气相色谱质谱法测定饮用水源中60种半挥发性有机物

在碱性、中性和酸性三种条件下,采用二氯甲烷萃取水中半挥发性有机物（SVOCs）,气相色谱质谱法测定饮用水源地水中的60种SVOCs。结果表明,60种SVOCs的回收率为88 %~96 %,相对标准偏差为1.1 %~4.8 %,具有较好的正确度和精密度。目标SVOCs在 0.05~1.00 μg/mL范围内有较好的线性关系,线性相关系数R2>0.99。     

Quantitative analysis of apoptotic cell death using proton nuclear magnetic resonance spectroscopy

Quantification of apoptotic cell death in vivo has become an important area of investigation in patients with acute lymphoblastic leukemia (ALL). We have devised a noninvasive analytical method to estimate the percentage of apoptotic lymphoblasts in doxorubicin-treated Jurkat T-cell ALL cultures, using proton nuclear magnetic resonance spectroscopy (1H NMR). We have found that the ratio of the methylene (CH2) resonance (at 1.3 ppm) to the methyl (CH3) resonance (at 0.9 ppm) signal intensity, as observed by 1H NMR, is directly proportional to the percentage of apoptotic lymphoblasts in vitro. The correlation between the CH2/CH3 signal intensity ratio and the percentage of apoptotic lymphoblasts was optimal 24 to 28 hours after doxorubicin treatment (r2 = .947, N = 27 samples). There was also a direct temporal relationship between an increase in the CH2/CH3 signal intensity ratio and the onset of apoptosis as detected by nuclear morphologic analysis, fluorescein-annexin V flow cytometry, and DNA gel electrophoresis. Thin-layer chromatography confirmed that a dynamic and/or compositional change of the plasma membrane, rather than increases in lipase activity or fatty acid production, appears to account for the increase in the CH2/CH3 signal intensity ratio during apoptosis. 1H NMR may have clinical utility for the early noninvasive assessment of chemotherapeutic efficacy in patients with ALL.     

Identification of 6-methylmercaptopurine derivative formed during acid hydrolysis of thiopurine nucleotides in erythrocytes, using liquid chromatography-mass spectrometry, infrared spectroscopy, and nuclear magnetic resonance assay

6-Thioguanine and 6-methylmercaptopurine (Me6-MP) nucleotides are the two major thiopurine metabolites of azathioprine found in erythrocytes. During the acid hydrolysis required for the conversion of thiopurine nucleotides into their free bases, Me6-MP was converted into a compound that could be analyzed on a Purospher RP18-e column with dihydrogen phosphate-methanol buffer as eluent. The pH of the acid extract strongly influenced the conversion of Me6-MP into its derivative. The Me6-MP derivative was identified using liquid chromatography-mass spectrometry and infrared and nuclear magnetic resonance spectrometric methods. During the acid hydrolysis of thiopurine nucleotides in erythrocytes, Me6-MP undergoes degradation, leading to 4-amino-5-(methylthio)carbonyl imidazole.     

Conformational analysis of potent sweet taste ligands by nuclear magnetic resonance, computer simulations and X-ray diffraction studies

Four potent sweet-tasting molecules, N-(3,3-dimethylbutyl)-L-aspartyl-L-phenylalanine methylester 1 (7000 times more potent than sucrose), N-(3,3-dimethylbutyl)-L-aspartyl-D-valine (S)-alpha-ethylbenzylamide 2 (3000 time more potent than sucrose), L-aspartyl-D-valine (R)-alpha-methoxymethylbenzylamide 3 (1350 times more potent than sucrose and L-aspartyl-(1R,2S,4S)-1-methyl-2-hydroxy-4-phenylhexylamide 4 (2500 times more potent than sucrose) were studied by 1H NMR and computer simulations. These flexible molecules adopt multiple conformations in solution. The "L-shaped" structure, which we believe to be responsible for sweet taste is accessible to all four compounds in solution. Extended conformations with the AH and B-containing moieties in the +y-axis and the hydrophobic group X pointing in the y-axis have also been observed for all four sweeteners. For compounds 1 and 3, the solid-state conformations were determined by X-ray diffraction studies. These results demonstrate that compounds 1 and 3 adopt an "L-shaped" structure even in the crystalline state. The extraordinary potency of the N-alkylated compound 1 compared with the unsubstituted Asp-Phe-OMe may be explained by the effect of a second hydrophobic binding domain in addition to interactions arising from the "L-shaped" structure.     

Direct analysis of several Fusarium mycotoxins in cereals by capillary gas chromatography-mass spectrometry

This article describes three experiments relating to the legibility of TV menus. Special emphasis is placed on the influence of a relatively new feature in TVs: the possibility to blend graphics and video. Three experiments are presented: one concentrating on the influences of blending level and video content; one concentrating on the influences of content and of colour combinations; and one concentrating on the influences of various font characteristics. The results are interpreted in terms of guidelines for blended TV menus.     

Analysis of nonpolar heterocyclic amines in cooked foods and meat extracts using gas chromatography-mass spectrometry

A fully automated method including column-switching and isocratic high-performance liquid chromatography (HPLC) was developed for simultaneous determination of the tricyclic antidepressant clomipramine and its metabolites demethylclomipramine, 2-, 8-, and 10-hydroxyclomipramine, 2-, and 8-hydroxydemethylclomipramine and didemethylclomipramine in serum. After serum injection into the HPLC system and on-line sample clean-up on a clean-up column (Hypersil CN; 10 x 4.6 mm) by an eluent consisting of 35% acetonitrile and 65% deionized water, the chromatographic separation was performed on an analytical column (LiChrospher CN; 250 x 4.6 mm I.D.) by an eluent consisting of 38% acetonitrile and 62% aqueous sodium perchlorate (0.02 M, pH 2.5). The UV detector was set at 260 nm. The limit of quantification was about 15 ng/ml for all analytes. The coefficients of variation ranged between 3 and 12% with recovery rates between 64 and 110%. Linear regression analyses revealed coefficients of correlation between 0.98 and 0.99. The method could be applied to therapeutic drug monitoring as well as metabolism studies in man and rat.     

Determination of gacyclidine enantiomers in human plasma by gas chromatography-mass spectrometry using selected-ion monitoring

A sensitive gas chromatographic assay using mass selective-detection has been developed for the simultaneous quantitation of the enantiomers of (+/-)-gacyclidine (a non competitive N-methyl-D-aspartate antagonist) in human plasma. Gacyclidine enantiomers and phencyclidine (PCP), the internal standard, were extracted using a single-step liquid-liquid extraction with hexane at pH 8.0. Each enantiomer was separated on a chiral gas chromatography capillary column and specifically detected by mass spectrometry (MS) in selected-ion monitoring (SIM) mode. Gacyclidine enantiomers and PCP were monitored using the fragment ions at m/z 206 and 200, respectively. No interference was observed from endogenous components. The limit of quantitation (LOQ) for each enantiomer of gacyclidine was 300 pg/ml by using plasma samples of 500 microl. The calibration curves were linear (r2=0.998) over a range of 0.3125 to 20 ng/ml. The extraction efficiency was higher than 95% for both enantiomers. Intra- and inter-day bias were less than 10% at every standard curve concentration. Intra-day precision was less than 19% for (-)-gacyclidine and 15% for (+)-gacyclidine. Inter-day precision was below 15% for both enantiomers. The assay was validated for an enantioselective pharmacokinetic study in healthy male volunteers.     

Phospholipid profiling of sediments using phosphorus-31 nuclear magnetic resonance

A phosphorus-31 nuclear magnetic resonance method has been developed for the determination of aquatic sediment phospholipid profiles that may be generally applied to all soils and deposits containing viable cellular material. A method of scrubbing chloroform/methanol extracts with potassium acid phosphate overcomes adverse signal broadening from the mineral component, permitting eleven sediment phospholipids to be determined at the quantitative level.     

Clinical magnetic resonance spectroscopy: feasibility and methods using whole body tomographs

Novel applications for cellulases have reinitiated interest in the regulation of production of these enzymes by the soft rot fungus Trichoderma reesei and related species. This paper reviews the current state of knowledge concerning the question "How can insoluble molecules like cellulose initiate their own breakdown by a microorganism?" The evidence available--based on biochemical as well as molecular biological approaches--favors a model in which conidial bound cellobiohydrolases carry out a first exo-exo-wise attack on the cellulose molecule. The disaccharides so formed (cellobiose, alpha-cellobiono-1,5-lactone) are then taken up by the mycelia and promote further cellulase biosynthesis. Evidence available suggests that they are further metabolized to, rather than being, the "true" inducer. Speculations on the nature of the inducer are presented. The roles of the beta-glucosidases of Trichoderma in this process are discussed. The pathway of cellulase secretion is discussed on the basis of electron microscopical as well as gene sequence information.     

气相色谱-质谱联用技术在砷、硒、汞和铅形态分析中的应用

气相色谱-质谱联用技术具有分析速度快、灵敏、准确的优点,其在元素形态分析中的应用越来越广泛。本文对气相色谱-质谱联用技术（GC-MS）的接口设计方法做一简要回顾。并介绍了砷、硒、汞和铅的存在形式及其形态分析的意义,综述了气相色谱与质谱联用技术在砷、硒、汞和铅四种元素的形态分析中的应用情况。展望了该联用技术的发展趋势,及其局限性。     

Spatial structure determination of antiarrhythmic peptide using nuclear magnetic resonance spectroscopy

The peptide AAP10 was synthesized according to the Merrifield technique following the Fmoc-strategy and its spatial structure in aqueous solution studied with NMR principles. It is known from previous studies that the peptide has antiarrhythmic activity and inhibits cardiac ischemia induced alterations of the activation pattern, decreases the activation-recovery interval (ARI) dispersion and improves cellular coupling via enhancement of gap junction conductance (2, 2, 3, 4). The peptide was synthesized as a peptide amide. Two different semi cyclic conformations were characterized.     

Multiresidue determination of pesticides in apples and pears by gas chromatography-mass spectrometry

This paper describes a rapid, specific and sensitive multiresidue method for the routine analysis of several classes of pesticides used for the treatment of apples and pears, involving a rapid extraction procedure at pH 4.5 with a mixture of acetone-dichloromethane-hexane (50:20:30, v/v/v) and gas chromatography coupled to mass-selective detection, in order to achieve quantitative analysis down to their respective maximum residue limit. Extraction recoveries were between 55 and 98%. Limits of detection and limits of quantitation ranged respectively, from 0.01 to 0.05 mg/kg and from 0.02 to 0.1 mg/kg. Intra-assay relative standard deviation was less than 19% for all compounds. An excellent linearity was observed from these LOQs up to 500 mg/kg. Intermediate (inter-assay) precision and accuracy were satisfactory. The method has been applied to many fruit samples intended for commercialisation.     

Automated procedure for determination of barbiturates in serum using the combined system of PrepStation and gas chromatography-mass spectrometry

A sensitive HPLC method with piroxicam as internal standard was developed for assaying amphotericin B in plasma and tissue. An Ultrabase-C18 column and a simple mobile phase consisting of an acetonitrile-acetic acid (10%)-water (41:43:16) mixture were used. The flow-rate was 1 ml/min and the effluent was monitored at 405 nm. The linearity of the assay method was up to 1000 ng/ml and 100 micrograms/g for plasma and tissue, respectively. Intra-day and inter-day RSDs were below 10% for all the sample types. This HPLC assay has been applied to determine amphotericin B in plasma and tissue samples taken during pharmacokinetic and tissue distribution studies in rats.     

The identification of metal-binding ligand residues in metalloproteins using nuclear magnetic resonance spectroscopy

The identification of metal-binding ligands in metalloproteins is an important step in gaining detailed information regarding the environment of the active site. Traditionally, techniques such as 13Cd-substitution for the active metal followed by isotope-filtered NMR techniques have been used to this end. However, for medium to high molecular weight proteins (>20 kDa), these experiments may not be beneficial due to extensive 1H spectral overlap. Here, we describe an alternative approach, where metal-binding ligands such as histidine and cysteine are specifically 15N backbone labeled, excess EDTA is added and changes to (1H-15N) HSQC spectra are followed. Under these conditions, the amide groups of all 15N labeled histidine and cysteine residues, which were either ligands or residues close to the active site, were identified unambiguously for metallo-beta-lactamase from Bacteroides fragilis.     

Diffusion tensor analysis with nuclear magnetic resonance in human central nervous system

Nuclear magnetic resonance has been used to measure the diffusivity of water molecules. In central nervous system, anisotropic diffusion, which is characterized by apparent diffusion tensor Dapp zeta, is thought to be related to neuronal fiber tract orientation. For precise observation of anisotropic diffusion, it is needed to determine the diagonal and off-diagonal elements of Dapp zeta. Once Dapp zeta is estimated from a series of diffusion weighted images, a tissue's orthotropic principal axes and diffusivity of each direction are determined from eigenvalues and eigenvectors of Dapp zeta. There are several methods to represent anisotropic diffusion with Dapp zeta. Examples are diffusion ellipsoids constructed in each voxel depicting both these principal axes and the mean diffusion length in these directions, trace invariant values and its mapping image, largest eigenvalue, and ratio of largest eigenvalue to the other eigenvalue. In this study, the author investigated practical procedure to analyze diffusion tensor Dapp zeta using both of spin-echo and echo-planer diffusion weighted imagings with 3-tesla magnetic resonance machine in human brain. The ellipsoid representation provided particularly useful information about microanatomy including neuronal fiber tract orientation and molecular mobility reflective of microstructure. Furthermore, in the lesion of Wallerian degeneration, the loss of anisotropy of local apparent diffusion was observed. It is suggested that the function of axons can be observed via degree of anisotropy of apparent diffusion. Consequently, diffusion tensor analysis is expected to be a powerful, noninvasive method capable of quantitative and functional evaluation of the central nervous system.     

Determination of butorphanol in horse race urine by immunoassay and gas chromatography-mass spectrometry

An analytical procedure to screen butorphanol in horse race urine using ELISA kits and its confirmation by GC-MS is described. Urine samples (5 ml) were subjected to enzymatic hydrolysis and extracted by solid-phase extraction. The residues were then evaporated, derivatized and injected into the GC-MS system. The ELISA test (20 microl of sample) was able to detect butorphanol up to 104 h after the intramuscular administration of 8 mg of Torbugesic, and the GC-MS method detected the drug up to 24 h in FULL SCAN or 31 h in the SIM mode. Validation of the GC-MS method in the SIM mode using nalbuphine as internal standard included linearity studies (10-250 ng/ml), recovery (+/-100%), intra-assay (4.1-14.9%) and inter-assay (9.3-45.1%) precision, stability (10 days), limit of detection (10 ng/ml) and limit of quantitation (20 ng/ml).     

Use of 13C nuclear magnetic resonance and gas chromatography to examine methionine catabolism by lactococci

Formation of methanethiol from methionine is widely believed to play a significant role in development of cheddar cheese flavor. However, the catabolism of methionine by cheese-related microorganisms has not been well characterized. Two independent methionine catabolic pathways are believed to be present in lactococci, one initiated by a lyase and the other initiated by an aminotransferase. To differentiate between these two pathways and to determine the possible distribution between the pathways, 13C nuclear magnetic resonance (NMR) performed with uniformly enriched [13C]methionine was utilized. The catabolism of methionine by whole cells and cell extracts of five strains of Lactococcus lactis was examined. Only the aminotransferase-initiated pathway was observed. The intermediate and major end products were determined to be 4-methylthio-2-oxobutyric acid and 2-hydroxyl-4-methylthiobutyric acid, respectively. Production of methanethiol was not observed in any of the 13C NMR studies. Gas chromatography was utilized to determine if the products of methionine catabolism in the aminotransferase pathway were precursors of methanethiol. The results suggest that the direct precursor of methanethiol is 4-methylthiol-2-oxobutyric acid. These results support the conclusion that an aminotransferase initiates the catabolism of methionine to methanethiol in lactococci.