Search Results (55)

Protein phosphorylation is a crucial post-translational modification that regulates protein activity, cellular signaling, transcriptional regulation, and cell cycle control. However, the analysis of phosphoproteins in biological samples is often compromised by complex sample matrices and interference from high-abundance proteins. While the top-down phosphoproteomics strategy enables comprehensive analysis of post-translational modifications based on intact proteins, its requirement for higher protein purity due to low protein ionization efficiency poses stern challenges. Consequently, developing appropriate enrichment methods for phosphoproteins in practical samples becomes essential. Immobilized metal ion affinity chromatography (IMAC) represents a common strategy for phosphorylated protein separation and enrichment. Among metal ions, Ti⁴⁺ has gained widespread application as IMAC chelating ligands due to its capacity to form multiple coordination networks and its high selectivity for phosphorylated protein enrichment, leveraging the strong chelating ability of phosphate groups toward metal ions. This paper presents the design and preparation of a novel magnetic Ti-IMAC nanocomposite, MNP@MPTMS–VPA–Ti(IV). The material is modified with phosphate groups via facile thiol-ene click chemistry and then immobilizes Ti⁴⁺, enabling selective enrichment of intact phosphoproteins through IMAC affinity. The efficiency of enrichment was evaluated using subsequent matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) for detection and analysis. This Ti-IMAC material-based magnetic solid-phase extraction (MSPE)-MALDI-TOF MS protocol has been successfully applied to enrich intact phosphoproteins in milk and eel mucus with high selectivity, sensitivity, and suitability. Full article

Although covalently crosslinked gelatin hydrogels have been investigated for use in 3D cell culture due to inherent bioactivity and proliferation within the denatured collagen precursor, the stability of the matrix, and relatively inexpensive synthesis, current systems lack precise control over mechanical properties, including homogeneity, stiffness, and efficient diffusion of nutrients to embedded cells. Difficulties in modifying gel matrix composition and functionalization have limited the use of covalently crosslinked gelatin hydrogels as a three-dimensional (3D) cell culture medium, lacking the ability to tailor the microenvironment for specific cell types. In addition, the currently utilized chain-growth photopolymerization mechanism for crosslinking hydrogels has a potential for side reactions between the matrix backbone and components of the cell surface, requires a high concentration of radicals for initiation, and only cures with long irradiation times, which could lead to cytotoxicity. To overcome these limitations, a superfast curing reaction mechanism, in which a thiol monomer reacts efficiently with non-homopolymerizable alkenes, is suggested. This mechanism reliably produces a well-defined matrix that does not require a high radical concentration for photoinitiation. Mechanical customization of the hydrogel is largely achievable through variation in degree of functionalization of the gelatin backbone, dependent on reaction conditions such as pH, allyl concentration, and time. This work provides a mechanistic framework for GelAGE hydrogel fabrication by elucidating the molecular mechanism of gelatin functionalization with AGE and the thiol-ene crosslinking reactions controlling network stiffness. These insights provide the foundation for engineering hydrogels that mimic the viscoelastic and structural characteristics of cartilage, enabling advanced in vitro models for osteoarthritis research. Full article

Click chemistry has become a powerful and flexible approach for designing hydrogels used in tissue engineering thanks to its high specificity, fast reaction rates, and compatibility with biological systems. In this review, we introduce the core principles of click chemistry, including efficiency, orthogonality, and modularity, and highlight the main types of reactions commonly used in hydrogel formation, such as azide-alkyne c-cloadditions, thiol-ene/yne reactions, Diels–Alder cycloadditions, and tetrazine–norbornene couplings. These chemistries allow researchers to create covalently crosslinked hydrogels that are injectable, responsive to environmental stimuli, biodegradable, or multifunctional. We also explore strategies to enhance bioactivity, such as incorporating peptides, growth factors, or extracellular matrix components, and enabling precise spatial and temporal control over biological cues. Click-based hydrogels have shown promise across a wide range of tissue engineering applications, from cartilage and skin repair to neural regeneration, corneal healing, and cardiovascular scaffolds, as well as in 3D bioprinting technologies. Despite the many advantages of click chemistry such as mild reaction conditions and customizable material properties, some challenges remain, including concerns around copper toxicity, the cost of specialized reagents, and scalability. Finally, we discuss the status of clinical translation, regulatory considerations, and future directions, including integration with advanced bio fabrication methods, the design of dual-click systems, and the emerging role of in vivo click chemistry in creating next-generation biomaterials. Full article

This study presents the synthesis of allyl-functionalized polysaccharide carbamates (AFCs) with tailored water solubility designed for use in responsive hydrogels and 3D printing applications. A modular one-pot approach was employed to produce cellulose- and xylan-based AFCs, utilizing polysaccharide phenyl carbonates as activated compounds. By fine-tuning the degree of substitution (DS) of functional groups, the water solubility and shear-thinning properties of AFCs were controlled to enhance the gelation and printability. AFC-based hydrogels could be obtained by rapid gelation induced without harmful catalysts through UV irradiation at 365 nm. The materials displayed highly porous and interconnected microstructures, as well as mechanical resilience and high swelling ratios. The hydrogel formation was characterized, and its crosslinking degree was calculated using HR-MAS NMR. The study demonstrated that gelation behavior was sensitive to the pH value, with optimal results under neutral or acidic conditions. Initial 3D printing trials confirmed the material’s rapid shaping capabilities, which is beneficial for biomedical applications and advanced manufacturing of stimuli-responsive materials. Full article

Since its conceptualization, click chemistry in all its variants has proven to be a superior synthesis protocol, compared to conventional methods, for forming new covalent bonds under mild conditions, orthogonally, and with high yields. If a term like reactive resilience could be established, click reactions would be good examples, as they perform better under increasingly challenging conditions. Particularly, highly hindered couplings that perform poorly with conventional chemistry protocols—such as those used to conjugate biomacromolecules (e.g., proteins and aptamers) or multiple drugs onto macromolecular platforms—can be more easily achieved using click chemistry principles, while also promoting high stereoselectivity in the products. In this review, three molecular platforms relevant in the field of nanomedicine are considered: polymers/copolymers, cyclodextrins, and fullerenes, whose functionalization poses a challenge due to steric hindrance, either from the intrinsic bulk behavior (as in polymers) or from the proximity of confined reactive sites, as seen in cyclodextrins and fullerenes. Their functionalization with biologically active groups (drugs or biomolecules), primarily through copper-catalyzed azide–alkyne cycloaddition (CuAAC), strain-promoted azide–alkyne cycloaddition (SPAAC), inverse electron-demand Diels–Alder (IEDDA) and thiol–ene click reactions, has led to the development of increasingly sophisticated systems with enhanced specificity, multifunctionality, bioavailability, delayed clearance, multi-targeting, selective cytotoxicity, and tracking capabilities—all essential in the field of nanomedicine. Full article

Graft copolymers have gained significant importance in various fields due to their tunable functionality and well-defined architecture. However, there are still limitations due to the compatibility of monomers and functional groups depending on the polymerization mode. Click chemistry has solved this problem through its ability to easily and quantitatively link a wide range of polymers and functional groups. The combination of click chemistry, including copper-catalyzed azide-alkyne cycloaddition (CuAAC), thiol-ene, and thiol-yne reactions, with various polymerization techniques offers a promising solution for the robust and efficient preparation of graft copolymers with the desired architecture and functionality. In this review, we present successful applications of click chemistry in the production of well-defined graft copolymers with diverse functionalities such as for electronics, energy devices, biomedical applications, and nanotechnology. Full article

Background: The incorporation of radionuclides into peptides and larger biomolecules requires efficient and sometimes biorthogonal reaction conditions, to which click chemistry provides a convenient approach. Methods: Traditionally, click-based radiolabeling techniques have focused on classical click chemistry, such as copper(I)-catalyzed alkyne-azide [3+2] cycloaddition (CuAAC), strain-promoted azide-alkyne [3+2] cycloaddition (SPAAC), traceless Staudinger ligation, and inverse electron demand Diels–Alder (IEDDA). Results: However, newly emerging click-based radiolabeling techniques, including tyrosine-click, sulfo-click, sulfur(VI) fluoride exchange (SuFEx), thiol-ene click, azo coupling, hydrazone formations, oxime formations, and RIKEN click offer valuable alternatives to classical click chemistry. Conclusions: This review will discuss the applications of these techniques in peptide radiochemistry. Full article

Antibiotic-resistant microorganisms have become a serious threat to public health, resulting in hospital infections, the majority of which are caused by commonly used urinary tract catheters. Strategies for preventing bacterial adhesion to the catheters’ surfaces have been potentially shown as effective methods, such as coating thesurface with antimicrobial biomolecules. Here, novel antimicrobial peptides (AMPs) were designed as potential biomolecules to prevent antibiotic-resistant bacteria from binding to catheter surfaces. Thiolated AMPs were synthesized using solid-phase peptide synthesis (SPPS), and prep-HPLC was used to obtain AMPs with purity greater than 90%. On the other side, the silicone catheter surface was activated by UV/ozone treatment, followed by functionalization with allyl moieties for conjugation to the free thiol group of cystein in AMPs using thiol-ene click chemistry. Peptide-immobilized surfaces were found to become more resistant to bacterial adhesion while remaining biocompatible with mammalian cells. The presence and site of conjugation of peptide molecules were investigated by immobilizing them to catheter surfaces from both ends (C-Pep and Pep-C). It was clearly demonstrated that AMPs conjugated to the surface via theirN terminus have a higher antimicrobial activity. This strategy stands out for its effective conjugation of AMPs to silicone-based implant surfaces for the elimination of bacterial infections. Full article

Thiols can react with readily available organic substrates under benign conditions, making them suitable for use in chemical, biological, physical, and materials and engineering research areas. In particular, the highly efficient thiol-based click reaction includes the reaction of radicals with electron-rich enes, Michael addition with electron-poor enes, carbonyl addition with isocyanate S_N2 ring opening with epoxies, and S_N2 nucleophilic substitution with halogens. This mini review provides insights into emerging venues for their industrial applications, especially for the applications of thiol-ene, thiol–isocyanate, and thiol–epoxy reactions, highlighting a brief chemistry of thiols as well as various approaches to polythiol synthesis. Full article

Fish skin is a by-product of the fishing industry, which has become a significant environmental pollutant in recent years. Therefore, there is an emerging interest in developing novel technologies to utilize fish skin as a versatile raw material for the clothing and biomedical industries. Most research on finishing procedures is conducted on cattle leather, and practically very limited information on fish leather finishing is found in the literature. We have developed three functional surface finishing treatments on chromium (CL)- and vegetable (VL)- tanned salmon leather. These treatments include hydrophobic, oil repellent, and electro-conductive ones. The hydroxyl functional groups present on the surface of the leather were covalently grafted with bi-functional aliphatic small molecule, 10-undecenoylchloride (UC), by esterification reaction forming hydrophobic coating. The surface hydrophobicity was further increased via covalent binding of perfluorodecanethiol (PFDT) to the double bond end-groups of the UC-modified leather via thiol-ene click chemistry conditions. The oleophobic coating was successfully developed using synthesized fluorinated silica nanoparticles (FSN) and polyvinylidene fluoride-co-hexafluoropropylene (PVDF-HFP), showing oil repellency with a contact angle of about 100° for soybean oil and n-hexadecane. The electrically conductive coating was realized by the incorporation of conjugated polymer, polyaniline (PANI), via in situ polymerization method. The treated leather exhibited surface resistivity of about 5.2 (Log (Ω/square)), much lower than untreated leather with a resistivity of 11.4 (Log (Ω/square)). Full article

Dielectric capacitors present many advantages for large-scale energy storage, but they presently require higher energy density. We demonstrate novel high energy density polymer-nanoparticle composite capacitors utilizing thiol-ene click chemistry surface groups to bond the nanoparticles covalently to the polymer matrix. Interfacial effects in composites cannot be observed directly, and in our previous work, we examined the nanoparticle–polymer interface in silico. In this work, we experimentally examine the five surface functionalizations modeled previously, fabricating high energy density thin film capacitors to test our predictions. Results from this study, in conjunction with properties previously determined in silico, further improve the understanding of the role of surface functionalizations in composites prepared using click chemistry. The coating density of the surface functionalizations is shown to be a key factor in relating our computational results to experimental results. We show how using both coating density and our previous modeling in combination allows for prescreening of surface functionalizations for future composites, reducing experimental cost. We also demonstrate high energy density capacitors with ~20 J/cm³. Full article

In this study, we elaborated new chitosan-based films reinforced by iron(III)-containing chitosan nanoparticles Fe(III)-CS-NPs at different concentrations. We found that the optimum concentration of Fe(III)-CS-NPs for the improvement of antibacterial and mechanical properties of the films was 10% (σ_b = ca. 8.8 N/mm², ε_b = ca. 41%, inhibition zone for S. aureus = ca. 16.8 mm and for E. coli = ca. 11.2 mm). Also, using the click-chemistry approach (thiol–ene reaction), we have synthesized a novel water-soluble cationic derivative of chitin. The addition of this derivative of chitin to the chitosan polymer matrix of the elaborated film significantly improved its mechanical (σ_b = ca. 11.6 N/mm², ε_b = ca. 75%) and antimicrobial (inhibition zone for S. aureus = ca. 19.6 mm and for E. coli = ca. 14.2 mm) properties. The key mechanism of the antibacterial action of the obtained films is the disruption of the membranes of bacterial cells. The elaborated antibacterial films are of interest for potential biomedical and food applications. Full article

Lignin and cellulose derivatives have vast potential to be applied in polymer materials. The preparation of cellulose and lignin derivatives through esterification modification is an important method to endow cellulose and lignin with good reactivity, processability and functionality. In this study, ethyl cellulose and lignin are modified via esterification to prepare olefin−functionalized ethyl cellulose and lignin, which are further used to prepare cellulose and lignin cross−linker polymers via thiol–ene click chemistry. The results show that the olefin group concentration in olefin−functionalized ethyl cellulose and lignin reached 2.8096 mmol/g and 3.7000 mmol/g. The tensile stress at break of the cellulose cross−linked polymers reached 23.59 MPa. The gradual enhancement in mechanical properties is positively correlated with the olefin group concentration. The existence of ester groups in the cross−linked polymers and degradation products makes them more thermally stable. In addition, the microstructure and pyrolysis gas composition are also investigated in this paper. This research is of vast significance to the chemical modification and practical application of lignin and cellulose. Full article

Temperature and light responsiveness are widely exploited stimuli to tune the physico-chemical properties of double network hydrogels. In this work, new amphiphilic poly(ether urethane)s bearing photo-sensitive moieties (i.e., thiol, acrylate and norbornene functionalities) were engineered by exploiting the versatility of poly(urethane) chemistry and carbodiimide-mediated green functionalization procedures. Polymers were synthesized according to optimized protocols maximizing photo-sensitive group grafting while preserving their functionality (approx. 1.0 × 10¹⁹, 2.6 × 10¹⁹ and 8.1 × 10¹⁷ thiol, acrylate and norbornene groups/g_polymer), and exploited to prepare thermo- and Vis-light-responsive thiol-ene photo-click hydrogels (18% w/v, 1:1 thiol:ene molar ratio). Green light-induced photo-curing allowed the achievement of a much more developed gel state with improved resistance to deformation (ca. 60% increase in critical deformation, γL). Triethanolamine addition as co-initiator to thiol-acrylate hydrogels improved the photo-click reaction (i.e., achievement of a better-developed gel state). Differently, L-tyrosine addition to thiol-norbornene solutions slightly hindered cross-linking, resulting in less developed gels with worse mechanical performances (~62% γL decrease). In their optimized composition, thiol-norbornene formulations resulted in prevalent elastic behavior at lower frequency compared to thiol-acrylate gels due to the formation of purely bio-orthogonal instead of heterogeneous gel networks. Our findings highlight that exploiting the same thiol-ene photo-click chemistry, a fine tuning of the gel properties is possible by reacting specific functional groups. Full article

This paper focuses on creating one-dimensional diffractive grooved structures of antigen proteins on glass substrates for the label-free detection of antibodies to dairy allergens. In particular, the fabrication of protein structures is carried out by combining microcontact printing with physisorption, imines coupling, and thiol-ene click chemistry. The work first sets up these patterning methods and discusses and compares the main aspects involved in them (structure, biolayer thickness, functionality, stability). Homogeneous periodic submicron structures of proteins are created and characterized by diffractive measurements, AFM, FESEM, and fluorescence scanning. Then, this patterning method is applied to proteins involved in cow milk allergy, and the resulting structures are implemented as optical transducers to sense specific immunoglobulins G. In particular, gratings of bovine serum albumin, casein, and β-lactoglobulin are created and assessed, reaching limits of detection in the range of 30–45 ng·mL⁻¹ of unlabeled antibodies by diffractive biosensing. Full article