Fluorescence spectroscopy of UV-MALDI matrices and implications of ionization mechanisms

Abstract

Matrix-assisted laser desorption ionization (MALDI) has been widely used in the mass analysis of biomolecules; however, there are a lot of debates about the ionization mechanisms. Previous studies have indicated that S1-S1 annihilation might be a key process in the generation of primary ions. This study investigates S1-S1 annihilation by examining the time-resolved fluorescence spectra of 12 matrices. No S1-S1 annihilation was observed in six of these matrices (3-hydroxy-picolinic acid, 6-aza-2-thiothymine, 2,4-dihydroxy-acetophenone, 2,6-dihydroxy-acetophenone, 2,4,6-trihydroxy-acetophenone, and ferulic acid). We observed two matrix molecules reacting in an electronically excited state (S1) in five of these matrices (2,5-dihydroxybenzoic acid, α-cyano-4-hydroxycinnamic acid, 2,5-dihydroxy-acetophenone, 2,3-dihydroxybenzoic acid, and 2,6-dihydroxybenzoic acid), and S1-S1 annihilation was a possible reaction. Among these five matrices, no S1-S1 annihilation was observed for 2,3-dihydroxybenzoic acid in typical peak power region of nanosecond laser pulses in MALDI, but a very small value of reaction rate constant was observed only in the high peak power region. The excited-state lifetime of sinapinic acid was too short to determine whether the molecules reacted in an electronically excited state. No correlation was observed between the ion generation efficiency of MALDI and S1-S1 annihilation. The results indicate that the proposal of S1-S1 annihilation is unnecessary in MALDI and energy pooling model for MALDI ionization mechanism has to be modified.

Reply to “Comment on: MALDI ionization mechanisms investigated by comparison of isomers of dihydroxybenzoic acid”

“…However, they showed only a short, noisy part of the decay. As we reported even at low fluence high‐quality decay curves demonstrate both non‐exponentiality at short times, and a pronounced biexponential decay at times beyond the cutoff in Lin et al .,. We also found that the purity of commercial THAP dramatically affected the decay curves, as did the solvent used to prepare the samples.…”

“…Chen and Ni note that the Ni group has reported pooling rate constants in a different ratio than we proposed . Currently, one of us (R. K.) is conducting a new study of DHB isomers, including fluence dependence of emission yields and photodynamics.…”

“…Related results and conclusions from Lin et al , and have previously required revision . It was reported that the MALDI matrix 2,4,6 trihydroxyacetophenone (THAP) exhibits only a pure exponential decay and that sample purity had no effect .…”

Ionization Mechanism of Matrix-Assisted Laser Desorption/Ionization

“…In this model, S 1 -S 1 annihilation is the first key step to ion generation. However, recent studies (59,60) have shown that numerous matrices do not undergo S 1 -S 1 annihilation. Another example is the contribution from preformed ions.…”

Ion Yields in the Coupled Chemical and Physical Dynamics Model of Matrix-Assisted Laser Desorption/Ionization

“…These and earlier values were used to argue that the CPCD cannot be correct because it is claimed to predict yields that are too high [4,[26][27][28]. Here, we use the newest data to revise the S n + S 1 pooling rate constants, and to test the revised model against the data for DHB and CHCA matrices.…”