Kinetics and Mechanism of Oxidation of N-acetyl-d-penicillamine in Acidified lodate and Aqueous Iodine

RESEARCH ARTICLE K. Chipiso, T. Duc and R.H. Simoyi, S. Afr. J. Chem., 2019, 72, 1–9, <http://journals.sabinet.co.za/content/journal/chem/>. 1 Kinetics and Mechanism of Oxidation of N-acetyl-d-penicillamine in Acidified Iodate and Aqueous Iodine a Kudzanai Chipiso , Trang Duc a § a a,b, and Reuben H. Simoyi * Department of Chemistry, Portland State University, Portland, OR 97207-0751, USA. b School of Chemistry and Physics, University of KwaZulu-Natal, Westville Campus, Durban, 4000, South Africa. Received 17 January 2018, revised 26 September 2018, accepted 26 September 2018. ABSTRACT The oxidation of the biologically-active thiol, N-acetyl-d-penicillamine (NDPen) by acidified iodate and aqueous iodine has been studied. The stoichiometry of the reaction is 1:1 in which the thiol (RSH) is oxidized to its sulfonic acid (RSO3H) without cleavage of the C-S bond. In excess acidified iodate the reaction displayed a short induction period, followed by formation of aqueous iodine. Overall stoichiometry in excess iodate was 6:5: 6 IO3– + 5 RSH + 6 H+ ® 5 RSO3H + 3 I2(aq) + 3 H2O. The direct reaction of aqueous iodine and was relatively fast, over within 100 ms and had a stoichiometry of 1:3: 3 I2(aq) + RSH + 3 H2O ® RSO3H + 6 I– + 6 H+. The reaction is essentially diffusion-controlled and our stopped-flow with a mixing time limitation of 1.00 ms could only catch the reaction of the last iodine molecule. This reaction is, however, strongly inhibited by the product of reaction, I–. This is due to the formation of the relatively inert triiodide anion: I2(aq) + I– ® I3–. Mass spectrometry results showed that the reaction proceeds through the sulfinic acid with negligible stabilization of the sulfenic acid. In excess of reductant, the dimeric species is the favoured product due to a rapid condensation-type reaction between the electrophilic unstable sulfenic acid and unreacted thiol. KEYWORDS Biological thiols, bioactivation, oxidations, oxyhalogen chemistry. 1. Introduction N-acetyl-d-penicillamine (NDPen), an acetylated analog of d-penicillamine (DPen), is one of the chelating agents used clinically as an antidote to metal toxicity.1,2 Heavy metals are ubiquitous in the environment and are associated with serious health effects which include neurological diseases such as Alzheimer ’s and Parkinson disease.3–5 Chelation therapy is one of the methods in clinical use to reduce those toxic effects.6,7 Some people receive exposure to mercury (Hg) from dental amalgam fillings, although the toxicological consequences of such exposure are still debatable.8,9 NDPen has been recommended and used in the treatment of mercury poisoning as it was found to be more effective than D-penicillamine (DPen) and other agents in mercury extraction.2,10 Chelating agents act by selectively binding toxic metals which are then excreted from the body11. However, some chelating agents are toxic and this limits their use and effectiveness12. For example, the toxicity of Dpen is characterized by alterations in dermal elastic tissue, hypersensitivity reactions such as urticarial and morbilliform, as well as lupus-like autoimmune reactions, which are not associated with NDPen.13 Apart from chelation therapy, NDPen has also been used in the treatment of cystinuria, a condition in which there is excess cysteine resulting in painful cystine stones in the bladder and kidney.14 NDPen also reacts with the free cysteine to form a mixed disulfide. This N-acetyl-d-penicillamine-cysteine disulfide is highly soluble in water; about 500 times that of cystine and 10 times that of D-penicillamine-cystine.15 Thus, it makes NDPen more effective in extracting free cysteine. Reactions of NDPen and acidified nitrites to form the thionitrite (SNAP) have been extensively studied and reported in * To whom correspondence should be addressed. E-mail: literarure.16,17 S-nitrosothiols have found relevance in biology of vascular homeostasis, neurotransmission and inflammation.18 S-nitroso-N-acetyl-penicillamine (SNAP) generates nitric oxide (NO) spontaneously, thus, it is an important NO donor and is involved in vasodilation of veins and arteries, along with inhibition of platelet aggregation.19 There are other metabolic reactions that take place in the complex physiological environment which can also reduce bioavailability of therapeutic agents.20 Thiols, for example that of DPen, are easily oxidized to a disulfide. Though minor, the extent to which such side-reactions take place may alter the therapeutic value of the chelating agents. Albeit in low concentrations, oxyhalogen species such as hypohalous acids; HOCl, HOBr and HOI, which are present in the physiological environment, have been observed to oxidize thiols. The kinetics and mechanism of oxidation of NDpen by acidified iodate and aqueous iodine is reported here. 2. Experimental Procedures 2.1. Materials The following reagent grade chemicals were used without further purification: sodium iodate, perchloric acid (70–72 %), ISSN 0379-4350 Online / ©2019 South African Chemical Institute / http://saci.co.za/journal DOI: https://doi.org/10.17159/0379-4350/2019/v72a1 RESEARCH ARTICLE K. Chipiso, T. Duc and R.H. Simoyi, S. Afr. J. Chem., 2019, 72, 1–9, <http://journals.sabinet.co.za/content/journal/chem/>. sodium iodide, iodine, sodium perchlorate, soluble starch, sodium thiosulfate (Fisher) and N-acetyl-d-penicillamine (Sigma). Iodine solutions, being volatile, were kept capped and standardized spectrophotometrically before each set of experiments. Stock solutions of N-acetyl-d-penicillamine were prepared just before use. 2.2. Methods The rapid reactions of NDPen with iodine were followed on a Hi-Tech Scientific™ SF61-DX2 double-mixing stopped-flow spectrophotometer. These reactions were monitored by following consumption of iodine at 460 nm (e = 770 M–1 cm–1). NDPen has no absorbance in the visible region, while aqueous iodine has an isolated peak at 460 nm, which is its isosbestic point with triiodide, I3– (see Fig. 1). Thus, absorbance at this peak was used for analytical determination of aqueous iodine. Slower reactions involving oxidation of NDPen by iodate were monitored on a conventional Perkin-Elmer Lambda 25 UV-Vis spectrophotometer. All kinetics experiments were performed at 25.0 ± 0.1 °C and at an ionic strength of 1.00 M (NaClO4). All solutions were prepared with doubly-distilled deionized water from a Barnstead Sybron Corporation water purification unit capable of producing both distilled and deionized water (Nanopure). Mass spectra of product solutions were taken on a Thermo Scientific LTQ-Orbitrap Discovery mass spectrometer (San Jose, CA) equipped with an electrospray ionization source operated in the positive mode. 2.2.1. Stoichiometric Determinations The stoichiometry of the reaction was determined both spectrophotometrically and titrimetrically. NDPen has no measurable absorbance in the visible region (spectrum (a), Fig. 1). Aqueous iodine has an (...truncated)