Rapid detection and identification of N-acetyl-L-cysteine thioethers using constant neutral loss and theoretical multiple reaction monitoring combined with enhanced product-ion scans on a linear ion trap mass spectrometer

Abstract

A sensitive and specific liquid chromatography-mass spectrometry (LC-MS) method based on the combination of constant neutral loss scans (CNL) with product ion scans was developed on a linear ion trap. The method is applicable for the detection and identification of analytes with identical chemical substructures (such as conjugates of xenobiotics formed in biological systems) which give common CNLs. A specific CNL was observed for thioethers of N-acetyl-L-cysteine (mercapturic acids, MA) by LC-MS/MS. MS and HPLC parameters were optimized with 16 MAs available as reference compounds. All of these provided a CNL of 129 Da in the negative-ion mode. To assess sensitivity, a multiple reaction monitoring (MRM) mode with 251 theoretical transitions using the CNL of 129 Da combined with a product ion scan (IDA thMRM) was compared with CNL combined with a product ion scan (IDA CNL). An information-dependent acquisition (IDA) uses a survey scan such as MRM (multiple reaction monitoring) to generate "informations" and starting a second acquisition experiment such as a product ion scan using these "informations." Th-MRM means calculated transitions and not transitions generated from an available standard in the tuning mode. The product ion spectra provide additional information on the chemical structure of the unknown analytes. All MA standards were spiked in low concentrations to rat urines and were detected with both methods with LODs ranging from 60 pmol/mL to 1.63 nmol/mL with IDA thMRM. The expected product ion spectra were observed in urine. Application of this screening method to biological samples indicated the presence of a number of MAs in urine of unexposed rats, and resulted in the identification of 1,4-dihydroxynonene mercapturic acid as one of these MAs by negative and positive product ion spectra. These results show that the developed methods have a high potential to serve as both a prescreen to detect unknown MAs and to identify these analytes in complex matrix. (J Am Soc Mass Spectrom 2005, 16, 1976 -1984

Quantifying the Metabolic Activation of Nevirapine in Patients by Integrated Applications of NMR and Mass Spectrometries

“…3B) that can be assigned to [M ϩ 1] ϩ of a mercapturate of NVP. At a higher cone voltage (70 V) this metabolite was found to produce a major fragment at m/z 299, which corresponds to the characteristic neutral loss of N-acetyldehydroalanine [129 atomic mass units (amu)] from a mercapturate (Scholz et al, 2005).…”

“…Neutral loss of 129 amu from parent anions is generally regarded as the generic fragmentation of mercapturates (Scholz et al, 2005), and thereby, as a ␤-elimination reaction, is obtained irrespective of the type of thioether linkage, i.e., (hetero) aryl, cycloalkyl, alkylaryl, and alkyl. Jian et al (2009) have developed LC/MS/MS screens for mercapturates based on the negative-ion neutral loss of 129 amu or MRM involving this loss as survey scans that trigger the acquisition of fragment-rich, positive-ion, product ion spectra.…”

“…6) and was derived from a difference between scissions of the thioether linkage. Whereas the prominent fragment at m/z 299 in the product ion spectrum of NVP-M2 corresponds to the characteristic ␤-elimination of N-acetyldehydroalanine (129 amu), NVP-M1 yielded an ion at m/z 265 that is attributed to neutral loss of NAC (163 amu) (Scholz et al, 2005). This retro-Michael elimination might be expected to be more favorable for an alky-linked thioether rather than an (hetero)aryl-linked thioether.…”

Specificity of Aminoacylase III-Mediated Deacetylation of Mercapturic Acids

“…cysteine, and N-acetyl-S-(4-methoxybenzyl)-L-cysteine were synthesized and characterized as described before Werner et al, 1997;Scholz et al, 2005)…”

Isolation and characterisation of 13 pterosins and pterosides from bracken (Pteridium aquilinum (L.) Kuhn) rhizome

“…Their presence in crude rhizome extracts, using MeOH and Et2O instead of chloroform, was confirmed by HPLC/ESI-MS n analysis. Both negative molecular ions (427 and 429 Da) of compounds 11, 13 and 35 showed a constant neutral loss of 162 Da to give ions at m/z 265 and 267, respectively, which corresponded to the loss of an O-linked hexose moiety (glucose) (Felipe et al, 2014;Levandi et al, 2014;Qu et al, 2004;Scholz et al, 2005) (Figs. SD1-4, Supplementary Data 2).…”