Enantioselective Analysis Of Bambuterol Using Microwave-assisted Chiral Derivatization Coupled With Ultra-performance Liquid Chromatography-tandem Mass Spectrometry And Its Application

Bambuterol is a β2-adrenoceptor stimulant and can relax airway smooth muscle,which has been widely used for the treatment of asthma and bronchospasm.The bambuterol enantiomers may possess the same physical and chemical properties,but their pharmacology and pharmacodynamic properties may have great difference.Therefore,it is most important to research its chiral analysis.In this thesis,a microwave-assisted chiral derivatization coupled with UPLC-MS/MS method has been developed for determination of bambuterol enantiomers and bambuterol in human clinical plasma samples,respectively.The stereoselective pharmacokinetics of bambuterol in human has been studied.The established method has been applied to stereoselective pharmacokinetics.Moreover,we have validated the feasibility of online microwave-assisted derivatization coupled with UPLC-MS/MS for the determination of bambuterol.Main contents of this study are as follows:1.A microwave-assisted chiral derivatization coupled with UPLC-MS/MS method has been developed for determination of bambuterol enantiomers.The methodology validation and chiral inversion was studied.The reaction conditions of chiral derivatization were optimized and compared to conventional derivatization method.Furthermore,we have studied the stability of chiral derivatives.Experimental results showed conventional derivatization cost 1 h,while microwave-assisted derivatization only take 3 min.The calibration curves for bambuterol enantiomers were linear in the range of 1.00~500 pg/mL(r = 0.9975),and the LLOQ was 1.00 pg/m L.Precision and accuracy meet the relevant regulations of guidance for bioanalytical method validation.The effect of acetic acid content,temperature,power and time on the chiral inversion has been studied for the first time.The chiral derivatives are stable after being kept in room temperature for 6 h and in-80 oC for 15 days.2.A microwave-assisted chiral derivatization coupled with UPLC-MS/MS method has been developed for determination of bambuterol enantiomer in human clinical plasma samples.Reaction vessel is optimized to be 1.5 m L tubes for easy to batch processing.The reaction conditions of chiral derivatization were optimized and compared to conventional derivatization method.The calibration curves for bambuterol enantiomers were linear in the range of 2.50~500 pg/mL(r = 0.9998),and the LLOQ was 2.500 pg/m L.The precision,matrix effect,stability and recovery are all meet the requirements.A stereoselecte pharmacokinetics of bambuterol in human after an oral administration with 10 mg bambuterol tablets has been studied.The results showed that stereoselective metabolism existed in the metabolic process of bambuterol enantiomers in human.3.The on-line microwave-assisted derivatization with UPLC-MS/MS system was designed for determinating bambuterol enantiomers.The reaction conditions were optimized.The calibration curves for bambuterol enantiomers were linear in the range of 5.000~1000 pg/m L(r = 0.9965),and the LLOQ was 5.000 pg/m L.The precision and accuracy are all meet the requirements.The results showed that the on-line microwave-assisted derivatization with UPLC-MS/MS can achieved by microwave-assisted chiral derivatization coupled with large volume injection.