Determination of beta-lactamase residues in milk using matrix-assisted laser desorption/ionization Fourier transform mass spectrometry.

A selective, fast, and effective enzyme assay based on matrix-assisted laser desorption/ionization Fourier transform mass spectrometry (MALDI-FTMS) for quantifying beta-lactamase, an illegal additive in milk products, has been reported. The strengths of the mass spectrometric assay are its response to all substrate and products, simple and direct detection of the conversion of substrate, and facile determination of enzyme activity. Also, MALDI MS is tolerant to many buffer salts and reagents without the requirement of complicated sample pretreatment procedures. In this study, the approach was used to detect the presence of beta-lactamases (BLA) in milk samples. The amount of BLA that could be determined in a milk sample is 6 x 10(-3) U x mL(-1) by this approach. To test the strategy, it has been applied to the fortified milk (adding a BLA product known as an antimicrobial destroyer). It is then tested whether the pasteurization procedure of the milk process affects the activity of BLA in milk samples. This study offers a perspective into the utility of MALDI-FTMS as an alternative detection tool for BLA screening in milk.

Electrophilic aromatic substitution and single-electron transfer (SET) by the phenylium ion in the gas phase: characterization of a long-lived SET intermediate.

Gas-phase mass spectrometric studies and calculations were performed for the reaction of naked phenylium ion with several benzene halides. From these reactions, the molecular ion for biphenyl as the predominant product was obtained only from the reaction of phenylium ions with iodobenzene and bromobenzene. Furthermore, through the collision-induced dissociation (CID) of the ion at m/z 281, the only dissociation observed is the loss of a phenyl radical, which indicates that a single-electron transfer (SET) mechanism might have occurred within the reaction. Additionally, according to the comparison between the CID experiments of those isomeric compounds of the sigma-complexes and the CID experiment of the ion at m/z 281 captured in the ion trap, we have also defined the captured ion at m/z 281 as an SET-intimate ion pair rather than those of sigma-complexes or the diphenyliodonium.

Monitoring enzyme reaction and screening of inhibitors of acetylcholinesterase by quantitative matrix-assisted laser desorption/ionization Fourier transform mass spectrometry.

A matrix-assisted laser desorption/ionization Fourier transform mass spectrometry (MALDI-FTMS)-based assay was developed for kinetic measurements and inhibitor screening of acetylcholinesterase. Here, FTMS coupled to MALDI was applied to quantitative analysis of choline using the ratio of choline/acetylcholine without the use of additional internal standard, which simplified the experiment. The Michaelis constant (K(m)) of acetylcholinesterase (AChE) was determined to be 73.9 micromol L(-1) by this approach. For Huperzine A, the linear mixed inhibition of AChE reflected the presence of competitive and noncompetitive components. The half maximal inhibitory concentration (IC(50)) value of galantamine obtained for AChE was 2.39 micromol L(-1). Inhibitory potentials of Rhizoma Coptidis extracts were identified with the present method. In light of the results the referred extracts as a whole showed inhibitory action against AChE. The use of high-resolution FTMS largely eliminated the interference with the determination of ACh and Ch, produced by the low-mass compounds of chemical libraries for inhibitor screening. The excellent correlation with the reported kinetic parameters confirms that the MS-based assay is both accurate and precise for determining kinetic constants and for identifying enzyme inhibitors. The obvious advantages were demonstrated for quantitative analysis and also high-throughput characterization. This study offers a perspective into the utility of MALDI-FTMS as an alternate quantitative tool for inhibitor screening of AChE.

High-sensitivity matrix-assisted laser desorption/ionization Fourier transform mass spectrometry analyses of small carbohydrates and amino acids using oxidized carbon nanotubes prepared by chemical vapor deposition as matrix.

In matrix-assisted laser desorption/ionization (MALDI) Fourier transform mass spectrometry (FTMS) analyses of small oligosaccharides and amino acids, high sensitivities for oligosaccharides (10 fmol) were obtained by introducing oxidized carbon nanotubes (CNTs) with short and open-end structure as valuable matrix. The CNTs were deposited in porous anodic alumina (PAA) templates by chemical vapor deposition. Transmission electron microscopy (TEM) images show that those CNTs include low levels of amorphous carbon. Thus, the background interference signals generally caused by amorphous carbon powder in CNTs can be reduced effectively. Experiments also confirmed that the FTMS signal intensity of CNTs prepared in PAA template is much lower than that of commercial multi-wall carbon nanotubes (MCNTs). Moreover, the purified process for CNTs with mixed acid (H2SO4 and HNO3) also contributed to the minimization of background. Intense signals corresponding to alkali cation adduct of neutral carbohydrates and amino acids have been acquired. In addition, reliable quantitative analyses for urine and corn root were also achieved successfully. The present work will open a new way to the application of oxidized CNTs as an effective matrix in MALDI MS research.

Study of the coupling reaction of pyridine in the gas phase.

The coupling reaction of pyridine in the gas phase to form bipyridyl and terpyridyl has been studied by electron ionization using an ion trap mass spectrometer. In contrast to the difficulty in carrying out electrophilic substitutions at carbon atoms in the pyridine ring under highly acidic solvent conditions, reactions in the gas phase overcame the conjugate acidification of pyridine in the solvent phase, thus decreasing the hardness of this electrophilic coupling. Through product ion mass spectra of the ion at m/z 157, we have shown that this ion was protonated bipyridyl rather than the ion/molecule adduct. A computational study of the heat of formation surface also supported the formation of polypyridyls through the electrophilic substitution of pyridine. We have confirmed the reaction through a study of pyridine-d(5) coupling in the gas phase.