Mass Spectrometric Analysis Of Aldehyde Disease Markers Using Chemical Isotope Labeling Technology Based On Bromine Isotope Characteristic

Metabolomics and exposomics play an important role in the study of human health and disease.Due to the diversity of physicochemical properties of metabolites and xenobiotics and the complex sample matrices,the comprehensive analysis of metabolome and exposome is a great challenge.Mass spectrometry（MS）has powerful qualitative and quantitative capabilities,but cannot provide sufficient sensitivity for all analytes when used alone.Chemical isotope labeling（CIL）technique combined with MS can not only make up for the lack of sensitivity,but also have significant advantages in improving selectivity,accuracy,omics coverage,and analysis throughput.Because of these advantages,MS-based CIL techniques have been increasingly used in metabolomics and exposomics studies,and have greatly promoted the progresses of related research fields.CIL technique based on MS offers methodological advantages in terms of sensitivity,accuracy,selectivity,stability,analytical throughput and application,et al.More importantly,CIL technique based on MS has produced a number of outstanding studies in targeted and nontargeted analysis of metabolome and exposome,focusing on five representative metabolites and xenobiotics containing chemical functional groups of amino,carboxyl,carbonyl,hydroxyl and sulfhydryl.Moreover,it is exciting that some emerging CIL techniques for MS,such as characteristic isotope pattern strategy,have begun to be used in the structural design of CIL reagents and mass spectrum analysis,which is of great help to the improvement of the analytical performance of CIL technique.The main work of this paper is as follows:Chapter One:The application advantages of CIL technique combined with MS in the metabolome and exposome,as well as its sensitivity,selectivity,accuracy,analytical throughput and other technical considerations and evaluations were reviewed.At the same time,the analysis methods of several common metabolites or xenobiotics such as amino,carboxyl,carbonyl,hydroxyl and sulfhydryl are succinctly described,and the related more commonly used CIL reagents were briefly listed.Finally,a new design idea of CIL reagents was proposed,which uses characteristic isotope pattern strategy to design CIL reagents.Chapter Two:Two bromine-containing phenanthrene quinones,3-bromophenanthrene-9,10-dione（Br-PQ）and 2,7-dibromophenanthrene-9,10-dione（Br₂-PQ）,were used as characteristic isotope CIL reagents combined with ultra-high performance liquid chromatography and mass spectrometry（UHPLC-MS/MS）to selectively analyze hexaldehyde and heptaldehyde which are potential biomarkers of lung cancer in human blood samples.The bromine isotopes contained in the labeled product showed ⁷⁹Br and ⁸¹Br with the characteristic isotope abundance ratio close to1:1 in MS,which improved the selectivity of MS analysis and helped to exclude the effects of false positives.The limit of detection（LODs）of this method is 0.36 n M,which can provide technical support for early screening of lung cancer.Chapter Three:4-Hydroxynonenal（4-HNE）is a kind of unsaturated fatty aldehyde with neurotoxicity,which has potential research value in a variety of neurodegenerative diseases.In combination with the characteristic reaction of phenoquinone with aldehydes,a strategy was established that used two bromine-containing phenanthrene quinones,Br-PQ and Br₂-PQ,as CIL reagents combined with UHPLC-MS/MS for highly selective and sensitive analysis of 4-HNE as a potential marker of Alzheimer’s disease（AD）in human blood samples.In this strategy,not only the characteristic reaction of phenanthrene quinones with ammonium acetate and aldehydes greatly reduce the side reactions during the labeling process,but also the characteristic isotopes of bromine can improve the selectivity of MS analysis.The LODs of this method is0.34 n M,provides a new method with highly sensitive for the early diagnosis of AD patients.