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Enzyme-mediator bioconjugation is emerging as a building block for designing electrode platforms for the construction of biosensors and biofuel cells. Here, we report a one-pot bioconjugation technique for flavin adenine dinucleotide-dependent glucose dehydrogenase (FAD-GDH) and thionine (TH) using a series of cross-linkers, including epoxy, N-hydroxysuccinimide (NHS), and aldehydes. In this technique, FAD-GDH and thionine are conjugated through an amine cross-linking reaction to generate a redox network, which has been successfully employed for the oxidation of glucose. The bioconjugation chemistry of cross-linkers with the amino groups on FAD-GDH and thionine plays a vital role in generating distinct network structures. The epoxy-type cross-linker reacts with the primary and secondary amines of thionine at room temperature, thereby producing an FAD-GDH-TH-FAD-GDH hyperbranched bioconjugate network, the aldehyde undergoes a rapid cross-linking reaction to produce a network of FAD-GDH-FAD-GDH, while the NHS-based cross-linker can react with the primary amines of both FAD-GDH and thionine, forming an FAD-GDH-cross-linker-TH polymeric network. This reaction has the potential to enable the conjugation of a redox mediator with a FAD-GDH network, which is particularly essential when designing an enzyme electrode platform. The data demonstrated that the polymeric cross-linked network based on the NHS cross-linker exhibited a considerable increase in electron transport while producing a catalytic current of 830 µA cm-2. The cross-linker spacer arm length also affects the overall electrochemical function of the network and its performance; an adequate spacer length containing a cross-linker is required, resulting in a faster electron transfer. Finally, a leaching test confirmed that the stability of the enzyme electrode was improved when the electrode was tested using the redox probe. This study elucidates the relationship between cross-linking chemistry and redox network structure and enhances the high performance of enzyme electrode platforms for the oxidation of glucose.

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A new series of flexible polyamide (PA) aerogels was synthesized using terephthaloyl chloride (TPC), 2,2'-dimethylbenzidine (DMBZ) and cross-linked with an inexpensive, commercially available tri-isocyanate (Desmodur N3300A) at polymer concentrations of 6-8 wt.% total solids and repeating units, n, from 30 to 60. The cross-linked DMBZ-based polyamide aerogels obtained, after supercritically drying using liquid CO2, had shrinkages of 19-27% with densities ranging from 0.12 g/cm3 to 0.22 g/cm3, porosity and surface areas up to 91% and 309 m2/g, respectively, and modulus values ranging from 20.6 to 109 MPa. Evidence suggests that a higher flexibility could be achieved using DMBZ in the polyamide backbone with N3300A as a cross-linker, when compared to previously reported TPC-mPDA-BTC PA aerogels, N3300A-polyimide aerogels, and N3300-reinforced silica aerogels.

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Natural-derived biomaterials can be used as substrates for the growth, proliferation, and differentiation of cells. In this study, bovine vitreous humor as a biological material was cross-linked to silk fibroin with different concentration ratios to design a suitable substrate for corneal tissue regeneration. The cross-linked samples were evaluated with different analyses such as structural, physical (optical, swelling, and degradation), mechanical, and biological (viability, cell adhesion) assays. The results showed that all samples had excellent transparency, especially those with higher silk fibroin content. Increasing the ratio of vitreous humor to silk fibroin decreased mechanical strength and increased swelling and degradation, respectively. There was no significant difference in the toxicity of the samples, and with the increase in vitreous humor ratio, adhesion and cell proliferation increased. Generally, silk fibroin with vitreous humor can provide desirable characteristics as a transparent film for corneal wound healing.

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Proton exchange membranes (PEMs) applied in fuel cell technology suffer from the trade-off between fast proton conduction and durable operation involving dimensional stability, mechanical strength, and oxidative resistance. To address this issue, a novel branched polybenzimidazole (brPBI) was synthesized, covalently cross-linked with (3-chloropropyl)triethoxysilane (CTS), and doped with a novel proton conductor FeATMP to prepare brPBI-CTS/FeATMP membranes. The branching degree of brPBI was optimized to achieve high molecular weight while the branching structure offered high free volume, abundant end-groups, and self-cross-linking moiety that enhanced proton conduction and dimensional/mechanical/oxidative stability. Covalent cross-linking with CTS enhanced the dimensional, mechanical, and oxidative stability while improving the water-assisted proton conduction owing to the hydrophilic nature of siloxane structure formed. At 180 ℃, the proton conductivity of the brPBI3-CTS/FeATMP composite membrane reached 0.136, 0.073, and 0.041 S cm-1 at 100 % RH, 50 % RH, and 0 % RH, respectively, while its swelling ratio after immersion in water at 90 ℃ for 24 h was 4.69 %. The performance of the membranes demonstrated that construction of hydrophilic structure by covalent cross-linking was a successful strategy to break the trade-off effect for PEMs.

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The traditional chemotherapeutic agents' disadvantages such as high toxicity, untargeting and poor water solubility lead to disappointing chemotherapy effects, which restricts its clinical application. In this work, novel size-appropriate and glutathione (GSH)-responsive nano-hydrogels were successfully prepared via the active ester method between chitosan (containing -NH2) and cross-linker (containing NHS). Especially, the cross-linker was elaborately designed to possess a disulfide linkage (SS) as well as two terminal NHS groups, namely NHS-SS-NHS. These functionalities endowed chitosan-based cross-linked scaffolds with capabilities for drug loading and delivery, as well as a GSH-responsive mechanism for drug release. The prepared nano-hydrogels demonstrated excellent performance applicable morphology, excellent drug loading efficiency (∼22.5%), suitable size (∼100 nm) and long-term stability. The prepared nano-hydrogels released over 80% doxorubicin (DOX) after incubation in 10 mM GSH while a minimal DOX release less than 25% was tested in normal physiological buffer (pH = 7.4). The unloaded nano-hydrogels did not show any apparent cytotoxicity to A 549 cells. In contrast, DOX-loaded nano-hydrogels exhibited marked anti-tumor activity against A 549 cells, especially in high GSH environment. Finally, through fluorescent imaging and flow cytometry analysis, fluorescein isothiocyanate-labeled nano-hydrogels show obvious specific binding to the GSH high-expressing A549 cells and nonspecific binding to the GSH low-expressing A549 cells. Therefore, with this cross-linking approach, our present finding suggests that cross-linked chitosan nano-hydrogel drug carrier improves the anti-tumor effect of the A 549 cells and may serve as a potential injectable delivery carrier.

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Background: Current drugs for treating osteoporosis may lead to toxic side effects. Echinacoside (ECH) is a natural small molecule drug. This study examined and compared the therapeutic effects of cross-linker (CL)-ECH and ECH-free nanoparticles on osteoporosis. Methods: Echinocandin-based CL-ECH nanoparticles were prepared, and the nanoparticle size and drug loading were optimized and characterized by adjusting the ratio. The antioxidant effect of CL-ECH nanoparticles on bone marrow-derived macrophages (BMDMs) was analyzed using flow cytometry, immunofluorescence staining and quantitative real-time polymerase chain reaction (qRT-PCR). Bone marrow stromal cells (BMSCs)-based detection of bone-producing effects was conducted using alkaline phosphatase (ALP), Alizarin Red S (ARS) and qRT-PCR. TRAP, phalloidin staining, and qRT-PCR was performed to detect osteogenesis-inhibiting effect on BMDMs. CL-ECH nanoparticles were applied to treat an ovariectomized (OVX) mouse model at low doses. Results: Compared to ECH, CL-ECH nanoparticles suppressed oxidative stress in BMDMs by promoting NRF-2 nuclear translocation, which inhibited the production of both reactive oxygen species (ROS) and osteoclast production through downregulating NF-κB expression, with limited effect on the osteogenesis of BMSCs. In vivo studies showed that low-dose CL-ECH nanoparticles markedly improved bone trabecular loss compared to ECH administration in the treatment of osteoporosis. Conclusions: The current discoveries provided a solid theoretical foundation for the development of a new generation of anti-bone resorption drugs and antiosteoporosis drugs.

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BCL9 is a key protein in Wnt signaling pathway. It acts as a transcriptional co-activator to ß-catenin, and dysregulation in this pathway leads to tumor growth. Inhibiting such a protein-protein interaction is considered as a therapeutic challenge. The interaction between ß-catenin and BCL9 is facilitated by a 23-residue helical domain from BCL9 and a hydrophobic groove of ß-catenin. To prevent this interaction, a peptide that mimics the alpha-helical domain of BCL9 can be designed. Stapling is considered a successful strategy in the pursuit of designing such peptides in which amino acids side are stitched together using chemical moieties. Among the various types of cross-linkers, triazole is the most rapid and effective one synthesized via click reaction. However, the underlying interactions behind maintaining the secondary structure of stapled peptides remain less explored. In the current work, we employed the molecular dynamics simulation to study the conformational behavior of the experimentally synthesized single and double triazole stapled BCL9 peptide. Upon the addition of a triazole staple, there is a significant reduction in the conformational space of BCL9. The helical character of the stapled peptide increases with an increase in separation between the triazole cross-linkers. Also, we encompassed the Replica Exchange with Solute Tempering (REST2) simulation to validate the high-temperature response of the stapled peptide. From REST2, the PCA and t-SNE show the reduction in distinct cluster formation on the addition of triazole staple. Our study infers further development of these triazole-stapled BCL9 peptides into effective inhibitors to target the interaction between ß-catenin and BCL9.

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The advancement of anion exchange membranes (AEMs) with superior ionic conductivity has been greatly hindered due to the inherent "trade-off" between membrane swelling and ionic conductivity. To resolve this dilemma, macromolecular covalently cross-linked C-FPVBC-x AEMs were fabricated by combining partially functionalized ether-bond-free polystyrene (FPVBC) with poly(arylene piperidinium). The results from atomic force microscopy reveal that an increase in the ratio of FPVBC promotes the fabrication of microphase separation morphology, resulting in a high ionic conductivity of 40.15 mS cm-1 (30 °C) for the C-FPVBC-1.7 membrane. Molecular dynamics simulations further examine the ionic conduction effect of cross-linked AEMs. Besides, the unique cross-linking structure significantly improves mechanical and alkaline stability. After treatment in 1 M KOH at 50 °C for 1200 h, the C-FPVBC-1.7 membrane shows only a 6.9% decrease in conductivity. The C-FPVBC-1.7 AEM-based water electrolyzer achieves a high current density of 890 mA cm-2 at 2.4 V (80 °C) and maintains good stability, enduring over 100 h at 100 mA cm-2 (50 °C). These results demonstrate the significant potential of macromolecularly cross-linked AEMs for practical applications in water electrolysis.

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Acidic residues (Asp and Glu) have a high prevalence on protein surfaces, but cross-linking reactions targeting these residues are limited. Existing methods either require high-concentration coupling reagents or have low structural compatibility. Here a previously reported "plant-and-cast" strategy is extended to develop heterobifunctional cross-linkers. These cross-linkers first react rapidly with Lys sidechains and then react with Asp and Glu sidechains, in a proximity-enhanced fashion. The cross-linking reaction proceeds at neutral pH and room temperature without coupling reagents. The efficiency and robustness of cross-linking using model proteins, ranging from small monomeric proteins to large protein complexes are demonstrated. Importantly, it is shown that this type of cross-linkers are efficient at identifying protein-protein interactions involving acidic domains. The Cross-linking mass spectrometry (XL-MS) study with p53 identified 87 putative binders of the C-terminal domain of p53. Among them, SARNP, ZRAB2, and WBP11 are shown to regulate the expression and alternative splicing of p53 target genes. Thus, these carboxylate-reactive cross-linkers will further expand the power of XL-MS in the analysis of protein structures and protein-protein interactions.

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Magnetite nanoparticles (Fe3O4 NPs) are among the most effective Fenton-Like heterogeneous catalysts for degrading environmental contaminants. However, Fe3O4 NPs aggregate easily and have poor dispersion stability because of their magnetic properties, which seriously decrease their catalytic efficiency. In this study, a novel environmentally friendly method for synthesising Fe3O4@CA was proposed. Fe3O4 NPs were immobilized on the 3D cellulose aerogels (CAs) in order to augment the degradation efficiency of p-nitrophenol (PNP) treatment and make the separation of the catalyst accessible by vacuum filtration method. Besides, CAs were fabricated from a cellulose source extracted from water hyacinth by using different cross-linking agents, such as kymene (KM) and polyvinyl alcohol-glutaraldehyde system (PVA-GA), and other drying methods, including vacuum thermal drying and freeze drying, were evaluated in the synthesis process. As-synthesized samples were analysed by various methods, including Powder X-ray diffraction, Fourier transform infrared spectroscopy, field emission scanning electron microscopy, energy dispersive X-ray analysis and Brunauer-Emmett-Teller. Then, using ultraviolet-visible spectroscopy, the difference in the degradability of PNP of the obtained material samples was also investigated to determine their potential applications. Results highlighted that the Fe3O4-3@CA-KF catalyst with an Fe3O4 loading of 0.40 g/gCA used KM as a cross-linker and the freeze-drying method demonstrated the highest PNP removal efficiency (92.5 %) in all Fe3O4@CA samples with a H2O2 content of 5 g/L. The degradation kinetics and well-fitted pseudo-first-order model were investigated. Notably, after five successive PNP degradation experiments, this catalyst retained ∼80 % of the ability to degrade PNP, indicating its outstanding reusability. In environmental remediation, this study provides valuable insights into the development of simply separated and high-efficiency catalysts for heterogeneous catalytic reactions.

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In recent years, nanotechnology has been the focus of study for the cure of different diseases, among which nanosponge delivery system is one of a kind. Nano sponges are tiny, highly porous, three-dimensional nanostructures with a size range of 250nm-1µm in an amorphous or crystalline structure. Nanosponges usually act as an excipient or carrier of a drug in the different delivery systems. The type of polymers and cross-linkers, along with their concentration ratio, causes variation in nanosponges's dimension and encapsulation efficiency. Nanosponges have gained prominence in recent times due to their distinct ability to encapsulate both hydrophilic and lipophilic drugs within their internal cavity, thereby improving the solubility of drugs that have low water solubility. Virus-like size helps the nanosponges to circulate within the body without getting eliminated by the immune system until they stick to the targeted part of the body, which makes it the perfect candidate for a targeted drug delivery system and controlled delivery system as well because of its slow drug release property for a more extended period. Cyclodextrin-based nanosponges are the best choice for anticancer drug delivery as their small virus-like diameter helps them in passive targeting by enhancing the enhanced permeability and retention effect, allowing the anticancer drug to stay inside the tumour cell to show more significant therapeutic action on cancer, while for active targeting to the cancerous cell, nanosponges are attached with a ligand on it for receptor binding purpose. It can be used for drug delivery in many major diseases like brain-related diseases, diabetes, cancer, fungal, hypertension, etc., in different dosage forms, like oral, topical, hydrogel, parenteral, etc.

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Chemical modification of the enzymes with biospecific macromolecules is used in various fields of biotechnology to impart new functions or improve their properties and is a fast and convenient way to get the final products. The preparation of highly active, stable, and functionally active conjugates of the thermostable luciferase through the NH2-groups or free SH-groups of the enzyme with target molecules of different molecular weight (albumin, avidin from chicken eggs, antibodies, and progesterone) is described. The obtained conjugates were successfully tested as a reporter in bioluminescent immunoassay for the detection of the molecules and pathogens. Thus, the luc-albumin (Luc-Alb) and luc-insulin (Luc-Ins) conjugates were used in competitive ELISA for the detection of an analyte (albumin or insulin) in the samples. Luc-progesterone (Luc-Pg) was used in the rapid homogeneous immunoassay of progesterone by the BRET technique with the detection limit of 0.5 ng/ml. Luciferase conjugates with avidin (Luc-Avi) and secondary and primary antibodies (Luc-RAM and Luc-Sal) were used for enzyme immunoassay detection of Salmonella paratyphi A cells with the cell detection limit of 5 × 104 CFU/ml. To reduce the detection limit of Salmonella cells, we developed a pseudo-homogeneous bioluminescent enzyme immunoassay of cells using a new matrix for the analyte capture-polystyrene microparticles coated with Pluronic F108, covalently labeled with Sal antibodies. This allowed to achieve efficient trapping of cells from solution, significantly reduced nonspecific sorption and decreased the cell detection limit to 2.7 × 103 CFU/ml without prior concentration of the sample. The methodology that was developed in this study can be applied for the development of novel bioanalytical systems based on firefly luciferases.

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Bio-based cross-linkers can fulfill the role of enhancing additives in bio-sourced curable materials that do not compare with artificial resin precursors. Isosorbide dimethacrylate (ISDMMA) synthesized from isosorbide (ISD) can serve as a cross-linker from renewable sources. Isosorbide is a bicyclic carbon molecule produced by the reaction modification of sorbitol and the optimal conditions of this reaction were studied in this work. The reaction temperature of 130 °C and 1% w/w amount of para-toluenesulfonic acid (p-TSA) were determined as optimal and resulted in a yield of 81.9%. Isosorbide dimethacrylate was synthesized via nucleophilic substitution with methacrylic anhydride (MAA) with the conversion of 94.1% of anhydride. Formed ISD and ISDMMA were characterized via multiple verification methods (FT-IR, MS, 1H NMR, and XRD). Differential scanning calorimetry (DSC) proved the curability of ISDMMA (activation energy Ea of 146.2 kJ/mol) and the heat-resistant index of ISDMMA (Ts reaching value of 168.9) was determined using thermogravimetric analysis (TGA). Characterized ISDMMA was added to the precursor mixture containing methacrylated alkyl 3-hydroxybutyrates (methyl ester M3HBMMA and ethyl ester E3HBMMA), and the mixtures were cured via photo-initiation. The amount of ISDMMA cross-linker increased all measured parameters obtained via dynamic mechanical analysis (DMA), such as storage modulus (E') and glass transition temperature (Tg), and the calculated cross-linking densities (νe). Therefore, the enhancement influence of bio-based ISDMMA on resins from renewable sources was confirmed.

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Cross-linking mass spectrometry (XL-MS) is widely used in the analysis of protein structure and protein-protein interactions (PPIs). Throughout the entire workflow, the utilization of cross-linkers and the interpretation of cross-linking data are the core steps. In recent years, the development of cross-linkers and analytical software mostly follow up on the classical models of non-cleavable cross-linkers such as BS3/DSS and MS-cleavable cross-linkers such as DSSO. Although such a paradigm promotes the maturity and robustness of the XL-MS field, it confines the innovation and flexibility of new cross-linkers and analytical software. This critical insight will discuss the classification, advantages, and disadvantages of existing data analysis search engines. Take the new platinum-based metal cross-linker as an example, potential pitfalls in characterization of cross-linked peptides using existing software are discussed. Finally, ideas on developing more flexible, comprehensive, and user-friendly cross-linkers and software tools are proposed.

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When it comes to mass spectrometry data analysis for identification of peptide pairs linked by N-hydroxysuccinimide (NHS) ester cross-linkers, search engines bifurcate in their setting of cross-linkable sites. Some restrict NHS ester cross-linkable sites to lysine (K) and protein N-terminus, referred to as K only for short, whereas others additionally include serine (S), threonine (T), and tyrosine (Y) by default. Here, by setting amino acids with chemically inert side chains such as glycine (G), valine (V), and leucine (L) as cross-linkable sites, which serves as a negative control, we show that software-identified STY-cross-links are only as reliable as GVL-cross-links. This is true across different NHS ester cross-linkers including DSS, DSSO, and DSBU, and across different search engines including MeroX, xiSearch, and pLink. Using a published data set originated from synthetic peptides, we demonstrate that STY-cross-links indeed have a high false discovery rate. Further analysis revealed that depending on the data and the search engine used to analyze the data, up to 65% of the STY-cross-links identified are actually K-K cross-links of the same peptide pairs, up to 61% are actually K-mono-links, and the rest tend to contain short peptides at high risk of false identification.

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Alginate-based hydrogels are promising smart materials widely employed in the food, bioengineering, and energy sectors. The development and optimization of their production require a thorough knowledge of gelation. In recent years, advanced experimental procedures have been developed for real-time cross-linking alginate reaction monitoring. Novel methods, such as customized rheometric setups, enable the recording of mechanical properties and morphological changes during hydrogel formation. These innovative techniques provide important insights into the gelation stages, the reaction rate, the diffusion of cross-linker to polymer chains, and the homogeneity of the gelling structures. Based on real-time experimental data, kinetic models are developed to enhance comprehension of the reaction mechanism and, eventually, to predict the gelation progress. The aim is to enable better control of the characterization of both the complex gelation and the propagated structures. This review aspires to present a comprehensive overview and evaluation of the breakthrough innovations of the real-time in situ recording of cross-linking alginate hydrogels and bead formation. A detailed analysis of the pioneering experimental developments provides a deep comprehension of the alginate gelation, including the parameters controlling the reaction.

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Selective delivery of anticancer drug molecules to the tumor site enhances local drug dosages, which leads to the death of cancer cells while simultaneously minimizing the negative effects of chemotherapy on other tissues, thereby improving the patient's quality of life. To address this need, we developed reduction-responsive chitosan-based injectable hydrogels via the inverse electron demand Diels-Alder reaction between tetrazine groups of disulfide-based cross-linkers and norbornene groups of chitosan derivatives, which were applied to the controlled delivery of doxorubicin (DOX). The swelling ratio, gelation time (90-500 s), mechanical strength (G'~350-850 Pa), network morphology, and drug-loading efficiency (≥92%) of developed hydrogels were investigated. The in vitro release studies of the DOX-loaded hydrogels were performed at pH 7.4 and 5.0 with and without DTT (10 mM). The biocompatibility of pure hydrogel and the in vitro anticancer activity of DOX-loaded hydrogels were demonstrated via MTT assay on HEK-293 and HT-29 cancer cell lines, respectively.

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Fabricating sustainable ionic skin with multi-functional outstanding performances using biocompatible natural polymer-based ionogel is highly desired but remains a great challenge up to now. Herein, a green and recyclable ionogel has been fabricated by in-situ cross-linking of gelatin with a green bio-based multifunctional cross-linker of Triglycidyl Naringenin in ionic liquid. Benefiting from the unique multifunctional chemical crosslinking networks along with multiple reversible non-covalent interactions, the as-prepared ionogels exhibit high stretchability (>1000 %), excellent elasticity, fast room-temperature self-healability (>98 % healing efficiency at 6 min), and good recyclability. These ionogels are also highly conductive (up to 30.7 mS/cm at 150 °C), and exhibit extensive temperature tolerance (-23 to 252 °C) and outstanding UV-shielding ability. As a result, the as-prepared ionogel can easily be applied as stretchable ionic skin for wearable sensors, which exhibits high sensitivity, fast response time (102 ms), excellent temperature tolerance, and stability over 5000 stretching-relaxing cycles. More importantly, the gelatin-based sensor can be used in signal monitor system for various human motion real-time detection. This sustainable and multifunctional ionogel provides a new idea for easy and green preparation of advanced ionic skins.

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Herein, we report biocompatible hydrogel for wound healing that was prepared using nature-sourced building blocks. For the first time, OCS was employed as a building macromolecule to form bulk hydrogels along with the nature-sourced nucleoside derivative (inosine dialdehyde, IdA) as the cross-linker. A strong correlation was obtained between the mechanical properties and stability of the prepared hydrogels with a cross-linker concentration. The Cryo-SEM images of IdA/OCS hydrogels showed an interconnected spongy-like porous structure. Alexa 555 labeled bovine serum albumin was incorporated into the hydrogels matrix. The release kinetics studies under physiological conditions indicated that cross-linker concentration could also control the release rate. The potential of hydrogels in wound healing applications was tested in vitro and ex vivo on human skin. Topical application of the hydrogel was excellently tolerated by the skin with no impairment of epidermal viability or irritation, determined by MTT and IL-1α assays, respectively. The hydrogels were used to load and deliver epidermal growth factor (EGF), showing an increase in its ameliorating action, effectively enhancing wound closure inflicted by punch biopsy. Furthermore, BrdU incorporation assay performed in both fibroblast and keratinocyte cells revealed an increased proliferation in hydrogel-treated cells and an enhancement of EGF impact in keratinocytes.

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Hydrogels are 3D crosslinking networks of hydrophilic biopolymers which can able to absorb and retain large amount of water. In this present study, the Sodium alginate (SA)- Galactoxyloglucan (GXG) blended hydrogel beads were prepared and optimized through two level optimization steps. Alginate and xyloglucan are the cell wall polysaccharides biopolymers obtained from the plant sources, Sargassum sp. and Tamarindus indica L. respectively. The extracted biopolymers were confirmed and characterized by UV-Spectroscopy, FT-IR, NMR and TGA analysis. Based on the hydrophilicity, non-toxicity and biocompatibility, SA-GXG hydrogel were prepared and optimized through two-level optimization steps. The optimized hydrogel bead formulation was characterized through FT-IR, TGA and SEM analysis. From the obtained result, it is found that the polymeric formulation GXG (2 % w/v)-SA (1.5 % w/v), cross-linker (CaCl2) concentration at 0.1 M and the cross-linking time at 15 Min showed significant swelling index. The optimized hydrogel beads are porous and show good swelling capacity and thermal stability. The optimized protocol of hydrogel beads may be useful in designing hydrogel beads for specific applications in agricultural, Biomedical and remediation sectors.

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