Increasing evidence has shown that abnormal metabolic phenotypes in body fluids reflect the pathogenesis and pathophysiology of Parkinson’s disease (PD). These body fluids include urine; however, the relationship between, specifically, urinary metabolic phenotypes and PD is not fully understood. In this study, urinary metabolites from a total of 401 clinical urine samples collected from 106 idiopathic PD patients and 104 normal control subjects were profiled by using high-performance liquid chromatography coupled to high-resolution mass spectrometry. Our study revealed significant correlation between clinical phenotype and urinary metabolite profile. Metabolic profiles of idiopathic PD patients differed significantly and consistently from normal controls, with related metabolic pathway variations observed in steroidogenesis, fatty acid beta-oxidation, histidine metabolism, phenylalanine metabolism, tryptophan metabolism, nucleotide metabolism, and tyrosine metabolism. In the fruit fly Drosophila melanogaster, the alteration of the kynurenine pathway in tryptophan metabolism corresponded with pathogenic changes in the alpha-synuclein overexpressed Drosophila model of PD. The results suggest that LC–MS-based urinary metabolomic profiling can reveal the metabolite signatures and related variations in metabolic pathways that characterize PD. Consistent PD-related changes across species may provide the basis for understanding metabolic regulation of PD at the molecular level.Keywords: drosophila; idiopathic Parkinson’s disease; metabolic phenotype; metabolite signatures; urinary metabolomics;

•The magnetic Fe3O4@GO nanocomposite was prepared and used as the sorbent of MSPE.•MSPE-HPLC method was developed for the analysis of flavors and fragrances.•The method was applied to analysis of real samples with high sensitivity.•The method has potential application in the analysis of food additives.In this article, a method for the analysis of flavors and fragrances including ethyl vanillin, trans-cinnamic acid, methyl cinnamate, ethyl cinnamate, and benzyl cinnamate in foods by HPLC with Fe3O4@GO nanocomposite as the adsorbent of magnetic solid-phase extraction was developed. The magnetic Fe3O4@GO nanocomposite was prepared and characterized by transmission electron microscope, infrared spectroscopy, X-ray diffraction and vibrating sample magnetometer. The as-prepared Fe3O4@GO nanocomposite was used as adsorbent for extraction and preconcentration of flavors and fragrances in foods. The extraction and desorption conditions including amount of materials, extraction time and desorption solvents were investigated. The analytes were separated within 10 min and detected with LODs ranged from 0.02 to 0.04 μg/mL. The recovery obtained from the analysis of spiked juice sample was between 71.5% and 112.4% with RSDs lower than 4.14%. The developed method was successfully applied to the analysis of real samples including orange juice, chocolate and fruit sugar.

Core–shell structured magnetic covalent organic frameworks (Fe3O4@COFs) were synthesized via a facile approach at room temperature. Combining the advantages of high porosity, magnetic responsiveness, chemical stability and selectivity, Fe3O4@COFs can serve as an ideal absorbent for the highly efficient enrichment of peptides and the simultaneous exclusion of proteins from complex biological samples.

Since the 2010s, the authorities of Guangdong province and local governments have enhanced law enforcement and environmental regulations to abolish open burning, acid washing, and other uncontrolled e-waste recycling activities. In this study, ambient air and indoor dust near different kinds of e-waste recycling processes were collected in Guiyu and Qingyuan to investigate the pollution status of particles and polychlorinated dibenzo-p-dioxins and furans (PCDD/Fs) after stricter environmental regulations. PM2.5 and PCDD/Fs both showed significantly reduced levels in the two regions compared with the documented data. The congener distribution and principal component analysis results also confirmed the significant differences between the current PCDD/Fs pollution characterizations and the historical ones. The estimated total intake doses via air inhalation and dust ingestion of children in the recycling region of Guiyu ranged from 10 to 32 pg TEQ/(kg•day), which far exceeded the tolerable daily intake (TDI) limit (1–4 pg TEQ/(kg•day). Although the measurements showed a significant reduction of the release of PCDD/Fs, the pollution status was still considered severe in Guiyu town after stricter regulations were implemented.Download high-res image (167KB)Download full-size image

Polybrominated diphenyl ethers (PBDE), as one kind of the major persistent organic pollutants (POP), have potential adverse effects on human health. 2,2′,4,4′-tetrabromodiphenyl ether (BDE-47) has been identified as one of the dominant PBDE congener in environmental and human samples. In this paper, liquid chromatography–orbitrap mass spectrometry (LC-Orbitrap MS) was applied for the profiling analysis of small metabolites in mice serum and striatum. The metabolic characteristics of adult male C57BL/6 mice exposed to BDE-47 were investigated with a non-targeted metabolomics method. The partial least-squares discriminant analysis (PLS-DA) indicated that the metabolites profile was significantly changed due to the exposure. Fifty-seven differential metabolites that significantly altered in mice serum and striatum were identified through databases searching and authentic standards confirmation. The related metabolic pathways mainly involved purine metabolism, alanine, aspartate and glutamate metabolism, tryptophan metabolism, phenylalanine metabolism, and glutathione metabolism. On the basis of metabolic variations, molecular mechanisms including disturbance of dopaminergic system, neurotransmitters regulation, DNA methylation, and oxidative stress were proposed. The obtained results suggested that LC-Orbitrap MS-based metabolomics had the great potential for molecular understanding of metabolic regulation linked to the exposure of environment pollutants.

The rising prevalence of obesity and diabetes cannot be fully explained by known risk factors, such as unhealthy diet, a sedentary lifestyle, and family history. This review summarizes the available studies linking persistent organic pollutants (POPs) to obesity and diabetes and discusses plausible underlying mechanisms.Increasing evidence suggest that POPs may act as obesogens and diabetogens to promote the development of obesity and diabetes and induce metabolic dysfunction. POPs are synthesized chemicals and are used widely in our daily life. These chemicals are resistant to degradation in chemical or biological processes, which enable them to exist in the environment persistently and to be bio-accumulated in animal and human tissue through the food chain. Increasingly, epidemiologic studies suggest a positive association between POPs and risk of developing diabetes.Understanding the relationship of POPs with obesity and diabetes may shed light on preventive strategies for obesity and diabetes.

•Exploring lung DNA damage effects exposed to 1-NP with different doses.•Clarifying lung DNA damage repair gene expression induced by 1-NP.•Investigating oxidative stress and metabolic enzyme change caused by 1-NP.•DNA damage exerts an important role in 1-NP-induced lung genotoxicity.1-Nitropyrene (1-NP) is a mutagenic and carcinogenic pollutant very widespread in the environment. However, the relative investigations on genotoxicity, oxidative stress and metabolic enzymes in lungs of mammalian caused by 1-NP have not been fully established. In this study, the 1-NP solutions at 3 dosages (1.0 × 10−5, 4.0 × 10−5 and 1.6 × 10−4 mg/kg body weight) were respectively given to rats by the intratracheal instillation. The responses of 1-NP on DNA damage and repair, oxidative stress and metabolism biomarkers in rat lungs after exposure to 1-NP were measured. The results showed 1-NP at three dosages induced obvious DNA strand breaks, 8-OH-dG formation and DNA-protein cross-link in rat lungs compared with the control. Higher dosage 1-NP (4.0 × 10−5 and 1.6 × 10−4 mg/kg body weight) greatly activated DNA repair gene OGG1 and inhibited MTH1 and XRCC1 expressions, and they significantly elevated the levels of GADD153, heme oxygenase-1 and malondialdehyde and decreased SOD activity, accompanied by the increases of CYP450, CYP1A1, CYP1A2 and GST levels. These results suggested the genotoxicity of 1-NP might rely on 1-NP-caused DNA damage and its combined effects on the suppression of DNA repair and the enhancement of oxidative stress and metabolic enzyme activity.Download high-res image (115KB)Download full-size image

•A total of 627 urine samples collected from 209 pregnant women were analyzed.•Seven out of 11 target contaminants were detected at > 50% frequency.•There was significant correlation between BP-1 and BP-3 and between MeP and PrP.•Levels of BP-1 and BP-3 were significantly higher in the first trimester than later stages.Humans are potentially exposed to many environmental pollutants, many of which may cause adverse health effects, especially to pregnant women and their fetuses. In this study, 11 environmental pollutants from three different chemical classes, including benzophenones, parabens and triclosan were measured in 627 urine samples collected from 209 pregnant women to evaluate exposure and trends as a function of pregnancy stage. Methylparaben (MeP), ethylparaben, propylparaben (PrP), butylparaben, 2,4-dihydroxybenzophenone (BP-1), 2-hydroxy-4-methoxybenzophenone (BP-3) and 4-hydroxybenzophenone were detected in > 50% samples. The concentrations of BP-1 and BP-3 (Spearman's r = 0.57, p < 0.01) and those of MeP and PrP (Spearman's r = 0.68, p < 0.01) were found to be correlated. The urinary concentrations of BP-1 and BP-3 in the first trimester were significantly higher than those in the second or third trimester (Mann-Whitney U test, p < 0.05). These findings provide valuable information for improving the prediction of maternal exposure to these emerging pollutants and for assessing their potential health risks to the mother as well as the offspring.Download high-res image (215KB)Download full-size image

•Review the recent applications of MS-based strategies in cooking oil assessment.•Introduce technical and instrumental improvements of MS for oil composition analysis.•Highlight computational approaches for oil aging, traceability and authentication.•Discuss technical challenges and future prospects of current MS-based strategies.Cooking oil, composed of great variety of chemical constitutes, has been increasingly prevailing in public diet due to its functional and nutritional benefits to consumers. The long-term storage, deep frying process and repeated use of cooking oil will release numerous deterioration products, such as aldehydes, ketones, epoxides, polymerides and cyclic aromatic hydrogen compounds, which influences the quality of cooking oil and has emerged safety concerns on used cooking oil. Analyzing these components with complex molecular structures and dynamic concentration ranges in oil samples poses a big challenge to researchers. In this article, we review the recent developments of mass spectrometry as a powerful tool for quality assessment of cooking oil and highlight its increasing applications in authentication, aging and marker-detection of used cooking oil. Additionally, the current technical challenges and future prospects associated with these methodologies are provided.

•A UHPLC-TQMS method was developed to analyze six BTRs and five BTHs in urine samples.•Precision, accuracy, recovery of the method were validated and found satisfactory.•The method was applied to analyze the targeted compounds in 22 urine specimens.•1-hydroxy-benzotriazole was detected in urine samples for the first time.Benzotriazole (BTR) and benzothiazole (BTH) derivatives are extensively applied in industrial processes and consumer products, and are thus frequently detected in the environmental matrices. Due to their potential estrogenic effects reported in animal studies, the assessment of human exposure to BTRs and BTHs is important. In this study, a method was developed and validated to determine six BTRs and five BTHs in human urine by using ultra-high performance liquid chromatography coupled with a triple quadrupole mass spectrometry (UHPLC-TQMS) in positive electrospray ionization mode. After de-conjugation by β-glucuronidase, urine samples were liquid-liquid extracted for the measurement of total concentrations of BTRs and BTHs. The linearity, detection limit, precision, recovery, accuracy and matrix effect were evaluated. The limits of detection were less than 0.5 ng/mL. The validated method demonstrated good precision (RSD% < 15%), linearity (R2 > 0.99), recovery (>80%) and accuracy (80–100%). The method was successfully applied for the analysis of 22 human urine samples. Four out of eleven targeted compounds were detected in more than half of participants at ng/mL levels.

Ambient fine particulate matter (PM2.5) is a complex mixture associated with lung cancer risk. PM2.5-bound nitro-polycyclic aromatic hydrocarbons (NPAHs) have been demonstrated to possess mutagenicity and carcinogenicity. Previous studies showed that PM2.5 induced DNA damage, whereas there is little knowledge of whether 9-nitroanthracene (9-NA), a typical compound of NPAHs in PM2.5, causes DNA damage. Also, the regulating mechanisms of PM2.5 and 9-NA in DNA damage and repair are not yet fully established. Here we sought to investigate the molecular mechanisms of DNA damage and repair in the lungs of male Wistar rats exposed to PM2.5 (1.5 mg per kg body weight) or three different dosages of 9-NA. And then DNA strand breaks, 8-OH-dG formation, DNA–protein crosslink and DNA repair gene expressions in rat lungs were analyzed. In addition, alteration in oxidative stress factors and metabolic enzymes were detected. The results showed that (1) PM2.5 and higher dosage 9-NA (4.0 × 10−5 and 1.2 × 10−4 mg per kg body weight) significantly caused lung DNA damage, accompanied by increasing OGG1 expression while inhibiting MTH1 and XRCC1 expression, elevating the levels of GADD153, hemeoxygenase-1 and malondialdehyde, and promoting the activities of CYP450 isozymes and glutathione S-transferase. (2) 1.3 × 10−5 mg kg−1 9-NA exposure couldn't cause DNA damage and oxidative stress. (3) At the approximately equivalent dose level, PM2.5-induced DNA damage effects were more obvious than 9-NA with positive correlation. It suggests that DNA damage caused by PM2.5 and 9-NA may be mediated partially through influencing the DNA repair capacity and enhancing oxidative stress and biotransformation, and this negative effect of 9-NA might be related to the PM2.5-induced lung genotoxicity.

Targeting protein–protein interactions (PPIs) offers tantalizing opportunities for therapeutic intervention for the treatment of human diseases. Modulating PPI interfaces with organic small molecules has been found to be exceptionally challenging, and few candidates have been successfully developed into clinical drugs. Meanwhile, the striking array of distinctive properties exhibited by metal compounds renders them attractive scaffolds for the development of bioactive leads. Here, we report the identification of iridium(III) compounds as inhibitors of the H-Ras/Raf-1 PPI. The lead iridium(III) compound 1 exhibited potent inhibitory activity against the H-Ras/Raf-1 interaction and its signaling pathway in vitro and in vivo, and also directly engaged both H-Ras and Raf-1-RBD in cell lysates. Moreover, 1 repressed tumor growth in a mouse renal xenograft tumor model. Intriguingly, the Δ-enantiomer of 1 showed superior potency in the biological assays compared to Λ-1 or racemic 1. These compounds could potentially be used as starting scaffolds for the development of more potent Ras/Raf PPI inhibitors for the treatment of kidney cancer or other proliferative diseases.

•Established a UHPLC-MS/MS method for amino acids and their derivatives analysis.•Traced the metabolic fate of 13C-labeled amino acids using 13C-glutamine as tracer.•Monitored alterations of amino acids and their derivatives in colon cancer cells.Rapid and simple quantitative analysis of intracellular metabolites is a critical tool for monitoring the alteration of biologically significant metabolites in cell lines or in vivo. We established an ultra-high performance liquid chromatography (UHPLC) method, equipped with hydrophilic interaction liquid chromatography (HILIC) column coupled to tandem mass spectrometry (MS/MS) for the simultaneous determination of 19 amino acids and 2 related derivatives in human cell lines. Chromatographic separation was achieved within 20 min using a BEH amide column, with aqueous mobile phase containing 20 mM ammonium acetate and 20 mM ammonium hydroxide, and acetonitrile as the organic mobile phase. Amino acids were analyzed in positive ion multiple reaction monitoring (MRM) mode without the need of derivatization. Intra- and inter-day precisions were less than 13.7%. The method was successfully applied to simultaneously detect the 21 compounds in a human colon cancer cell line DLD1. Moreover, metabolite fate of glutamine-derived carbons into amino acids in DLD1 cells was successfully traced by using [U-13C5] glutamine as the isotope tracer. Metabolic consequences of glutaminolysis inhibition on amino acid metabolism were evaluated. Analysis of 12C- and U-13C-labeled amino acids revealed the significantly decreased incorporation of [U-13C5]-glutamine derived carbons into aspartate, alanine and ornithine, indicating impaired metabolic flux via the tricarboxylic acid cycle and the urea cycle.

N-doping carbon dots (N-CDs) were prepared by microwave-assisted pyrolysis of dl-malic acid and ethanolamine as precursors. The material served as an excellent matrix for the detection of the environmental pollutants hydroxy-polycyclic aromatic hydrocarbons (OH-PAHs) by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) in negative ion mode. The obtained N-CDs exhibited good UV absorption capacity and favorable solubility. The use of the N-CDs matrix exhibited low matrix background interference and was beneficial to improve the signal response due to the specific π-conjugated polyaromatic structure and the doping of nitrogen atoms. The developed method was found to have good reproducibility and sensitivity. The N-CDs as a new matrix also were employed for the detection of OH-PAHs in real PM2.5 samples. The mass concentrations of Σ-hydroxy-pyrene, Σ-dihydroxy-anthraquinone, and Σ-dihydroxy-benzo(a)pyrene on the collected PM2.5 samples ranged from 0.125 to 0.136 ng/m3, 0.039 to 0.052 ng/m3, and 0.053 to 0.072 ng/m3, respectively. This work extends the application field of N-CDs and provides a good candidate of matrix for MALDI-TOF MS detection of environmental pollutants.

Exposure to bisphenol A (BPA), an environmental contaminant, has been linked to metabolic disorders. However, there are no reports describing the effects of BPA on the profiling of cis-diol metabolites. It is challenge to detect these metabolites in biological samples because of their low abundance, high polarity and serious matrix interference. In this study, a chemical isotope-labeling method was applied to solve these problems. Acetone and deuterated acetone (acetone-d6) were used as chemical tags to label the rat urine samples, respectively. The light and heavy labeling products were recognized using the ShiftedIonsFinder software. The selected cis-diol metabolite signals were used to build a data set. The data set was applied to evaluate the changes in the urinary profiling of cis-diol-containing metabolites in rats with BPA exposure. The results showed that chromatographic separation and mass spectrometry detection of cis-diol metabolites were improved after acetone labeling. Using this method, the cis-diol metabolites were recognized easily from the urine samples. By comparing different dose administration on rats, the influence of BPA exposure on cis-diol metabolites was investigated. The analytes showing noticeable differences were identified. It was found that high-dose BPA exposure had strong effects on the cis-diol compound metabolism. The influences were mostly related to the metabolism of galactose and nucleoside and its analogues. The disturbance of the galactose metabolism by BPA is reported for the first time, to the best of our knowledge. This may have some implications for exploring the toxic effects of BPA exposure.

Zeolite imidazole framework (ZIF) nanocrystals serve as sorbents and matrices for negative ion surface assisted laser desorption/ionization mass spectrometry. The unique properties of ZIFs make them suitable for analyzing a series of environmental pollutants with high signal intensity and a clean background.

•Methods for the analysis of intracellular metabolites of E. coli are reviewed.•Interference and variations during cell sampling are discussed.•Sample loss during quenching is discussed and the possible solution is suggested.•Different MS-based analytical methods are reviewed.Understanding the mechanism of metabolic network response to extracellular changes has become one of the primary objectives of systems biology. Quantitative metabolic network profiling of model organisms such as E. coli is the principal approach. Well-designed analytical protocols for the determination of intracellular metabolites may provide a crucial premise to quantitative profiling of cellular metabolism. This article reviews and evaluates the existing methods for determining intracellular metabolites of E. coli in terms of sample preparation and mass spectrometry (MS) analysis. Critical issues including the variation of metabolic status and interference of external pools during cell sampling, sample loss during quenching procedures, degradation of metabolites and efficiency of extraction, and matrix effects in separation and quantification are discussed. Moreover, suggestions are provided to solve the analytical problems based on the results from the recent studies in order to establish a precise and reliable analytical platform for the intracellular metabolites.

•An UHPLC-MS/MS-based metabolomics was established to elucidate altered metabolism.•Metabolic perturbations in EIF5A2-transfected LO2 cells were figured out.•Levels of lactate and ATP in EIF5A2-transfected LO2 cells were dramatically altered.•Patterns of amino acid imbalance in EIF5A2-transfected LO2 cells were observed.Elucidation of altered metabolic pathways by using metabolomics may open new avenues for basic research on disease mechanisms and facilitate the development of novel therapeutic strategies. Here, we report the development of ultrahigh performance liquid chromatography-tandem mass spectrometry-based metabolomics platform with capability of measuring both cationic and anionic intermediates in cellular metabolism. The platform was established based on the hydrophobic ion-pairing interaction chromatography coupled with tandem mass spectrometry in multiple reaction monitoring (MRM) mode. The MRM transitions were created and optimized via energy-resolved collision-induced dissociation experiments, serving as an essential reference point for the quantification and identification. For chromatographic separation, application of hydrophobic ion-pairing interaction led to dramatic enhancement on retention of water-soluble metabolites and provision of good peak shapes. Two volatile ion-pairing reagents, namely heptafluorobutyric acid and tributylamine, were used with dedicated C18 columns as complementary separation systems coupled with the MRM analysis, allowing measurement of the metabolites of interest at nanomolar levels. The developed platform was successfully applied to investigate the altered metabolism in hepatic cells with over-expression of an oncogene, thus can provide important information on the rewired metabolism.

•Localization of ginsenosides in root tissue of Panax ginseng.•Identification of ginsenosides by MALDI-TOF-MSI and MALDI-TOF-MS/MS.•Principal component analysis of data collected from specific area in tissue.•Rapid differentiation of Panax ginseng of three ages using MALDI imaging.The root of Panax ginseng C.A. Mey. (P. ginseng) is one of the most popular traditional Chinese medicines, with ginsenosides as its main bioactive components. Because different ginsenosides have varied pharmacological effects, extraction and separation of ginsenosides are usually required for the investigation of pharmacological effects of different ginsenosides. However, the contents of ginsenosides vary with the ages and tissues of P. ginseng root. In this research, an efficient method to explore the distribution of ginsenosides and differentiate P. ginseng roots with different ages was developed based on matrix assisted laser desorption/ionization time-of-flight mass spectrometry imaging (MALDI-TOF-MSI). After a simple sample preparation, there were 18 peaks corresponding to 31 ginsenosides with distinct localization in the mass range of m/z 700–1400 identified by MALDI-TOF-MSI and MALDI-TOF-MS/MS. All the three types of ginsenosides were successfully detected and visualized in images, which could be correlated with anatomical features. The P. ginseng at the ages of 2, 4 and 6 could be differentiated finely through the principal component analysis of data collected from the cork based on the ion images but not data from the whole tissue. The experimental result implies that the established method for the direct analysis of metabolites in plant tissues has high potential for the rapid identification of metabolites and analysis of their localizations in medicinal herbs. Furthermore, this technique also provides valuable information for the component-specific extraction and pharmacological research of herbs.

Bromodomain-containing protein 4 (BRD4) has recently emerged as an attractive epigenetic target for anticancer therapy. In this study, an iridium(III) complex is reported as the first metal-based, irreversible inhibitor of BRD4. Complex 1a is able to antagonize the BRD4-acetylated histone protein–protein interaction (PPI) in vitro, and to bind BRD4 and down-regulate c-myc oncogenic expression in cellulo. Chromatin immunoprecipitation (ChIP) analysis revealed that 1a could modulate the interaction between BRD4 and chromatin in melanoma cells, particular at the MYC promoter. Finally, the complex showed potent activity against melanoma xenografts in an in vivo mouse model. To our knowledge, this is the first report of a Group 9 metal complex inhibiting the PPI of a member of the bromodomain and extraterminal domain (BET) family. We envision that complex 1a may serve as a useful scaffold for the development of more potent epigenetic agents against cancers such as melanoma.

Zeolitic imidazolate framework-8 coated magnetic nanocomposites (Fe3O4@ZIF-8 MNCs) served as an absorbent and a matrix for negative-ion MALDI-TOF MS. The host–guest property and interference-free background made them an ideal dual platform for the sensitive analysis of small molecules.

One kind of surface protein imprinting method was developed by a more convenient, simpler and cheaper approach based on vinyl-functionalized magnetic nanofibers (NFs).

Ultrathin graphitic carbon nitride (g-C3N4) nanosheets served as a novel matrix for the detection of small molecules by negative ion matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) was described for the first time. In comparison with conventional organic matrices and graphene matrix, the use of g-C3N4 nanosheet matrix showed free matrix background interference and increased signal intensity in the analysis of amino acids, nucleobases, peptides, bisphenols (BPs), and nitropolycyclic aromatic hydrocarbons (nitro-PAHs). A systematic comparison of g-C3N4 nanosheets with positive and negative ion modes revealed that mass spectra produced by g-C3N4 nanosheets in negative ion mode were featured by singly deprotonated ion without matrix interference, which was rather different from the complicated alkali metal complexes in positive ion mode. Good salt tolerance and reproducibility allowed the determination of 1-nitropyrene (1-NP) in sewage, and its corresponding detection limit was lowered to 1 pmol. In addition, the ionization mechanism of the g-C3N4 nanosheets as matrix was also discussed. The work expands its application scope of g-C3N4 nanosheets and provides an alternative approach for small molecules.

•A simple, robust and low-cost derivatization method was reported for ribonucleoside determination for the first time.•Improvement of separation and enhancement of sensitivity were achieved by using the derivatization approach.•Isotope labeling method with acetone-d6 and multivariate statistical analysis facilitated ribonucleoside identification.•Application of the method enabled the positive identification of 56 ribonucleosides.Ribonucleosides are the end products of RNA metabolism. These metabolites, especially the modified ribonucleosides, have been extensively evaluated as cancer-related biomarkers. However, the determination of urinary ribonucleosides is still a challenge due to their low abundance, high polarity and serious matrix interferences in urine samples. In this study, a derivatization method based on a chemical reaction between ribonucleosides and acetone to form acetonides was developed for the determination of urinary ribonucleosides. The derivative products, acetonides, were detected by using liquid chromatography–tandem mass spectrometry (LC–MS/MS). The methodological evaluation was performed by quantifying four nucleosides for linear range, average recovery, precision, accuracy and stability. The validated procedures were applied to screen modified ribonucleosides in urine samples. Improvement of separation and enhancement of sensitivity were obtained in the analysis. To identify ribonucleosides, inexpensive isotope labeling acetone (acetone-d6) and label-free acetone were applied to form ordinary and deuterated acetonides, respectively. The two groups of samples were separated with orthogonal partial least squares (OPLS). The ordinary and deuterated pairs of acetonides were symmetrically distributed in the S-plot for easy and visual signal identification. After structural confirmation, a total of 56 ribonucleosides were detected, 52 of which were modified ribonucleosides. The application of derivatization, deuterium-labeling and multivariate statistical analysis offers a new option for selective detection of ribonucleosides in biological samples.

Combining free radical polymerization with click chemistry via a copper-mediated azide/alkyne cycloaddition (CuAAC) reaction in a “one-pot” process, a facile approach was developed for the preparation of a poly(3′-azido-3′-deoxythymidine-co-propargyl methacrylate-co-pentaerythritol triacrylate) (AZT-co-PMA-co-PETA) monolithic column. The resulting poly(AZT-co-PMA-co-PETA) monolith showed a relatively homogeneous monolithic structure, good permeability and mechanical stability. Different ratios of monomers and porogens were used for optimizing the properties of a monolithic column. A series of alkylbenzenes, amides, anilines, and benzoic acids were used to evaluate the chromatographic properties of the polymer monolith in terms of hydrophobic, hydrophilic and cation-exchange interactions, and the results showed that the poly(AZT-co-PMA-co-PETA) monolith exhibited more flexible adjustment in chromatographic selectivity than that of the parent poly(PMA-co-PETA) and AZT-modified poly(PMA-co-PETA) monoliths. Column efficiencies for toluene, DMF, and formamide with 35000–48000 theoretical plates per m could be obtained at a linear velocity of 0.17 mm s−1. The run-to-run, column-to-column, and batch-to-batch repeatabilities of the retention factors were less than 4.2%. In addition, the proposed monolith was also applied to efficient separation of sulfonamides, nucleobases and nucleosides, anesthetics and proteins for demonstrating its potential.

CuFe2O4 magnetic nanocrystal clusters (CuFe2O4 MNCs) were proposed as a new matrix for small molecule analysis by negative ion matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) for the first time. We demonstrated its advantages over conventional organic matrices in the detection of small molecules such as amino acids, peptides, nucleobases, fatty acids, and steroid hormones. A systematic comparison of CuFe2O4 MNCs with different ionization modes revealed that MS spectra obtained for the CuFe2O4 MNC matrix in the negative ion mode was only featured by deprotonated ion peaks with a free matrix background, which was different from the complicated alkali metal adducts produced in the positive ion mode. The developed method was found relatively tolerant to salt contamination and exhibited good reproducibility. A detection limit down to the subpicomolar level was achieved when testosterone was analyzed. In addition, by comparison of the MS spectra obtained from bare Fe3O4 and MFe2O4 MNC (M = Co, Ni, Cu, Zn) matrices, two main factors of MFe2O4 MNC matrices were revealed to play a vital role in assisting the negative ion desorption/ionization (D/I) process: doping transition metals into ferrite nanocrystals favoring laser absorption and energy transfer and a good match between the UV absorption of MFe2O4 MNCs and the excitation of nitrogen laser source facilitating LDI efficiency. This work creates a new branch of application for MFe2O4 MNCs and provides an alternative solution for small molecule analysis.

Type 2 diabetes (T2DM) is caused by a complex set of interactions between genetic modifications and life styles. This complexity creates challenges for a full understanding of the altered metabolic pathways that contribute to the development of T2DM, which needs a comprehensive metabolic analysis. Exercise training is a common therapeutic approach known to antagonize the metabolic consequences of T2DM. However, the metabolic phenotypes of exercise effected in T2DM have not been clearly characterized. Here, we present the effect of physical activity on biochemical changes in diabetic db/db mice. An untargeted metabolomics study based on liquid chromatography coupled with high resolution mass spectrometry was carried out to delineate the plasma metabolic signatures in conjunction with a multivariate statistical analysis. As a result, a total of 24 differential metabolites were identified, covering amino acids, organic acids and lipids. Three biomarkers, including lysine, creatine and uridine, were significantly reversed by exercise training in db/db diabetic mice groups compared to lean db/m+ groups. Of note, pantothenic acid and palmitoylcarnitine, which are involved in fatty acid β-oxidation (FAO), were promoted by exercise training in diabetic mice rather than in lean mice. These findings indicated that diabetic mice might be more susceptible to exercise for energy expenditure. Together, the results might demonstrate that exercise could mitigate insulin resistance in T2DM through improving FAO and that uridine in blood might be an important indicator to reflect insulin sensitivity promoted by exercise training in T2DM mice.

Exposure to ambient fine particulate matter (PM2.5) increases the risk of respiratory disease. Although previous mitochondrial research has provided new information about PM toxicity in the lung, the exact mechanism of PM2.5-mediated structural and functional damage of lung mitochondria remains unclear. In this study, changes in lung mitochondrial morphology, expression of mitochondrial fission/fusion markers, lipid peroxidation, and transport ATPase activity in SD rats exposed to ambient PM2.5 at different dosages were investigated. Also, the release of reactive oxygen species (ROS) via the respiratory burst in rat alveolar macrophages (AMs) exposed to PM2.5 was examined by luminol-dependent chemiluminescence (CL). The results showed that (1) PM2.5 deposited in the lung and induced pathological damage, particularly causing abnormal alterations of mitochondrial structure, including mitochondrial swelling and cristae disorder or even fragmentation in the presence of higher doses of PM2.5; (2) PM2.5 significantly affected the expression of specific mitochondrial fission/fusion markers (OPA1, Mfn1, Mfn2, Fis1, and Drp1) in rat lung; (3) PM2.5 inhibited Mn superoxide dismutase (MnSOD), Na+K+-ATPase, and Ca2+-ATPase activities and elevated malondialdehyde (MDA) content in rat lung mitochondria; and (4) PM2.5 induced rat AMs to produce ROS, which was inhibited by about 84.1% by diphenyleneiodonium chloride (DPI), an important ROS generation inhibitor. It is suggested that the pathological injury observed in rat lung exposed to PM2.5 is associated with mitochondrial fusion–fission dysfunction, ROS generation, mitochondrial lipid peroxidation, and cellular homeostasis imbalance. Damage to lung mitochondria may be one of the important mechanisms by which PM2.5 induces lung injury, contributing to respiratory diseases.

Hepatocellular carcinoma (HCC) is one of the most common cancers in human beings and not well treated. Due to its blurred biomarkers, lipidomics can be considered as a new diagnostic tool. In this study, mass spectrometry-based lipidomics analysis coupled with multivariate statistical analysis was applied to profile twelve pairs of human hepatocellular carcinoma tissues (HCT) and matched adjacent non-tumour tissues (ANT). As a result, perturbation of lipid biosynthesis was observed in HCT compared to ANT. Lipid species were profiled and 28 significant lipids were identified based on high resolution mass spectrometry with mass error less than 3 ppm, covering phosphatidylcholine (PC), phosphatidylethanolamine (PE), phosphatidylglycerol (PG), phosphatidylinositol (PI), sphingomyelin (SM) and triglyceride (TG). The high mass accuracy as well as the tandem MS (MS/MS) technique provides high confidence of lipid identification. Interestingly, decreased PGs and PIs were observed in this study, indicating that lipolysis of PGs and PIs might play a crucial role in HCC development. The results indicated that mass spectrometry-based lipidomics analysis could serve as an effective means to diagnose canceration.

In this work, we developed a novel quantitative analysis method for detecting nitropolycyclic aromatic hydrocarbons (nitro-PAHs) in PM2.5 samples with graphene as a matrix using MALDI-TOF MS. A good linearity was obtained for the typical four nitro-PAHs, namely 1-nitropyrene (1-NP), 2-nitrofluorene (2-NFL), 6-nitrochrysene (6-NC) and 9-nitroanthracene (9-NA) with the correlation coefficient better than 0.985 by means of 9-nitroanthracene-d9 as the internal standard. The average recoveries of this method were from 69.2% to 119.4%, and the inter-day precisions were less than 12.3% with intra-day precisions less than 20.7%. The limits of detection for 1-NP, 2-NFL, 6-NC and 9-NA were 0.74, 8.04, 2.67 and 2.31 ng μL−1, respectively. We have validated the practicality by the analysis of nitro-PAHs in PM2.5 samples collected during haze weather in Taiyuan of China. The mass concentrations of Σ-nitropyrene, Σ-nitrofluorene, Σ-nitrochrysene and Σ-nitroanthracene on the collected PM2.5 samples were detected at levels of 0.38 to 3.04 ng m−3, 0.21 to 0.43 ng m−3, 0.19 to 2.38 ng m−3, and 9.55 to 16.52 ng m−3, respectively. The present method proved to be a good candidate for simple, fast and eco-friendly determination of nitro-PAHs by using MALDI-TOF MS coupled with an internal standard method.

A gas chromatography/mass spectrometry (GC/MS)-based method was developed for simultaneous determination of triclosan (TCS) and its degradation products including 2,4-dichlorophenol (2,4-DCP), 2,8-dichlorodibenzo-p-dioxin (2,8-DCDD), and methyl triclosan (MTCS) in wastewater and sludge samples. The method provides satisfactory detection limit, accuracy, precision and recovery especially for samples with complicated matrix such as sewage sludge. Liquid-liquid extraction and accelerated solvent extraction (ASE) methods were applied for the extraction, and column chromatography was employed for the sample cleanup. Analysis was performed by GC/MS in the selected ion monitoring (SIM) mode. The method was successfully applied to wastewater and sludge samples from three different municipal wastewater treatment plants (WWTPs). Satisfactory mean recoveries were obtained as 91(±4)–106(±7) %, 82(±3)–87(±4) %, 86(±6)–87(±8) %, and 88(±4)–105(±3) % in wastewater and 88(±5)–96(±8) %, 84(±2)–87(±3) %, 84(±7)–89(±4) %, and 88(±3)–97(±5) % in sludge samples for TCS, 2,4-DCP, 2,8-DCDD, and MTCS, respectively. TCS degradation products were detected based on the type of the wastewater and sludge treatment. 2,8-DCDD was detected in the plant utilizing UV disinfection at the mean level of 20.3(±4.8) ng/L. 2,4-DCP was identified in chemically enhanced primary treatment (CEPT) applying chlorine disinfection at the mean level of 16.8(±4.5) ng/L). Besides, methyl triclosan (MTCS) was detected in the wastewater collected after biological treatment (10.7 ± 3.3 ng/L) as well as in sludge samples that have undergone aerobic digestion at the mean level of 129.3(±17.2) ng/g dry weight (dw).

•Peptidomics, a new omics technique, was applied for analysis of elderberry juice.•Solid phase extraction method development separated anthocyanins from peptides and improved mass spectrometry analysis.•Novel application of peptidomics identified an abundance of peptides in elderberry juice.•Mass spectrometry-based peptidomics was demonstrated valuable in food nutrient and safety study.Biologically active peptides play a role in plant signaling and defense. Elderberry juice is known to contain a variety of anthocyanin compounds, a sub-set of polyphenols, which are responsible for the deep purple color of the juice. In this paper, we describe a method utilizing solid phase extraction (SPE) to remove anthocyanins from peptides. Liquid chromatography coupled with tandem mass spectrometry was used to separate and identify the peptides. The results showed that the use of SPE was an effective method to separate peptides from anthocyanins and other background compounds including high polyphenol content in the juice samples. More than 1000 peptides present in elderberry juice were successfully identified.

Metabolic variations occur during normal pregnancy to provide the growing fetus with a supply of nutrients required for its development and to ensure the health of the woman during gestation. Mass spectrometry-based metabolomics was employed to study the metabolic phenotype variations in the maternal plasma that are induced by pregnancy in each of its three trimesters. Nontargeted metabolomics analysis showed that pregnancy significantly altered the profile of metabolites in maternal plasma. The levels of six metabolites were found to change significantly throughout pregnancy, with related metabolic pathway variations observed in biopterin metabolism, phospholipid metabolism, amino acid derivatives, and fatty acid oxidation. In particular, there was a pronounced elevation of dihydrobiopterin (BH2), a compound produced in the synthesis of dopa, dopamine, norepinephrine, and epinephrine, in the second trimester, whereas it was markedly decreased in the third trimester. The turnover of BH2 and tryptophan catabolites indicated that the fluctuations of neurotransmitters throughout pregnancy might reveal the metabolic adaption in the maternal body for the growth of the fetus. Furthermore, 11 lipid classes and 41 carnitine species were also determined and this showed variations in the presence of long-chain acylcarnitines and lysophospholipids in later pregnancy, suggesting changes of acylcarnitines and lysophospholipids to meet the energy demands in pregnant women. To our knowledge, this work is the first report of dynamic metabolic signatures and proposed related metabolic pathways in the maternal plasma for normal pregnancies and provided the basis for time-dependent metabolic trajectory against which disease-related disorders may be contrasted.

Dioxin exposure tends to accumulate in adipose tissue and alters metabolism in mammals. In this study, gas chromatography coupled with mass spectrometry (GC-MS) in conjunction with multivariate statistical analysis was applied to profile small molecular metabolites in adipose tissue of aryl hydrocarbon receptor (AhR)-high affinity wild-type C57BL/6J mice exposed to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). A partial least squares discriminant analysis model was also constructed to map the discrimination between TCDD dosages and the control group. As a result, a total of 16 differential metabolites were identified in the high-dose TCDD group compared to the control group, and 12 free fatty acids (FFAs) were highlighted among them. Both saturated and unsaturated FFA levels were significantly elevated in adipose tissues of TCDD-exposed mice. This promising tool for global characterization highlights FFAs which could be served as indicators for understanding the toxic responses to TCDD exposure in a dose-dependent manner. The data indicated that the use of GC-MS coupled with multivariate statistical analysis could provide new insight for fatty acid biosynthesis on AhR activation with TCDD exposure in wild-type mice.

Metabolism study was carried out on 12β-hydroxylveratroylzygadenine (VOG) that is a cevine-type alkaloid existing in Veratrum nigrum L. and a neurotoxic component. In order to better understand the potential mechanism of neurotoxicity of VOG, this study measured VOG-induced DNA damage in the cerebellum and cerebral cortex of mice after 7 days repetitive oral dose by using single-cell gel electrophoresis (Comet assay). High performance liquid chromatography-tandem mass spectrometry (LC–MS/MS) was developed and applied to separate and identify in vitro and in vivo metabolites of VOG for investing the possible relationship of metabolism and neurotoxicity. In vitro experiment was carried out using rat liver microsomes, while the in vivo study was conducted on rats. The obtained results indicated that VOG might cause DNA damage in cerebellum and cerebral cortex of mice in a dose-dependent manner. Hydrolysis of ester bond and O-demethylation were proposed to be the main in vivo metabolic pathways of VOG, while the major in vitro metabolic pathways were proposed as methyl oxidation to aldehyde, dehydrogenation, hydrolysis of ester bond, hydrolysis of ester bond together with acetylation, and methoxylation. O-Demethylation reaction was likely to be associated with reactive oxygen species production, leading to the DNA damage.A HPLC–MS/MS was applied to identify in vitro and in vivo metabolites of VOG for investigating their relationship with the neurotoxicity. O-Demethylation reaction was likely to be associated with the DNA damage in mouse brain.

The pollution characteristics of ambient fine particulate matter (PM2.5) containing polycyclic aromatic hydrocarbons (PAHs) and nitrated PAHs (NPAHs) in samples collected during a typical winter time period in Taiyuan of China were investigated. The obtained results revealed that the mean mass concentrations of PM2.5, ΣPAHs (sum of 16 PAHs) and ΣNPAHs (sum of 3 NPAHs) on PM2.5 were 161.4 μg/m3, 119.8 ng/m3 and 0.446 ng/m3, respectively. Diagnostic ratios of PAHs and NPAHs implied that coal consumption might be the main source of the PM2.5 pollution. The measured PM2.5 mass concentrations, BaP equivalent toxicity (28.632 ng/m3) and individual carcinogenicity index (3.14 × 10−5) were much higher than those of the recommended safety standards.The main source of serious PM2.5 pollution in a typical Taiyuan's winter time period was coal combustion. PAHs and NPAHs on the PM2.5 may have a significant impact on human health and cancer risk.

•Ultrahigh resolution mass spectrometry was performed to profile the metabolites.•Metabolic alterations in cerebellum of mice were figured out.•The levels of amino acids and fatty acids were enhanced in Alzheimer's disease.•The reduced amino acids and fatty acids were observed in mice exposed to TCDD.•Tissue-specific metabolomics revealed that cerebellar metabolism could be disturbed.In the previous reports about cognitive dysfunction, cerebellum was thought to be a less affected tissue by genetic or environmental alterations in comparison to other tissues in the brain including hippocampus under the same conditions. In this work, we investigated two types of metabolomic alterations inside the cerebellum tissue. The first one addressed the differences in the metabolomics profiles between Transgenic (Tg) CRND8 of Alzheimer's disease mice and non-transgenic (non-Tg) littermates. The second one addressed the metabolic differences between wild type mice exposed to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) and wild type mice which are not exposed to this toxic compound. For these two investigations, ultrahigh resolution Fourier transform ion cyclotron resonance mass spectrometry (FT-ICR/MS) was implemented. As a result, the significant changes of each comparison were tentatively annotated by the high mass accuracy generated from the measurements in the negative ion mode. The biosynthesis of amino acids was also enhanced pronouncedly, and perturbation of purine metabolism was also observed in Tg mice compared to non-Tg littermates. In another animal model, the reduced levels of amino acids were found whereas the intermediate levels in purine metabolism and fatty acids including fatty acid conjugated metabolites were elevated in cerebellar tissues of mice exposed to TCDD compared to control group. Collectively, it was demonstrated that FT-ICR/MS was a powerful tool for interpretation of the elemental compositions of the peaks, revealing that the metabolic perturbations in cerebellar tissues of mice were induced by either genetic manipulation or environmental factor. Therefore, the non-targeted approach, alternatively, provides various metabolic phenotypes for the systems-level mirror of the complex etiology of neurotoxicity in the cerebellum.

•An ultrasensitive immunosensor for H1N1 influenza virus was designed.•A new technology using Ag–S covalent binding was applied for antibody labeling.•Signal amplification is based on fluorescent produced by Ag+ catalyzing OPDA.•The proposed sensor maybe also used to detect DNA, proteins and small molecules.A versatile, ultrasensitive immunosensor for detection of influenza virus was designed by combining silver nanoparticles (Ag NPs) labeled antibodies with indirect fluorescence. A new technology using Ag–S covalent binding was applied for antibody labeling. Influenza A (H1N1) virus, as a subtype of influenza A virus that was the most common cause of human influenza (flu), was acted as the target antigen using sandwich type-immunoreactions on the high binding ELISA plates. The antibody-labeled Ag NPs were then released by acid solution to produce Ag+ which can catalyze o-phenylenediamine (OPDA) oxidation to produce fluorescence for highly sensitive detection. Under the optimal conditions, it shows good linear relationship between fluorescence intensity and the logarithm of the concentration of H1N1 over the range of 1.0×10−12–1.0×10−8 g mL−1 with a detection limit (LOD, 3σ) of 1.0×10–13 g mL−1. Results indicated that the proposed method give a good sensitivity and simple operation for detecting the influenza virus. This work also provided a promising potential for antigen detection by Ag NPs labeled, and the steps were easy to handle.

A versatile, ultrasensitive chemiluminescent metalloimmunoassay method for detection of H1N1 influenza virus was designed by using silver nanoparticles as an anti-H1N1 labeling tag to strongly amplify the CL signal of luminol.

N-phosphorylation labeling was utilized to analyze the low molecular weight compounds by using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). A wide range of natural amino acids and short peptides was successfully analyzed by MALDI-TOF MS without matrix background interferences. The N-phosphorylation labeling reaction was carried out easily within 30 min in a one-pot reaction under mild reaction conditions. The phosphoryl derivatization reaction is a global labeling approach with high selectivity and high specificity with targeting only on the N-terminal and ε-amino group of Lys. The incorporation of a neutral phosphoryl group with high gas-phase affinity of protons not only improves the ionization efficiency of target molecules and simultaneously decreases the ion suppression effects in MALDI-TOF MS analysis, but also greatly reduces or eliminates the matrix background interferences by suppressing the matrix signals and increasing the molecular weight of the targeted compounds. By applying the N-phosphorylation labeling approach, many amino acids could be detected in serum samples by using MALDI-TOF MS.

2,3,7,8-Tetrachlorodibenzo-p-dioxin (2,3,7,8-TCDD) is known as a highly toxic environmental contaminant and poses potential risks to human and animal health. As a persistent organic pollutant, 2,3,7,8-TCDD is hard to metabolize and thus gets accumulated in the human and animal body. Literature reports on the metabolism study of 2,3,7,8-TCDD are rare. The traditional method of GC-MS for 2,3,7,8-TCDD analysis might not be amenable for the direct analysis of its polar metabolites. In this study, in vitro metabolism of 2,3,7,8-TCDD with rat liver microsomes by using LC-MS and MS/MS was investigated and two hydroxylated metabolites of 2,3,7,8-TCDD and four trichloro-dihydroxydibenzo-p-dioxins were identified from the direct LC-MS and MS/MS analyses of the incubated samples.

Aconitine, hypaconitine, mesaconitine, bulleyaconitine and lappaconitine are Aconitum alkaloids that have been proved to be neurotoxic and cardiotoxic. A method with liquid–liquid extraction and liquid chromatography coupled with electrospray ionization-tandem mass spectrometry (LC/ESI-MS/MS) was developed and validated for the quantification of the five Aconitum alkaloids in human urine. Upon the successful chromatographic separation, the alkaloids were determined by ion trap tandem mass spectrometry. The precursor and major product ions of aconitine at m/z 646 → 586, mesaconitine at m/z 632 → 572, hypaconitine at m/z 616 → 556, bulleyaconitine at m/z 644 → 586 and lappaconitine at m/z 585 → 535 were monitored in positive ion mode. Linear calibration curves were generated from 0.02–5 μg mL−1 with coefficients of greater than 0.99. The method was validated with the intra-day and inter-day precisions represented by relative standard deviation of less than 7% in urine, and the recoveries from spiked urine samples varied from 89.8% to 96.4% for lappaconitine, 87.4–96.3% for hypaconitine, 81.5–90.6% for mesaconitine, 82.6–90.0% for aconitine and 82.2–88.7% for aconitine. Stability of aconitine was assessed under acidic, neutral and alkaline conditions. Aconitine hydrolysis was not observed during the urine sample pretreatment under neutral and weak acidic conditions.

•Current analytical techniques for POPs in environment and biota are reviewed.•The review covers most updated literatures reports on POPs analysis.•For the first time, analysis of new POPs under Stockholm Convention is reviewed.•Future perspectives on POPs, especially the potential POPs, are discussed.Persistent organic pollutants (POPs) are major environmental concern due to their persistence, long-range transportability, bio-accumulation and potentially adverse effects on living organisms. Analytical chemistry plays an essential role in the measurement of POPs and provides important information on their distribution and environmental transformations. Much effort has been devoted during the last two decades to the development of faster, safer, more reliable and more sensitive analytical techniques for these pollutants. Since the Stockholm Convention (SC) on POPs was adopted 12 years ago, analytical methods have been extensively developed. This review article introduces recent analytical techniques and applications for the determination of POPs in environmental and biota samples, and summarizes the extraction, separation and instrumental analyses of the halogenated POPs. Also, this review covers important aspects for the analyses of SC POPs (e.g. lipid determination and quality assurance/quality control (QA/QC)), and finally discusses future trends for improving the POPs analyses and for potential new POPs.

•The latest developments in mass spectrometry (MS) are reviewed.•Recent MS applications in food safety and food quality are introduced.•Different MS techniques are described and compared.•The novel characteristics of the newly-developed ambient MS are discussed.This review introduces fundamental principles and applications of different types of mass detector and highlights the novel advances of newly developed mass-spectrometry (MS) methods, including MALDI-TOF-MS imaging and ambient ionization MS for direct food analysis. We also discuss and compare the advantages and the limitations of different MS techniques in their applications to food safety and quality, and comment on the future outlook.

N-Phosphorylation labeling was utilized to analyze small metabolites using matrix-assisted laser desorption–ionization time-of-flight mass spectrometry (MALDI-TOF MS). The incorporation of a neutral phosphoryl group with high gas-phase proton affinity not only improved the ionization efficiency of the target molecules, but also greatly reduced the matrix background interference.

In this paper, we describe the development of a novel stable isotope N-phosphorylation labeling (SIPL) strategy for peptide de novo sequencing and protein quantification based on organic phosphorus chemistry. The labeling reaction could be performed easily and completed within 40 min in a one-pot reaction without additional cleanup procedures. It was found that N-phosphorylation labeling reagents were activated in situ to form labeling intermediates with high reactivity targeting on N-terminus and ε-amino groups of lysine under mild reaction conditions. The introduction of N-terminal-labeled phosphoryl group not only improved the ionization efficiency of peptides and increased the protein sequence coverage for peptide mass fingerprints but also greatly enhanced the intensities of b ions, suppressed the internal fragments, and reduced the complexity of the tandem mass spectrometry (MS/MS) fragmentation patterns of peptides. By using nano liquid chromatography chip/time-of-flight mass spectrometry (nano LC-chip/TOF MS) for the protein quantification, the obtained results showed excellent correlation of the measured ratios to theoretical ratios with relative errors ranging from 0.5% to 6.7% and relative standard deviation of less than 10.6%, indicating that the developed method was reproducible and precise. The isotope effect was negligible because of the deuterium atoms were placed adjacent to the neutral phosphoryl group with high electrophilicity and moderately small size. Moreover, the SIPL approach used inexpensive reagents and was amenable to samples from various sources, including cell culture, biological fluids, and tissues. The method development based on organic phosphorus chemistry offered a new approach for quantitative proteomics by using novel stable isotope labeling reagents.

Perfluorinated compounds (PFCs) are ubiquitous in the environment and are becoming a public health concern. It is desirable to develop sensitive and accurate methods to measure PFCs in non-invasive matrices such as hair and nail for biomonitoring of body burden. Different extraction methods coupled with solid phase extraction were investigated for extraction efficiency. The extracts were separated, identified and quantified by liquid chromatography-tandem mass spectrometry. Extraction with acetonitrile proved to be the most efficient extraction method for human hair sample, while extraction by methanol with alkaline digestion performed best for human nail sample. The matrix recoveries of the optimized methods ranged from 78% to 116% for hair and from 87% to 126% for nail sample. The ranges of the limit of detection (LOD) were 0.026–0.069 ng/g and 0.023–0.094 ng/g for hair and nail, respectively. These methods were validated by evaluating LOD, accuracy and precision and were proven to be useful for measuring paired human hair and nail samples collected from the general population.Highlights► The methods for determination of PFCs in human hair and nail are developed. ► The methods provide new matrixes for biomonitoring of human PFCs exposure. ► In this study, the data of PFCs in paired human hair and nail is presented firstly.

Hydroxylated polybrominated diphenyl ethers (OH-PBDEs) have emerged as contaminants of environmental concerns because they pose potential risks to human and animal health. The purpose of this study was to investigate the in vitro metabolism of OH-PBDEs and their potential inhibition against 17β-estradiol (E2) metabolism.Rat liver microsomes were used as a source of P450 enzymes in an in vitro metabolism study of OH-PBDEs. Inhibition of E2 metabolism and kinetic study were performed by incubating with rat liver microsomes in the presence of OH-PBDEs.The obtained data clearly demonstrated that OH-PBDEs, especially those congeners with lower bromination, could be metabolized to bromophenol and diOH-PBDEs. The less metabolic rate of OH-PBDEs was observed with the increasing number of bromine substituents. OH-PBDEs with hydroxyl group and bromine adjacent to the ether bridge showed faster metabolic rates. In addition, the results showed non-competitive inhibition of E2 metabolism by OH-PBDEs with IC50 values in the range from 13.7 to 55.2 μM. The most potent OH-PBDE inhibitor was found to be 3′-OH-BDE-100. The inhibitory potencies for OH-PBDEs were significantly higher than those of parent PBDE and methoxylated metabolites, providing the evidence that PBDEs exerted estrogenic activity in part by their hydroxylated metabolites.OH-PBDEs exhibited large differences in their capacity to be metabolized and to inhibit E2 metabolism in rat liver microsomes. The finding might increase our understanding of healthy risk associated with PBDEs in human and wildlife.

A novel method for the characterization of polymers by laser desorption/ionization on the layer of graphene nanoparticles coupled with time-of-flight mass spectrometry was demonstrated. Various polymers including polypropylene glycol, polystyrene and polymethyl methacrylate with average molecular weights from 425 to 3500 Da were analyzed.

This work presents a new approach for the analysis of small molecules with direct negative ion laser desorption/ionization (LDI) on graphene flakes. A series of matrix interference-free mass spectra were obtained for the analysis of a wide range of small molecules including peptides, amino acids, fatty acids, as well as nucleosides and nucleotides. The mixture of analytes and graphene flakes suspension were directly pipetted onto a sample plate for LDI-time-of-flight mass spectrometry (TOFMS) analysis. Deprotonated monomeric species [M−H]− ions were homogeneously obtained on uniform graphene flakes film when negative ion mode was applied. In positive ion mode, the analytes were detected in form of multiple adduct ions such as sodium adduct [M+Na]+, potassium adduct [M+K]+, double sodium adduct [M+2Na−H]+, double potassium adduct [M+2K−H]+, as well as sodium and potassium mixed adduct [M+Na+K−H]+. Better sensitivity and reproducibility were achieved in negative ion mode compared to positive ion mode. It is believed that the new method of matrix interference-free negative ion LDI on graphene flakes may be expanded for LDI-MS analysis of various small molecules.

Perfluorooctane sulfonyl fluoride (PFOSF) is a main precursor of environmentally ubiquitous perfluorooctanesulfonate (PFOS), and the quantity released to the environment is substantial. Determination of PFOSF, particularly at low concentrations, presents significant challenges for high-performance liquid chromatography and liquid chromatography/mass spectrometry (LC/MS) analyses due to the lack of chromophore and ionizable functional group, respectively. In this study, a new method was developed by derivatizing PFOSF with benzylamine to allow rapid quantitative analysis by using LC/MS. The method demonstrated good linearity in the range from 2 to 80 ng mL–1 with r2 > 0.994 for the derivatization product while the absolute detection limit was 2.5 pg. Liquid–liquid and liquid–solid extraction procedures were established for analysis of water and soil samples, and recoveries were in the range of 51–128%. In addition, the derivatization was selective for PFOSF, whereas PFOS did not nearly react. The developed simple analytical method with good reproducibility might not only be applied for analysis of PFOSF in the environment but also be applicable for supporting investigations on environmental fate of PFOSF, particularly its environmental and biotransformation to PFOS.

This study investigated the formation of DNA adducts of polybrominated diphenyl ethers (PBDEs) and the possible mechanisms. DNA adduction was conducted by in vitro reaction of deoxyguanosine (dG) and DNA with PBDE-quinone (PBDE-Q) metabolites, and DNA adducts were characterized by using electrospray ionization tandem mass spectrometry. The results suggested DNA adduction involved Michael Addition between the exocyclic NH2 group at the N-2 position of dG and the electron-deficient carbon of quinone, followed by reductive cyclization with loss of (bromo-)1-hydroperoxy-benzene or water to form a type I or type II adduct. PBDE-Q with substituted bromine on the quinone ring was proven to be a favorable structure to form a type I adduct, while the absence of bromine on the quinone ring resulted in a type II adduct. Lower reactivity of adduction was also observed with increasing the number of bromine atoms on the phenoxyl ring. Our data clearly demonstrated PBDEs could covalently bind to DNA mediated by quinone metabolites, depending on the degree of bromine substitution. This study opened a new view on the mechanism of toxicity of PBDEs and reported the structure of PBDE–DNA adducts, which might be valuable for the evaluation on potential in vivo formation of PBDE–DNA adducts.

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) has been demonstrated to have the adverse effects on human health. In this study, we applied a metabolomic approach in conjunction with unsupervised and supervised machine learning methods to investigate the toxic effects of TCDD. By using liquid chromatography/quadrupole time-of-flight mass spectrometry, non-targeted metabolomic analysis revealed the metabolic signatures of the toxicity in aryl hydrocarbon receptor (AhR)-high affinity C57BL/6J (C6) mice as well as low affinity strain-DBA/2J (D2) mice. Lysophospholipids and long chain fatty acids were strikingly elevated in the C6 mice exposed to TCDD in both liver and skeletal muscle tissues. Meanwhile, the level of palmitoylcarnitine, which is one of the important indicators in fatty acid β-oxidation, increased significantly. Moreover, several nucleosides and amino acids decreased markedly. On the other hand, much less differentiating metabolites were highlighted in another strain-D2 mouse model. Taking liver and skeletal muscle tissues together, the levels of inosine, valine and glutamine decreased significantly. One lysophospholipid and two fatty acids were found to be enhanced. The principal components analysis and support vector machine clustering results also exhibited discriminations in the liver and skeletal muscle tissues of the mice. The obtained results indicated that TCDD could disrupt several metabolic pathways, including fatty acid biosynthesis and amino acid metabolism in both C6 and D2 mice. The increased rate of fatty acid beta-oxidation, however, was only observed in the liver and skeletal muscle tissues of C6 mice. The perturbation of the tricarboxylic acid (TCA) cycle was testified in two strains but the change was much slighter in D2 mice. It was of particular interest to note that the succinate level was enhanced in the liver tissues of both strains, and particularly, the change was up to 11.49-fold in the liver of C6 mice treated with TCDD. Collectively, the discrimination of D2 mice was not as distinct as that of C6 mice when exposed to the same dosage. Furthermore, D2 was confirmed to be less-sensitive rather than resistant to a high dose of TCDD.

High-fructose diet-fed rats as one of the insulin resistant models was used widely for understanding the mechanisms of type 2 diabetes mellitus. Systems-level metabolic profiling of the rat model, however, has not been deciphered clearly. To address this issue, mass spectrometry-based metabolomics was employed to unlock the metabolic snapshots of the oral glucose tolerance test (oGTT) effect in either healthy or diabetic rats, as well as to delineate the metabolic signatures in tissues of rats fed with high-fructose diet. Several differentiating metabolites were highlighted to reveal the metabolic perturbation of the oGTT effects in healthy and diabetic rats, which involved amino acid biosynthesis, polyunsaturated fatty acids, phospholipids and purine metabolism. Surprisingly, the patterns of relationships for the metabolic phenotypes by using data mining revealed that glucose ingestion might induce the healthy group to display its trajectory towards diabetic status, while only a very slight influence was observed on the high-fructose diet-fed rats 120 min after glucose ingestion. The data treatment for liver, skeletal muscle and brain tissues suggested that oxidative stress, such as lipid peroxidation and the declined antioxidant, the elevated amino acids and the perturbation of fatty acids, were caused by the high-fructose diet in liver and skeletal muscle tissues. On the other hand, the up-regulation in purine biosynthesis and the decreased concentrations for amino acids were observed in the cerebral cortex and hippocampus tissues. Collectively, the obtained results might provide a new insight not only for the impairment of glucose tolerance but also for the dietary style in rats.

Although numerous studies have been performed for the toxicological mechanisms of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), the metabolic changes of TCDD toxicity is less well understood. In this study, liquid chromatography/quadrupole time-of-flight mass spectrometry (LC/QTOFMS) was used for non-targeted metabolomics for understanding the different metabolic patterns associated with TCDD exposure in aryl hydrocarbon receptor (AhR) sensitive C57BL/6J (C6) and less sensitive DBA/2J (D2) mouse strains. The serum samples were analyzed and treated with metabolomic analysis in conjunction with multivariate data analysis. Metabolite identification was performed with interpreting high resolution MS data and MS/MS fragmentation, searching against databases and comparing with authentic compounds. Twelve differentiating metabolites (defined as a ≥1.5-fold change with a P ≤ 0.001) were highlighted in C6 mice versus control group, revealing lipid accumulation, fatty acid beta-oxidation, inflammation and alteration of amino acids as well as phase II drug-like metabolism. In contrast, only 2 differentiating metabolites were detected in D2 mouse model.Non-targeted metabolomics was applied for investigating different metabolic patterns associated with TCDD exposure in aryl hydrocarbon receptor sensitive C57BL/6J and less sensitive DBA/2J mouse strains.

Ultra performance liquid chromatography (UPLC) provides improved resolution, speed and sensitivity compared to conventional high performance liquid chromatography (HPLC). In this study, a robust UPLC–ESI–MS/MS method was developed for the rapid determination of nine hydroxylated polybrominated diphenyl ethers (OH-PBDEs) in rat plasma. Under the optimized conditions, the OH-PBDE congeners were eluted within 7.0 min. The limits of quantification defined at the signal-to-noise ratio of 10 were 0.17–2.78 ng/mL in rat plasma. The method provided good linearity for the calibration curves with recoveries of 93.3–114.0% and repeatability with relative standard deviation (RSD) of 0.6–5.8% for intra-day and 3.2–10.4% for inter-day measurements. The developed method was applied for supporting the pharmacokinetics investigation of 6-OH-BDE-47 in two groups of Sprague–Dawley rats that received, respectively a single dose of 0.60 mg/kg (high dose) and 0.15 mg/kg (low dose) by intravenous injection. The results showed that plasma levels of 6-OH-BDE-47 declined bi-exponentially with elimination half-life of 71.7 and 85.6 min for lower and higher dose group, respectively. The obtained results of short elimination half-life suggested that 6-OH-BDE-47 might not accumulate significantly in rat.

Aristolochic acid nephropathy (AAN) is associated with the prolonged exposure to nephrotoxic and carcinogenic aristolochic acids (AAs). DNA adducts induced by AAs have been proven to be critical biomarkers for AAN. Therefore, accurate and specific quantification of AA–DNA adducts is important. In this study, a specific method using ultra performance liquid chromatography–tandem mass spectrometry (UPLC–MS/MS) was developed and applied for the determination of 7-(deoxyadenosin-N6-yl)aristolactam I (dA-AAI) in exfoliated urothelial cells of AA-dosed rats. After the isolation from urine samples, DNA in urothelial cells were subjected to enzymatic digestion and solid-phase extraction on a C18 Sep-Pak cartridge for the enrichment of DNA adducts. The sample extracts were analyzed by reverse-phase UPLC–MS/MS with electrospray ionization in positive ion mode. The quantification of the AA–DNA adduct was performed by using multiple reaction monitoring with reserpine as internal standard. The method provided good accuracy and precision with a detection limit of 1 ng/ml, which allowed the detection of trace of dA-AAI in exfoliated urothelial cells. After one-month oral dose of AAI at 10 mg/kg/day, 2.1 ± 0.3 dA-AAI per 109 normal dA was detected in exfoliated urothelial cells of rats. Compared to the traditional methods such as 32P-postlabelling and HPLC with fluorescence detection, the developed UPLC–MS/MS method is more specific and rapid with a retention time of 4 min. The outcome of this study may have clinical significance for diagnosing and monitoring AA-associated disease because detection of DNA adducts in exfoliated urothelial cells is non-invasive and convenient.

Metabolomics, the exponentially developing technique, could provide a systemic mapping in toxicology by directly measuring small molecular metabolites. 2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) was found to be neurotoxic in mammalian animals. In this study, we employed liquid chromatography/quadrupole time-of-flight mass spectrometry for non-targeted analysis of metabolic profiling in hippocampal sample sets of the rats exposed to TCDD. Hippocampal metabolome from different ages of the healthy rats (4-week, 12-week and 20-week) was also deciphered. The relationship between the two tested cases was unlocked to delineate TCDD toxicity associated with ageing. Tandem mass spectrometry fragmentation in conjunction with metabolic database searching and compared to authentic standards was utilized for metabolite identification. As a consequence, the reduced levels of phenylalanine and leucine/isoleucine as well as the up-regulation of inosine and hypoxanthine were highlighted for understanding of TCDD toxicity related to age in rats and the trajectory was depicted by principal components analysis.

Prolonged intake of aristolochic acid (AA) has been shown to be associated with the development of certain renal disorders. Renal tubular atrophy and interstitial fibrosis are the early symptoms of AA nephropathy. The symptoms were observed in rats that were dosed with AA at a dosage of 10 mg/kg/day for 1 month. Apart from the renal tubular atrophy and interstitial fibrosis, AA–DNA adducts were detected in the rat kidney tissue. Differentiated proteins were identified in the kidney tissues from proteomics investigations. The upregulated proteins identified included ornithine aminotransferase, sorbitol dehydrogenase, actin, aspartoacylase, 3-hydroxyisobutyrate dehydrogenase, and peroxiredoxin-1. Downregulated proteins such as ATP synthase subunit β, glutamate dehydrogenase 1, regucalcin, glutamate–cysteine ligase regulatory subunit, dihydropteridine reductase, hydroxyacyl-coenzyme A dehydrogenase, voltage-dependent anion-selective channel protein 1, prohibitin, and adenylate kinase isoenzyme 4 were also identified. Several identified protein markers were found to have biological and medical significance.

Oxygen exchange may occur at carboxyl groups catalyzed by acid. The reaction, however, takes at least several days at room temperature. The long-time exchanging reaction often prevents its application from protein analysis. In this study, an 18O-labeling method utilizing microwave-assisted acid hydrolysis was developed. After being dissolved in 16O/18O (1:1) water containing 2.5% formic acid, protein samples were exposed to microwave irradiation. LC-MS/MS analysis of the resulted peptide mixtures indicated that oxygen in the carboxyl groups from glutamic acid, aspartic acid, and the C-terminal residues could be efficiently exchanged with 18O within less than 15 min. The rate of back exchange was so slow that no detectable back exchange could be found during the HPLC run.

A new pressure-assisted capillary electrochromatography coupled with electrospray ionization-mass spectrometry method using a silica-based monolithic column as separation media was developed for the analysis of β2-agonists in human urine. Experimental conditions including the mobile phase, separation voltage, assisted pressure, and sheath liquid were optimized for the analysis: mobile phase composed of 82% (v/v) ACN and 18% (v/v) 20 mmol/L ammonium acetate (pH 6.0); separation voltage 25 kV; assisted pressure 2 bar; and the sheath liquid consisting of 7.5 mmol/L acetic acid in isopropanol/water 50/50% (v/v) that was delivered at a flow rate of 3.0 μL/min. Six β2-agonists were separated within 12.5 min with LODs (defined as S/N = 3) in the range of 0.25–2.0 ng/mL. The absolute LODs of the developed method for analyzing six β2-agonists ranged from 5.75 to 46.0 fg. Method repeatability of run-to-run and column-to-column was satisfactory. The recovery obtained from the analysis of spiked urine samples was between 88.2% and 106% with RSDs lower than 6.68%. The method was successfully applied to the analysis of real urine sample from volunteers.

Metabolomics is the downstream of systems biology and has drawn significant interest for studying the metabolic networks from cells to organisms. To profile the metabolites in two different cell lines (A549 and AGS) infected with influenza A virus, gas chromatography coupled with mass spectrometry (GC/MS) was employed. Some differentiating metabolites in the cell lines were tentatively identified using reference library, interpreted and visualized by applying principal components analysis (PCA) and cluster heat map. Consequently, metabolic flux profiling allowed the differentiation of fatty acid biosynthesis and cholesterol metabolism during viral replication in the cell lines. The change in fatty acid turnover was also observed. Metabolomics investigation also revealed the different responses between A549 and AGS cell lines to the virus infection. From the pattern recognition results, AGS cell line might be more susceptible to influenza A virus. Regarding the fact that AGS is a poorly differentiated gastric adenocarcinoma cell line whereas A549 is a relatively differentiated lung tumor one, it is speculated that viral replication might be associated with the cell differentiations.

A simple and effective method was developed for peptide sequencing and protein identification through the determination of its N-terminal residue. The method of N-terminal carbamidomethylation with iodoacetamide could specifically and remarkably enhance the intensity of a1 ions in the tandem mass spectra of the peptide derivatives without significantly altering their fragmentation pattern, thus allowing determination of their N-terminal residues. The effectiveness and specificity of the method was demonstrated by confirming and extending sequence interpretation of several model peptides and proteins. The developed method was then applied in the LC-MS/MS analysis of the tryptic digests of myoglobin and a whole protein extract from rat heart tissues. The results from database searches were well validated with the enhancement of a1 ions in tandem mass spectra and the specificity of protein identification was obtained when the information of N-terminal residues was included in the database search.N-terminal carbamidomethylation with iodoacetamide provided significant enhancement of a1 ions in MS/MS analysis of tryptic peptide (ALELFR) from myoglobin and allowed the identification of the N-terminal amino acids.Figure optionsDownload full-size imageDownload high-quality image (95 K)Download as PowerPoint slide

Nucleoprotein (NP), the structural component of ribonucleoprotein complex of avian influenza virus, performs multiple essential functions in the regulation of viral RNA synthesis and in the control of nuclear traffic of viral proteins. Mutations have often been found in NP, some of which are relevant to viral survival strategies. In this study, we used nanospray-MS/MS to analyze tryptic digestion of nucleoprotein of avian influenza virus (H5N1) and to identify three mutated peptides. The MS/MS analyses allowed the confident determination of the three mutated amino acid residues F313Y, I194V and V408I/L in the mutated peptides of LLQNSQVYSLIRPNENPAHK, GVGTMVMELVR and ASAGQI/LSVQPTFSVQR, respectively.

Determination of O-glycosylation sites in glycopeptides was developed by using two model compounds designed from mucin2 tandem repeat motif and erythropoietin. β-Elimination/addition reaction using dimethylamine on glycosylated site through a Michael-type condensation produced efficient deglycosylation with appropriate chemical modification. The use of dimethylamine was efficient to release the O-linked glycan in a reaction time period of 2–6 h at 55 °C. Peptide sequencing was then performed using the liquid chromatography/quadrupole time-of-flight mass spectrometry and MS–MS experiments. Interpretation of fragmentation pathways of the β-elimination/addition products enabled straightforward recognition of glycosylation site. Compared to the fragmentation of corresponding native peptides, mass shift of −18 Da or +27 Da was clearly observed for the two kinds of β-elimination/addition products of the glycosylated threonine. Dimethylamine was found to provide higher efficiency of β-elimination/addition than methylamine and ammonia.

Aristolochic acid (AA), a mixture of structure-related nitrophenanthrene carboxylic acid derivatives derived from Aristolochia spp, is associated with nephrotoxin and carcinogen. AA-DNA adducts induced by reductive metabolic activation of AA were detected in tissues of animals and in patients exposed to AA. The DNA adducts were generally used as biomarkers in toxicological study of AA. In this short review, quantitative analysis of AA-DNA adducts in various in vitro and in vivo systems by using 32P-postlabelling assay, HPLC-UV, HPLC-radiation monitor, HPLC-FLD, HPLC-ESI/MS and UPLC-MS/MS methods is discussed. The distribution of AA-DNA adducts in various tissues is also summarized.

Matrix protein 1 (M1), the major structural protein of the avian influenza virus, plays a critical role in regulation of viral RNA transcription via interaction with RNA and transportation of RNP cores. Mutations in M1 have been frequently observed in the highly virulent avian influenza H5N1 virus, which might be crucial to the pathogenic function. Here we report the characterization of mutated peptides in M1 purified from highly pathogenic avian influenza virus H5N1 by nanoelectrospray MS and MS/MS analyses on a quadrupole-time-of-flight mass spectrometer (Q-TOFMS). The specificity of tandem mass spectrometry allowed the identification of six amino acid (AA) substitutions in M1, including R95K, A166V, I168T, N207S, N224S, and R230K. Two commonly observed modifications such as oxidation and deamidation were accurately assigned in the protein. Bioinformatics analysis suggested some relationship between the amino acid substitution and structural property of M1 protein. Discussions on de novo sequencing of MS/MS spectra, especially in dealing with the AA substitutions, were provided.

A high performance liquid chromatography coupled with electrospray ionization/mass spectrometry method was developed for the determination of adenosine 5′-monophosphate (AMP), adenosine 5′-diphosphate (ADP), and adenosine 5′-triphosphate (ATP) in the extract of HepG-2 cells. The chromatographic conditions were optimized by using porous graphitic carbon as the stationary phase for the retention and separation of the AMP, ADP and ATP. Negative-ion mode ESI-MS in basic mobile phase was applied to improve the method sensitivity. An external calibration method with linear ranges from 0.22 to 57.80 μM for AMP, from 0.59 to 117.37 μM for ADP, and from 0.49 to 98.81 μM for ATP was used for quantitative analysis. The levels of ATP, ADP, and AMP in HepG-2 cells treated with benzo[a]pyrene with different time periods were determined. Total adenine nucleotides and the energy charge potential were calculated for the investigation of the effect of benzo[a]pyrene on cell energy metabolism.

A sensitive high-performance liquid chromatography method with fluorescence detection (HPLC-FLD) for the determination of DNA adducts induced by nephrotoxic and carcinogenic aristolochic acid (AA) is presented. The DNA adduct of AAII (dA-AAII) was synthesized by in vitro incubation, purified by preparative HPLC, characterized using fluorescence spectroscopy and high-resolution mass spectrometry, and was used as the biomarker for AA exposure in rats. The developed HPLC-FLD method was validated and applied for the determination of dA-AAII in rat kidney tissues. The method provided a detection limit of 18.3 fmol, which allowed the detection of dA-AAII in the rat kidney tissue samples collected after a single oral dose of AA. dA-AAII was detected in the kidney DNA digestion extracts of the rats that were dosed with AA at 5 mg/kg and 30 mg/kg at concentrations of 6.2 ± 1.1 and 41.3 ± 8.0 dA-AAII per 109 normal dA, respectively.

A sensitive and rapid method of high-performance liquid chromatography with fluorescence detection (HPLC-FLD) was developed and applied for the determination of aristolochic acid I (AAI) in rat urine and plasma. Prior to the HPLC analysis, the samples were derivatized to increase fluorescence character. The linear ranges of the calibration curves were 0.48–48 ng in urine and 0.23–69 ng in plasma. The intra- and inter-day precisions referred by relative standard deviation (RSD) were less than 3.2% and 4.0% for urine samples as well as less than 9.4% and 10.8% for plasma samples. The limits of quantification were 0.09 and 0.07 ng in urine and plasma, respectively. The developed method was successfully applied for the determination of AAI in rat urine and plasma samples collected after the oral administrations of AAI standard and Aristolochia contorta Bge. (Madouling) or Aristolochia manshuriensis (Guanmutong) herbal extracts. The ratios of the detected AAI amount in urine compared to the dosing amount of AAI were approximately constant. The concentrations of AAI in rat plasma were much lower than those in urine. The obtained results indicated that the metabolism of the AAI standard and AAI-containing herbs might be different, probably due to the complicated and multiple components in the herbs.

Highly active antiretroviral therapy (HAART) is the common treatment strategy for human immunodeficiency virus (HIV)-infected patients at present. Generally, HAART regimens apply multitherapy drugs that contain nucleoside/nucleotide reverse transcriptase inhibitors (NRTIs), non-nucleoside/nucleotide reverse transcriptase inhibitors (NNRTIs) and protease inhibitors (PIs). Unlike NNRTIs and PIs, the active form of NRTIs is not the drug itself but its triphosphorylated (TP) metabolites in intracellular medium. Analysis of both the prodrugs or NRTIs and their intracellular metabolites is needed to provide overall information in pharmacokinetic and therapeutic effects to HIV-infected patients. Numerous publications have reported the assays for NRTIs and their phosphorylated metabolites in various biological matrices. The methods involved liquid chromatography (LC) with UV detection (LC-UV), LC with tandem mass spectrometry (LC–MS/MS), capillary electrophoresis/electrochromatography (CE/CEC) with UV detection (CE/CEC-UV) or/and MS/MS detection (CE-MS/MS). Due to the extremely low concentration of NRTIs and the phosphorylated metabolites as well as the complex biological matrices, sample pretreatment methods such as protein precipitation (PP), liquid–liquid extraction (LLE) and solid-phase extraction (SPE) have played important role in the successful analytical method development.

We demonstrated that anionic porphyrins could be stacked and separated in micellar electrokinetic chromatography (MEKC) and microemulsion electrokinetic chromatography (MEEKC) by applying acetonitrile and high salt content in human urine sample matrix. The introduction of sample containing acetonitrile and sodium chloride into the CE capillary at more than 10% of the total capillary volume resulted in the improvement of peak resolution and the enhancement of detection sensitivity. The achieved acetonitrile stacking enrichment factors of six porphyrins ranged from 12 to 32 in MEKC and from 28 to 33 in MEEKC, respectively. The stacking technique was successfully applied for analyzing porphyrins present in urine samples that were deproteinized with acetonitrile. For the analysis of coproporphyrin isomers, addition of the sodium cholate (SC) into micelle and microemulsion solutions provided adequate resolution. Calibration curves obtained for the determination of coproporphyrin isomers were found linear between 30 and 400 nmol L−1, and the limit of detection (LOD) was 20 nmol L−1 in MEEKC. Intra- and interday precisions (n = 11) in the microemulsion separation system for the isomers at spiked concentrations of 40–400 nmol L−1 in urine were in the range of 0.1–0.4% and 0.7–7.6% for migration time and peak area, respectively. Coproporphyrin III, coproporphyrin I and uroporphyrin were detected at levels of 80.7 nmol L−1, 32.3 nmol L−1 and 19.8 nmol L−1, respectively, in the urine samples collected from healthy individuals. Different porphyrin profiles, however, were observed in urine samples from porphyria cutanea tarda (PCT) patients.

Metabolism and pharmacokinetic studies on rat were conducted for lithospermic acid B, one of the components from Radix Salviae Miltiorrhizae (danshen) that shows many bioactivities. Liquid chromatography–electrospray ionization mass spectrometry method was applied for the determination of lithospermic acid B and its metabolites in samples from in vitro and in vivo metabolism studies. Rat plasma samples collected after intravenous administration were analyzed for obtaining pharmacokinetic data of lithospermic acid B. Four O-methylated metabolites, namely one monomethyl-, two dimethyl- and one trimethyl-lithospermic acid B, were detected when lithospermic acid B was incubated in rat hepatic cytosol. These four metabolites were also detected in rat bile, plasma and feces samples after intravenous administration of lithospermic acid B. The in vitro and in vivo results indicate that the methylation is the main metabolic pathway of lithospermic acid B. The danshen component and its methylated metabolites were excreted to rat bile and feces.

Electrophilic attack of aristolactam-nitrenium ion by the C7 position to the exocyclic amino group in the DNA bases led to the formation of the major adducts. In this study, liquid chromatography coupled with electrospray ionization tandem mass spectrometry was applied to the study of DNA adducts of aristolochic acid (AA). When DNA (bases and CT-DNA) was incubated with AA, dG-AAI, dG-AAII, dA-AAI, dA-AAII, dC-AAI, and dC-AAII were detected and characterized. The dC adducts of AA were identified for the first time. The soft ionization technology allowed detection of the intact DNA adducts. High-resolution MS and MS-MS capabilities of a quadrupole time-of-flight mass spectrometer were shown to be efficient for DNA adducts analysis. DNA-AA adducts showed characteristic fragmentation patterns in MS-MS analysis. The dissociative loss of 116 Da from the DNA-AA adducts, which resulted from internal hydrogen transfer and cleavage at the C—N glycosidic bond, provided a characteristic fragment for the structural elucidation.

A LC–MS/MS method was developed for the separation and simultaneous determination of phenolic components including danshensu, protocatechuic acid, protocatechuic aldehyde and caffeic acid as well as tanshinones including cryptotanshinone, tanshinone I and tanshinone IIA in samples of Radix Salviae Miltiorrhizae and Salviae Miltiorrhizae tablet. Triple quadrupole mass spectrometry was optimized in both positive and negative ion multiple reaction monitoring modes for the simultaneous quantitative analysis of the two different types of active components by using a time-segment program. The method gave recoveries of 85.4–106.4% with relative standard deviations of 2.4–8.0% for the spiked herb samples. The limits of detection were 0.30–0.83 μg/g for the analysis of 1.0 g Radix Salviae Miltiorrhizae or tablet samples.

Analytical method using mass spectrometric techniques was applied for the determination of polybrominated diphenyl ethers (PBDEs) in freshwater fishes. Fish samples collected from Nanyang River contaminated by the recycling electron-wastes (e-wastes) materials were prepared by using Soxhlet extraction and multiple-step column chromatographic clean-up. PBDEs were determined by gas chromatography (GC) coupled with ion trap mass spectrometry (for mono- to hepta-BDEs) and quadrupole mass spectrometry (for BDE-209). The method performance was evaluated with the recovery of 13C-labeled internal standards and with the analysis of certified reference biota. The obtained recoveries ranged from 75 to 125% with a relative standard deviation of lower than 10% for 16 PBDE congeners. The total PBDE (ΣPBDE) concentrations in fishes showed the following trend: grass carp < mud carp < crucian carp < silver carp < carp. ΣPBDE concentrations in the abdomen, back and tail muscles of carp ranged from 766, 458 and 530 ng/g w.w., and 53, 52, 45 ng/g w.w. in grass carp, respectively. The ΣPBDE concentrations in abdomen muscles were no significantly higher than in back and tail muscles in carp, crucian carp, grass carp and mud carp. PBDE congener concentrations in muscles correlated well with their lipid content. BDE-47 and BDE-28 were the most abundant congeners followed by BDE-17, BDE-15, BDE-66, BDE-154 and BDE-153 in fishes collected from Guiyu.

Triclosan in the waste, river and sea water samples collected in Hong Kong was analyzed by using gas chromatography–ion trap mass spectrometry method. 13C12-triclosan was used as internal standard for the quantitative analysis. Water samples were prepared and cleaned-up by using a C18 solid-phase extraction cartridge. The recoveries of triclosan in spiked coastal water at three different concentrations ranged from 83 to 110%. The method detection limit was 0.25 ng/L for triclosan in 1-L water and the relative standard deviations and relative error were less than 11.0 and 12.3%, respectively (n = 3). The method was successfully applied to analyze water samples collected from rivers, coastal water bodies and wastewater treatment plants at ng/L levels.

A column chromatography procedure was developed for the clean-up of solvent-extracted sediment samples for the fractionation of polybrominated diphenyl ethers (PBDEs) and polybrominated dibenzo-p-dioxins and dibenzofurans (PBDD/Fs). The procedure included multiple column chromatography steps for clean-up for the separation of PBDEs from PBDD/Fs. The separation of the two chemical groups overcame the mutual interfering problem during the GC–ion trap MS analysis. The method was validated with the analysis of quality control samples. The method accuracy represented with relative error was less than 16% for all targeted PBDEs and PBDD/Fs congeners. Recoveries of the 13C-labeled standards ranged from 64% to 117% with relative standard deviation from 7.3% to 15%. Results from the analysis of environmental sediment samples collected in the vicinity of a recycling site for electronic wastes showed high levels of PBDEs (1.5–12 ng/g, dry weight), trace levels of PBDFs (0.025–0.92 ng/g, dry weight) and non-detectable PBDDs.

Liquid chromatography coupled with mass spectrometry and tandem mass spectrometry has been applied to investigate the in vivo metabolism of ginsenoside Rb1 in rat. Both positive electrospray ionization mass spectrometry and negative electrospray ionization mass spectrometry were used to identify the Rb1 and its metabolites in rat plasma, urine, and feces samples. Oxygenation and deglycosylation were found to be the major metabolic pathways of Rb1 in rat. A total of nine metabolites were detected in urine and feces samples collected after intravenous and oral administration. Deglycosylated metabolism of Rb1 generated other ginsenosides as the major metabolites, such as Rd, Rg3 or F2, Rh2, or C-K. This result indicates that the ginsenoside Rb1 has many pharmacological activities and could be used as a prodrug.

A comprehensive method was developed for quantitative analysis of polybrominated diphenyl ethers (PBDEs), polychlorinated biphenyls (PCBs) and polychlorinated dibenzo-p-dioxins and dibenzo-furans (PCDD/Fs) in one single extract of environmental samples. The sample preparation procedure included two fractionation steps using silver nitrate silica chromatography to separate PBDEs from PCBs and PCDD/Fs and florisil column to separate PCBs from PCDD/Fs. Acidic silica, acidic alumina and gel permeation chromatography (GPC, for PCBs) or activated carbon column (for PCDD/Fs) were used for further clean-up. The sample extracts were analyzed by using high-resolution gas chromatography/high-resolution mass spectrometry. The entire method was validated from the analysis of mixed standards of PBDEs, PCBs and PCDD/Fs (n = 3); the analysis of certified reference biota (WMF-01). The method was applied for the analysis of 10 sediment samples collected from Haihe River and Dagu Drainage River in Tianjin City. No significant PBDEs pollution was found in the areas.

A method for the trace analysis of antifouling agent Irgarol 1051 and its decyclopropylated degradation product in seawater was developed by using reversed-phase LC–MS with quadrupole time-of-flight mass spectrometry detection. Atrazine-d5 was used as the internal standard for the LC–MS determination. Irgarol 1051 and the major degradation product were detected at trace levels in seawater samples collected from several marinas in Hong Kong. The water samples were pre-concentrated by solid-phase extraction with a polystyrene-divinylbenzene cartridge. Quadrupole time-of-flight tandem mass spectrometry in positive ion electrospray ionization was used for the identification and quantitation. The recoveries of Irgarol 1051 and degradation product were >85%. Method detection limits were 0.2 ng l−1 and 0.8 ng l−1 for the antifouling agent and the decyclopropylated degradation product, respectively.

Soil samples collected from an electronic waste recycling site were prepared by using Soxhlet extraction and multiple-step column chromatographic clean-up. Gas chromatography/ion trap mass spectrometry method was developed to determine polybrominated diphenyl ethers (PBDEs) in the sample extracts. The method performance was evaluated by the recovery of 13C-labeled internal standards and by analyzing quality assurance and quality control samples. Relative error and relative standard deviation obtained from the analysis of duplicated samples and spiked matrix were better than 10%. PBDEs were detected in the field soil samples collected from an e-wastes disposal site at levels from low parts-per-billions to 600 parts-per-billions.

Three pairs of ginsenoside isomers (Rg2 and Rg3, Rg1 and F11 as well as Rd and Re) were differentiated and identified through accurate mass measurement of mass spectrometry (MS) and MS–MS. [M + Li]+ and [M − H]− ions were detected in full-scan MS analyses and selected for the MS–MS experiments using positive and negative ion electrospray ionizations (ESI), respectively. The structures of aglycone and α- and β-saccharide sugars in various ginsenosides were determined from the spectrum interpretation and accurate mass measurement. Z and C type ions were predominantly observed in the MS–MS spectra of [M + Li]+ ions, while Y type ions were the most abundant ions in the spectra obtained from the negative ion mode analysis. Furthermore, X and A ions resulted from cross-ring cleavage on the sugar directly connected to aglycone were detected in both positive and negative ion spectra, which provided the site information of the saccharide chains. The obtained MS–MS profiles were used for the structural confirmation of ginsenoside Rg2 collected from column chromatography separation of a Chinese Panax ginseng extract. The ESI–MS data with accurate mass assignment suggested that a co-eluted ginsenoside also existed in the sample fraction. The interpretation of its MS–MS spectrum and fragmentation pathways allowed the detection of the ginsenoside Rf, differentiating from its isomers Rg1 and F11.

Metabolism of an anti-tumor active component of Panax ginseng, ginsenoside (20R)-Rg3, was studied for better understanding its pharmacokinetics in rat. LC–MS was used to determine Rg3 and its metabolites in rat plasma, urine and feces samples. An average half-life of 18.5 min was obtained after the ginsenoside was intravenously dosed at 5 mg/kg. However, Rg3 was not detected in rat plasma collected after oral administration at 100 mg/kg. Only 0.97–1.15% Rg3 of the dosed amount was determined in feces. Hydrolysis and oxygenated metabolites were detected and identified in feces collected after oral administration by using LC–MS and MS–MS.

A method using ion-pairing liquid chromatography–mass spectrometry (MS) was developed for analyzing adenosine 5′-monophosphate (AMP), adenosine 5′-diphosphate (ADP), and adenosine 5′-triphosphate (ATP) in cellular extracts. Dimethylhexylamine (DMHA) was used as ion-pairing agent to retain and separate the analytes on a reversed-phase microbore column with a gradient program. Positive-ion electrospray ionization–MS was applied for the detection because of the use of the ion-pairing agent. Adduct ions of DMHA with AMP, ADP, and ATP were found to be the most intensive peaks and thus selected as quantitative ions. An external calibration method with linear ranges from 0.1 to 20 μM for AMP, 2 to 20 μM for ADP, and 2.5 to 20 μM for ATP was used for the quantitation. The method was applied to determine concentrations of AMP, ADP, and ATP in extracts of cultured rat C6 glioma cells that were pretreated with various concentrations of Zn. The detected levels of the adenosine nucleotides have been used to calculate total adenosine nucleotide and energy charge potential. Changes in cellular energy status upon exposure to increasing concentration of Zn in the culture medium were analyzed. The results indicated that the addition of Zn in a range of 40 to 120 μg/ml cause a gradual increased in energy charge potential of the cells.

A sensitive and specific liquid chromatography (LC)–electrospray ionization (ESI) mass spectrometry (MS) method with quadrupole-time-of-flight detection was applied for the determination of ginsenoside Rg3 in rat plasma and its major metabolites in samples from in vitro metabolism studies. The biological samples were prepared by protein precipitation with methanol and the sample extracts were directly injected into a reversed-phase HPLC column with solvent gradient program. Pharmacokinetic study of Rg3 on rat gave a half-life of 14 min after the ginsenoside was intravenously dosed. Metabolite identification and confirmation from the MS–MS and high-resolution MS analyses indicated that the short half-life might be due to the high rate of metabolic clearance of the ginsenoside under gastrointestinal conditions. Hydrolysis products of Rg3 such as ginsenoside Rh2 and protopanaxadiol were identified as the major metabolites from the incubation with 0.1 M HCl, and a hydrated product of protopanaxadiol was detected as the metabolite with artificial gastric juice, while a monooxygenated metabolite was identified in the incubated samples with rat liver S9 fraction.

A method of capillary HPLC–high-resolution MS was developed for the trace analysis of ATP, GTP, dATP and dGTP. Dimethylhexylamine (DMHA) was used as ion-pairing agent for the HPLC retention and separation of the nucleotides and positive ion electrospray time-of-flight MS was used for the detection. The application of capillary HPLC allowed minimal usage of DMHA while providing excellent peak retention and resolution, which significantly reduced the ion suppression in electrospray ionization-MS analysis and thus increased the sensitivity. Adduct ions of nucleotides and DMHA were used as quantitative ions in order to achieve the best sensitivity. DMHA concentration at 5 mM in the aqueous mobile phase at pH 7 was found to be the optimal conditions for the C18 capillary column. The method was applied to determine ATP level in cultured C6 glioma cells that were treated with toxic concentrations of Zn. The results showed that the cellular ATP level decreased from 2.7 pmol/cell (<10% cell death) in average control cell samples to 0.36 pmol/cell as the concentration of Zn increased to 120 mg/l (>35% cell death) in culture medium.

Matrix protein 1 (M1), the major structural protein of the avian influenza virus, plays a critical role in regulation of viral RNA transcription via interaction with RNA and transportation of RNP cores. Mutations in M1 have been frequently observed in the highly virulent avian influenza H5N1 virus, which might be crucial to the pathogenic function. Here we report the characterization of mutated peptides in M1 purified from highly pathogenic avian influenza virus H5N1 by nanoelectrospray MS and MS/MS analyses on a quadrupole-time-of-flight mass spectrometer (Q-TOFMS). The specificity of tandem mass spectrometry allowed the identification of six amino acid (AA) substitutions in M1, including R95K, A166V, I168T, N207S, N224S, and R230K. Two commonly observed modifications such as oxidation and deamidation were accurately assigned in the protein. Bioinformatics analysis suggested some relationship between the amino acid substitution and structural property of M1 protein. Discussions on de novo sequencing of MS/MS spectra, especially in dealing with the AA substitutions, were provided.Six amino acid substitutions in a major structural protein of avian influenza virus were identified by nanoelectrospray Q-TOFMS and MS/MS. Oxidation at methionines and deamidation at asparagines were determined.Download high-res image (100KB)Download full-size image

•Adsorption of triclosan and methyl triclosan to biological sludge was investigated.•The Linear and Freundlich isotherms were able to describe the sorption system well.•The adsorption systems followed pseudo second order kinetic model.•Inactivated dried sludge showed promising results for removal of TCS and MTCS.Biosorption equilibrium and kinetics of TCS and MTCS sorption to inactivated dried sludge were studied in a batch system with respect to pH, temperature and sludge concentration. Linear and Freundlich isotherms were able to describe the adsorption system well. Sorption capacity increased from 4.80 to 7.36 μg/g and 6.28 to 7.76 μg/g for TCS and MTCS, respectively with an increase in temperature from 15 to 35 °C. As pH decreased from 11.0 to 5.0, the sorption capacity of TCS and MTCS enhanced from 4.36 to 7.92 μg/g and 5.52 to 7.52 μg/g, respectively. The distribution coefficient for MTCS was higher than that for TCS due to hydrophobicity. The calculated Kf and 1/n implied that the adsorption process was physiosorption and exothermic in nature involving weak forces such as Van der Waal’s interactions. In the kinetic study, adsorption of TCS and MTCS to dried sludge predominantly followed a pseudo-second order kinetic over the range of applied initial concentrations based on regression coefficients and the relative error for the calculated equilibrium sorption capacity. IR analysis of sludge showed the presence of various polar groups that could highly affect the sorption of organic pollutants.Download high-res image (148KB)Download full-size image

Electrophilic attack of aristolactam-nitrenium ion by the C7 position to the exocyclic amino group in the DNA bases led to the formation of the major adducts. In this study, liquid chromatography coupled with electrospray ionization tandem mass spectrometry was applied to the study of DNA adducts of aristolochic acid (AA). When DNA (bases and CT-DNA) was incubated with AA, dG-AAI, dG-AAII, dA-AAI, dA-AAII, dC-AAI, and dC-AAII were detected and characterized. The dC adducts of AA were identified for the first time. The soft ionization technology allowed detection of the intact DNA adducts. High-resolution MS and MS-MS capabilities of a quadrupole time-of-flight mass spectrometer were shown to be efficient for DNA adducts analysis. DNA-AA adducts showed characteristic fragmentation patterns in MS-MS analysis. The dissociative loss of 116 Da from the DNA-AA adducts, which resulted from internal hydrogen transfer and cleavage at the CN glycosidic bond, provided a characteristic fragment for the structural elucidation.

•Fate of triclosan was investigated in different wastewater and sludge treatments.•TCS was considerably biodegraded in aerobic/anaerobic sludge digestion process.•TCS transformation products were observed based on the treatment practice.•Kd values were estimated for TCS, 2,4-DCP and MTCS in primary and secondary sludge.•TCS temporal variations as well as few relationships were investigated.Triclosan (TCS) as an antimicrobial agent has been ubiquitously found in wastewater and sewage sludge. TCS may undergo transformation/degradation during wastewater treatment. Some of the resulted products such as 2,4-dichlorophenol (2,4-DCP), 2,8-dichlorodibenzoparadioxin (2,8-DCDD) and methyl triclosan (MTCS) are presumed toxic/persistent compounds. In this study, fate of TCS and the probability of formation of important degradation products were investigated in three susceptible wastewater/sludge treatment practices. 24.1% and 27.2% of the loading TCS was adsorbed to the generated sludge, whereas up to 60% of the loading TCS was biotransformed. Up to 9.9% and 13.0% of TCS loss was attributed to the formation of 2,4-DCP and 2,8-DCDD in chlorination and UV disinfection, respectively. Anaerobic and aerobic sludge digestion processes eliminated up to 23.0% and 56.0% of TCS, respectively. About 7.4% of TCS in aerobic digestion was transformed to methyl triclosan (MTCS). Significant temporal variation of TCS was observed in primary sedimentations, except for chemically enhanced primary treatment that was suggested to be governed by chemical-forced sedimentation. Distribution coefficient (Kd) of TCS was directly correlated to the total organic carbon of the sludge (TOC). Moreover, strong correlation was observed between elimination efficiency in primary sedimentation and loading concentration.Download high-res image (100KB)Download full-size image

•BPAQ was capable of binding directly to the nucleophilic site of ribonucleosides/deoxyribonucleosides in vitro via a Michael addition.•BPAQ produced depurinating adducts that was lost from deoxyribonucleosides, generating apurinic sites in the deoxyribonucleosides.•ESI–MS/MS is a powerful analytical tool for the detection of suspected ribonucleosides/deoxyribonucleosides and GSH adducts.Bisphenol A is a monomer used in the manufacture of polycarbonate plastic products, epoxy resin-based food can liners and flame retardants. To determine the genotoxic potential of bisphenol A, the mechanism of the reactions between the reactive electophilic bisphenol A 3,4-quinone (BPAQ) with glutathione and ribonucleosides/deoxyribonucleosides were studied. The obtained results demonstrated that BPAQ reacted with 2′-deoxyguanosine (dG)/guanosine (G), 2′-deoxyadenosine (dA)/adenosine (A), but not with 2′-deoxycytidine (dC)/cytidine (C) and thymidine (T)/uridine (U) in aqueous acetic acid. The reactions were accompanied by loss of deoxyribose, and the rate of depurination by deoxyribonucleoside adducts were faster than that of ribonucleoside adducts. In mixtures of ribonucleosides and deoxyribonucleosides treated with BPAQ, reactions occurred more readily with dG/G than dA/A. The structures of the modified bases were confirmed by electrospray ionization tandem mass spectrometry (ESI–MS/MS). We also found that BPAQ reacted readily with glutathione (GSH) in aqueous acetic acid, and characterized the BPAQ-GSH conjugate by ESI–MS/MS. The in vitro data of depurinating DNA/RNA adducts and BPAQ-GSH adducts may provide appropriate reference for the identification of BPAQ adducts in environmental and biological systems.

•PM2.5 induces heart mitochondrial morphological damage of rats.•Mitochondrial fission/fusion gene expression is important regulation mechanism.•Proinflammatoy cytokine level changes are accompanied with mitochondrial damage.•Alterations in oxidative stress and calcium homeostasis are focused on.Epidemiological studies suggested that ambient fine particulate matter (PM2.5) exposure was associated with cardiovascular disease. However, the underlying mechanism, especially the mitochondrial damage mechanism, of PM2.5-induced heart acute injury is still unclear. In this study, the alterations of mitochondrial morphology and mitochondrial fission/fusion gene expression, oxidative stress, calcium homeostasis and inflammation in hearts of rats exposed to PM2.5 with different dosages (0.375, 1.5, 6.0 and 24.0 mg/kg body weight) were investigated. The results indicated that the PM2.5 exposure induced pathological changes and ultra-structural damage in hearts such as mitochondrial swell and cristae disorder. Furthermore, PM2.5 exposure significantly increased specific mitochondrial fission/fusion gene (Fis1, Mfn1, Mfn2, Drp1 and OPA1) expression in rat hearts. These changes were accompanied by decreases of activities of superoxide dismutase (SOD), Na+K+-ATPase and Ca2+-ATPase and increases of levels of malondialdehyde (MDA), inducible nitric oxide synthase (iNOS) and nitric oxide (NO) as well as levels of pro-inflammatory mediators including TNF-α, IL-6 and IL-1β in rat hearts. The results implicate that mitochondrial damage, oxidative stress, cellular homeostasis imbalance and inflammation are potentially important mechanisms for the PM2.5-induced heart injury, and may have relations with cardiovascular disease.Download full-size image

•An asymmetric labeling strategy and PCI-GC–MS–MS approach for determination of non-amino organic acid and amino acid for the first time, as well as short chain fatty acid.•This asymmetric labeling strategy and PCI-GC–MS–MS provided the special neutral loss can be useful in the selective identification for qualitative analysis of organic acids and amino acid derivatives.•We provide a new alternative for the separation and identification of non-amino organic acid and amino acid.Alkyl chloroformate have been wildly used for the fast derivatization of metabolites with amino and/or carboxyl groups, coupling of powerful separation and detection systems, such as GC–MS, which allows the comprehensive analysis of non-amino organic acids and amino acids. The reagents involving n-alkyl chloroformate and n-alcohol are generally employed for providing symmetric labeling terminal alkyl chain with the same length. Here, we developed an asymmetric labeling strategy and positive chemical ionization gas chromatography—tandem mass spectrometry (PCI-GC–MS–MS) approach for determination of non-amino organic acids and amino acids, as well as the short chain fatty acids. Carboxylic and amino groups could be selectively labelled by propyl and ethyl groups, respectively. The specific neutral loss of C3H8O (60 Da), C3H5O2 (74 Da) and C4H8O2 (88 Da) were useful in the selective identification for qualitative analysis of organic acids and amino acid derivatives. PCI-GC–MS–MS using multiple reaction monitoring (MRM) was applied for semi-quantification of typical non-amino organic acids and amino acids. This method exhibited a wide range of linear range, good regression coefficient (R2) and repeatability. The relative standard deviation (RSD) of targeted metabolites showed excellent intra- and inter-day precision (<5%). Our method provided a qualitative and semi-quantitative PCI-GC–MS–MS, coupled with alkyl chloroformate derivatization.

•BPS induced immunotoxicity in macrophages by metabolomics and toxicological analysis.•BPS exposure varied expression and secretion of cytokines, and macrophage polarization.•BPS exposure disrupted glycolysis and the metabolism of GSH and lipids in macrophages.•Omics study plays a significant role in exploring the toxicity mechanism of BPS.As an important structural analogue of bisphenol A (BPA), bisphenol S (BPS) has been used as alternatives to BPA in industrialized production. However, the immunotoxicity of BPS remains poorly understood. As a critical model in inflammatory responses, macrophages are used to explore the immunotoxic potential and mechanisms of BPS at environmentally relevant concentrations in our study. Here, we are combining molecular toxicology and mass spectrometry (MS)-based global metabolomics and lipidomics study together to estimate the variation of cytokines profiling and metabolism characteristic following BPS exposure. Our results demonstrated that BPS exposure induced pro-inflammatory phenotype by activating the immuno-related cytokines which include TNF-α, IL-1β and IL-6, modulating metabolic pathways which include glycolytic, glutathione (GSH), sphingomyelin (SM)-ceramide (Cer), glycerophospholipids (GPs) and glycerolipids (GLs). These toxicological mechanisms are providing us with a deeper understanding of the critical role of metabolites and lipids reprogramming in immunotoxicity of BPS.Download high-res image (143KB)Download full-size image

Aristolochic acid (AA), derived from the herbal genus Aristolochia and Asarum, has recently been shown to be associated with the development of nephropathy. Upon enzyme activation, AA is metabolized to the aristolactam-nitrenium ion intermediate, which reacts with the exocyclic amino group of the DNA bases via an electrophilic attack at its C7 position, leading to the formation of the corresponding DNA adducts. The AA-DNA adducts are believed to be associated with the nephrotoxic and carcinogenic effects of AA. In this study, liquid chromatography coupled with electrospray ionization mass spectrometry (LC–MS) was used to identify and quantify the AA–DNA adducts isolated from the kidney and liver tissues of the AA-dosed rats. The deoxycytidine adduct of AA (dC–AA) and the deoxyadenosine–AA adduct (dA–AA) were detected and quantified in the tissues of rats with one single oral dose (5 mg or 30 mg AA/kg body weight). The deoxyguanosine adduct (dG–AA), however, was detected only in the kidney of rats that were dosed at 30 mg AA/kg body weight for three consecutive days. The amount of AA–DNA adducts found in the rats correlated well with the dosage.

•Evaluation of electron ionization and negative chemical ionization under selected ion monitoring or multiple reaction monitoring mode.•A simple, accurate and sensitive GC-MS/MS assay for the determination of OCPs.•The method was validated and was successfully used for the analysis of real human samples.Gas chromatography-triple quadrupole mass spectrometry (GC-QqQMS) was applied for the determination of eight organochlorine pesticides (OCPs) in human serum. OCPs were extracted from the serum sample by solid phase extraction (SPE) and analyzed by gas chromatography mass spectrometry (GC–MS) or gas chromatography tandem mass spectrometry (GC–MS/MS). Electron ionization (EI) and negative chemical ionization (NCI) under two data acquisition modes, namely selected ion monitoring (SIM) and multiple reaction monitoring (MRM), were compared. The use of MRM generally provided higher selectivity and sensitivity because less interference from the sample matrix existed. The EI mode is more suitable for less electronegative compounds such as dichlorodiphenyldichloroethanes (DDDs) with detection limits ranging from 0.0060 to 0.060 ng/mL. In the NCI mode, MRM analysis provided good and lower detection limits (0.0011–0.0030 ng/mL) for pesticides containing more chlorines. The methods were validated by analyzing the pesticides in spiked serum at different levels with recoveries ranged from 83% to 116% and relative standard deviations of less than 10%. The developed method was applied for the determination of the OCPs in real human serum samples.

2,3,7,8-Tetrachlorodibenzo-p-dioxin (2,3,7,8-TCDD) is known as a highly toxic environmental contaminant and poses potential risks to human and animal health. As a persistent organic pollutant, 2,3,7,8-TCDD is hard to metabolize and thus gets accumulated in the human and animal body. Literature reports on the metabolism study of 2,3,7,8-TCDD are rare. The traditional method of GC-MS for 2,3,7,8-TCDD analysis might not be amenable for the direct analysis of its polar metabolites. In this study, in vitro metabolism of 2,3,7,8-TCDD with rat liver microsomes by using LC-MS and MS/MS was investigated and two hydroxylated metabolites of 2,3,7,8-TCDD and four trichloro-dihydroxydibenzo-p-dioxins were identified from the direct LC-MS and MS/MS analyses of the incubated samples.

Hepatocellular carcinoma (HCC) is one of the most common cancers in human beings and not well treated. Due to its blurred biomarkers, lipidomics can be considered as a new diagnostic tool. In this study, mass spectrometry-based lipidomics analysis coupled with multivariate statistical analysis was applied to profile twelve pairs of human hepatocellular carcinoma tissues (HCT) and matched adjacent non-tumour tissues (ANT). As a result, perturbation of lipid biosynthesis was observed in HCT compared to ANT. Lipid species were profiled and 28 significant lipids were identified based on high resolution mass spectrometry with mass error less than 3 ppm, covering phosphatidylcholine (PC), phosphatidylethanolamine (PE), phosphatidylglycerol (PG), phosphatidylinositol (PI), sphingomyelin (SM) and triglyceride (TG). The high mass accuracy as well as the tandem MS (MS/MS) technique provides high confidence of lipid identification. Interestingly, decreased PGs and PIs were observed in this study, indicating that lipolysis of PGs and PIs might play a crucial role in HCC development. The results indicated that mass spectrometry-based lipidomics analysis could serve as an effective means to diagnose canceration.

Dioxin exposure tends to accumulate in adipose tissue and alters metabolism in mammals. In this study, gas chromatography coupled with mass spectrometry (GC-MS) in conjunction with multivariate statistical analysis was applied to profile small molecular metabolites in adipose tissue of aryl hydrocarbon receptor (AhR)-high affinity wild-type C57BL/6J mice exposed to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). A partial least squares discriminant analysis model was also constructed to map the discrimination between TCDD dosages and the control group. As a result, a total of 16 differential metabolites were identified in the high-dose TCDD group compared to the control group, and 12 free fatty acids (FFAs) were highlighted among them. Both saturated and unsaturated FFA levels were significantly elevated in adipose tissues of TCDD-exposed mice. This promising tool for global characterization highlights FFAs which could be served as indicators for understanding the toxic responses to TCDD exposure in a dose-dependent manner. The data indicated that the use of GC-MS coupled with multivariate statistical analysis could provide new insight for fatty acid biosynthesis on AhR activation with TCDD exposure in wild-type mice.

Core–shell structured magnetic covalent organic frameworks (Fe3O4@COFs) were synthesized via a facile approach at room temperature. Combining the advantages of high porosity, magnetic responsiveness, chemical stability and selectivity, Fe3O4@COFs can serve as an ideal absorbent for the highly efficient enrichment of peptides and the simultaneous exclusion of proteins from complex biological samples.

Targeting protein–protein interactions (PPIs) offers tantalizing opportunities for therapeutic intervention for the treatment of human diseases. Modulating PPI interfaces with organic small molecules has been found to be exceptionally challenging, and few candidates have been successfully developed into clinical drugs. Meanwhile, the striking array of distinctive properties exhibited by metal compounds renders them attractive scaffolds for the development of bioactive leads. Here, we report the identification of iridium(III) compounds as inhibitors of the H-Ras/Raf-1 PPI. The lead iridium(III) compound 1 exhibited potent inhibitory activity against the H-Ras/Raf-1 interaction and its signaling pathway in vitro and in vivo, and also directly engaged both H-Ras and Raf-1-RBD in cell lysates. Moreover, 1 repressed tumor growth in a mouse renal xenograft tumor model. Intriguingly, the Δ-enantiomer of 1 showed superior potency in the biological assays compared to Λ-1 or racemic 1. These compounds could potentially be used as starting scaffolds for the development of more potent Ras/Raf PPI inhibitors for the treatment of kidney cancer or other proliferative diseases.

Aconitine, hypaconitine, mesaconitine, bulleyaconitine and lappaconitine are Aconitum alkaloids that have been proved to be neurotoxic and cardiotoxic. A method with liquid–liquid extraction and liquid chromatography coupled with electrospray ionization-tandem mass spectrometry (LC/ESI-MS/MS) was developed and validated for the quantification of the five Aconitum alkaloids in human urine. Upon the successful chromatographic separation, the alkaloids were determined by ion trap tandem mass spectrometry. The precursor and major product ions of aconitine at m/z 646 → 586, mesaconitine at m/z 632 → 572, hypaconitine at m/z 616 → 556, bulleyaconitine at m/z 644 → 586 and lappaconitine at m/z 585 → 535 were monitored in positive ion mode. Linear calibration curves were generated from 0.02–5 μg mL−1 with coefficients of greater than 0.99. The method was validated with the intra-day and inter-day precisions represented by relative standard deviation of less than 7% in urine, and the recoveries from spiked urine samples varied from 89.8% to 96.4% for lappaconitine, 87.4–96.3% for hypaconitine, 81.5–90.6% for mesaconitine, 82.6–90.0% for aconitine and 82.2–88.7% for aconitine. Stability of aconitine was assessed under acidic, neutral and alkaline conditions. Aconitine hydrolysis was not observed during the urine sample pretreatment under neutral and weak acidic conditions.

One kind of surface protein imprinting method was developed by a more convenient, simpler and cheaper approach based on vinyl-functionalized magnetic nanofibers (NFs).

Zeolitic imidazolate framework-8 coated magnetic nanocomposites (Fe3O4@ZIF-8 MNCs) served as an absorbent and a matrix for negative-ion MALDI-TOF MS. The host–guest property and interference-free background made them an ideal dual platform for the sensitive analysis of small molecules.

A versatile, ultrasensitive chemiluminescent metalloimmunoassay method for detection of H1N1 influenza virus was designed by using silver nanoparticles as an anti-H1N1 labeling tag to strongly amplify the CL signal of luminol.

N-Phosphorylation labeling was utilized to analyze small metabolites using matrix-assisted laser desorption–ionization time-of-flight mass spectrometry (MALDI-TOF MS). The incorporation of a neutral phosphoryl group with high gas-phase proton affinity not only improved the ionization efficiency of the target molecules, but also greatly reduced the matrix background interference.

A novel method for the characterization of polymers by laser desorption/ionization on the layer of graphene nanoparticles coupled with time-of-flight mass spectrometry was demonstrated. Various polymers including polypropylene glycol, polystyrene and polymethyl methacrylate with average molecular weights from 425 to 3500 Da were analyzed.

Bromodomain-containing protein 4 (BRD4) has recently emerged as an attractive epigenetic target for anticancer therapy. In this study, an iridium(III) complex is reported as the first metal-based, irreversible inhibitor of BRD4. Complex 1a is able to antagonize the BRD4-acetylated histone protein–protein interaction (PPI) in vitro, and to bind BRD4 and down-regulate c-myc oncogenic expression in cellulo. Chromatin immunoprecipitation (ChIP) analysis revealed that 1a could modulate the interaction between BRD4 and chromatin in melanoma cells, particular at the MYC promoter. Finally, the complex showed potent activity against melanoma xenografts in an in vivo mouse model. To our knowledge, this is the first report of a Group 9 metal complex inhibiting the PPI of a member of the bromodomain and extraterminal domain (BET) family. We envision that complex 1a may serve as a useful scaffold for the development of more potent epigenetic agents against cancers such as melanoma.