Fragmentation of N‐oxides (deoxygenation) in atmospheric pressure ionization: Investigation of the activation process

Abstract

The diagnostic fragmentation of N-oxides resulting from loss of the oxygen atom (MH+ --> MH+-O) in electrospray and atmospheric pressure chemical ionization (APCI) mass spectra was investigated. When the temperature of the heated capillary tube was elevated, the ratio of the intensity of the [MH+ -16] fragment to the precursor ion (MH+) increased. This 'deoxygenation' process was associated with thermal activation and did not result from collisional activation in the desolvation region of the API source. Although the extent of 'deoxygenation' is compound-dependent, it can provide evidence for the presence of an N-oxide in a sample and can be used to distinguish N-oxides from hydroxylated metabolites (Ramanathan et al. Anal. Chem. 2000; 72: 1352). To demonstrate the practical application of thermal fragmentation of N-oxides, liquid chromatography (LC)/APCI-MS was used to distinguish an N-oxide drug from its hydroxylated metabolite in an unprocessed rat urine sample, despite the fact that the drug and its metabolite were not fully resolved by HPLC.

Flavin-Containing Monooxygenase Activity in Hepatocytes and Microsomes: In Vitro Characterization and In Vivo Scaling of Benzydamine Clearance

“…2). Whereas other metabolites were apparently formed from P450 and UGT activity, it is clear that It is well known that N-oxide metabolites can undergo in-source breakdown to the parent amine under some mass spectrometry conditions (Ramanathan et al, 2000;Tong et al, 2001). If this occurred with benzydamine N-oxide, it would complicate the determination of benzydamine lability and its quantitation in various matrices.…”

“…1) It is unknown if enterohepatic recirculation of benzydamine occurs in humans, although N-oxides and glucuronides are primary candidates for recirculation due to enzymes in gut flora with potent reductase and glucuronidase activity, respectively (Rowland and Tozer, 1995;Cashman, 1999). 2) The mass spectral source-induced and/or thermal deoxygenation of N-oxides is well documented and has recently been studied as a diagnostic tool for N-oxide structural assignment (Ramanathan et al, 2000;Tong et al, 2001). Mass spectral source-induced decomposition could easily result in misquantitation of N-oxide metabolite as unchanged parent compound in pharmacokinetic studies and thus an artificially increased area under the curve for parent and reduced clearance value.…”

An experimental and computational study on the dissociation behavior of hydroxypyridine N‐oxides in atmospheric pressure ionization mass spectrometry

“…[5] Therefore, a detailed understanding of the ionization and fragmentation of heterocyclic N-oxides would be very valuable. [6] While the hydroxypyridines have been extensively studied by mass spectrometry, [7,8] to our knowledge their N-oxides have thus far escaped examination.…”

Metabolism and Excretion of Capravirine, a New Non-Nucleoside Reverse Transcriptase Inhibitor, Alone and in Combination With Ritonavir in Healthy Volunteers

“…ϩ under atmospheric pressure chemical ionization (APCI) conditions (Ramanathan et al, 2000;Tong et al, 2001). This methodology was used to identify N-oxide metabolites of capravirine in humans.…”