The Effects of Antioxidant Nutraceuticals on Cellular Sulfur Metabolism and Signaling

Abstract

Significance: Nutraceuticals are ingested for health benefits, in addition to their general nutritional value. These dietary supplements have become increasingly popular since the late 20th century and they are a rapidly expanding global industry approaching a half-trillion U.S. dollars annually. Many nutraceuticals are promulgated as potent antioxidants. Recent Advances: Experimental support for the efficacy of nutraceuticals has lagged behind anecdotal exuberance. However, accumulating epidemiological evidence and recent, well-controlled clinical trials are beginning to support earlier animal and in vitro studies. Although still somewhat limited, encouraging results have been suggested in essentially all organ systems and against a wide range of pathophysiological conditions. Critical Issues: Health benefits of "antioxidant" nutraceuticals are largely attributed to their ability to scavenge oxidants. This has been criticized based on several factors, including limited bioavailability, short tissue retention time, and the preponderance of endogenous antioxidants. Recent attention has turned to nutraceutical activation of downstream antioxidant systems, especially the Keap1/Nrf2 (Kelch like ECH associated protein 1/nuclear factor erythroid 2-related factor 2) axis. The question now becomes, how do nutraceuticals activate this axis? Future Directions: Reactive sulfur species (RSS), including hydrogen sulfide (H₂S) and its metabolites, are potent activators of the Keap1/Nrf2 axis and avid scavengers of reactive oxygen species. Evidence is beginning to accumulate that a variety of nutraceuticals increase cellular RSS by directly providing RSS in the diet, or through a number of catalytic mechanisms that increase endogenous RSS production. We propose that nutraceutical-specific targeting of RSS metabolism will lead to the design and development of even more efficacious antioxidant therapeutic strategies. Antioxid. Redox Signal. 38, 68-94.

Keywords: Nrf2; ROS; RSS; garlic; lipoic acid; oxidative stress; polyphenols; sulfur metabolism.

FIG. 1.

Pathways for H₂S and per- polysulfide production in cells. (A) H₂S is primarily derived from l-CysS (S indicates reactive sulfur atoms) or to a lesser extent from l-MetS by the enzymes CSE, CBS, CAT, and 3-MSTS. CAT transfers the sulfur from l-CysS to pyruvate producing 3-MPS, which then forms 3-MSTSS, 3-MPS can also be produced from d-CysS by DAO. H₂S is released from 3-MSTSS by AA, DHLA, or Trx. H₂S can also be produced from COS by CA (although see Steiger et al, 2018), from thiosulfate by Trx or DHLA, from l-CysS by ST1, or by PLP plus free or heme-bound iron (Fe³⁺). H₂S can also be released from a variety of persulfides by numerous nucleophiles. (B) Per- and polysulfides are produced by direct reaction with NO, from Fe3⁺, Hb, Mb, SOD, and Cat, often with initial formation of a thiyl radical (HS^•). (C) Per- and polysulfides produced by exchange reactions with 3-MSTSS. (D) Per- and polysulfides produced by CBS and CSE metabolism of cystine transported into cells by X_c- and AT2. (E) Mitochondrial CARS2 produces CysSS that is translocated to the cytosol, where CARS1 affixes it to tRNA for incorporation into a protein (Prot). CARS 1 may also form additional Cys persulfides. (F) Hydrolysis equilibrium of polysulfides forms both neutrophilic and electrophilic compounds that may rearrange to form a variety of other compounds. Electrophiles {E}, required for these reactions shift the equilibrium to the right. (G) Polysulfide oxidation produces H₂S₂O₃ and S⁰, and the latter can react with H₂S to produce other polysulfides. 3-MPS, 3-mercaptopyruvate; 3-MSTS, 3-mercaptopyruvate sulfur transferase; AA, ascorbate; AT2, sodium-coupled neutral amino acid transporter; CA, carbonic anhydrase; CARS1, cytosolic cysteinyl-tRNA synthetase; CARS2, mitochondrial cysteinyl-tRNA synthetase; CAT, cysteine aminotransferase; Cat, catalase; CBS, cystathionine β-synthase; COS, carbonyl sulfide; CSE, cystathionine γ-lyase; CysSSCys, cystine; DAO, d-amino acid oxidase; d-CysS, d-cysteine; DHLA, dihydrolipoic acid; GSHS, glutathione; H₂S, hydrogen sulfide; H₂S₂O₃, thiosulfate; Hb, methemoglobin; l-CysS, l-cysteine; l-MetS, l-methionine; Mb, metmyoglobin; NO, nitric oxide; PLP, pyridoxyl phosphate; ProtS, protein reactive sulfur; S⁰, elemental sulfur; SOD, superoxide dismutase; ST1, sulfur transferase 1, tRNA, transfer RNA; Trx, thioredoxin; X_c-, cystine/glutamate antiporter. Bold S indicates reactive sulfur.

FIG. 2.

Metabolism of ROS (H₂O₂) and RSS (H₂S_n) by endogenous antioxidants. (A) GSH is sulfenylated by H₂O₂ to form GSOH or GSSG. (B) SOD dismutes O₂^•− to H₂O₂ and O₂ whereas it oxidizes H₂S to polysulfides (H₂S_n; n = 2–5). (C) Catalase dismutes H₂O₂ to H₂O and O₂; in the presence of O₂, it oxidizes H₂S to H₂S_n (C), whereas in hypoxia (D) it uses NADPH to produce H₂S from select sulfur-containing substrates (RS). (E) Thiol switch pathways, such as the Trx/TrxR pathway, transfer electrons from NADPH to reduce H₂O₂ or sulfenylated proteins (Prot) or Prx or to produce H₂S from inorganic or organic per and polysulfides (H₂S_n, Prot-S_n). GSH, glutathione; GSSG, oxidized glutathione dimer; H₂O₂, hydrogen peroxide; H₂S_n, hydrogen polysulfide; O₂^•−, superoxide; Prx, peroxiredoxin; ROS, reactive oxygen species; RSS, reactive sulfur species; TrxR, thioredoxin reductase.

FIG. 3.

Reactive sulfur and signaling by Allium and Brassica species. (A) Alliinase is released by damaged garlic cells and catalyzes the formation of allicin from alliin. Allicin then decomposes into mono-, di-, and tri-diallyl sulfides (DAS, DADS, and DATS, respectively). (B) DATS may react with low-molecular-weight thiols (GSH, Cys) to produce a variety of glutathione and cysteine allylsulfides and allylpersulfides, and these may then be reduced by glutathione reductase and NADPH to produce H₂S, or they may exchange the sulfane sulfur with other thiols (R and R′ signifying H, GSH or Cys) to produce a variety of polysulfides. (C) Persulfidation of Keap1 disassociates it from Nrf2, and the latter initiates the ARE. (D) Sulforaphane from Brassica also binds to Keap1 and liberates Nrf2. (E) Ajoene, another sulfur molecule found in Allium. ARE, antioxidant response elements; DADS, diallyl disulfide; DAS, diallyl sulfide; DATS, diallyl trisulfide; Keap1, Kelch like ECH associated protein 1; Nrf2, nuclear factor erythroid 2-related factor 2.

FIG. 4.

Possible mechanisms of antioxidant activity by LA and DHLA. (1) LA taken up by cells (on right) can oxidize cysteine on Keap1, thereby liberating Nrf2 and allowing it to activate ARE in the cell nucleus. (2) LA taken up by cells can be reduced by Trx, Grx, and Prx to DHLA. DHLA can release H₂S from thiosulfate (S₂O₃²⁻) produced during the course of protein (Prot) catabolism and cysteine degradation to sulfate (SO₄²⁻). The H₂S can then be oxidized to persulfide (H₂S₂), which will oxidize Keap1 cysteine and liberate NrF2. (3) DHLA can also liberate H₂S from 3-MST-SS. Other possible, but yet to be identified mechanisms include intracellular recycling of LA to DHLA (4), or oxidation of Keap1 (5), or oxidation of H₂S to persulfide (H₂S₂) (6), which will also liberate Nrf2 from Keap1. 3-MST-SS, 3-mercaptopyruvate sulfur transferase persulfide; Grx, glutaredoxin; H₂S₂, hydrogen persulfide; LA, lipoic acid.

FIG. 5.

(A–R) Nutraceutical polyphenols and quinones described in this review. Panel A shows the basic structure of the flavonoid and the three rings, A, B and C; R indicates substitution sites for H, OH, or OCH₃.

FIG. 6.

Suggested mechanisms for H₂S oxidation by polyphenols and quinones and formation of various polysulfides and thiosulfate from thiyl radicals and superoxide. (A) A quinol is autoxidized to a semiquinone, which then oxidizes H₂S to a thiyl radical, reducing the semiquinone back to the quinol. The semiquinone may also undergo a second autoxidation to the quinone, and this could oxidize another H₂S. It is also possible that the quinone undergoes a two-electron oxidation of H₂S to produce elemental sulfur. Single-electron autoxidation of the quinol and semiquinone will also produce superoxide. (B) A quinone is reduced by a hydropersulfide-forming persulfide and semiquinone radicals, and the semiquinone is reoxidized by oxygen-producing superoxide. (C) Two semiquinones can be formed by comproportionation of a quinone and hydroquinone in buffer under physiological conditions. (D) A few of the possible reactions catalyzed by quinones. Two thiyl radicals can combine to form persulfide (1); a thiyl radical can combine with a hydrosulfide anion to produce a persulfide radical (2), which can then combine with another thiyl radical to produce a 3-sulfur polysulfide (3). The persulfide radical can also react with oxygen to produce persulfide and superoxide (4) or persulfide can react with oxygen to produce thiosulfate and water (5).

FIG. 7.

Phase I and II transformations of resveratrol, epicatechin ,and quercetin. (A) In Phase I biotransformation, resveratrol can be hydroxylated at the 3′ position by CYP450 to form the hydroquinone piceatannol or in a Phase II biotransformation it may be sulfated by SULT to 3- and 4′-sulfated isomers. (B) Epicatechin undergoes Phase II transformations of both methylation by COMT and sulfation by SULT on the B-ring. (C) In Phase II biotransformation, quercetin can be sulfated by SULT on the 3′ position of the B-ring. COMT, catechol-O-methyltransferase; SULT, sulfate transferase.

FIG. 8.

EGCG is susceptible to a wide variety of Phase I and II biotransformations. The gallate ester is cleaved by esterases to form EGC. Both EGCG and EGC can be glucuronidated or methylated. EGC is also susceptible to oxidation, which forms a Michael acceptor that reacts with Michael donors such as glutathione or cysteine (RS). EGC, epigallocatechin; EGCG, epigallocatechin gallate.