Chemical Modification Of Carboxyl Groups Research Articles

Polyacrylic Acid Nanoplatforms: Antimicrobial, Tissue Engineering, and Cancer Theranostic Applications.

Polyacrylic acid (PAA) is a non-toxic, biocompatible, and biodegradable polymer that gained lots of interest in recent years. PAA nano-derivatives can be obtained by chemical modification of carboxyl groups with superior chemical properties in comparison to unmodified PAA. For example, nano-particles produced from PAA derivatives can be used to deliver drugs due to their stability and biocompatibility. PAA and its nanoconjugates could also be regarded as stimuli-responsive platforms that make them ideal for drug delivery and antimicrobial applications. These properties make PAA a good candidate for conventional and novel drug carrier systems. Here, we started with synthesis approaches, structure characteristics, and other architectures of PAA nanoplatforms. Then, different conjugations of PAA/nanostructures and their potential in various fields of nanomedicine such as antimicrobial, anticancer, imaging, biosensor, and tissue engineering were discussed. Finally, biocompatibility and challenges of PAA nanoplatforms were highlighted. This review will provide fundamental knowledge and current information connected to the PAA nanoplatforms and their applications in biological fields for a broad audience of researchers, engineers, and newcomers. In this light, PAA nanoplatforms could have great potential for the research and development of new nano vaccines and nano drugs in the future.

The attachment opioid antagonist drug of Naltrexone to multi-walled carbon nanotubes by different kinds of reagents carbodiimides

Surface functionalization of multi-walled carbon nanotubes(MWCNTs) by opioid antagonist drug of Naltrexone via chemical modification of carboxyl groups,using of reagents di cyclohexyl carbodiimide(DCC),1-ethyl-3[3-dimethylaminopropyl] carbodiimide hydrochloride(EDC),N-hydroxysuccinimide(NHS),1-hydroxybenzotriazolemonohydrate(HOBT),O-(Benzotriazole-1-yl)-1,1,3,3-tetramethyluraniumtetrafluoroborate(TBTU),were performed. In synthetic organic chemistry,compounds containing the carbodiimide functionality are dehydration agents and are often used to activate carboxylic acids towards amide or ester formation. Carboxylic acids will react with the carbodiimide to produce the key intermediate as O-acylisourea which can be considered as a carboxylic ester with an activated leaving group.We have used of five carbodiimide reagent to synthesis twelve sample in this work.The resulting materials were characterized by different techniques,such as Fourier transform infraredspectroscopy (FT-IR), Thermogravimetricanalysis (TGA),Raman spectroscopy,Scanning Electron Microscopy (SEM),Field Emission Scanning Electron Microscopy(FESEM),Atomic Force Microscopy(AFM),High performance liquid chromatography(HPLC).

Preparation, Morphology and Properties of Acyl Azide-Functionalized Carbon Nanotubes

Multi-walled carbon nanotubes (MWNTs) functionalized with acyl azide groups were prepared via chemical modification of carboxyl groups introduced on the carbon nanotube surface. The acyl azides MWNTs have potential application in chemistry functionalization of MWNTs and energetic materials. The functionalizations of the MWNTs were characterized in detail using scanning electron microscope (SEM), transmission electron microscopy (TEM), Fourier Transform Infrared Spectroscopy (FTIR), X-ray photoelectron spectroscopy (XPS), Raman spectroscopy and thermogravimetry (TG). The results confirm the carboxylic acid groups converted to the desired acid azide functionalization of MWNTs.

Solubilization of Multi Walled Carbon Nanotubes Under a Facile and Mild Condition

Multi-walled carbon nanotubes (MWCNTs) functionalized with amino groups was prepared via chemical modification of carboxyl groups introduced on the carbon nanotube surfaces. Oxidation of MWCNTs was performed with ozone in aqueous phase and amidation of generated carboxylic groups, was occurred with amines in the presence of HATU as a coupling agent. Obtained functionalized MWCNTs are soluble in many common organic solvents. The functionalized MWCNTs were characterized in detail using FTIR-ATR, Raman CHN and SEM methods.

Direct electrochemistry of catalase at amine-functionalized graphene/gold nanoparticles composite film for hydrogen peroxide sensor

Direct electrochemistry and electrocatalysis of catalase (Cat) was studied based on a nano-composite film consisting of amine functionalized graphene and gold nanoparticles (AuNPs) modified glassy carbon electrode. Graphene was synthesized chemically by Hummers and Offeman method and then was functionalized with amino groups via chemical modification of carboxyl groups introduced on the graphene surface. The nano-composite film showed an obvious promotion of the direct electron transfer between Cat and the underlying electrode, which attributed to the synergistic effect of graphene-NH2 and AuNPs. The resultant bioelectrode retained its biocatalytic activity and offered fast and sensitive H2O2 quantification. Under the optimized experimental conditions, hydrogen peroxide was detected in the concentration range from 0.3 to 600μM with a detection limit of 50nM at S/N=3. The biosensor exhibited some advantages, such as short time respond (2s), high sensitivity (13.4μA/mM) and good reproducibility (RSD=5.8%).

Immobilization–stabilization of glucoamylase: Chemical modification of the enzyme surface followed by covalent attachment on highly activated glyoxyl-agarose supports

Commercial glucoamylase immobilizes very slowly on highly activated glyoxyl-agarose supports. The resulting derivatives were only 6-fold more stable than soluble enzyme. The unmodified glucoamylase, highly glycosylated, seems to have a low number of Lys able to react with glyoxyl groups on the support. Thus, the enzyme surface was highly enriched in amino groups by chemical modification of carboxyl groups (activated with carbodiimide) with ethylenediamine. The aminated enzyme preserves a good percentage of activity (80%) and it exhibits the same stability than unmodified enzyme. The aminated enzyme was immobilized very rapidly on highly activated glyoxyl-agarose support. The new resulting derivatives preserved 50% of activity and were more than 500-fold more stable than soluble enzyme in experiments of thermal inactivation.

Attachment of biomolecules (protein and DNA) to amino-functionalized carbon nanotubes

An efficient method for the attachment of biomolecules [e.g. bovine serum albumin (BSA) protein and deoxyribonucleic acid (DNA)] to amino-group-functionalized multiwalled carbon nanotubes (f-MWCNTs) was reported. MWCNTs were prepared by spray pyrolysis of a benzene-ferrocene solution in argon atmosphere at ∼850 °C followed by functionalization with an amino group by chemical modification of carboxylic groups introduced on the nanotube surface. This process involves a direct coupling of ethylenediamine with carboxylic groups to introduce amino groups by amide formation. The as-synthesized MWCNTs, f-MWCNTs, and amino f-MWCNTs with BSA protein and DNA were characterized by scanning and transmission electron microscopy, and Fourier transform infrared spectroscopy, which confirm the attachment of biomolecules (BSA protein and DNA) to the amino f-MWCNTs.

Removal of cadmium from aqueous solutions by oxidized and ethylenediamine-functionalized multi-walled carbon nanotubes

Surface functionalization of multi-walled carbon nanotubes (MWCNTs) by ethylenediamine, via chemical modification of carboxyl groups, using O-(7-azabenzotriazol-1-yl)-N,N,N′,N′-tetramethyluronium hexafluorophosphate, was performed. The resulting materials were characterized by different techniques, such as FTIR, TGA and elemental analysis. Biocompatibility studies showed that the functionalized MWCNTs, at concentrations between 1 and 50 μg mL−1, were not cytotoxic for the fibroblast L929 cell line. In batch tests, the influences of solution pH, contact time, initial metal ion concentration and temperature on the sorption of Cd2+ ions onto raw-MWCNTs (raw-MWCNT), oxidized MWCNTs (o-MWCNT) and ethylenediamine-functionalized MWCNTs (e-MWCNT) were studied. The adsorption of Cd2+ ions by o-MWCNT and e-MWCNT was strongly pH dependent. The time dependent Cd2+ sorption onto raw-MWCNT, o-MWCNT and e-MWCNT can be described by a pseudo-second-order kinetic model. The Langmuir isotherm model agrees well with the equilibrium experimental data. The maximum capacity was obtained for e-MWCNT, 25.7 mg g−1, at 45 °C. The thermodynamic parameters were also deduced for the adsorption of Cd2+ ions on raw-MWCNT, o-MWCNT and e-MWCNT and the results showed that the adsorption was spontaneous and endothermic.

Synthesis, characterization and cytotoxicity of surface amino-functionalized water-dispersible multi-walled carbon nanotubes

Surface functionalization of multi-walled carbon nanotubes (MWCNTs), with amino groups via chemical modification of carboxyl groups introduced on the nanotube surface, using O-(7-azabenzotriazol-1-yl)- N, N, N′, N′-tetramethyluronium hexafluorophosphate (N-HATU) and N, N-diisopropylethylamine (DIEA) is reported. The N-HATU coupling agent provides faster reaction rate and the reaction occurs at lower temperature compared to amidation and acylation–amidation chemistry. The amines, 1,6-hexanediamine (HDA), diethylenetriamine (DETA), triethylenetetramine (TETA) and 1,4-phenylenediamine (PDA) were used. The resulting materials were characterized with different techniques such as FTIR, XRD, elemental analysis, TGA, TEM, UV–vis spectroscopy and cyclic voltammetry. MWCNTs functionalized with PDA posses the best dispersibility and electron transfer properties in comparison to the others amines. Functionalized MWCNTs, at the concentrations between 1 and 50 μg ml −1, were not cytotoxic for the fibroblast L929 cell line. However, the concentrations of MWCNTs higher of 10 μg ml −1 reduced cell growth and this effect correlated positively with the degree of their uptake by L929 cells.

A novel stationary phase based on amino derivatized nanotubes for HPLC separations: Theoretical and practical aspects

A novel column based on silica containing immobilized carbon nanotubes (CNTs) was developed and evaluated in terms of its binding efficiency and resolution. First, CNT functionalized with amino groups (CNT-NH(2)) were prepared via chemical modification of carboxylic groups introduced on the CNT surface. Secondly the covalent immobilization of CNT-NH(2) was carried out by using glutardialdehyde activating agent on aminopropyl (AP)-silica surface. This CNT stationary phase was applied to the HPLC separation of two molecule series, i.e. polychlorinated biphenyl (PCB) isomers with different degrees of substitution in the ortho-position (non-ortho to tetra-ortho substituted) and terpenes (linalool, geraniol, thymol, alpha-terpineol). The retention behavior of these solute molecules was measured under isocratic conditions with different mobile phase compositions, ranging from 0.05-0.70 v/v of toluene in cyclohexane. The retention factors of the solute molecule do not depend linearly on the toluene fraction but follow a quadratic relationship. This CNT stationary phase was a very useful column for the separation of PCB congeners and terpenes. It was demonstrated that a planar conformation of the solute molecule enhanced the solute retention on this CNT stationary phase. As well, a quantitative structure relationship derived, demonstrated the significant input to retention was due to the structurally selective dipole-dipole and charge transfer interactions with the solutes. These results were compared with those obtained on the AP stationary phase. The proposed CNT stationary phase for the separation possess distinctive and interesting retentive properties, and chemometric analysis of retention data of appropriate designed series of test solutes appears to be a convenient, objective and quantitative method to prove a new phase specificity.

The Studies on Reaction Control and Development of New Practical Reagents Based on Characteristics of Reaction Field

Reaction rates and selectivities can be critically affected by the reaction field. Using a diverse set of reagents and reaction systems, the author reviews a variety of ways to control the reaction field. In the first example, we discuss 4-(4,6-dimethoxy-1,3,5-triazin-2-yl)-4-methylmorpholinium chloride (DMT-MM), which serves as an exceptionally convenient reagent for dehydrocondensation. In particular, formation of carboxamide by DMT-MM was found to take place even if water or alcohol were used as a reaction medium. Thus, chemical modification of carboxyl groups and/or amino groups of highly polar substrates, such as amino acid derivatives, peptides, glyco-chains, and nucleotides, can be simply effected by mixing them with DMT-MM in aqueous or alcoholic solvents. The author also found that a tertiary amine catalyzes the activation step of carboxylic acid with 2-chloro-4,6-dimethoxy-1,3,5-triazine (CDMT) via in situ generation of coupling reagents. Proceeding further in this direction, we determined that artificial acyltransferase and cyclotransferase could be formed by conjugation of a tertiary amine catalyst to host-molecules to mimic a substrate binding site. Finally, the micellar interface, well known for promoting hydrolysis, clearly provides a superior reaction field for dehydrocondensation. When a 1,3,5-triazine-type amphiphilic dehydrocondensing agent was used, bimolecular dehydrocondensation between amphiphilic carboxylate and amines was highly accelerated (2000-fold) in a micellar system. Spontaneous membrane fusion was induced by adoption of the micellar reaction in the ceramide synthesis at the surface of membranes. All together, these diverse findings strongly support the central importance of the reaction field in controlling reaction rates and selectivity.

Chemical modification results in hyperactivation and thermostabilization ofFusarium solaniglucoamylase

Chemical modification of carboxyl groups of glucoamylase from a mesophilic fungus, Fusarium solani, was carried out using ethylenediamine as nucleophile in the presence of water-soluble 1-ethyl-3(3-dimethylaminopropyl)carbodiimide. Modification brought about a dramatic enhancement of catalytic activity and thermal stability of glucoamylase. Temperature and pH optima of ethylenediamine-coupled glucoamylase (ECG) increased as compared with those of native enzyme. The specificity constant (k(cat)/K(m)) of native, ECG-2, ECG-11, and ECG-17 was 136, 173, 225, and 170, respectively, at 55 degrees C. The enthalpy of activation (Delta H*) and free energy of activation (Delta G*) for soluble starch hydrolysis were lower for the chemically modified forms. All of the modified forms were stable at higher temperatures and possessed high Delta G* against thermal unfolding. The effects of alpha-chymotrypsin and subtilisin on the modified forms were activating as compared with native. Moreover, denaturation of ECG-2, ECG-11, and ECG-17 in urea at 4 mol x L(-1) also showed an activation trend. A possible explanation for the thermal denaturation of native and increased thermal stability of ECG-2, ECG-11, and ECG-17 at higher temperatures is also discussed.

Chemically modified thrombin and anhydrothrombin that differentiate macromolecular substrates of thrombin

Thrombin is a primary inducer of thrombus formation by activations of coagulation cascade and platelet aggregation. Hitherto, several types of thrombin inhibitors have been developed for therapeutic purpose. We prepared modified thrombin (M-thrombin) and modified anhydrothrombin (M-anhydrothrombin) by chemical modification of carboxyl groups of thrombin and anhydrothrombin, respectively, to present a new strategy for a potent antiplatelet-anticoagulant agent and new tools for investigation of thrombin functions. M-anhydrothrombin retained high affinity for factor VIII (FVIII), but demonstrated lower affinity than anhydrothrombin for fibrinogen and factor V (FV). Both M-anhydrothrombin and anhydrothrombin prolonged activated partial thromboplastin time (APTT) without affecting prothrombin time, and M-anhydrothrombin prolonged APTT much more than anhydrothrombin. M-anhydrothrombin also retained affinity for the recombinant extracellular domain peptide of protease-activated receptor 1 (PAR1). M-thrombin exhibited marginal clotting activity (4% of thrombin), but induced platelet aggregation in platelet-rich plasma without forming a fibrin clot, which was completely suppressed by anti-PAR1 antibody (ATAP2) and by M-anhydrothrombin, but not by anhydrothrombin. These results indicate that M-thrombin induced platelet aggregation through the activation of PAR1, and M-anhydrothrombin inhibited this process completely. In contrast, neither M-anhydrothrombin nor anhydrothrombin apparently inhibited thrombin-induced platelet aggregation. Only in the presence of the Gly-Pro-Arg-Pro (GPRP) peptide that inhibits polymerization of fibrin, M-anhydrothrombin completely inhibited thrombin-induced platelet aggregation. M-thrombin is PAR1-specific and M-anhydrothrombin is FVIII- and PAR1-specific derivatives, and thereby, are new tools as specific agonist and antagonist, respectively, of PAR1. Furthermore, M-anhydrothrombin may be an attractive model for development of a potent anticoagulant-antiplatelet agent.

“M-AAA-Thrombin”, a New Thrombin Mutant with Potent Anticoagulant and Antiplatelet Properties.

Thrombosis is a major cause of morbidity and mortality, and thrombin is a major inducer of thrombus formation. Thus several antithrombotic agents targeting thrombin have been developed. We previously reported an anticoagulant and antiplatelet thrombin derivative, ‘M-anhydrothrombin' prepared by chemical modifications. In this study, we prepared a new thrombin mutant, specificity of which was highly modulated with substantially improved antithrombotic efficacy. The thrombin mutant designated “AAA-Thrombin” in which Lys65, His43 and Ser205 in B-chain have been replaced by Ala revealed higher affinity and specificity for factor VIII with no enzymatic activity. AAA-Thrombin prolonged APTT much more than anhydrothrombin in a dose dependent manner without affecting PT and TT. Platelet aggregation induced by activation of PAR-1 was also effectively suppressed by AAA-Thrombin. “M-AAA-Thrombin” prepared by further chemical modification of carboxyl groups in AAA-Thrombin enhanced its antithrombotic efficacy. M-AAA-Thrombin (250nM) prolonged APTT approx. two times, and suppressed platelet aggregation by PAR-1 activation, while AAA-Thrombin did not at the same concentration. M-AAA-Thrombin also suppressed ristocetin-induced platelet aggregation. In vivo experiments, M-AAA-Thrombin demonstrated significant antithrombotic property in the arterio-venous shunt thrombosis model and the FeCl3-induced carotid artery thrombosis model in guinea pigs. These results indicate that M-AAA-Thrombin would be a candidate for quite an innovative anticoagulant and antiplatelet agent for both arterial and venous thromboses. Further optimization of mutagenesis and modification, in terms of efficacy and safety is in progress in our laboratory.

Amino-Functionalized Carbon Nanotubes for Binding to Polymers and Biological Systems

Single-walled carbon nanotubes (SWCNT) functionalized with amino groups were prepared via chemical modification of carboxyl groups introduced on the carbon nanotube surface. Two different approaches (amide and amine-moieties) were used to produce the amino-functionalized nanotubes. The amino-termination allows further chemistry of the functionalized SWCNTs and makes possible covalent bonding to polymers and biological systems such as DNA and carbohydrates. The functionalization of the SWCNTs was characterized in detail using FTIR and XPS.

Preparation of potent cytotoxic ribonucleases by cationization: enhanced cellular uptake and decreased interaction with ribonuclease inhibitor by chemical modification of carboxyl groups.

Carboxyl groups of bovine RNase A were amidated with ethylenediamine (to convert negative charges of carboxylate anions to positive ones), 2-aminoethanol (to eliminate negative charges), and taurine (to keep negative charges), respectively, by a carbodiimide reaction. Human RNase 1 was also modified with ethylenediamine. Surprisingly, the modified RNases were all cytotoxic toward 3T3-SV-40 cells despite their decreased ribonucleolytic activity. However, their enzymatic activity was not completely eliminated by the presence of excess cytosolic RNase inhibitor (RI). As for native RNase A and RNase 1 which were not cytotoxic, they were completely inactivated by RI. More interestingly, within the cytotoxic RNase derivatives, cytotoxicity correlated well with the net positive charge. RNase 1 and RNase A modified with ethylenediamine were more cytotoxic than naturally occurring cytotoxic bovine seminal RNase. An experiment using the fluorescence-labeled RNase derivatives indicated that the more cationic RNases were more efficiently adsorbed to the cells. Thus, it is suggested that the modification of carboxyl groups could change complementarity of RNase to RI and as a result endow RNase cytotoxicity and that cationization enhances the efficiency of cellular uptake of RNase so as to strengthen its cytotoxicity. The finding that an extracellular human enzyme such as RNase 1 could be effectively internalized into the cell by cationization suggests that cationization is a simple strategy for efficient delivery of a protein into cells and may open the way of the development of new therapeutics.

Effects of chemical modification of carboxyl groups in the hemolytic lectin CEL-III on its hemolytic and carbohydrate-binding activities.

Effects of chemical modification of carboxyl groups in the hemolytic lectin CEL-III on its activities were investigated. When carboxyl groups were modified with 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC) and glycine methyl ester, hemolytic activity of CEL-III decreased as the EDC concentration increased, accompanied by reduction of oligomerization ability and hemagglutinating activity. However, binding ability of CEL-III for immobilized lactose was retained fairly well after modification, suggesting that one of two carbohydrate-binding sites might be responsible for such inactivation of CEL-III.

Effect of chemical modification of carboxyl groups in apple residues on metal ion binding

AR (apple residue) was used as an alternative and cheap material for binding metal ions due to the presence of carboxyl and phenolic functional groups. The binding capacity of copper, lead, and cadmium by AR was pH dependent. Carboxyl groups of AR were esterified by acidic methanol to determine the contribution of carboxyl groups to metal ions binding. The extent of esterification was determined by analyzing the amount of methanol released in the sample hydrolysates by gas chromatography. The effect of esterification on binding metal ions was investigated in batch experiments by unmodified and modified AR. All esterified AR showed significant decreases in binding copper, lead and cadmium. The loss in the capacity of metal ion binding was proportional to the extent of esterification. The capacity of metal binding decreased with increase in the concentration of methanol in the respective hydrolysates or the modified AR. The data indicate that carboxyl groups on AR play an important role in the metal ion binding.

Effects of Drugs, Salts, and Phospholipid Vesicles on Hemoglobin Release from Hydrostatic Pressure-Treated Human Erythrocytes

When human erythrocytes subjected to a pressure of 2.0 kbar were incubated at 0 degree C and atmospheric pressure, hemoglobin was released from the membrane. Effects of drugs, salts, and phospholipid vesicles on hemoglobin release were examined. The hemoglobin release was suppressed by the addition of amphiphathic drugs such as chlorpromazine, trifluoperazine, tetracaine, indomethacin, and phenylbutazone or by a hypertonic condition. The suppressive effect induced by these drugs or hypertonic buffer was enhanced by the addition of aminophospholipid vesicles containing phosphatidylserine (PS) and/or phosphatidylethanolamine (PE) but decreased by vesicles composed of phosphatidylcholine (PC) and sphingomyelin. In the isotonic buffer without these drugs, such effects of vesicles were small. The suppressive effects of PS and PE were decreased by chemical modification of carboxylic groups and amino groups with 1-ethyl-3-(3-dimethylaminopropyl)carbodimide and 2,4,6-trinitrobenzenesulfonate, respectively. The size of vesicles as well as phospholipid composition was also an important factor in the suppression of hemoglobin release. The ESR spectrum of pressure-treated membrane spin-labeled with a maleimide was changed by PC vesicles but not by PS vesicles. These results indicate that the resealing of pressure-induced membrane holes by amphipathic drugs or hypertonic medium is synergistic with aminophospholipid vesicles and the hole size is affected by the vesicle-membrane protein interaction.

Reduction of Ricin Toxicity without Impairing the Saccharide-Binding Properties by Chemical Modification of the Carboxyl Groups

The usefulness of ricin as a research tool is handicapped by its extremely biohazardous nature. In this work, ricin toxicity has been reduced by chemical modification of carboxyl groups using 1-ethyl-3(3-dimethylaminopropyl) carbodiimide and [ 14C]glycine methyl ester. The reaction was carried out in 8 M urea and in the presence of 0.1 M lactose to protect the groups involved in saccharide binding. Together with carboxyl groups, tyrosine residues were also modified. The maximum modification achieved was 13 carboxyl groups and 7 tyrosines per molecule (about 30% of total carboxyls and tyrosines). The modification did not alter substantially the strength and specificity of the carbohydrate-binding ability of the lectin, as observed by hemagglutination tests and by inhibition assays with different carbohydrate structures. However, the LD 50 decreased 90-fold when the highest modification was achieved. Therefore, the modified lectin can be used more safely in the study of galactose-containing carbohydrates.