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SEM images of lyophilized collagen/elastin gel. a 95Coll-5El, b 95Coll-5El-5DAS, c 95Coll-5El-10DAS, d 90Coll-10El, e 90Coll-10El- 5DAS, f 90Coll-10El-10DAS
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Collagen and elastin are the main structural proteins in mammal bodies. They provide mechanical support, strength, and elasticity to various organs and tissues, e.g. skin, tendons, arteries, and bones. They are readily available, biodegradable, biocompatible and they stimulate cell growth. The physicochemical properties of collagen and elastin-base...
Citations
... Furthermore, only 5% DAS addition to 1% collagen films caused a small shift of the Amide II band (which provides information regarding the conformation of proteins) to a higher wavenumber [41]. Moreover, it can be noticed that the increased intensity of the band at 1030 cm −1 might be correlated with DAS compound addition (the highest intensity was for 5% DAS addition), which is a characteristic band for dialdehyde starch corresponding to C-O stretching vibrations [41], while for 2% collagen samples with 5% DAS addition, bands at 1030 cm −1 presented slightly different shapes and were wider. ...
... For non-cross-linked and DAS-cross-linked collagen samples, wavenumbers for characteristic bands remained almost at the same positions; thus, it might indicate that the collagen secondary structure was not influenced; however, changes can be observed in characteristic band intensities [38]. Based on the assessment of the Amide I band, which allows for protein secondary structure analysis [39][40][41], changes in its intensity for cross-linked samples might reflect created interactions between a cross-linking agent and the collagen matrix [38]. Moreover, it can be also observed that all 1% and 2% collagen films with cross-linking agents showed alternations of the band corresponding to a symmetrical stretch of COO-to lower wavenumbers, which also might be associated with carboxyl groups' involvement in new interactions. ...
... Moreover, it can be also observed that all 1% and 2% collagen films with cross-linking agents showed alternations of the band corresponding to a symmetrical stretch of COO-to lower wavenumbers, which also might be associated with carboxyl groups' involvement in new interactions. Furthermore, only 5% DAS addition to 1% collagen films caused a small shift of the Amide II band (which provides information regarding the conformation of proteins) to a higher wavenumber [41]. Moreover, it can be noticed that the increased intensity of the band at 1030 cm −1 might be correlated with DAS compound addition (the highest intensity was for 5% DAS addition), which is a characteristic band for dialdehyde starch corresponding to C-O stretching vibrations [41], while for 2% collagen samples with 5% DAS addition, bands at 1030 cm −1 presented slightly different shapes and were wider. ...
The aim of this research was the modification of fish collagen films with various amounts of dialdehyde starch (DAS). Film properties were examined before and after the cross-linking process by DAS. Prepared biopolymer materials were characterized by Fourier Transform Infrared Spectroscopy and Atomic Force Microscopy. Moreover, the mechanical, thermal and swelling properties of the films were evaluated and the contact angle was measured. Research has shown that dialdehyde starch applied as a cross-linking agent influences collagen film properties. Mechanical testing indicated a decrease in Young’s Modulus and an increase in breaking force, elongation at break, and tensile strength parameters. Results for contact angle were significantly higher for collagen films cross-linked with DAS; thus, the hydrophilicity of samples decreased. Modified samples presented a lower swelling degree in PBS than native collagen films. However, the highest values for the degree of swelling among the modified specimens were obtained from the 1% DAS samples, which were 717% and 702% for 1% and 2% collagen, respectively. Based on AFM images and roughness values, it was noticed that DAS influenced collagen film surface morphology. The lowest value of Rq was observed for 2%Coll_2%DAS and was approximately 10 nm. Analyzing thermograms for collagen samples, it was observed that pure collagen samples were less thermally stable than cross-linked ones. Dialdehyde starch is a promising cross-linking agent for collagen extracted from fish skin and may increase its applicability.
... Also, PEG-A and PEG-NHS are amine-reactive compounds. They are formed as a result of modification of the hydroxyl end groups of the polyethylene glycol chain-the water-soluble, neutral, and biocompatible polymer widely used in pharmacy [18,19]. We also used squaric acid (SQ) (3,4-dihydroxy 3-cyclobutene 1,2-dione), which is a strong aromatic acid. ...
... This is proved by the increase in the thermal degradation temperature of cross-linked materials, as well as the changes observed in the FTIR spectra. The increase in Amide I and Amide II intensity indicates the formation of new amide bonds as a result of reactions between aldehyde groups of DAS, PEG-A, and amino groups of hAM [18,19], and also mediated by EDC/NHS and PEG-NHS [4]. As a result, the mechanical strength of cross-linked human amniotic membranes is improved. ...
... Interestingly, elongation at the breaking point increases after cross-linking with most of the reagents used. As it is well known, aldehydes easily react with amino groups under mild conditions [14,19]. Hence, the high effectiveness of crosslinking using DAS and PEG-A is observed. ...
Human amniotic membranes (hAMs) obtained during cesarean sections have proven to be clinically useful as an interesting biomaterial in a wide range of tissue engineering applications such as ocular surface reconstruction, burn treatments, chronic wounds, or bedsore ulcers. It presents antimicrobial properties, promotes epithelization, reduces inflammation and angiogenesis, contains growth factors, and constitutes the reservoir of stem cells. However, variability in hAM stiffness and its fast degradation offers an explanation for the poor clinical applications and reproducibility. In addition, the preparatory method of hAM for clinical use can affect its mechanical properties, and these differences can influence its application. As a directly applied biomaterial, the hAM should be available in a ready-to-use manner in clinical settings. In the present study, we performed an analysis to improve the mechanical properties of hAM by the addition of various reagents used as protein cross-linkers: EDC/NHS, PEG-dialdehyde, PEG-NHS, dialdehyde starch, and squaric acid. The effect of hAM modification using different cross-linking agents was determined via infrared spectroscopy, thermal analyses, mechanical properties analyses, enzymatic degradation, and cytotoxicity tests. The use of PEG-dialdehyde, PEG-NHS, dialdehyde starch, and squaric acid increases the mechanical strength and elongation at the breaking point of hAM, while the addition of EDC/NHS results in material stiffening and shrinkage. Also, the thermal stability and degradation resistance were evaluated, demonstrating higher values after cross-linking. Overall, these results suggest that modification of human amniotic membrane by various reagents used as protein cross-linkers may make it easier to use hAM in clinical applications, and the presented study is a step forward in the standardization of the hAM preparation method.
... Several groups have used covalent crosslinking strategies in collagen to improve gel mechanics, through free-radical photochemistry [49], various enzymatic routes, [50][51][52] and non-enzymatic chemistry [53][54][55]. While these strategies have been somewhat effective in improving the moduli and strength of the resultant materials, they generally result in decreased extensibility and disrupt native structural and functional properties, [56][57][58] namely the formation of a uniform fibrillar network structure and thermal gelation. ...
Current auricular cartilage replacements for pediatric microtia fail to address the need for long-term integration and neocartilage formation. While collagen hydrogels have been successful in fostering neocartilage formation, the toughness and extensibility of these materials do not match that of native tissue. This study used the N-terminal functionalization of collagen with alginate oligomers to improve toughness and extensibility through metal–ion complexation. Alginate conjugation was confirmed via FTIR spectroscopy. The retention of native collagen fibrillar structure, thermal gelation, and helical conformation in functionalized gels was confirmed via scanning electron microscopy, oscillatory shear rheology, and circular dichroism spectroscopy, respectively. Alginate–calcium complexation enabled a more than two-fold increase in modulus and work density in functionalized collagen with the addition of 50 mM CaCl2, whereas unmodified collagen decreased in both modulus and work density with increasing calcium concentration. Additionally, the extensibility of alginate-functionalized collagen was increased at 25 and 50 mM CaCl2. Following 2-week culture with auricular chondrocytes, alginate-functionalization had no effect on the cytocompatibility of collagen gels, with no effects on cell density, and increased glycosaminoglycan deposition. Custom MATLAB video analysis was then used to quantify fracture toughness, which was more than 5-fold higher following culture in functionalized collagen and almost three-fold higher in unmodified collagen.
... Collagen is present in all multicellular animals, providing mechanical support, resistance, and elasticity to several organs and tissues (1). Collagen is the most abundant extracellular matrix protein and represents one third of all proteins in humans. ...
The need to fully exploit fishing resources due to increasing production and consequent waste generation requires research to promote the sustainability of the fishing industry. Fish waste from the industry is responsible for relevant environmental contamination. However, these raw materials contain high amounts of collagen and other biomolecules, being attractive due to their industrial and biotechnological applicability. Thus, to reduce the waste from pirarucu (Arapaima gigas) processing, this study aimed to obtain collagen from pirarucu skin tissue. The extraction process used 0.05 M sodium hydroxide, 10% butyl alcohol, and 0.5 M acetic acid, with extraction temperature of 20°C. The obtained yield was 27.8%, and through sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), it was determined that the collagen obtained was type I. This study showed that collagen solubility was highest at pH 3 and the lowest solubility was at concentrations of 3% sodium chloride. The denaturation temperature of collagen was 38.1°C, and its intact molecular structure was observed using the Fourier transform infrared spectrophotometry technique with an absorption radius of 1. The results showed that it was possible to obtain collagen from pirarucu skin at 20°C, which has the typical characteristics of commercial type I collagen. In conclusion, the procedures used may be considered to be an interesting alternative for collagen extraction, a new product obtained from the processing of fish waste.
... The main structural proteins in mammals are collagen and elastin, which provide mechanical support, strength, and elasticity in various tissues and organs. Collagen and elastin are readily available, biodegradable, biocompatible, and can stimulate cell growth [36]. In the anti-enzymatic hydrolysis experiments (collagenase I and collagenase IV) of fixed samples in vitro, the ability of GNP-fixed DPP to resist collagenase digestion is better than that of GA-fixed DPP at 72 h, and OKGM-fixed DPP is equivalent to GA-fixed DPP (Fig. 7). ...
The use of natural polysaccharide crosslinkers for decellularized matrices is an effective approach to prepare cardiovascular substitute materials. In this research, NaIO 4 was applied to oxidize konjac glucomannan to prepare the polysaccharide crosslinker oxidized konjac glucomannan (OKGM). The as-prepared crosslinker was then used to stabilize collagen-rich decellularized porcine peritoneum (DPP) to construct a cardiovascular substitute material (OKGM-fixed DPP). The results demonstrated that compared with GA-fixed DPP and GNP-fixed DPP, 3.75% OKGM [1:1.5 (KGM: NaIO 4 )]-fixed DPP demonstrated suitable mechanical properties, as well as good hemocompatibility, excellent anti-calcification capability, and anti-enzymolysis in vitro. Furthermore, 3.75% OKGM [1:1.5 (KGM: NaIO 4 )]-fixed DPP was suitable for vascular endothelial cell adhesion and rapid proliferation, and a single layer of endothelial cells was formed on the fifth day of culture. The in vivo experimental results also showed excellent histocompatibility. The current results demonstrted that OKGM was a novel polysaccharide cross-linking reagent for crosslinking natural tissues featured with rich collagen content, and 3.75% OKGM [1:1.5 (KGM: NaIO 4 )]-fixed DPP was a potential cardiovascular substitute material.
Graphical Abstract
... During the cross-linking process, the structure of the protein's triple helix does not change and their biological properties remain the same as they were before. During the process, dialdehyde starch reacts with the amino acid residues to form intermolecular bonds [166]. ...
The main aim of this review is to assess the potential use of natural cross-linking agents, such as genipin, citric acid, tannic acid, epigallocatechin gallate, and vanillin in preparing chemically cross-linked hydrogels for the biomedical, pharmaceutical, and cosmetic industries. Chemical cross-linking is one of the most important methods that is commonly used to form mechanically strong hydrogels based on biopolymers, such as alginates, chitosan, hyaluronic acid, collagen, gelatin, and fibroin. Moreover, the properties of natural cross-linking agents and their advantages and disadvantages are compared relative to their commonly known synthetic cross-linking counterparts. Nowadays, advanced technologies can facilitate the acquisition of high-purity biomaterials from unreacted components with no additional purification steps. However, while planning and designing a chemical process, energy and water consumption should be limited in order to reduce the risks associated with global warming. However, many synthetic cross-linking agents, such as N,N’-methylenebisacrylamide, ethylene glycol dimethacrylate, poly (ethylene glycol) diacrylates, epichlorohydrin, and glutaraldehyde, are harmful to both humans and the environment. One solution to this problem could be the use of bio-cross-linking agents obtained from natural resources, which would eliminate their toxic effects and ensure the safety for humans and the environment.
... Non-crosslinked single-component or blended materials typically have low elastic stability in aqueous environments [227,228]. However, cross-linking improves the mechanical properties and increases the aqueous stability and degradation resistance [229]. The amine and carboxylic acid groups of SF are easily accessible, allowing it to be easily modified [103]. ...
... Chemical cross-linking is thought to be the most controllable and efficient strategy for producing extremely strong bonds [229]. Although more expensive compared to physical cross-linking, but it is often preferred for engineering of weight bearing tissues. ...
Cartilage tissue engineering is becoming increasingly popular for repairing cartilage defects. However, lack of nerves and blood vessels, as well as low cell density cause inadequate cartilage tissue regeneration. As a result, it is critical to create an appropriate microenvironment for chondrocyte proliferation. Hydrogels, as three-dimensional cross-linked polymeric networks, are considered promising alternatives to cartilage tissue due to their high-water content, viscoelasticity, and resemblance to the articular cartilage extracellular matrix. Silk fibroin hydrogels of Bombyx mori, a natural polymer, have recently been widely used to regenerate cartilage due to their ability to mimic the extracellular matrix of cartilage and endurability. For this purpose, silk fibroin is frequently combined with other materials. Composite silk fibroin hydrogels have shown strong potential for cartilage regeneration. This review explains various composite silk fibroin hydrogels studied for cartilage regeneration, as well as their advantages over single SF scaffolds.
... Both Glutaraldehyde and dialdehyde starch function by two highly reactive aldehydic groups forming covalent bonds with free amine groups on adjacent collagen peptide chains [28,37]. Glutaraldehyde is noted to be one of the most effective crosslinkers; however, unreacted molecules result in cytotoxic responses [38] when compared to dialdehyde starch, which has good biocompatibility [39]. Genipin is naturally found in Genipa americana fruit extract, with lower toxicity compared to Glutaraldehyde [40]. ...
As the most prevalent structural protein in the extracellular matrix, collagen has been extensively investigated for biofabrication-based applications. However, its utilisation has been impeded due to a lack of sufficient mechanical toughness and the inability of the scaffold to mimic complex natural tissues. The anisotropic alignment of collagen fibres has been proven to be an effective method to enhance its overall mechanical properties and produce biomimetic scaffolds. This review introduces the complicated scenario of collagen structure, fibril arrangement, type, function, and in addition, distribution within the body for the enhancement of collagen-based scaffolds. We describe and compare existing approaches for the alignment of collagen with a sharper focus on electro-compaction. Additionally, various effective processes to further enhance electro-compacted collagen, such as crosslinking, the addition of filler materials, and post-alignment fabrication techniques, are discussed. Finally, current challenges and future directions for the electro-compaction of collagen are presented, providing guidance for the further development of collagenous scaffolds for bioengineering and nanotechnology.
... The frequency of vibrations could indeed be contingent on the intensity of hydrogen bonds which stabilize the structure of blended polymers, and this analysis enables us to see whether functional groups are present in the CMCTS/HA crosslinked as well as their shift patterns, which might specify hydrogen interactions (1167 cm -1 region, C-O stretching harmonic resonance peak) [40]. The CMCTS and HA characteristic bands were seen: The amide A bond, which is close to 3355 cm -1 , has an NHstretching vibration [41]. In the combined polymers, the band positions are not significantly different. ...
... Collagen and elastin are the major structural proteins in mammals, and they provide mechanical support, strength, and elasticity for various organs and tissues. 23 Both peritoneum and pericardium are rich in collagen and elastin. In vitro enzymatic degradation experiments could provide insight into the contents of collagen and elastin in different tissues, and their ability to resist enzymatic digestion. ...
Decellularized porcine pericardium has many applications in the cardiovascular field for its excellent properties. The peritoneum is a single-layer bio-dialysis membrane with many similarities and differences in physical characteristics, biochemical composition and structure to the pericardium. The limited available literature suggests that, like the pericardium, the peritoneum has good application potential in the field of cardiovascular substitute materials. This research focused on comparing the differences between decellularized peritoneum and decellularized pericardium in microstructure, biochemical composition, mechanical properties, hemocompatibility, in vitro enzymatic degradation, in vitro calcification, cytocompatibility, and other vital indicators. The peritoneum was consistent with pericardium in terms of fibrous structure, hemocompatibility, in vitro calcification, and cytocompatibility. The peritoneal elastic fiber content (219 μg/mg) was significantly higher than that of the pericardium (66 μg/mg), resulting in 2-3 times higher maximum load (21.1 N) and burst pressure (1309 mmHg), and better performance than the pericardium in terms of in vitro resistance to enzymatic degradation. In the cardiovascular field, decellularized peritoneum can be used as vascular substitute material.
