Antiradical Activity of Dopamine, L-DOPA, Adrenaline, and Noradrenaline in Water/Methanol and in Liposomal Systems

Abstract

Catecholamines play a crucial role in signal transduction and are also expected to act as endogeneous antioxidants, but the mechanism of their antioxidant action is not fully understood. Here, we describe the impact of pH on the kinetics of reaction of four catecholamines (L-DOPA, dopamine, adrenaline, and noradrenaline) with model 2,2-diphenyl-1-picrylhydrazyl radical (dpph • ) in methanol/water. The increase in pH from 5.5 to 7.4 is followed by a 2 order of magnitude increase in the rate constant, e.g., for dopamine (DA) k pH5.5 = 1,200 M –1 s –1 versus k pH7.4 = 170,000 M –1 s –1 , and such rate acceleration is attributed to a fast electron transfer from the DA anion to dpph • . We also proved that at pH 7.0 DA breaks the peroxidation chain of methyl linoleate in liposomes assembled from neutral and negatively charged phospholipids. In contrast to no inhibitory effect during peroxidation in non-ionic emulsions, in bilayers one molecule of DA traps approximately four peroxyl radicals, with a rate constant k inh >10 3 M –1 s –1 . Our results from a homogeneous system and bilayers prove that catecholamines act as effective, radical trapping antioxidants with activity depending on the ionization status of the catechol moiety, as well as microenvironment: organization of the lipid system (emulsions vs bilayers) and interactions of catecholamines with the biomembrane.

Concentration-dependent HAT/ET mechanism of the reaction of phenols with 2,2-diphenyl-1-picrylhydrazyl (dpph˙) in methanol

“…However, in ionization supporting solvents (water and alcohols) the HAT mechanism is “contaminated” by electron transfer (ET) from the ionized fraction of ArOH. This two-step mechanism is described as Sequential Proton-Loss Electron Transfer (SPLET), 3,4 with k ET >> k HAT , see Chart 1A , and even traces of ArO − causing an enormous increase of overall rate of reaction 1 . 2 Mixed mechanism in alcohols excludes dpph˙ as mimetic to peroxyl radicals but, regardless of this controversy, 5 reaction 1 is one of the most frequently used colorimetric assays employed for quick assessment of the antiradical abilities of natural and synthetic compounds, with more than 40 thousand papers published during the last decade (SCOPUS, keywords “dpph” and “antioxidant”).…”

Mutual Activation of Two Radical Trapping Agents: Unusual “Win–Win Synergy” of Resveratrol and TEMPO during Scavenging of dpph• Radical in Methanol

“…Another explanation for the enhanced antiradical reactivity of nitroxyl mixed with RSV is an acid–base equilibrium, which is immediately established after mixing both compounds in methanol, reaction . Due to large differences in p K a for RSV and TEMPOH •+ (Table ), the equilibrium is strongly shifted to the left; however, even traces of the products are kinetically significant, as previously described for phenols reacting via the SPLET mechanism; ,,, thus, electron transfer (reaction ) would be responsible for a great acceleration of the reaction rate. RSV − + dpph · → ET RSV · + dpph − …”

“…Reaction was monitored for series of concentrations of RTA being always in stoichiometric excess over the constant initial concentration of dpph • . Pseudo-first order experimental rate constants were converted into bimolecular k S from the linear least-square slopes derived from the plots of k exp versus [RTA], eq . , k exp = k normalS [ RTA ] + const …”

“…The large acceleration for RSV cannot be justified as an effect of increased fraction of non-H bonded RSV in methanol compared to ethyl acetate because the ratio k MeOH / k EtOAc calculated from empirical equation for Δβ 2 H = 0.04 (see footnote b in Table ) could not be greater than 2. Therefore, acceleration observed for RSV in methanol can be assigned to another mechanism, the Sequential Proton-Loss Electron Transfer (SPLET), which is typical for dpph • reacting with phenols in polar, ionization supporting solvents like water and alcohols. ,,, SPLET includes three steps: (i) ionization of a phenol and (ii) electron transfer from phenoxide anion to the dpph • radical to form dpph – , which undergoes protonation (iii) to give dpph-H. The addition of acetic acid to methanol reduces the reaction rate exactly to the level for the pure HAT process, and k MeOH ≈ 2 k EtOAc , as predicted for Δβ 2 H = 0.04, see Table and ref .…”

“…Stopped-flow measurements were made following the procedure described elsewhere. , The decay of dpph • (concentration between 6 and 9 × 10 –5 M) in the stoichiometric excess of the tested compounds (concentration between 1 and 2 × 10 –2 M) was monitored at 517 nm on an Applied Photophysics SX 20 stopped-flow apparatus, equipped with a xenon arc lamp source and photodiode array detector. The mixing cell (2 mm optical path length, dead-time of mixing was 1.1 ms) and the tubes with the reactants were thermostated at 25 °C.…”