Heteroatom Doped-graphene Quantum Dots As A Matrix For MALDI-TOF MS Probing Of Small Molecules And MALDI MS Imaging Of Tissue Samples

Matrix assisted laser desorption ionization time of flight mass spectrometry(MALDI-TOF)has a series of advantages,such as fast and high throughput data acquisition,soft ionization,easy sample preparation,low sample consumption and high sensitivity.It has already been widely used in the field of biological analysis,especially on the detection of biological macromolecules,including proteins,peptides,oligosaccharides,synthetic polymers,etc.However,the application of MALDI is still limited for small molecules characterizations(<500 Da),because the traditional matrix will generate strong endogenous background in the low-mass region,suppressing the target analyte signals and making spectra complex and difficult to parse.Small molecules(including amino acids,fatty acids and peptides)play a crucial role in regulating cell growth,signal transfer,and other physiological activities.Many reports have verified that varied spatial distribution of functional small biomolecules was tightly associated with cancer,cardiovascular disease,nervous system,etc.,highlighting the importance of identification and detection of these small molecules.In this work,we adopt heteroatom doped-graphene quantum dots(HD-GQDs)as the novel matrix for MALDI-TOF MS analysis,and investigate the mechanism how the carbon-based materials assist desorption/ionization(D/I)process.By virtue of the high D/I efficiency and good homogeneity of HD-GQDs,high sensitive detection of small molecule and mass spectrometry imaging of tissue samples have been successfully accomplished.1.HD-GQDs as the novel matrix for highly efficient MALDI-TOF MS detection of small molecules.Three graphene quantum dots(NGQDs,BNGQDs,BGQDs)were derived from the precursor graphene(NG,BNG,BG)by microwave chemical cleavage strategy,and used as the matrixs for MALDI-TOF MS analysis of small molecules in the negative ion mode.The results showed that NGQDs had the best effect,and the mechanism of laser desorption and ionization was discussed,the N elements doped in play a crucial role.Unlike the complicated ion signals of adducts that emerge in the positive ion mode,only the characteristic peaks of the deprotonated analytes were observed in the negative ion mode,rendering the spectrum easy to interpret.Based on the excellent D/I efficiency of NGQDs,a series of low molecular weight analytes can be detected efficiently,including amino acids,fatty acids,peptides,nucleic acid bases,sucrose,anti-cancer drugs,environmental pollutants and so on.Different from traditional matrix DHB,which suffers "sweet spot" effect,NGQDs have high salt tolerance,good dispersion,and good reproducibility.The above performances prove that NGQDs are an efficient matrix which provide a good solution for the detection of small molecules by MALDI-TOF MS.2.NGQDs as a novel matrix for MALDI-TOF MS analysis and imaging of biomolecule in tissue samples and evaluation of enzyme Michaelis-Menten constantDue to its strong ultraviolet absorption,low background interference in the small molecular range,and good salt tolerance,NGQDs were employed as a novel matrix for analysis and imaging of small molecules in tissue samples by MALDI-TOF MS.In this experiment,more than 60 substances,including amino acids,fatty acids,ceramides,phosphatidic acids were successfully detected and imaged in the mouse rat tissue sections with good signal reproducibility.This method is of great significance to unveil the possible pathological characters and mechanism of many diseases on the molecular level.In addition,we have also determined the Michaelis-Menten constant(Km)of trypsin by quantitatively monitoring the small molecule enzymatic products via internal standard method.