Photoreduction and photophysical characterization of hydroxynitrobenzoic acids as potential UV MALDI matrices maldi

OSCAR E. SILVA RODRIGUEZ; GABRIELA PETROSELLI; HERNÁN B. RODRÍGUEZ; ROSA ERRA-BALSELLS

Río de Janeiro

Conferencia; III Iberoamerican Conference on Mass Spectrometry; 2022

Introduction: Hydroxy-nitrobenzoic acids (HNBA) have been proposed as oxidizing MALDI matrices that induce analyte fragmentation in the ionization source (ISD) in the UV-MALDI process, generating ions with a predictable structure, different from those obtained with matrices classics by collision induced dissociation (CID). Its use as a MALDI matrix is mainly focused on the analysis of peptide molecules. 1,2 However, these compounds have not been studied from the photochemical point of view as an auxiliary tool to give an explanation of its behavior as a MALDI matrix.Methods: Photochemical stability study (366 nm; 254 nm) was carried out in solution (MeCN; MeOH) of eight isomers of the HNBA family (3H4NBA; 2H4NBA; 2H5NBA; 3H2NBA; 5H2NBA; 2H3NBA; 4H3NBA; 3H5NBA), monitoring kinetic by UV-vis spectroscopy. It was observed that at 366 nm they presented a remarkable stability or very minor changes, while at 254 nm they presented significant changes. Two photo-products obtained at 254 nm in MeOH (hydroxy-aminobenzoics, HABA) were characterized by High Performance Liquid Chromatography with UV diode array (HPLC-DAD) detection: 2H4ABA from 2H4NBA and 3H2ABA from 3H2NBA. On the other hand, with the purpose of carrying out the photophysical characterization of the matrices, a detailed study of fluorescence in solid systems was conducted: (i) pure solids and (ii) matrices immersed in a non-emitting solid support, BaSO4, in proportions 1:10 in mass, with the purpose of evaluating the effect that the substituent position plays on the competition between the different deactivation pathways, because their efficiency as MALDI matrix is different for each isomer. Besides, the diffuse reflectance spectra (solid state) were obtained.Results: The photoreduction of HNBA compounds in MeOH would occurs: (1) under open atmosphere with the formation of 1O2 (2) under Ar atmosphere by taking an electron from MeOH. Both pathways lead to the oxidation of MeOH to HCOOH (HPLC-DAD). (1) requires a time delta that allows HCOOH to accumulate in sufficient quantity to start the process, HNBA → HABA. Under these conditions, the mechanism would be similar to that occurs during the preparation of references. The HNBAs that perform most efficiently as UV-MALDI matrices, in order of decreasing efficiency are: 2H5NBA, 2H4NBA, 2H3NBA, and 3H4NBA.Conclusions: None of the HNBA emit radiation efficiently, so their deactivation would occur by non-radiatively process as rapid heat released to the medium. On the other hand, the diffuse reflectance studies revealed that the HNBA in solid state absorb efficiently till near by 400 nm; Thus the 355 nm UV-MALDI laser source is suitable for population of the electronic excited states and from them induce the H /electrons exchange as a competitive deactivation process, before and during ablation/desorptionAcknowledgments: The authors thank the UBA, CONICET and MinCyt for their financialsupport. MALDI MS/MS Ultraflex II (Bruker) TOF/TOF was obtained with a grant fromANPCYT, PME2003 No.125, CEQUIBIEM, DQB, FCEN, UBA. Also thanks to the IMSF forawarding the Nico Nibbering awardReferences: [1] Fukuyama,Yetal.(2018). J.Am.Soc.MassSpectrom,29(3),2227-2236.[2]Asakawa,Detal.(2020).J.MassSpectrom,19,1-11.