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J. Mass Spectrom. Soc. Jpn., 46(3), 157-172 (in Japanese), 1998
Deuterium substitution of certain hydrogen atoms in a molecule may change its metabolism rate in various biological systems. Some of the deuterium labeled pesticides were metabolized in some in vitro systems, such as rat liver microsomes and purified P450 reconstituted systems, in a slower rate than the unlabeled counterpart. This sometimes caused a substantial bioactivity - insecticidal activity, etc.- enhancement. Metabolic reactions of lindane, DDT analogs, diuron analogs and methoxychlor are discussed. Measurement of isotope effect values would provide us various information on the biological and biochemical reactions. There are examples of metabolic reaction mechanism studies of oxidative O-demethylations of methoxy-compounds, oxidative N-demethylations of N-methylamine derivatives and benzene ring hydroxylations using specifically deuteriated aromatic compounds. These studies revealed some details of relevant metabolic reaction pathways, the information that cannot be obtained by other methods. Analysis of very large isotope effect values observed in some metabolic reactions of xenobiotics, such as indene derivatives, revealed a branched metabolic pathway. Also, a tunnel effect in metabolic reactions was proposed for some xenobiotic metabolism that exhibited a very large observed isotope effect value, for example metabolic oxygenation of linoleic acid and methane. Mass spectrometric analysis is an essential means in all these studies.
J. Mass Spectrom. Soc. Jpn., 46(3), 173-177 (in Japanese), 1998
Chemical and ecological investigations on the bioactive constituents of Vernonia amygdalina (Compositae) were carried out as a case study of the medicinal plant-use of chinpanzee in the wild. The sesquiterpene lactones and steroid related compounds were isolated as the constituents of biological significance. Based on the in vitro activity-tests together with chimpanzee's behavioral analyses, the steroid related constituents were hypothesized to be used for his self-medication to parasite related diseases. Further medicinal plants possibly used by chimpanzee are reviewed.
J. Mass Spectrom Soc. Jpn., 46(3), 178-183 (in Japanese), 1998
Among various methods, hyphenated mass spectrometry combined with gas chromatography and liquid chromatography has been well recognized as a powerfull tool for the separatory determination of biologically active substances. A review is presented of the clean-up methods for sensitive determinataion of drugs and their metabolites in complex matrics such as blood and urine. The state of the art of solid-phase extraction using traditional sorbents and immunoaffinity extraction by an immobilized antobody column as well as chromatographic separation by lipophilic ion-exchange gel are discussed.
J. Mass Spectrom. Soc. Jpn., 46(3), 184-190 (in Japanese), 1998
The aim of drug metabolism study is to elucidate the phenomena associated with the efficacy or the toxicity of a drug by tracing its biotransformation after administration to an organism. The structure determination of metabolites in plasma, urine and bile and the clarification of metabolic pathways have been considered as the important tasks indispensabe to the development of a new drug. However the structure determination is not a final object, but the investigation of pharmacological activity of determined metabolites and species differences in metabolism is important for application of the drug candidate to human. In particular, mass spectrometry (MS) has played a major role in this field. This study expalined two examples of such elucidation of chemical structure of metabolites and metabolic pathways using MS. In the structural elucidation of metabolites, the most suitable ionization method should be selected for each drug and metabolite from various ionization modes of EI, FAB and ESI.
J. Mass Spectrom. Soc. Jpn., 46(3), 191-195 (in Japanese), 1998
In the development of a drug candidate, radioisotope (RI) technique has been extensively used for investigating the absorption, distribution, metabolism, and excretion using experimental animals in preclinical stage due to detection with excellent specificity. However, the use of RI is limited for the protection of the radiation. In contrast to RI, stable isotope (SI) is safe for the human body, and mass spectrometry is most suitable for the detection of SI. Moreover, the combination of the SI technique with mass spectrometry makes it possible to acquire more rapid and abundant information concerning the structure of the compound to be ienterest. Recently, the sensitivity of SI technique has been more improved by the remarkable advance of ionization interfaces in mass spectrometry. This paper describes an application of SI technique for quantitative analysis by mass spectrometry with regard to the measurement of fecal excretion ratio in rats by LC/MS/MS method utilizing a mixed double labeling technique with [14C] and [2H6] variants.
J. Mass Spectrom. Soc. Jpn., 46(3), 196-203 (in Japanese), 1998
Precise and rapid glyco-mapping of a mouse monoclonal immunoglobulin G2b (IgG2b) was carried out by liquid chromatography-electrospray ionization ion trap mass spectrometry/mass spectrometry (LC/ESI IT-MS/MS). It was possible to obtain spectra of minor glycopeptides with a quantity as low as 1.8 pmol. Reduced and carboxymethylated mouse antidansyl monoclonal IgG2b (RCM-IgG2b) was digested with Lys-C. Proteolytic peptides were subjected to capillary HPLC separation followed by analysis with an ion trap mass spectrometer. The structures of twelve different types of O-linked oligosaccharides attached to Thr-221AH in the hinge region and those of three major types of N-linked oligosaccharides attached to Asn-297H have been characterized.
J. Mass Spectrom. Soc. Jpn., 46(3), 204-210 (in Japanese), 1998
As a part of our search for functional natural molecules we have been investgating the extract of an edible fungi (bunashimeji). In this manuscript we describe structural elucidation of complex lipids and polyisoprenepolyols isolated from the mushuroom bunashimeji by FAB mass spectrometry.
J. Mass Spectrom. Soc. Jpn., 46(3), 211-218 (in Japanese), 1998
The detection method of chiral discrimination ability of some macrocycles toward amino acid ester hydrochlorides using FAB mass spectrometry has been explained in this paper. The FABMS enantiomer labeled (EL) guest method, which was recently proposed by Sawada et al., was an useful and facile technique to detect chiral discrimination ability of new chiral hosts. A host (H) was complexed with 1: 1 amino acid guest mixture of (R)-enantiomer (GR+) and a deuterium labeled (S)-enantiomer (GS-dn+). The enantioselectivity of the host for the guest was quantitatively estimated from the relative peak intensity (I(H-GR)+/I(H-GS-dn)+)=IRIS) values of the two host-guest complex ion peaks at a difference of m/z = n.
J. Mass Spectrom. Soc. Jpn., 46(3), 219-227 (in Japanese), 1998
A number of polynuclear ruthenium(II), rhodium(III), and cobalt(III) bipyridine complexes have been examined by electrospray ionization mass spectrometry (ESI-MS). ESI-MS is shown to be a powerful tool for the identification of polymetallic complexes and for detecting the contamination, because ESI mass spectra for the complexes showed a simple enough mass pattern for easy structural assignment. Multiply charged ions with the charge state from 8+ to 3+ were observed in the full scan ESI mass spectra of star-burst type tetranuclear complexes denoted by [Met(II)(bpy)2B]3RuX8 (Met = Ru, Os, and bpy = 2,2'-bipyridine, B = bridging ligands, X = ClO4-). Different charged ions were found to be generated by the combination of counterion loss and protonation/deprotonation at the proton site of the bridging ligand B. These ion intensities are qualitatively explained in terms of acidity of metal complexes depending upon bridging ligand structures and the charge of metal ions. Ions produced by a removal of ligands were not observed in the ESI spectra unlike the FAB spectra. For tetranuclear complexes, collision-induced dissociation (CID) of the multiply charged ions has made it clear that loss of one or more HX neutral is a major process for lower charge states, while complete fragmentation takes place for higher charge states and the fragments are based on a subunit group of [Met(II)(bpy)2B]2+. Insertion of formal O2- produced from ClO4- counterion was observed for the complexes without active protons even if low energy CID experiments, but not for the complexes with active protons. Advantages of ESI mass spectrometry are demonstrated for these polynuclear compounds and CID chemistry of the investigated ions will be discussed.
J. Mass Spectrom. Soc. Jpn., 46(3), 228-234 (in Japanese), 1998
In contrast to low energy CID, high energy CID produces the characteristic fragmentations which yield to give a wealth of structural information. The side-chain cleavage of the C-terminal amino acid of an an ion leads to a dn ion, which is useful for the differentiation of Leu and Ile in the analysis of peptide sequences. In the CID spectrum of [M+Na]+ for peptides even those lacking basic amino acid in the sequence, the [dn+ Na-H]+ ions allow for the differentiation of Leu and Ile, though observation of the corresponding [an+Na-H]+ ions, whereas no an ions were seen in its [M+H]+. The charge remote fragmentations indicate the location of branching points and substituents, as well as of double bonds of fatty acid without any derivatization. Although the charge remote fragmentations arise from closed-shell precursor ions, they show in the CID spectrum of radical cation, M+. of tocopherol, which reveal direct structural information about the isoprene chain. The MS/MS method has also been applied to the determination of the structure of natural products, such as pittosporumxanthin B1, which feature the partial structures of antheraxanthin and tocopherol, and to the differentiation of glucose and mannose.
J. Mass Spectrom. Soc. Jpn., 46(3), 235-239, 1998
Assays that combine liquid chromatography and tandem mass spectrometry (LC-MS/MS) for measurement of drug and metabolite concentrations in physiological specimens have proliferated dramatically in recent years. The popularity of LC-MS/MS for these applications is primarily due to the sensitivity and versatility offered by electrospray and atmospheric pressure chemical ionization and the specificity provided by tandem mass spectrometry. However, during the development and application of LC-MS/MS methods we have encountered a number of problems that adversely affected the precision and accuracy of the assays. These include: 1) electrospray formation of dimers which can cause nonlinear calibration curves when a co-eluting internal standard is used; 2) interference from unsuspected metabolites; 3) ion suppression causing a reduction in sensitivity; 4) "charging" of components of the mass analyzer which also can cause a reduction in sensitivity; and 5) difficulties in converting an assay from one matrix to another. Procedures for overcoming each of these problems are discussed in this report.
J. Mass Spectrom. Soc. Jpn., 46(3), 246-249, 1998
The method for observing molecular related ions of methyl hydroperoxyoleates (OMH) by atmospheric pressure chemical ionization (APCI) mass spectrometry (MS) was investigated. Since thermally labile OMH is decomposed in the APCI nebulizer and/or desolvation chamber heated to a high temperature, it is difficult to observe significant peaks of their protonated molecules. The addition of ammonia water to the mobile phase (ethanol) was quite effective to suppress the thermal decomposition of OMH. As the result the mass spectra exhibited the intense peaks of ammonium adduct ions of OMH, [M+NH4]+, at m/z 346. The nebulizer temperature at 170oC and the drift voltage at 170V were adequate for observing the molecular ion species of OMH.
J. Mass Spectrom. Soc. Jpn., 46(3), 250-254, 1998
Applicability of FAB high-energy-CID MS/MS to structural characterization of two types of tetraphenylporphyrin(TPP)-derivatives was examined. These tetraphenylporphyrin(TPP)-derivatives, 20-hydroxy-2,2-dimethylicosanoyl picket-fence porphyrinatoiron and 20-cholinephosphoryl-2,2-dimethylicosanoyl picket-fence porphyrinatoiron have four 2,2-dimethylicosanamido long-chains in common in their structures, which keep a balance between hydrophobic and hydrophilic properties. The structural difference between the two compounds is that the former has a hydroxyl group at the remote end of the 2,2-dimethylicosanamido long-chain, while the latter has a phosphocholine ester functional group at the remote-end. This structural difference gives rise to different features as well as some analogies in their FAB and the high-energy-CID MS/MS spectra. Differences and analogies in those spectra were also noted between the long-chain porphyrin derivatives we are describing here and the pivaloyl-type porphyrins we reported previously [T, Shibue, H. Kambe, H. Nishide, S. Akashi, N. Dohmae, Y. Ohashi, Eur. Mass Spectrom., 3, 291-299 (1997)]. By selecting a precursor ion at m/z 476, which corresponds to the 20-cholinephosphoryl-2,2-dimethylicosanoyl group, charge-remote fragmentation was observed. This observation reminded us our previous study on the imidazole-tailed picket-fence porphyrin fragmentaions.
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