Oxidation of Purine Nucleotides by Triplet 3,3′,4,4′-Benzophenone Tetracarboxylic Acid in Aqueous Solution: pH-Dependence

Abstract

The photo-oxidation of purine nucleotides adenosine-5′-monophosphate (AMP) and guanosine-5′-monophosphate (GMP) by 3,3′,4,4′-benzophenone tetracarboxylic acid (TCBP) has been investigated in aqueous solutions using nanosecond laser flash photolysis (LFP) and time-resolved chemically induced dynamic nuclear polarization (CIDNP). The pH dependences of quenching rate constants and of geminate polarization are measured within a wide range of pH values. As a result, the chemical reactivity of reacting species in different protonation states is determined. In acidic solution (pH < 4.9), the quenching rate constant is close to the diffusion-controlled limit: kq = 1.3 × 109 M–1 s–1 (GMP), and kq = 1.2 × 109 M–1 s–1 (AMP), whereas in neutral and basic solutions it is significantly lower: kq = 2.6 × 108 M–1 s–1 (GMP, 4.9 < pH < 9.4), kq = 3.5 × 107 M–1 s–1 (GMP, pH > 9.4), kq = 1.0 × 108 M–1 s–1 (AMP, pH > 6.5). Surprisingly, the strong influence of the protonation state of the phosphoric group on the oxidation of adenosine-5′-monophosphate is revealed: the deprotonation of the AMP phosphoric group (6.5) decreases the quenching rate constant from 5.0 × 108 M–1 s–1 (4.9 < pH < 6.5) to 1.0 × 108 M–1 s–1 (pH > 6.5).

Kinetics of Reversible Protonation of Transient Neutral Guanine Radical in Neutral Aqueous Solution

“…Furthermore, we have demonstrated that based on the difference in g-factor, it is possible to differentiate between the two radicals utilizing only spectral and kinetic CIDNP data, despite of the close similarity in spin density distribution in the two investigated guanine radicals as discussed in Ref. [25] To identify the protonation position, heteronuclear CIDNP experiments are planned, namely, 15 N and 13 C CIDNP. The information presented can now be used to aid in the identification of transient guanine radicals involved in nucleic acid damage.…”

“…[25] In the present study, measurements were performed at pH 6.2 and higher to exclude contributions of the reaction between GMP and triplet excited TCBP having two protonated carboxylic groups (pK a~4 .9). [25] In the present study, measurements were performed at pH 6.2 and higher to exclude contributions of the reaction between GMP and triplet excited TCBP having two protonated carboxylic groups (pK a~4 .9).…”

“…The hyperpolarization formed is sensitive to the magnetic resonance parameters of the radicals and therefore allows for determination of structure of shortlived radicals. [25] A detailed analysis of the pH-dependence of the geminate CIDNP intensity allowed us to reveal the mechanism of the reaction between triplet-excited TCBP and GMP in its dependence on the protonation state of the reacting species. [20] Moreover, photo-CIDNP in its time-resolved version opens the possibility to follow the kinetics of radical transformations with microsecond time resolution: CIDNP effects arising in primary radical pairs on a nanosecond time scale can be separated from the influence of radical reactions on a microsecond time scale.…”

Time‐Resolved Chemically Induced Dynamic Nuclear Polarization of Biologically Important Molecules

“…By using a photo‐sensitized chemical reaction one can also polarize purine nucleotides. In adenosine, the H2 and H8 protons can be efficiently polarized,, whereas in guanosine only the H8 proton exhibits CIDNP ,, . NMR detection for different nuclei, e. g., 13 C and 15 N, is also feasible: in studies of CIDNP of such hetero‐nuclei one can obtain detailed information on the spin density distribution in transient radicals ,.…”

“…As photosensitizers, which are used to initiate a photo‐reaction giving rise to CIDNP, various dyes are used, which are listed in Scheme . These dyes are 2,2‐dipyridyl (DP),– 4‐carboxy‐benzophenone (4‐CBP),,, 3,3′,4,4′‐tetracarboxybenzophenone (TCBP),, and flavins with different substituents (flavin mononucleotide, FMN, is shown in Scheme ) ,,,. For running experiments under special conditions, i. e., at very low concentrations, one can also use other dyes, such as fluorescein .…”