Photochemical Reaction of 2-Nitrobenzaldehyde by Monitoring the Diffusion Co Jungkwon Choi and Masahide Terazima* Department of Chemistry, Graduate School of Science, Kyoto UniVersity, Kyoto 606, Japan ReceiVed: January 25, 2003; In Final Form: June 2, 2003 The photochemical reaction of 2-nitrobenzaldehyde (NBA) in various electrolyte solutions as well as in a solution with a protein is investigated from the viewpoint of the diffusion coefficient by using the time- resolved transient grating (TG) method. For studying reaction with optically silent materials such we found that the diffusion coefficient is useful for identifying chemical species involved in the re TG signal that represents the diffusion process was dramatically changed by adding electrolytes s - or Br - , and this result indicates that these electrolytes should participate in the photochemical reaction of NBA. It was found that the time profile of the grating signal also depends on the addition of guani or apomyoglobin. The structural change of the protein associated with the NBA reaction is discussed 1.Introduction Since caged compounds can control the concentration of a bioactive substance such as proton, 1-3 ATP, 4 GTP, 5 and Ca 2+ , 6 etc., spatially as well as temporally by photoirradiation, a variety of caged compounds have been developed and widely utilized in biophysical, biochemical, physiological, or cellbiological systems. Understanding the substance-releasing process is essential to correctly interpret observations in these studies. In many cases, caged compounds have a 2-nitrobenzyl group, which is a major photolabile precursor. For example, a photo- dissociation reaction of 2-nitrobenzaldehyde (NBA) can change the proton concentration. 1-3 Since thisis the simplest and fundamental caged compound, the reaction of NBA should be clarified in order to elucidate the mechanism of the substance releasing of the other caged compounds. In particular, the caged reactions are frequently used with the presence of ions or proteins for biological research, and, therefore, the study of the reaction under these conditions is unavoidable. For studying chemical reactions, the transient absorption method has been frequently used and it is certainly a very powerful and informative technique. The fundamental reaction scheme of NBA has been reported by the transient absorption method. 1 Upon the photoexcitation of NBA, the aci-form is rapidly produced through the intramolecular proton-transfer reaction and a proton is released to yield the nitronate anion in aqueous solution (pH jump). Consequently the nitronate anion is converted to 2-nitrosobenzoic anion (NS - ) with a quantum yield of about 0.4 (Scheme 1). When this reaction takes place in a buffer solution for a biological application, many ions or proteins should be contained in the solution and the reaction mechanism could be different from that in aqueous solution. The contribution of these ions to the reaction scheme should be clarified. However, it is rather difficulto detect the dynamics of such ions by the transient absorption method because these species absorb light in the far- UV region, and the absorption measurement in this region is frequently disturbed by the presence of many absorption ban from many species including NBA itself. Therefore it is desirable if there is another independent method that can de such optically silent materials. In this respect, the detection o molecular volume change or diffusion coefficient (D) could provide a very unique method to clarify the reaction involving such ions. One prominent example recently shown is a study of the kinetics of a proton released from NBA. 2,7 There is no light absorption band of proton, and the presence is very diff to detect without adding a pH sensitive dye, which could cha the kinetics of the proton. However, even in such a case, we found that the presence of the proton can be identified by the diffusion coefficient and that the time development of the pro concentration can be monitored by the volume contribution without any pH indicator reagent. In this paper, we investiga the reaction of NBA in various solutions with the presence of ions and a protein, apomyoglobin (Mb), by monitoring D. The diffusion coefficient was sensitively detected in a time domain by the transient grating (TG) technique. We found evidence f the D measurement that the electrolyte ions are involved in t NBA reaction. Previously the NBA photoreaction was used to study the protein folding process induced by the pH jump in the presen of guanidine hydrochloride (GdnHCl). 3,8 Protein folding of * To whom correspondence should be addressed. Telephone: +81-75-753-4026. FAX: +81-75-753-4000. E-mail: mterazima@ kuchem.kyoto-u.ac.jp. SCHEME 1. Photoreaction of NBA 9552 J. Phys.Chem. B 2003, 107, 9552-9557 10.1021/jp0342071 CCC: $25.00 © 2003 American Chemical Society Published on Web 08/05/2003