Gas chromatography-tandem mass spectrometry analysis of gabapentin in serum

A new highly sensitive analytical method for determining gabapentin [1-(aminomethyl) cyclohexaneacetic acid; Neurontin] in serum using gas chromatography/tandem mass spectrometry (GC-MS/MS) was developed. GC-MS/MS was applied to determine the levels of gabapentin in serum samples of mice at 1 and 6 h after oral or intraperitoneal treatment (300 mg/kg). At 1 h, the concentrations of the drug were 4.02 +/- 0.42 and 4.32 +/- 0.28 microg/mL in mice treated orally and intraperitoneally, respectively. At 6 h, drug levels decreased by about 66% in both groups. The method, coupling two stages of mass analysis, could be very useful in identifying the drug in complex mixtures such as blood and urine. Moreover, it is easy and rapid to perform, and sensitive enough to allow the presence of the drug to be determined at very low detection limits. It is a very reliable method for both clinical and experimental monitoring of gabapentin.     

Development and application of a high-performance liquid chromatography-mass spectrometry method to determine alendronate in human urine

Despite the high potential offered by electrospray ionization on highly polar compounds like biphosphonates, few applications have been developed. High-performance liquid chromatography (HPLC) separation methods suitable for such molecules cannot be used in tandem with mass spectrometry (MS) due to high non-volatile salt content; at the same time the sample preparation, in biological fluids, is also a challenging problem. In the past ion-pair chromatography was mainly used in the case of HPLC-MS of biphosphonates, but no application to quantitative pharmacokinetic (PK) studies has been presented. In this study, after preliminary tests with ion-pair chromatography showing a poor sensitivity, a combined derivatization of the amino group and the biphosphonate has been developed and tested in a PK study. Using this analytical approach we were able to fully validate the quantitation of alendronate in the range of 6.667-4860.0 ng/ml in urine (sample volume 2.0 ml); each analytical run was 5.0 min long. The sensitivity achieved permitted a correct evaluation of the alendronate urinary excretion over the full period of urine collection. Sample preparation despite its complexity permitted to process and analyze up to 200 samples in a working day.     

Gas chromatography-mass spectrometry and high-performance liquid chromatography-tandem mass spectrometry in quantifying fatty acids

This critical overview covers current analytical methods and future developments in quantitative determination of fatty acids (FAs), emphasizing sample extraction, derivatization and instrumental analysis with gas chromatography/mass spectrometry (GC/MS) and high-performance liquid chromatography/tandem mass spectrometry (HPLC/MS²). We compare the benefits and the drawbacks of these two analytical techniques.We consider the well-established GC/MS method with pre-derivatization to be a traditional technique in terms of highly standardized sample-preparation procedures, affordability and readily available library searching for compound identification. However, the complicated derivatization steps required prior to instrumental analysis with GC/MS take a long time, with loss and transformation of FAs, low recovery and poor reproducibility.HPLC/MS² without derivatization shows the benefits of simple, mild sample-processing conditions, satisfactory recovery, short running time and high selectivity and sensitivity, which may allow it to become a viable alternative to GC/MS for the analysis of FAs in the years ahead.     

Quantitation of anabolic hormones and their metabolites in bovine serum and urine by liquid chromatography-tandem mass spectrometry

A specific and sensitive method based on tandem mass spectrometry with on-line high-performance liquid chromatography using atmospheric pressure chemical ionisation (LC–APCI-MS–MS) for the quantitation of anabolic hormone residues (17β-19-nortestosterone, 17β-testosterone and progesterone) and their major metabolites (17-19-nortestosterone and 17-testosterone) in bovine serum and urine is reported. [²H₂]17β-Testosterone was used as internal standard. The analytes were extracted from urine (following enzymatic hydrolysis) and serum samples by liquid–liquid extraction and purified by C₁₈ solid-phase extraction. Ionisation was performed in a heated nebulizer interface operating in the positive ion mode, where only the protonated molecule, [M+H]⁺, was generated for each analyte. This served as precursor ion for collision-induced dissociation and two diagnostic product ions for each analyte were identified for the unambiguous hormone confirmation by selected reaction monitoring LC–MS–MS. The overall inter-day precision (relative standard deviation) ranged from 6.37 to 2.10% and from 6.25 to 2.01%, for the bovine serum and urine samples, respectively, while the inter-day accuracy (relative error) ranged from −5.90 to −3.18% and from −6.40 to −2.97%, for the bovine serum and urine samples, respectively. The limit of quantitation of the method was 0.1 ng/ml for all the hormones in bovine serum and urine. On account of its high sensitivity and specificity the method has been successfully used to confirm illegal hormone administration for regulatory purposes.     

Combination of liquid chromatography/tandem mass spectrometry and gas chromatography/mass spectrometry for the detection of 21 anabolic steroid residues in bovine urine

For the detection of anabolic steroid residues in bovine urine, a highly sensitive liquid chromatographic/electrospray ionization tandem mass spectrometric (LC/ESI-MS/MS) method was developed using both positive and negative ionization. For four compounds the ESI mode was not sensitive enough and gas chromatographic/mass spectrometric GC/MS detection was therefore still necessary as a complementary method. The sample clean-up consisted of solid-phase extraction (SPE) on a C(18) column followed by enzymatic hydrolysis and a second solid-phase extraction on a combination of a C(18) and a NH(2) column. After this last SPE clean-up, the eluate was split into two equal aliquots. One aliquot was further purified and after derivatization used for GC/MS analysis. The other aliquot was analyzed with LC/MS/MS in both ESI+ and ESI- modes. The method was validated according to the European Commission Decision 2002/657/EC. Decision limits (CCalpha) were between 0.16 and 1 ng ml(-1) for the compounds detected with the LC/MS/MS method. The developed method is used in routine analysis in our laboratory.     

Detection of trace levels of trichothecene mycotoxins in human urine by gas chromatography-mass spectrometry

A method is described for the simultaneous detection of the trichothecene mycotoxins T-2, HT-2, T-2 tetraol, diacetoxyscirpenol, 15-monoacetoxyscirpendiol, scirpentriol, nivalenol and deoxynivalenol, in human urine. Samples were extracted from Clin Elut columns and cleaned up using reversed-phase Sep-Pak C18 cartridges. Trichothecenes were derivatised as their heptafluorobutyryl esters, and detected by gas chromatography-mass spectrometry-selected-ion monitoring using electron impact ionisation. The method was validated by the analysis of 22 urine samples, spiked and submitted "blind" for analysis by another laboratory. An alternative gas chromatography-mass spectrometry method using negative ion chemical ionisation is also described and a preliminary comparison of the two methods made. The methods enabled levels down to 1 ppb to be detected, with confirmation of identity at levels between 2 and 5 ppb, depending on the toxin.     

Determination of beta-agonist residues in bovine urine using liquid chromatography-tandem mass spectrometry

A multiresidue method was developed and validated to screen bovine urine samples for 10 beta-2-adrenergic agonistic drugs--brombuterol, cimaterol, clenbuterol, clenpenterol, isoxsuprine, mabuterol, ractopamine, ritodrine, salbutamol, and tulobuterol--at the 2 microg/L level. The method is also quantitative in the range of 1 to 4 microg/L for all analytes except salbutamol. The procedure uses enzymatic digestion, liquid-liquid extraction, and cleanup on solid-phase extraction columns, followed by detection using a liquid chromatograph-tandem quadrupole mass spectrometer operated in the positive-ion atmospheric pressure chemical ionization multiple-reaction monitoring mode. Method validation included assessment of recoveries, repeatabilities, linearity of responses, decision limits, and detection capabilities. Overall average recoveries ranged from 70-91%; recoveries were generally lower for salbutamol. The decision limits ranged from 0.4-1.0 microg/L, and detection capabilities from 0.6-1.7 microg/L.     

Determination of amino acid enantiomers in human urine and blood serum by gas chromatography-mass spectrometry

Amino acid (AA) enantiomers were determined as N(O)-pentafluoropropionyl-(2)-propyl esters by chiral gas chromatography-mass spectrometry (GC-MS) in 24 h samples of the urine of three healthy volunteers and in their blood sera. In urine the largest amounts were determined for D-Ser (64-199 micromol/day) and D-Ala (24-138 micromol/day). In blood sera, D-Ala (2.3-4.2 micromol/L) and D-Ser (1.0-2.9 micromol/L) were most abundant. Varying amounts of the D-enantiomers of Thr, Pro, Asx, Glx, Phe, Tyr, Orn and Lys were also found, albeit not in all urines and sera. Further, enantiomers were quantified in urine samples of two volunteers fasting for 115 h. Quantities of renally excreted D-AAs decreased in fasting, although amounts of D-Ser (69 and 77 micromol/L urine) as well as other D-AAs were still detectable. Time-dependent analyses of urine showed that D-AAs are continuously excreted. Copyright -Copyright 2001 John Wiley & Sons, Ltd.     

Simultaneous analysis of fourteen tertiary amine stimulants in human urine for doping control purposes by liquid chromatography-tandem mass spectrometry and gas chromatography-mass spectrometry

A method for the simultaneous screening and confirmation of the presence of fourteen tertiary amine stimulants in human urine by gas chromatography-mass spectrometry (GC-MS) in combination with liquid chromatography-tandem mass spectrometry (LC-MS/MS) has been developed and validated. Solid phase extraction (SPE) and liquid-liquid extraction (LLE) approaches were utilized for the pre-treatment of the urine samples. The study indicated that the capillary temperature played a significant role in the signal abundances of the protonated molecules of cropropamide and crotethamide under positive ion electrospray ionization (ESI) conditions. In addition, comparison studies of two different pre-treatment approaches as well as the two ionization modes were conducted. The LODs of the developed method for all the analytes were lower than the minimum required performance limit (MRPL) as set forth in the World Anti-Doping Agency (WADA) technical document for laboratories. The human urine sample obtained after oral administration of prolintane·HCl was successfully analyzed by the developed method, which demonstrated the applicability and reliability of the method for routine doping control analysis.     

Confirmatory analysis of residues of stanozolol and its major metabolite in bovine urine by liquid chromatography-tandem mass spectrometry

A reliable method for the confirmation of the synthetic hormone stanozolol and its major metabolite, 16beta-hydroxystanozolol, in bovine urine by liquid chromatography coupled with tandem mass spectrometry has been developed. [2H3]Stanozolol was used as internal standard. Sample preparation involved enzymatic hydrolysis, liquid-liquid extraction and purification on an amino solid-phase extraction column. The analytes were ionized using atmospheric pressure chemical ionization with a heated nebulizer interface operating in the positive ion mode, where only the protonated molecules, [M+H]+, at m/z 329 and m/z 345, for stanozolol and 16beta-hydroxystanozolol, respectively, were generated. These served as precursor ions for collision-induced dissociation and three diagnostic product ions for each analyte were identified for the unambiguous hormone confirmation by selected reaction monitoring liquid chromatography-tandem mass spectrometry. The accuracy ranged from 19.7 to 14.9% and from 18.9 to 13.2% for stanozolol and 16beta-hydroxystanozolol, respectively. The precision ranged from 12.4 to 2.4% and from 13.1 to 1.8% for stanozolol and 16beta-hydroxystanozolol, respectively. The limit of quantification of the method was 1 ng/ml in the bovine urine for both stanozolol and 16beta-hydroxystanozolol. The developed method fulfils the European Union requirements for confirmatory methods.     

Rapid and high-performance analysis of thyreostatic drug residues in urine using gas chromatography-mass spectrometry

A more sensitive method was developed using the hyphenated technique of gas chromatography-mass spectrometry (GC-MS) supplementary to the official high-performance thin-layer chromatography (HPTLC) method. Even combined with less efficient extraction and clean-up methods, GC-MS is able to lower the detection limit to less than 50 ppb. The powerful technique of GC-MS-MS is tried out to reduce the detection limit even more, in combination with simplified extraction methods. This time-saving approach combined with the increase in sensitivity is of great importance for a routine technique.     

超高效液相色谱-串联质谱法测定牛尿液中的甲状腺抑制剂(英文)

Five thyreostats(TSs),namely tapazole,thiouracil,methylthiouracil,propylthiouracil,and phenylthiouracil,were determined in bovine urine using ultra-high performance liquid chromatography-tandem mass spectrometry(UHPLC-MS/MS)in positive electrospray ionization mode.Extraction and clean-up were achieved using a ChemElut cartridge with tert-butyl methyl ether,without a derivatization step.Separation was achieved on an Acquity UPLC SS T3 column.The mobile phase was acetonitrile and water containing 0.2%(v/v)formic acid.The mass spectrometer was operated in multiple reaction monitoring mode.Urine samples were spiked with TS solution at levels corresponding to 5,10,15,and 20μg/L.The accuracy(internal standard corrected)ranged from 92%to 107%,with a repeatability precision(relative standard deviation,RSD)less than 15%for all five analytes.The RSDs within-laboratory reproducibility was less than 26%.The decision limits(CCα)and detection capabilities(CCβ)were obtained from a calibration curve and were in the ranges of 3.1-6.1μg/L and 4.0-7.4μg/L,respectively.The CCαand CCβvalues were below the recommended concentration,which was set at 10μg/L.The results show that the described method is suitable for the direct detection of TSs in bovine urine.This method can also be used to determine TSs in porcine urine.     

Gas chromatography-mass spectrometry and gas chromatography-tandem mass spectrometry of cyclic fatty acid monomers isolated from heated fats

The analysis of hydrogenated cyclic fatty acid monomers isolated from heated linseed and sunflower oils is achieved by gas chromatography-tandem mass spectrometry of their pentafluorobenzyl esters. Collisionally activated dissociation of the carboxylate anions produced by electron-capture ionization shows remote charge-site fragmentation that allows location of cyclopentane and cyclohexane rings by examining the resulting mass-analysed ion kinetic energy spectra. Oxidative ozonolysis of the methyl esters of the unsaturated cyclic fatty acid monomers allows location of some double bonds. However, preliminary results obtained with remote charge fragmentation of synthetic unsaturated models make this approach an alternative for double bond location in the cyclic fatty acid monomers isolated from heated fats.     

Determination of zopiclone in urine by gas chromatography-mass spectrometry

A procedure is described for the quantitative determination of zopiclone and the sum of its metabolites in urine using gas chromatography with the mass-spectrometric detection of their common hydrolysis product, 6-(5-chloro-2-pyridyl)-7-hydroxy-6,7-dihydro-5H-pyrrolo[3,4-b]pyrazine-5-on. The procedure is very sensitive. The detection limit for ions with a mass of 45–450 au detected in the full scanning mode is 70 ng/mL. The data of gas chromatography-mass spectrometry are presented for different derivatives of the hydrolysis product of zopiclone; these data can be used for the qualitative identification of zopiclone. The stability of zopiclone and its metabolites upon time was studied by analyzing urine samples from patients receiving therapeutic doses of this substance stored for 1, 3, and 6 months.     

Confirmatory and quantitative analysis using experimental design for the extraction and liquid chromatography-UV, liquid chromatography-mass spectrometry and liquid chromatography-mass spectrometry/mass spectrometry determination of quinolones in turkey muscle

The aim of this work is to established methods for determination of quinolones (ciprofloxacin, danofloxacin, enrofloxacin, difloxacin and flumequine), regulated by European Union, and sarafloxacin in turkey muscle. An experimental design has been applied for the optimization of the factors that influence the extraction of quinolones from turkey muscle in order to determine the experimental conditions for their extraction with high recoveries. Liquid chromatography with ultraviolet detection (LC-UV), liquid chromatography-mass spectrometry (LC-MS) and liquid chromatography tandem mass spectrometry (LC-MS/MS) have been used for the simultaneous quantification of quinolones antibiotics in turkey muscle. The proposed methods have been validated according to the Food Drugs Administration guideline and presents the limit of quantification below the maximum residue limits established by the European Union for quinolones in turkey muscle. The methods developed have been applied to quantification of enrofloxacin and its main metabolite ciprofloxacin in samples of turkey muscle obtained from animals treated with enrofloxacin.     

Comparison of microextraction procedures to determine pesticides in oranges by liquid chromatography-mass spectrometry

A liquid chromatographic–mass spectrometric method has been developed for the determination of bitertanol, carbendazim, fenthion, flusilazole, hexythiazox, imidacloprid, methidathion, methiocarb, pyriproxyfen and trichlorfon. Two procedures, based on stir bar sorptive extraction (SBSE) and matrix solid-phase dispersion (MSPD), have been evaluated for the extraction of these compounds in oranges. Their respective advantages and disadvantages are also discussed. The recoveries obtained by MSPD ranged from 47 to 96% and the relative standard deviations (RSDs) ranged from 1 to 15%, whereas with the SBSE method the recoveries were between 8 and 84% and the RSDs between 4 and 16%. Although, the limits of quantitation of most compounds are much better (0.001–0.05 mg kg⁻¹) by SBSE, it is not suitable to determine some polar pesticides as carbendazim, imidacloprid and trichlorfon. Results obtained by both methods were compared, in terms of sensitivity and selectivity, with a classical ethyl acetate extraction method, and the three methods were applied to analyze real samples. As MSPD is easier to perform, faster than the organic solvent extraction, and shows equal accuracy and resolution, its application for analyzing pesticides in oranges is recommended.