This Article Full Text Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by van Maanen, M. J. Articles by Beijnen, J. H. Search for Related Content PubMed PubMed Citation Articles by van Maanen, M. J. Articles by Beijnen, J. H.

A Search for New Metabolites of N,N',N''-Triethylenethiophosphoramide

Departments of Pharmaceutical Analysis [M. J. v. M., I. M. T., J. H. B.] and Biomolecular Mass Spectrometry [J. M. A. D., C. V., J. J. K. v. d. B., A. J. R. H.], University of Utrecht, 3584 CA Utrecht; The Netherlands Cancer Institute/Slotervaart Hospital, 1066 EC Amsterdam [M. J. v. M., J. H. B.]; and The Netherlands Cancer Institute/Antoni van Leeuwenhoek Hospital, 1066 CX Amsterdam [S. R.], the Netherlands

An attempt was made to unravel the metabolic profile of the alkylating agent N,N',N''-triethylenethiophosphoramide (thioTEPA). thioTEPA and its metabolite N,N',N''-triethylenephosphoramide (TEPA) were quantified in urine of treated patients by gas chromatography with selective nitrogen/phosphorous detection. Total alkylating activity was assessed by p-nitrobenzylpyridine reactivity. The total alkylating activity exceeded the amount of thioTEPA and TEPA, indicating the presence of other alkylating metabolites. Solid-phase extraction and liquid-liquid extractions followed by gas chromatography-mass spectrometry analysis revealed the conversion of an aziridinyl function of TEPA into a ß-chloroethyl moiety. This metabolite, N,N'-diethylene-N''-2-chloroethylphosphoramide, was quantified by gas chromatography with selective nitrogen/phosphorous detection and accounted for only 0.69% of the administered dose. Large volumes of urine were concentrated with solid-phase extraction and fractionated with high-performance liquid chromatography. Alkylating activity was determined for each 2-ml fraction and showed the presence of an alkylating compound eluting between 8 and 12 ml. The fractions with alkylating activity were collected, evaporated under a stream of nitrogen at room temperature to dryness, reconstituted in methanol, and subjected to fast atom bombardment-mass spectrometry and fast atom bombardment-tandem mass spectrometry. A new metabolite was found with a molecular mass of 352 Da, the same as that of thioTEPA-mercapturate. thioTEPA-mercapturate is likely the result of glutathione conjugation, after which the glutathione adduct loses two amino acid residues in separate stages. The fragmentation pattern and chromatographic properties of this new metabolite were identical to those of the reference, thioTEPA-mercapturate, which was obtained by incubation of thioTEPA with N-acetylcysteine at pH 11 and 95°C for 30 min. Quantification of thioTEPA-mercapturate was carried out by liquid chromatography-mass spectrometry. The thioTEPA-mercapturate levels in urine accounted for 12.3% of the administered dose and exceeded the amount of TEPA, which was previously assumed to be the main metabolite of thioTEPA. The total excreted amount of thioTEPA and its metabolites accounts for 54–100% of the total alkylating activity, indicating the presence of still other alkylating metabolites.

This article has been cited by other articles:

				A. D. R. Huitema, M. Spaander, R. A. A. Mathot, M. M. Tibben, M. J. Holtkamp, J. H. Beijnen, and S. Rodenhuis Relationship between exposure and toxicity in high-dose chemotherapy with cyclophosphamide, thiotepa and carboplatin Ann. Onc., March 1, 2002; 13(3): 374 - 384. [Abstract] [Full Text] [PDF]