The effect of feeding methods on fecal metabolites of infants delivered by cesarean section based on non-targeted metabolomics / 中华实用儿科临床杂志

Objective:To explore the effect of breast feeding versus mixed feeding on fecal metabolites of infants delivered by cesarean section.Methods:This was a cross-sectional study.Fecal samples were collected from 23 healthy 1-month-old infants delivered by cesarean section from autumn 2021 and winter 2022 in two maternal and infant care facilities in the North and South of Xi′an city.The samples were divided into the breast feeding group (11 cases) and mixed feeding group (12 cases). Fecal metabolites were analyzed by the non-targeted metabolomic approach and gas chromatography-mass spectrometry coupling, and differentially expressed fecal metabolites between groups were screened using the non-parametric Mann- Whitney U test.Metabolic pathways enriched in them were further analyzed. Results:A total of 155 metabolites were characterized, including 57 sugars and sugar derivatives, 34 fatty acids, 25 organic acids, 22 amino acids, 8 esters, 4 nucleosides, 3 vitamins and 2 other substances.The relative contents of the differentially expressed fecal metabolites were measured, and it was found that some types of sugars and sugar derivatives were highly expressed in the fecal samples of breast feeding group, while amino acids, organic acids and fatty acids were highly expressed in those of the mixed feeding group.A total of 28 metabolic pathways enriched in differentially expressed fecal metabolites were obtained.Among them, alanine, aspartic acid and glutamic acid metabolism, valine, leucine and isoleucine metabolism, arginine metabolism and the tricarboxylic acid (TCA) cycle influenced infant health.Conclusions:Feeding methods have an effect on the fecal metabolites in infants delivered by cesarean section born infants, and mixed feeding may speed up the process of TCA cycle and amino acid metabolism in the intestine of infants delivered by cesarean section to a certain extent.

Differential analysis of coumarins in Angelica dahurica from different origins / 中国药房

OBJECTIVE To screen the differential components of coumarins in Angelica dahurica from two origins （A. dahurica cv.‘ Hangbaizhi’； A. dahurica cv.‘ Qibaizhi’）. METHODS Non-targeted metabolomics technique of UPLC-Q-Exactive- MS/MS was used to analyze the coumarins in 6 batches of A. dahurica cv. ‘Hangbaizhi’ and 12 batches of A. dahurica cv. ‘Qibaizhi’. The differential components were screened by principal component analysis， partial least squares discriminant analysis and orthogonal partial least squares discriminant analysis. Cluster analysis was performed on differential components. RESULTS A total of 41 coumarins were identified in 18 batches of samples， in which 23 coumarins were differential components. Therein， 6 differential components were higher in content in A. dahurica cv.‘ Hangbaizhi’， while 17 differential components were higher in content in A. dahurica cv.‘ Qibaizhi’. The content of marmesin galactoside in A. dahurica cv.‘ Hangbaizhi’ was significantly higher than that in A. dahurica cv.‘ Qibaizhi’. Based on 23 differential components， A. dahurica cv.‘ Hangbaizhi’ and A. dahurica cv. ‘Qibaizhi’ could be grouped into one category， respectively. CONCLUSIONS The screened differential components of coumarins can be used to distinguish A. dahurica cv. ‘Hangbaizhi’ from A. dahurica cv. ‘Qibaizhi’， especially marmesin galactoside contributed the most， which can be used to identify A. dahurica cv.‘ Hangbaizhi’ and A. dahurica cv.‘ Qibaizhi’.

Non-targeted screening of emerging contaminants in drinking water based on high-resolution mass spectrometry / 上海预防医学

ObjectiveTo establish a non-targeted screening method for emerging contaminants in drinking water based on high-resolution mass spectrometry and apply it to actual water samples. MethodsA total of 9 drinking water samples collected from 3 reservoirs in Shanghai were purified and concentrated by HLB solid phase extraction column， then separated and analyzed by liquid chromatography high-resolution mass spectrometer and gas chromatography high⁃resolution mass spectrometer. The acquired data were analyzed by Thermo Tracefinder， Excel and other software combined with mzCloud and NIST databases. The methodology was verified with representative compound standards. Pesticide and perfluorinated compounds were taken as examples to analyze their pollution status. ResultsA non-targeted analysis strategy based on liquid chromatography and gas chromatography tandem high-resolution mass spectrometry was established. The pollution level of 20 kinds of pesticides and 4 kinds of perfluorinated compounds identified in 9 drinking water samples were higher in the Huangpu River than in the Yangtze River estuary. ConclusionThe established non-targeted screening method by high-resolution mass spectrometry can detect potential emerging contaminants in drinking water without relying on the standards， which provides a powerful technical means for water quality monitoring and risk assessment.

Discovery of biomarkers related to abnormal lipid metabolism in liver and serum and intervention mechanism of ginsenoside Rb_1 in hyperlipidemia rats based on non-targeted metabolomics / 中国中药杂志

Through the non-targeted metabolomics study of endogenous substances in the liver and serum of hyperlipidemia rats, the biomarkers related to abnormal lipid metabolism in hyperlipidemia rats were found, and the target of ginsenoside Rb_1 in improving hyperlipidemia was explored and its mechanism was elucidated. The content of serum biochemical indexes of rats in each group was detected by the automatic biochemical analyzer. The metabolite profiles of liver tissues and serum of rats were analyzed by HPLC-MS. Principal component analysis(PCA) and orthogonal partial least squares-discriminant analysis(OPLS-DA) were used to compare and analyze the metabolic data in the normal group, the hyperlipidemia group, and the ginsenoside Rb_1 group, and screen potential biomar-kers. The related metabolic pathways were further constructed by KEGG database analysis. The results showed that hyperlipemia induced dyslipidemia in rats, which was alleviated by ginsenoside Rb_1. The non-targeted metabolomics results showed that there were 297 differential metabolites in the liver tissues of hyperlipidemia rats, 294 differential metabolites in the serum samples, and 560 diffe-rential metabolites in the hyperlipidemia rats treated by ginsenoside Rb_1. Perillic acid and N-ornithyl-L-taurine were common metabolites in the liver and serum samples, which could be used as potential biomarkers for ginsenoside Rb_1 in the improvement of hyperlipidemia. As revealed by pathway enrichment in the liver and serum, ginsenoside Rb_1 could participate in the metabolic pathway of choline in both the liver and serum. In addition, ginsenoside Rb_1 also participated in the ABC transporter, alanine, aspartic acid, and glutamate metabolism, protein digestion and absorption, β-alanine metabolism, taurine and hypotaurine metabolism, caffeine metabolism, valine, leucine, and isoleucine biosynthesis, arachidonic acid metabolism, and methionine and cysteine metabolism to improve dyslipidemia in rats.

Effect of Shaoyaotang on Fecal Metabolites in Rats with Ulcerative Colitis Based on Liquid Chromatography-mass Spectrometry / 中国实验方剂学杂志

ObjectiveTo investigate the effect of Shaoyaotang on fecal metabolites in rats with ulcerative colitis （UC） induced by 2，4，6-trinitrobenzenesulfonic acid （TNBS） based on liquid chromatography-mass spectrometry （LC-MS）. MethodMale SPF SD rats were randomly divided into normal group， model group and Shaoyaotang group （11.1 g·kg-1）. Except for normal group， UC rat model was induced by TNBS， and each group was given normal saline except Shaoyaotang group. All groups were treated for 7 days， and the general condition and disease activity index （DAI） were observed. Hematoxylin-eosin （HE） staining was used to observe the histopathological changes of colon， and the protein expressions of interleukin-8 （IL-8） and interleukin-22 （IL-22） in colon tissue were detected by immunohistochemistry （IHC）. Rat fecal samples were detected by LC-MS， and the data were analyzed by principal component analysis （PCA） and partial least squares discriminant analysis （PLS-DA）. Human Metabolome Database （HMDB） and Kyoto Encyclopedia of Genes and Genomes （KEGG） were searched to screen differential metabolites in combination with literature reference. Then， pathway enrichment analysis was conducted using Metabo Analyst 5.0. ResultShaoyaotang （ig） decreased the DAI of UC rats. Compared with the normal group， the model group had damaged colonic mucosa structure， submucosal inflammatory cell infiltration， increased protein expressions of IL-8 （P<0.01） and IL-22 （P<0.05） in colon tissue. Compared with the conditions in the model group， the colonic damage was alleviated in the Shaoyaotang group， and the protein expressions of IL-8 and IL-22 in colon tissue were decreased （P<0.01）. After screening， 15 differential metabolites were identified from the Shaoyaotang group， and the involved pathways mainly included biosynthesis of unsaturated fatty acids， linoleic acid metabolism， terpenoid backbone biosynthesis， porphyrin and chlorophyll metabolism， amino sugar and nucleotide sugar metabolism， pyrimidine metabolism and steroid hormone biosynthesis. ConclusionShaoyaotang has a therapeutic effect on UC， and its anti-inflammatory effect may be related to improving lipid metabolism and regulating the metabolism of cofactors and vitamins as well as the abnormal carbohydrate metabolism.

The toxic effects of imidacloprid exposure on HepG2 cell based on non-targeted metabolomics / 环境与职业医学

Background Imidacloprid is a neonicotinoid insecticide that is widely used in agricultural production, with a high detection rate in human biological samples. Previous studies have shown a high correlation between imidacloprid exposure and liver injury, but the specific mechanism is still unknown. Objective To observe potential toxic effects of HepG2 cells and its perturbation of non-targeted metabolic profile after imidacloprid exposure, and to explore possible molecular mechanisms of hepatotoxicity of imidacloprid by analyzing invovlved biological processes and signaling pathways. Methods HepG2 cell suspension was prepared and seeded in a 96-well plate, which was divided into blank control group, dimethyl sulfoxide (DMSO) solvent control group and imidacloprid exposure groups with multiple concentrations. Each group was set with 5 parallel samples. The viability of HepG2 cells viability were determined after 8 h of exposure to different concentrationsof imidacloprid (1, 2.5, 5, 7.5, 10 mmol·L−1), and the dose-effect relationship was analyzed. A proper concentration (3 mmol·L−1 with 80% viability) was chosen for imidacloprid exposure, non-targeted metabolomic analysis was applied to the cultivated HepG2 cells using UHPLC-Q-TOF/MS technology, the differential metabolites between groups were screened, and the bioprocess and related signaling pathways of their enrichment were annotated using the Kyoto Encyclopedia of Genes and Genomes (KEGG) database. Results Compared to the other two groups, the survival rates of HepG2 cells in the imidacloprid exposure groups decreased. A survival rate of about 86% of HepG2 cells was found in HepG2 cells exposed to 2.5 mmol·L−1 imidacloprid exposure. The non-targeted metabolomics studies showed that 61 metabolites were significantly affected in HepG2 cells after 3 mmol·L−1 imidacloprid exposure, including creatine (variable importance in projection VIP=1.11, P<0.001), arginine (VIP=1.47, P=0.048), taurine (VIP=4.28, P=0.001), and α-D-glucose (VIP=1.90, P=0.006). The differential metabolites enriched in bioprocess and related signaling pathways were mainly directed to mTOR signaling pathways (P<0.001), arginine and proline metabolism (P=0.002), and galactose metabolism (P=0.015). Conclusion Imidacloprid exposure can significantly inhibit the survival rate of HepG2 cells, and interfere with the mTOR signaling pathway, arginine and proline metabolism, galactose metabolism, and so on.

Research Progress on Machine Learning Assisted Non-Targeted Screening Strategy for Identification of Fentanyl Analogs / 法医学杂志

In recent years, the types and quantities of fentanyl analogs have increased rapidly. It has become a hotspot in the illicit drug control field of how to quickly identify novel fentanyl analogs and to shorten the blank regulatory period. At present, the identification methods of fentanyl analogs that have been developed mostly rely on reference materials to target fentanyl analogs or their metabolites with known chemical structures, but these methods face challenges when analyzing new compounds with unknown structures. In recent years, emerging machine learning technology can quickly and automatically extract valuable features from massive data, which provides inspiration for the non-targeted screening of fentanyl analogs. For example, the wide application of instruments like Raman spectroscopy, nuclear magnetic resonance spectroscopy, high resolution mass spectrometry, and other instruments can maximize the mining of the characteristic data related to fentanyl analogs in samples. Combining this data with an appropriate machine learning model, researchers may create a variety of high-performance non-targeted fentanyl identification methods. This paper reviews the recent research on the application of machine learning assisted non-targeted screening strategy for the identification of fentanyl analogs, and looks forward to the future development trend in this field.

Study on biomarkers of acteoside in treating puromycin aminonucleoside nephropathy in young rats based on non-targeted urine metabolomics technology / 中国中药杂志

This study aims to reveal the endogenous metabolic characteristics of acteoside in the young rat model of purinomycin aminonucleoside nephropathy(PAN) by non-targeted urine metabolomics and decipher the potential mechanism of action. Biochemical indicators in the urine of rats from each group were determined by an automatic biochemical analyzer. The potential biomarkers and related core metabolic pathways were identified by ultra-high performance liquid chromatography coupled with linear ion trap-Orbitrap mass spectrometry(UHPLC-LTQ-Orbitrap MS) combined with principal component analysis(PCA) and orthogonal partial least squares-discriminant analysis(OPLS-DA). MetaboAnalyst 5.0 was used to establish the receiver operating characteristic(ROC) curve for evaluating the clinical diagnostic performance of core metabolites. The results showed that acteoside significantly decreased urinary protein-to-creatinine ratio in PAN young rats. A total of 17 differential metabolites were screened out by non-targeted urine metabolomics in PAN young rats and they were involved in phenylalanine metabolism and phenylalanine, tyrosine and tryptophan biosynthesis. Thirtten differential metabolites were screened by acteoside intervention in PAN young rats, and they were involved in phenylalanine metabolism and arginine and proline metabolism. Among them, leucylproline and acetophenone were the differential metabolites that were significantly recovered after acteoside treatment. These pathways suggest that acteoside treats PAN in young rats by regulating amino acid metabolism. The area under the curve of two core biomarkers, leucylproline and acetophenone, were both greater than 0.9. In summary, acteoside may restore amino acid metabolism by regulating endogenous differential metabolites in PAN young rats, which will help to clarify the mechanism of acteoside in treating chronic glomerulonephritis in children. The characteristic biomarkers screened out have a high diagnostic value for evaluating the treatment of chronic glomerulonephritis in children with acteoside.

Therapeutic effect and mechanism of Mailuo Shutong Pills on posterior limb swelling caused by femur fracture in rats based on intestinal flora and intestinal metabolism / 中国中药杂志

This study aimed to investigate the protective effect and underlying mechanism of Mailuo Shutong Pills(MLST) on posterior limb swelling caused by femur fracture in rats. The rats were randomly divided into a sham operation group, a model group, a low-dose MLST group(1.8 g·kg~(-1)·d~(-1)), a high-dose MLST group(3.6 g·kg~(-1)·d~(-1)), and a positive drug group(60 mg·kg~(-1)·d~(-1) Maizhiling Tablets). The femur in the sham operation group was exposed and the wound was sutured, while the other four groups underwent mechanical damage to cause femur fracture. The rats were treated with corresponding drugs by gavage 7 days before modeling and 5 days after modeling, while those in the sham operation group and the model group were given an equivalent dose of distilled water by gavage. Hematoxylin-eosin(HE) staining was used to detect the pathological injury of the posterior limb muscle tissues in rats, and the degree of hind limb swelling was measured. The enzyme-linked immunosorbent assay(ELISA) kit was used to detect the expression levels of interleukin-6(IL-6), interleukin-1β(IL-1β), and tumor necrosis factor-α(TNF-α) in the serum of rats in each group. The activity of superoxide dismutase(SOD), malondialdehyde(MDA), catalase(CAT), and glutathione peroxidase(GSH-Px) in rat serum was also measured. Western blot was used to detect the protein expression levels of heme oxygenase 1(HO-1), NAD(P)H quinone oxidoreductase 1(NQO1), and nuclear transcription factor E2-related factor 2(Nrf2) in rat posterior limb muscle tissues. The changes in the intestinal flora and intestinal metabolites in rats were detected by 16S rDNA sequencing and ultra-performance liquid chromatography-tandem mass spectrometry(UPLC-MS/MS), respectively, to explore the underlying mechanism of MLST in treating posterior limb swelling caused by femur fracture in rats. Compared with the model group, MLST significantly improved the degree of posterior limb swelling in rats, reduced the levels of serum inflammatory factors, and alleviated oxidative stress injury. The HE staining results showed that the inflammatory infiltration in the posterior limb muscle tissues of rats in the MLST groups was significantly improved. Western blot results showed that MLST significantly increased the protein expression of HO-1, NQO1, and Nrf2 in rat posterior limb muscle tissues compared with the model group. The 16S rDNA sequencing results showed that MLST improved the disorder of intestinal flora in rats after femur fracture. The UPLC-MS/MS results showed that MLST significantly affected the bile acid biosynthesis and metabolism pathway in the intestine after femur fracture, and the Spearman analysis confirmed that the metabolite deoxycholic acid involved in bile acid biosynthesis was positively correlated with the abundance of Turicibacter. The metabolite cholic acid was positively correlated with the abundance of Papilibacter, Staphylococcus, and Intestinimonas. The metabolite lithocholic acid was positively correlated with Papilibacter and Intestinimonas. The above results indicated that MLST could protect against the posterior limb swelling caused by femur fracture in rats. This protective effect may be achieved by improving the pathological injury of the posterior limb muscle, reducing the expression levels of inflammatory and oxidative stress-related factors in serum, reducing the oxidative injury of the posterior limb muscle, improving intestinal flora, and balancing the biosynthesis of bile acids in the intestine.

UPLC-Q-TOF-MS metabolomic study on improvement of acute myocardial ischemia in rats by Dalbergia cochinchinensis heartwood / 中国中药杂志

This paper aimed to study the effect of Dalbergia cochinchinensis heartwood on plasma endogenous metabolites in rats with ligation of the left anterior descending coronary artery, and to analyze the mechanism of D. cochinchinensis heartwood in improving acute myocardial ischemic injury. The stability and consistency of the components in the D. cochinchinensis heartwood were verified by the establishment of fingerprint, and 30 male SD rats were randomly divided into a sham group, a model group, and a D. cochinchinensis heartwood(6 g·kg~(-1)) group, with 10 rats in each group. The sham group only opened the chest without ligation, while the other groups established the model of ligation. Ten days after administration, the hearts were taken for hematoxylin-eosin(HE) staining, and the content of heart injury indexes in the plasma creatine kinase isoenzyme(CK-MB) and lactate dehydrogenase(LDH), energy metabolism-related index glucose(Glu) content, and vascular endothelial function index nitric oxide(NO) was determined. The endogenous metabolites were detected by ultra-high-performance liquid chromatography-time-of-flight-mass spectrometry(UPLC-Q-TOF-MS). The results showed that the D. cochinchinensis heartwood reduced the content of CK-MB and LDH in the plasma of rats to relieve myocardial injury, reduced the content of Glu in the plasma, improved myocardial energy metabolism, increased the content of NO, cured the vascular endothelial injury, and promoted vasodilation. D. cochinchinensis heartwood improved the increase of intercellular space, myocardial inflammatory cell infiltration, and myofilament rupture caused by ligation of the left anterior descending coronary artery. The metabolomic study showed that the content of 26 metabolites in the plasma of rats in the model group increased significantly, while the content of 27 metabolites decreased significantly. Twenty metabolites were significantly adjusted after the administration of D. cochinchinensis heartwood. D. cochinchinensis heartwood can significantly adjust the metabolic abnormality in rats with ligation of the left anterior descending coronary artery, and its mechanism may be related to the regulation of cardiac energy metabolism, NO production, and inflammation. The results provide a corresponding basis for further explaining the effect of D. cochinchinensis on the acute myocardial injury.

Antipyretic activity and potential mechanism of Indigo Naturalis on 2,4-dinitrophenol-induced fever rat model / 中国中药杂志

As an effective antipyretic medicine,Indigo Naturalis has a long history of application in the field of Chinese medicine.The content of organics,mainly indigo and indirubin,is about 10%. However,the active ingredients and mechanism of its antipyretic effect have not yet been fully elucidated. In view of this,they were investigated in this study with the rectal temperature change as an indicator and 2,4-dinitrophenol-induced fever rats as subjects. The content of PGE2 and c AMP in the hypothalamus and the serum levels of TNF-α,IL-1β and IL-6 were determined by ELISA. Moreover,the plasma samples of fever rats were analyzed by metabonomics in combination with UPLC-Q-TOF-MS for the exploration of potential biomarkers and the discussion on the antipyretic mechanism of Indigo Naturalis and its active ingredients. The results showed that the rising trend of rectal temperature in rats was suppressed 0. 5 h after the treatment with Indigo Naturalis,organic matter,indigo or indirubin as compared with the rats of model group( P < 0. 05),among which Indigo Naturalis and organic matter had better antipyretic effect. ELISA results showed that organic matter and indigo can inhibit the expression of PGE2 and c AMP( P<0. 01),while Indigo Naturalis and organic matter were effective in curbing the increase in TNF-α( P<0. 05). A total of 21 endogenous metabolites were identified from the plasma samples of the Indigo Naturalis,organic matter,indigo and indirubin groups,which were mainly involved in glycerophospholipid metabolism.

Screening of serum susceptibility biomarkers in patients with mild liver function abnormalities based on non-targeted metabolomics / 中国中药杂志

In the present study,non-targeted metabolomics technique was used to screen potentially susceptibility biomarkers in patients with mild liver function abnormalities during long-term use of Chinese herbal compound. According to the inclusion and exclusion criteria,we collected 7 cases of patients with abnormal liver function during the period of complete taking Chinese herbal medicine( 60 days),and 18 cases of patients with normal liver function in re-examination from the reproductive medicine center in our hospital. Ultra performance liquid chromatography coupled with time-of-flight mass spectrometry( UPLC-Q-TOF/MS~E) technique combined with Progenesis QI software was used to analyze the differential biomarkers in serum of patients with wild liver function abnormalities and normal liver function. 11 potential biomarkers such as bilirubin,pantothenic acid,hippuric acid,sphingomyelin,palmitic acid,and oleic acid were tentatively identified. Metabolic disorders in patients with herbal-induced mild liver abnormality were mainly related to two pathways: pantothenic acid and coenzyme A biosynthesis and linoleic acid metabolism. It could provide a reference for the early warning of mild liver function abnormalities of patients that may be caused by long-term use of Chinese medicine compound in clinical application,and will lay a foundation for further understanding the endogenous substance changes in different levels of liver injury.

Effect of Manual and Machine Decocting on Chemical Constituents in Bazhentang Based on Non-targeted Metabolomics / 中国实验方剂学杂志

Objective: To evaluate the effect of manual decocting and machine decocting on the chemical constituents in Bazhentang based on non-targeted metabolomics, and to find the differential chemical constituents of these two decocting methods. Method: Bazhentang was boiled by standardized manual decocting and machine decocting methods,respectively. Orthogonal partial least square-discriminate analysis(OPLS-DA) and other multivariate statistical methods, combined with variable importance in the projection(VIP) value and t-test, were employed to analyze the effect of two decocting methods on the chemical constituents in Bazhentang. The differential chemical constituents were analyzed by UPLC-LTQ-Orbitrap MS under positive and negative ion modes,mobile phase was acetonitrile-0.1%formic acid aqueous solution for gradient elution,the scanning range was m/z 50-1 500. Result: Under the positive and negative ion modes of high-resolution mass spectrometry, a total of 87 differential components were found,40 of them were identified according to the mass spectrometry data and literature reports, including senkyunolide A, glycyrrhizin, ferulic acid, etc. Conclusion: Based on the analysis of color and chemical compositions of Bazhentang, there are obvious differences between the standardized manual decocting and machine decocting. If the advantages of these two methods are combined,a standardized decoction process can be established on the basis of maintaining the advantages of manual decocting, the effectiveness of traditional Chinese medicine decoction will be maximized and it will be convenient for patients to take it.