Wan Chan

Proteomics analysis of differential expression of cellular proteins in response to avian H9N2 virus infection in human cells

We present the first proteomic analysis on the cellular responses to avian influenza virus (H9N2) infection in a human cell line in different time courses in order to search for target proteins for viral pathogenesis/adaptation studies. By using 2‐DE coupled with MALDI‐TOF MS and nano‐ESI‐MS/MS, we identified a set of differentially expressed cellular proteins, including cytoplasmic actin, cytokeratin, prohibitin, enoyl‐CoA hydratase, peptide‐prolyl cis–trans isomerase A (PPIase A), chloride intracellular channel protein 1, pyruvate dehydrogenase E1 component subunit beta, adenine phosphoribosyltransferase, guanine nucleotide‐binding protein subunit beta, nucleoside diphosphate kinase A, elongation factor 1‐beta and splicing factor, arginine/serine rich 1. The most significant changes in different time courses were found in cytoplasmic actin and cytokeratin, both of which constituted the major components of cytoskeleton network in the cells. The obtained data suggested a possible role of the cytoskeleton during avian influenza virus infection of mammalian cells, which might help for better understanding of the dynamics of avian influenza virus and host interaction in mammalian cell setting.

One Pot Phase Transfer Synthesis of Trithiocarbonates from Carbon Bisulphide and Alkyl Halides

Abstract Symmetrical and cyclic alkyl trithiocarbonates were synthesized by reacting CS2 and 33% aqueous NaOH with alkyl hal-ides under phase transfer catalysis conditions

Investigation of the Metabolism and Reductive Activation of Carcinogenic Aristolochic Acids in Rats

The metabolic activation of aristolochic acids (AAs) that have been demonstrated to be mutagenic and carcinogenic was investigated. In vitro metabolism study indicated that AAs were metabolized to N-hydroxyaristolactam, which could be either reduced to aristolactams or rearranged to 7-hydroxyaristolactams via the Bamberger rearrangement. In vivo metabolism study is important because the intermediates (aristolactam-nitriumion) of the nitroreduction process are thought to be responsible for the carcinogenicity of AAs. Liquid chromatography-mass spectrometry and liquid chromatography-tandem mass spectrometry (MS/MS) were applied to the analyses of a series of positional isomers of hydroxyaristolactams in rat urine samples after the in vivo study of AAs. Three hydroxylated metabolites of aristolactam II and two hydroxylated metabolites of aristolactam I were identified. The structures of the positional isomers were elucidated from the interpretation of MS/MS spectra and theoretical calculations. In addition, several new metabolites were detected in the rat urine by high-resolution mass spectrometry and MS/MS, including those from the decarboxylation of AAs and the conjugations of acetylation, glucuronidation, and sulfation of aristolochic acid Ia.

One Pot Phase Transfer Synthesis of O-Alkyl, S-Methyl Dithiocarbonates (Xanthates)

Abstract O-Alkyl, S-methyl dithiocarbonates of phenol, benzyl, primary and secondary alcohols were prepared in a one pot procedure under phase transfer catalysis conditions.

Liquid chromatography/mass spectrometry for metabonomics investigation of the biochemical effects induced by aristolochic acid in rats: The use of information-dependent acquisition for biomarker identification

The toxic effects of oral administrations of nephrotoxic and carcinogenic aristolochic acid (AA) to male Sprague-Dawley rats were investigated by using high-performance liquid chromatography coupled with a quadrupole time-of-flight mass spectrometer. Analysis of the urine and plasma samples revealed distinct changes in the biochemical patterns in the AA-dosed rats. After peak finding and alignment, principal component analysis (PCA) and partial least-squares discriminant analysis (PLS-DA) were used for multivariate data analysis. Potential biomarkers were studied by high-resolution mass spectrometry (MS) and tandem mass spectrometry (MS/MS) analyses. The MS/MS spectra of all endogenous metabolites satisfying the pre-defined criteria were acquired in a single information-dependent acquisition (IDA) analysis, demonstrating that IDA was an efficient approach for structural elucidation in metabonomic studies. Citric acid and a glucuronide-containing metabolite were observed as potential biomarkers in rat urine. A significant increase in plasma creatinine concentration was also observed in the AA-dosed rats, which indicated that AA induced an adverse effect on the renal clearance function.

New chiral sultam auxiliaries: preparation and their application in asymmetric Diels-Alder reactions

Abstract Enantiomerically pure sultams (+)- 4 and (−)- 4 were synthesized from tricyclic sultone 1 via a four-step reaction sequence. Their uses as new chiral auxiliaries in asymmetric Diels-Alder reactions with diastereoselectivity up to 94:6 are presented.

Capillary electrophoretic study of amine/carboxylic acid-functionalized carbon nanodots

Capillary zone electrophoresis (CZE) coupled with UV absorption and laser-induced fluorescence detections has been applied to study the complexity of carbon nanodots (C-dots) products synthesized with microwave-assisted pyrolysis of citric acid (CA) and 1,2-ethylenediamine (EDA). The effects of pH and concentration of run buffer on the CZE separation of C-dots are studied in detail. The optimal acetate run buffer (30mM, pH 3.6) is subsequently employed to investigate the effect of reaction time and mole ratio of amine (NH2) to carboxylic acid (COOH) moieties of the precursors on the C-dots species present in C-dots products. Our results confirm that the synthesis of C-dots could be improved by lengthening the microwave irradiation time and optimizing the initial mole ratio of NH2/COOH in the precursors. Negatively charged C-dots are obtained only when the amount of CA exceeds that of EDA, i.e., the mole ratio of NH2/COOH is 0.25-0.80. By contrast, when the quantity (mole) of NH2 in EDA is equal to or larger than that of COOH in CA, only positively charged and neutral C-dots species are formed, inferring that the C-dots species are predominantly covered by the surface-attached ammonium and amido moieties. This work highlights the merit of CZE to identify the composition of an as-prepared C-dots product which is pretty much dependent on the mole ratio of NH2/COOH. It is anticipated that our CZE methodology will open a new avenue in optimizing the synthetic conditions for producing specific C-dots of desired composition.

Rapid screening method for intact glucosinolates in Chinese medicinal herbs by using liquid chromatography coupled with electrospray ionization ion trap mass spectrometry in negative ion mode.

An optimized method using liquid chromatography coupled with electrospray ionization ion trap mass spectrometry (LC/ESI-ITMS) in negative ion mode has been developed for screening different structural classes of intact glucosinolates in six Chinese medicinal herbs. The glucosinolates were extracted with hot methanol/water (70:30 v/v) and separation of the individual glucosinolates was achieved using a reversed-phase C18 column with an aqueous ammonium acetate/methanol gradient. Identification of the intact glucosinolates was based on the detection of compounds with a constant neutral loss of 242 Da corresponding to the combined loss of anhydroglucose (162 Da) and sulfur trioxide (80 Da) in collision-induced dissociation. The structures of the identified glucosinolates were confirmed with the use of group-specific product ions at m/z 195, 241, 259, 275 in their corresponding MS/MS product ion spectra. Differentiation of intact glucosinolates was achieved through their respective retention times and molecular masses as well as the characteristic product ions. The limits of detection were at the low nanogram level per injection, based on constant neutral loss scans. Significant variation in the compositions of intact glucosinolates was identified in the cruciferous herbs. This method was applied in the differentiation and quality control of two pairs of easily confused herbs.

Aristolochic acid induced changes in the metabolic profile of rat urine.

Prolonged exposure to aristolochic acid (AA) has shown to pose rapid progressive renal fibrosis in Belgium women in a slimming regimen in the early 90s. We hypothesize that changes in metabolic profile could have occurred before symptoms were observed, which may allow early treatment. In this study, metabonomics was used for toxicology study of AA in rats. Liquid chromatography coupled with a hybrid quadrupole time-of-flight mass spectrometry (Qq-TOF) was used for the analysis of endogenous metabolites in rat urine samples. The difference in metabolic profiles between the control and the dosed rats was well observed by the principal component analysis (PCA) of the MS data. Significant changes of two metabolite markers, kynurenic acid and hippuric acid, were detected in the rat urine samples. The identification of potential biomarkers was performed by high-resolution mass measurement and MS-MS analyses on a Qq-TOF. We believe that metabolic profiling may act as a preclinical protocol for AA exposure before symptoms are observed.

Quantification of Aristolochic Acid-Derived DNA Adducts in Rat Kidney and Liver by Using Liquid Chromatography-Electrospray Ionization Mass Spectrometry

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 (5mg or 30mg AA/kg body weight). The deoxyguanosine adduct (dG-AA), however, was detected only in the kidney of rats that were dosed at 30mg AA/kg body weight for three consecutive days. The amount of AA-DNA adducts found in the rats correlated well with the dosage.