Determination of tetrahydrothiophene formation as a probe of in vitro busulfan metabolism by human glutathione S-transferase A1-1: use of a highly sensitive gas chromatographic-mass spectrometric method.
A method for the sensitive determination of tetrahydrothiophene (THT) in cytosolic incubation mixtures was developed. Busulfan conjugation with glutathione was predominantly catalysed by glutathione S-transferase A1-1 (GST A1-1) and THT was released from the primary metabolite by alkalization. After liquid-liquid extraction using n-pentane separation and quantification of the product was performed by gas chromatography with a mass-selective detector. The method showed good sensitivity, accuracy and reproducibility with a detection limit of 2 ng ml(-1) and a limit of quantification of 5 ng ml(-1). The suitability of the method is shown for enzyme kinetic studies in human liver cytosol as well as for determination of GST A1-1 activity.
Ritter
C A
CA
Institut für Pharmakologie, Ernst-Moritz-Arndt-Universität, Greifswald, Germany.
Bohnenstengel
F
F
Hofmann
U
U
Kroemer
H K
HK
Sperker
B
B
D018906:Antineoplastic Agents, Alkylating / Q000378:metabolism*; D015415:Biomarkers / Q000032:analysis; D002066:Busulfan / Q000378:metabolism*; D002138:Calibration; D008401:Gas Chromatography-Mass Spectrometry / Q000379:methods*; D005982:Glutathione Transferase / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D008099:Liver / Q000201:enzymology / Q000378:metabolism; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D013876:Thiophenes / Q000494:pharmacology*
D018906:Antineoplastic Agents, Alkylating; D015415:Biomarkers; D013876:Thiophenes; C121030:tetrahydrothiophene; D005982:Glutathione Transferase; D002066:Busulfan