Role of thiols (cysteine and teiourea) in the mechanism of the periodic decomposition reaction of hydrogen peroxide by potassium iodate in an acid medium

We have investigated the kinetics of the oxidation of cysteine and thiourea by an excess of potassium iodate under conditions corresponding to the production of auto-oscillations of concentration. First-order rate constants have been calculated for the disulfide oxidation of cystine by potassium iodate and of the disulfide of formamidine, the primary oxidation product of thiourea. It is shown that thorough oxidation of disulfides by iodate can occur and its stoichiometry is investigated. A new oscillating chemical system has been discovered, containing potassium iodate, hydrogen peroxide, thiourea, and a mixture of sulfuric and hydrochloric acids.Translated from Teoreticheskaya i Eksperimental'naya Khimiya, Vol. 26, No. 1, pp. 51–56, January–February, 1990.     

Oxidation and decomposition kinetics of thiourea oxides

In this report the decomposition and oxidation kinetics of thiourea dioxide and thiourea trioxide are investigated with a reversed-phase ion-pair high-performance liquid chromatography (HPLC) method. The HPLC method allows us to simultaneously determine and quantify several sulfur-containing reagents such as (NH2)2CSO2, (NH2)2CSO3, (NH2)2CO, and SO3(2-) (HSO3-). Experiments illustrate that the decomposition of thiourea oxides is a first-order reaction, in which the rate constants increase with the pH of the solution. Oxidations of thiourea oxides by hydrogen peroxide are first order with respect to both reagents within the studied pH range between 4.0 and 8.0. Oxidation rate constants are measured under different pH conditions, which show that increasing the pH of the reaction solution significantly accelerates the oxidation process.     

S-oxygenation of thiocarbamides II: Oxidation of trimethylthiourea by chlorite and chlorine dioxide

The kinetics of the oxidation of a substituted thiourea, trimethylthiourea (TMTU), by chlorite have been studied in slightly acidic media. The reaction is much faster than the comparable oxidation of the unsubstituted thiourea by chlorite. The stoichiometry of the reaction was experimentally deduced to be 2ClO2- + Me2N(NHMe)C=S + H2O --> 2Cl- + Me2N(NHMe)C=O + SO4(2-) + 2H+. In excess chlorite conditions, chlorine dioxide is formed after a short induction period. The oxidation of TMTU occurs in two phases. It starts initially with S-oxygenation of the sulfur center to yield the sulfinic acid, which then reacts in the second phase predominantly through an initial hydrolysis to produce trimethylurea and the sulfoxylate anion. The sulfoxylate anion is a highly reducing species which is rapidly oxidized to sulfate. The sulfinic and sulfonic acids of TMTU exists in the form of zwitterionic species that are stable in acidic environments and rapidly decompose in basic environments. The rate of oxidation of the sulfonic acid is determined by its rate of hydrolysis, which is inhibited by acid. The direct reaction of chlorine dioxide and TMTU is autocatalytic and also inhibited by acid. It commences with the initial formation of an adduct of the radical chlorine dioxide species with the electron-rich sulfur center of the thiocarbamide followed by reaction of the adduct with another chlorine dioxide molecule and subsequent hydrolysis to yield chlorite and a sulfenic acid. The bimolecular rate constant for the reaction of chlorine dioxide and TMTU was experimentally determined as 16 +/- 3.0 M(-1) s(-1) at pH 1.00.     

Electrooxidation of Formamidine Disulfide Simultaneously Investigated by On-Line High Performance Liquid Chromatography and Cyclic Voltammetry

The electro-oxidation of formamidine disulfide, an important sulfur-containing compound, was simultaneously investigated with on-line high-performance liquid chromatography and cyclic voltammetry. Using a home-made microporous sampler located at the electrode interface, the solution on the electrode surface was in situ sampled and analyzed. The electrochemical scanning was synchronously performed, which allowed the electro-oxidation products to be detected at a given potential. The main products on the surface of platinum electrode were found to be thiourea, formamidine sulfinic acid, cyanamide, and elemental sulfur. Forced convection arising from the sampling played an important role in the electrochemical oxidation. The extraction of electrode surface solution promoted the renewal of reactant and its intermediates, which induced the change of cyclic voltammetry curve. The forced convection also contributed to the redox peak current of the species on the cyclic voltammetry curves through the change of concentration of reactant and its intermediates. This technique can help to explore the reaction mechanism of complex electrochemical reactions.     

Oxydationsprodukte von Thioharnstoff

Oxidation Products of Thiourea The oxidation of thiourea with hydrogen peroxide forms in dependence of test conditions thiourea S-dioxide (H₂N)₂C? SO₂ or in the presence of HCl dithiobisformamidinium dichloride [(H₂N)₂C? S? S? C(NH ₂)₂]Cl₂. Their electron absorption, infrared, nuclear magnetic resonance, and mass spectra are communicated. Both compounds as well as unstable bis(formamidine)disulfane do not react with carbon disulfide to produce the corresponding dithiocarbamate neither in the presence of help-bases. Also no reaction happens with alkyl esters of chlorodithioformic acid.     

S-oxygenation of thiocarbamides. 3. Nonlinear kinetics in the oxidation of trimethylthiourea by acidic bromate

The oxidation of trimethylthiourea (TMTU) by acidic bromate has been studied. The reaction mimics the dynamics observed in the oxidation of unsubstituted thiourea by bromate with an induction period before formation of bromine. The stoichiometry of the reaction was determined to be 4:3, thus 4BrO(3)- + 3R(1)R(2)C=S+ 3H(2)O --> 4Br- + 3R(1)R(2)C=O + 3SO(4)(2-) + 6H+. This substituted thiourea is oxidized at a much faster rate than the unsubstituted thiourea. The oxidation mechanism of TMTU involves initial oxidations through sulfenic and sulfinic acids. At the sulfinic acid stage, the major oxidation pathway is through the cleavage of the C-S bond to form a reducing sulfur leaving group, which is easily oxidized to sulfate. The minor pathway through the sulfonic acid produces a very stable intermediate that is oxidized only very slowly to urea and sulfate. The direct reaction of aqueous bromine with TMTU was faster than reactions that form bromine, with a bimolecular rate constant of (1.50 +/- 0.04) x 10(2) M(-1) s(-1). This rapid reaction ensured that no oligooscillatory bromine formation was observed. The oxidation of TMTU was modeled by a simple reaction scheme containing 20 reactions.     

Characterization by tandem mass spectrometry of stable cysteine sulfenic acid in a cysteine switch peptide of matrix metalloproteinases

Cysteine sulfenic acid (Cys-SOH) is an elusive intermediate in reactive oxygen species-induced oxidation reactions of many proteins such as peroxiredoxins and tyrosine phosphatases. Cys-SOH is proposed to play a vital role in catalytic and signaling functions. The formation of cysteine sulfinic acid (Cys-SO(2)H) and cysteine sulfonic acid (Cys-SO(3)H) has been implicated in the activation of matrix metalloproteinase-7 (MMP-7) and oxidation of thiol to cysteine sulfinic acid has been associated with the autolytic cleavage of MMP-7. We have examined the formation of cysteine sulfenic acid in a synthetic peptide PRCGVPDVA, which is a cysteine switch domain of MMP-7 and other matrix metalloproteases. We have prepared the cysteine sulfenic acid containing peptide, PRC(SOH)GVPDVA, by reaction with hydroxyl radicals generated by the Fenton reaction (Fe(+2)/H(2)O(2)). We characterized this modified peptide by tandem mass spectrometry and accurate mass measurement experiments. In addition, we used 7-chloro-4-nitrobenzo-2-oxa-1,3-diazol (NBD-Cl) reagent to form an adduct with PRC(SOH)GVPDVA to provide additional evidence for the viability of PRC(SOH)GVPDVA in solution. We also characterized an intramolecular cysteine sulfinamide cross-link product PRC[S(O)N]GVPDVA based on tandem mass spectrometry and accurate mass measurement experiments. These results contribute to the understanding of a proteolytic cleavage mechanism that is traditionally associated with MMP activation.     

Kinetics and mechanism of the [Ru(III)(edta)(H2O)](-)-mediated oxidation of cysteine by H2O2

The kinetics and mechanism of the [Ru(III)(edta)(H(2)O)](-)-mediated oxidation of cysteine (RSH) by hydrogen peroxide (edta(4-) = ethylenediaminetetraacetate), were studied in detail as a function of both the hydrogen peroxide and cysteine concentrations at pH 5.1 and room temperature. The kinetic traces reveal clear evidence for a catalytic process in which hydrogen peroxide reacts directly with cysteine coordinated to the Ru(III)(edta) complex in the form of [Ru(III)(edta)SR](2-). A parallel process in which [Ru(III)(edta)(H(2)O)](-) first reacts with H(2)O(2) to produce [Ru(V)(edta)O](-) and subsequently oxidizes cysteine, is orders of magnitude slower than the [Ru(III)(edta)(H(2)O)](-)-mediated oxidation in which cysteine rapidly coordinates to [Ru(III)(edta)(H(2)O)](-) prior to the reaction with H(2)O(2). HPLC product analyses revealed the formation of cystine (RSSR) as major product along with cysteine sulfinic acid (RSO(2)H) in the reaction system, and established the catalytic role of [Ru(III)(edta)(H(2)O)](-). Simulations were performed to account for the rather complex kinetic traces in terms of the suggested reaction mechanism. The results of the simulations support the proposed reaction mechanism that involves the oxidation of coordinated cysteine to cysteine sulfenic acid (RSOH), which subsequently rapidly reacts with H(2)O(2) and RSH to form RSO(2)H and RSSR, respectively.     

Glutathione oxidation in real time by thermospray liquid chromatography-mass spectrometry

The oxidation and reduction of glutathione and oxidized glutathione were studied in real time by liquid chromatography-mass spectrometry during exposure to hydrogen peroxide and mercaptoethanol. By mass spectrometry mixed disulfides and both reversible and irreversible oxidations of sulfur to higher states (sulfinic and sulfonic acids) were directly observed during exposure to hydrogen peroxide. The irreversible oxidation of glutathione to glutathione sulfonic acid could be detected after 30 min exposure of glutathione to 40 mM H2O2 at 20 degrees C. A peak consistent with glutathione-sulfinic acid was transiently present, suggesting this compound behaved as an oxygen consuming antioxidant. Liquid chromatography-mass spectrometry appears to be an excellent method to study oxidation and reductions of sulfur containing peptides and amino acids.     

Kinetics and mechanism of the oxidation of thiourea by chlorine dioxide

The stoichiometry and kinetics of the oxidation of thiourea (SC(NH₂)₂) by chlorine dioxide (ClO₂) have been studied by uv-vis spectrophotometry using conventional and stopped-flow mixing techniques at 25.0 ± 0.1°C, pH 0.3–4.8. In high acid and initial 10:1 molar ratio of thiourea to chlorine dioxide, thiourea is oxidized relatively rapidly to dithiobisformamidine ion ((NH₂)₂CSSC(NH₂)₂²⁺), which slowly decomposes to thiourea, sulfur, and cyanamide (NCNH₂). In high acid and excess ClO₂, thiourea is oxidized to relatively stable formamidine sulfinic acid ((NH) (NH₂)CSO₂H). In high acid and molar ratios of ClO₂ to thiourea of 5:1 and higher, some oxidation to formamidine sulfonic acid ((NH) (NH₂)CSO₃H) occurs. At lower acidity, along with Cl^?, the major ClO₂ reduction product, byproduct sulfate is detected and, at pH < 3, ClO₂^?, also, appears. Kinetics data were collected for high excess thiourea with varying pH. The [ClO₂]-time curves are straight lines with negative slopes that increase in magnitude with increasing [thiourea]. The dependence on [thiourea] is first-order; the dependence on [ClO₂] is zero-order for 90% of reaction. With decreasing pH, the rate increases and the disappearance of ClO₂ becomes autocatalytic. Studies of the effects of reaction products on the rate of reaction lead to the conclusion that autocatalysis at low pH is due to the greater reactivity of HClO₂ compared with ClO₂^?. A 10-step mechanism incorporating a slow one-electron transfer from thiourea to ClO₂ to generate the (NH) (NH₂)CS · radical and subsequent more rapid reactions has been constructed and implemented in a computer simulation which provides a reasonably accurate fit to the observed kinetics curves. © 1993 John Wiley & Sons, Inc.     

Effect of thiourea on the radical polymerization of vinyl monomers. Part II. Effect of diphenyl thiourea on the polymerization of methyl methacrylate with benzoyl peroxide as initiator

The polymerization of methyl methacrylate (MMA) initiated by benzoyl peroxide (BPO) in the presence of diphenyl thiourea (DPTU) has been studied. It was found that the BPO–DPTU catalyst system was not an effective accelerating system but showed a relatively strong retarding effect. With DPTU derivatives, the polymerization rate was found to decrease with the increase in the electron-attracting forces of substituents attached to the phenyl groups of DPTU. In the polymerization of MMA initiated by AIBN, the addition of DPTU to the reaction systems affected neither the polymerization rate nor the degree of polymerization. From this fact, it might be concluded that DPTU itself serves as a radical scavenger. It seems most probable from the results of kinetic studies, iodometric titration, and from the effect of an oxidation product of DPTU (diphenyl formamidine disulfide) that the retardation effect observed is attributable to the action of the disulfide (DPFDS). By extending the Alfrey-Price scheme for the copolymerization reactions to the chain-transfer reactions, the Q and e values of DPFDS were determined. The apparent chain-transfer constants for DPTU and its derivatives were calculated by means of rate measurements and were correlated with substituent constants. The mechanism of the polymerization is discussed on the basis of these results.     

Reactive sulfur species: aqueous chemistry of sulfenyl thiocyanates

Sulfenyl thiocyanate (RSSCN) derivatives of penicillamine (PENSCN) and glutathione (GSSCN) have been synthesized in situ at pH = 0 from equilibrium mixtures that consists of hypothiocyanous acid (HOSCN), thiocyanogen ((SCN)2), and trithiocyanate ((SCN)3-). The electrophilic thiocyanating agent N-thiocyanatosuccinimide (NTS) also reacts with PEN and GSH to yield the corresponding RSSCN derivatives. PENSCN and GSSCN were characterized by NMR, ES-MS, and IR spectroscopy. While stable at pH = 0, at higher pH the RSSCN derivatives decompose to give products that are consistent with hydrolysis and formation of reactive sulfenic acids.     

Thermal rearrangement of 1,3-thiazolidine sulfoxides: Thiolsulfinate and thioaldehyde intermediates

Stereospecific ring opening of the sulfoxides cis- 13 and trans- 14 in refluxing toluene gave the corre sponding sulfenic acids 9 , 10 intermediates respectively. The sulfenic acid 9 dimerized to the thiolsulfinate 17 by dual function of the sulfenic acid as S-nucleophile/S-electrophile with loss of water while the sulfenic acid 10 was unchanged. The stereospecific recyclization of 10 to the parent sulfoxide 14 increases the higher pi-electron density of the double. The thermolysis of the thiolsulfinate 17 gave the transient sulfenic acid 9 , which dimerized again to repeat the process and unisolable thioaldehyde 21 . The thioaldehyde 21 was con verted to either pyrrole 15 by the action of a sulfinic acid 20 catalyst formed inevitably by hydrolysis of 17 under the reaction conditions, or thiazole 18 under neutral conditions. In these rearrangements, the amide carbonyl group facilitated the elimination of a neighboring hydrogen.     

Microwave Activation of Processes in Mesopores: The Thiourea Electrooxidation at Mesoporous Platinum

In situ microwave activation is applied to the electrochemical oxidation of thiourea at low surface area (polished polycrystalline) and at high surface area (electrodeposited mesoporous platinum coated) platinum microelectrodes. The one electron oxidation of thiourea to formamidine disulfide is monitored as a function of the different activation parameters. In the absence of microwaves (ambient temperature, low mass transport) increasing the surface area (roughness) increases the thiourea oxidation response predominantly due to adsorption effects. In the presence of high microwave intensities, high mass transport and thermal effects further increase the oxidation current at mesoporous platinum. The most effective thickness of the mesoporous platinum film for the thiourea oxidation process is estimated as 3 μm independent of electrode diameter, temperature, or mass transport effects.     

Norbornene probes for the study of cysteine oxidation

Cysteine residues on proteins can react with cellular oxidants such as hydrogen peroxide. While this process is important for scavenging excess reactive oxygen species, the products of this oxidation may also mediate cell signalling. To understand the role of cysteine oxidation in biology, selective probes are required to detect and quantify its occurrence. Cysteine oxidation products such as sulfenic acids are sometimes unstable and therefore short-lived. If such cysteine derivatives are to be analysed, rapid reaction with the probe is required. Here we introduce norbornene derivatives as probes for cysteine oxidation, and demonstrate their ability to trap sulfenic acids. The synthesis of norbornene derivatives containing alkyne or biotin affinity tags are also reported to facilitate the use of these probes in chemical biology and proteomics.     

Fragmentation of protonated ions of peptides containing cysteine,cysteine sulfinic acid,and cysteine sulfonic acid

The oxidation of the sulfhydryl group in cysteine to sulfenic acid, sulfinic acid, and sulfonic acid in proteins is important in a number of enzymatic processes. In this study we examined the fragmentation of four peptides containing cysteine, cysteine sulfinic acid (Cys-SO(2)H), and cysteine sulfonic acid (Cys-SO(3)H) in an ion-trap mass spectrometer. Our results show that the presence of a Cys-SO(2)H in a peptide leads to preferential cleavage of the amide bond at the C-terminal side of the oxidized cysteine residue. The results are important for the determination of the site of the cysteine oxidation and might be useful for the sequencing of cysteine-containing peptides.     

Reactive sulfur species: kinetics and mechanisms of the oxidation of cysteine by hypohalous acid to give cysteine sulfenic acid

Cysteine sulfenic acid has been generated in alkaline aqueous solution by oxidation of cysteine with hypohalous acid (HOX, X = Cl or Br). The kinetics and mechanisms of the oxidation reaction and the subsequent reactions of cysteine sulfenic acid have been studied by stopped-flow spectrophotometry between pH 10 and 14. Two reaction pathways were observed: (1) below pH 12, the condensation of two sulfenic acids to give cysteine thiosulfinate ester followed by the nucleophilic attack of cysteinate on cysteine thiosulfinate ester and (2) above pH 10, a pH-dependent fast equilibrium protonation of cysteine sulfenate that is followed by rate-limiting comproportionation of cysteine sulfenic acid with cysteinate to give cystine. The observation of the first reaction suggests that the condensation of cysteine sulfenic acid to give cysteine thiosulfinate ester can be competitive with the reaction of cysteine sulfenic acid with cysteine.     

Simultaneous tracking of sulfur species in the oxidation of thiourea by hydrogen peroxide

We employ reversed-phase ion-pair high-performance liquid chromatography to quantitatively characterize the oxidation kinetics of thiourea oxidation by hydrogen peroxide. The HPLC technique makes it possible to monitor the concentrations of a variety of sulfur-containing species with different oxidation states and to elucidate the relative phase relations among them. The experimental results are in good agreement with simulations from an 8-step reaction mechanism.     

Post-translational modification of Rhodococcus R312 and Comamonas NI1 nitrile hydratases

Nitrile hydratases (NHases) are industrially significant iron- and cobalt-containing enzymes used in the large-scale synthesis of acrylamide. Previous reports have shown that the active site peptides of NHases are post-translationally modified by oxidation of cysteine residues, and that these modifications are essential for catalysis. We report mass spectrometric evidence of the oxidation states of the active site cysteines in the iron coordination spheres of two iron-containing nitrile hydratases, namely R312 NHase from Rhodococcus rhodochrous strain R312 and NI1 NHase from Comamonas testosteroni. At least one of these cysteines is oxidised to a sulfinic acid (SO(2)H) and there is also evidence suggesting an additional oxidation to a sulfenic acid (SOH). This is the first evidence for the presence of these oxidation states for full-length NHases and for Fe-NHases from different microorganisms. The presence of these covalent modifications was confirmed by performing mass spectrometry on the active site peptide of R312 NHase, under native, reduced and carboxymethylated conditions. We also show the nitrosylation of the iron by mass spectrometry, as well as the release of NO by photoirradiation.     

Degradation kinetics of telithromycin determined by HPLC method

The degradation kinetics of the antibiotic telithromycin using a stability-indicating high-performance liquid chromatography (HPLC) method is demonstrated. The photodegradation is performed by UVC lamp-254 nm (15W), installed in a chamber internally coated with mirrors, where telithromycin solutions prepared from coated tablets are placed in quartz cells. To promote oxidation, the reaction between the telithromycin solution and 3% hydrogen peroxide solution is carried out. The kinetics parameters of order of reaction and the rate constants of the degradation are determined for both conditions. The degradation process of telithromycin can be described by first-order kinetics under both experimental conditions used in this study. The results reveal the photo and oxidation lability of the drug and confirm the reliability of HPLC method for telithromycin in the presence of its degradation products.