ObjectiveTo analyze the processing mechanism underlying the enhanced effect of invigorating spleen and stopping diarrhea of soil-fried Atractylodis Macrocephalae Rhizoma（AMR） by analyzing the changes of microstructure， chemical composition and anti-ulcerative colitis（UC） activity before and after soil stir-frying. MethodsThe microstructure and elemental composition of AMR before and after soil stir-frying were analyzed by scanning electron microscopy-energy dispersive spectroscopy（SEM-EDS）， to investigate the differences in microstructure and the underlying causes. Ultra-high performance liquid chromatography-quadrupole-time-of-flight mass spectrometry（UPLC-Q-TOF-MS） coupled with UNIFI 1.9.2 natural product analysis platform were used to analyze and identify the chemical constituents in raw and soil-fried products， and multivariate statistical methods including principal component analysis（PCA） and orthogonal partial least squares-discriminant analysis（OPLS-DA） were used to explore the differences and sources of chemical constituents between them. A dextran sulfate sodium（DSS）-induced UC mouse model was established. The method of disease activity index（DAI） was used to evaluate the severity of intestinal inflammation. Hematoxylin-eosin（HE） staining was used to observe the pathological changes of colon tissue， enzyme-linked immunosorbent assay（ELISA） was used to detect the levels of inflammatory factors， Real-time quantitative polymerase chain reaction（Real-time PCR） and Western blot were used to analyze the expressions of key genes and proteins involved in the intestinal mucosal barrier. The 16S rRNA sequencing was used to evaluate the diversity of intestinal flora， headspace gas chromatography-mass spectrometry（HS-GC-MS） was used to explore the levels of short-chain fatty acids（SCFAs） in feces. Base on the above findings， this paper investigated the effects of raw and soil-fried AMR on the biological， chemical， mechanical and immune barriers of model animals， and the differences in pharmacological effects and underlying mechanisms from the perspective of regulating the intestinal mucosal barrier in UC mice. ResultsSEM observation revealed numerous hearth soil particles on the surface of soil-fried AMR， accompanied by bubble-like bulges. At the same time， there were many cracks and folds on the surface of the hearth soil. EDS analysis revealed that the contents of Si， Al， Mg and Ca in soil-fried AMR were significantly higher than those of raw products， and these elements constituted the primary components of hearth soil. UPLC-Q-TOF-MS combined with database comparison was used to identify the chemical constituents of raw and soil-fried AMR. In positive ion mode， a total of 132 components were identified， primarily comprising three categories of terpenoids， polyphenols and amino acids. In negative ion mode， a total of 40 components were identified， primarily polyphenolic and glycoside compounds. Among them， the contents of sesquiterpenes and polyphenolic acids were changed significantly before and after processing. Soil-fried AMR could reduce the DAI score of UC mice， alleviate the shortening of colon length， reduce the levels of pro-inflammatory factors such as interleukin（IL）-17， IL-18， γ-interferon（IFN-γ） and tumor necrosis factor（TNF）-α in serum， increase the levels of anti-inflammatory factors such as secretory immunoglobulin A（sIgA）， IL-10， IL-4 and transforming growth factor-β（TGF-β） in serum， increase the expressions of key genes and proteins of intestinal mucosal barrier such as tight junction protein-1（ZO-1）， Occludin， Claudin-1 and mucin 2（MUC2） in colonic mucosa， and improve the disorders of intestinal flora diversity and the levels of SCFAs（P<0.05， P<0.01）. The raw and stir-fried products of AMR also exhibited the aforementioned effects， but they were weaker than the soil-fried products. Additionally， the auxiliary material hearth soil also had a certain pharmacodynamic effect. ConclusionSoil-fried AMR can enhance the protective effect on intestinal mucosal barrier in UC mice. These changes or heating-induced alterations in the microscopic structure and chemical composition of AMR may be attributed to the dual effects of adsorption of hearth soil.

ObjectiveTo analyze the processing mechanism underlying the enhanced effect of invigorating spleen and stopping diarrhea of soil-fried Atractylodis Macrocephalae Rhizoma（AMR） by analyzing the changes of microstructure， chemical composition and anti-ulcerative colitis（UC） activity before and after soil stir-frying. MethodsThe microstructure and elemental composition of AMR before and after soil stir-frying were analyzed by scanning electron microscopy-energy dispersive spectroscopy（SEM-EDS）， to investigate the differences in microstructure and the underlying causes. Ultra-high performance liquid chromatography-quadrupole-time-of-flight mass spectrometry（UPLC-Q-TOF-MS） coupled with UNIFI 1.9.2 natural product analysis platform were used to analyze and identify the chemical constituents in raw and soil-fried products， and multivariate statistical methods including principal component analysis（PCA） and orthogonal partial least squares-discriminant analysis（OPLS-DA） were used to explore the differences and sources of chemical constituents between them. A dextran sulfate sodium（DSS）-induced UC mouse model was established. The method of disease activity index（DAI） was used to evaluate the severity of intestinal inflammation. Hematoxylin-eosin（HE） staining was used to observe the pathological changes of colon tissue， enzyme-linked immunosorbent assay（ELISA） was used to detect the levels of inflammatory factors， Real-time quantitative polymerase chain reaction（Real-time PCR） and Western blot were used to analyze the expressions of key genes and proteins involved in the intestinal mucosal barrier. The 16S rRNA sequencing was used to evaluate the diversity of intestinal flora， headspace gas chromatography-mass spectrometry（HS-GC-MS） was used to explore the levels of short-chain fatty acids（SCFAs） in feces. Base on the above findings， this paper investigated the effects of raw and soil-fried AMR on the biological， chemical， mechanical and immune barriers of model animals， and the differences in pharmacological effects and underlying mechanisms from the perspective of regulating the intestinal mucosal barrier in UC mice. ResultsSEM observation revealed numerous hearth soil particles on the surface of soil-fried AMR， accompanied by bubble-like bulges. At the same time， there were many cracks and folds on the surface of the hearth soil. EDS analysis revealed that the contents of Si， Al， Mg and Ca in soil-fried AMR were significantly higher than those of raw products， and these elements constituted the primary components of hearth soil. UPLC-Q-TOF-MS combined with database comparison was used to identify the chemical constituents of raw and soil-fried AMR. In positive ion mode， a total of 132 components were identified， primarily comprising three categories of terpenoids， polyphenols and amino acids. In negative ion mode， a total of 40 components were identified， primarily polyphenolic and glycoside compounds. Among them， the contents of sesquiterpenes and polyphenolic acids were changed significantly before and after processing. Soil-fried AMR could reduce the DAI score of UC mice， alleviate the shortening of colon length， reduce the levels of pro-inflammatory factors such as interleukin（IL）-17， IL-18， γ-interferon（IFN-γ） and tumor necrosis factor（TNF）-α in serum， increase the levels of anti-inflammatory factors such as secretory immunoglobulin A（sIgA）， IL-10， IL-4 and transforming growth factor-β（TGF-β） in serum， increase the expressions of key genes and proteins of intestinal mucosal barrier such as tight junction protein-1（ZO-1）， Occludin， Claudin-1 and mucin 2（MUC2） in colonic mucosa， and improve the disorders of intestinal flora diversity and the levels of SCFAs（P<0.05， P<0.01）. The raw and stir-fried products of AMR also exhibited the aforementioned effects， but they were weaker than the soil-fried products. Additionally， the auxiliary material hearth soil also had a certain pharmacodynamic effect. ConclusionSoil-fried AMR can enhance the protective effect on intestinal mucosal barrier in UC mice. These changes or heating-induced alterations in the microscopic structure and chemical composition of AMR may be attributed to the dual effects of adsorption of hearth soil.

Objective To optimize the wine steamed Radix Rehmanniae processing technology.Methods The single factor experiment method was used to investigate the wine steamed Rehmannia glutinosa.Ultra-performance liquid chromatography-triple quadrupole tandem mass spectrometry(UPLC-QqQ-MS/MS)was used to determine the content of active ingredients such as stachyose tetrahydrate,manninotriose,melibiose,fructose,echinacoside,rhmannioside C,rhmannioside D,verbascoside,verbascoside,melittoside,monomelittoside,D(+)-raffinose pentahydrate,8-epicarnolic acid,catalpol,dihydrocatalpol,ajugol and so on.Through analytic hierarchy process and criteria importance through inter-criteria correlation(AHP-CRITIC),the content of the above chemical components and the color and sweetness values of rehmannia decoction pieces were used as evaluation indexes,and the amount of wine,steaming time and stuffiness time of steamed Rehmannia were investigated by a single-factor test method.The Box-Behnken response surface method was used to carry out the experimental design,and the divine Rehmannia processing process optimized.Results The optimal processing technology of wine steamed Radix Rehmanniae was as followed:20.6 g of alcohol per 100 g of herbs,1.5 h of stuffiness,and 6.5 h of steaming time.Conclusion The process of wine steamed rehmannia determined in this experiment is feasible,which can provide a reference for the research of processed rehmannia and provide certain guiding significance for the processed variety in industrial production.

Objective To investigate the effects of Rehmanniae Radix before and after processing on the intestinal flora of rats with kidney yin deficiency syndrome.Methods SD rats were randomly divided into blank group,model group,probiotic group(0.35 g·kg-1),high-/medium-/low-dose groups of Rehmanniae Radix Praeparata(3.5,1.75,0.875 g·kg-1),and high-/medium-/low-dose groups of Rehmanniae Radix(3.5,1.75,0.875 g·kg-1),with 9 rats in each group.Except for the blank group,rats in each group were injected intramuscularly with Dexamethasone Sodium Phosphate Injection(0.35 mg·kg-1)once a day for 21 days.The drug was administered by gavage once a day on the seventh day of modelling for 14 days.The adrenal histopathological changes were observed by using HE staining;the levels of serum cyclic adenosine monophosphate(cAMP),cyclic guanosine monophosphate(cGMP),corticotropin-releasing hormone(CRH),adrenocorticotropic hormone(ACTH),and corticosterone(CORT)were detected by ELISA;and the levels of short-chain fatty acids in the feces and changes in the diversity of intestinal flora were detected by a targeted metabolomic approach in conjunction with 16SrRNA sequencing in the rats in each group.Results(1)Compared with the blank group,the body mass of rats in the model group was significantly decreased on days 7,14 and 21(P＜0.05,P＜0.01);serum levels of cAMP,CRH,ACTH,CORT and the cAMP/cGMP ratio were all significantly increased(P＜0.01),and the cGMP content was significantly decreased(P＜0.01);and the adrenal cortex was thinned,with the boundaries of various layers of the cortex unclear.Compared with the model group,the body mass of rats in the Rehmanniae Radix Praeparata administration group on day 21 were all significantly increased(P＜0.05,P＜0.01),and the serum content of cAMP,CRH,ACTH,CORT and the cAMP/cGMP ratio were all significantly decreased(P＜0.05,P＜0.01),and the content of cGMP was significantly increased(P＜0.05,P＜0.01)in the rats;body mass of rats in the Rehmanniae Radix administration group did not change significantly(P＞0.05),CRH and CORT contents in serum of rats in the high-dose group of Rehmanniae Radix were significantly reduced(P＜0.01),and ACTH contents in serum of rats in the medium-dose group of Rehmanniae Radix were significantly reduced(P＜0.05);the adrenal cortex of rats in all the administration groups were improved,in particular,the thickening of the adrenal cortex layers was obvious in the Rehmanniae Radix Praeparata group,and the improvement effect was superior to that in the Rehmanniae Radix group.(2)Compared with the blank group,the difference of Coverage index was not statistically significant(P＞0.05),and the coverage of each group was good;the abundance index(Sobs,Ace,Chao)and diversity index(Shannon)of the model group were significantly increased(P＜0.01),and the Simpson index was significantly decreased(P＜0.01).Compared with the model group,Sobs index was significantly decreased in the medium-and high-dose groups of Rehmanniae Radix Praeparata(P＜0.05),Chao index was significantly decreased in the administered groups of Rehmanniae Radix Praeparata and high-dose group of Rehmanniae Radix(P＜0.05,P＜0.01),and Simpson index was significantly increased in the high-dose group of Rehmanniae Radix Praeparata(P＜0.05).The changes of Rehmanniae Radix on the richness and diversity of intestinal microbial community in kidney yin-deficient rats were small,while Rehmanniae Radix Praeparata could better maintain the stability of the richness and diversity of intestinal microbial community in kidney yin-deficient rats.(3)Compared with the blank group,the abundance of phylum firmicutes in the feces of the model group was significantly decreased,while the abundance of bacteroides and actinomycetes was significantly increased.The abundance of Lactobacillus was significantly decreased(P＜0.01),while the abundance of norank_f__Muribaculaceae and Bifidobacterium was significantly increased(P＜0.01).Compared with the model group,the trend of recovery of bacterial abundance in the probiotic group and the high-dose group of Rehmanniae Radix Praeparata was more similar to that of the blank group,which showed that it had the best regulating effect on the ratio of bacterial flora;the abundance of lactobacillus in all administered groups was increased,with that of the probiotic group was significantly increased(P＜0.01);the abundance of norank__f__Muribaculaceae and Bifidobacterium were all decreased,among which the probiotic group and the medium-and high-dose groups of Rehmanniae Radix Praeparata were significantly decreased(P＜0.01),and the effect was significantly superior to that of Rehmanniae Radix.The COG functions of the samples in each group were mainly focused on amino acid transport and metabolism,carbohydrate transport and metabolism,translation,ribosomal structure and biogenesis,replication,recombination and repair,but the abundance information of each function was different between groups,which may be due to the differences caused by dysbiosis of intestinal flora.(4)Compared with the blank group,the levels of acetic acid,butyric acid and propionic acid in the faeces of rats in the model group were significantly decreased(P＜0.05,P＜0.01),and the level of isobutyric acid was significantly increased(P＜0.01).Compared with the model group,the levels of acetic acid,butyric acid and propionic acid in the faeces of rats in the probiotic group and the low-,medium-and high-dose groups of Rehmanniae Radix Praeparata were significantly increased(P＜0.05,P＜0.01)and the levels of isobutyric acid were significantly decreased(P＜0.05,P＜0.01);although the above indexes in the Rehmanniae Radix group were improved,the difference was not statistically significant(P＞0.05).Conclusion The enhanced therapeutic effect of Rehmanniae Radix after processing on rats with kidney yin deficiency syndrome may be related to its adjusting effect on intestinal flora.

OBJECTIVE：To establish a method for the determination of 8 glycosides（astragaloside Ⅰ，Ⅱ，Ⅲ，Ⅳ and calycosin glucopyranoside ，2′-hydroxy-3′，4′-dimethoxy-isoflavan-glucoside and 9，10-dimethoxy-pterocarpan-glucoside） and 4 aglycones（calycosin，formononetin，7，2′-dihydroxy-3′，4′-dimethoxy-isoflavan and 3-hydroxy-9，10-dimethoxy-pterocarpan） in Astragalus membranaceus ，and to investigate the effects of different processing temperatures on the contents of above 12 components. METHODS ：The contents of 12 components in A. membranaceus and samples processed under different temperatures（120，140，160，180，200 ℃）were determined by UPLC-MS/MS. The determination was performed on ACQUITY UPLC HSS T 3 column with mobile phase consisted of 0.1 mol/L formic acid water solution -0.1 mol/L formic acid acetonitrile solution （gradient elution ）at the flow rate of 0.5 mL/min. The column temperature was 30 ℃. The detection wavel ength was 260 nm，and sample size was 2 μL. Electrospray ion source（ESI）was used under positive ion mode （ESI+）. The mass scanning range was mass ratio （m/z）of 50-1 500，with capillary voltage of 2 000 V and ion source temperature of 100 ℃. The desolvation temperature was 400 ℃；flow rate of atomizing gas （N2） was 40 L/h，and that of desolvation was 800 L/h；collision energy （CE）was 20-30 V；data acquisition rate was 0.5 s/scan. RESULTS：The linear range of astragaloside Ⅰ，astragaloside Ⅱ，astragaloside Ⅲ，astragaloside Ⅳ，calycosin-glucopyranoside， calycosin，ononin，formononetin，2′-hydroxy-3′，4′-dimethoxy-isoflavan-glucoside，7，2′-dihydroxy-3′，4′-dimethoxy-isoflavan，9， 10-dimethoxy-pterocarpan-glucoside and 3-hydroxy-9，10-dimethoxy-pterocarpan were 0.001 16-0.232 0，0.000 276-0.055 2， 0.000 22-0.044 0，0.000 225-0.045 0，0.000 734-0.587 0，0.001 17-0.234 0，0.000 742- 0.148 0，0.001 30-0.260，0.003 98-0.795 0， 0.000 476-0.476 0，0.001 89-0.378 0，0.000 336-0.336 0 μg（all R2≥0.999 2），respectively. The limits of detection were 6.2×10－6， 4.8×10－6，3.8×10－6，3.4×10－6，5.8×10－6，4.8×10－6，4.2×10－6，3.2×10－6，5.8×10－6，2.6×10－6，4.2×10－6，6.4×10－6 μg，respectively. The limits of quantitation were 12.6×10－6，16.2×10－6，14.4×10－6，14.8×10－6，18.8×10－6，16.4×10－6，15.4×10－6，10.8×10－6，20.2×10－6， 12.4×10－6，14.6×10－6，23.4×10－6 μg，respectively. RSDs of precision ，stability（24 h）and repetition tests were all lower than 3.0％（n＝6）. The average recoveries were 99.1％，100.2％，98.7％，101.9％，98.6％，102.1％，99.2％，100.3％，98.7％， 99.2％，99.3％ and 100.8％，with the RSDs of 1.9％，2.2％，2.4％，1.8％，2.1％，1.7％，2.3％，1.9％，2.4％，1.8％，2.2％ and 1.9％（n＝6），respectively. The results showed that the contents of astragaloside Ⅰ，Ⅱ and Ⅲ decreased gradually with the increase of processing temperature ；the content of astragaloside Ⅳ increased gradually with the increase of temperature. The content of flavonoid glycosides ，such as calycosin glucopyranoside ，ononin，2′-hydroxy-3′，4′-dimethoxy-isoflavan-glucoside and 9， 10-dimethoxy-pterocarpan-glucoside decreased with the increase of temperature ；the corresponding aglycone components as flavonoid glycosides ，formononetin，3-hydroxy-9，10-dimethoxy- pterocarpan increased firstly and then decreased with the increase ； the content of 7，2′-dihydroxy-3′，4′- dimethoxy-isoflavan decreased with the increase of temperature. CONCLUSIONS ：Established UPLC-MS/MS method can be used for determination of 12 components in A. membranaceus . After processed under different temperature，the contents of glycosides decreased in general ，while the contents of aglycones increased in general.