Application of EPR spectroscopy to examination of gentamicin and kanamycin binding to DOPA-melanin

Abstract

Electron paramagnetic resonance (EPR) spectroscopy was applied to measure the influence of two aminoglycoside antibiotics: gentamicin and kanamycin on free radical properties of DOPAmelanin. DOPA-melanin was formed by oxidative polymerization of 3,4-dihydroxyphenylalanine. Different concentrations of gentamicin and kanamycin (from 1 9 10 -4 to 1 9 10 -2 M) were used. o-Semiquinone free radicals with a g factor of 2.0043 were found in all studied melanin samples. Their concentrations in the DOPA-melanin-drug complexes were higher than in DOPA-melanin, and increased with the increase of gentamicin and kanamycin concentration. A single EPR line of the analyzed samples (ABp~, 0.48-0.52 mT) indicates that aminoglycoside antibiotics do not create a new type of free radicals in DOPA-melanin. Microwave saturation behavior of the experimental lines indicates the homogeneous broadening of resonance absorption curves for DOPA-melanin and its complexes with aminoglycosides. The EPR lines saturate at low microwave powers. Slow spin-lattice relaxation processes were characteristic for all studied melanin samples.

Influence of Microwave Power on EPR Signal of Melanin Radical and Copper(II) Ions in DOPA-Melanin Complexes

“…Adding of drug into melanin causes the increase of amplitude A of EPR signal. Drugs (112) cause higher concentration of melanin paramagnetic centers [11,12]. Intensity I of EPR line is proportional to concentration of paramagnetic centers in sample and intensity I is proportional to amplitude A and for symmetric EPR line we can write…”

Paramagnetic Centers in DOPA-Melanin-Dihydrostreptomycin Complexes

“…Eumelanins with characteristic single lines mainly exist in living organisms. Free radicals play an important role in biochemical reac-tions with drugs [2][3][4]. Application of aminoglycoside antibiotics therapy may be accompanied by toxic effects resulting from free radicals generation in melanin.…”

Triplet States in DOPA-Melanin and in Its Complexes with Kanamycin and Copper Cu(II) Ions

“…The EPR signal of melanin radicals undergoes microwave saturation [19][20][21], therefore the linewidth ∆B pp of EPR signal is determined by spin-lattice relaxation time T 1 and by spin-spin relaxation time T 2 [22,23]. The relaxation time T 1 is dependent on temperature of measurement, however, the relaxation time T 2 changes only slightly with temperature.…”

“…On this ground, we can say that kanamycin added to DOPA-melanin polymer increases the number of paramagnetic centres with spin S = 1. Because the mechanism describing how drugs are bound to melanin is not well recognised yet [19,20,25], it is difficult to explain fully this effect. It is known that intensity of EPR line of melanin increases after the addition of drugs [19,20,26].…”