Signal enhancement of glucuronide conjugates in LC‐MS/MS by derivatization with the phosphonium propylamine cation tris(trimethoxyphenyl) phosphonium propylamine, for forensic purposes

Abstract

Although chemical derivatization for signal enhancement in drug testing is most often associated with gas chromatography, it also has the potential to improve the detection of analytes poorly ionized by atmospheric pressure ionization techniques, such as electrospray ionization used in liquid chromatography-mass spectrometry. A number of acidic compounds, namely drug glucuronides (e.g. conjugates of temazepam, oxazepam, lorazepam, morphine, testosterone, epitestosterone, 5-α-dihydrotestosterone, androsterone, p-nitrophenol, and paracetamol) were successfully derivatized with tris(trimethoxyphenyl) phosphoniumpropylamine to introduce a quaternary cation functionality to the analytes. Benzodiazepine glucuronides were more specifically investigated, and following positive mode electrospray ionization mass spectrometry, average improvements to peak areas as a result of derivatization were 67-, 6-, and 7- fold for temazepam, oxazepam, and lorazepam glucuronides. Average improvements to the signal-to-noise ratios for temazepam, oxazepam, and lorazepam glucuronides were 1336-, 371- and 217-fold, respectively. The values obtained for the derivatized conjugate were also typically higher than those for the underivatized parent drug. Urine containing benzodiazepine glucuronides was also successfully derivatized. The data indicates potential for the use of charge derivatization to improve the detection of molecules with acidic functionalities by liquid chromatography-mass spectrometry (LC-MS) techniques in certain scenarios.

Evaluation of the accuracy of protein quantification using isotope TMPP‐labeled peptides

“…Compared with the above‐mentioned labeling reagents, TMPP derivatization of peptides imparts some potentially favorable properties due to the introduction of a positively charged group, which increases hydrophobicity and retention time of peptides, improves ionizability, and simplifies the analysis of peptide fragmentation patterns. These properties have enabled the detection of N‐terminal peptides and the confirmation of translation initiation codons and translation start sites . TMPP‐labeled peptides have been selectively captured using anti‐TMPP antibodies coupled to magnetic beads, which was shown as a rapid immunoaffinity approach for the purification of N‐terminal peptides and improves yield .…”

The metabolic profile of the synthetic cannabinoid receptor agonist ADB-HEXINACA using human hepatocytes, LC–QTOF-MS and synthesized reference standards

“…This is an important consideration in clinical and forensic laboratories, as β-glucuronidase is typically used in urine analysis procedures to deconjugate any glucuronides formed during phase II metabolism, in order to increase phase I metabolite or parent drug concentration and hence aid detection ( 12 , 36 ). It should be noted that glucuronides are poorly ionized and undergo in-source fragmentation in ESI mass spectrometry, resulting in reduced signal ( 37–39 ). It is therefore possible that M2 (RT 5.21 min, m/z 551.2723), a mono-hydroxylated + glucuronidated hexyl tail product, is of higher concentration than its abundance would otherwise suggest.…”

Investigation into Small Molecule Isomeric Glucuronide Metabolite Differentiation Using In Silico and Experimental Collision Cross-Section Values

“…In addition, the difference between CCS values within isomer pairs was assessed to evaluate if the difference was sufficient for structural identification through in silico prediction. Although some isomer pairs cannot be distinguished through determination of their CCS values directly, several groups have derivatized metabolites including isomeric glucuronides , and hydroxyls with higher CCS value conjugates potentially providing supportive evidence for metabolites which cannot be purified in sufficient quantity for NMR analysis. The present study deals with isomers, that is ions with identical exact mass, not isobars where exact masses are different, but the nominal masses are the same .…”