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Effects of pepsin digestion at different temperatures and times on properties of telopeptide-poor collagen from bird feet
Abstract
This study was conducted to evaluate the effect of pepsin digestion at different temperatures (4, 12, 18 and 24°C) and times (24, 36, 48 and 72h) on properties of telopeptide-poor collagen from bird feet (TPCBF). The yield, SDS-PAGE, in vitro fibrillogenesis and denaturation temperature of the TPCBF from different treatments were determined. The results showed that the TPCBF yield increased as temperature and time increased. The yield of lots treated at 4°C were significantly lower than those of the treatments at 12, 18 and 24°C. In the SDS-PAGE electrophoretogram, the low molecular fragments of samples treated at 12, 18 and 24°C were higher than those at 4°C. The fibrillogenesis rate of lots at different temperatures decreased as treated time increased. The denaturation temperatures of the samples treated at 24°C for 48 and 72h were significantly lower than those of the treatments at 4, 12 and 18°C, for all times. These changes could be related to telopeptide removal and lost molecular integrity of collagen by pepsin digestion at a high temperature and long time of treatment. However, the optimum conditions of pepsin treatment should be controlled at 12°C for 24h due to the best fibril-forming capacity, high denaturation temperature and an acceptable yield shown in this study.
... It has been reported that functional properties of collagen, such as gelling and film-forming properties, surface behavior, and microencapsulation, differs from collagen types (Gómez-Guillén et al., 2011). Thus, numerous previous studies have been conducted to characterize the collagen type of novel sources and to evaluate its functionality with various extraction methods (Lin and Liu, 2006a;Lin and Liu, 2006b;Wang et al., 2014). In particular, enzymatic digestion, using pepsin or papain, could improve the extraction yield of collagen and its functionality, such as thermal stability (Hashim et al., 2014;Matmaroh et al., 2011;Wang et al., 2008). ...
... Thermal stability of PSCs from duck feet was determined in triplicates by using differential scanning calorimetry (DSC, thermal analyzer, TGA 2050, Mettler Toledo, USA). The heat rate and range were 5°C/min and 5-150°C (Lin and Liu, 2006b). ...
... According to Lin and Liu (2006a), Tp of PSC from chicken broiler feet was 88.77°C, which was similar to our result. The thermal stability of collagen is generally affected by amino acid composition, growing temperature of collagen source, and extracting methods (Lin and Liu, 2006a;Lin and Liu, 2006b). In particular, pepsin digestion could lead to a decrease in denaturation temperature due to decreased molecular weight of collagen subunits by the removal of telopeptides region (Nalinanon et al., 2007). ...
The objectives of this study were conducted to characterize pepsin-soluble collagen (PSC) extracted from bones (PSC-B), skins (PSC-S), and tendons (PSC-T) of duck feet and to determine their thermal and structural properties, for better practical application of each part of duck feet as a novel source for collagen. PSC was extracted from each part of duck feet by using 0.5 M acetic acid containing 5% (w/w) pepsin. Electrophoretic patterns showed that the ratio between α1 and α2 chains, which are subunit polypeptides forming collagen triple helix, was approximately 1:1 in all PSCs of duck feet. PSC-B had slightly higher molecular weights for α1 and α2 chains than PSC-S and PSC-T. From the results of differential scanning calorimetry (DSC), higher onset (beginning point of melting) and peak temperatures (maximum point of curve) were found at PSC-B compared to PSC-S and PSC-T (p<0.05). Fourier transform infrared spectroscopy (FT-IR) presented that PSC-S and PSC-T had similar intermolecular structures and chemical bonds, whereas PSC-B exhibited slight difference in amide A region. Irregular dense sheet-like films linked by random-coiled filaments were observed similarly. Our findings indicate that PSCs of duck feet might be characterized similarly as a mixture of collagen type I and II and suggest that duck feet could be used for collagen extraction without deboning and/or separation processes.
... The amide group III is associated with N-H bending and C-N stretching (Kong & Yu, 2007). According to Lin & Liu (2006), the wave spectrum between 1200-1300 cm -1 is the signature of the collagen molecule, which is related to specific tripeptides (Gly-Pro-Hyp). The triple helix structure of collagen could be seen from the Amide III wavenumber. ...
... According to Chi et al. Figure 1 Infrared spectra of salmon skin collagen extracted from acetic acid. Lin and Liu (2006) reported electrophoretic patterns of collagen with molecular weight distribution in the 25-150 kDa range. The result had similar characteristics with albacore tuna (Hema et al., 2013), dog shark (Hema et al., 2013), rohu (Hema et al., 2013) and black ruff (Bhuimbar et al., 2019) that showed collagen 1, 2 and band. ...
Collagen is a type of protein that is plentiful and has a wide variety of applications. Collagen is often derived from porcine and bovine, however, both sources are considered to transmit disease, particularly porcine sources, which haram to Moslem. There are some methods for collagen extraction, on of them using acid method. The purpose of this study was to obtain the best method of extraction and analyze collagen from alternative sources, specifically salmon skin, which is abundant. This study included two stages, i.e. pretreatment with sodium hydroxide for eliminating non-collagen proteins prior from salmon skin and extraction with acetic acid for collagen extraction. Pretreatment was carried out utilizing a split-plot design with parameters for determining the levels of non-collagen protein. The extraction step used a factorial completely randomized design to observe yields, amino acids, functional groups, and molecular weights. The results indicated that the extract was collagen. It was indicated by high glycine, alanine, and proline amino acid content. The result of SDS-PAGE showed the extracted was type I collagen was characterized by two identical 1 chains and one similar, but 2 different chains. The propeptides of type I procollagen have a similar composition. The most effective method for eliminating non-collagen proteins prior to extracting collagen from salmon skin was pretreatment with a 0.05 M NaOH solution for 10 hours. Collagen extraction using acetic acid at concentration of 0.5 M for 48 hours resulted in a yield of 12.79%.
... The amide group III is associated with N-H bending and C-N stretching (Kong & Yu, 2007). According to Lin & Liu (2006), the wave spectrum between 1200-1300 cm -1 is the signature of the collagen molecule, which is related to specific tripeptides (Gly-Pro-Hyp). The triple helix structure of collagen could be seen from the Amide III wavenumber. ...
... According to Chi et al. (2014), the structure of â (á chain dimers) indicates the presence of covalent crosslinks in the collagen molecule. Lin and Liu (2006) reported electrophoretic patterns of collagen with molecular weight distribution in the 25 -150 kDa range. The result had similar characteristics with albacore tuna (Hema et al., 2013), dog shark (Hema et al., 2013), rohu (Hema et al., 2013) and black ruff (Bhuimbar et al., 2019) that showed collagen á1, á2 and â band. ...
... However, low-temperature extraction is a time-consuming process and generates substantial undissolved residues. Previous studies reported that the recovery rate and extraction efficiency could be increased by using pepsin or extracting at a higher temperature [19,20]. Limited pepsin digestion could cleave the cross-linked collagen molecules at telopeptide regions without impairing the triple-helical structure, thus, increasing the recovery rate and further removing the antigenic P determinant located on the non-helical sections [21]. ...
... The fibril formation degree of extracted collagen was 94.42% (ASC-4), 89.62% (PSC-4), 84.36% (ASC-12), 72.80% (PSC-12), 48.31% (ASC-20), and 27.14% (PSC-20), respectively. It has been reported that fibril formation ability is related to molecular integrity [19]. ...
Acid-solubilized (ASC) and pepsin-solubilized collagen (PSC) extracted at 4 °C (ASC-4 and PSC-4), 12 °C (ASC-12 and PSC-12), and 20 °C (ASC-20 and PSC-20) from the skin of farmed pufferfish (Takifugu obscurus) was characterized by SDS-polyacrylamide gel electrophoresis (SDS-PAGE), Fourier-transform infrared spectroscopy (FTIR), and fibril-forming tests. The results indicate that extraction at 12 °C can effectively improve the extraction efficiency of natural collagen compared with extraction at 4 °C. However, extraction at 20 °C results in a decrease in molecular integrity, thus, inducing the resultant collagen to degrade or even lose fibril-forming ability. Transmission electron microscope (TEM) images revealed that ASC-4, PSC-4, ASC-12, and PSC-12 can assemble into fibrils with D-periodicities, and ASC-20 associated into molecular aggregates alongside partial D-banded fibrils, while no well-defined fibrils were observed in PSC-20. Scanning electron microscope (SEM) analysis confirmed the well-defined fibril morphologies of ASC-4, PSC-4, ASC-12, and PSC-12 with imino acid contents between 190.0 and 197.8 residues/1000 residues. The denaturation temperature of ASC-4, PSC-4, ASC-12, and PSC-12 was 30.0, 27.6, 25.9, and 22.7 °C, respectively. This study indicates that ASC and PSC extracted at 4 °C and 12 °C could be alternatives to terrestrial collagens for industrial applications.
... The first way is calculation on the basis of ratio of the lyophilisate and raw skin, showing yields on wet basis (Lin and Liu, 2006b;Loo Kiew and Mat Don, 2013;Singh et al., 2011) or dry basis (Duan et al., 2009;Nagai and Suzuki, 2000) according to following formulas: Y (wet basis) = lyophilisate (g) /raw skin mass (g) × 100, Y (dry basis) = dry lyophilisate (g)/ dry skin mass (g) × 100 and the second way is calculation on the basis of ratio of Hyp content in lyofilizate and raw skin (Kittiphattanabawon et al., 2005;Nalinanon et al., 2007;Skierka and Sadowska, 2007). ...
... They are able to bind both water and cations and form viscoelastic structures (Angele et al., 2004). It is known that enzyme cleavage sites may be blocked as a consequence of higher GAG content and collagens with higher GAG contents may be more resistant to proteolysis (Lin and Liu, 2006b). ...
The aim of this pilot study is to compare two extraction processes tested in our lab with intention to preserve the biological structure and functions of collagen-based isolate for biomedical purposes during its processing. Collagen type I was isolated from various animal sources from fish, porcine and rat skins. The resulting isolates, along with commercial calf collagens as control, were subsequently characterised and their composition examined by means of several analytical methods such as amino acid analysis, FTIR, SDS-PAGE. Moreover, the moisture, lipid, glycosaminoglycans, hydroxyproline and N contents were determined. We found that both the factors considered, the source of the collagen and the process employed, exerted effects on the final composition of the isolate and we monitored the key differences, in particular, with respect to the protein and lipid contents. Moreover, the study provides a discussion of issues surrounding the assessment of the actual pure collagen content determined on the basis of the content of hydroxyproline and the purity of the protein fraction.
... Studies investigating the preparation of polymerizable pepsin-soluble collagen 1, have demonstrated that digestion time and temperature effect the gelation behavior, physical structure, and mechanical properties of the resulting collagen 1 hydrogels. 78 Notably, studies investigating the effect of pepsin digestion on decellularized cardiac scaffolds found that the composition of the soluble ECM fraction was digestion time-dependent and that cardiomyocytes proliferated faster on tissue culture plastic coated with ECM digested for shorter times. 79 However, there is no comparable available information with respect to the effects of pepsin digestion time on subsequent properties of decellularized lung hydrogels. ...
... Several studies have demonstrated that pepsin isolation of soluble collagen and the resulting hydrogel properties are tied to digestion parameters, including time, temperature, and enzyme to substrate ratio. 78,92 In this work, we hypothesized that like collagen 1, digestion parameters play an important role in determining the properties of ECM hydrogels as well, and digestion time was selected for further evaluation. Currently, the majority of ECM hydrogels are prepared by adherence to standard 48 to 72 h long digestion, a strategy that asserts that maximizing the proportion of soluble ECM should be the main focus for preparing hydrogels. ...
Hydrogels derived from decellularized lungs are promising materials for tissue engineering, in the development of clinical therapies, and for modeling the lung extracellular matrix (ECM) in vitro. Characterizing and controlling the resulting physical, biochemical, mechanical, and biologic properties of decellularized ECM (dECM) after enzymatic solubilization and gelation is thus of key interest. As the role of enzymatic pepsin digestion in effecting these properties has been understudied, we investigated the digestion time-dependency on key parameters of the resulting ECM hydrogel. Using resolubilized homogenized decellularized pig lung dECM as a model system, significant time-dependent changes in protein concentration, turbidity, and gelation potential were found to occur between the 4 hour (h) and 24 h digestion time points, and plateauing with longer digestion times. These results correlated with qualitative SEM images and quantitative analysis of hydrogel interconnectivity and average fiber diameter. Interestingly, the time-dependent changes in the storage modulus tracked with the hydrogel interconnectivity results, while the Young's modulus values were more closely related to average fiber size at each time point. The structural and biochemical alterations correlated with significant changes in metabolic activity of several representative lung cells seeded onto the hydrogels with progressive decreases in cell viability and alterations in morphology observed in cells cultured on hydrogels produced with dECM digested for greater than 12 and up to 72 h of digestion. These studies demonstrate that 12 h pepsin digest of pig lung dECM provides an optimal balance between desirable physical ECM hydrogel properties and effects on lung cell behaviors.
... Studies investigating the preparation of polymerizable pepsin-soluble collagen 1, have demonstrated that digestion time and temperature effect the gelation behavior, physical structure, and mechanical properties of the resulting collagen 1 hydrogels. 78 Notably, studies investigating the effect of pepsin digestion on decellularized cardiac scaffolds found that the composition of the soluble ECM fraction was digestion time-dependent and that cardiomyocytes proliferated faster on tissue culture plastic coated with ECM digested for shorter times. 79 However, there is no comparable available information with respect to the effects of pepsin digestion time on subsequent properties of decellularized lung hydrogels. ...
... Several studies have demonstrated that pepsin isolation of soluble collagen and the resulting hydrogel properties are tied to digestion parameters, including time, temperature, and enzyme to substrate ratio. 78,92 In this work, we hypothesized that like collagen 1, digestion parameters play an important role in determining the properties of ECM hydrogels as well, and digestion time was selected for further evaluation. Currently, the majority of ECM hydrogels are prepared by adherence to standard 48 to 72 h long digestion, a strategy that asserts that maximizing the proportion of soluble ECM should be the main focus for preparing hydrogels. ...
... Limited studies on gelatins produced from broiler chicken feet have been reported. For example, Lin and Liu (2006) reported that collagen extracted from chicken broiler feet had higher hyproxyproline (Hyp) and proline (Pro) content and exhibited higher thermal stability. (Wangtueai and Noomhorm, 2009). ...
... From the table, it is similar with previous 43 studies on amino acids content of collagen. It was reported that aquatic animal collagen such as fish collagen contains lower imino acids compared to mammal's collagen Lin and Liu, 2006). Cheng et al. (2009) noted that silky fowl feet is a useful industrial by-product, as the collagen extraction yield is 7.3% and collagen content is 516.6 mg/g. ...
Global demand for gelatin industry is increasing especially in food and pharmaceutical industry. Major productions of gelatin are from mammalian (porcine and bovine) and the rest are from fish. There are halal issues and health related concerns associated with mammalian gelatin, while for fish gelatin is about allergic reactions. New potential gelatin sources such as poultry skin, feet, and bone has raised to replace mammalian resources. The objective of this review is to present the potentials of gelatin extracted from poultry as an alternative sources of gelatin. Poultry as an alternative doesn't have any dietary concern and can be accepted if it follows the religious requirement. Limited studies on gelatin produced from chicken show that it has higher glycine, hyproxyproline and proline content and exhibited higher thermal stability compared to mammalian and fish gelatin. Poultry gelatin from skin and feet are considered as by-products from poultry, so apart from gelatin extraction, it can minimize the product from poultry waste. This review focuses on current study of gelatin, alternatives sources of gelatin and the challenges of poultry gelatin for future commercial use.
... Conventionally, quantification of collagen extracted from various sources is done either by determining the content of one of the imino acids (Muyonga et al., 2004) known as hydroxyproline (Kolodziejska et al., 1999;Sadowska et al., 2003;Nalinanon et al., 2007) or yield by dry weight fraction resulting from weight of lyophilised collagen over dry weight of the raw material used (Lin and Liu, 2006;Wang et al., 2009;Aukkanit and Garnjanagoonchorn, 2010). Even though these methods are reported as reliable and sensitive, they are inappropriate for routine measurements since hydrolysis and drying of collagen samples are required (Komsa-Penkova et al., 1996). ...
... According to several researchers, photometry is another typical routine method used to determine the concentration of collagen in a substance (Komsa-Penkova et al., 1996;Shormanov and Bulatnikov, 2006). Measurement of UV absorbance for collagen was normally done between 230 to 280 nm wavelengths (Komsa-Penkova et al., 1996) due to the presence of phenylalanine and tyrosine which absorbed ultra-violet light at 251 and 283 nm wavelengths, respectively (Lin and Liu, 2006). In order to determine an increase of collagen concentration in extracting medium, quantitative measurements of bovine tendon collagen were conducted at the wavelength of 280 nm . ...
Acid soluble collagen (ASC) and pepsin soluble collagen (PSC) were extracted from the muscles of selected cultured freshwater fishes that are widely consumed as food in Malaysia. The species tested were Clarias species (hybrid of Clarias gariepinus × C. macrocephalus), black tilapia (Oreochromis mossambicus), pangasius catfish (Pangasius sutchi), and sultan fish (Leptobarbus hoevenii). The concentrations of extracted collagen were determined by using Lowry’s method with slight modification. Results showed that such modification was possible for measurement of collagen concentration directly from the extracting medium. The highest difference between the yields of ASC and PSC was found in Clarias spp. which was approximately 3.8-fold, though no significant increase in the yields of both collagens was achieved after 12 hours of the extraction period. At the end of the extraction process, the range of yields based on dry weight of muscles of ASC and PSC for all of the selected fish muscles were between 97.52 ± 0.97 to 139.71 ± 0.18 mg/g and 299.93 ± 0.45 to 368.36 ± 1.05 mg/g, respectively. The presence of collagen in fish muscles as reported in this study showed that these Malaysian freshwater fishes have a potential to be utilized as alternative sources of mammalian collagens.
... The collagen extracted from Heterotis niloticus has a greater denaturation temperature, which indicates that it is more stable in a high-temperature environment [52]. Nonetheless, the study's findings regarding the denaturation temperature of collagen derived from skin and scale of Heterotis niloticus and Lates niloticus were marginally lower than those of pork collagen (66.4°C) and calf skin collagen (67.84°C) as well as Nile tilapia fish (41.09°C) [53][54][55][56][57][58][59][60][61][62][63][64]. ...
Collagen is the principal segment of the extracellular grid that constitutes roughly 30% of the aggregate protein in vertebrates. It can be generally utilized as part of sustenance, medication, beauty care products and biomedical materials. There is developing enthusiasm for the procedures used to extricate collagen and its subsidiaries, because of the growing inclination to utilize this protein. A set up of manufactured operators of different modern procedures have been put in place. The aim of this study was to extract and characterize collagens from wastes (skin and scale) of two (2) selected Nigerian freshwater fish species (Heterotis niloticus and Lates niloticus) using either pepsin or acid soluble extraction. Proximate analysis of the waste (skin and scales) of both fishes were conducted, collagen was extracted using 0.5M acetic acid and pepsin. The collagen yield was determined and characterized by Sodium Dodecyl Sulfate Polyacrylamide Gel Electrophoresis (SDS PAGE), amino acid profile and Fourier Transform-Infra Red Spectrum (FTIR). Proximate composition of the samples revealed that at p<0.05 significant level; Ash content was higher in the scales (24.80±3.45% and 28.68±2.34%) than the skin (2.31±0.12%-3.03±0.35%). while fiber content was higher in the scales (0.45±0.02%-0.25±0.01%) than the skin (2.31±0.12%-3.03±0.35%). However, the proteins were higher in the skins (30.44±3.43%-44.47±4.32%) than the scale (22.39±1.23 % - 18.44±0.34%). Also fat were higher in the skin (14.91±1.34%-8.66±1.03%) than the scale 0.81±0.19% - 5.77±0.56%. Collagen extraction yields varied with the extraction process, yield was significantly (p<0.05) higher in skin (5.08±0.34% - 33.97±1.78%) than the scale (1.76 %-8.05%). However, acid soluble collagen isolated from the scale of Heterotis niloticus had the highest (p<0.05) amino acid profile content of 74.95±2.84 %, followed by pepsin soluble collagen isolated from the skin of Lates niloticus (57.09±2.65 %), while acid soluble collagen isolated from skin of Heterotis niloticus and skin of Lates niloticus had the least amino acid contents of 31.60±0.52% and 29.48±1.03% respectively. FTIR spectra of the collagens shows characteristic peaks of amides A, B, I, II and III. The result of the present study reveals the existence of helical arrangements of collagen and thus may be deduced that the skin and scale of Lates niloticus and Heterotis niloticus appears to be a good alternative source of high-quality collagen for industries.
... The physicochemical properties of various collagen samples depend on the sources [38]. Collagen obtained from sea animals possesses differing molecular structure to those obtained from domestic and land animals [39]. Collagen, primarily obtained from animal connective tissues, can be processed into nanofibers through electrospinning or self-assembly methods. ...
The potentials of nanofiber in food technology cannot be over-emphasized. Nanomaterials have become a versatile resource of multiple applications. The various adaptability of nanofibers is attributable to its special qualities including controlled shape, high porosity, large surface area, high encapsulation efficiency, and high chemical and thermal stability. Functionalization adds more value to a nanofiber through the introduction of additional moieties to it. Food technology is a wide field of research due to its importance in the society. In recent years, new measures are being introduced to produce new smart food resources. This chapter provides a comprehensive overview of how nanofibers are being prepared, functionalized, and applied within the realm of food technology. The various techniques and reagents used for nanofiber functionalization suitable for food technology are discussed. The advantages inherent in functionalized nanofibers are also elucidated. The chapter closes with future recommendations on functionalized nanofibers for food technology.
... In this method, when radiation is emitted on extracts, a vibration of atoms and their groups of organic compounds occurs, facilitating the identi cation of the bands. This happens in each change in the level of vibrational energy, in uencing changes in rotational level causing the printing and constitution of the data and bands observed in the spectrum (35,36). ...
The aim of the study was to extract and characterize collagen from solid untanned skin waste from the processing of leather in a Capra aegagrus tannery. Using pepsin (SPC), 37 grams of collagen were obtained from 100 grams of dry weight skin. Characterization took place using SDS-PAGE, FTIR and UV absorption techniques, identifying it as type I collagen. The ultraviolet (UV) absorption spectrum showed a peak at 238 nm. In the thermogram, the maximum transition temperature was 56º C. Using the electrophoresis technique, it was observed that SPC consists of band patterns formed by a γ chain, a β chain and two distinct α chains (α1 and α2). In the FTIR analysis, the collagen showed the absorption peaks for the amides, showing that the SPC extraction process maintained the integrity of the molecule. To observe the effect of NaCl concentration on the solubility of SPC, the collagen showed high solubility, up to a concentration of 2% NaCl. The solubility peak was observed at pH 4.0, with a sharp drop until pH 7.0, reaching its minimum point at pH 10. Scanning microscopy showed some irregular surfaces, cavities and fibrous structures, which may favor the application of collagen as a biomaterial. The zeta potential found the isoelectric point of collagen at pH = 6.61. These results indicate that the collagen obtained has a high level of structural integrity and can be applied as an alternative source, as well as adding value to a waste product that is often discarded in the environment.
... Another problem is the heat stability of the particular collagen type and its denaturation temperature, above which the structure of the collagen molecule is no longer preserved, and durability and scaffold function cannot be granted [90,91]. Fish-derived collagen generally has a lower denaturation temperature than mammalian products [3]. ...
Bone defects resulting from trauma, surgery, and congenital, infectious, or oncological diseases are a functional and aesthetic burden for patients. Bone regeneration is a demanding procedure, involving a spectrum of molecular processes and requiring the use of various scaffolds and substances, often yielding an unsatisfactory result. Recently, the new collagen sponge and its structural derivatives manufactured from European carp (Cyprinus carpio) were introduced and patented. Due to its fish origin, the novel scaffold poses no risk of allergic reactions or transfer of zoonoses and additionally shows superior biocompatibility, mechanical stability, adjustable degradation rate, and porosity. In this review, we focus on the basic principles of bone regeneration and describe the characteristics of an “ideal” bone scaffold focusing on guided bone regeneration. Moreover, we suggest several possible applications of this novel material in bone regeneration processes, thus opening new horizons for further research.
... Pepsin, depending on the pH of its surrounding environment, can engage in a variety of actions, and its capacity to digest ACM can also change [39]. Although enzymatic digestion was more complete in 1 N HCl than in the acid, experimental data showed that there were still particles in 0.01 N HCl and 0.1 N HCl that have not been entirely digested. ...
Decellularized matrices can effectively reduce severe immune rejection with their cells and eliminated nucleic acid material and provide specific environments for tissue repair or tissue regeneration. In this study, we prepared acellular cartilage matrix (ACM) powder through the decellularization method and developed ACM hydrogels by physical, chemical, and enzymatic digestion methods. The results demonstrated that the small size group of ACM hydrogels exhibited better gel conditions when the concentration of ACM hydrogels was 30 and 20 mg/mL in 1N HCl through parameter adjustment. The data also confirmed that the ACM hydrogels retained the main components of cartilage: 61.18% of glycosaminoglycan (GAG) and 78.29% of collagen, with 99.61% of its DNA removed compared to samples without the decellularization procedure (set as 100%). Through turbidimetric gelation kinetics, hydrogel rheological property analysis, and hydrogel tissue physical property testing, this study also revealed that increasing hydrogel concentration is helpful for gelation. Besides, the ex vivo test confirmed that a higher concentration of ACM hydrogels had good adhesive properties and could fill in cartilage defects adequately. This study offers useful information for developing and manufacturing ACM hydrogels to serve as potential alternative scaffolds for future cartilage defect treatment.
... Collagen obtained from different sources differs in their physicochemical properties slightly [125]. Collagens obtained from frog skin, bird feet, shark skin, and sea urchin has a molecular structure that is different from those obtained from domestic animals [126][127][128]. Furthermore, their thermal property, peptide constitution, amino acid composition, and content of glycosaminoglycan are significantly different from collagen isolated from land animals [126]. ...
Skin regeneration after an injury is very vital, but this process can be impeded by several factors. Regenerative medicine is a developing biomedical field with the potential to decrease the need for an organ transplant. Wound management is challenging, particularly for chronic injuries, despite the availability of various types of wound dressing scaffolds in the market. Some of the wound dressings that are in clinical practice have various drawbacks such as poor antibacterial and antioxidant efficacy, poor mechanical properties, inability to absorb excess wound exudates, require frequent change of dressing and fails to offer a suitable moist environment to accelerate the wound healing process. Collagen is a biopolymer and a major constituent of the extracellular matrix (ECM), making it an interesting polymer for the development of wound dressings. Collagen-based nanofibers have demonstrated interesting properties that are advantageous both in the arena of skin regeneration and wound dressings, such as low antigenicity, good biocompatibility, hemostatic properties, capability to promote cellular proliferation and adhesion, and non-toxicity. Hence, this review will discuss the outcomes of collagen-based nanofibers reported from the series of preclinical trials of skin regeneration and wound healing.
... However, it should be emphasized that the amount of pepsin used in collagen extraction and raw materials rich in collagen was quite large, because for poultry or fish skins, smaller amounts of enzyme are to be used, e.g., about 100 mg/g [8,20,21]. Additional enzyme (300 mg/g) was used in the process of collagen recovery from broiler legs [22], and for mineralized tissues such as bone elements, it was even over 1000 mg/g [23]. Significant differences were found in the amount of obtained fibers depending on the method of proteoglycan removal ( Table 3). ...
The aim of the presented research was to obtain reconstituted atelocollagen fibers after extraction from poultry cartilage using the pepsin-acidic method in order to remove telopeptides from the tropocollagen. Firstly, we examined the extraction of collagen from the cartilage extracellular matrix (ECM) after proteoglycans (PG) had been removed by the action of salts, i.e., NaCl or chaotropic MgCl2. Additionally, the effects of the salt type used for PG and hyaluronic acid removal on the properties of self-assembled fibers in solutions at pH 7.4 and freeze-dried matrices were investigated. The basic features of the obtained fibers were characterized, including thermal properties using scanning calorimetry, rheological properties using dynamic oscillatory rheometry, and the structure by scanning electron microscopy. The fibers obtained after PG removal with both analyzed types of salts had similar thermal denaturation characteristics. However, the fibers after PG removal with NaCl, in contrast to those obtained after MgCl2 treatment, showed different rheological properties during gelatinization and smaller diameter size. Moreover, the degree of fibrillogenesis of collagens after NaCl treatment was complete compared to that with MgCl2, which was only partial (70%). The structures of fibers after lyophilization were fundamentally different. The matrices obtained after NaCl pretreatment form regular scaffolds in contrast to the thin, surface structures of the cartilage matrix after proteoglycans removal using MgCl2.
... The degree of telopeptide variation is much greater, which is considered to be the main determinant of Col II immunogenicity. Some studies have tried to reduce the immunogenicity by using proteolytic enzymes (e.g., pepsin) to remove the terminal telopeptides, resulting in what is known as Atelocollagen (Lin and Liu, 2006;Jeevithan et al., 2015; Figure 2B). However, there is still a lack of scientific evidence regarding the effect of this treatment on collagen immunogenicity. ...
Biomaterials play a core role in cartilage repair and regeneration. The success or failure of an implanted biomaterial is largely dependent on host response following implantation. Host response has been considered to be influenced by numerous factors, such as immune components of materials, cytokines and inflammatory agents induced by implants. Both synthetic and native materials involve immune components, which are also termed as immunogenicity. Generally, the innate and adaptive immune system will be activated and various cytokines and inflammatory agents will be consequently released after biomaterials implantation, and further triggers host response to biomaterials. This will guide the constructive remolding process of damaged tissue. Therefore, biomaterial immunogenicity should be given more attention. Further understanding the specific biological mechanisms of host response to biomaterials and the effects of the host-biomaterial interaction may be beneficial to promote cartilage repair and regeneration. In this review, we summarized the characteristics of the host response to implants and the immunomodulatory properties of varied biomaterial. We hope this review will provide scientists with inspiration in cartilage regeneration by controlling immune components of biomaterials and modulating the immune system.
... Therefore, in most cases, enzymes are used to obtain specific protein products, high yield and reduced wastes, as well as a decrease in Mar. Drugs 2020, 18, 214 7 of 23 the antigenicity caused by telopeptides [68][69][70]. However, when a high amount of pepsin is used for long time, PSC yield may be lower because the collagen is likely cleaved, impairing the triple helix's integrity [62]. ...
Due to its unique properties, collagen is used in the growing fields of pharmaceutical and biomedical devices, as well as in the fields of nutraceuticals, cosmeceuticals, food and beverages. Collagen also represents a valid resource for bioplastics and biomaterials, to be used in the emerging health sectors. Recently, marine organisms have been considered as promising sources of collagen, because they do not harbor transmissible disease. In particular, fish biomass as well as by-catch organisms, such as undersized fish, jellyfish, sharks, starfish, and sponges, possess a very high collagen content. The use of discarded and underused biomass could contribute to the development of a sustainable process for collagen extraction, with a significantly reduced environmental impact. This addresses the European zero-waste strategy, which supports all three generally accepted goals of sustainability: sustainable economic well-being, environmental protection, and social well-being. A zero-waste strategy would use far fewer new raw materials and send no waste materials to landfills. In this review, we present an overview of the studies carried out on collagen obtained from by-catch organisms and fish wastes. Additionally, we discuss novel technologies based on thermoplastic processes that could be applied, likewise, as marine collagen treatment.
... Fingertips of chicken feet were excised, then the feet were skinned, washed three times using tap water, cut into 2 cm pieces, soaked in 1.5 L water at room temperature (25 ± 2 ºC) for 3 hr to remove blood. Then, ground by meat grinder and kept frozen at -20ºC in polyethylene bags until further processing according to the method described by Lin & Liu (2006). ...
Poultry by-products are not often processed into high-value products. One important application is the production of high quality chicken gelatin to meet the needs of markets that are not amenable to beef gelatin. So, the present study deals with the production of gelatin from chicken skin, feet and bone. The extraction process of gelatin from chicken by-products was optimized through the use of alkali (NaOH) and its effects on the protein yields and the physicochemical properties of the produced gelatin were investigated. As soaking period proceeded as yield decreased. The optimum sodium hydroxide concentration and liming period for gelatin production of chicken tissues were defined as 2.5 % for 60 h. Yield of gelatin of chicken skin, feet and bone reached to the maximum at 60°C and 6h extraction time. Skin, feet and bone gelatins had similar amino acid composition, with a total imino acid content of about (12.92-16.99%). Amino acid contents of chicken tissue gelatins were different from that of commercial gelatin due to the differences in raw materials and production process.
... This has indicated that the in vitro application of GSC will be superior to the application of GCC and CCC. The intertwined fibril network can be widely used in biological and pharmaceutical applications [44][45][46]. ...
Collagens were extracted from grass carp skin (GCC), grass carp scales (GSC), and crucian carp skin (CCC) using an acid-enzyme combination method, and their characteristics and self-assembly properties were analyzed. Electrophoretic patterns characterized all three as type I collagens. An ultraviolet analysis identified the optimal wavelengths for collagen detection, while a Fourier transform infrared spectroscopy analysis confirmed the triple-helical structure of the collagens. The GCC, GSC, and CCC had denaturation temperatures of 39.75, 34.49, and 39.05 °C, respectively. All three were shown to self-assemble into fibrils at 30 °C in the presence of NaCl, but the fibril formation rate of CCC (40%) was slightly higher than those of GCC (28%) and GSC (27%). The GSC were shown to form a more strongly intertwined fibril network with a characteristic D-periodicity. The fish collagens extracted in this study have potential applications in the development of functionalized materials.
... Among these enzymes, pepsin is the most commonly used enzyme for MC extraction. In addition, enzyme treatment can reduce the antigenicity caused by telopeptides [52]. Besides telopeptides, antigenicity related to noncollagenous proteins, cells, and cell remnants can be removed by the above-mentioned method of NaOH treatment [53]. ...
This review focuses on the expanding role of marine collagen (MC)-based scaffolds for biomedical applications. A scaffold—a three-dimensional (3D) structure fabricated from biomaterials—is a key supporting element for cell attachment, growth, and maintenance in 3D cell culture and tissue engineering. The mechanical and biological properties of the scaffolds influence cell morphology, behavior, and function. MC, collagen derived from marine organisms, offers advantages over mammalian collagen due to its biocompatibility, biodegradability, easy extractability, water solubility, safety, low immunogenicity, and low production costs. In recent years, the use of MC as an increasingly valuable scaffold biomaterial has drawn considerable attention from biomedical researchers. The characteristics, isolation, physical, and biochemical properties of MC are discussed as an understanding of MC in optimizing the subsequent modification and the chemistries behind important tissue engineering applications. The latest technologies behind scaffold processing are assessed and the biomedical applications of MC and MC-based scaffolds, including tissue engineering and regeneration, wound dressing, drug delivery, and therapeutic approach for diseases, especially those associated with metabolic disturbances such as obesity and diabetes, are discussed. Despite all the challenges, MC holds great promise as a biomaterial for developing medical products and therapeutics.
... From the above results concerning the higher yield and characterization of the PSC extracts, conjugated with the potential low immune responses upon in-vivo implantation when removed the telopeptides [74], this material was selected to proceed with the study for evaluation of biomedical potential as component on the development of implanted scaffolds for histological analyses in tissue engineering strategies. Additionally, there are studies that claim atelocollagens, due to removal of telopeptides through pepsin digestion, present a negligible level of immunogenicity as the majority of the antigenic sites were believed to be present on the telopeptide chains [39]. ...
Aquatic origin collagen (Aqua-collagen) has been pursued as an alternative to mammalian origins. Acid-soluble collagen (ASC) and pepsin-soluble collagen (PSC) were successfully extracted from the skin of tilapia with the yield about 19–20% (basis of lyophilized dry weight), and examined for their physico-functional and structural properties. Both ASC and PSC containing α1, α2 chains were characterized to be type I collagen and had lower denaturation temperature compared to mammalian origins. PSC (low telopeptides collagen) was selected for biomedical scaffolds construction due to its low immunogenicity. SEM analysis of fish collagen scaffolds showed a wide range of pore size distribution, high porosity, and high surface area-to-volume ratios. The tilapia collagen microfiber matrix scaffolds were grafted beneath the dorsal skin in 96 mice towards tissue regeneration, with bovine collagen microfiber matrix scaffolds (Avitene™ UltraFoam™ sponge) serving as control. Biocompatibility evaluation in the dorsal tissue showed that implanted scaffolds degraded completely after 20 days with no pathological inflammatory tissue responses. These findings indicated that aqua-collagens microfiber matrix scaffolds were highly biocompatible in nature, exploring its feasibility for the development of scaffolds in tissue engineering.
... Thus, pepsin provokes not only the cleavage of the collagens teleopeptide region maximising their solubility, but also the hydrolysis of non-collagenous proteins increasing collagen purity. In this case, pepsin maximise the extraction yield of collagens while reducing their antigenicity [26,27]. ...
The acid solubilised collagen (ASC) and pepsin solubilised collagen (PSC) were extracted from the by-products (skin) of a cartilaginous fish (Mustelus mustelus). The ASC and PSC yields were 23.07% and 35.27% dry weight, respectively and were identified as collagen Type I with the presence of α, β and γ chains. As revealed by the Fourier Transform Infrared (FTIR) spectra analysis, pepsin did not alter the PSC triple helix structure. Based on the various type of collagen yield, only PSC was used in combination with chitosan to produce a composite film. Such film had lower tensile strength but higher elongation at break when compared to chitosan film; and lower water solubility and lightness when compared to collagen film. Equally, FTIR spectra analysis of film composite showed the occurrence of collagen-chitosan interaction resulting in a modification of the secondary structure of collagen. Collagen-chitosan-based biofilm showed a potential UV barrier properties and antioxidant activity, which might be used as green bioactive films to preserve nutraceutical products.
... Pepsin is necessary to hydrolyze protein in cross-linking sites because the collagen triple helix structure protects of the enzymatic action (Simões et al., 2014). Enzyme activity can be increased at low pH and temperature close to the human body, but at low temperature it is possible to obtain native collagen (Lin & Liu, 2006b). ...
The objective of this study was to extract collagen from chicken feet, determining optimal extraction conditions according to acetic acid concentration, pepsin content and time of enzymatic hydrolysis. A factorial design 23 was used, with three replications at the central point, totaling 11 experiments. The response variable studied was the collagen content of the isolate obtained. In addition to the optimization, the characterization of the isolates with higher and lower collagen content, in relation to the amino acid profile, electrophoretic profile, peptide hydrophobicity and functional properties, such as water solubility, water retention capacity and emulsifying activity, were carried out. The proposed model was statistically significant, with conditions of higher collagen content of 0.3 mol/L of acetic acid, 0.2% of pepsin and 12 hours of hydrolysis. The collagen isolate under these conditions showed higher iminoacids content, higher sum of peptide areas, higher solubility in water and water retention at 60 °C. The treatment with lower collagen content showed high emulsifying activity. The collagen isolate of the chicken feet presented characteristics makes it suitable for application in the food industry.
... The papain hydrolysis showed the presence of dispersed molecular weight bands which was due to cleavage of telopeptide regions by enzyme, leading to slightly lower molecular weight components of collagen (Chuaychan, Benjakul, & Kishimura, 2015). Lin and Liu (2006) reported the electrophoretic pattern of collagen with molecular weight distribution in the range of 25-150 kDa. ...
The extraction of collagen from chicken feet by papain hydrolysis was optimized at various temperatures, time and solid to solvent ratios. The optimum extraction condition (with highest yield, 32.16%, w/w) was found after 28 h of enzymatic hydrolysis at 30 ⁰C. The chicken feet collagen was found rich in glycine (16.30 %), hydroxyproline (14.15 %) and proline (8.70 %). The molecular weight distribution of isolated collagen was found in the range of 25-150 KDa. The functional characteristics of chicken feet collagen such as solubility, emulsification, foaming, water and oil holding capacity indicate its potential application in food, pharmaceuticals and cosmetics products. The fibres were also developed by electrospinning from chicken feet extracted collagen and polycaprolactone. Scanning electron micrographs demonstrated the interconnected network of porous structure of collagen and its bio-polymeric fibres. The slight shifting and sharp absorption of peaks obtained from biopolymeric collagen presented the evidence of interactions, occurred between the polycapralactone and the collagen.
... Poultry by-products including skin and feet contains large amounts of collagen. Several studies were done on poultry such as chicken skin [7][8][9][10], chicken feet [8,11,12], bird feet [13], silky fowl feet [14] and duck feet [3,15,16]. Huda et al. [17] reported that duck feet collagen can improve the physicochemical properties of sardine surimi. ...
The original version of this article unfortunately contained a mistake. The first author name was incorrectly submitted and published as Nik Aisyah, N.M. instead of Nik Muhammad, N. The original article has been corrected.
... Poultry by-products including skin and feet contains large amounts of collagen. Several studies were done on poultry such as chicken skin [7][8][9][10], chicken feet [8,11,12], bird feet [13], silky fowl feet [14] and duck feet [3,15,16]. Huda et al. [17] reported that duck feet collagen can improve the physicochemical properties of sardine surimi. ...
This study was conducted to characterize the physicochemical properties of duck feet gelatin extracted by using different acid solutions which is hydrochloric acid (DFCl), acetic acid (DFAa), lactic acid (DFLa), and citric acid (DFCa). Proximate analysis, bloom strength, amino acid composition and sensory profiles were evaluated. Duck feet gelatin had higher bloom strength with 225.53, 334.17, 322.17 and 322.63 g for DFCl, DFAa, DFLa and DFCa, respectively, compared to commercial bovine gelatin with 216.63 g. DFCa had the highest imino acid with value of 23.01% compared to other gelatin sample. Sensory profiles in powder and gels form of gelatin were evaluated by 12 trained panellists using quantitative descriptive analysis (QDA). Overall, gelatin powder of DFCl had the highest degree of acceptability with average score of 11.15, but there’s no significant different between DFCa. While for gelatin gel, DFCa had the highest intensity of fracturability, firmness and the overall degree of acceptability score is 10.48, but there is no significant difference among other acid treatments. It was found that all extracted duck feet gelatin has high potential for application as an alternative to commercial gelatin that already available in market.
... Extraction of gelatin from chicken feet has been done by using alkaline treatment (Rahman and Jamlulail, 2012) and acid treatment (Sarbon et al., 2013). Collagen extracted from chicken broiler feet was reported to have higher hyproxyproline and proline content and exhibited higher thermal stability (Lin and Liu, 2006). ...
Fat reduction has been performed on crude gelatin extracted from chicken feet (shank) through curing process using sodium hydroxide and acetic acid in various proportions. This study was conducted to evaluate the best fat reduction condition by the addition of acetic acid at concentrations of 0, 2.5 and 5% (v/v) with decantation time 0, 6 and 12 hours on crude gelatine from curing process at a ratio of 1% acetic acid; 1% acetic acid with 1% NaOH; 1% acetic acid and 3% NaOH and without curing (initial gelatine). The analysis was carried out to determine the composition of gelatin and their byproducts to total solids, total fat and fat binding capacity. The results showed that the variation in the ratio of the addition of acetic acid and decantation time affect the composition of fat and fat-binding ability. Based on fat binding capacity (FBC), treatment of chicken feet gelatin with curing of 1% acetic acid and 3% NaOH and continued by the addition of 5% (v/v) acetic acid with a 12-hour decantation time resulted in gelatin with the best FBC of 7.4 g/g, fat content 1.0248% and total solids 2.7575%.
DOI : 10.15408/jkv.v0i0.3149
... (3) reduce antigenicity cause by telopeptide in the collagen which serve as the major problem in food and pharmaceutical applications (Werkmeister and Ramshaw 2000, Lee et al. 2001, Lin and Liu 2006, Cao and Xu 2008. Pepsin, particularly pepsin from porcine stomach, has been used to maximize the extraction efficiency of collagen from several species such as largefin (Nagai and Suzuki 2002), and common minke whale (Nagai et al. 2008). ...
Fish collagen has gained increasing interest as the alternative for mammalian counterpart. It can be generally produced from by-products generated during processing of fish and invertebrates. The potential raw materials include skin, bone, scale, and so on. Types and molecular properties of collagen vary with the source, habitat of fish, extraction process, and other factors. In general, collagen can be extracted from collagenous materials at low temperature with the aid of various acids to avoid thermal denaturation. To increase the extraction yield, pepsins from mammalian and fish origins, which specifically cleave at telopeptide region, have been used successfully without the changes in molecular properties. Fish collagen can be of food, biomedical, and pharmaceutical applications.
... Severe conditions will impair the integrity of the collagen; therefore, most studied of collagen extraction have been limited to low temperature (Lin, 2006). From extracted collagen, it was expected to see the col1α1 and col1α2 strands running separately on the gel by using denaturing sample buffer (containing DTT) for running gel, as it promotes the collagen synthesis, but since there are two col1α1 and strands and one col1α2 strand in each collagen molecule, col1α1 is more expected to be present, as in be darker and thicker. ...
... This treatment is very useful, since it cleaves peptides specifically in telopeptide region of collagen, which are non-helical ends, and thus, by hydrolyzing some non-collagenous proteins, increases the purity of collagen. It results in a much more efficient collagen extraction, since it makes the sample ready to solubilize while reducing, at the same time, the antigenicity caused by telopeptides [43,44]. For this reason, it is common to use this proteolytic procedure after the extraction of ASC, obtaining, thus, the abovementioned PSC. ...
Collagens are the most abundant high molecular weight proteins in both invertebrate and vertebrate organisms, including mammals, and possess mainly a structural role, existing different types according with their specific organization in distinct tissues. From this, they have been elected as one of the key biological materials in tissue regeneration approaches. Also, industry is constantly searching for new natural sources of collagen and upgraded methodologies for their production. The most common sources are from bovine and porcine origin, but other ways are making their route, such as recombinant production, but also extraction from marine organisms like fish. Different organisms have been proposed and explored for collagen extraction, allowing the sustainable production of different types of collagens, with properties depending on the kind of organism (and their natural environment) and extraction methodology. Such variety of collagen properties has been further investigated in different ways to render a wide range of applications. The present review aims to shed some light on the contribution of marine collagens for the scientific and technological development of this sector, stressing the opportunities and challenges that they are and most probably will be facing to assume a role as an alternative source for industrial exploitation.
... This was most likely due to the formation of low molecular weight fragments at the higher extraction temperature. Lin and Liu (2006) observed higher amounts of lower molecular weight fragments when the digestion temperature increased due to a loss of integrity. ...
This study was conducted to evaluate the effects of soaking pH and extraction temperature on the physicochemical properties of chicken skin gelatin. In order to extract gelatin from chicken skin, the chicken skin was soaked at various pH ranges (1-13) and was extracted at 75 and . For the rate of weight increase, the highest value was obtained from two pH ranges (1-2 and 12-13). In addition, the rate of weight increase was affected by soaking time. The alkali treatments had greater crude protein content as well as total extraction yield compared to the acid process (p
... At a higher temperature and longer time of pepsin digestion, larger amounts of telopeptide of tropocollagen would be digested and resulted in collagen with less fibril forming capacity. The fibril forming capacity is an important index of collagen molecular integrity and the denaturation of collagen caused a reduction in fibril-forming capacity (Lin and Liu, 2006). Higher extraction temperature and longer extraction time would lead to severe and serious pepsin digestion which caused the possibility of collagen to lose all their fibril forming capacity. ...
Extraction of collagen from muscles of cultured catfish (Clarias gariepinus × C. macrocephalus) with the aid of pepsin digestion was investigated using a statistical tool. Fractional factorial design (FFD) was applied to evaluate the effects of eight process parameters: acetic acid concentration, acid extraction time, acid extraction temperature, acetic acid to muscles ratio, NaOH concentration, NaOH to muscles ratio, NaOH treatment time, and stirring speed. Contribution of every parameter in influencing the extraction efficiency was evaluated and factors that significantly affected the extraction were elucidated by employing experimental design and analysis of variance in FFD. The result of first order factorial design showed that acetic acid concentration, acid extraction time, acid to muscles ratio, and stirring speed had significant effect (P < 0.05) to the yield of pepsin soluble collagen (PSC) obtained at the end of the experiment. Effects of these process factors on the efficiency of collagen extraction were investigated, and are discussed in detail. Optimum conditions were found at 0.5 M acetic acid, 16 hr extraction period, solvent to muscles ratio at 25 ml/g, and stirring speed of 400 rpm, resulting in yield of PSC as high as 211.49 ± 15.51 mg/g.
Collagen is a key component of connective tissue and has been frequently used in the fabrication of medical devices for tissue regeneration. Human-originated collagen is particularly appealing due to its low immune response as an allograft biomaterial compared to xenografts and its ability to accelerate the regeneration process. Ethically and economically, adipose tissues available from liposuction clinics are a good resource to obtain human collagen. However, studies are still scarce on the extraction and characterization of human collagen, which originates from adipose tissue. The aim of this study is to establish a novel and simple method to extract collagen from human adipose tissue, characterize the collagen, and compare it with commercial-grade porcine collagen for tissue engineering applications.
We developed a method to extract the collagen from human adipose tissue under quasi-Good Manufacturing Practice (GMP) conditions, including freezing the tissue, blood removal, and ethanol-based purification. Various techniques, including protein quantification, decellularization assessment, SDS-PAGE, FTIR, and CD spectroscopy analysis, were used for characterization. Amino acid composition was compared with commercial collagen. Biocompatibility and cell proliferation tests were performed, and in vitro tests using collagen sponge scaffolds were conducted with statistical analysis.
Our results showed that this human adipose-derived collagen was equivalent in quality to commercially available porcine collagen. In vitro testing demonstrated high cell attachment and the promotion of cell proliferation.
In conclusion, we developed a simple and novel method to extract and characterize collagen and extracellular matrix from human adipose tissue, offering a potential alternative to animal-derived collagen for xeno-free tissue engineering applications.
Rhizostoma pulmo is a jellyfish from the Black Sea basin that can be a source of natural bioactive compounds with substantial beneficial implications. It is important to use under-exploited marine resources in areas such as pharmaceuticals industry, medicine, cosmetics and dermatology. Marine collagen can be obtained from various sources. Several studies have focused on marine collagen, namely its extraction from alternative sources: fish, invertebrate marine animals such as sea sponges or jellyfish. The novelty is the extraction of marine collagen obtained from jellyfish of the species Rhizostoma pulmo found in the Black Sea and along the coast, the physico-chemical characterization, comparison with other types of collagenic extracts from fish and finally the formulation of a pharmaceutical preparation with medical applications.
Tissues rely on collagen for structural and biological integrity, as well as for function and strength. Several collagen sources have been investigated due to its wide variety of applications, such as collagen from cows and pigs. However, mammalian-based collagen has been limited by diseases like bovine spongiform encephalopathy (BSE) and other religious limitations. Hence, fish collagen has caught the attention of the research community because it is easy to extract, has a high level of collagen content, excellent absorption properties, a low molecular weight, biocompatibility, little risk of disease transmission from animals to humans, negligible environmental contamination, and fewer ethical and religious concerns, posing as an ideal resource for product development. This review focuses on the growing role of marine collagen in the advances of various biomedical applications, such as drug delivery, tissue engineering, regeneration, and wound healing, which will be covered in depth.
Decellularized extracellular matrix (dECM) hydrogels are being increasingly investigated for use in bio-inks for three-dimensional cell printing given their good cytocompatibility and biomimetic properties. The osmotic pressure and stiffness of bio-ink are important factors affecting the biological functions of printed cells. However, little attention has been given to the osmotic pressure and stiffness of the dECM bio-inks. Here, we compared three types of commonly used acidic solutions in the bio-fabrication of a tendon derived dECM bio-ink for 3D cell printing (0.5 M acetic acid, 0.1 M hydrochloric acid and 0.02 M hydrochloric acid). We found that low pH value of 0.1 M hydrochloric acid could accelerate the digestion process for dECM powders. This could lead to a much softer dECM hydrogel with storage modulus less than 100 Pa. This soft dECM hydrogel facilitated the spreading and proliferation of stem cells encapsulated within it. It also showed better tendon-inducing ability compared with two others much stiffer dECM hydrogels. However, this over-digested dECM hydrogel was more unstable as it could shrink with the culture time going on. For 0.5 M acetic acid made dECM bio-ink, the hyperosmotic state of the bio-ink led to much lower cellular viability rates. Postprocess (Dilution or dialysis) to tailor the osmotic pressure of hydrogels could be a necessary step before mixed with cells. Thus, kindly choosing the type and concentration of acidic solution is necessary for dECM bio-ink preparation. And a balance should be made between the digestion period, strength of acidic solution, as well as the size and concentration of the dECM powders.
Statement of significance
The dECM bio-ink has been widely used in 3D cell printing for tissue engineering and organ modelling. In this study, we found that different types of acid have different digestion and dissolution status for the dECM materials. A much softer tendon derived dECM hydrogel with lower stiffness could facilitate the cellular spreading, proliferation and tendon differentiation. We also demonstrated that the osmotic pressure should be taken care of in the preparation of dECM bio-ink with 0.5 M acetic acid. Thus, kindly choosing the type and concentration of acidic solution is necessary for dECM bio-ink preparation.
Collagen type II (CT-II) has unique biological activities and functions, yet the knowledge on amphibian-derived CT-II is rare. Herein, acid-soluble (ASC) and pepsin-soluble collagen (PSC) were successfully isolated and characterized from the cartilage of Chinese Giant Salamander (CGS). The in vitro immunogenicity of collagen was then evaluated and compared with that of the standard bovine CT-II (SCT-II) by T-lymphocyte cell proliferation activity. Results demonstrated that ASC and PSC were predominantly CT-II along with minor collagen type I and maintained intact triple-helical structure of nature collagen. Compared with SCT-II, higher glycine content (337.80 and 339.93 residues/1000 residues) and lower degree of proline hydroxylation (51.81% and 52.52%) were observed in ASC and PSC. Additionally, PSC showed comparable Td (63 °C) and higher Tm (109 °C) than SCT-II (64 °C and 103 °C, respectively), indicating its high thermal and structural stability. SEM revealed that the lyophilized ASC and PSC had interconnected porous network structures of collagen-based materials. Moreover, different from SCT-II, both ASC and PSC presented no immunogenicity because they did not cause obvious proliferation of murine T-lymphocyte regardless of the induced concentration of collagen increased from 8 to 417 μg/mL. These data suggested that the amphibian-derived CGS cartilage collagens avoid the immunogenic risk of terrestrial animal collagen, and show high thermal stability and potential advantage in biomedical application.
In recent years, tissue engineering has helped to reduce hospital stays and deaths caused by skin wounds. Scaffolds are one of the main factors that influence the success of any tissue graft. Collagen is one of the main components of the extracellular matrix, and there has been much interest in new sources for application as a biomaterial. In this work, a tissue engineering scaffold was developed using the electrospinning technique. The chicken skin was used as an alternative source to obtain collagen. The combination of this collagen with elastin was successfully electrospun, and a distribution of diameters was obtained, less than 100 nm. In vitro tests showed the adhesion and proliferation of the cells, as well as an absence of cytotoxicity from non–cross-linked scaffolds and scaffolds that were cross-linked with carbonyldiimidazole. The structure and composition of the developed scaffolding provide a favorable environment for cell growth and generating a skin substitute.
Decellularized extracellular matrices (dECMs) have demonstrated excellent utility as bioscaffolds in recapitulating the complex biochemical microenvironment, however, their use as bioinks in 3D bioprinting to generate functional biomimetic tissues has been limited by their printability and lack of tunable physical properties. Here, we describe a method to produce photocrosslinkable tissue-specific dECM bioinks for fabricating patient-specific tissues with high control over complex microarchitecture and mechanical properties using a digital light processing (DLP)-based scanningless and continuous 3D bioprinter. We demonstrated that tissue-matched dECM bioinks provided a conducive environment for maintaining high viability and maturation of human induced pluripotent stem cell (hiPSC)-derived cardiomyocytes and hepatocytes. Microscale patterning also guided spontaneous cellular reorganization into predesigned striated heart and lobular liver structures through biophysical cues. Our methodology enables a light-based approach to rapidly bioprint dECM bioinks with accurate tissue-scale design to engineer physiologically-relevant functional human tissues for applications in biology, regenerative medicine, and diagnostics.
Using decellularized extracellular matrix (dECM) hydrogels as bioinks has been an important step forward for bioprinting of functional tissue constructs, considering their rich microenvironment and their high degree of biomimicry. However, directly using dECM hydrogels as bioinks may not be suitable for bioprinting processes because of the loss of shape fidelity and geometrical precision of bioprinted structure due to their slow gelation kinetics. In this article, the development and direct bioprinting of dECM hydrogel bioink from bovine Achilles tendon were presented. The developed bioink is used for a microcapillary‐based bioprinting process without any support structure and/or any additional cross‐linker components. The reported decellularization and solubilization methods yield dECM pre‐gels which turn into stable hydrogels in a short time at physiological conditions. The gelation kinetics and mechanical strength of bioinks with different concentrations and digestion times are characterized. A support structure‐free 3D bioprinting of the developed bioink is shown by aspirating dECM bioinks and then in situ gelation and extrusion through a fine microcapillary nozzle. The viability assays indicate that the developed dECM bioink has no cytotoxic effect on encapsulated NIH 3T3 cells and the cells show lineage‐specific morphology in the early days of culture as well.
Slaughterhouse waste from chicken processing industries includes skin, tendon, cartilage, bones and other connective tissues that are the source materials for collagen extraction. Burning, incineration, composting and using for animal feed supplements are the different traditional applications of those waste. Waste utilization is a great challenge for the growth of chicken and egg processing industries. By-products from the chicken meat industries have high nutritional value, comprising of high-quality proteins and peptides. The utilization of these by-products for the extraction of proteins and peptides is more beneficial than discarding them as waste. However, the eggshells, mainly the eggshell membrane, comprise of organic matter such as proteins, peptides and amino acids that can be utilized for the creation of value-added products from waste material. Suitable environmentally-friendly methods are needed for waste disposal and utilization. Biotechnological methods, such as enzymatic hydrolysis and fermentation, are found to be better than chemical ones.
The objective of this research is to evaluate alternative source for collagen. The common source of collagen is from bovine and pig. There is need to find an alternative source of collagen due to its demand. Broiler Chicken feet waste is one of the alternative sources because of its abundant collagen protein content. On an average each chicken shop generates minimum of 30-40 kg waste every day in Hyderabad. Collagen is a fibrillar protein composing different forms of the connective tissue: bone, cartilage, tendon and skin. Collagen is the major component of the extracellular matrix, and more than 27 genetic isoforms have been identified. Collagens type I, II and III are the most abundant and well investigated for biomedical applications. Chicken feet, rather than being transformed into meal for animal feed, a large quantity of chicken feet could be used to produce collagen, which is valued for its unique functional properties. The purpose of this research project was to extract and characterize collagen from chicken feet. Biochemical composition of chicken feet collagen such as moisture, protein, fat and ash content was 5.85, 29.11, 35.43 and 28.60%, respectively.Acid-solubilized collagen (ASC) was isolated from the chicken feet. Molecular weight of ASC was analysed and confirmed by SDSPAGE. Secondary structure of ASC was confirmed by FT-IR spectrum.Chicken feet thus appear to be a good alternative source of high-quality collagen.
Many connective tissue diseases and defects are associated with poor synthesis or excessive degradation of collagen. The modern tissue engineering approach is to replace the defective site via the implantation of a biocompatible scaffold which serves as a carrier for cell incorporation, proliferation, and growth. Collagen is widely used in the field of clinical medicine in connection with both hard and soft tissue applications. However, certain collagen properties such as poor dimensional stability, poor in vivo mechanical strength, low degree of elasticity, variable nature in terms of enzymatic degradation, crosslinking density, fiber size, trace impurities, and side effects frequently limit both its analysis and application. This review focuses particularly on the processing and modification of collagen type I with respect to its biological and mechanical properties. The processing of collagen into scaffolds is crucial to mimic successfully the extracellular matrices. Moreover, the review suggests several ways in which the most common problems related to the isolation, handling, electrospinning, and crosslinking of collagen can be overcome while maintaining its native character as much as possible. Further, the review provides a summary of the analytical methods available for the physicochemical characterization of collagen with respect to both its molecular and submolecular structure.
This paper reviews the structure, function and applications of collagens in food industry. Collagen is the most abundant protein in animal origin. It helps maintaining the structure of various tissues and organs. It is a modern foodstuff and widely used in food and beverage industries to improve the elasticity, consistency and stability of products. Furthermore, it also enhances the quality, nutritional and health value of the products. Collagen has been applied as protein dietary supplements, carriers, food additive, edible film and coatings. Therefore, this paper will review the functions and applications of collagen in the food and beverage industries. The structure and composition of collagen are also included.
Fish gelatin is an important alternative gelatin which can be considered as Halal free and acceptable by all religions. It is made from fish by-products of which fish skin is the most widely used part. The collogan and gelatin-like property of fish bones and scales coupled with their readily availability make it a potential source for development into gelatin products. This review discusses the potentials for the development and utilization of fish bones and scales in the production of gelatins. It also looks at the raw materials, processes, properties and the improvement of fish gelatins for future commercial use.
Type I collagen was prepared from tilapia (Oreochromis niloticus) skin by acetic acid and pepsin process at 4 °C, respectively (ASC and PSC), and hot-water method separately at 25, 35 and 45 °C (C-25, C-35 and C-45). Their structure and self-assembly property were discussed. SDS-PAGE patterns suggested that pepsin hydrolysis and the 35 and 45 °C extraction produced collagen with much reduced proportions of α- and β-chains. Fourier transform infrared spectroscopy spectra revealed that pepsin hydrolysis did not change the conformation of collagen, but higher extraction temperature did. Self-assembly curves and atomic force microscopy (AFM) observations showed that only ASC, PSC and C-25 could self-assemble into fibrils with D-periodicity, but the reconstruction rate of C-25 was lower. Besides, PSC had relatively higher resolution ratio compared with others. Overall, pepsin-extracted collagen displayed higher solubility and better fibril-forming capacity, having the potential of applying in biomaterials and food-packaging materials.
Acid-soluble collagen (ASC) and pepsin-soluble collagen (PSC) were extracted from the muscles of selected cultured catfish (hybrid of Clarias gariepinus x Clarias macrocephalus), red tilapia (Oreochromis niloticus), black tilapia (Oreochromis mossambicus), pangasius catfish (Pangasius sutchi), sultan fish (Leptobarbus hoevenii) and labyrinth fish (Trichogaster trichopterus), freshwater fishes that are widely consumed in Malaysia. The extracted yields for the tested species were higher for PSC as compared with ASC. The catfish contained the highest amount of collagen (PSC) at 368.36 ± 12.05 mg collagen/g muscles. A mathematical description of the principle kinetics involved in collagen extraction was attained using four two-parametric models, namely the power law, parabolic diffusion, Peleg's and Elovich's models. The power law model was found to be the best model capable of predicting the extraction data with R2 = 0.993, P value = 0.033% and root mean square deviation = 2.605%, respectively.
Utilization of fish collagen has been favorable over that of mammalian sources, mainly in making more effective use of underutilized resources. The application of acid-extraction technology to recover and isolate collagen from Malaysian freshwater fish muscles would be able to boost up the commercial values of these cheap and underutilized protein sources while assisting in conservation of these natural resources in Malaysia. Mathematic models proposed in this study were also proven to be successful in governing and describing the kinetics of collagen extraction process.
The present study focused on the recovery of collagen and its hydrolysate from a homogenate obtained by enzymatic hydrolysis of bovine hide. The influence of Bat enzyme, protease and pepsin on the hydrolysis of hide material is investigated. Aqueous two-phase systems (ATPS) having a composition of 14% polyethylene glycol (PEG) 600 and 15% potassium phosphate was loaded with 20% w/w of collagen homogenate. Such loaded systems yielded a two phase system, a top phase (97% recovery yield of collagen) and a bottom phase (94% recovery yield of pepsin enzyme). The study indicated that it is possible to selectively obtain collagen and its hydrolysed product in one phase and the enzyme used for the hydrolysis in another phase in ATPS partition.
The effects of concentration, pH value and ionic strength on the kinetic self-assembly of acid-soluble collagen from walleye pollock (Theragra chalcogramma) skin were investigated. A two-phase kinetic process was provided which included the formation of nucleus center and nucleus growth, the first phase being the controlled step for collagen self-assembly. Collagen showed marked assembly behavior when concentration reaching and above 0.6 mg/mL, and higher concentration could accelerate collagen self-assembly. Rate constants of the first and second assembly phase both increased with pH to a maximum around pH 7.2 and then decreased, indicating that pH 7.2 was the optimum pH value for collagen self-assembly. The kinetics of collagen self-assembly could be modulated by NaCl concentration. The concentration of NaCl from 30 to 60 mM was more suitable to self-assemble for pollock skin collagen.
Soluble collagen from fish (sardine) scales was yielded at about 5% with 0.5 M acetic acid after demineralization with EDTA, while a great portion of the collagen remained insoluble. The solubility of this insoluble collagen was about 20% at 45 degrees C (denaturation temperature of soluble collagen) for 24 h. The remaining 80% of the insoluble collagen was denatured in the form of insoluble gelatin, and that may be an interesting food material.
The objective of this research was to evaluate alternative treatments for the best extraction condition for collagen from chicken feet. Various properties such as chemical composition, amino acid, pH, swelling percentage, yield and pure collagen, collagen loss, color (Hunter L, a and b) and electrophoresis of collagen from chicken feet treated by 5% acids (acetic acid, citric acid, hydrochloric acid and lactic acid) and soaking times (12, 24, 36 and 48 h) were evaluated. The crude protein, fat, ash and moisture contents of chicken feet was 17.42, 12.04, 5.98 and 62.05%, respectively. Amino acid composition of collagen from chicken feet indicated that the protein of collagen was markedly hydrolized by the hydrochloric acid treatment. The result of electrophoresis also supported this phenomenon. Both the swelling percentage of lactic acid and citric acid treatments were significantly higher than that of acetic acid and HCl treatment. The pH of the acid treatments ranged from 2.43-3.62. According to the result of yield, pure collagen and loss of collagen, the best condition of extracting collagen from chicken feet was soaked in 5% lactic acid for 36 h. However, a brighter yellow color of collagen from all treatments was observed with a longer soaking time. (Asian-Aust. J. Anim. Sci. 2001. Vol 14, No. 11 : 1638-1644).
As a part of the study into the potential development of unused and under-used resources, collagen was isolated from the mesogloea of the rhizostomous jellyfish, Rhopilema asamushi, by limited pepsin digestion and characterized. The yield of this collagen was high (35.2% on a dry weight basis). The primary structure was very similar to that of pepsin-solubilized collagen from edible jellyfish mesogloea, but it was different from those of the collagen from edible jellyfish exumbrella and the acid-soluble collagen from its mesogloea. The denaturation temperature (Td) of 28.8°C. This collagen contained a large amount of a fourth subunit that was provisionally designated α4. This collagen may have the chain composition of an α1α2α3α4 heterotetramer.
The mesogloea and skin of a common edible jellyfish, Stomolophus nomurai, were characterized with respect to amino acid composition and compared with a commercially salted jellyfish. Then the mesogloea was digested with pepsin at 3°C for 48 hr and its solubilized protein was isolated and subjected to biochemical analyses. These composite results have shown that the major edible component of jellyfish was the connective tissue protein, collagen, characterized by its high content of hydroxylysine and its glycosides.
1. Collagen was extracted from chick skin with dilute acetic acid followed by dilute acetic acid containing pepsin. 2. The solubilized collagens were purified and portions subjected to further digestion by pepsin. 3. This treatment decreased the aldehyde content but contamination by hexosamine was not diminished. 4. Pepsin treatment converted practically all the acid-soluble collagen into monomeric subunits (alpha-chains), but the pepsinsolubilized material retained a significant amount of higher subunits (beta- and gamma-chains). 5. Treatment lowered the rate of fibrillogenesis by acid-soluble collagen, but was without effect on pepsin-solubilized collagen.
Using an improved method of gel electrophoresis, many hitherto unknown
proteins have been found in bacteriophage T4 and some of these have been
identified with specific gene products. Four major components of the
head are cleaved during the process of assembly, apparently after the
precursor proteins have assembled into some large intermediate
structure.
This chapter presents methods generally applicable to the preparation of native collagen from a variety of sources. The preparation of a collagen sample generally involves several different steps. These include acquisition and preliminary processing of an appropriate tissue or organ, extraction of the collagen, and its purification. The latter process requires not only elimination of the noncollagenous components that are present in the extract, but may also require selective removal of alternative collagen types. This can usually be accomplished by a judicious use of selective precipitation techniques accompanied ultimately by one or two chromatographic steps. Given the extreme diversity of the tissues or organs in which collagen occurs as well as the multiplicity of collagen types that may be present in a given source, there understandably exists no single standard or preferred method for the preparation of collagen. Unless otherwise indicated, all procedures are conducted at relatively low temperatures, in the range of 4-8 °. This minimizes bacterial growth, enhances the solubility of native collagens, and ensures the retention of native conformation on the part of the solubilized collagens.
In this paper, a chromatographic method for the purification of native types I, II, and III collagen is described. The method consists of two consecutive gel permeation chromatography steps, followed by anion-exchange chromatography. The two consecutive gel permeation chromatography steps take advantage of the fact that collagens like other asymmetric molecules, elute anomalously late from gel permeation columns, thus allowing one to separate collagens from less asymmetric proteins of comparable molecular weight, notably gelatin, procollagen and higher molecular weight oligomers of collagen. The anion-exchange chromatography separates types I, II, and III collagens from each other with baseline resolution. The collagen products obtained from these procedures are at least 99% pure by a variety of criteria, and in the native state by the traditional criteria of optical rotation, intrinsic viscosity, solubility properties and resistance to non-collagenase proteases. Rat skin type I collagen prepared by this chromatographic method exhibits a higher and sharper thermal transition temperature than an otherwise identical sample of rat skin type I collagen prepared by fractional salt precipitation. In addition, the latter collagen is more susceptible to digestion by trypsin at 37 degrees C. We conclude that salt precipitation of the collagen per se is responsible for a lowering of the Tm values. Our observations indicate that the chromatographic purification of collagen preserves the native structure at a few select sites where high salt concentrations induce irreversible local imperfections of the three-dimensional structure.
A unique collagen with three distinct chains, was purified from the cranial cartilage of the squid Sepia officinalis, by pepsinisation and salt precipitation and compared with shark cartilage collagen. These chains, which were different from the known cartilage collagen chains, were referred as C1, C2 and C3, had approximate molecular weights of 105 kDa, 115 kDa and 130 kDa, respectively, and were present in a ratio of 3:2:1, suggestive of two molecules of composition, [(C1)2C2] and [C1C2C3]. These collagens were purified by fractionation at acid and neutral pH, and by ammonium sulfate precipitation. Solubility data indicated that this collagen was more crosslinked than the type I collagen isolated from cartilage of shark, Carcharius acutus. In vitro fibrillogenesis revealed that the sepia collagen formed denser aggregates, as compared to shark collagen, and was stabilised by a higher degree of carbohydrate association. Polyclonal antisera raised against shark collagen was also reactive against the sepia collagens, while the converse was not true, indicating the high immunospecificity of the latter. These results demonstrate collagen polymorphism in an invertebrate cartilage and may hold significance in understanding tissue calcification and molecular evolution. Further, these collagens may represent ancestral forms of vertebrate minor collagens like typeV/XI.
The current paper pertains to the study of frog skin, more specifically Rana tigerina skin collagen, which is a major extracellular matrix protein known to play an important role in the wound-healing process. This study revealed interesting differences in the frog skin collagen when compared to the hitherto known vertebrate collagens. This could probably be attributed to the position of the amphibians in the vertebrate hierarchy. Therefore, detailed investigations on the various physico-chemical properties, such as reconstitution, redissolution, viscosity and denaturation were carried out. The study confirms the structural relationship of collagen to habitat and function.
Collagen is regarded as one of the most useful biomaterials. The excellent biocompatibility and safety due to its biological characteristics, such as biodegradability and weak antigenecity, made collagen the primary resource in medical applications. The main applications of collagen as drug delivery systems are collagen shields in ophthalmology, sponges for burns/wounds, mini-pellets and tablets for protein delivery, gel formulation in combination with liposomes for sustained drug delivery, as controlling material for transdermal delivery, and nanoparticles for gene delivery and basic matrices for cell culture systems. It was also used for tissue engineering including skin replacement, bone substitutes, and artificial blood vessels and valves. This article reviews biomedical applications of collagen including the collagen film, which we have developed as a matrix system for evaluation of tissue calcification and for the embedding of a single cell suspension for tumorigenic study. The advantages and disadvantages of each system are also discussed.
Collagen-based scaffolds are appealing products for the repair of cartilage defects using tissue engineering strategies. The present study investigated the species-related differences of collagen scaffolds with and without 1-ethyl-3-(3-dimethyl aminopropyl) carbodiimide (EDC)/N-hydroxysuccinimide (NHS)-crosslinking. Resistance against collagenase digestion, swelling ratio, amino acid sequence, shrinkage temperature, ultrastructural matrix morphology, crosslinking density and stress-strain characteristics were determined to evaluate the physico-chemical properties of equine- and bovine-collagen-based scaffolds. Three-factor ANOVA analysis revealed a highly significant effect of collagen type (p=0.0001), crosslinking (p=0.0001) and time (p=0.0001) on degradation of the collagen samples by collagenase treatment. Crosslinked equine collagen samples showed a significantly reduced swelling ratio compared to bovine collagen samples (p< 0.0001). The amino acid composition of equine collagen revealed a higher amount of hydroxylysine and lysine. Shrinkage temperatures of non-crosslinked samples showed a significant difference between equine (60 degrees C) and bovine collagen (57 degrees C). Three-factor ANOVA analysis revealed a highly significant effect of collagen type (p=0.0001), crosslinking (p=0.0001) and matrix condition (p=0.0001) on rupture strength measured by stress-strain analysis. The ultrastructure, the crosslinking density and the strain at rupture between collagen matrices of both species showed no significant differences. For tissue engineering purposes, the higher enzymatic stability, the higher form stability, as well as the lower risk of transmissible disease make the case for considering equine-based collagen. This study also indicates that results obtained for scaffolds based on a certain collagen species may not be transferable to scaffolds based on another, because of the differing physico-chemical properties.
Preparation of telopeptide-poor collagen from pig skin
- A B Wu
- H W Cheng
- S J Chen
- L H Lin
- H K Lu
- C W Lin
Wu, A. B., Cheng, H. W., Chen, S. J., Lin, L. H., Lu, H. K., Lin, C. W., et al. (1999). Preparation of telopeptide-poor collagen from pig skin. Journal of Agricultural Association of China, 187, 93–100.
Preparation and character-ization of different types collagen
- E J Moller
- R K Rhode
Moller, E. J., & Rhode, R. K. (1982). Preparation and character-ization of different types collagen. Method Enzymology, 82, 33–64.
Preparation of telopeptide-poor collagen from pig skin
- Wu
Pepsin treatment of avian skin collagen
- Bannister