Electron Paramagnetic Resonance Study on Free Radical Scavenging and/or Generating Activity of Dopamine-4-O-sulfate.

Abstract

With the increasing understanding of the cytotoxicity of free radicals and other reactive oxygen species as a major cause of human disease and aging, the roles of endogenous antioxidants have drawn attentions. Uric acid, 1) vitamin E, 2) vitamin C, 3) ubiquinone, 4) SH-compounds, 5) bilirubin, 6) and pyruvate 7) are recognized as endogenous non-enzymatic antioxidants, while catalase, superoxide dismutase and glutathione peroxidase are known to be enzymatic antioxidants.Dopamine is the immediate metabolic precursor of norepinephrine and epinephrine. Dopamine interacts with dopamine receptors, then exerts its pharmacological effects.In human and experimental animals, dopamine is present almost entirely as a sulfate conjugated form in the circulation, 8,9) and very little of the free form is presented in the plasma.10) The physiological roles of sulfate conjugated dopamine are still unclear. Pharmacologically, conjugated dopamine has convulsive effects, 11) inhibits aldosterone secretion from cultured bovine adrenal cells, 12) and induces a constriction of some vascular beds. 13) One study, however, found that sulfate conjugated dopamine has no physiological activity. 14)From the viewpoint of free radical chemistry, dopamine can serve electrons via its oxidation pathway (Chart 1). This enables dopamine to act as both an antioxidant and a prooxidant. It acts as an antioxidant by scavenging free radicals. [15][16][17] Recently, it was found that dopamine spontaneously react with molecular oxygen to produce reactive oxygen species and semiquinone dopamine radical, 18,19) which are toxic to catecholamine neurons. [20][21][22][23] However, there is no evidence that sulfate conjugated dopamine acts as an antioxidant or a prooxidant. Therefore, in the present report, we compared dopamine and dopamine-4-O-sulfate for susceptibility to enzymatic and non-enzymatic oxidization in vitro using alkaline solution, horseradish peroxidase, UV irradiation, hydroxyl radical, superoxide anion radical, and sodium periodate. ExperimentalChemicals Dopamine, hypoxanthine, xanthine oxidase (from butter milk), and peroxidase (from horseradish) were purchased from Sigma Chemical Co., (MO, U.S.A.), hydrogen peroxide from Wako Pure Chemical Industries Ltd. (Osaka, Japan), and sodium metaperiodate from Acros Organics (NJ, U.S.A.). Dopamine-4-O-sulfate was kindly donated by Kyowa Hakko Kogyo Co., Ltd. (Tokyo, Japan). Other reagents were of the highest quality available from Wako Pure Chemical Industries Ltd.Free Radical Analysis Using EPR Spectroscopy The free radical metabolites of dopamine and dopamine-4-O-sulfate were examined with an EPR spectrometer (JES-TE 300, JEOL Co., Ltd., Tokyo, Japan) equipped with a cavity and an aqueous quartz flat cell (JEOL Co., Ltd., Tokyo, Japan) at X-band (9.5 GHz). EPR data were analyzed by a computer (HP Apollo 9000 Series 400) with the software, ESPRIT 432 ( JEOL Co., Ltd., Tokyo, Japan). Typical operating conditions of the EPR spectrometer were: power, 5 mW; center field, 3360 G; sweep width, 20 G; modulat...

Oxidation of 3,4-Dihydroxyphenylacetaldehyde, a Toxic Dopaminergic Metabolite, to a Semiquinone Radical and an ortho-Quinone

“…The hyperfine splitting constants, reported for the first time for DOPAL, are shown in Table 1. The coupling constants determined for DA and DOPAC are in good agreement with those reported in the literature (6,36). The g values for DA, DOPAL, and DOPAC are reported in Table 1.…”

Stability of baicalin in biological fluids in vitro

“…Some authors reported that phenoxyl radicals were transient, except at higher pH [10]. Partly because of the short lives of radical species [12] and less radicals formed, no obvious EPR signal was observed in the aqueous buffer at pH 7.4. Due to the similarity of their structures, scutellarin and baicalein (the aglycone of baicalin) could also undergo this degradation reaction at pH 9.0 (data not shown).…”

Oxidation and reactivity of 3,4-dihydroxyphenylacetaldehyde, a reactive intermediate of dopamine metabolism

“…The experimentally determined g-value for benzosemiquinone was 2.00677 G. Compared to the literature [176] value of 2.00469 G, this resulted in a correction factor of -0.00208 G. This corrective factor was then applied to all experimentally measured g-values. The corrected g-values are summarized in (Table 3) are in good agreement with the literature [105,175]. Figure 35.…”

“…The g-value for a DOPAL radical has not been previously reported. The experimentally determined g-values for DA and DOPAC match those reported in the literature for similar compounds under similar experimental conditions[105,175,177,178].In summary, the dopaminergic radicals formed (DOPAL, DOPAC, DA) from the experimental methods utilized (Mg 2+ -stabilized oxidation with tyrosinase, auto-oxidation with pH 10.5) were successfully simulated using WinSim and WinSim2002 software.…”