Carbodiimides are important crosslinkers in organic synthesis and are used in the isocyanate industry as modifier additives. Therefore, the understanding of their formation is of high importance. In this work, we present a theoretical B3LYP/6-31G(d) and SMD solvent model and experimental investigation of the formation of diphenylcarbodiimide (CDI) from phenyl isocyanate using a phosphorus-based catalyst (MPPO) in ortho-dichlorobenzene (ODCB) solvent. Kinetic experiments were based on the volumetric quantitation of CO2 evolved, at different temperatures between 40 and 80 °C. Based on DFT calculations, we managed to construct a more detailed reaction mechanism compared to previous studies which is supported by experimental results. DFT calculations revealed that the mechanism is composed of two main parts, and the rate determining step of the first part, controlling the CO2 formation, is the first transition state with a 52.9 kJ mol-1 enthalpy barrier. The experimental activation energy was obtained from the Arrhenius plot (ln k vs. 1/T) using the observed second-order kinetics, and the obtained 55.8 ± 2.1 kJ mol-1 was in excellent agreement with the computational one, validating the complete mechanism, giving a better understanding of carbodiimide production from isocyanates.

BACKGROUND: This study investigated the effect of carbodiimide (EDC) combined with Clearfil SE self-etch adhesive on the shear bond strength (SBS), crosslinking degree, denaturation temperature, and enzyme activity of dentin in vitro.
METHODS: Collected human sound third molars were randomly divided into different groups with or without EDC treatment (0.01-1 M). The specimens (n = 16)were stored for 24 h (immediate) or 12 months (aging) before testing the SBS. Fine dentin powder was obtained and treated with the same solutions. Then the crosslinking degree, denaturation temperature (Td), and enzyme activity were tested. Statistical analysis was performed using a one-way analysis of variance (ANOVA) to compare the differences of data between groups (α = 0.05).
RESULTS: There was a significant drop in immediate SBS and more adhesive fracture of 1.0 M EDC group, while there were no significant differences among the other groups. SEM showed a homogeneous interface under all treatments. After 12 months of aging, the SBS significantly decreased. Less decreases of SBS in the 0.3 and 0.5 M groups were found. Due to thermal and enzymatical properties consideration, the 0.3 and 0.5 M treatments also showed higher cross-link degree and Td with lower enzyme activity.
CONCLUSIONS: 0.3 and 0.5 M EDC may be favorable for delaying the aging of self-etch bond strength for 12 months. But it is still needed thoroughly study.

The process of crosslinking improves the physicochemical properties of biopolymer-based composites, making them valuable for biomedical applications. EDC/NHS-crosslinked collagen materials have a significant potential for tissue engineering applications, due to their enhanced properties and biocompatibility. Chemical crosslinking of samples can be carried out in several ways, which is crucial and has a direct effect on the final properties of the obtained material. In this study, the effect of crosslinking conditions on the properties of collagen films using EDC and NHS was investigated. Studies included FTIR spectroscopy, AFM, swelling and degradation tests, mechanical testing and contact angle measurements. Evaluation of prepared collagen films indicated that both crosslinking agents and crosslinking conditions influenced film properties. Notable alternations were observed in the infrared spectrum of the sample, to which EDC was added directly to the fish collagen solution. The same sample indicated the lowest Young modulus, tensile strength and breaking force parameters and the highest elongation at break. All samples reached the maximum swelling degree two hours after immersion in PBS solution; however, the immersion-crosslinked samples exhibited a significantly lower degree of swelling and were highly durable. The highest roughness was observed for the collagen film crosslinked with EDC, whereas the lowest was observed for the specimen crosslinked with EDC with NHS addition. The crosslinking agents increased the surface roughness of the collagen film, except for the sample modified with the addition of EDC and NHS mixture. All films were characterized by hydrophilic character. The films\' modification resulted in a decrease in their hydrophilicity and wettability. Our research allows for a comparison of proposed EDC/NHS crosslinking conditions and their influence on the physicochemical properties of fish collagen thin films. EDC and NHS are promising crosslinking agents for the modification of fish collagen used in biomedical applications.

The problem of antimicrobial resistance is becoming a daunting challenge for human society and healthcare systems around the world. Hence, there is a constant need to develop new antibiotics to fight resistant bacteria, among other important social and economic measures. In this regard, murepavadin is a cyclic antibacterial peptide in development. The synthesis of murepavadin was undertaken in order to optimize the preparative protocol and scale-up, in particular, the use of new activation reagents. In our hands, classical approaches using carbodiimide/hydroxybenzotriazole rendered low yields. The use of novel carbodiimide and reagents based on OxymaPure® and Oxy-B is discussed together with the proper use of chromatographic conditions for the adequate characterization of peptide crudes. Higher yields and purities were obtained. Finally, the antimicrobial activity of different synthetic batches was tested in three Pseudomonas aeruginosa strains, including highly resistant ones. All murepavadin batches yielded the same highly active MIC values and proved that the chiral integrity of the molecule was preserved throughout the whole synthetic procedure.

Biochemical systems make use of out-of-equilibrium polymers generated under kinetic control. Inspired by these systems, many abiotic supramolecular polymers driven by chemical fuel reactions have been reported. Conversely, polymers based on transient covalent bonds have received little attention, even though they have the potential to complement supramolecular systems by generating transient structures based on stronger bonds and by offering a straightforward tuning of reaction kinetics. In this study, we show that simple aqueous dicarboxylic acids give poly(anhydrides) when treated with the carbodiimide EDC. Transient covalent polymers with molecular weights exceeding 15,000 are generated which then decompose over the course of hours to weeks. Disassembly kinetics can be controlled using simple substituent effects in the monomer design. The impact of solvent polarity, carbodiimide concentration, temperature, pyridine concentration, and monomer concentration on polymer properties and lifetimes has been investigated. The results reveal substantial control over polymer assembly and disassembly kinetics, highlighting the potential for fine-tuned kinetic control in nonequilibrium polymerization systems.

A key to understanding the roles of RNA in regulating gene expression is knowing their structures in vivo. One way to obtain this information is through probing the structures of RNA with chemicals. To probe RNA structure directly in cells, membrane-permeable reagents that modify the Watson-Crick (WC) face of unpaired nucleotides can be used. Although dimethyl sulfate (DMS) has led to substantial insight into RNA structure, it has limited nucleotide specificity in vivo, with WC face reactivity only at adenine (A) and cytosine (C) at neutral pH. The reagent 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC) was recently shown to modify the WC face of guanine (G) and uracil (U). Although useful at lower concentrations in experiments that measure chemical modifications by reverse transcription (RT) stops, at higher concentrations necessary for detection by mutational profiling (MaP), EDC treatment leads to degradation of RNA. Here, we demonstrate EDC-stimulated degradation of RNA in Gram-negative and Gram-positive bacteria. In an attempt to overcome these limitations, we developed a new carbodiimide reagent, 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide methiodide (ETC), which we show specifically modifies unpaired Gs and Us in vivo without substantial degradation of RNA. We establish ETC as a probe for MaP and optimize the RT conditions and computational analysis in Escherichia coli Importantly, we demonstrate the utility of ETC as a probe for improving RNA structure prediction both alone and with DMS.

Crosslinking is usually required to improve the mechanical properties and stability of collagen-based scaffolds. Introducing exogenous crosslinks into collagen may however affect the collagen structure. Since the architecture of collagen is tied to its functionality, it is important to study the effect of crosslinking and to select a crosslinking method that preserves both the collagen structure and mechanical properties. The objective of this study is to compare the effect of various crosslinking methods on the structure and mechanical properties of bioartificial tendon-like materials (collagen multifilament bundles) fabricated by contact drawing. We examine both physical (ultraviolet light, UVC) and chemical (genipin, carbodiimide (EDC), and glutaraldehyde) crosslinking methods. The presence of collagen and the formation of well-ordered collagen structures are confirmed by attenuated total reflectance Fourier-transform infrared spectromicroscopy and wide-angle X-ray scattering for all crosslinking methods. The morphology of the collagen multifilament bundles is similar across crosslinking methods. Swelling of the multifilament bundles is dramatically reduced following crosslinking and varies by crosslinking method, with genipin- and carbodiimide-crosslinked specimens swelling the least. Ultimate tensile strength (UTS) and Young\'s modulus significantly improve for all crosslinked specimens compared to non-crosslinked specimens. Glutaraldehyde crosslinked collagen multifilament bundles display the highest UTS values ranging from 33.82±0.0 MPa to 45.59±0.76 MPa.

Particle-based systems have become a state-of-the-art method for in vitro expanding cytotoxic T cells by tailoring their surface with activating molecules. However, commonly used methods utilize facile carbodiimide chemistry leading to uncontrolled orientation of the immobilized antibodies on the particle surface that can lead to poor binding to target cells. To address this, selective coupling strategies utilizing regioselective chemical groups such as disulfide bridges offer a simple approach. In this work we present a set of methods to investigate the effect of polymeric nanoparticles, conjugated with either regioselective- or randomly-immobilized antiCD3 and antiCD28 antibodies, on the activation potential, expansion and expression of activation markers in T cells. We show that nanoparticles with well-oriented monovalent antibodies conjugated via maleimide require fewer ligands on the surface to efficiently expand T cells compared to bivalent antibodies randomly-immobilized via carbodiimide conjugation. Analysis of the T cell expression markers reveal that the T cell phenotype can be fine-tuned by adjusting the surface density of well-oriented antibodies, while randomly immobilized antibodies showed no differences despite their ligand density. Both conjugation techniques induced cytotoxic T cells, evidenced by analyzing their Granzyme B secretion. Furthermore, antibody orientation affects the immunological synapse and T cell activation by changing the calcium influx profile upon activation. Nanoparticles with well-oriented antibodies showed lower calcium influx compared to their bivalent randomly-immobilized counterparts. These results highlight the importance of controlling the antibody density and orientation on the nanoparticle surface via controlled coupling chemistries, helping to develop improved particle-based expansion protocols to enhance T cell therapies.

A pH-responsive amphiphilic chitosan derivative, N-lauric-O-carboxymethyl chitosan (LA-CMCh), is synthesized. Its molecular structures are characterized by FTIR, 1H NMR, and XRD methods. The influencing factors are investigated, including the amount of lauric acid (LA), carboxymethyl chitosan (CMCh), N-(3-dimethylaminopropyl)-N\'-ethylcarbodiimide hydrochloride (EDC), and N-hydroxysuccinimide (NHS), and their molar ratio, reaction time, and reaction temperature on the substitution. The degrees of substitution (DS) of the lauric groups on the -NH2 groups are calculated based on the integrated data of 1H NMR spectra. The optimum reaction condition is obtained as a reaction time of 6 h, a reaction temperature of 80 °C, and a molar ratio of lauric acid to O-carboxymethyl chitosan to N-(3-dimethylaminopropyl)-N\'-ethylcarbodiimide hydrochloride to N-hydroxysuccinimide of 1:3:4.5:4.5, respectively. The crystallinity and initial decomposition temperature of LA-CMCh decrease, but the maximum decomposition temperature increases. The crystallinity is reduced due to the introduction of LA and the degree of hydrogen bonding among LA-CMCh molecules. LA-CMCh could self-aggregate into particles, which size and critical aggregation concentration depend on the degree of substitution and medium pH. LA-CMCh aggregates could load curcumin up to 21.70 %, and continuously release curcumin for >200 min. LA-CMCh shows nontoxicity to fibroblast HFF-1 cells and good antibacterial activity against S. aureus and E. coli, indicating that it could be used as an oil-soluble-drug carrier.

The utilization of contrast agents in magnetic resonance imaging (MRI) has become increasingly important in clinical diagnosis. However, the low diagnostic specificity of this technique is a limiting factor for the early detection of tumors. To develop a new contrast agent with a specific target for early stage tumors, we present the synthesis and characterization of a nanocontrast composed of gold nanoparticles (AuNPs), gadopentetic acid (Gd-DTPA), and epidermal growth factor (EGF). Carbodiimide-based chemistry was utilized to modify Gd-DTPA for functionalization with AuNPs. This resulted in the formation of the Au@Gd-EGF nanocontrast. The relaxation rate (1/T1) of the nanocontrast was analyzed using MRI, and cytotoxicity was determined based on cell viability and mitochondrial activity in a human breast adenocarcinoma cell line. Fourier-transform infrared spectroscopy analysis confirmed the effectiveness of carbodiimide in the formation of the Gd-DTPA-cysteamine complex in the presence of bands at 930, 1042, 1232, 1588, and 1716 cm-1. The complexes exhibited good interactions with the AuNPs. However, the signal intensity of the Au@Gd-EGF nanocontrast was lower than that of the commercial contrast agent because the r1/r2 relaxivities of the Gd-DTPA-based contrast agents were lower than those of the gadoversetamide-based molecules. The Au@Gd-EGF nanocontrast agent exhibited good biocompatibility, low cytotoxicity, and high signal intensity in MRI with active targeted delivery, suggesting significant potential for future applications in the early diagnosis of tumors.