in the raw product group,raw product aglycone group and raw product glycoside group.2 Pharmacodynamic effects of raw and processed Belamcanda chinecsis products on ALI rats.The results showed that in the HE staining of lung pathology in ALI rats,the low-dose Belamcanda chinecsis processed products group was better than other Belamcanda chinecsis administration groups.The lung pathological score was consistent with the result of HE staining.In addition,no significant difference was found in the effects of dried raw and processed products on lung D/W weight ratio.In the comparison of serum metabolomics results between raw and processed Belamcanda chinecsis products,there are six Final differential components:N-acetylneuraminic acid,Muramic acid,Arachidic acid,L-methionine s-oxide,7-oxodeoxycholate,Dehydroepiandrosterone.In addition,among the total 92 different metabolic components compared between dried raw products and processed products,7 metabolic pathways involved had the largest influence values,namely Beta alanine metabolism,Arginine and Proline metabolism,Arginine biosynthesis,Pyrimidine metabolism,Phospholipase D signaling pathway,Prostate cancer,Linoleic acid metabolism Pantothenate and COA biosynthesis.3 Comparing 16 batches of Belamcanda chinecsis raw products from different producing areas with their rice swill processed products:PCA,PLS-DA and OPLS-DA analysis showed that there were significant differences in the composition between Belamcanda chinecsis raw products and rice swill processed products.Screening of differential compounds:the differential compounds in positive and negative ion modes were preliminarily identified in combination with VIP value and loading map,and compared with the map of the control group,they were preliminarily determined as iristectorigenin A、irigenin、Noririsflorentin、tectorige-origenin、iristectorigenin B.The contents of iristectorigenin A、Noririsflorentin in processed products decreased;The contents of tectorigenin、iristectorigenin B、irigenin increased.Conclusion:1 Based on the above conclusions,we analyzed that the general state,gastrointestinal hormones,digestive enzymes,liver and kidney function and intestinal flora environment of rats were changed due to the"bitter cold injuring the spleen"of ejaculation.After processed with rice swill,the toxic and side effects of Belamcanda chinecsis were reduced by improving the"spleen injury"effect of Belamcanda chinecsis.These changes may be caused by glycosides in Belamcanda.This effect may be due to the changes of gastrointestinal hormones,digestive enzymes,liver function and intestinal flora.It is suggested that the processed products of Belamcanda chinecsis rice swill have a certain detoxification effect.The above conclusions show that the processing of Belamcanda chinecsis can reduce the risk of clinical medication and increase the safety of medication.2 Among the pharmacodynamic indexes of Belamcanda chinecsis raw products and processed products in the intervention of ALI rats,Belamcanda chinecsis processed products were better than raw products group in terms of HE staining and pathological score of lung pathology in rats(P<0.05);In terms of improving pulmonary edema in rats,the processed group was better than the raw group.At the level of serum metabolomics,the small molecule metabolites in Belamcanda chinecsis changed after processing.The differential metabolic components were N-acetylneuraminic acid,Dehydroepiandrosterone,et al.These differential component-s are involved in the process of antiviral and anti-inflammatory in vivo,and can affect the metabolism of Amino acids and Unsaturated fatty acids.The differential components involve amino acid metabolism,pyrimidine metabolism and other metabolic pathways.It indicates that there may be differences in the regulation of these metabolic pathways between raw products and processed products,and the specific quantitative experiment needs to be further studied.The above suggests that the processed rice swill products at the metabolomic level play a better pharmacodynamic role through the regulation of small molecular metabolic components in the body,which provides important significance for further clinical application.3 The application of UPLC-Q-TOF-MS~Etechnology and multivariate analysis method,the results showed that there were differences in chemical composition between 16batches of Shegan raw products and rice swill processed products from different places of origin.The difference of these components after processed with rice swill is the change of the content of aglycones,among which the content of tectorigenin and irigenin,which play the main anti-inflammatory and antitussive effects.Therefore,it is inferred that the medicinal effect of rice swill may be increased after it is processed and dried.However,according to the previous research results in the laboratory,aglycones are difficult to dissolve in water.Whether the medicinal effect of rice swill processed and dried with water decoction is better than that of raw products remains to be further discussed. |
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