Cystine Derivatives Research Articles

Smart Hydrogel Based on Derivatives of Natural α-Amino Acids for Efficient Removal of Metal Ions from Wastewater.

A novel class of hydrogels, rich in a variety of functional groups capable of interacting/complexing with metal ions was successfully synthesized. This was achieved by using acryloyl derivatives of natural α-amino acids, specifically ornithine and cystine. The δ-amino group of ornithine was modified with an acryloyl group to facilitate its attachment to the polymer chain. Additionally, N,N'-bisacryloylcystine, derived from cystine, was employed as the cross-linker. The hydrogel was obtained through a process of free radical polymerization. This hydrogel, composed only from derivatives of natural amino acids, has proven to be a competitive sorbent and has been effectively used to remove heavy metal pollutants, mainly lead, copper, and silver ions, from aqueous media. The maximum sorption capacities were ca. 155 mg·g-1, 90 mg·g-1, and 215 mg·g-1, respectively for Pb(II), Cu(II), and Ag(I). The material was characterized by effective regeneration, maintaining the sorption capacity at around 80%, 85%, and 90% for Cu(II), Ag(I), and Pb(II), respectively, even after five cycles. The properties of sorption materials, such as sorption kinetics and the effect of pH on sorption, as well as the influence of the concentration of the examined metal ions on the swelling ratio and morphology of the gel, were investigated. The EDS technique was employed to investigate the composition and element distribution in the dry gel samples. Additionally, IR spectroscopy was used to identify the functional groups responsible for binding the studied metal ions, providing insights into their specific interactions with the hydrogel.

Pnictogen bonding in imide derivatives for chiral folding and self-assembly.

Pnictogen bonding (PnB) is an attraction interaction that originates from the anisotropic distribution of electron density of pnictogen elements, which however has been rarely found in nitrogen atoms. In this work, for the first time, we unveil the general presence of N-involved PnB in aromatic or aliphatic imide groups and reveal its implications in chiral self-assembly of folding. This long-neglected interaction was consolidated by Cambridge structural database (CSD) searching as well as subsequent computational studies. Though the presence of PnB has limited effects on spectroscopic properties in the solution phase, conformation locking effects are sufficiently expressed in the chiral folding and self-assembly behavior. PnB anchors the chiral conformation to control the emergence and inversion of chiroptical signals, while intramolecular PnB induces the formation of supramolecular tilt chirality. It also enables the chiral folding of imide-containing amino acid or peptide derivatives, which induces the formation of unique secondary structural sequences such as β-sheets. Finally, the effects of PnB in directing folded helical structures were revealed. Examples of cysteine and cystine derivatives containing multiple N⋯O and N⋯S PnBs constitute an α-helix like secondary structure with characteristic circular dichroism. This work discloses the comprehensive existence of imide-involved PnB, illustrates its important role in folding and self-assembly, and sheds light on the rational fabrication of conformation-locked compounds and polymers with controllable chiroptical activities.

Reduction-triggered Disassembly of Organic Cationic Nanorods Produces a Cysteine Protease Mimic (Miravet et al.).

The emergence of catalytic activity associated with a disassembly process is reported, reminiscent of complex biological systems. A cystine derivative with pendant imidazole groups self-assembles into cationic nanorods in the presence of the cationic surfactants cetylpyridinium chloride (CPC) or cetyltrimethylammonium bromide (CTAB). Disulfide reduction triggers nanorod disassembly and the generation of a simple cysteine protease mimic, which shows a dramatically improved catalytic efficiency in the hydrolysis of p-nitrophenyl acetate (PNPA).

Degradable nanohydrogel with high doxorubicin loadings exhibiting controlled drug release and decreased toxicity against healthy cells.

A new nanogel/drug carrier of 100-150nm size, based on poly(N-isopropylacrylamide-co-sodium acrylate) and degradable crosslinker (cystine derivative), was synthesized. Using the electrostatic interactions between the carboxylic groups in the polymer network and the protonated amine groups of doxorubicin it was possible to load the drug into the carrier to a very high level of 28-30% relative to the dry mass of the polymer. The presence of the -S-S- groups made the polymer network susceptible to degradation by glutathione. The size of the nanoparticles was small enough to enable them to easily penetrate the cells. The MTT assay indicated that compared to free doxorubicin the nanogel particles loaded with doxorubicin were more cytotoxic against the MCF-7 and A2780 cancer cells, while they were 150 times less toxic against the MCF-10A healthy cells. The new carrier nanoparticles appeared also to be useful for prolonged drug delivery.

Optimized one-pot derivatization and enantioseparation of cysteine: Application to the study of a dietary supplement

Cysteine is a sulfur-containing amino acid which plays an outstanding role in many biological pathways in mammals. The analysis and quantification of native cysteine remains a critical issue due to its highly reactive thiol group evolving to the disulfide cystine derivative through oxidation reaction. Aimed at improving the derivative stability, cysteine was labelled with 4-fluoro-7-nitro-2,1,3-benzoxadiazole (NBD-F), which reacts with both amino and thiol groups. The derivatization was optimized and the chemical identity of the reaction product was assessed via high-resolution mass spectrometry. The NBD-cysteine derivative resulted stable for 10 days. This derivative was enantioresolved (α and RS equal to 1.25 and 2.70, respectively) thanks to a (R,R)-Whelk-O1 phase with the following chromatographic setting: eluent, MeOH/water-90/10 (v/v) with 15 mM ammonium formate (pwsH 6.0); column temperature, 35 °C; flow rate, 1.0 mL/min. The developed method was validated following the ICH guidelines and applied for the quality control of a L-cysteine containing dietary supplement.

Tunable environmental sensitivity and degradability of nanogels based on derivatives of cystine and poly(ethylene glycols) of various length for biocompatible drug carrier

Nanogels are promising carriers for drug delivery, especially when they degrade under conditions typical for cancer cells to release a drug. By using the precipitation polymerization we synthesized a biocompatible and degradable nanogel based on poly(ethylene glycol) methyl ether methacrylate (OEGMA) and di(ethylene glycol) methyl ether methacrylate (MEO2MA) cross-linked with a redox-sensitive linker, N,N′-bis(methacryloyl)cystine – (mBISS). The application of appropriate proportions of monomers during the nanogel synthesis allowed us to set its volume-phase-transition temperature to the body temperature while the aggregation of the nanogel in physiological conditions was eliminated. The presence of carboxylic groups in mBISS made the nanogel sensitive to pH, gave it stability under conditions of high ionic strength and allowed it to bind the anticancer drug – doxorubicin (DOX). The degradation of nanogel was examined by using electron microscopies. In the presence of a high concentration of glutathione (a reducing agent for the -S-S- bridges) which is the case for most cancer cells, the spherical shape of the nanogel and correspondingly its structure were destroyed. The obtained profiles for the release of DOX from the nanogel revealed that the smallest amount of DOX was released from the nanogel at pH 7.4, while under conditions present in most cancer cells, this is at pH 5.0 and cGSH = 40 mM, the highest cumulative release of DOX was observed. The DOX loaded nanogels were cytotoxic against MCF-27 and HT-29 cancer cells similarly to DOX alone. The unloaded gel nanoparticles did not inhibit proliferation of the cells.

Chemical fingerprints of cold physical plasmas \u2013 an experimental and computational study using cysteine as tracer compound

Reactive oxygen and nitrogen species released by cold physical plasma are being proposed as effectors in various clinical conditions connected to inflammatory processes. As these plasmas can be tailored in a wide range, models to compare and control their biochemical footprint are desired to infer on the molecular mechanisms underlying the observed effects and to enable the discrimination between different plasma sources. Here, an improved model to trace short-lived reactive species is presented. Using FTIR, high-resolution mass spectrometry, and molecular dynamics computational simulation, covalent modifications of cysteine treated with different plasmas were deciphered and the respective product pattern used to generate a fingerprint of each plasma source. Such, our experimental model allows a fast and reliable grading of the chemical potential of plasmas used for medical purposes. Major reaction products were identified to be cysteine sulfonic acid, cystine, and cysteine fragments. Less-abundant products, such as oxidized cystine derivatives or S-nitrosylated cysteines, were unique to different plasma sources or operating conditions. The data collected point at hydroxyl radicals, atomic O, and singlet oxygen as major contributing species that enable an impact on cellular thiol groups when applying cold plasma in vitro or in vivo.

Bioreducible amino acid-derived polymeric nanoparticles for delivery of functional proteins

We report the preparation of protein encapsulated amino acid derived redox-responsive nanoparticles (NPs) as effective nanocarriers for intracellular delivery of proteins. More specifically, acryloyl derivatives of lysine, ornithine, cystine and cystamine, were employed as monomers and disulfide crosslinkers for non-covalent encapsulation of model protein bovine serum albumin (BSA) and were interfacially crosslinked via free radical polymerization to form redox-responsive protein NPs. Notably, prepared NPs exhibited high protein loading content between 37 and 45%, averaged ∼400 nm in hydrodynamic size and possessed a mean surface charge of -15 mV. Furthermore, blank polymeric NPs displayed exceptional cytocompatibility with cell viability exceeding 92% at concentrations as high as 4 mg/mL, while redox-responsive protein NPs displayed glutathione (GSH)-dependent BSA release behavior in vitro. Additionally, cellular uptake studies confirmed that protein NPs entered MDA-MA-231 cells predominantly via the endocytic pathway. Upon cellular internalization, redox-responsive NPs delivered protein into cytosol of cells within 60 min demonstrating intrinsic endosomolytic characteristics and efficient protein release under cytoplasmic high GSH conditions. Most importantly, insulin analog-loaded NPs significantly increased glucose consumption in HepG2 cultures confirming protein stability and retention of protein function. Cumulatively, our approach presents a simple yet effective strategy for intracellular delivery of biologically active proteins for various biomedical applications.

Double-Tailed Cystine Derivatives as Novel Substitutes of Phospholipids with Special Reference to Liposomes.

Cystine-based gemini surfactants with dodecyl, tetradecyl, hexadecyl, and octadecyl hydrocarbon chains were synthesized, and their interactions with unsaturated (soy phosphatidylcholine, SPC)/saturated (hydrogenated SPC, HSPC) soy phosphatidylcholines in the forms of a monolayer and a model liposome were estimated for different combinations of the components in the mixed systems. Studies of Langmuir monolayers at the air-aqueous buffer interface revealed condensation of the monomolecular films with the addition of surfactants. The effect of surfactants decreased according to the following order: octadecyl > hexadecyl > tetradecyl > dodecyl homologs. The nonideal mixing between the components was estimated using the deviation of the experimental molecular area from the ideal area per molecule. The excess molecular area increased with the increase in the surfactant chain length and phospholipid saturation. The 50 mol % mixture of cystine derivatives and phospholipids formed thermodynamically stable monolayers. The surfactants increased the rigidity of SPC monolayers and decreased that of HSPC monolayers, as observed by the studies of surface dialational rheology. The film structure at the air-water interface could differentiate the SPC- and HSPC-comprising systems through the formation of organized regions, especially at a higher surface pressure. The constriction of surfactant/phospholipid hybrid vesicles was observed with an increase in the length of surfactant hydrocarbon chains. The negative zeta potential of vesicles took the highest values and did not change with time for 20 and 50 mol % surfactant. The spherical shape of the vesicles was confirmed by transmission electron microscopy. Differential scanning calorimetry revealed an increase in fluidity of HSPC bilayers and rigidity of SPS bilayers under the influence of surfactants. These effects were confirmed by fluorescence spectroscopy. All of the vesicle formulations were found to be nontoxic from the 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl-tetrazolium bromide assay, suggesting their potential as a novel membranous system for the delivery of drugs, genetic materials, vaccines, and other therapeutic agents.

ChemInform Abstract: One‐Pot Synthesis of Unsymmetrical Disulfides Using 1‐Chlorobenzotriazole as Oxidant: Interception of the Sulfenyl Chloride Intermediate.

Abstract A high-yielding and low temperature one-pot procedure is described for unsymmetrical disulfide synthesis from two different thiols using 1-chlorobenzotriazole (BtCl) as oxidant. The mechanism of the coupling involves in situ trapping of the sulfenyl chloride intermediate R 1 SCl by nucleophilic benzotriazole (BtH) to form R 1 SBt, which protects R 1 SCl from forming the homodimer R 1 SSR 1 . The methodology is applicable to all types of thiol (aliphatic, aromatic, heteroaromatic), with a variation developed for aliphatic–aliphatic couplings. Differentially N-protected cysteines couple to afford the unsymmetrical cystine derivatives in high yield (90%), which serves as a model for the one-pot intermolecular coupling of cysteine-containing peptides to form peptide disulfide heterodimers. Minimal exchange in aromatic–aromatic disulfide synthesis is noted on account of the mild conditions.

One-pot synthesis of unsymmetrical disulfides using 1-chlorobenzotriazole as oxidant: Interception of the sulfenyl chloride intermediate

A high-yielding and low temperature one-pot procedure is described for unsymmetrical disulfide synthesis from two different thiols using 1-chlorobenzotriazole (BtCl) as oxidant. The mechanism of the coupling involves in situ trapping of the sulfenyl chloride intermediate R 1SCl by nucleophilic benzotriazole (BtH) to form R 1SBt, which protects R 1SCl from forming the homodimer R 1SSR 1. The methodology is applicable to all types of thiol (aliphatic, aromatic, heteroaromatic), with a variation developed for aliphatic–aliphatic couplings. Differentially N-protected cysteines couple to afford the unsymmetrical cystine derivatives in high yield (90%), which serves as a model for the one-pot intermolecular coupling of cysteine-containing peptides to form peptide disulfide heterodimers. Minimal exchange in aromatic–aromatic disulfide synthesis is noted on account of the mild conditions.

Preparation of Arylmercapturic Acids by S‐Arylation of N,N′‐Diacetylcystine

AbstractA simple convenient method has been developed for the preparation of N‐acetyl‐S‐arylcysteines based on the Chan–Lam–Evans arylation of N,N′‐diacetylcystine dimethyl ester with arylboronic acids and used to synthesize a series of arylmercapturic acids. Unlike copper‐mediated N‐arylation, the S‐arylation of neither cysteine nor cystine derivatives proceeded satisfactorily in the presence of air.(© Wiley‐VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2009)

Facile Synthesis of β-Amino Disulfides, Cystines, and Their Direct Incorporation into Peptides

Herein, we report a simple and efficient methodology for the synthesis of beta-amino disulfides by regioselective ring opening of sulfamidates with benzyltriethylammonium tetrathiomolybdate [BnNEt3](2)MoS4. Stability and reactivity of different protecting groups under the reaction conditions have been discussed. This methodology has also been extended to serine and threonine derived sulfamidates to furnish cystine and 3,3'-dimethyl cystine derivatives.

Simple, Rapid, and Accurate RP-HPLC Method for Determination of Cystine in Human Urine after Derivatization with Dansyl Chloride

An accurate, simple, and sensitive reversed-phase high-performance liquid chromatographic method, with loratadine as internal standard (IS) and UV detection at 286 nm, has been developed for deterination of cystine in human urine. The major innovations of the method include use of acrylonitrile to protect cysteine from oxidization to cystine, separation of cysteine, as the dansyl derivative, from cystine, and use of isocratic elution instead of gradient elution to reduce the time and cost of serial analysis. The mobile phase was 0.05 M sodium acetate–methanol, 35:65 (v/v), adjusted to pH 3.5 with 2.5 M citric acid, at a flow rate of 1.0 mL min−1. The retention times of cystine and the IS were 16.6 and 19.9 min, respectively. The limit of detection for cystine was 0.3 mg L−1. Extraction recovery of cystine was >85.6%. Intra-day and inter-day precision (RSD) for cystine were below 4.3 and 8.5%, respectively. There was no chromatographic interference from other α-amino acids present in mammalian proteins, or from other urine components. The calibration plot for the cystine derivative was linear in the range 1–500 mg L−1 and the correlation coefficient was 0.9992. The method was validated appropriately and successfully used for determination of cystine in human urine.

Reactive Sulfur Species: Kinetics and Mechanism of the Oxidation of Cystine by Hypochlorous Acid to Give N,N‘-Dichlorocystine

Cystine and HOCl (a neutrophil-derived oxidant) react to form an intermediate that has a half-life of ca. 5 min at pH 7.5. The intermediate subsequently decomposes to eventually yield a mixture of cystine, higher oxides of Cys, and other uncharacterized species. Spectral titrations, transitory (1)H NMR and UV-vis spectra, and the reaction properties of the intermediate are consistent with a formulation of N,N'-dichlorocystine {NDC = [-SCH(2)CH(NHCl)(CO(2)H)](2)}. The reaction of equimolar amounts of HOCl with cystine at pH 11.3 does not yield N-chlorocystine [NCC = (-O2C)(H3N+)CHCH(2)SSCH(2)CH(NHCl)(CO(2)H)] but rather a 1:1 mixture of NDC and cystine. This result could be explained by two mechanisms: rapid disproportionation of NCC to produce NDC and cystine or a faster reaction of the second equivalent of HOCl with NCC than the first equivalent of HOCl reacts with cystine. The latter mechanism is favored because of our observation by NMR spectroscopy that NDC decomposes via a species that we have assigned as NCC. Thus, disproportionation of NCC is apparently a relatively slow process. The rates of reaction of cystine(0) = [-SCH(2)CH(NH(3)(+))(CO(2)(-))](2) degrees , cystine(1-) = [((-)O(2)C)(H(2)N)CHCH(2)SSCH(2)CH(NH(3)(+))(CO(2)(-))](-), and cystine(2-) = [-SCH(2)CH(NH2)(CO2)(-))]2(2-) have been investigated, and it is clear that cystine(0) is unreactive, whereas cystine(2-) is about four times more reactive than cystine(1-). Accordingly, the following mechanism is proposed (constants for 5 degrees C): HOCl = H+ + OCl-, pK1 = 7.47; cystine(0) = cystine(1-) + H+, pK2 = 8.15; cystine(1-) = cystine(2-) + H+, pK3 = 9.00; cystine(1-) + HOCl --> NCC(1-) + H2O, k4 = 4.3(2) x 10(6) M(-1) s(-1); cystine(2-) + HOCl --> NCC(2)(-) + H2O, k5 = 1.6(2) x 10(7) M(-1) s(-1); NCC(1-) --> NCC(2-) + H+, k6 = fast; NCC(2-) + HOCl --> NDC(2-) + H2O, k7 = fast. At physiologic pH, the k4 pathway dominates. The generation of long-lived chloramine derivatives of cystine may have physiological consequences, since such compounds are known to react with nucleophiles via mechanisms that are also characteristic of HOCl, electrophilic transfer C+.

L-Cystine inhibits aspartate-β-semialdehyde dehydrogenase by covalently binding to the essential 135Cys of the enzyme

Aspartate-β-semialdehyde dehydrogenase (ASADH) from Escherichia coli is inhibited by l- and d-cystine, and by other cystine derivatives. Enzyme inhibition is quantitatively reversed by addition of dithiothreitol (DTT), dithioerythrytol, β-mercaptoethanol, di-mercaptopropanol or glutathione to the cystine-inactivated enzyme. Cystine labeling of the enzyme is a pH dependent process and is optimal at pH values ranging from 7.0 to 7.5. Both the cysteine incorporation profile and the inactivation curve of the enzyme as a function of pH suggest that a group(s) with pK a of 8.5 could be involved in cystine binding. Stoichiometry of the inactivation reaction indicates that one cysteine residue from the enzyme subunit is reactive against cystine, as found by direct incorporation of radioactive cystine into the enzyme and by free-thiol titration of the enzyme with 5,5′-dithiobis-2-nitrobenzoic acid (DTNB) before and after the cystine treatment. One mole of cysteine is released from each mol of cystine after reaction with the enzyme. ASA, NADP and NADPH did not prevent cystine inhibition. The [ 35S]cysteine-labelled enzyme can be visualized after electrophoresis in polyacrylamide gels and further detection by autoradiography. After pepsin treatment of the [ 35S]cysteine-inactivated enzyme, a main radioactive peptide was isolated by HPLC. The amino acid sequence of this peptide was determined as FVGGN(Cys) 2TVSL, thus demonstrating that the essential 135Cys is the amino acid residue modified by the treatment with cystine.

Comparison of different chloroformates for the derivatisation of seleno amino acids for gas chromatographic analysis

Three chloroformate reagents, ethyl chloroformate (ECF), methyl chloroformate (MCF) and menthyl chloroformate (MenCF), have been used for the derivatisation of seleno amino acids and their performance was compared. Chromatographic parameters and the inertness of the different instrumental configurations used (gas chromatography–atomic emission detection (GC–AED), and GC–MS) were shown to have a significant influence on the detection of various seleno amino acids (selenomethione, selenoethione and selenocysteine) and some sulphur-containing amino acids (methionine, cysteine, cystine and methylcysteine) which were included in the experiments for comparison. Methyl chloroformate was the preferred derivatisation reagent, since it generally performed best in terms of derivatisation yield and reproducibility and also showed less significant conditioning effects than ethyl chloroformate. Methyl and ethyl chloroformate derivatives of selenomethionine, selenoethionine, cysteine and methionine were detectable, while the detection of the menthyl chloroformate derivatives of selenocystine and cystine was not reproducible. Overall efficiencies for the determination of selenomethionine and selenoethionine from aqueous extracts ranged from 40 to 100% for methyl chloroformate, over 30–75% for ethyl chloroformate to 15–70% for menthyl chloroformate for different series measured over a period of months. The relative standard deviation of the method for the methyl and menthyl chloroformate derivatisation ranged from 7 to 13% without internal standard and was improved to 2% for the determination of selenomethionine using selenoethionine as internal standard. This indicates that, despite the limited reproducibility of the method, its repeatability is good enough to allow accurate determination of seleno amino acids, which was also demonstrated by the analysis of selenium supplementation tablets for human diet that contained selenomethionine.

Intracellular redox regulation by a cystine derivative suppresses UV-induced NF-κB activation

Nuclear factor (NF)-κB pathways are influenced by the intracellular reduction–oxidation (redox) balance. While NF-κB is activated through inhibitor (I)-κB degradation by oxidative stress, its DNA binding is accelerated in the reduced state. We found that N, N′-diacetyl- L-cystine dimethylester (DACDM) suppressed the UVB-induced NF-κB binding activity at a much lower concentration (50–100 μM) than N-acetyl- L-cysteine (NAC, 10–30 mM). While NAC suppressed the I-κB degradation but not the DNA binding, DACDM prevented the activated NF-κB from binding DNA, without influencing the I-κB degradation. These properties of DACDM make it possible to effectively regulate the intracellular redox balance.

A new and versatile method for determination of thiolamines of biological importance

A method for the separation and quantitation of several important biological thiolamines is described. The procedure employs a C 18 reversed-phase HPLC system to separate the dinitrophenyl derivatives of reduced and oxidized glutathione and cysteine and relies on an internal standard, N ϵ-methyllysine, to minimize experimental error. The method was validated in three matrices (water, HepG2 cell lysates, and mouse liver homogenates) using several criteria. The detector response was linear for the dinitrophenyl derivatives of glutathione, glutathione disulfide, cysteine, and cystine in the concentrations ranging from 10 to 50 nmol/ml. Inter- and intra-day variation, percent recovery in the biological matrices, and limits of detection and quantitation were determined. For the most accurate determination, it is essential that standard curves be produced daily and in the same matrix as that being analyzed.

Chiral hydroxythiols as catalysts for enantioselective borane ketone reductions

Several chiral 1,2- and 1,3-hydroxythiols derived from ( R)-camphor, (1 S)-(+)-10-camphorsulfonyl chloride, cysteine and cystine derivatives were prepared and evaluated as catalysts in borane ketone reductions. Under the best experimental conditions (10 mol% catalyst, THF, 35°C), a 95% yield of ( R)-1-phenylethanol of 64% ee was obtained in the borane reduction of acetophenone.