Differentiation of the anomeric configuration and ring form of glucosyl-glycolaldehyde anions in the gas phase by mass spectrometry: isomeric discrimination between m/z 221 anions derived from disaccharides and chemical synthesis of m/z 221 standards.

Mass spectrometry of disaccharides in the negative-ion mode frequently generates product anions of m/z 221. With glucose-containing disaccharides, dissociation of isolated m/z 221 product ions in a Paul trap yielded mass spectra that easily differentiated between both anomeric configurations and ring forms of the ions. These ions were shown to be glucosyl-glycolaldehydes through chemical synthesis of their standards.

A phase I and pharmacokinetic study of silybin-phytosome in prostate cancer patients.

Silibinin is a polyphenolic flavonoid isolated from milk thistle with anti-neoplastic activity in several in vitro and in vivo models of cancer, including prostate cancer. Silybin-phytosome is a commercially available formulation containing silibinin. This trial was designed to assess the toxicity of high-dose silybin-phytosome and recommend a phase II dose.

The bioreduction of a series of benzoquinone ansamycins by NAD(P)H:quinone oxidoreductase 1 to more potent heat shock protein 90 inhibitors, the hydroquinone ansamycins.

We have previously evaluated the role of NAD(P)H:quinone oxidoreductase 1 (NQO1) in the bioreductive metabolism of 17-(allylamino)-demethoxygeldanamycin (17AAG) to the corresponding hydroquinone, a more potent 90-kDa heat shock protein (Hsp90) inhibitor. Here, we report an extensive study with a series of benzoquinone ansamycins, which includes gel-danamycin, 17-(amino)-17-demethoxygeldanamycin, and 17-demethoxy-17-[[2-(dimethylamino)ethyl]amino]-geldanamycin. The reduction of these benzoquinone ansamycins by recombinant human NQO1 to the corresponding hydroquinone ansamycins was monitored by high-performance liquid chromatography (HPLC) and confirmed by liquid chromatography/mass spectrometry.

Tissue distribution and metabolism of the tyrosine kinase inhibitor ZD6474 (Zactima) in tumor-bearing nude mice following oral dosing.

ZD6474 [N-(4-bromo-2-fluorophenyl)-6-methoxy-7-[(1-methylpiperidin-4-yl)methoxy]-quinazolin-4-amine; Zactima] is a tyrosine kinase inhibitor with antiangiogenic and antitumor activity currently undergoing human trials for cancer treatment. Pharmacokinetic studies in animal models are an important component in the clinical development of this agent to relate preclinical studies to patient treatment. In the studies presented here, the pharmacokinetics of ZD6474 was determined in plasma and tissues of MCF-7 tumor-bearing nude mice following single p.

Protonation of crotonyl-CoA dienolate by human glutaryl-CoA dehydrogenase occurs by solvent-derived protons.

The protonation of crotonyl-CoA dienolate following decarboxylation of glutaconyl-CoA by glutaryl-CoA dehydrogenase was investigated. Although it is generally held that the active sites of acyl-CoA dehydrogenases are desolvated when substrate binds, recent evidence has established that water has access to the active site in these binary complexes of glutaryl-CoA dehydrogenase. The present investigation shows that the dehydrogenase catalyzes (a) a rapid exchange of C-4 methyl protons of crotonyl-CoA with bulk solvent and (b) protonation of crotonyl-CoA dienolate by solvent-derived protons under single turnover conditions.

Rapid and sensitive LC/MS/MS analysis of the novel tyrosine kinase inhibitor ZD6474 in mouse plasma and tissues.

ZD6474 (N-(4-Bromo-2-fluorophenyl)-6-methoxy-7-[(1-methylpiperidin-4-yl)methoxy] quinazolin-4-amine) is a tyrosine kinase inhibitor with anti-angiogenic and anti-tumor activity that is currently undergoing human trials for cancer treatment. Pharmacokinetic studies in animal models are an important component in clinical development of this agent to relate pre-clinical models to patient treatment. A liquid chromatography tandem mass spectrometry method was developed for the determination of ZD6474 levels in mouse plasma and tissues.

Analysis of docetaxel pharmacokinetics in humans with the inclusion of later sampling time-points afforded by the use of a sensitive tandem LCMS assay.

Purpose: Docetaxel is a semisynthetic taxane derived from the needles of the European yew ( Taxus baccata) and it is an important chemotherapeutic agent in the treatment of recurrent ovarian, breast and non-small-cell lung cancers. Traditional dosing regimens with docetaxel involve doses of 60-100 mg/m(2) by infusion every 3 weeks. Now weekly low-dose (30-36 mg/m(2)) regimens are being evaluated in phase I trials.

Electrospray ionization and low energy tandem mass spectrometry of polyhydroxy unsaturated fatty acids.

Negative ion electrospray ionization, fast-atom bombardment, and low energy tandem mass spectrometry were used for the analysis of dihydroxy-eicosatrienoic acids, which contain a vicinol diol and three nonconjugated double bonds, dihydroxy-eicosatetraenoic acids, which contain a conjugated triene structure, and trihydroxy-eicosatetraenoic acids, which contain a vicinol diol and a conjugated tetraene structure. In general, the product ion spectra were qualitatively similar for both modes of ionization, but electrospray ionization was strikingly more efficient in generation of abundant carboxylate anions that could be collisionally activated to yield product ion spectra. Collision-induced dissociation fragmentation mechanisms were described generally by α-hydroxy fragmentations directed by relative positions of double bonds and were consistent with stable isotope labeling studies.

Negative ion electrospray tandem mass spectrometric structural characterization of leukotriene B4 (LTB 4) and LTB 4-derived metabolites.

The low energy collision induced dissociation (CID) of the carboxylate anions generated by electrospray ionization of leukotriene B4 (LTB4) and 16 of its metabolites was studied in a tandem quadrupole mass spectrometer. LTB4 is a biologically active lipid mediator whose activity is terminated by metabolism into a wide variety of structural variants. The collision-induced dissociation spectra of the carboxylate anions revealed structurally informative ions whose formation was determined by the position of hydroxyl substituents and double bonds present in the LTB4 metabolite.

Analysis of hydroxy fatty acids as pentafluorobenzyl ester, trimethylsilyl ether derivatives by electron ionization gas chromatography/mass spectrometry.

Electron ionization (EI) gas chromatography/mass spectrometry (GC/MS) analysis of pentafluorobenzyl ester-trimethyl sllyl ether (PFB-TMS) derivatives of hydroxy-subshtuted fatty acids provides structural information comparable to that obtained in analysis of methyl ester-trimethyl silyl ether (Me-TMS) derivatives. Use of this derivative eliminates the need to prepare two separate derivatives, the PFB-TMS derivative for molecular weight determination by electron capture ionization (negative ions) analysis and the Me-TMS derivative for structural determination by EI GC/MS analysis. The relative abundance of fragment ions observed during EI GC/MS analysis of these derivatized unsaturated fatty acids indicates the location of the -OTMS substituents relative to double bond positions in those cases studied.

Analysis of long-chain fatty acyl coenzyme a thioesters by negative ion fast-atom bombardment mass spectrometry and tandem mass spectrometry.

Long-chain acyl Coenzyme A (CoA) is essentially composed of three major chemical groups, fatty acyl-, phosphopantetheino-, and 3', 5',-adenosine diphospho-moieties. The negative ion fast-atom bombardment mass spectrometry spectra of long-chain acyl CoA thioesters were characterized by the formation of abundant [M - H](-) and two distinct classes of fragment ions, one class which retained the acyl group and another class which is related to CoA that contains the phosphopantethene and adenine. The ions which retained the acyl group in the spectrum of palmitoyl CoA appeared at m/z 675, 657, 595, and 577 and were found to decompose by loss of alkylketene observed at m/z 357 and 339.

Metabolism of leukotriene B4 by cultured human keratinocytes. Formation of glutathione conjugates and dihydro metabolites.

Six previously unidentified leukotriene (LT) B4 metabolites formed during incubation of LTB4 with human keratinocytes in primary culture indicate the importance of the 12-hydroxyeicosanoid dehydrogenase pathway in LTB4 metabolism. The ultraviolet absorption spectra obtained for all keratinocyte metabolites revealed the presence of a conjugated diene structural moiety rather than the conjugated triene structure of LTB4. Metabolites were characterized using fast atom bombardment-mass spectrometry, gas chromatography-mass spectrometry of the pentafluorobenzyl ester, trimethylsilyl ether derivatives and specific degradation reactions.

Low-energy fast atom bombardment tandem mass spectrometry of monohydroxy substituted unsaturated fatty acids.

The low-energy collision-induced dissociation (CID) of the carboxylate anions generated by fast atom bombardment ionization of monohydroxy unsaturated fatty acids derived from oleic, linoleic, linolenic and arachidonic acids were studied in a tandem quadrupole mass spectrometer. The collisional activation spectra revealed structurally informative ions as to the position of the hydroxyl substituent in relationship to the sites of unsaturation. Five mechanisms are proposed for the fragmentation of hydroxyl substituted unsaturated fatty acids and are dependent upon the presence of alpha- or beta-unsaturation sites.

Low-energy tandem mass spectrometry of the molecular ion derived from fatty acid methyl esters: A novel method for analysis of branched-chain fatty acids.

Low-energy collision-induced dissociation (CID) of the molecular ions of fatty acid methyl esters obtained by electron ionization (70 eV) decompose in the tandem quadrupole mass spectrometer to yield a regular homologous series of carbomethoxy ions. Even at energies up to 200 eV (E lab), primarily carbomethoxy ions are present, with the most abundant found at m/z 101 at hi her energies. The lack of any other CID ions, including m/z 74 (McLafferty rearrangement) or m/z 87, suggest a rearranged molecular ion structure on leaving the first quadrupole mass analyzer.

Collision-induced dissociation of carboxylate anions from derivatized 5-lipoxygenase metabolites of arachidonic acid.

The low-energy collision-induced dissociations (CID) of carboxylate anions derived from pentafluorobenzyl ester, trimethylsilyl ether derivatives of four arachidonic acid metabolites of the 5-lipoxygenase pathway have been determined. These molecules include leukotriene B4 (LTB4), a potent chemotactic factor for the human neutrophil; 20-carboxy-LTB4, an inactive metabolite; 5-hydroxyeicosatetraenoic acid (5-HETE), a useful marker of 5-lipoxygenase activity within cells; and 5-hydroxyeicosanoic acid (5-HEA), which has been previously used for the quantitation of leukotriene E4. The carboxylate anion of 5-HEA (m/z 399) was found to decompose by the loss of trimethylsilanol as well as the loss of 146 u corresponding to the loss of trimethylsilanol followed by acrolein, a process specific for 5-hydroxy-containing saturated fatty acids.

General anesthetics potentiate gamma-aminobutyric acid actions on gamma-aminobutyric acidA receptors expressed by Xenopus oocytes: lack of involvement of intracellular calcium.

Potentiation of the gamma-aminobutyric acid (GABAA) receptor-gated Cl- channel response has been suggested to be a primary action of some anesthetic agents. We asked whether the GABAA receptor is a target site common for general anesthetics that are chemically and structurally diverse. This hypothesis was tested in Xenopus oocytes expressing mouse cortical mRNA, and GABA-activated Cl- currents were measured using two-electrode voltage clamping.

Analysis of glycerophosphocholine molecular species as derivatives of 7-[(chlorocarbonyl)-methoxy]-4-methylcoumarin.

A method has been developed for the analysis of derivatized diradylglycerols obtained from glycerophosphocholine (GPC) of transformed murine bone marrow-derived mast cells that provided high performance liquid chromatography (HPLC) separation of GPC subclasses and molecular species separation with on-line quantitation using UV detection. In addition, the derivatized diradylglycerol species were unequivocably identified by continuous flow fast-atom bombardment mass spectrometry. GPC was initially isolated by thin-layer chromatography (TLC), the phosphocholine group was hydrolyzed, and the resultant diradylglycerol was derivatized with 7-[(chlorocarbonyl)-methoxy]-4-methylcoumarin (CMMC).

Tandem mass spectrometry of negative ions from choline phospholipid molecular species related to platelet activating factor.

Fast atom bombardment mass spectrometry of choline phospholipids produces negative ions characteristic of the intact molecule and tandem mass spectrometry of collision-induced decomposition of M-15 anions characterizes both the identity and substituent position of radyl groups. Certain choline phospholipid molecular species which may be of special interest in the generation of platelet activating factor contain a highly unsaturated fatty acyl substituent at sn-2 and an ether radyl group at sn-1; other choline phospholipid molecular species which contain esterified arachidonic acid are of interest as potential sources of arachidonate for eicosanoid biosynthesis. Collisional activated decomposition of 1-hexadecanoyl-2-arachidonoyl-sn-glycero-phosphocholine produce abundant carboxylate anions at m/z 303 (arachidonate) and m/z 255 (hexadecanoate) in a ratio of 3:1, diagnostic for the sn-2 arachidonoyl position.

Stereospecific gas chromatographic/mass spectrometric assay of the chiral labetalol metabolite 3-amino-1-phenylbutane.

We previously identified 3-amino-1-phenylbutane (APB) as an oxidative N-dealkylated, metabolite of the antihypertensive agent labetalol. Labetalol has two asymmetric centers and is used clinically as a mixture of the four possible stereoisomers; APB has one asymmetric center. We now report an enantiospecific gas chromatographic/mass spectrometric assay for APB in urine.

Metabolism of 12(R)-hydroxy-5,8,10,14-eicosatetraenoic acid (12(R)-HETE) in corneal tissues: formation of novel metabolites.

12(R)-Hydroxy-5,8,10,14-eicosatetraenoic acid [12(R)-HETE], a cytochrome P450 arachidonate metabolite, is metabolized by corneal tissues via three distinct metabolic pathways: beta-oxidation, omega-hydroxylation, and keto-reduction. The major metabolite released from the intact rabbit corneal epithelium or cultured cells was identified by mass spectrometric analysis as 8-hydroxy-4,6,10-hexadecatrienoic acid, the tetranor metabolite derived following two steps of beta-oxidation from the carboxy terminus. The beta-oxidation pathway was expressed in both microsomes and mitochondria isolated from bovine corneal epithelium and was dependent on the addition of oxidizing equivalents.

Negative ion tandem mass spectrometry of leukotriene E4, and LTE 4, metabolites: Identification of LTE 4, in human urine.

The sulfidopeptide leukotrienes, leukotriene E4, (LTE4,) and its N-acetyl derivative and several ω- and β-oxidized metabolites of LTE4, have been analyzed by tandem mass spectrometry. [M-H](-) ions were produced by continuous flow fast atom bombardment, and collision-induced dissociation of these ions was studied by using a triple quadrupole instrument. The product ion spectra obtained were characteristic of the structure of LTE4, and mechanisms of ion formation were investigated by using deuterated compounds.

Fast atom bombardment and collision-induced dissociation of prostaglandins and thromboxanes: Some examples of charge remote fragmentation.

The mass spectra of products found by collisional activation of selected prostaglandins and thromboxanes were studied by tandem mass spectrometry as barium carboxylate salts and as carboxylate anions. Collision-induced dissociation (CID) of these closed shell ions generated by fast atom bombardment mass spectrometry reveals a wealth of structural information for these hydroxy acids. Decomposition reactions were found to be dependent upon the eicosanoid ring structure and the type of ion being studied, either positive or negative ion.