UHPLC-MS/MS method for the simultaneous determination of nicotine and tobacco-specific nitrosamines NNN and NNK for use in preclinical studies.

A new method was developed and validated for the simultaneous determination of nicotine and tobacco-specific nitrosamines (TSNAs) 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) and N-nitrosonornicotine (NNN) in two different tests matrices: porcine buccal epithelium tissue and phosphate buffered saline (PBS) extracts of smokeless tobacco products. The novelty of this work is in the development of a liquid chromatography tandem mass spectrometry method that can provide simultaneous quantification of trace levels of TSNAs and high concentrations of nicotine in biological media. Precision, accuracy, and stability were evaluated during method validation to ensure the method was fit for purpose. Several sample preparation and extraction methods were evaluated to minimize matrix effects and maximize analyte recoveries. The method was accurate in the range of 81.1% - 117%; repeatability was estimated in the range of 1.5% - 13.6% across multiple concentrations. The linear regression correlation coefficient (R2) was greater than 0.9959 for all analytes, and the limit of detection (LOD) was determined for nicotine, NNK, and NNN at 1 ng/mL 0.005 ng/mL, and 0.006 ng/ mL, respectively. Our method was found to be appropriate for the analysis of nicotine, NNN, and NNK in the porcine buccal epithelium and PBS extracts of smokeless tobacco products.

Impact of exercise on fecal and cecal metabolome over aging: a longitudinal study in rats.

AIM: Physical exercise can reduce adverse conditions during aging, while both exercise and aging act as metabolism modifiers. The present study investigates rat fecal and cecal metabolome alterations derived from exercise during rats' lifespan. METHODS & RESULTS: Groups of rats trained life-long or for a specific period of time were under study. The training protocol consisted of swimming, 15-18 min per day, 3-5 days per week, with load of 4-0% of rat's weight. Fecal samples and cecal extracts were analyzed by targeted and untargeted metabolic profiling methods (GC-MS and LC-MS/MS). Effects of exercise and aging on the rats' fecal and cecal metabolome were observed. CONCLUSION: Fecal and cecal metabolomics are a promising field to investigate exercise biochemistry and age-related alterations.