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Accurate and reliable quantification of endogenous lipid mediators in complex biological samples is a daunting challenge. In this study, a robust and direct endogenous quantitative method using background subtracting calibration curves by liquid chromatography-tandem mass spectrometry was first developed for the determination of endogenous lipid mediators in ischemic stroke rats. Absolute quantification without surrogate matrix could be achieved by using background subtracting calibration curves, which were corrected and verified from standard curves constructed on original matrix. The recoveries of this method were in the range of 50.3-98.3%, the precision with the relative standard deviation was less than 13.8%, and the accuracy with the relative error was within ± 15.0%. In addition, background subtracting calibration curves were further verified by validation factors ranging from 90.3 to 110.9%. This validated method has been successfully applied to the analysis of seven endogenous inflammation-related lipid mediators in the brain tissues of ischemic stroke rats. The results indicated that prostaglandins as inflammatory factors and some lipid mediators with neuroprotective effects increased apparently (p < 0.05) in the stroke groups compared with the normal rats. Besides, the two drugs (isosteviol sodium and edaravone) could significantly reduce (p < 0.05) the levels of prostaglandin E and prostaglandin F of stroke rats to inhibit inflammation. Based on the results, it is strongly believed that this approach can be readily generalized as a new reference for the quantification of endogenous compounds in the complex biological samples. Graphical abstract The analysis procedure of determining endogenous inflammation-related lipid mediators using BSCC by LC-MS/MS.

R)-Salbutamol is a selective β2-adrenoreceptor agonist, which produces a shortacting bronchodilator effect and is widely used for the treatment of respiratory diseases in humans. Drug metabolism and identification of the metabolites play an essential role in the evaluation of the overall efficacy and safety of the drugs in clinical practices. There are few reports on the identification of major metabolites of (R)salbutamol in humans, and the number of identified metabolites is very limited. In this research, a method of ultra-high performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry was developed for the discovery and identification of (R)-salbutamol and its major metabolites in human biological samples. Totally, twelve metabolites of (R)-salbutamol were found and identified and all the metabolites could be found in urine, one metabolite in plasma and two metabolites in feces. Among all the metabolites, eight metabolites have never been reported before. The results indicated that (R)-salbutamol was mainly metabolized through isomerization, oxidation, reduction, glucuronidation, and sulfation pathways in vivo. The possible metabolic pathways of (R)-salbutamol were subsequently presented in this study, which contribute to a better understanding of the metabolism of (R)-salbutamol in humans. K E Y W O R D S biological samples, metabolic pathways, quadrupole time-of-flight mass spectrometry, (R)-salbutamol, ultra-high performance liquid chromatography Article Related Abbreviations: CID, collision-induced dissociation; M, metabolite; PIS, product ion scanning; RA, relative abundance. Fengying Ye and Shan Liu contributed equally to this work.and (S)-salbutamol. It is well-known that the enantiomers of a chiral drug have different pharmacodynamics and pharmacokinetic behaviors [3,4]. (R)-Salbutamol is the active β2 receptor stimulant while (S)-salbutamol is presumed to be inert. Thus, (R)-salbutamol carries most of the therapeutically bronchoprotective and bronchodilator effects and has been approved as a new drug for the treatment of pulmonary diseases [5].The investigation of drug metabolism is of great significance to the evaluation of the efficacy and safety of clinical drugs. Some early researches reported the major metabolites 3200

In this study, an enantioselective analytical method based on microwave-assisted chiral derivatization coupled with ultra high performance liquid chromatography and tandem mass spectrometry was developed for the determination of bambuterol enantiomers in human plasma. The chiral derivatization reaction was greatly accelerated by microwave irradiation. Under the optimized conditions, both the derivatization time and separation time on column was only 3 min, and the lower limit of quantification was 2.5 pg/mL. The recoveries were in the range of 90.1-93.0% without significant matrix effect. Compared with the conventional heating chiral derivatization, microwave-assisted chiral derivatization obtained higher chiral derivatization yields with much shorter time due to the effect of microwave irradiation. Furthermore, the racemization during the derivatization reaction was systematically investigated. The results showed the concentration of acetic acid and the reaction time had significant effects on the racemization, which could be well controlled during microwave-assisted chiral derivatization for the short reaction time. Finally, this novel approach was demonstrated by determining bambuterol in human plasma of a clinical pharmacokinetic study in eight healthy volunteers. On the basis of the results, microwave-assisted chiral derivatization coupled with ultra high performance liquid chromatography and tandem mass spectrometry as a simple and effective enantioselective analysis technique for the determination of chiral drugs in complex biological samples showed great promise.