Redox and Nucleophilic Reactions of Naphthoquinones with Small Thiols and Their Effects on Oxidization of H2S to Inorganic and Organic Hydropolysulfides and Thiosulfate

Abstract

Naphthoquinone (1,4-NQ) and its derivatives (NQs, juglone, plumbagin, 2-methoxy-1,4-NQ, and menadione) have a variety of therapeutic applications, many of which are attributed to redox cycling and the production of reactive oxygen species (ROS). We previously demonstrated that NQs also oxidize hydrogen sulfide (H2S) to reactive sulfur species (RSS), potentially conveying identical benefits. Here we use RSS-specific fluorophores, mass spectroscopy, EPR and UV-Vis spectrometry, and oxygen-sensitive optodes to examine the effects of thiols and thiol-NQ adducts on H2S-NQ reactions. In the presence of glutathione (GSH) and cysteine (Cys), 1,4-NQ oxidizes H2S to both inorganic and organic hydroper-/hydropolysulfides (R2Sn, R=H, Cys, GSH; n = 2–4) and organic sulfoxides (GSnOH, n = 1, 2). These reactions reduce NQs and consume oxygen via a semiquinone intermediate. NQs are also reduced as they form adducts with GSH, Cys, protein thiols, and amines. Thiol, but not amine, adducts may increase or decrease H2S oxidation in reactions that are both NQ- and thiol-specific. Amine adducts also inhibit the formation of thiol adducts. These results suggest that NQs may react with endogenous thiols, including GSH, Cys, and protein Cys, and that these adducts may affect both thiol reactions as well as RSS production from H2S.

Reaction Mechanisms of H2S Oxidation by Naphthoquinones

“…Replacing the methoxy groups with chlorine in 2,3-dichloro-1,4-NQ (DCNQ, Figure 1H) consumed H 2 S faster than any of the other NQs. These results are similar to what we demonstrated for NQs with a single substitution on the quinoid group [13] and extend our findings to the more substituted DMNQ and DCNQ. HMNQ (2-hydroxy-3-methoxy-NQ) did not consume H 2 S, suggesting that the 2-hydroxy-3-methoxy-NQ has effects similar to the 2C hydroxyl in lawsone that does not react with H 2 S [13].…”

“…These results are similar to what we demonstrated for NQs with a single substitution on the quinoid group [13] and extend our findings to the more substituted DMNQ and DCNQ. HMNQ (2-hydroxy-3-methoxy-NQ) did not consume H 2 S, suggesting that the 2-hydroxy-3-methoxy-NQ has effects similar to the 2C hydroxyl in lawsone that does not react with H 2 S [13]. Our results also illustrate the utility of measuring H 2 S consumption to provide a more comprehensive appreciation of H 2 S metabolism by NQs.…”

“…We have previously shown that the amount of polysulfide production (measured by SSP4 fluorescence) and the rate of oxygen consumption by a variety of NQs with open positions on the C2 or C3 quinoid carbon are dependent on the concentrations of both NQ and H 2 S [12,13]. Qualitatively similar results for polysulfide production (SSP4 fluorescence) were observed with DCNQ and DMNQ (Figure 2).…”

“…Thiol and amine adducts were prepared as described previously [13]. Briefly, 1 mM GSH or Cys was added to equimolar concentrations of NQs in open containers and incubated for 1 h at room temperature to allow adduct formation and autoxidation of the NQ adduct.…”

“…We recently demonstrated that NQs may use hydrogen sulfide (H 2 S) as the reductant and oxidize it to hydroper-and hydropolysulfides (H 2 S n where n = 2-5), sulfite, and thiosulfate [12], or, in the presence of glutathione or Cys (GSH-S and Cys-S where S denotes the additional, reactive sulfur), to organic hydroper-and hydropolysulfides (GSH-S n and Cys-S n , where n = 2-4) as well as GSH-S 2 OH [13]. We also observed that GSH, Cys, and propylamine readily formed adducts with NQs and these, in turn, variously affected H 2 S oxidation by NQs.…”