Simultaneous GC-MS/MS measurement of malondialdehyde and 4-hydroxy-2-nonenal in human plasma: Effects of long-term L-arginine administration.

Here, we report the simultaneous derivatization and quantification of malondialdehyde (MDA) and 4-hydroxy-2-nonenal (HNE) in human plasma by GC-MS/MS using [1,3-H]-MDA (d-MDA) and [9,9,9-H]-HNE (d-HNE) as the internal standards, respectively. MDA, d-MDA, HNE and d-HNE were converted to their pentafluorobenzyl oximes (PFBOX) by pentafluorobenzyl hydroxylamine. Subsequently, the hydroxyl groups of the PFBOX of HNE and d-HNE were trimethylsilylated with N,O-bis(trimethylsilyl)trifluoroacetamide/1% trimethylchlorosilane.

GC-MS and GC-MS/MS measurement of ibuprofen in 10-μL aliquots of human plasma and mice serum using [α-methylo-H]ibuprofen after ethyl acetate extraction and pentafluorobenzyl bromide derivatization: Discovery of a collision energy-dependent H/D isotope effect and pharmacokinetic application to inhaled ibuprofen-arginine in mice.

GC-MS and GC-MS/MS methods were developed and validated for the quantitative determination of ibuprofen (d-ibuprofen), a non-steroidal anti-inflammatory drug (NSAID), in human plasma using α-methyl-H-4-(isobutyl)phenylacetic acid (d-ibuprofen) as internal standard. Plasma (10μL) was diluted with acetate buffer (80μL, 1M, pH 4.9) and d- and d-ibuprofen were extracted with ethyl acetate (2×500μL).

GC-MS and GC-MS/MS measurement of the cardiovascular risk factor homoarginine in biological samples.

L-Homoarginine (hArg) has recently emerged as a novel cardiovascular risk factor and to herald a poor prognosis in heart failure patients. Here, we report on the development and thorough validation of gas chromatography-mass spectrometry (GC-MS) and gas chromatography-tandem mass spectrometry (GC-MS/MS) methods for the quantitative determination of hArg in biological samples, including human plasma, urine and sputum. For plasma and serum samples, ultrafiltrate (10 µL; cutoff, 10 kDa) was used.

GC-MS/MS and LC-MS/MS studies on unlabelled and deuterium-labelled oleic acid (C18:1) reactions with peroxynitrite (O=N-O-O⁻) in buffer and hemolysate support the pM/nM-range of nitro-oleic acids in human plasma.

Oleic acid (cis-9,10-octadecenoic acid) is the most abundant monounsaturated fatty acid in human blood. Peroxynitrite (ONOO(-)) is a short-lived species formed from the reaction of nitric oxide (NO) and superoxide (O2(-)). Peroxynitrite is a potent oxidizing and moderate nitrating agent.

UPLC-MS/MS measurement of S-nitrosoglutathione (GSNO) in human plasma solves the S-nitrosothiol concentration enigma.

We developed and validated a fast UPLC-MS/MS method with positive electrospray ionization (ESI+) for the quantitative determination of S-nitrosoglutathione (GSNO) in human plasma. We used a published protocol for the inactivation of plasma γ-glutamyltransferase (γGT) activity by using the γGT transition inhibitor serine/borate and the chelator EDTA for the stabilization of GSNO, and N-ethylmaleimide (NEM) to block SH groups and to avoid S-transnitrosylation reactions which may diminish GSNO concentration. S-[(15)N]Nitrosoglutathione (GS(15)NO) served as internal standard.

A validated, rapid UPLC-MS/MS method for simultaneous ivabradine, reboxetine, and metoprolol analysis in human plasma and its application to clinical trial samples.

A recent clinical trial assessing human autonomic cardiovascular regulation applied pacemaker channel inhibition with ivabradine, norepinephrine transporter blockade with reboxetine, and beta-adrenoreceptor blockade with metoprolol. To verify patient adherence, we developed and validated a fast UPLC-MS/MS assay measuring all three compounds simultaneously. Deuterium-labeled drugs, d3-ivabradine, d5-reboxetine and d7-metoprolol, served as internal standards.

Glutathione promotes prostaglandin H synthase (cyclooxygenase)-dependent formation of malondialdehyde and 15(S)-8-iso-prostaglandin F2α.

Prostaglandin (PG) H synthases (PGHS) or cyclooxygenases (COX) catalyse the peroxidation of arachidonic acid (AA) to PGG(2) and PGH(2) which are further converted to a series of prostaglandins and thromboxane A(2). Here, we report that GSH promotes concomitant formation of the current oxidative stress biomarkers malondialdehyde (MDA) and 15(S)-8-iso-prostaglandin F(2α) from AA via PGHS. This illustrates an uncommon interplay of enzymatic and chemical reactions to produce species that are considered to be exclusively produced by free-radical-catalysed reactions.

Stable isotope gas chromatography-tandem mass spectrometry determination of aminoethylcysteine ketimine decarboxylated dimer in biological samples.

Aminoethylcysteine ketimine decarboxylated dimer (AECK-DD; systematic name: 1,2-3,4-5,6-7,8-octahydro-1,8a-diaza-4,6-dithiafluoren-9(8aH)-one) is a previously described metabolite of cysteamine that has been reported to be present in mammalian brain, urine, plasma, and cells in culture and vegetables and to possess potent antioxidative properties. Here, we describe a stable isotope gas chromatography-tandem mass spectrometry (GC-MS/MS) method for specific and sensitive determination of AECK-DD in biological samples. (13)C(2)-labeled AECK-DD was synthesized and used as the internal standard.

Stable-isotope dilution GC-MS method for ethanol in vapour ethanol and microdialysis systems based on carbonate-catalyzed extractive pentafluorobenzoylation.

Common ethanol detection methods are not applicable to cell culture media and microdialysates due to interference with medium constituents including amino acids and pH indicators. We present a novel GC-MS method for the accurate and precise analysis of ethanol in cell cultures and microdialysates. The method is based on the carbonate-catalyzed extractive pentafluorobenzoylation of ethanol and deuterium-labelled ethanol serving as the internal standard and on their GC-MS analysis in the electron-capture negative-ion chemical ionization mode.

Renal carbonic anhydrases are involved in the reabsorption of endogenous nitrite.

Nitrite (ONO(-)) exerts nitric oxide (NO)-related biological actions and its concentration in the circulation may be of particular importance. Nitrite is excreted in the urine. Hence, the kidney may play an important role in nitrite/NO homeostasis in the vasculature.

Measurement of nitrite in urine by gas chromatography-mass spectrometry.

Nitric oxide (NO) is enzymatically produced from L-arginine and has a variety of biological functions. Autoxidation of NO in aqueous media yields nitrite (O = N-O(-)). NO and nitrite are oxidized in erythrocytes by oxyhemoglobin to nitrate (NO(3)(-)).

¹⁸O-Labeled nitrous acid and nitrite: Synthesis, characterization, and oxyhemoglobin-catalyzed oxidation to ¹⁸O-labeled nitrate.

We describe a simple laboratory method for specific labeling of nitrite with ¹⁸O for use in chemical and biochemical studies in the area of nitric oxide research. NaNO₂ (0.1 mmol) is diluted in H₂¹⁸O (45 μl) and acidified with HCl (1 μl, 5 M), and the solution is allowed to equilibrate.

Measurement of 3-nitro-tyrosine in human plasma and urine by gas chromatography-tandem mass spectrometry.

Reaction of reactive nitrogen species (RNS), such as peroxynitrite and nitryl chloride with soluble tyrosine and tyrosine residues in proteins produces soluble 3-nitro-tyrosine and 3-nitro-tyrosino-proteins, respectively. Regular proteolysis of 3-nitro-tyrosino-proteins yields soluble 3-nitro-tyrosine. 3-Nitro-tyrosine circulates in plasma and is excreted in the urine.

Quantification of endocannabinoids in biological systems by chromatography and mass spectrometry: a comprehensive review from an analytical and biological perspective.

The endocannabinoids anandamide (arachidonoyl ethanolamide, AEA) and 2-arachidonoyl glycerol (2AG) are physiologically occurring, biologically active compounds on CB(1) and CB(2) receptors with multiple physiological functions. AEA and 2AG have been identified and quantified in many mammalian biological fluids and tissues, such as human plasma, adipocytes, tissues and tissue microdialysates, at concentrations in the picomolar-to-nanomolar range under basal conditions. In this article, recently published chromatographic and mass spectrometric analytical methods, i.

Simultaneous UPLC-MS/MS quantification of the endocannabinoids 2-arachidonoyl glycerol (2AG), 1-arachidonoyl glycerol (1AG), and anandamide in human plasma: minimization of matrix-effects, 2AG/1AG isomerization and degradation by toluene solvent extraction.

Analysis of the endocannabinoid (EC) system's key molecules 2-arachidonoyl glycerol (2AG) and arachidonoyl ethanolamide (anandamide, AEA) is challenging due to several peculiarities. 2AG isomerizes spontaneously to its biologically inactive analogue 1-arachidonoyl glycerol (1AG) by acyl migration and it is only chromatographically distinguishable from 1AG. Matrix-effects caused primarily by co-extracted phospholipids may further compromise analysis.

Quantification of acetaminophen (paracetamol) in human plasma and urine by stable isotope-dilution GC-MS and GC-MS/MS as pentafluorobenzyl ether derivative.

We report on the quantitative determination of acetaminophen (paracetamol; NAPAP-d(0)) in human plasma and urine by GC-MS and GC-MS/MS in the electron-capture negative-ion chemical ionization (ECNICI) mode after derivatization with pentafluorobenzyl (PFB) bromide (PFB-Br). Commercially available tetradeuterated acetaminophen (NAPAP-d(4)) was used as the internal standard. NAPAP-d(0) and NAPAP-d(4) were extracted from 100-μL aliquots of plasma and urine with 300 μL ethyl acetate (EA) by vortexing (60s).

In-source formation of N-acetyl-p-benzoquinone imine (NAPQI), the putatively toxic acetaminophen (paracetamol) metabolite, after derivatization with pentafluorobenzyl bromide and GC-ECNICI-MS analysis.

Pentafluorobenzyl (PFB) bromide (PFB-Br) is a versatile derivatization reagent for numerous classes of compounds. Under electron-capture negative-ion chemical ionization (ECNICI) conditions PFB derivatives of acidic compounds readily and abundantly ionize to produce intense anions due to [M-PFB](-). In the present article we investigated the PFB-Br derivatization of unlabelled acetaminophen (N-acetyl-p-aminophenol, NAPAP-d(0); paracetamol; MW 151) and tetradeuterated acetaminophen (NAPAP-d(4); MW 155) in anhydrous acetonitrile and their GC-ECNICI-MS behavior using methane as the buffer gas.

Simultaneous gas chromatography-tandem mass spectrometry quantification of symmetric and asymmetric dimethylarginine in human urine.

Previously, we demonstrated the utility of a gas chromatography-tandem mass spectrometry (GC-MS/MS) method for the quantitative determination of asymmetric dimethylarginine (ADMA) in biological samples. Here we report the extension of this method to symmetric dimethylarginine (SDMA) in human urine. SDMA and ADMA were simultaneously quantitated in urine by using their in situ prepared trideuteromethyl esters as internal standards.

Stable-isotope dilution GC-MS approach for nitrite quantification in human whole blood, erythrocytes, and plasma using pentafluorobenzyl bromide derivatization: nitrite distribution in human blood.

Previously, we reported on the usefulness of pentafluorobenzyl bromide (PFB-Br) for the simultaneous derivatization and quantitative determination of nitrite and nitrate in various biological fluids by GC-MS using their (15)N-labelled analogues as internal standards. As nitrite may be distributed unevenly in plasma and blood cells, its quantification in whole blood rather than in plasma or serum may be the most appropriate approach to determine nitrite concentration in the circulation. So far, GC-MS methods based on PFB-Br derivatization failed to measure nitrite in whole blood and erythrocytes because of rapid nitrite loss by oxidation and other unknown reactions during derivatization.

GC-MS determination of creatinine in human biological fluids as pentafluorobenzyl derivative in clinical studies and biomonitoring: Inter-laboratory comparison in urine with Jaffé, HPLC and enzymatic assays.

In consideration of its relatively constant urinary excretion rate, creatinine in urine is a useful biochemical parameter to correct the urinary excretion rate of endogenous and exogenous biomolecules. Assays based on the reaction of creatinine and picric acid first reported by Jaffé in 1886 still belong to the most frequently used laboratory approaches for creatinine measurement in urine. Further analytical methods for creatinine include HPLC-UV, GC-MS, and LC-MS and LC-MS/MS approaches.

Evidence by gas chromatography-mass spectrometry of ex vivo nitrite and nitrate formation from air nitrogen oxides in human plasma, serum, and urine samples.

Nitrite and nitrate in body fluids and tissues result from dietary source, endogenous nitric oxide (NO) production and from NO and its higher oxides (NO(x)) present as pollutants in the atmosphere. Nitrite and nitrate in human blood serum and plasma or urine are commonly used as biomarkers and measures of endogenous NO synthesis. In addition to dietary intake of nitrite and nitrate, our study indicates that NO(x) naturally present in the laboratory air may be an abundant source for nitrite and nitrate in human serum, plasma, and urine ex vivo.

Nitro-fatty acids occur in human plasma in the picomolar range: a targeted nitro-lipidomics GC-MS/MS study.

First studies on the occurrence of nitrated fatty acids in plasma of healthy subjects revealed basal concentrations of 600 nM for free/nonesterified nitro-oleic acid (NO(2)-OA) as measured by liquid chromatography tandem mass spectrometry (LC-MS/MS). We recently showed by a gas chromatography tandem mass spectrometry (GC-MS/MS) method the physiological occurrence of two isomers, i.e.

Specific GC-MS/MS stable-isotope dilution methodology for free 9- and 10-nitro-oleic acid in human plasma challenges previous LC-MS/MS reports.

Nitrated unsaturated fatty acids including nitro-oleic acid (NO(2)-OA) have been measured in human blood samples in their free and esterified forms. Plasma concentrations in healthy subjects have been reported to be of the order of 600 nM for free NO(2)-OA and 300 nM for esterified NO(2)-OA, as measured by LC-MS/MS. In the present article we report a GC-MS/MS method for the specific and accurate quantification of two NO(2)-OA isomers, i.

Application of a stable-isotope dilution technique to study the pharmacokinetics of human 15N-labelled S-nitrosoalbumin in the rat: possible mechanistic and biological implications.

In the year 1992, S-nitrosoalbumin (SNALB) has been proposed to be the most abundant physiological carrier and pool of nitric oxide (NO) activity in human circulation, by which NO-dependent biological functions are regulated. The concentration, the metabolism and the mechanisms of the biological actions of SNALB are controversial and still incompletely understood. Moreover, the suitability of SNALB as a biomarker of diseases associated with altered NO bioactivity in human circulation has not been demonstrated convincingly so far.