The autoderivatization of 4‐ketocyclophosphamide during capillary gas chromatography

Abstract

The selectivity of a capillary gas chromatographic assay for the anti‐neoplastic and immunosuppressive agent cyclophosphamide (CPA) towards one of its naturally occurring metabolites, i.e. 4‐ketocyclophosphamide (4‐keto CPA), has been studied. Mass spectrometry studies showed that a cyclization product of 4‐keto‐CPA can form using the same chromatographic conditions as those under which CPA was determined. However, the amount produced is relatively small (less than 10%) compared with the percentage produced from CPA. Furthermore, both cyclization products, formed by loss of HCI and intraalkylation of the parent compounds, can be separated well under suitable chromatographic conditions.

Determination of ifosfamide by gas chromatography—mass spectrometry

“…The gas chromatography of oxazaphosphorines generally requires derivatization with trifluoroacetic or heptafluorobutyric anhydride, which is time-consuming and laborious [S-12]. In a series of papers, De Bruijn et al [4][5][6]131 demonstrated that oxazaphosphorines can be handled reliably in GC without derivatization by exploiting the oncolumn intra-alkylation of cyclophosphamide. In contrast, unchanged ifosfamide was not subject to intra-alkylation during GC and it was demonstrated that this drug could be determined by GC in the presence of its naturally occurring metabolites without any derivatization [4].…”

“…The determination of underivatized ifosfamide by GC appeared to be possible with detection limits at the pg level when electron capture detection (ECD) was used [4]. The higher sensitivity obtained for ifosfamide than for cyclophosphamide by GC-ECD can in part be explained by the absence of intra-alkylation; approximately half the signal for cyclophosphamide is lost because two peaks are observed [4][5][6]131. The concept was exploited in the present study.…”

“…without derivatization [4]. Mass spectrometry (MS) showed that intact ifosfamide (m/z 260) was eluted in the temperature range 150-300" C whereas an isomer of ifosfamide, cyclophosphamide underwent partial degradation under similar conditions [5,6]. Data from these studies, however, indicated the possibility of using cyclophosphamide as an internal or external (chromatographic conditions) standard because the formation of the degradation product, intra-alkylated cyclophosphamide, was linear over a sufficient range and did not interfere with the determination of ifosfamide [4-61.…”