Evaluation of A Liver Yin Deficiency Mouse Model Based on Untargeted Metabolomics / 中国实验方剂学杂志

ObjectiveBased on ultra performance liquid chromatography-quadrupole-time-of-flight tandem mass spectrometry（UPLC-Q-TOF-MS）， to evaluate the establishment of a mouse model of liver Yin deficiency by thyroid tablet suspension combined with 10% carbon tetrachloride（CCl4） from the perspective of non-targeted metabolomics， in order to lay the foundation for the establishment of a traditional Chinese medicine（TCM） syndrome model. MethodA total of 24 mice were randomly divided into blank group and model group. The model group was given thyroid tablet suspension（0.003 2 g·kg-1） by gavage for 14 consecutive days， and 10% CCl4（5 mL·kg-1） was intraperitoneally injected once a week to establish a liver Yin deficiency model， while the blank group was injected with an equal amount of olive oil intraperitoneally and gavaged with an equal amount of distilled water， and was fed with normal feed. After the modeling was completed， 6 mice in each group were randomly selected， the levels of alanine aminotransferase（ALT）， aspartate aminotransferase（AST）， cyclic adenosine monophosphate（cAMP）， cyclic guanosine monophosphate（cGMP）， interleukin（IL）-6， IL-10， tumor necrosis factor-α（TNF-α）were measured in the mice serum， and malondialdehyde（MDA）， superoxide dismutase（SOD）， total protein（TP）， hydroxyproline（HYP） and other indicators were measured in the mice liver. Liver tissue sections were taken for hematoxylin-eosin（HE） staining and observing pathological changes. The remaining 6 mice in each group were subjected to UPLC-Q-TOF-MS combined with principal component analysis（PCA） and orthogonal partial least squares-discriminant analysis（OPLS-DA） were used to screen differential metabolites in the liver Yin deficiency mouse model， Kyoto Encyclopedia of Genes and Genomes（KEGG） database was used to analyze the corresponding metabolic pathways of differential metabolites. ResultCompared with the blank group， mice in the model group showed liver Yin deficiency manifestations such as reduced body weight， fatigue and sleepiness， disheveled and lusterless hair， irritability. The levels of ALT， cAMP/cGMP， IL-6， AST， MDA， cAMP， TNF-α significantly increased（P<0.05， P<0.01）， while the levels of SOD， IL-10 and cGMP significantly decreased（P<0.05， P<0.01）， and the changes of HYP and TP were not statistically significant. Hepatic steatosis and distortion of the radial arrangement of the liver plate cells were seen in the section images of the model group， endogenous substances were clearly separated， and 252 differential metabolites were identified in the serum samples， which were mainly involved in the metabolic pathways of purine metabolism， steroid hormone biosynthesis and pyrimidine metabolism. A total of 229 differential metabolites were identified in the liver samples， mainly involving nucleotide metabolism， purine metabolism， steroid hormone biosynthesis， pyrimidine metabolism， antifolate resistance， insulin resistance， primary bile acid biosynthesis， prostate cancer， sulfur relay system， arachidonic acid metabolism and other metabolic pathways. ConclusionThe successful establishment of liver Yin deficiency model in mice by CCl4 combined with thyroid hormone is evaluated through the investigation of serum and liver metabolomics， combined with biochemical indicators， which provides a biological basis and experimental foundation for the Yin deficiency syndrome model of TCM.

Main ways and target prediction of detoxification mechanism of Euphorbia kansui after stir-frying with vinegar / 中国药房

OBJECTIVE To study main way and target of Euphorbia kansui after stir-frying with vinegar. METHODS Twenty-four SPF grade SD rats were randomly divided into blank group， E. kansui group （850 mg/kg） and vinegar stir-fried E. kansui group （850 mg/kg）， with 8 rats in each group. Blank group was given 0.5% sodium carboxymethyl cellulose solution intragastrically， and E. kansui group and vinegar stir-fried E. kansui group were given relevant test sample for consecutive 20 d. The rats’ urine of 12 hours was collected on the 20th day. The urine samples of rats in each group were determined by UPLC-Q- Exactive-MS. The data was pre-processed by Compound Discoverer 3.0 software， and the metabolite structure was identified by BioCyc， HMDB and other databases. Whether different groups presented their own clustering phenomenon was observed by principal component analysis （PCA） and orthogonal partial least squares-discriminant analysis （OPLS-DA）， etc. Based on the pathway analysis of MetaboAnalyst， the potential targets of detoxification mechanism of E. kansui after stir-frying with vinegar were predicted. RESULTS Twenty significantly differential endogenous metabolites were identified， of which 10 target metabolites， such as N-acetyl-L-aspartate and 3-phosphonooxypyruvic acid， were targets of detoxification mechanism of E. kansui after stir- frying with vinegar. The main metabolic pathways included arginine biosynthesis， alanine， aspartic acid and glutamic acid metabolism， cysteine and methionine metabolism， and arginine and D-ornithine metabolism. The biological significance of all related metabolites in the pathways was analyzed and speculated； after stir-frying with vinegar， E. kansui may alleviate neurotoxicity by reducing the level of N-acetyl-L-aspartic acid； E. kansui had a protective effect on cardio-cerebrovascular system by increasing the level of L-high arginine. CONCLUSIONS After stir-frying with vinegar， E. kansui can significantly improve the adverse factors in terms of nervous system， cardio-cerebrovascular system， immune system and energy metabolism. The most concentrated metabolic pathway related to its detoxification mechanism is arginine biosynthesis.

Combined analysis of transcriptomics and metabolomics revealed complex metabolic genes for diterpenoids biosynthesis in different organs of Anoectochilus roxburghii / 中草药·英文版

OBJECTIVE@#Diterpenoids with a wide variety of biological activities from Anoectochilus roxburghii, a precious medicinal plant, are important active components. However, due to the lack of genetic information on the metabolic process of diterpenoids in A. roxburghii, the genes involved in the molecular regulation mechanism of diterpenoid metabolism are still unclear. This study revealed the complex metabolic genes for diterpenoids biosynthesis in different organs of A. roxburghii by combining analysis of transcriptomics and metabolomics.@*METHODS@#The differences in diterpenoid accumulation in roots, stems and leaves of A. roxburghii were analyzed by metabonomic analysis, and its metabolic gene information was obtained by transcriptome sequencing. Then, the molecular mechanism of differential diterpenoid accumulation in different organs of A. roxburghii was analyzed from the perspective of gene expression patterns.@*RESULTS@#A total of 296 terpenoid metabolites were identified in the five terpenoid metabolic pathways in A. roxburghii. There were 38, 34, and 18 diterpenoids with different contents between roots and leaves, between leaves and stems, and between roots and stems, respectively. Twenty-nine metabolic enzyme genes with 883 unigenes in the diterpenoid synthesis process were identified, and the DXS and FDPS in the terpenoid backbone biosynthesis stage and CPA, GA20ox, GA3ox, GA2ox, and MAS in the diterpenoid biosynthesis stage were predicted to be the key metabolic enzymes for the accumulation of diterpenoids. In addition, 14 key transcription factor coding genes were predicted to be involved in the regulation of the diterpenoid biosynthesis. The expression of genes such as GA2ox, MAS, CPA, GA20ox and GA3ox might be activated by some of the 14 transcription factors. The transcription factor NTF-Y and PRE6 were predicted to be the most important transcription factors.@*CONCLUSION@#This study determined 29 metabolic enzyme genes and predicted 14 transcription factors involved in the molecular regulation mechanism of diterpenoid metabolism in A. roxburghii, which provided a reference for the further study of the molecular regulation mechanism of the accumulation of diterpenoids in different organs of A. roxburghii.

Chromatin Remodeling Factor SMARCA5 is Essential for Hippocampal Memory Maintenance via Metabolic Pathways in Mice / 神经科学通报·英文版

Gene transcription and new protein synthesis regulated by epigenetics play integral roles in the formation of new memories. However, as an important part of epigenetics, the function of chromatin remodeling in learning and memory has been less studied. Here, we showed that SMARCA5 (SWI/SNF related, matrix-associated, actin-dependent regulator of chromatin, subfamily A, member 5), a critical chromatin remodeler, was responsible for hippocampus-dependent memory maintenance and neurogenesis. Using proteomics analysis, we found protein expression changes in the hippocampal dentate gyrus (DG) after the knockdown of SMARCA5 during contextual fear conditioning (CFC) memory maintenance in mice. Moreover, SMARCA5 was revealed to participate in CFC memory maintenance via modulating the proteins of metabolic pathways such as nucleoside diphosphate kinase-3 (NME3) and aminoacylase 1 (ACY1). This work is the first to describe the role of SMARCA5 in memory maintenance and to demonstrate the involvement of metabolic pathways regulated by SMARCA5 in learning and memory.

Mechanism of famous classical formula Huaihua Powder in treatment of ulcerative colitis based on metabonomics / 中国中药杂志

Ultra-high performance liquid chromatography-quadrupole-time of flight tandem mass spectrometry(UHPLC-Q-TOF-MS) was employed in this study to observe the effect of Huaihua Powder on the serum metabolites of mice with ulcerative colitis and reveal the mechanism of Huaihua Powder in the treatment of ulcerative colitis. The mouse model of ulcerative colitis was established by dextran sodium sulfate salt(DSS). The therapeutic effect of Huaihua Powder on ulcerative colitis was preliminarily evaluated based on the disease activity index(DAI), colon appearance, colon tissue morphology, and the content of inflammatory cytokines such as tumor necrosis factor-α(TNF-α), interleukin-6(IL-6), and interleukin-1β(IL-1β). UHPLC-Q-TOF-MS was employed to profile the endogenous metabolites of serum samples in blank control group, model group, and low-, medium-, and high-dose Huaihua Powder groups. Multivariate analyses such as principal component analysis(PCA), partial least squares discriminant analysis(PLS-DA), and orthogonal partial least squares discriminant analysis(OPLS-DA) were performed for pattern recognition. Potential biomarkers were screened by Mass Profiler Professional(MPP) B.14.00 with the thresholds of fold change≥2 and P<0.05. The metabolic pathways were enriched by MetaboAnalyst 5.0. The results showed that Huaihua Powder significantly improved the general state and colon tissue morphology of mice with ulcerative colitis, reduced DAI, and lowered the levels of TNF-α, IL-6, and IL-1β in serum. A total of 38 potential biomarkers were predicted to be related to the regulatory effect of Huaihua Powder, which were mainly involved in glycerophospholipid metabolism, glycine, serine, and threonine metabolism, mutual transformation of glucuronic acid, and glutathione metabolism. This study employed metabolomics to analyze the mechanism of Huaihua Powder in the treatment of ulcerative colitis, laying a foundation for the further research.

Phenylpropanoid pathway in plants and its role in response to heavy metal stress: a review / 生物工程学报

Phenylpropanoid metabolic pathway is one of the most important secondary metabolic pathways in plants. It directly or indirectly plays an antioxidant role in plant resistance to heavy metal stress, and can improve the absorption and stress tolerance of plants to heavy metal ions. In this paper, the core reactions and key enzymes of the phenylpropanoid metabolic pathway were summarized, and the biosynthetic processes of key metabolites such as lignin, flavonoids and proanthocyanidins and relevant mechanisms were analyzed. Based on this, the mechanisms of key products of phenylpropanoid metabolic pathway in response to heavy metal stress were discussed. The perspectives on the involvement of phenylpropanoid metabolism in plant defense against heavy metal stress provides a theoretical basis for improving the phytoremediation efficiency of heavy metal polluted environment.

Investigation on the mechanism of Compound zaoren granules in improving insomnia based on serum meta-bonomics / 中国药房

OBJECTIVE To study the mechanism of Compound zaoren granule in improving insomnia. METHODS Forty-nine mice were divided into blank group， model group， positive control group 1 （Estazolam tablets 0.5 mg/kg），control group 2 （Shumian capsule 0.6 g/kg）， Compound zaoren granule low-dose， medium-dose and high-dose groups （2.5， 5， 10 g/kg）， with 7 mice in each group. The insomnia model was established by chronic unpredictable mild stress combined with 4-chloro-DL- phenylacetic acid. The behavioral changes of mice were investigated through open field test and pentobarbital sodium synergistic hypnosis experiment， as well as the pathomorphology of mice hypothalamus tissue was observed by HE staining. The metabonomics analysis and multivariate statistical analysis of serum in mice were performed by UHPLC-Q-TOF-MS/MS， and the differential metabolites were screened out； the metabolic pathway analysis was conducted based on MetaboAnalyst 5.0 database. RESULTS Compared with blank group， the total travelling distance， the number of entering the central region and the moving distance in the central region of the model group were significantly reduced （P＜0.05）， the proportion of total rest time was significantly increased （P＜0.05）， the sleep duration of mice was significantly shortened （P＜0.05）， and hypothalamic nerve cells damaged and severely vacuolated. Compared with model group， the total travelling distance of Compound zaoren granule low-dose and medium-dose groups were increased significantly and the proportions of total rest time of those groups were decreased significantly （P＜0.05）， and the sleep duration of mice in Compound zaoren granule high-dose group was prolonged significantly （P＜0.05）； the hypothalamic nerve cells of mice in each administration group recovered to varying degrees， and the hypothalamus histiocytes of mice in the Compound zaoren granules high-dose group were closer to those in the blank group. A total of 18 differential metabolites （such as phenylalanine， taurine， norvaline， methionine） and 4 important amino acid metabolic pathways （L-phenylalanine， tyrosine and tryptophan biosynthesis； taurine and hypotaurine metabolism； L-phenylalanine metabolism； cysteine and methionine metabolism） were identified through metabolomics analysis. CONCLUSIONS Compound zaoren granules can normalize the disordered metabolism in vivo by regulating differential metabolites such as phenylalanine， taurine， and four amino acid metabolic pathways， so as to improve insomnia.

Research progress on metabonomics of traditional Chinese medicine in the treatment of ulcerative colitis / 中国药房

OBJECTIVE Ulcerative colitis （UC）， as a common and refractory disease of the digestive system， has always been a hot and difficult point in medical research. Traditional Chinese medicine has the advantages of good efficacy， high safety and not easy to relapse after drug withdrawal in the treatment of UC， but the mechanism has not been fully elucidated. Metabonomics looks for potential biomarkers and metabolic pathways from the point of view of the endogenous dynamic metabolism of the whole body， which is helpful to evaluate the efficacy of drugs and explore related mechanisms. Metabolomics studies on the treatment of UC with traditional Chinese medicine have shown that traditional Chinese medicine formulas， single herbs and herbs monomers act on various related pathways such as amino acid metabolism， lipid metabolism and energy metabolism by regulating endogenous metabolites in the body， thereby inhibiting immune inflammatory reactions， improving oxidative stress， reducing intestinal sensitivity， regulating intestinal microbiota， repairing intestinal mucosal damage， and restoring normal metabolic activity in the body. However， further screening and validation of relevant metabolic markers are needed.

Metabolomics study on occupational acute methyl acetate poisoning using patient plasma / 中国职业医学

Objective: To investigate the specificity of endogenous metabolic profile in plasma of patients with occupational acute methyl acetate poisoning using non-targeted metabolomics. Methods: A total of six patients with occupational acute methyl acetate poisoning were selected as the poisoning group, while 10 healthy workers without occupational exposure history of chemical hazards in the same industry were selected as the control group using the judgment sampling method. Metabolites in patient plasma of the two groups were detected using ultra-high performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry, and non-targeted metabolomics analysis was performed. Principal component analysis and partial least squares discriminant analysis were used to identify differential metabolites and analyze their metabolic pathways. Results: There were significant differences in metabolite profiles in patient plasma between poisoning group and control group. A total of 195 differentially expressed metabolites were screened in plasma of patients in poisoning group, including 119 upregulated and 76 downregulated metabolites. Lipid substances (lipids and lipid-like molecules) accounted for the highest proportion (21.5%). The differential metabolites of poisoning group were related to folate biosynthesis, amino acid metabolism, pyrimidine metabolism, sphingolipid biosynthesis and other metabolic pathways in plasma compared with the control group (all P<0.05). Conclusion: Occupational acute methyl acetate poisoning affects metabolism of the body. The folic acid biosynthesis, amino acid and lipid metabolism and other pathways may be involved in the occurrence and development of poisoning.

Preparation of Fe3O4@SW-MIL-101-NH2 for selective pre-concentration of chlorogenic acid metabolites in rat plasma,urine,and feces samples / 药物分析学报

An innovative sandwich-structural Fe-based metal-organic framework magnetic material(Fe3O4@SW-MIL-101-NH2)was fabricated using a facile solvothermal method.The characteristic properties of the material were investigated by field emission scanning electron microscopy,transmission electron mi-croscopy(TEM),energy-dispersive X-ray spectroscopy,Fourier transform infrared spectroscopy,X-ray powder diffraction,vibrating sample magnetometry,and Brunauer-Emmett-Teller measurements.Fe3O4@SW-MIL-101-NH2 is associated with advantages,such as robust magnetic properties,high specific surface area,and satisfactory storage stability,as well as good selective recognition ability for chlorogenic acid(CA)and its metabolites via chelation,hydrogen bonding,and π-interaction.The results of the static adsorption experiment indicated that Fe3O4@SW-MIL-101-NH2 possessed a high adsorption capacity toward CA and its isomers,cryptochlorogenic acid(CCA)and neochlorogenic acid(NCA),and the adsorption behaviors were fitted using the Langmuir adsorption isotherm model.Then,a strategy using magnetic solid-phase extraction(MSPE)and ultra-performance liquid chromatography coupled with quadrupole time-of-flight tandem mass spectrometry(UPLC-Q-TOF MS/MS)was developed and suc-cessfully employed for the selective pre-concentration and rapid identification of CA metabolites in rat plasma,urine,and feces samples.This work presents a prospective strategy for the synthesis of magnetic adsorbents and the high-efficiency pretreatment of CA metabolites.

Investigation of metabolites of synthetic cannabinoid 4CN-MDMB-BUTINAC in vitro by liquid-chromatography-quadruple/orbitrap mass spectrometry / 药学学报

To investigate the metabolites of a new synthetic cannabinoid 3,3-dimethyl-2-[1-(4-cyanobutyl)indazole-3-formamimino]methyl butyrate (4CN-MDMB-BUTINACA) in vitro, a human liver microsome incubation model was established to analyze the metabolic biotransformation of synthetic cannabinoids using ultra-high performance liquid chromatography coupled to quadrupole-orbitrap high-resolution mass spectrometry. Nontarget metabolomic results showed that the metabolites of 4CN-MDMB-BUTINACA included hydroxylation, ester hydrolysis, ester hydrolysis with hydroxylation reaction, pentane oxidation and ester hydrolysis with pentane oxidation reaction, among which M1-a, M2 and M4 were potential metabolic markers. The research results provide a theoretical basis and technical support for the biomonitoring and metabolic characterisation of the cannabinoid 4CN-MDMB-BUTINACA.

Anaerobic digestion pathways in biochemistry: a teaching practice / 生物工程学报

Anaerobic digestion is another important anaerobic catabolism pathway besides lactic acid and ethanol fermentation, which is of great significance for recycling resources, maintaining the ecological balance, optimizing the energy structure, alleviating the energy crisis, and promoting the implementation of the "Carbon Peaking and Carbon Neutrality" strategy. However, such an important metabolic process has not been involved in the current textbooks and teaching of biochemistry courses, making the teaching system incomplete. The anaerobic digestion process involves many reactions and complex metabolic pathways. In order to improve the students' understanding to this process, we created a full chart of the whole anaerobic digestion process based on systemic literature review and integrated it into the classroom teaching through the BOPPPS teaching mode. It was found that the classroom teaching assisted by this metabolic chart could help students establish the structural framework of the anaerobic digestion process and enrich the knowledge system of metabolism, achieving a good teaching effect. This paper introduces the content of the metabolic pathways of anaerobic digestion and the design of the teaching process, which would facilitate the teaching reforms and perfection of textbooks for related courses, such as Biochemistry, Environmental Engineering Microbiology and New Energy Engineering.

Microorganisms capable of degrading neonicotinoids and their metabolic pathways: a review / 生物工程学报

Neonicotinoid compounds are usually considered harmless and eco-friendly in terms of their targeted toxicity compared to that of pyrethroids and phosphorus-containing pesticides. However, overuse of neonicotinoid insecticides resulted in the accumulation of its residuals or intermediates in soil and water, which consequently affected beneficial insects as well as mammals, yielding pollution and secondary risks. This review summarized the recent advances in neonicotinoid degrading microorganisms and their metabolic diversity, with the aim to address the urgent need for degrading these insecticides. These advances may facilitate the development of controllable and reliable technologies for efficiently transforming neonicotinoid insecticides into value-added products by synthetic biology and metagenomics.

Research progress of metabolic flux analysis technology in glucose metabolism pathway research / 药学学报

With the wide application of stable isotope tracer metabolomics technology, its comprehensive analysis and in-depth mining of data are particularly important, and metabolic flux analysis is one of the main technical means, especially in the study of glucose metabolism. Metabolic flux analysis technology combines isotope tracing with mathematical models to deduce and calculate the metabolic flux between metabolites. The metabolic flux provides more information for research and reflects a dynamic metabolic process more clearly and specifically. This paper reviews the basic process, precautions, and application examples of metabolic flux analysis in glucose metabolism research, and provides a reference for the application of metabolic flux analysis based on stable isotope tracer metabolomics in glucose metabolism research.

Effect of Kansui Radix stir-fried with vinegar on fecal metabolomics in Walker-256 rats with ascites / 中国中药杂志

Utilizing metabolomics technology, this study explored the change of fecal endogenous metabolites in Walker-256 rats with malignant ascites after the administration with Kansui Radix(KR) stir-fried with vinegar(VKR), sought the potential biomarkers in feces which were related to the treatment of malignant ascites by VKR and revealed the biological mechanism of water-expelling effect of VKR. Ultra-fast liquid chromatography-quadrupole-time-of-flight mass spectrometry(UFLC-Q-TOF-MS) was employed to detect the feces of rats in all groups. Principle component analysis(PCA) and partial least squares discriminant analysis(PLS-DA) were conducted to achieve pattern recognition. Combining t-test and variable importance in the projection(VIP) enabled the screening of potential biomarkers for the malignant ascites. Metabolic pathway analysis was accomplished with MetaboAnalyst. Correlation analysis was finally conducted integrating the sequencing data of gut microbiota to elucidate the mechanism underlying the water-expelling effect of VKR. The results showed that both KR and VKR could restore the abnormal metabolism of model rats to some extent, with VKR being inferior to KR in the regulation. Eleven potential biomarkers were identified to be correlated with the malignant ascites and five metabolic pathways were then enriched. Four kinds of gut microbiota were significantly related to the potential biomarkers. The water-expelling effect of VKR may be associated with the regulation of phenylalanine metabolism, biosynthesis of phenylalanine, tyrosine and tryptophan, tryptophan metabolism, glycerophospholipid metabolism, and glycosylphosphatidylinositol(GPI)-anchor biosynthesis. This study can provide a scientific basis for comprehensive understandings of the interaction between gut microbiota and host which has relation to the water-expelling effect of VKR and guide the reasonable clinical application of VKR.

Application of metabonomics in study of traditional Chinese medicine syndrome:a review / 中国中药杂志

Syndrome is a nonlinear "internal-excess external-deficiency", "dynamic spatial-temporal" and "multi-dimensional" complex system and thus only by using a versatile method can the connotation be expounded. Metabonomics, which is dynamic, holistic, and systematic, is consistent with the overall mode of traditional Chinese medicine(TCM)(holistic view and syndrome differentiation and treatment). Therefore, metabonomics is very important for the research on the differentiation, material basis, and metabolic pathways of syndromes, and efficacy on syndromes. This study reviewed the application of metabonomics in the study of TCM syndromes in recent years, which is expected to objectify the research on TCM syndromes.

Microbial synthesis of monoterpenoids: a review / 生物工程学报

Monoterpenoids that belong to the terpenoids family are usually volatile and have strong aroma. Some monoterpenoids also have antioxidant, antibacterial and anti-inflammatory activities, which make them important raw materials for medicine, food and cosmetics industry. In recent years, the heterologous synthesis of monoterpenoids by microorganisms has attracted extensive attention. However, its large-scale application is greatly hampered by the low yield and high production cost. Nowadays, the rapid development of synthetic biology provides new approaches for enhancing the production of monoterpenoids by microorganisms. Different kinds of recombinant strains can be obtained via engineering of microbial cells to produce a variety of monoterpenoids with different properties. This paper summarized the latest strategies and progress in the application of synthetic biology to produce monoterpenoids by microorganisms, including the design and modification of biosynthetic pathway, as well as the design and optimization of high-yield monoterpenoids producing chassis cells.

Metabolism of New Psychoactive Substances 4F-MDMB-BUTINACA in Zebrafish / 法医学杂志

Objective To study the metabolic transformation pathways of 4F-MDMB-BUTINACA in vivo by establishing zebrafish models. Methods Six adult zebrafish were randomly divided into blank control group and experimental group, with three fish in each group. After the zebrafish in the experimental group were exposed to 1 μg/mL 4F-MDMB-BUTINACA for 24 h, they were transferred to clean water and cleaned three times, then pretreated for instrumental analysis. The zebrafish in blank control group were not exposed to 4F-MDMB-BUTINACA. Mass spectrometry and structural analysis of 4F-MDMB-BUTINACA and its metabolites were conducted by liquid chromatography-high resolution mass spectrometry and Mass Frontier software. Results A total of twenty-six metabolites of 4F-MDMB-BUTINACA were identified in zebrafish, including eighteen phase Ⅰ metabolites and eight phase Ⅱ metabolites. The main metabolic pathways of phase Ⅰ metabolites of 4F-MDMB-BUTINACA in zebrafish were ester hydrolysis, N-dealkylation, oxidative defluorination and hydroxylation, while the main metabolic pathway of phase Ⅱ metabolites was glucuronidation. Conclusion Metabolite Md24 （ester hydrolysis） and Md25 （ester hydrolysis combined with dehydrogenation） would be recommended to be potentially good biomarkers for abuse of 4F-MDMB-BUTINACA.

Analysis of the metabolic characteristics in rat plasma after total body irradiation / 中华放射医学与防护杂志

Objective:To investigate the metabolite changes in rat plasma after total body irradiation (TBI) and to explore dose classification based on radiation sensitive metabolites.Methods:The differential metabolites induced by radiation were screened and verified by metabolomics. In the discovery stage, 50 SD rats were irradiated with 0, 1, 2, 3, 5 and 8 Gy of 60Co γ-rays. In the verification stage, 25 rats were irradiated with 0, 0.5, 2.5, 4 and 6 Gy. Peripheral blood samples were collected 4 h after irradiation, and plasma was separated. Radiation-induced differential metabolites were identified and their concentrations were determined. Receiver operating characteristic (ROC) curve of the differential metabolites was used to classify dose range. Results:In the discovery stage, 8 radiation-induced differential metabolites in rat plasma were identified and four of them (cytosine, L-hexylcarnitine, Linoelaidylcarnitine and L-palmitylcarnitine) were upregulated, which was confirmed in the verification stage. The area under the curve (AUC) for the specific dose was >0.75. After combining these four metabolites, the AUC value to classify the radiation dose of 0 Gy versus >0 Gy, <2 Gy versus ≥2 Gy, <5 Gy versus ≥5 Gy were 0.96, 1 and 0.94, respectively.Conclusions:The metabolites in rat plasma changed significantly at 4 h after TBI, where 8 differential metabolites were identified. Cytosine, L-hexylcarnitine, linoelaidylcarnitine and L-palmiylcarnitine were stably over-expressed in the plasma after irradiation. The combination of these four compounds had high classification accuracy and thus may applicable as radiation sensitive biomarkers for dose classification.

Screening of potential biomarkers and metabolic pathways of knee osteoarthritis in rats / 中华地方病学杂志

Objective:To screen differential metabolites and metabolic pathways which are related to knee osteoarthritis in rats, and to provide clues for further study of biomarkers of osteoarthritis.Methods:Sixty SPF male SD rats were divided into model and control groups according to their body weight (300 - 350 g) by random number table method, with 30 rats in each group. The experimental periods were 4, 8 and 12 weeks, each period included 10 rats in each group. The left knee joints of rats in the model group were operated by the modified Hulth method. After 5 d, rats in the model group were driven to move for 30 min every day. All rats were fed ordinary solid fodder and drank tap water. At the end of the experimental period, the knee joints and blood samples of rats were collected. Hematoxylin-eosin (HE) staining was used to observe histopathological changes of knee joint, ultra-performance liquid chromatography quadrupole time-flight mass spectrometry (UPLC-Q-TOF-MS) was used to detect small molecule metabolites in serum, and multivariate statistical analysis and database comparison were used to screen the differential metabolites and related metabolic pathways associated with osteoarthritis.Results:In the model group, the articular cartilage of knee was thinned, the surface was roughness, defect or peeling, chondrocytes were degeneration, necrosis and deletion, and the lesions were aggravated with prolonged experimental period. A total of 11 serum differential metabolites related to osteoarthritis were screened out, including selenocysteine, 6-hydroxymelatonin, γ-glutamylcysteine, arachidonic acid, sphinganine, leukotriene A4, leukotriene B4, 11,12-epoxy-eicostrienoic acid (11,12-EpETrE), lysopc, ceramide and N-arachidonoyl glycine. Among them, 9 metabolites were screened out at 4 weeks, compared with the control group, 5 metabolites were increased and 4 metabolites were decreased in the model group; 8 metabolites were screened out at 8 weeks, compared with the control group, 2 metabolites were increased and 6 metabolites were decreased in the model group; 8 metabolites were screened out at 12 weeks, compared with the control group, 5 metabolites were increased and 3 metabolites were decreased in the model group. The most relevant metabolic pathways related to osteoarthritis were sphingolipid metabolic pathway and arachidonic acid metabolic pathway. Sphinganine and ceramide were belonged to sphingolipid metabolic pathway, whereas arachidonic acid, leukotriene A4 and leukotriene B4 were belonged to arachidonic acid metabolic pathway.Conclusions:The progression of osteoarthritis can affect the composition and level of serum metabolite profile. Eleven serum differential metabolites are involved in sphingolipid metabolic pathway and arachidonic acid metabolic pathway, which are related to the occurrence and development of osteoarthritis.