Intact pZ189 DNA was replicated in monkey kidney vero cells which had been pretreated with Nmethyl-N’-nitro-N-nitrosoguanidine (MNNG) . The mutants were selected in E. coli MBM7070 and the mutation frequencies involv...Intact pZ189 DNA was replicated in monkey kidney vero cells which had been pretreated with Nmethyl-N’-nitro-N-nitrosoguanidine (MNNG) . The mutants were selected in E. coli MBM7070 and the mutation frequencies involving mutants with unchanged electrophoretic mobilrty of their plasmid DNA were scored. When compared to the spontaneous mutation frequency. the mutation frequencies were increased by 5.8 and 2.9-fold in cells pretreated with 0. 2 and 2μmol/L MNNG, respectively. The supF genes of these mutants were sequenced. and it was found that the types of base substitution and the sites of frameshifts differed from findings in studies of spontaneous and targeted mutagenesis. The results suggest that nontargeted mutagenesis occurs in mammalian cells and may have a sequence specificity.展开更多
The combination of hydrogen/deuterium(H/D)formaldehyde-based isotopic methyl labeling with solid-phase extraction and high-performance liquid chromatography–high resolution mass spectrometry(HPLC-HRMS)is a powerful a...The combination of hydrogen/deuterium(H/D)formaldehyde-based isotopic methyl labeling with solid-phase extraction and high-performance liquid chromatography–high resolution mass spectrometry(HPLC-HRMS)is a powerful analytical solution for nontargeted analysis of trace-level amino-containing chemicals in water samples.Given the huge amount of chemical information generated in HPLC-HRMS analysis,identifying all possible H/Dlabeled amino chemicals presents a significant challenge in data processing.To address this,we designed a streamlined data processing pipeline that can automatically extract H/D-labeled amino chemicals from the raw HPLC-HRMS data with high accuracy and efficiency.First,we developed a cross-correlation algorithm to correct the retention time shift resulting from deuterium isotopic effects,which enables reliable pairing of H-and D-labeled peaks.Second,we implemented several bioinformatic solutions to remove false chemical features generated by in-source fragmentation,salt adduction,and natural13C isotopes.Third,we used a data mining strategy to construct the AMINES library that consists of over 38,000 structure-disjointed primary and secondary amines to facilitate putative compound annotation.Finally,we integrated these modules into a freely available R program,HDPairFinder.R.The rationale of each module was justified and its performance tested using experimental H/D-labeled chemical standards and authentic water samples.We further demonstrated the application of HDPairFinder to effectively extract N-containing contaminants,thus enabling the monitoring of changes of primary and secondary N-compounds in authentic water samples.HDPairFinder is a reliable bioinformatic tool for rapid processing of H/D isotopic methyl labeling-based nontargeted analysis of water samples,and will facilitate a better understanding of N-containing chemical compounds in water.展开更多
BACKGROUND The relationship between metabolic syndrome(MetS)and gastric cancer(GC),which is a common metabolic disease,has attracted much attention.However,the specific metabolic characteristics of MetS in elderly pat...BACKGROUND The relationship between metabolic syndrome(MetS)and gastric cancer(GC),which is a common metabolic disease,has attracted much attention.However,the specific metabolic characteristics of MetS in elderly patients with GC remain unclear.AIM To investigate the differentially abundant metabolites and metabolic pathways between preoperative frailty and MetS in elderly patients with GC based on nontargeted metabolomics techniques.METHODS In this study,125 patients with nonfrail nonmeal GC were selected as the control group,and 50 patients with GC in the frail group were selected as the frail group.Sixty-five patients with GC combined with MetS alone were included in the MetS group,and 50 patients with GC combined with MetS were included in the MetS group.Nontargeted metabolomics techniques were used to measure plasma metabolite levels by ultrahigh-performance liquid chromatography-mass spectrometry.Multivariate statistical analysis was performed by principal component analysis,orthogonal partial least squares,pattern recognition analysis,cluster analysis,and metabolic pathway annotation.RESULTS A total of 125 different metabolites,including amino acids,glycerophospholipids,sphingolipids,fatty acids,sugars,nucleosides and nucleotides,and acidic compounds,were identified via nontargeted metabolomics techniques.Compared with those in the control group,there were 41,32,and 52 different metabolites in the MetS group,the debilitated group,and the combined group,respectively.Lipid metabolites were significantly increased in the MetS group.In the weak group,amino acids and most glycerol phospholipid metabolites decreased significantly,and fatty acids and sphingosine increased significantly.The combined group was characterized by significantly increased levels of nucleotide metabolites and acidic compounds.The alanine,aspartic acid,and glutamate metabolic pathways were obviously enriched in the asthenic group,and the glycerol and phospholipid metabolic pathways were obviously enriched in the combined group.CONCLUSION Elderly GC patients with simple frailty,simple combined MetS,and frailty combined with MetS have different metabolic characteristics,among which amino acid and glycerophospholipid metabolite levels are significantly lower in frail elderly GC patients,and comprehensive supplementation of fat and protein should be considered.Many kinds of metabolites,such as amino acids,lipids,nucleotides,and acidic compounds,are abnormally abundant in patients with MetS combined with fthenia,which may be related to tumor-related metabolic disorders.展开更多
目的研究高原性高血压(altitude-related hypertension,ARH)患者和健康个体之间血浆代谢产物的差异,并分析ARH的潜在发病机制。方法选取2020年7月在进驻海拔4200 m处的健康平原成年男性官兵中按简单随机抽样法选取20名ARH患者为ARH组,3...目的研究高原性高血压(altitude-related hypertension,ARH)患者和健康个体之间血浆代谢产物的差异,并分析ARH的潜在发病机制。方法选取2020年7月在进驻海拔4200 m处的健康平原成年男性官兵中按简单随机抽样法选取20名ARH患者为ARH组,30名血压正常者为对照(Control)组。在测量血压、体质指数(body mass index,BMI)、血氧饱和度和心率后,采集研究对象空腹静脉血样本。基于超高效液相色谱-质谱联用(ultra-high performance liquid chromatography-tandem mass spectrometry,UHPLC-MS/MS)技术,对研究对象血浆中的代谢产物进行筛选和鉴定。使用无监督主成分分析(principal component analysis,PCA)和有监督的正交偏最小二乘判别分析(orthogonal partial least squares discrimination analysis,OPLS-DA)模型对代谢产物进行指纹图谱分析,以辅助生物标志物的筛选,并对OPLS-DA模型进行质量考查和验证,以确保模型的稳定性和可靠性。使用独立样本t检验和差异倍数(fold change,FC)分析,绘制火山图,筛选有差异的血浆代谢物。使用京都基因与基因组百科全书(Kyoto Encyclopedia of Genes and Genomes,KEGG)富集通路分析对筛选出的差异性代谢物进行功能途径的富集分析和拓扑学分析。结果与Control组比较,ARH组的收缩压、舒张压和心率显著升高,动脉血氧饱和度显著降低(P<0.05)。PCA分析显示,正离子模式下,解释了81.96%的变异;负离子模式下,解释了79.25%的变异,表明2组之间存在显著的代谢差异。OPLS-DA模型分析显示,正离子模式下,PC1解释了77.36%的变异,PC2解释了12.25%的变异,R^(2)Y=0.96,Q^(2)Y=0.91;负离子模式下,PC1解释了84.15%的变异,PC2解释了17.24%的变异,R^(2)Y=0.99,Q^(2)Y=0.86。组间差异均超过75%,组内差异均小于20%。正离子模式下,R2拟合直线的Y轴截距为0.58,Q^(2)拟合直线的Y轴截距为-0.48;负离子模式下,R2拟合直线的Y轴截距为0.93,Q^(2)拟合直线的Y轴截距为-0.41。共筛选出32种具有显著差异的代谢产物,包括氨基酸、核苷、脂肪酸和有机生物碱。KEGG分析显示,10条代谢通路中有4条为氨基酸代谢通路,氨酰-tRNA生物合成途径富集的代谢产物最多。结论基于UHPLC-MS/MS技术,运用非靶向代谢组学筛选出的32种具有明显差异的代谢产物可能作为ARH的特征性生物标志物,而氨酰-tRNA生物合成途径可能与ARH的发病机制相关。展开更多
A broad range of organic compounds are known to exist in drinking water sources and serve as precursors of disinfection byproducts（DBPs）.Epidemiological findings of an association of increased risk of bladder cancer...A broad range of organic compounds are known to exist in drinking water sources and serve as precursors of disinfection byproducts（DBPs）.Epidemiological findings of an association of increased risk of bladder cancer with the consumption of chlorinated water has resulted in health concerns about DBPs.Peptides are thought to be an important category of DBP precursors in water.However,little is known about the actual presence of peptides and their DBPs in drinking water because of their high sample complexity and low concentrations.To address this challenge and identify peptides and non-chlorinated/chlorinated peptide DBPs from large sets of organic compounds in water,we developed a novel high throughput analysis strategy,which integrated multiple solid phase extraction（SPE）,high performance liquid chromatography（HPLC）separation,and non-target identification using precursor ion exclusion（PIE）high resolution mass spectrometry（MS）.After MS analysis,structures of candidate compounds,particularly peptides,were obtained by searching against the Human Metabolome Database（HMDB）.Using this strategy,we successfully detected 625 peptides（out of 17,205 putative compounds）and 617 peptides（out of 13,297）respectively in source and finished water samples.The source and finished water samples had 501 peptides and amino acids in common.The remaining 116 peptides and amino acids were unique to the finished water.From a subset of 30 putative compounds for which standards were available,25 were confirmed using HPLC-MS analysis.By analyzing the peptides identified in source and finished water,we successfully confirmed three disinfection reaction pathways that convert peptides into toxic DBPs.展开更多
It is thought that there are many unregulated anthropogenic chemicals in the environment.For risk assessment of chemicals, it is essential to estimate the predicted environmental concentrations. As an effort of identi...It is thought that there are many unregulated anthropogenic chemicals in the environment.For risk assessment of chemicals, it is essential to estimate the predicted environmental concentrations. As an effort of identifying residual organic contaminants in air and water in Korea, nontarget screening using two-dimensional gas chromatography time-of-flight mass spectrometry（GC × GC-TOFMS） was conducted at 10 sites using polyurethane foam passive air sampler and at 6 sites using polydimethyl siloxane（PDMS） passive water sampler in three different seasons in 2014. More than 600 chemical peaks were identified satisfying the identification criteria in air and water samples, respectively, providing a list for further investigation. Chemical substances with reported national emission rates in2014（n = 149） were also screened for potential existence in the environment using a level Ⅱ fugacity model. Most of chemical substances classified as not detectable were not identified with detection frequency greater than 20% by nontarget screening, indicating that a simple equilibrium model has a strong potential to be used to exclude chemicals that are not likely to remain in the environment after emissions from targeted monitoring.展开更多
Among the expressed sequence tags (ESTs) which were isolated from genetically instable monkey kidney vero cells induced by N-methyI-N’-nitro-N-nitrosoguanidine (MNNG), a fragment 9 was isolated. When the expression o...Among the expressed sequence tags (ESTs) which were isolated from genetically instable monkey kidney vero cells induced by N-methyI-N’-nitro-N-nitrosoguanidine (MNNG), a fragment 9 was isolated. When the expression of fragment 9 was blocked through transcripting antisense RNA, MNNGinduced nontargeted mutation frequency enhanced significantly compared with control ( P 【 0.05). The result showed that the relevant gene of fragment 9 might participate in maintaining cellular genetic stability and be involved in inhibition of nontargeted mutagenesis.展开更多
Fungicides are used to control pathogenic fungi of crop species,but they have also been shown to alter behavioral,life history and fitness related traits of nontarget insects.Here,we tested the fungicide effects on fe...Fungicides are used to control pathogenic fungi of crop species,but they have also been shown to alter behavioral,life history and fitness related traits of nontarget insects.Here,we tested the fungicide effects on feeding behavior,survival and physiology of the nontarget pest insect,the Colorado potato beetle(CPB)(Leptinotarsa decemlineata).Feeding behavior was studied by a choice test of adult beetles,which were allowed to choose between a control and a fungicide(fluazinam)treated potato leaf.Larval survival was recorded after 24 and 72 h exposure to control and fungicide-treated leaves with 2 different concentrations.The adults did not show fungicide avoidance behavior.Similarly,survival of the larvae was not affected by the exposure to fungicides.Finally,to understand the effects of fungicides at the physiological level(gene expression),we tested whether the larval exposure to fungicide alter the expression of 5 metabolic pathway and stress associated genes.Highest concentration and 72-h exposure caused upregulation of 1 cytochrome P450(CYP9Z14v2)and 1 insecticide resistance gene(Ldace1),whereas metabolic detoxification gene(Ugt1)was downregulated.At 24-h exposure,highest concentration caused downregulation of another common detoxification gene(Gs),while both exposure times to lowest concentration caused upregulation of the Hsp70 stress tolerance gene.Despite these overall effects,there was a considerable amount of variation among different families in the gene expression levels.Even though the behavioral effects of the fungicide treatments were minor,the expression level differences of the studied genes indicate changes on the metabolic detoxifications and stress-related pathways.展开更多
文摘Intact pZ189 DNA was replicated in monkey kidney vero cells which had been pretreated with Nmethyl-N’-nitro-N-nitrosoguanidine (MNNG) . The mutants were selected in E. coli MBM7070 and the mutation frequencies involving mutants with unchanged electrophoretic mobilrty of their plasmid DNA were scored. When compared to the spontaneous mutation frequency. the mutation frequencies were increased by 5.8 and 2.9-fold in cells pretreated with 0. 2 and 2μmol/L MNNG, respectively. The supF genes of these mutants were sequenced. and it was found that the types of base substitution and the sites of frameshifts differed from findings in studies of spontaneous and targeted mutagenesis. The results suggest that nontargeted mutagenesis occurs in mammalian cells and may have a sequence specificity.
基金supported by grants from the Natural Sciences and Engineering Research Council of Canada,Alberta Innovatesthe Canada Research Chairs Program。
文摘The combination of hydrogen/deuterium(H/D)formaldehyde-based isotopic methyl labeling with solid-phase extraction and high-performance liquid chromatography–high resolution mass spectrometry(HPLC-HRMS)is a powerful analytical solution for nontargeted analysis of trace-level amino-containing chemicals in water samples.Given the huge amount of chemical information generated in HPLC-HRMS analysis,identifying all possible H/Dlabeled amino chemicals presents a significant challenge in data processing.To address this,we designed a streamlined data processing pipeline that can automatically extract H/D-labeled amino chemicals from the raw HPLC-HRMS data with high accuracy and efficiency.First,we developed a cross-correlation algorithm to correct the retention time shift resulting from deuterium isotopic effects,which enables reliable pairing of H-and D-labeled peaks.Second,we implemented several bioinformatic solutions to remove false chemical features generated by in-source fragmentation,salt adduction,and natural13C isotopes.Third,we used a data mining strategy to construct the AMINES library that consists of over 38,000 structure-disjointed primary and secondary amines to facilitate putative compound annotation.Finally,we integrated these modules into a freely available R program,HDPairFinder.R.The rationale of each module was justified and its performance tested using experimental H/D-labeled chemical standards and authentic water samples.We further demonstrated the application of HDPairFinder to effectively extract N-containing contaminants,thus enabling the monitoring of changes of primary and secondary N-compounds in authentic water samples.HDPairFinder is a reliable bioinformatic tool for rapid processing of H/D isotopic methyl labeling-based nontargeted analysis of water samples,and will facilitate a better understanding of N-containing chemical compounds in water.
文摘BACKGROUND The relationship between metabolic syndrome(MetS)and gastric cancer(GC),which is a common metabolic disease,has attracted much attention.However,the specific metabolic characteristics of MetS in elderly patients with GC remain unclear.AIM To investigate the differentially abundant metabolites and metabolic pathways between preoperative frailty and MetS in elderly patients with GC based on nontargeted metabolomics techniques.METHODS In this study,125 patients with nonfrail nonmeal GC were selected as the control group,and 50 patients with GC in the frail group were selected as the frail group.Sixty-five patients with GC combined with MetS alone were included in the MetS group,and 50 patients with GC combined with MetS were included in the MetS group.Nontargeted metabolomics techniques were used to measure plasma metabolite levels by ultrahigh-performance liquid chromatography-mass spectrometry.Multivariate statistical analysis was performed by principal component analysis,orthogonal partial least squares,pattern recognition analysis,cluster analysis,and metabolic pathway annotation.RESULTS A total of 125 different metabolites,including amino acids,glycerophospholipids,sphingolipids,fatty acids,sugars,nucleosides and nucleotides,and acidic compounds,were identified via nontargeted metabolomics techniques.Compared with those in the control group,there were 41,32,and 52 different metabolites in the MetS group,the debilitated group,and the combined group,respectively.Lipid metabolites were significantly increased in the MetS group.In the weak group,amino acids and most glycerol phospholipid metabolites decreased significantly,and fatty acids and sphingosine increased significantly.The combined group was characterized by significantly increased levels of nucleotide metabolites and acidic compounds.The alanine,aspartic acid,and glutamate metabolic pathways were obviously enriched in the asthenic group,and the glycerol and phospholipid metabolic pathways were obviously enriched in the combined group.CONCLUSION Elderly GC patients with simple frailty,simple combined MetS,and frailty combined with MetS have different metabolic characteristics,among which amino acid and glycerophospholipid metabolite levels are significantly lower in frail elderly GC patients,and comprehensive supplementation of fat and protein should be considered.Many kinds of metabolites,such as amino acids,lipids,nucleotides,and acidic compounds,are abnormally abundant in patients with MetS combined with fthenia,which may be related to tumor-related metabolic disorders.
文摘目的研究高原性高血压(altitude-related hypertension,ARH)患者和健康个体之间血浆代谢产物的差异,并分析ARH的潜在发病机制。方法选取2020年7月在进驻海拔4200 m处的健康平原成年男性官兵中按简单随机抽样法选取20名ARH患者为ARH组,30名血压正常者为对照(Control)组。在测量血压、体质指数(body mass index,BMI)、血氧饱和度和心率后,采集研究对象空腹静脉血样本。基于超高效液相色谱-质谱联用(ultra-high performance liquid chromatography-tandem mass spectrometry,UHPLC-MS/MS)技术,对研究对象血浆中的代谢产物进行筛选和鉴定。使用无监督主成分分析(principal component analysis,PCA)和有监督的正交偏最小二乘判别分析(orthogonal partial least squares discrimination analysis,OPLS-DA)模型对代谢产物进行指纹图谱分析,以辅助生物标志物的筛选,并对OPLS-DA模型进行质量考查和验证,以确保模型的稳定性和可靠性。使用独立样本t检验和差异倍数(fold change,FC)分析,绘制火山图,筛选有差异的血浆代谢物。使用京都基因与基因组百科全书(Kyoto Encyclopedia of Genes and Genomes,KEGG)富集通路分析对筛选出的差异性代谢物进行功能途径的富集分析和拓扑学分析。结果与Control组比较,ARH组的收缩压、舒张压和心率显著升高,动脉血氧饱和度显著降低(P<0.05)。PCA分析显示,正离子模式下,解释了81.96%的变异;负离子模式下,解释了79.25%的变异,表明2组之间存在显著的代谢差异。OPLS-DA模型分析显示,正离子模式下,PC1解释了77.36%的变异,PC2解释了12.25%的变异,R^(2)Y=0.96,Q^(2)Y=0.91;负离子模式下,PC1解释了84.15%的变异,PC2解释了17.24%的变异,R^(2)Y=0.99,Q^(2)Y=0.86。组间差异均超过75%,组内差异均小于20%。正离子模式下,R2拟合直线的Y轴截距为0.58,Q^(2)拟合直线的Y轴截距为-0.48;负离子模式下,R2拟合直线的Y轴截距为0.93,Q^(2)拟合直线的Y轴截距为-0.41。共筛选出32种具有显著差异的代谢产物,包括氨基酸、核苷、脂肪酸和有机生物碱。KEGG分析显示,10条代谢通路中有4条为氨基酸代谢通路,氨酰-tRNA生物合成途径富集的代谢产物最多。结论基于UHPLC-MS/MS技术,运用非靶向代谢组学筛选出的32种具有明显差异的代谢产物可能作为ARH的特征性生物标志物,而氨酰-tRNA生物合成途径可能与ARH的发病机制相关。
基金supported by grants from the Natural Sciences and Engineering Research Council of Canada,Alberta Health,and Alberta Innovates-Energy and Environment Solutions
文摘A broad range of organic compounds are known to exist in drinking water sources and serve as precursors of disinfection byproducts（DBPs）.Epidemiological findings of an association of increased risk of bladder cancer with the consumption of chlorinated water has resulted in health concerns about DBPs.Peptides are thought to be an important category of DBP precursors in water.However,little is known about the actual presence of peptides and their DBPs in drinking water because of their high sample complexity and low concentrations.To address this challenge and identify peptides and non-chlorinated/chlorinated peptide DBPs from large sets of organic compounds in water,we developed a novel high throughput analysis strategy,which integrated multiple solid phase extraction（SPE）,high performance liquid chromatography（HPLC）separation,and non-target identification using precursor ion exclusion（PIE）high resolution mass spectrometry（MS）.After MS analysis,structures of candidate compounds,particularly peptides,were obtained by searching against the Human Metabolome Database（HMDB）.Using this strategy,we successfully detected 625 peptides（out of 17,205 putative compounds）and 617 peptides（out of 13,297）respectively in source and finished water samples.The source and finished water samples had 501 peptides and amino acids in common.The remaining 116 peptides and amino acids were unique to the finished water.From a subset of 30 putative compounds for which standards were available,25 were confirmed using HPLC-MS analysis.By analyzing the peptides identified in source and finished water,we successfully confirmed three disinfection reaction pathways that convert peptides into toxic DBPs.
基金supported by the National Institute of Environmental Research(No.NIER-RP-2014-335)
文摘It is thought that there are many unregulated anthropogenic chemicals in the environment.For risk assessment of chemicals, it is essential to estimate the predicted environmental concentrations. As an effort of identifying residual organic contaminants in air and water in Korea, nontarget screening using two-dimensional gas chromatography time-of-flight mass spectrometry（GC × GC-TOFMS） was conducted at 10 sites using polyurethane foam passive air sampler and at 6 sites using polydimethyl siloxane（PDMS） passive water sampler in three different seasons in 2014. More than 600 chemical peaks were identified satisfying the identification criteria in air and water samples, respectively, providing a list for further investigation. Chemical substances with reported national emission rates in2014（n = 149） were also screened for potential existence in the environment using a level Ⅱ fugacity model. Most of chemical substances classified as not detectable were not identified with detection frequency greater than 20% by nontarget screening, indicating that a simple equilibrium model has a strong potential to be used to exclude chemicals that are not likely to remain in the environment after emissions from targeted monitoring.
文摘Among the expressed sequence tags (ESTs) which were isolated from genetically instable monkey kidney vero cells induced by N-methyI-N’-nitro-N-nitrosoguanidine (MNNG), a fragment 9 was isolated. When the expression of fragment 9 was blocked through transcripting antisense RNA, MNNGinduced nontargeted mutation frequency enhanced significantly compared with control ( P 【 0.05). The result showed that the relevant gene of fragment 9 might participate in maintaining cellular genetic stability and be involved in inhibition of nontargeted mutagenesis.
基金supported by the Finish Cultural foundation and the Academy of Finland(Grant NO.308302 to LL and 322980 to MK).
文摘Fungicides are used to control pathogenic fungi of crop species,but they have also been shown to alter behavioral,life history and fitness related traits of nontarget insects.Here,we tested the fungicide effects on feeding behavior,survival and physiology of the nontarget pest insect,the Colorado potato beetle(CPB)(Leptinotarsa decemlineata).Feeding behavior was studied by a choice test of adult beetles,which were allowed to choose between a control and a fungicide(fluazinam)treated potato leaf.Larval survival was recorded after 24 and 72 h exposure to control and fungicide-treated leaves with 2 different concentrations.The adults did not show fungicide avoidance behavior.Similarly,survival of the larvae was not affected by the exposure to fungicides.Finally,to understand the effects of fungicides at the physiological level(gene expression),we tested whether the larval exposure to fungicide alter the expression of 5 metabolic pathway and stress associated genes.Highest concentration and 72-h exposure caused upregulation of 1 cytochrome P450(CYP9Z14v2)and 1 insecticide resistance gene(Ldace1),whereas metabolic detoxification gene(Ugt1)was downregulated.At 24-h exposure,highest concentration caused downregulation of another common detoxification gene(Gs),while both exposure times to lowest concentration caused upregulation of the Hsp70 stress tolerance gene.Despite these overall effects,there was a considerable amount of variation among different families in the gene expression levels.Even though the behavioral effects of the fungicide treatments were minor,the expression level differences of the studied genes indicate changes on the metabolic detoxifications and stress-related pathways.
