Benzo Acridone Is Dual-sensitive Probe Preparation And Its Application In The Reversed-phase High Performance Liquid Chromatography - Mass Spectrometry

For the powerful separation and analytical capacities, reversed-phase high performance liquid chromatographic method has been widely used in many fields of scientific research and application, such as biology, chemistry, toxicology, clinical medicine, food hygiene and environmental monitoring. It is also one of the fastest developing and the most widely used methods in analytical chemistry. However, not all compounds are accessible to be analyzed by chromatography due to the limitation of the compounds'properties and the low content of detected components. Therefore, developing testing methods with high sensitivity has very important significance. In recent years, derivatization of analytes with fluorescent labeling reagent has been widely adopted by many researchers. This derivatization method which adopt fluorescence detection can enhance detective sensitivity in a large extent. In this paper, three dual-sensitive benzoacridinone probes which were sensitive not only to fluorescence but also to MS ionizable potential were designed, prepared and characterized. The spectral properties, optimal derivatization conditions, chromatographic separation conditions and MS conditions were investigated. Pre-column derivatization methods based on reversed-phase high-performance liquid chromatography were established and evaluated for the analysis of real samples.Chapter One:The developing progress and basic theory about high performance liquid chromatography and high performance liquid chromatography coupled with mass spectrometry were simply presented. The principles of fluorescence and chemical derivatization were introduced. The widely used fluorescence derivatization reagents were summarized.Chapter Two:The syntheses of three fluorescent probes: 2-(benzoacridin)ethyl-imidazole-1-carboxylate(BAEIC), 5-(2-(1H-imidazol-1-yl)-2-oxoethyl)benzo[b]acridin-12(5H)-one(IEBA)and 2-(12-oxobenzo[b]acridin-5(12H)-yl)ethyl 4-methylbenzenesulfonate(BAETS) were described followed with their structural characterization.Chapter Three:Pre-column derivatization method for the sensitive determination of amine compounds using BAEIC as labeling reagent followed by HPLC-MS analysis has been developed. The optimal derivatization was evaluated. The proposed method showed wide linear range, excellent reproducibility, and simple derivatization for the determination of aliphatic amines. The suitability of the developed method for the analysis of real sample was satisfactory.Chapter Four:Pre-column derivatization methods for the sensitive determination of aliphatic amine, aromatic amines, short-chain aliphatic alcohols and policosanol with 5-(2-(1H-imidazol-1-yl)-2-oxoethyl)benzo[b]acridin-12(5H)-one (IEBA) as labeling reagent followed by HPLC-MS analysis have been developed. The optimization of derivatization and chromatographic separation conditions using IEBA as labeling reagent was evaluated. Then, this method was applied to the determination of different real samples, the results suggested that the established methods have great feasibility.Chapter Five:Pre-column derivatization method for the sensitive determination of bile acids with 2-(12-oxobenzo[b]acridin-5(12H)-yl)ethyl 4-methylbenzenesulfonate(BAETS) as derivative reagents followed by HPLC-MS analysis has been developed. The optimization of derivatization and chromatographic separation conditions using BAETS as labeling reagent was evaluated. Then, this method was applied to the determination of actual sample, the results suggested that the established method has great feasibility.