Development Of C60-based Labeling Reagents For The Determination Of Low-molecular-weight Compounds By MALDI-TOF-MS

Matrix assisted laser desorption ionization mass spectrometry(MALDI-TOF MS)based on its easy operation,fast detection,high-throughput,strong anti-interference ability,less organic solvent consumption,and low sample consumption and other advantages,has been widely used in the analysis of the structure and molecular weight determination of protein,sugars,and nucleic acids,peptides,biological macromolecules and polymer.But,due to the conventional MALDI-TOF-MS matrix are mainly low molecular weight organic acids such as CHCA,DHB,SA,nicotinic acid,etc.,they are easily to produce a large number of matrix signal peak in the region of the low molecular weight(<800 Da),affect the target molecular mass spectrum peak identification.Moreover,due to the uneven distribution of substrate-sample points on the target board,make MALDI-TOF-MS reproducibility is poor,interfere with the identification and quantitative analysis of small molecular detection.Since fullerenes was discovered in 1985,we opened a new chemical world,with its representatives of C60,is a series of unique cage structure of three-dimensional aromatic compounds.The three-dimensional structure and a large number of double bond of fullerenes,which determine its unique physical and chemical properties,have provided a broad space for fullerenes science development.Fullerene functionalization chemistry remains one of the hottest topics in fullerene chemistry,and tremendous efforts are underway to obtain various fullerene derivatives.After years of intense studies,fullerene functionalization approaches is now mature.Various functional groups including amino,carboxyl,hydrazide,succinimide etc.,are easily incorporated into the fullerene structure as desired by nucleophilic addition,pericyclic reaction,oxidation or hydroxylation.The resulting fullerene derivatives are chemically reactive,and can be used to readily derive a variety of low molecular weight(LMW)compounds.Here,a C60 labeling strategy was proposed to couple with MALDI-TOF-MS for the analysis of LMW compounds.After derivatization,the LMW compounds can be labeled with a large fullerene tag to add 720 Da to the molecular weight.Therefore,the application of fullerene derivatives as the labeling reagents of LMW compounds can shift the LMW weight from a low m/z range to a high m/z reign.In addition,C60-labeled LMW product retains the main properties of the original C60,which makes sure that all C60-labeled products have similar MALDI-TOF-MS behavior and could provide easily approached C60 analogue as IS.Based on the issue above,some work are carried as follows:1.Synthetic fullerene derivatives with the chemical activity,such as NHS-C60 and Br-C60,and to investigate whether the derivatization reaction with small molecular compounds,validation the possibility of fullerene derivatives as MALDI-TOF-MS m/z displacement reagent;2.As a proof-of concept,a N-hydroxysuccinimide functionalized C60(NHS-C60)was synthesized as the labeling reagent,and a carboxyl acid functionalized C60(COOH-C60)was employed as the IS.They were applied to the MALDI MS analysis of amino acids(AAs)in biofluids,which are the basic building blocks of proteins and play a critical role in maintaining human physiology.