Effects of miR-722 on gene expression and alternative splicing in the liver of half-smooth tongue sole after infection with Vibrio anguillarum.

MicroRNAs play crucial roles in various biological processes, including but not limited to differentiation, development, disease, and immunity. However, their immunoregulatory roles in half-smooth tongue sole are lacking. Our previous studies indicated that miR-722 could target C5aR1 to modulate the complement pathway to alleviate inflammatory response and even affect the mortality after the bacterial infection with Vibrio anguillarum. Driven by the purpose of revealing the underlying mechanisms, in this study, we investigated the effects of miR-722 on the gene expression and alternative splicing (AS) in the liver of half-smooth tongue sole after Vibrio anguillarum infection, with the approach of miR-722 overexpression/silencing and subsequent RNA-seq. Among the different comparisons, the I group (miR-722 inhibitor and V. anguillarum) versus blank control (PBS) exhibited the highest number of differentially expressed genes (DEGs), suggesting that the immune response was overactivated after inhibiting the miR-722. In addition, enrichment analyses were performed to reveal the functions of DEGs and differential AS (DAS) genes, reflecting the enrichment of RNA splicing and immune-related pathways including NF-κB and T cell receptor signaling pathway. Comparing the M group (miR-722 mimic and V. anguillarum) with the negative control (random sequence and V. anguillarum), two immune-related genes, cd48 and mapk8, were differentially expressed, of which mapk8 was also differentially spliced, indicating their importance in the immune response. Furthermore, representative gene analysis was performed, suggesting their corresponding functional changes due to AS. To verify the RNA-seq data, quantitative real-time PCR was employed with twenty pairs of primers for DEGs and DAS events. Overall, our results demonstrated that miR-722 could mediate the transcriptome-wide changes of gene expression and AS in half-smooth tongue sole, and provided insights into the regulatory role of miR-722 in immune responses, laying the foundation for further functional analyses and practical applications in aquaculture.

Role of Akkermansia muciniphila in insulin resistance.

Insulin resistance (IR) is a pathogenic factor in numerous metabolic diseases. The gut microbiota plays a crucial role in maintaining the function of the intestinal barrier and overall human health, thereby influencing IR. Dysbiosis of the gut microbiota can contribute to the development of IR. Therefore, it is essential to maintain a balanced and diverse gut microbiota for optimal health. Akkermansia muciniphila, a widely present microorganism in the human intestine, has been shown to regulate gastrointestinal mucosal barrier integrity, reduce endotoxin penetration, decrease systemic inflammation levels, and improve insulin sensitivity. Reduced abundance of A. muciniphila is associated with an increased risk of IR and other metabolic diseases, highlighting its correlation with IR. Understanding the role and regulatory mechanism of A. muciniphila is crucial for comprehending IR pathogenesis and developing novel strategies for preventing and treating related metabolic disorders. Individual variations may exist in both the gut microbiota composition and its impact on IR among different individuals. Further investigation into individual differences between A. muciniphila and IR will facilitate advancements in personalized medicine by promoting tailored interventions based on the gut microbiota composition, which is a potential future direction that would optimize insulin sensitivity while preventing metabolic disease occurrence. In this review, we describe the physiological characteristics of A. muciniphila, emphasize its roles in underlying mechanisms contributing to IR pathology, and summarize how alterations in its abundance affect IR development, thereby providing valuable insights for further research on A. muciniphila, as well as new drug development targeting diabetes.

Mass spectrometry-based metabolomics reveals metabolism of molnupiravir may lead to metabolic disorders and hepatotoxicity.

Molnupiravir (MO) is a pyrimidine nucleoside anti-SARS-CoV-2 drug. MO treatment could cause mild liver injury. However, the underlying mechanism of MO-induced liver injury and the metabolic pathway of MO in vivo are unclear. In this study, metabolomics analysis and molecular biology methods were used to explore these issues. Through metabolomics analysis, it was found that the homeostasis of pyrimidine, purine, lysophosphatidylcholine (LPC), and amino acids in mice was destroyed after MO treatment. A total of 80 changed metabolites were detected. Among these changed metabolites, 4-ethylphenyl sulfate, dihydrouracil, and LPC 20:0 was related to the elevation of alkaline phosphatase (ALP), interleukin-6 (IL6), and nuclear factor kappa-B (NF-κB). The levels of 4-ethylphenyl sulfate, dihydrouracil, and LPC 20:0 in plasma were positively correlated with their levels in the liver, suggesting that these metabolites were associated with MO-induced liver injury. MO treatment could increase NHC and cytidine levels, activate cytidine deaminase (CDA), and increase LPC levels. CDA and LPC could increase the mRNA expression level of toll-like receptor (TLR). The current study indicated that the elevation of hepatic TLR may be an important reason for MO leading to the liver injury.

Based on Serum Pharmacochemistry and Metabolomics Studied the Pharmacodynamic Material Basis and Mechanism of Rubi Fructus (Fupenzi) in Improving the Symptom of Kidney-Yang Deficiency.

Rubi fructus (Fupenzi) is the Chinese medicine for both food and medicine, which can be used to tonify kidney yang, strengthen essence and shrink urine, but its effective components and mechanism are not clear. In this paper, the active components of Fupenzi in vivo and in vitro were detected. Adenine was used to replicate the model of kidney yang deficiency, and organ index, biochemical index and histopathology were used to evaluate the effect of different doses of Fupenzi on tonifying kidney yang. Metabonomics technique was used to analyze the metabolic regulation mechanism of Fupenzi in improving kidney yang deficiency syndrome. The results showed that 61 chemical constituents of Fupenzi were identified in vitro. A total of 51 chemical components were identified, including 30 prototype components and 21 metabolic components, which may be theeffective components of Fupenzi. The results of pharmacodynamics showed that Fupenzi can effectively improve the symptom of kidney-yang deficiency, which may be mainly through primary bile acid biosynthesis, linoleic acid metabolism, steroid hormone biosynthesis, ß-alanine metabolism, glutathione metabolism, porphyrin and chlorophyll metabolism, unsaturated fatty acid biosynthesis, arachidonic acid metabolism, arginine and proline metabolism and other metabolic pathways to improve adenine-induced metabolic disorders in rats with kidney-yang deficiency syndrome.

[Effects and mechanisms of Gegen Qinlian Decoction in inhibiting M1 polarization of macrophages under inflammatory hypoxia microenvironment].

To explore the effect and mechanism of Gegen Qinlian Decoction(GQD) in inhibiting M1 polarization of macrophages under inflammatory hypoxia by simulating intestinal hypoxia microenvironment in vitro. A tri-gas incubator was used to simulate normal physiological hypoxia of the colon and inflammatory hypoxia microenvironments of ulcerative colitis(UC). RAW264.7 macrophages were divided into 18.5% O_(2 )(normoxia group), 4% O_2(physiological hypoxia group), and 1% O_2(inflammatory hypoxia group), and they were induced by lipopolysaccharide(LPS) for 24 h. M1 polarization was detected by flow cytometry. Under the condition of 1% inflammatory hypoxia, they were divided into blank group, model group, and GQD-containing serum low, medium, and high dose groups. Flow cytometry was used to detect M1 polarization marker CD86, and ELISA was used to detect the expression of tumor necrosis factor-α(TNF-α) and interleukin-1ß(IL-1ß) in cell supernatant. The mRNA expression of hypoxia-inducible factor-1α(HIF-1α), TNF-α, and IL-1ß was detected by qRT-PCR. Western blot was used to detect the expression of HIF-1α/nuclear transcription factor-κB(NF-κB) signaling pathway-related proteins. The nuclear translocation of NF-κB p65 was detected by immunofluorescence. The results showed that the positive rate of CD86 in the 1% O_2 group was the highest. Under the condition of 1% inflammatory hypoxia, compared with the blank group, the expression of CD86, TNF-α, IL-1ß, and HIF-1α in the model group increased. Compared with the model group, each group of GQD could reduce the expression of CD86, TNF-α, IL-1ß, and HIF-1α. Compared with the blank group, the protein expression of HIF-1α, NF-κB p65, p-IKKα/ß, and p-IκBα in the model group increased. Compared with the model group, the protein expression of HIF-1α, NF-κB p65, p-IKKα/ß, and p-IκBα in GQD groups was significantly decreased. Compared with the blank group, NF-κB p65 in the model group entered the nucleus significantly. Compared with the model group, the nuclear expression of NF-κB p65 was decreased in each GQD group. Studies have shown that GQD may protect the intestine by down-regulating the HIF-1α/NF-κB signaling pathway to inhibit M1 polarization of macrophages and secretion of related inflammatory factors under 1% inflammatory hypoxia.

[Guidelines for traditional dietary care in autumn and winter].

The National Nutrition Plan(2017-2030) and the Healthy China Action Plan(2019-2030) propose to vigorously develop traditional dietary care services, fully leverage the role of traditional dietary care in modern nutrition, and guide citizens to develop dietary habits that are in line with the dietary characteristics of different regions in China. Traditional dietary care has a long history in China and is one of the brilliant treasures of Chinese cuisine and traditional Chinese medicine(TCM) culture. It has played an important role in disease prevention, treatment, and health preservation and longevity. To promote the traditional culture of TCM, and guide and standardize the application and promotion of dietary care, it is necessary to develop a dietary care guideline with TCM characteristics. Based on the theories and practices of TCM, the China Academy of Chinese Medical Sciences(CACMS) has developed this guideline, which is tailored to local conditions and combined with modern nutrition, and targets people with different physical constitutions. According to the principles of dialectical diet, tailored to people, times, and local conditions, reinforcing healthy qi, correction, the combination of meat and vegetables, and the combination of four qi and five flavors, suitable ingredients are recommended(including TCM materials that are both food and medicinal materials). By promoting the popularization and development of traditional dietary care, this guideline contributes to integrating the strength of TCM into a unique nutritional and health model with Chinese characteristics.

Inflammatory reaction and immune response of half-smooth tongue sole (Cynoglossus semilaevis) after infection with Vibrio anguillarum.

Frequently occurred bacterial diseases have seriously affected the aquaculture industry of half-smooth tongue sole (Cynoglossus semilaevis). Notably, vibriosis, with Vibrio anguillarum as one of the causative pathogens, is the most severe bacterial disease with severe inflammatory response of the host, leading to high mortality rates. In the present study, we explored the relationship between bacterial concentrations and host mortality, inflammatory reaction, and immune response in half-smooth tongue sole after infection with V. anguillarum at different concentrations (Treatment 1, 6.4 × 105 CFU/mL; Treatment 2, 6.4 × 106 CFU/mL). The mortality of Treatment 2 (77.5%) was significantly higher than that of Treatment 1 (10%), corresponding with bacterial concentrations. Although the number of deaths varies, intensive deaths were observed within 24 h post infection (hpi) in both bacterial concentration groups. Histopathological analyses revealed that fish tissues were most severely damaged at 24 or 48 hpi, and Treatment 2 was more severe than Treatment 1. A qRT-PCR-based detection method with virulence factor gene empA was established to quantify the bacterial loads in various tissues, and the bacterial loads were the highest at 24 hpi in Treatment 2, and at 48 hpi in Treatment 1. Additionally, the expression levels of complement genes (C5a, C3, C5, and C6), inflammatory factors (IL-1ß, TNF-α, and IL-10), and other immune-related genes (jak2, NF-κB1, stat3, and tlr3) were increased in various tissues after infection in both treatment groups, with most genes being most expressed at 24 or 48 hpi, and expression levels of inflammatory factors in Treatment 2 were higher than those in Treatment 1. Moreover, the expression of C5a was positively correlated with that of proinflammatory cytokines in both bacterial concentration groups. According to the results of this study, 24-48 hpi was a key node for early vibriosis detection and intervention. Compared with the low mortality of Treatment 1, the mass death of fish in Treatment 2 was suggested to be caused by uncontrolled excessive inflammatory reaction induced by the overactivation of complement system, especially C5a. We believe these results could provide theoretical basis for prevention, evaluation, and treatment of vibrio disease in tongue sole aquaculture, and lay a solid foundation for future functional analyses.

C5a drives the inflammatory response with bacterial dose effect by binding to C5aR1 in zebrafish infected with Aeromonas hydrophila.

The complement system is essential to host defense, but its excessive activation caused by severe pathogen invasion is a driving force in adverse inflammatory. The binding of complement component 5a (C5a) and complement component 5a receptor 1 (C5aR1) is the key to trigger complement-mediated inflammatory response in mammals. However, the role of C5a-C5aR1 axis in fish immune response remains obscure. In this study, the role of C5a-C5aR1 axis of zebrafish (Danio rerio) after serious infection with Aeromonas hydrophila was investigated. C5a and C5aR1 of zebrafish were cloned, with CDS sequences of 228 and 1041 bp, respectively, and they were widely expressed in various tissues with the highest expression in the liver and spleen, respectively. The survival of zebrafish was closely correlated to the dose of A. hydrophila. The cytokine storm occurred at high concentrations of A. hydrophila infection. At 24 h post infection (hpi), the expression of C5a and C5aR1 in the spleen increased 26.8-fold and 9.9-fold in treatment group 1 (TG1, 3.0 × 107 CFU/mL) (P < 0.01), and 4.7-fold and 3.4-fold in treatment group 2 (TG2, 1.0 × 107 CFU/mL) (P < 0.05), respectively. Correspondingly, proinflammatory cytokines interleukin-1ß (IL-1ß), interleukin-8 (IL-8), and interleukin-17 (IL-17) were positively correlated to C5a and C5aR1 at mRNA and protein expression levels. The expression of IL-1ß was significantly increased in the spleen at 6 hpi, with a 599.2-fold and 203.2-fold upregulation in TG1 and TG2 (P < 0.001), respectively. Moreover, after inhibition of C5a-C5aR1 binding treated with C5aR1 antagonist (W-54011), zebrafish showed lower expression of C5a, C5aR1, and cytokines, less intestinal damage, and significantly enhancement of survival (P < 0.05) after A. hydrophila challenge. This study revealed that the inflammatory effect of C5a was achieved by binding to C5aR1 in zebrafish, providing novel insights into using C5a-C5aR1 axis as an effective target to reduce bacterial inflammation and disease in fish.

Dendrobium officinale polysaccharide Converts M2 into M1 Subtype Macrophage Polarization via the STAT6/PPAR-r and JAGGED1/NOTCH1 Signaling Pathways to Inhibit Gastric Cancer.

Dendrobium officinale polysaccharide (DOP) has shown various biological activities. However, the ability of DOP to participate in immune regulation during anti-gastric cancer treatment has remained unclear. In this study, the in vitro results showed that DOP has the potential to polarize THP-1 macrophages from the M2 to the M1 phenotype, downregulate the STAT6/PPAR-r signaling pathway and the protein expression of their down-targeted ARG1 and TGM2, and further decrease the main protein and mRNA expression in the JAGGED1/NOTCH1 signaling pathway. DOP suppressed the migration of gastric cancer cells by decreasing the protein expression of N-cadherin and Vimentin and increasing E-cadherin. In addition, CM-DOP promoted the apoptosis of gastric cancer cells by upregulating Caspase-3 and increasing the ratio of Bax/Bcl-2. In vivo, DOP effectively inhibited the growth of tumors and the expression of Ki-67. In summary, these findings demonstrated that DOP converted the polarization of M2 subtype macrophages into M1 subtypes via the STAT6/PPAR-r and JAGGED1/NOTCH1 signaling pathways in order to reduce apoptosis and prevent migration, thus indicating the potential of DOP as an adjuvant tumor therapy in preclinical and clinical trials.

[Effect of guarana on lipid metabolism in obese rats: a study based on lipidomics].

This study investigated the effect of guarana on plasma lipid metabolites in obese rats and analyzed its mechanism in the treatment of dyslipidemia in obesity. High-fat diet was used to establish obese rat models, and the therapeutic effect of guarana on obese rats was evaluated by measuring body weight, white fat, liver weight, and lipid content, as well as observing liver histomorphology. Lipid metabolites in plasma of rats in each group were detected by UHPLC-Q-TOF-MS lipidomics. The protein expressions of fatty acid synthase, acetyl-CoA carboxylase 1, triglyceride synthesis enzyme, carnitine palmitoyltransferase Ⅰ, and acetyl-coenzyme A acyltransferase 2 in rat liver were detected using Western blot. The results revealed that guarana significantly reduced body weight, white fat, and liver weight of obese rats due to high-fat diet, and alleviated dyslipidemia and liver steatosis. Lipidomics showed that some triglycerides and phospholipids were significantly elevated in the high-fat model group, and part of them was reduced after guarana treatment. Western blot found that guarana inhibited the expression of hepatic fatty acid and triglyceride synthesis-related proteins and increased the expression of fatty acid ß-oxidation-related proteins. Abnormalities in triglyceride and phospholipid metabolism are the main characteristics of plasma lipid metabolism in obese rats induced by high-fat diet. Guarana may regulate partial triglyceride and phospholipid metabolism by inhibiting hepatic fatty acid and triglyceride synthesis and increasing fatty acid ß-oxidation, thereby improving rat obesity and dyslipidemia.

[Metabolomic study on urine of chronic inflammation rats treated with Buyang Huanwu Decoction based on UPLC-Q-TOF-MS].

The study investigated the effect of Buyang Huanwu Decoction(BYHWD) on endogenous biomarkers in the urine of rats with chronic inflammation induced by lipopolysaccharide(LPS) using ultra-high performance liquid chromatography-quadrupole-time-of-flight-mass spectrometry(UPLC-Q-TOF-MS), aiming to elucidate the molecular mechanism underlying the therapeutic effect of BYHWD on chronic inflammation from a metabolomics perspective. Male SD rats were randomly divided into a normal group, a model group, and low-, medium-, and high-dose BYHWD groups(7.5, 15, and 30 g·kg~(-1)). The model group and BYHWD groups received tail intravenous injection of LPS(200 µg·kg~(-1)) on the first day of each week, followed by oral administration of BYHWD once a day for four consecutive weeks. Urine samples were collected at the end of the administration period, and UPLC-Q-TOF-MS was used to analyze the metabolic profiles of the rat urine in each group. Multivariate statistical analysis methods such as principal component analysis(PCA), partial least squares-discriminant analysis(PLS-DA), and orthogonal partial least squares-discriminant analysis(OPLS-DA) were used to analyze the effect of BYHWD on endogenous metabolites. One-way ANOVA and variable importance for the projection(VIP) were used to screen for potential biomarkers related to chronic inflammation. The identified biomarkers were subjected to pathway and enrichment analysis using MetaboAnalyst 5.0. A total of 25 potential biomarkers were screened and identified in the rat urine in this experiment. Compared with the normal group, the model group showed significant increases in the levels of 14 substances(P<0.05) and significant decreases in the levels of 11 substances(P<0.05). BYHWD was able to effectively reverse the trend of most endogenous biomarkers. Compared with the model group, BYHWD significantly down-regulated 13 biomarkers(P<0.05) and up-regulated 10 biomarkers(P<0.05). The metabolic products were mainly related to the biosynthesis of pantothenic acid and coenzyme A, tryptophan metabolism, retinol metabolism, and propionate metabolism. BYHWD has therapeutic effect on chronic inflammation induced by LPS, which may be related to its ability to improve the levels of endogenous metabolites, enhance the body's anti-inflammatory and antioxidant capabilities, and restore normal metabolic activity.

The Effect of Dried Ginger (Gan Jiang) on Stomach Energy Metabolism and the Related Mechanism in Rats Based on Metabonomics.

Dried ginger is a commonly used stomachic. Dried ginger is often used as a gastric protector to treat stomach-related diseases. However, the effect of dried ginger on energy metabolism in stomach tissue of rats under physiological condition has not been studied. In this study, different doses of water extract of dried ginger were given to rats for 4 weeks. The activity of Na+ -K+ -ATPase, Ca2+ -Mg2+ -ATPase, SDH (succinate dehydrogenase) enzyme, ATP content, mitochondrial metabolic rate and mitochondrial number in stomach tissue of rats were measured. Analysis of potential biomarkers related to the effect of dried ginger on energy metabolism in stomach tissue of rats by metabonomics, and their metabolic pathways were also analyzed. The results revealed that there was no significant difference in Na+ -K+ -ATPase in high-dose group (GJH), medium-dose group (GJM) and low-dose group (GJL) compared to the Control group. The Ca2+ -Mg2+ -ATPase activity was significantly increased in stomach tissue of GJH group and GJM group, but there were no significant changes in stomach tissue of GJL group. The SDH activity and the ATP levels were significantly increased in stomach tissue of GJH group, GJM group and GJL group. The mitochondrial metabolic rate was significantly increased in GJL group, but there was no significant change in GJM group and was inhibited in GJH group. These effects might be mediated by arginine biosynthesis, glutathione metabolism, arachidonic acid metabolism, glycerophospholipid metabolism, arginine and proline metabolism, purine metabolism pathway.

Fabrication and Characterization of ß-Cyclodextrin/Mosla Chinensis Essential Oil Inclusion Complexes: Experimental Design and Molecular Modeling.

Essential oils (EOs) are primarily isolated from medicinal plants and possess various biological properties. However, their low water solubility and volatility substantially limit their application potential. Therefore, the aim of the current study was to improve the solubility and stability of the Mosla Chinensis (M. Chinensis) EO by forming an inclusion complex (IC) with ß-cyclodextrin (ß-CD). Furthermore, the IC formation process was investigated using experimental techniques and molecular modeling. The major components of M. Chinensis 'Jiangxiangru' EOs were carvacrol, thymol, o-cymene, and terpinene, and its IC with ß-CD were prepared using the ultrasonication method. Multivariable optimization was studied using a Plackett-Burman design (step 1, identifying key parameters) followed by a central composite design for optimization of the parameters (step 2, optimizing the key parameters). SEM, FT-IR, TGA, and dissolution experiments were performed to analyze the physicochemical properties of the ICs. In addition, the interaction between EO and ß-CD was further investigated using phase solubility, molecular docking, and molecular simulation studies. The results showed that the optimal encapsulation efficiency and loading capacity of EO in the ICs were 86.17% and 8.92%, respectively. Results of physicochemical properties were different after being encapsulated, indicating that the ICs had been successfully fabricated. Additionally, molecular docking and dynamics simulation showed that ß-CD could encapsulate the EO component (carvacrol) via noncovalent interactions. In conclusion, a comprehensive methodology was developed for determining key parameters under multivariate conditions by utilizing two-step optimization experiments to obtain ICs of EO with ß-CD. Furthermore, molecular modeling was used to study the mechanisms involved in molecular inclusion complexation.

Elucidation of the Transport Mechanism of Puerarin and Gastrodin and Their Interaction on the Absorption in a Caco-2 Cell Monolayer Model.

Puerarin (PUR) and gastrodin (GAS) are often used in combined way for treating diseases caused by microcirculation disorders. The current study aimed to investigate the absorption and transportation mechanism of PUR and GAS and their interaction via Caco-2 monolayer cell model. In this work, the concentration in Caco-2 cell of PUR and GAS was determined by HPLC method. The bidirectional transport of PUR and GAS and the inhibition of drug efflux including verapamil and cyclosporine on the transport of these two components were studied. The mutual influence between PUR and GAS, especially the effect of the latter on the former of the bidirectional transport were also investigated. The transport of 50 µg·mL-1 PUR in Caco-2 cells has no obvious directionality. While the transport of 100 and 200 µg·mL-1 PUR presents a strong directionality, and this directionality can be inhibited by verapamil and cyclosporine. When PUR and GAS were used in combination, GAS could increase the absorption of PUR while PUR had no obvious influence on GAS. Therefore, the compatibility of PUR and GAS is reasonable, and GAS can promote the transmembrane transport of PUR, the effect of which is similar to that of verapamil.

A strategy combining solid-phase extraction, multiple mass defect filtering and molecular networking for rapid structural classification and annotation of natural products: characterization of chemical diversity in Citrus aurantium as a case study.

Medicinal plants are complex chemical systems containing thousands of secondary metabolites. The rapid classification and characterization of the components in medicinal plants using mass spectrometry (MS) remains an immense challenge. Herein, a novel strategy is presented for MS through the combination of solid-phase extraction (SPE), multiple mass defect filtering (MMDF) and molecular networking (MN). This strategy enables efficient classification and annotation of natural products. When combined with SPE and MMDF, the improved analytical method of MN can perform the rapid annotation of diverse natural products in Citrus aurantium according to the tandem mass spectrometry (MS/MS) fragments. In MN, MS2LDA can be initially applied to recognize substructures of natural products, according to the common fragmentation patterns and neutral losses in multiple MS/MS spectra. MolNetEnhancer was adopted here to obtain chemical classifications provided by ClassyFire. The results suggest that the integrated SPE-MMDF-MN method was capable of rapidly annotating a greater number of natural products from Citrus aurantium than the classical MN strategy alone. Moreover, SPE and MMDF enhanced the effectiveness of MN for annotating, classifying and distinguishing different types of natural products. Our workflow provides the foundation for the automated, high-throughput structural classification and annotation of secondary metabolites with various chemical structures. The developed approach can be widely applied in the analysis of constituents in natural products.

Discovery and validation of quality markers of Fructus Aurantii against acetylcholinesterase using metabolomics and bioactivity assays.

Fructus Aurantii is a traditional medicated diet in East Asia. To determine the underlying chemical markers responsible for the quality and efficacy of Fructus Aurantii, a sensitive metabolomic method was applied to distinguish Fructus Aurantii in Jiangxi Province from other two geographical locations (Hunan Province and Chongqing City) in China. In the present study, multivariate analyses were adopted to compare chemical compositions in 21 batches of Fructus Aurantii samples. Among three geographical origins, 23 differential compounds were structurally identified. Serum pharmacochemistry exhibited that 22 components could be detected in rat serum. Six differential and absorbed components were selected as six potential markers. Statistical analysis revealed that the content of six markers varied widely in three origins of Fructus Aurantii. Six differential and absorbed components were evaluated further by biological activity. Neohesperidin, naringin, and meranzin showed inhibitory effect on acetylcholinesterase that regulates gastrointestinal motility in vitro and in silico, suggesting that these three components may be determined as the active biomarkers of Fructus Aurantii. These findings demonstrate the potential of biomarkers for identification and quality control of Fructus Aurantii.

[Molecular mechanism analysis of Erzhi Wan in treatment of aging based on network pharmacology].

This present study aimed to explore the molecular mechanism of Erzhi Wan(a prescription of nourishing Yin and toni-fying liver and kidney) in treatment of aging by network pharmacology. The active constituents and target proteins of Erzhi Wan were searched from Traditional Chinese Medicine Systems Pharmacology Database(TCMSP) and PubChem databases respectively. Aging-related genes were searched from Gene and HAGR databases. Based on the Ingenuity Pathway Analysis(IPA), we analyzed the common molecular network, biological pathway and interaction sites between these two parts, and verified some of them by Western blot. Twelve active constituents of Erzhi Wan were screened by TCMSP databases, 69 protein targets were predicted through PubChem, and 148 aging-related genes were found in Gene and HAGR databases. IPA comparison showed that the molecular networks of these two were complex, with diversity of biological functions. The common pathways involved 292 pathways, mainly related to tumors. They acted on hypoxia inducible factor-1α gene(HIF1α), nuclear factor-E2 related factor(Nrf2/NFE2 L2), tumor necrosis factor(TNF) and other sites. Western blot results suggested that Erzhi Wan could down-regulate the expression of HIF1α, with statistical difference(P<0.05). It was concluded that, Erzhi Wan could intervene aging through improving pseudo-hypoxic microenvironment and inflammation. The molecular mechanism of Erzhi Wan in delaying aging was preliminarily revealed, which laid a foundation for further stu-dying the anti-aging mechanism of Erzhi Wan, and also provided a reference for the compatibility mechanism and extended application of Chinese medicine compounds.

A Comprehensive and Rapid Quality Evaluation Method of Traditional Chinese Medicine Decoction by Integrating UPLC-QTOF-MS and UFLC-QQQ-MS and its Application.

Decoction is one of the oldest forms of traditional Chinese medicine and it is widely used in clinical practice. However, the quality evaluation and control of traditional decoction is a challenge due to the characteristics of complicated constituents, water as solvent, and temporary preparation. ShenFu Prescription Decoction (SFPD) is a classical prescription for preventing and treating many types of cardiovascular disease. In this article, a comprehensive and rapid method for quality evaluation and control of SFPD was developed, via qualitative and quantitative analysis of the major components by integrating ultra-high-performance liquid chromatography equipped with quadrupole time-of-flight mass spectrometry and ultra-fast-performance liquid chromatography equipped with triple quadrupole mass spectrometry. Consequently, a total of 39 constituents were tentatively identified in qualitative analysis, of which 21 compounds were unambiguously confirmed by comparing with reference substances. We determined 13 important constituents within 7 min by multiple reaction monitoring. The validated method was applied for determining five different proportion SFPDs. It was found that different proportions generated great influence on the dissolution of constituents. This may be one of the mechanisms for which different proportions play different synergistic effects. Therefore, the developed method is a fast and useful approach for quality evaluation of SFPD.

Dendrobium officinale Polysaccharides Inhibit 1-Methyl-2-Nitro-1-Nitrosoguanidine Induced Precancerous Lesions of Gastric Cancer in Rats through Regulating Wnt/ß-Catenin Pathway and Altering Serum Endogenous Metabolites.

Dendrobium officinale is a herb in traditional Chinese medicine where D. officinale polysaccharides (DOP) are the main active ingredient. This study aimed at evaluating DOP efficiency at inhibiting 1-Methyl-2-nitro-1-nitrosoguanidine (MNNG) induced precancerous lesions of gastric cancer (PLGC) in rats through the Wnt/b-catenin pathway and analyzing the variations of serum endogenous metabolites. PLGC was established in male Sprague-Dawley (SD) rats by administering 150 µg/mL MNNG in drinking water for 7 months and giving 0.1 mL of 10% NaCl once weekly during the initial 20 weeks. Treatment with DOP inhibited the progress of PLGC through decreasing the expression of ß-catenin by immunohistochemical analysis. The futher study indicated DOP downregulated gene expression of Wnt2ß, Gsk3ß, PCNA, CyclinD1, and ß-catenin, as well as protein expression of Wnt2ß, PCNA, and ß-catenin. On the other hand, there were nine endogenous metabolites identified after the DOP treatment. Among these, the most significant one is betaine because of its strong antioxidant activity, leading to an anti-tumor effect. DOP can inhibit MNNG-induced PLGC models via regulating Wnt/ß-catenin pathway and by changing endogenous metabolites.

[Optimized model for formulation prescription of traditional Chinese medicine buccal tablets based on temporal dominant description of sensations combined with multivariate statistical analysis:an example of Compound Caoshanhu Buccal Tablets].

The taste is the key to the research and design for formulation prescription of traditional Chinese medicine buccal tablets( TCMBTs). TCMBTs are dissolved in the oral cavity in parallel with the release of the drug,providing a sufficient " time window" for the perception of various basic taste,tactile and retention. The overall taste is the result of competitive inhibition,comprehensive superposition and dynamic change of multiple tastes. Traditional evaluation methods rely mostly on recalled experience score,difficult to reflect the dynamic changes of taste for buccal tablets. Therefore,it is urgent to establish a new optimized model for taste evaluation and formulation prescription according to the formulation characteristics of TCMBTs. To this end,this paper proposed for the first time to construct a research method for the optimal formulation of TCMBTs based on temporal dominant description of sensations combined with multivariate statistical analysis: the sensory test of volunteers was carried out by temporal dominant description analysis method,and elements separation was conducted for the time and taste in the process of inclusion to form a temporal dominant descriptive score of multi-time points and multi-sensory attributes. Finally,the optimal formulation was obtained by multivariate statistical analysis. Taking the formulation prescription of Compound Caoshanhu Buccal Tablets as an example,the research ideas of this method were explained,and the optimal formulation prescription was obtained as follows,Glabrous Sarcandra Extract of 20. 0 g,mannitol of 24. 0 g,microcrystalline cellulose of 12. 0 g,aspartame of 1. 0%,menthol of 0. 7%,and menthol oil of 0. 7%. This study provides a new method for the taste evaluation and formulation research of TCMBTs,providing a new idea for the fine manufacturing and innovative development of TCM buccal tablets in the new era.