Anna Koziołowa

SOLVENT AND METHYL SUBSTITUENT EFFECT ON PHOTOTAUTOMERISM AND IONIZATION OF ALLOXAZINES

Abstract— The absorption and emission spectra of alloxazine and its methyl derivatives have been measured in solvents of different polarity. From roughly linear correlation between two long wavelength absorption maxima and the emission maxima positions vs solvent polarities. systematic deviations have been observed for alloxazines in pyridine, acetic acid and water. The deviations depend on the methylation site of alloxazine in a way suggesting formation of hydrogen bond in the ground state and phototautomerism in the excited state. As phototautomerism of alloxazines in the presence of pyridine and water may involve ionic equilibria, the pKa values of alloxazines have been measured (8.11–8.58) and the pKB* estimated from the Förster cycle (2.3–6.6 for N‐1 monoanion and 7.4–7.8 for N‐3 monoanion). The phototautomeric efficiency of alloxazines in pyridine, acetic acid and water depend on the acidity of the N‐1 proton and on the steric effects of methyl substituents, mainly at C‐9. The anionic forms of alloxazines differ from the phototautomeric species in emission maxima positions, radiative lifetimes and they are dissimilarly influenced by the methylation site and medium viscosity.

Photophysical properties of lumichromes in water.

The photophysics of lumichrome, 1-methyllumichrome, and lumiflavin in water solutions have been investigated. Fluorescence lifetimes of 2.7 and 2.2 ns were observed for lumichrome and 1-methyllumichrome, respectively, the corresponding triplet state lifetimes of 17 and 18 micros have been obtained from the transient absorption spectra. Evidence for long lived species with absorption maxima near 450 nm and lifetimes of ca. 400 micros has been found in the transient absorption spectra of both lumichromes. Quantum yields for the sensitised production of singlet oxygen, phi(Delta), are 0.36 and 0.41 for lumichrome and 1-methyllumichrome, respectively, in D(2)O.

Spectroscopy and photophysics of alloxazines studied in their ground and first excited singlet states

The acid–base properties of alloxazine (All) and its methyl derivatives have been studied in their ground and first excited singlet states. The concept of an effective electronic valence potential was applied to predict the changes in basicity and acidity of heteroatoms upon excitation and substitution. Changes in the acid–base properties of N(1) and N(10) nitrogen atoms are particularly important from the point of view of the excited state proton transfer in alloxazines from N(1) to N(10) to form isoalloxazinic structures. A good linear correlation was obtained between the calculated electronic potentials of N(1) and N(3) nitrogen atoms and the experimental pKa values for ground and excited state deprotonation.

The solvent effect on the excited-state proton transfer of lumichrome

Abstract The acetic acid (AA)-catalysed tautomerism of lumichrome (Lc) was investigated in 1,2-dichloroethane, acetonitrile and pure AA. The interactions between Lc and AA were studied by means of UV-Vis and fluorescence spectroscopy. The results suggest the formation of 1:1 Lc–AA hydrogen-bonded complexes in the ground state of Lc. The apparent equilibrium constants were one order of magnitude higher in 1,2-dichloroethane than in acetonitrile, indicating a solvent effect on the ground-state interactions. The dynamics of exited-state processes were studied using time-resolved methods. The results show that the mechanism of tautomerism depends on the solvent.

Phototautomerism of lumichrome in methanol-acetic acid mixtures: a steady-state and time-resolved fluorescence study

Abstract— Fluorescence lifetimes of 3‐methyllumichrome dual fluorescence at 440 nm (alloxazinic) and 520 nm (isoalloxazinic) and of 1,3‐dimethyllumichrome, which is unable to phototautomerize, at the same wavelengths have been measured in methanol‐acetic acid mixtures. The fluorescence decays of both lumichromes studied are exponential and the phototautomeric fluorescence of 3‐methyllumichrome is created within approximately 50 ps. From the initial values of about 0.9 ns (alloxazinic) and 6.4 ns (isoalloxazinic) in 5% acetic acid the lifetimes shorten considerably with increasing acid concentration reaching 0.2 ns for 1,3‐dimethyllumichrome and 80 ps for the alloxazinic and 2.4 ns for the isoalloxazinic form of 3‐methyllumichrome in pure acid. Static and dynamic fluorescence quenching constants were estimated. The differences in the quenching rate and equilibrium constants of both lumichromes are interpreted in terms of different equilibria of hydrogen bond formation in ground and excited state as influenced by steric effects of the methyl substituent at the N‐l position in 1,3‐dimethyl. The hydrogen bonding atN–10 of 3‐methyllumichrome with acetic acid is a prerequisite for additional hydrogen bonding at N‐l enabling excited state proton transfer.

Phototautomerism of 5-deazalumichrome (in the presence of acetic acid)

Abstract The phototautomerism of 5-deazalumichrome was studied in 1,2-dichloroethane in the presence of acetic acid. In comparison with 3-methyllumichrome, 5-deazalumichrome undergoes excited state proton transfer with higher efficiency, manifested by its fluorescence spectrum and lifetimes. The replacement of N-5 by carbon in the pyrazine ring of alloxazine restricts the electron density redistribution involved in the phototautomerism to the region of N-1:C-10a:N-10, and eliminates the competitive contribution of the rest of the conjugated double bond system. Semiempirical calculations confirm the experimental observations.

A new method of isolation of natural flavins using phenoltype resins.

It was shown that riboflavin (as lumiflavin and lumichrome) is sorbed by all resins with free phenolic groups, e.g. phenolsulphonic and phenolcarboxylic ion exchangers or a simple resin of resorcinol and formaldehyde. FMN and FAD, in aqueous solutions, passed through these columns without sorption, but can be retained in salt solutions, especially by a resorcinol resin. Flavins can be eluted from phenol resins by a mixture of water and acetone, propanol or dioxane and saturated aqueous solutions of urea with ethanol or cellosolve. The ability of phenol resins to form complexes with flavins can be used both for analytical and preparative purposes.

A fluorescence study of lumichrome phototautomerism in dodecylammonium propionate reversed micelles.

Abstract Spectral and time-resolved polarized fluorescence experiments were carried out on N(3)-undecyllumichrome and N(3)-undecyllumiflavin encapsulated in reversed micelles of dodecylammonium propionate in cyclohexane. The compounds were equipped with a long chain to anchor them to the interphase boundary between the water droplet and the organic phase. The spectral experiments revealed that the amphiphilic compounds were located at the interphase. The observation that the propionate counter-ion of the miceIlar ammonium head group enables phototautomerism to yield isoalloxazinic fluorescence provided evidence for the close contact of lumichrome with propionate. Dynamic information was obtained from a two-dimensional maximum entropy method of analysis of the time-resolved fluorescence data of flavin and phototautomeric lumichrome. On the basis of these results it was clear that the fluorophores were distributed differently within the micellar boundary region.

The effect of hydrogen bonding on the electron transfer reactions of the excited singlet and triplet states of flavins

Abstract The effects of hydrogen bonding of riboflavin tetrabutyrate to acetic and trichloroacetic acids in 1,2-dichloroethane were studied. The reactivity of the excited singlet and triplet states shows dramatic increases in the presence of trichloroacetic acid and to a lesser extent acetic acid. However, it is shown that the effects of trichloroacetic acid on the excited triplet state are partly due to protonation in addition to hydrogen bonding. The effects of hydrogen bonding confirm earlier molecular orbital calculations predicting an increase in electrophilicity of flavin excited states upon hydrogen bonding.