RESULTS: Analytes were gradiently separated on a C18 column and detected with a Sciex API 4000 MS/MS with an ESI source operated in the positive ion mode with deuterated PQ as internal standard. The response was linear in the range 3.9-2508nM with a runtime of 7.0min per sample. The method was applied to clinical samples from healthy volunteers.
CONCLUSION: This LC-MS/MS method for the simultaneous quantitation of PQ and two of its metabolites in plasma may prove helpful for assessment of metabolite safety issues in vivo.
METHODS: The mass spectral characterization of the proposed NMs-GSH conjugates was performed with liquid chromatography high-resolution tandem mass spectrometry (LC-HRMS/MS). The final reaction mixtures were analysed in positive electrospray ionisation (ESI) at different incubation times.
RESULTS: This study identified three types of conjugates in addition to ethanolamines, the hydrolysis products of NMs. Monoglutathionyl, diglutathionyl and phosphorylated conjugates were produced for each of the NMs, bis(2-chloroethyl)ethylamine (HN1), bis(2-chloroethyl)methylamine (HN2) and tris(2-chloroethyl)amine (HN3). The monoglutathionyl conjugates consisted of HN1-GSH, HN2-GSH and HN3-GSH. The spontaneous and primary conjugates of diglutathionyl were HN1-GSH2, HN2-GSH2 and HN3-GSH2. These included phosphorylated conjugates, namely HN1-GSH-PO4 , HN2-GSH-PO4 and HN3-GSH-PO4 , as might have formed due to hydrolysis in phosphate buffer.
CONCLUSIONS: The mass spectral data of all conjugates formed in the presence of all NMs and GSH are reported in this study. These GSH metabolites can be used to confirm NMs toxicity in biological samples such as urine.
METHODS: Pooled urine samples of patients with BTG (n=10), patients with PTC (n=9) and healthy controls (n=10) were subjected to iTRAQ analysis and immunoblotting.
RESULTS: The ITRAQ analysis of the urine samples detected 646 proteins, 18 of which showed significant altered levels (p<0.01; fold-change>1.5) between patients and controls. Whilst four urinary proteins were commonly altered in both BTG and PTC patients, 14 were unique to either BTG or PTC. Amongst these, four proteins were further chosen for validation using immunoblotting, and the enhanced levels of osteopontin in BTG patients and increased levels of a truncated gelsolin fragment in PTC patients, relative to controls, appeared to corroborate the findings of the iTRAQ analysis.
CONCLUSION: The data of the present study is suggestive of the potential application of urinary osteopontin and gelsolin to discriminate patients with BTG from those with PTC non-invasively. However, this needs to be further validated in studies of individual urine samples.