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Non-alcoholic fatty liver disease （NAFLD） is a chronic liver disease with complex and diverse pathogenesis， and there is no effective treatment or specific drugs for its clinical treatment. In recent years， its incidence has been on the rise， and it has become the earnest expectation of medical researchers in China and abroad that related patients could be treated. AMP-activated protein kinase （AMPK） functions to regulate cellular energy homeostasis and mitochondrial homeostasis. When activated， it has a good intervention effect on NAFLD progression with lipid metabolism disorders and mitochondrial homeostasis disorders. For NAFLD， the activation of AMPK can inhibit the production of new lipogenesis in the liver， promote the oxidation of fatty acids in the liver， and enhance the mitochondrial function of adipose tissues. As a key target of metabolic diseases， AMPK can also improve apoptosis， liver fibrosis， autophagy， and inflammation. Traditional Chinese medicine （TCM） is good at treating diseases from multiple targets and multiple pathways and is also commonly used in the treatment of chronic liver disease in clinical practice. A large number of in vitro and in vivo experimental studies on NAFLD have shown that TCM monomers have good prospects for the treatment of NAFLD through the AMPK signaling pathway， including glycosides， phenols， alkaloids， flavonoids， quinones， terpenoids， and lignans， which are natural activators of AMPK. This study reviewed the research progress on TCM monomers in regulating the AMPK pathway to prevent and treat NAFLD， providing a broader perspective for TCM treatment of NAFLD.
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Diabetic nephropathy （DN）， the most feared microvascular complication of diabetes and one of the most common and serious complications of diabetes， is the major cause of end-stage renal disease worldwide， leading cause of morbidity and mortality in patients with diabetes， and the main non-communicable cause of death worldwide. There are many types of saponins， which are the main bioactive components of various Chinese medicinals. They have various pharmacological activities such as lowering blood glucose and blood lipids， improving insulin resistance， anti-inflammation， anti-oxidative stress， anti-tumor， and immune modulation. In recent years， it has been frequently verified that the saponins in Chinese medicinals have definite effect in regulating DN， showing multi-target， multi-pathway， multi-system， multi-effect characteristics. Thus， they have broad prospects in the prevention and treatment of this disease. There has been an explosion of research on the treatment of DN with saponins in Chinese medicinals in vivo and in vitro， but there is a lack of systematic and comprehensive summary. Therefore， this study summed up the studies of saponins in Chinese medicinals in the intervention of DN and summarized the mechanisms such as improving glucolipid metabolism， inhibiting oxidative stress， anti-inflammation， anti-apoptosis， regulating autophagy， anti-fibrosis， and protecting podocytes， with a view to providing ideas and references for the development of drugs related to DN.
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Embryo implantation is one of the most critical steps in the reproductive process. The failure of embryo implantation to continue development is one of the important reasons leading to infertility. The success of embryo implantation depends on the high receptivity of endometrium and the embryo with implantation ability. Autophagy is a process in which cytoplasm， organelles， and inclusions are absorbed by double-membrane vesicles and transported to lysosomes for degradation and recycling， which is a way to maintain the homeostasis. A large amount of evidence have shown that autophagy plays an important role in all aspects of embryo implantation. Based on this， this paper explored the relationship between autophagy and endometrial receptivity and embryo implantation ability. According to the latest research progress， this paper combed 5 mechanisms （promotion of decidualization of endometrial stromal cells， promotion of apoptosis， regulation of hormone levels， coordination of inflammation， and promotion of ovulation） of 14 kinds of Chinese medicine monomers， including emodin， catalpol， paeoniflorin， resveratrol， folic acid， zearalenone， curcumin， wogonin， quercetin， chrysin， berberine， apigenin， phisetine， and kaempferol， in regulating different links of autophagy intervention in embryo implantation. This paper is expected to provide references and ideas for future Chinese medicine monomers to improve the success rate of embryo implantation.
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Colorectal cancer （CRC） is one of the refractory malignant tumors of the digestive system worldwide. With limitations， the available clinical therapies are usually suspended or show unsatisfactory effect. Therefore， it is urgent to find and develop new candidate drugs specifically targeting the cancer with ideal efficacy， low toxicity， and low cost， and the solutions can be found in traditional Chinese medicine （TCM） which has a long history. In TCM， sovereign， ministry， assistant， and guiding medicinals are selected based on the syndrome differentiation， and it has shown remarkable efficacy on CRC in recent years. In particular， Chinese medicinal compounds and monomers from Chinese medicinals which have been applied in clinical settings are advantageous in the treatment of CRCs， as they improve the quality of life， alleviate clinical symptoms and toxic and side effects of chemotherapy， and prolong the survival of patients. Therefore， we retrieved the English and Chinese articles with "CRC"， "TCM"， "compound" and "monomer" as keywords， and summarized the progress in the treatment of CRC with Chinese medicinal compounds and monomers from Chinese medicinals from four aspects of "replenishing Qi and invigorating spleen"， "clearing heat and removing toxin"， "nourishing liver and kidney"， and "tonifying Qi and nourishing blood". However， Chinese medicine features multiple components， multiple targets， and multiple pathways， and in-depth research should be carried out on the application of Chinese medicinal compounds and monomers from Chinese medicinals in the treatment of CRC， in an attempt to minimize the pain and side effects and maximize the therapeutic effect. This study is expected to provide new insight into the treatment of CRC and a reference for further research on the efficacy and mechanism of Chinese medicine.
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Diabetic kidney disease （DKD） is characterized by the hyperfiltration and albuminuria in the early phase which are followed by progressive renal function decline， renal tubular epithelial cell hypertrophy， and tubulointerstitial fibrosis. Thus， it is one of the leading causes of chronic kidney diseases. The currently available therapies mainly aim to control the primary diseases and reduce the risk of kidney injury. Based on syndrome differentiation， traditional Chinese medicine （TCM） relieves the symptoms by excreting water and alleviating edema and eliminates the root cause by tonifying deficiency and supplementing the original Qi， thereby showing therapeutic effect and delaying the progression of DKD. It excels in comprehensively regulating the constitution of patients with little side effects. Among the Zang-fu organs， kidney takes the second place in the content of mitochondria which participate in the metabolism of water and fluid and are the foundation of kidney Yin and kidney Yang. Mitochondria are energy producers within a cell， which carry out cellular respiration， produce reactive oxygen species， and generate adenosine triphosphate by oxidative phosphorylation. Mitochondrial quality control （MQC） is an effective way to maintain mitochondrial dynamic balance， whose imbalances， such as mitochondrial oxidative stress， mitophagy， mitochondrial dynamic changes， and abnormal calcium regulation， are related to the occurrence and development of DKD. It is generally believed that the destruction of mitochondrial structure in the case of metabolic disorder is the main cause of the disease. In recent years， TCM has attracted the attention of both Chinese and foreign researchers for the unique advantages of treating both symptoms and root cause at the same time and multi-target synergy in the treatment of DKD. However， the specific mechanism is still unclear. It has been frequently verified that mitochondria may be one of the targets of TCM in the treatment of DKD. At the moment， no review on the treatment of DKD by TCM through the intervention of MQC is available. Therefore， this paper aims to summarize the research on TCM treatment of DKD by regulating MQC in the past 10 years， which is expected to provide a new direction for the treatment of DKD by TCM.
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Diabetic nephropathy （DN）， as one of the common chronic microvascular complications of diabetes， has become the main cause of renal failure in end-stage renal disease in China， increasing the risks of renal dialysis and kidney transplantation in diabetes patients. It is the leading cause of death in people with diabetes. The latest research on DN has focused on the gene level. microRNAs （miRNAs） are a family of endogenous accessible short-chain non-coding RNA molecules. By acting on a particular target， they activate or inhibit its mediated signaling pathways and related molecules， playing an important role in the occurrence and development of DN. They have become microeconomic factors for the prevention and treatment of DN. Traditional Chinese medicine （TCM） has a long history in the diagnosis and treatment of DN and has unique advantages such as significant curative effect and few side effects. A large number of studies have proved that TCM can target miRNA to affect multiple signaling pathways， participate in the regulation of inflammatory response， pyroptosis， mesenchymal transdifferentiation， and other pathological changes， and delay the further development of DN. Therefore， this study discusses the biogenesis mechanism of miRNA and its action mechanism in disease-related signaling pathways based on TCM diagnosis and treatment approaches from the perspective of miRNA， and summarizes the effect of TCM targeting miRNA on the disease-related signaling pathways and on DN. Thus， this study is expected to provide a theoretical reference for exploring the progress of TCM intervention in DN from the perspective of genes.
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Gastric cancer is one of the malignancies with high incidence in the world. Xiangsha Liu Junzitang，a common prescription for the prevention and treatment of gastric cancer，has the effects of moving Qi to relieve pain，drying dampness， and invigorating the spleen. It is especially indicated for gastric cancer of the spleen and stomach qi deficiency syndrome. Based on the databases such as CNKI，Wanfang Data，and PubMed，the clinical efficacy and experimental studies of Xiangsha Liu Junzitang for the prevention and treatment of gastric cancer were summarized and sorted out，and the mechanism of Xiangsha Liu Junzitang for the prevention and treatment of gastric cancer was elaborated in order to provide useful references for the clinical and basic research on Xiangsha Liu Junzitang in the field of gastric cancer in the future. In clinical practice，Xiangsha Liu Junzitang can treat gastric precancerous lesions，increase the body immunity of patients with gastric cancer，improve the symptoms of spleen and stomach weakness after gastric cancer surgery，and reduce the adverse reactions of the digestive tract after chemotherapy for gastric cancer. Its clinical efficacy is superior to that of western medicine alone whether it is combined with western medicine or used alone. In the experimental research，Xiangsha Liu Junzitang has the effects of regulating inflammatory factors，inhibiting the proliferation of gastric cancer cells，promoting the apoptosis of gastric cancer cells，and improving the activity of pepsin. Modern pharmacological research has shown that Xiangsha Liu Junzitang can conduct a comprehensive intervention with multiple components and multiple targets. The main components of a single drug contained include saponins，polysaccharides，lactones，volatile oils，organic acids，and others， with the effects of protecting gastric mucosa，regulating endocrine，and promoting apoptosis of epithelial cells in gastric mucosal dysplasia，reflecting the advantages and values of traditional Chinese medicine in the prevention and treatment of gastric cancer.
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By reviewing the relevant literature of ancient herbal works and modern codices， this paper sorted out the historical evolution and developmental venation of processing of Notoginseng Radix et Rhizoma. On this basis， the modern research of processed products of Notoginseng Radix et Rhizoma was used as the breakthrough point to analyze the literature in terms of processing technology， chemical composition changes and changes in pharmacological effects before and after processing. According to the research status of processing of Notoginseng Radix et Rhizoma， some existing problems were analyzed in this paper， such as not many ancient processing methods used in modern time， lack of standardized research on processing technology. And saponins， polysaccharides， amino acids， flavonoids and other chemical components in Notoginseng Radix et Rhizoma may change to different degrees before and after processing， which was the main reason for the difference of efficacy before and after processing. However， the current research on the pharmacological effects of Notoginseng Radix et Rhizoma mainly focuses on raw products， resulting in a lack of in-depth research on the transformation mechanism of Notoginseng Radix et Rhizoma in processing difference， and the scientific connotation of "Shengxiao Shubu" has not been clearly elaborated， which is not conducive to the standardized clinical use of drugs. Therefore， it is necessary to further analyze the material basis of Notoginseng Radix et Rhizoma and its processed products， and to explore the change rule of chemical components before and after processing and its correlation with pharmacodynamic activity， so as to clarify the processing mechanism for providing scientific basis for its standardized processing， quality control and clinical rational use.
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Vascular dementia （VaD） is a common disease that affects the health of the elderly. Due to the aging of the social population， the incidence of VaD is increasing year by year. There have been no officially approved treatments for this disease， mainly because its pathogenesis is complex， and the mechanism of action of effective drugs is not yet clear， which hinders drug research for the treatment of VaD. Therefore， by reviewing the available literature related to VaD， this study sorted out the pathogenesis of VaD from traditional Chinese medicine （TCM） and western medicine， and concluded that in TCM， VaD was characterized by the deficiency of the spleen and kidney （deficiency） and combination of phlegm and blood stasis （excess）， while in western medicine， the pathogenesis of VaD is mainly inflammatory response， oxidative stress， abnormal expression of related proteins， and dysfunction of signaling pathways. On this basis， this study also summarized the research on the mechanism of action of commonly used single Chinese herbal medicine and Chinese herbal medicine compound， western medicine， and the combination of Chinese herbal medicine and western medicine in the treatment of VaD in recent years. The commonly used single Chinese herbal medicine Ginkgo Folium and Chinese herbal medicine compound Dihuang Yinzi have the multi-component and multi-target characteristics and few adverse reactions in the treatment of VaD， while the commonly used western medicines such as donepezil and memantine have the characteristics of the clear target and quick onset. The combination of Chinese herbal medicine and western medicines can achieve a better effect. This study summarized the research on the pathogenesis and treatment of VaD， aiming to link the pathogenesis of VaD with the mechanism of effective drug therapy， and provide an important reference for future drug development for the treatment of VaD.
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A total of 290 prescriptions in Treatise on Typhoid and Miscellaneous Diseases （《伤寒杂病论》） and Synopsis of the Golden Chamber （《金匮要略》） and 191 prescriptions in Prescriptionology were selected as representatives of classical prescriptions of traditional Chinese medicine （TCM）， which were translated into Chinese and Japanese. The prescription names were used as keywords for search， and the patent application data up to June 2022 were retrieved. The retrieved results underwent data cleaning and manual noise reduction. Taking the important applicants as the starting point， the key technologies were divided and indexed according to the technical improvement characteristics of classic TCM prescriptions， and the intelligence information of patent applications for classic TCM prescriptions in the two countries was deeply excavated. The differences in the patent applications and layout strategies of classic TCM prescriptions between China and Japan were further compared. This paper drew up an accurate and targeted search strategy for more accurate and comprehensive retrieval， collation， and statistics of patent data of classic TCM prescriptions of representative patent applicants in China and Japan. At the same time， this paper deeply explored the information on Chinese patent applications in Japan and compared the differences in patent application strategies and patent protection of classic TCM prescriptions and Chinese prescriptions in Japan taking the layout of classic TCM prescriptions of representative patent applicants in China and Japan as the entry point. On one hand， it can accurately grasp the current patent application status of classic TCM prescriptions in the two countries， and on the other hand， it can also provide some references for the secondary development and research， patent application， and layout of classic prescriptions of enterprises， universities， and research institutions in China.
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Renal fibrosis is the main pathological foundation of chronic kidney diseases progressing to end-stage renal diseases. With complex pathogenic factors and prolonged disease course， it threatens the quality of life of patients and brings about heavy financial burden to medical care. In the instance of intestinal flora disturbance， the internal homeostasis is broken， resulting in various "imbalances". The "combination of state and target" endows the syndrome differentiation-based treatment of renal fibrosis with new connotation from the perspective of intestinal flora reconstruction and microbial diversity restoration. In addition， traditional Chinese medicine （TCM）-targeted intervention of intestinal microecology has unique advantages under the principle of "treating different diseases with the same method"， which can guide the diagnosis and treatment of renal fibrosis. To be specific， TCM emphasizes macroscopic regulation of state and microscopic targeting. In view of the inflammatory response， accumulation of endotoxin， and excessive deposition of extracellular matrix （ECM） in the process of renal fibrosis， the strategies for treating this disease have been developed， such as alleviating dampness，removing turbid toxin， and relieving deficiency and stasis. Famous prescriptions in ancient books or compound Chinese medicine prescriptions， classical formulas， Chinese medicine monomers， or active components of Chinese medicine target intestinal microecology. Therefore， from the perspective of common pathogenic factors of renal diseases （renal fibrosis） or pathological product-intestinal microecological imbalance， this article combines TCM basic theory with modern medical pathogenesis， and summarizes the research on TCM intervention of renal fibrosis by regulating intestinal microecology and the scientific connotation of renal fibrosis， which is expected to provide ideas and methods for the product development and related preparations and in-depth molecular biological research.
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Safety is the core of the quality of Chinese materia medica products， and microbial pollution caused by medicinal materials， decoction pieces， intermediate products and others can bring certain impact on the quality and safety of Chinese materia medica products. The reasons for this are not only the problems of medicinal materials themselves， but also the exogenous pollution introduced in the production process. How to effectively use microbial detection technology and establish an appropriate microbial quality control strategy in the whole process of Chinese materia medica production is of great significance to improve the quality of Chinese materia medica products. Therefore， the authors put forward a microbial quality control strategy in the whole process of Chinese materia medica production based on the guidance of quality by design （QbD） concept， emphasizing the scientific linkage between the internal and external microbial quality control systems to jointly ensure the quality of products in all aspects. Among them， the internal microbial quality control system includes the control of the whole chain of Chinese materia medica-decoction pieces-intermediate products-excipitents-packaging materials-final products， which should be carried out by stages and characteristics， while the external microbial quality control system includes the control of personnel-equipment and facilities-pharmaceutical water-environment， emphasizing the principle of quality risk management and the development of monitoring programs， aiming to closely integrate microbial quality risk management with the production process of Chinese materia medica products， and to classify and develop microbial control strategies in order to minimize the impact of contaminating microorganisms and effectively guarantee the quality of Chinese materia medica products.
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ObjectiveTo understand the current situation and hot spots of research on the treatment of constipation by traditional Chinese medicine （TCM） in China and foreign countries， and provide references and directions for future research. MethodIn this study， the data analysis software CiteSpace 5.8.R3C was used to conduct bibliometrics analysis of the relevant papers on the treatment of constipation by TCM from January 1， 2000 to December 31， 2021 in China National Knowledge Infrastructure （CNKI） and Web of Science core database. The visual display was carried out. ResultAfter retrieval， 2 133 papers were included in CNKI database and 224 papers were included in Web of Science core database. The statistical analysis showed that the overall annual publication volume in China and foreign countries was on the rise， but the publication volume in China had been declining for the past two consecutive years. The author co-occurrence analysis showed that the authors with the largest number of publications in China and foreign countries were LIU Chunqiang and CHUNGWAH CHENG， respectively. The institution co-occurrence analysis showed that the institutions with the largest number of publications in China and foreign countries were the Anorectal Department of Longhua Hospital Affiliated to Shanghai University of TCM and Beijing University of Chinese Medicine， respectively. The keyword analysis showed that research in this field in China and foreign countries mainly focused on TCM therapy， clinical research， etiology and pathogenesis， etc. ConclusionThe research on the treatment of constipation by TCM is mainly concentrated in China， but there is no closely-related scientific research team. The research in this field in China and foreign countries is still developing. With the continuous deepening of research， all research institutions should pay attention to interdisciplinary intersection， highlight technological innovation， and strive to achieve greater breakthroughs in this field.
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ObjectiveTo clone coumarate-3-hydroxylase gene （C3H） from Angelica sinensis， and analyze the correlation between its bioinformatics， expression patterns and content of ferulic acid， and to explore the functions of ASC3H. MethodReal-time polymerase chain reaction （Real-time PCR） was used to clone the full-length cDNA of ASC3H based on the transcriptome dataset of A. sinensis， and the bioinformatics analysis of the gene sequence was carried out. Real-time PCR and high performance liquid chromatography （HPLC） were used to determine relative expression of ASC3H and content of ferulic acid in different root tissues of A. sinensis （periderm， cortex and stele）. ResultThe open reading frame （ORF） of ASC3H （GenBank accession number： MN2550298） was 1 530 bp， encoding 509 amino acids， with a theoretical molecular weight of 57.86 kDa and an isoelectric point of 8.36. It was a hydrophilic protein that was located in the chloroplast with multiple phosphorylation sites and a transmembrane region， and contained a conserved domain CGYDWPKGYGPIINVW_P450 （383-399 aa） in cytochrome P450. Multiple amino acid sequence alignment analysis showed that ASC3H had high similarity with C3H from other plants， especially Ammi majus in Umbelliferae. The Real-time PCR revealed that ASC3H had different expressions in periderm， cortex and stele tissues of A. sinensis roots. It was found from HPLC that the cortex tissues had the highest content of ferulic acid， and the stele tissues had the lowest. ConclusionASC3H was successfully cloned from A. sinensis， and its sequence characteristics were understood more clearly， suggesting that ASC3H might be involved in the ferulic acid biosynthesis pathway of A. sinensis. This paper provided a basis for further studying the functions of the gene and exploring the biosynthesis and regulation mechanism of ferulic acid in A. sinensis， while laying the foundation for the genetic improvement of A. sinensis．
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ObjectiveTo study the effect of a microbial compound fertilizer on the growth， quality， and soil properties of Andrographis paniculata and provide a basis for its application in the cultivation and production of A. paniculata. MethodThrough five treatment tests in pots， the control group （CK， no fertilization）， the treatment A group （chemical fertilization）， the treatment B group （microbial fertilization at 2.5 g·kg-1 soil）， the treatment C group （microbial fertilization at 7.5 g·kg-1 soil）， and the treatment D group （microbial fertilization at 12.5 g·kg-1 soil） were established. The effects of a compound microbial fertilizer on the agronomic properties， the content of active ingredients， the number of rhizosphere soil bacteria， Actinomycetes， and fungi， and soil physicochemical properties of A. paniculata were studied. ResultAs compared with the CK group and the treatment A group， the plant height， leaf number， leaf area， and above-ground and below-ground fresh weight of A. paniculata were significantly increased after microbial fertilization， with differences in different growth periods. The effect of the compound microbial fertilizer on the content of andrographolide， dehydrated andrographolide， neandrographolide， and 14-deoxyandrographolide differed. The content of andrographolide significantly increased by 26.13% and 13.23% in the treatment C group， respectively， as compared with the CK group and the treatment A group （P<0.05）. The content of neandrographolide increased with the increasing amount of microbial fertilizer， and the content in the treatment D group increased by 9.06% and 50.33%， respectively， as compared with the CK group and the treatment A group （P<0.05）， which was the most obvious. The content of 14-deoxyandrographolide in the treatment B group significantly increased by 42.04% and 1.74%， respectively， as compared with the treatment A group and the CK group （P<0.05）. The content of dehydrated andrographolide content in the treatment B group increased most significantly， significantly increasing by 11.73% and 27.74%， respectively， as compared with the CK group and the treatment A group （P<0.05）. The soil pH of the treatment B， C， and D groups was higher than that of the CK group and the treatment A group. The content of total nitrogen and effective phosphorus increased most significantly in the treatment B group， and the content of fast-acting potassium increased most significantly in the treatment D group. The application of the compound microbial fertilizer increased the number of rhizosphere soil bacteria and Actinomycetes， which was significantly higher than that in the CK group and the treatment A group （P<0.05）， and increased with the increase of applied amount. The number of fungi decreased with the increase of the compound microbial fertilizer application， which was significantly different from that in the CK group （P<0.05）. The fungi/bacteria of all treatment groups were significantly lower than those of the CK group （P<0.05） and increased with the increase of microbial fertilizer application. ConclusionThe application of the compound microbial fertilizer transforms the plant-soil-microbial system in a benign direction， which is beneficial to the growth of A. paniculata and the accumulation of active ingredients.
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ObjectiveTo clone squalene epoxidase （SE）， a potential key rate-limiting enzyme involved in the synthesis pathway of Poria cocos triterpenes， from P. cocos and analyze for bioinformatics and expression. MethodThe total RNA was extracted by the kit and reverse-transcribed to cDNA. Specific primers were designed， and the cDNA was used as a template for cloning the SE gene， which was analyzed for bioinformatics. The expression of P. cocos qualene epoxidase（PcSE） was examined by Real-time polymerase chain reaction（Real-time PCR） in P. coco Shenzhou No. 10， Xiangjing 28， and 5.78 strains. ResultThe full length of PcSE is 1 571 bp， containing four exons and three introns. The obtained CDS sequence is 1 413 bp， encoding 470 amino acids. This protein is a hydrophobic protein with no signal peptide structure and has two transmembrane structural domains with a FAD/NAD （P） binding domain and SE structural domain localized to the mitochondrial membrane and the plasma membrane. The homologous sequence alignment with fungi of the Poriferae family is 80.92%， and the phylogenetic tree shows that PcSE protein is most closely related to P. cocos from the US. The results of Real-time PCR showed that the PcSE was expressed in all three strains， with the highest expression in 5.78 strain， and there was no significant difference in PcSE expression among the three strains. ConclusionFor the first time， the PcSE gene was cloned and analyzed from P. cocos， providing a basis for further research on the function of PcSE and the analysis of P. cocos triterpene biosynthesis pathway.
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ObjectiveTo investigate the relative content changes of differential metabolites and reducing sugars during the processing process of Rehmanniae Radix Praeparata （RRP） processed with Amomi Fructus （AF） and Citri Reticulatae Pericarpium （CRP）， and to lay the foundation for revealing the processing principle of this characteristic variety. MethodThe samples of the 0-54 h processing process of RRP processed with AF and CRP were taken as the research object， and their secondary metabolites were detected by ultra performance liquid chromatography tandem quadrupole time-of-flight mass spectrometry （UPLC-Q-TOF-MS）. The 0.1% formic acid aqueous solution （A）-acetonitrile （B） was used as the mobile phase for gradient elution （0-1 min， 1%-3%B； 1-10 min， 3%-9%B； 10-15 min， 9%-12%B； 15-22 min， 12%-18%B； 22-31 min， 18%-24%B； 31-35 min， 24%-100%B； 35-36 min， 100%-5%B； 36-40 min， 5%-1%B； 40-45 min， 1%B）， column temperature was 40 ℃， injection volume was 3 μL， flow rate was 0.3 mL·min-1. Electrospray ionization （ESI） was used to scan and collect MS data in the negative ion mode， the scanning range was m/z 50-1 250. Data analysis was carried out using PeakView 1.2 software， and the chemical composition of RRP processed with AF and CRP was identified by combining the literature information and chemical composition databases. The MS data were normalized by MarkerView 1.2， and then the multivariate statistical analysis was applied to screen the differential metabolites， and the changes of the relative contents of the differential metabolites with different processing times was analyzed， finally， correlation analysis was performed between the differential metabolites， the change of the reducing sugar content was combined to determine the most suitable processing time of RRP processed with AF and CRP. ResultA total of 121 compounds were identified from RRP processed with AF and CRP at different processing times， and 12 differential metabolites were screened out by multivariate statistical analysis， including catalpol， hesperidin， isoacteoside， acteoside， narirutin， echinacoside， isomartynoside， decaffeoylacteoside， 6-O-E-feruloylajugol， dihydroxy-7-O-neohesperidin， jionoside D， and rehmapicroside. With the prolongation of processing time， the relative contents of these 12 differential metabolites and reducing sugars changed slightly at 52-54 h. ConclusionUPLC-Q-TOF-MS can comprehensively and accurately identify the chemical constituents of RRP processed with AF and CRP at different processing times， and the suitable processing time of 52-54 h is determined according to the content changes of different metabolites and reducing sugars， which provides a basis for revealing the scientific connotation of the processing principle of this variety.
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ObjectiveAgrobacterium tumefaciens-mediated transformation （ATMT） of Clonostachys rosea， an endophytic fungus of Glycyrrhiza uralensis seeds， was established and optimized， and orthogonal test was designed to optimize the colonization conditions of C. rosea for G. uralensis seeds， so as to lay foundation for the development of biofertilizer and the breeding of high-quality G. uralensis. MethodThe conditions of ATMT were optimized from three aspects， including the concentration of acetosyringone， co-culture time and the concentration of conidia of recipient fungi. Then， high-quality transformants were selected. Orthogonal test was used to optimize the colonization conditions by taking co-culture temperature， co-culture time and spore concentration as factors and colonization rate as index. ResultWhen spore concentration was 1×107 cfu·mL-1， acetosyringone concentration was 150 μmol·L-1 and the co-culture time was 60 h， the transformation efficiency of C. rosea was the highest， which was 135 transformants per 1×107 recipient fungal spores. The accuracy and stability of the transformations were tested by cloning the marker gene green fluorescent protein （GFP） and β-glucuronidase （GUS） staining. When co-culture temperature was 25 ℃， co-culture time was 36 h and the spore concentration was 1×106 cfu·mL-1， the colonizing rate for C. rosea back dyeing into G. uralensis seeds by seed soaking method was the highest， which was 71.11%. ConclusionThis study successfully establishes stable and efficient technical systems not only of ATMT in C. rosea， but also of colonization of the transformants into G. uralensis seeds， which can lay a foundation for the development of biofertilizer of G. uralensis.
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ObjectiveThrough the targeted lipidomics， we explored the intervention mechanism of Kaixuan Bushen method on psoriasis vulgaris （PV） from the perspective of lipid metabolism， providing reference for the diagnosis and treatment of PV. MethodTwenty-six patients with PV admitting the outpatient clinic of the Department of Dermatology， Guang'anmen Hospital， China Academy of Chinese Medical Sciences from September 2019 to November 2020 were selected as the research object （observation group）， and 26 sex- and age-matched healthy volunteers in the same period were recruited as control group. Venous blood was collected for lipid index and targeted lipidomics detection in the control and observation groups at inclusion. After 12 weeks of continuous treatment of Kaixuan Bushen method， the psoriasis area and severity index （PASI） was measured and compared before and after treatment to assess the clinical efficacy， while venous blood was collected again in the observation group to compare the blood lipid level and lipid metabolism of patients before and after treatment. Targeted lipidomics analysis was performed by ultra performance liquid chromatography tandem mass spectrometry （UPLC-MS/MS） on an ACQUITY UPLC BEH C8 column （2.1 mm×100 mm， 1.7 μm） with mobile phase of 5 mmol∙L-1 ammonium formate in acetonitrile-water （6∶4， A）-5 mmol∙L-1 ammonium formate in acetonitrile-isopropanol （1∶9， B） for gradient elution and flow rate of 0.26 mL∙min-1. Conditions of MS were electrospray ionization （ESI）， positive and negative ion modes， and scanning range of m/z 50-1 200. Then principal component analysis （PCA） and partial least squares-discriminant analysis （PLS-DA） models were developed to screen differential metabolites， and the differential metabolites were identified and the pathways were enriched. ResultAfter 12 weeks of treatment with Kaixuan Bushen method， PASI score decreased by more than 50% in a total of 22 out of 26 patients with PV， suggesting the total effective rate was 84.62%. The serum triglyceride level of patients with PV was significantly higher than that of healthy individuals （P<0.05）， and the triglyceride level was significantly reduced after treatment （P<0.05）. Targeted lipidomics analysis screened a total of 43 potential biomarkers for PV， of which 42 were up-regulated and 1 was down-regulated， involving 7 signaling pathways such as linoleic acid metabolism， glycerophospholipid metabolism and unsaturated fatty acid biosynthesis. Moreover， there were 14 response makers for clinical efficacy of Kaixuan Bushen method on PV， of which 6 were up-regulated and 8 were down-regulated， involving five signaling pathways such as linoleic acid metabolism， glycerophospholipid metabolism and sphingolipids metabolism. In a comparison between healthy individuals and patients with PV and PV before and after treatment， the common differential metabolites were screened as phosphatidylcholine （PC） 38∶0 and ceramide （Cer） 42∶1， and the common pathways were linoleic acid and glycerophospholipid metabolic pathways. ConclusionThe disorder of lipid metabolism in PV are largely due to abnormal sphingolipid， glycerophospholipid and linoleic acid metabolic pathways， of which Kaixuan Bushen method can regulate the glycerophospholipid and linoleic acid metabolism， thereby improving psoriatic lesions.
ABSTRACT
ObjectiveTo confirm the clinical efficacy and safety of Yishen Yangxin Anshen tablets in the treatment of insomnia （heart-blood deficiency and kidney-essence insufficiency syndrome）. MethodA randomized block， double-blind， placebo-controlled， multi-center clinical trial design method was adopted， and a total of 480 patients with insomnia due to deficiency of heart blood and insufficiency of kidney essence （treatment group-control group 3∶1） from seven hospitals （Guang'anmen Hospital， China Academy of Chinese Medical Sciences， The First Clinical Hospital， Jilin Province Academy of Traditional Chinese Medicine（TCM）， The Second Affiliated Hospital of Liaoning University of TCM， The First Affiliated Hospital of Henan University of Chinese Medicine， Henan Province Hospital of TCM， Hebei General Hospital， The First Hospital of Hunan University of Chinese Medicine） were enrolled. The treatment group was given Yishen Yangxin Anshen tablets and the control group received placebo tablets （4 tablets/time， 3 times/day， 4 weeks of administration， 4 weeks of follow-up after drug withdrawal）. The sleep dysfunction rating scale （SDRS） score， pittsburgh sleep quality index （PSQI） score， TCM， polysomnography （PSG） indicators from four hospital （Guang'anmen Hospital， China Academy of Chinese Medical Sciences， Henan Province Hospital of TCM， Hebei General Hospital， The First Hospital of Hunan University of Chinese Medicine）， and other efficacy indicators were compared between the two groups before and after treatment. Through general physical examination， laboratory examination， and observation of adverse events， the safety of the drugs was evaluated. ResultThe baseline indexes of the two groups showed no significant difference and thus the two groups were comparable. After treatment， the total score of SDRS in the treatment group was lower than that in the control group （P<0.01）. After drug withdrawal for 4 weeks， the total score of SDRS demonstrated no significant change in the treatment group as compared with that at the end of treatment， indicating that the rebound change of curative effect was not obvious. After treatment， the total score of PSQI in the treatment group decreased as compared with that in the control group （P<0.01）， and the change of total score of PSQI in the treatment group was statistically significant （P<0.05） after drug withdrawal for 4 weeks but small， indicating that the rebound change of curative effect was not obvious. After treatment， the total effective rate about the TCM symptoms in the treatment group was higher than that in the control group （χ2=137.521，P<0.01）. After treatment， the disappearance rates of single indexes in the treatment group， such as difficulty in falling asleep， easily waking up after sleeping， early awakening， short sleep time， dreamfulness， palpitation， forgetfulness， dizziness， mental fatigue， and weakness of waist and knee， increased compared with those in the control group （P<0.01）. After treatment， the treatment group demonstrated fewer awaking times （AT）， longer total sleep time （TST）， lower ATA/TST ratio， and higher sleep efficiency （%） than the control group （P<0.05）. No abnormal value or aggravation related to drugs was observed in either group. The incidence of adverse events in the treatment group and the control group was 5.57% and 8.40% respectively. No serious adverse events or adverse events leading to withdrawal happened in either group. ConclusionYishen Yangxin Anshen tablets is effective and safe for patients with insomnia of deficiency of heart-blood and insufficiency of kidney-essence.