Avian-keratin refinement and application in biomaterials

Pourjavaheri, F 2017, Avian-keratin refinement and application in biomaterials, Doctor of Philosophy (PhD), Science, RMIT University.

Document type: Thesis
Collection: Theses

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Title Avian-keratin refinement and application in biomaterials
Author(s) Pourjavaheri, F
Year 2017
Abstract The aim of this research project was to curb the environmental impact of chicken feathers, a waste from the poultry industry, by value-adding and development of bio-composites with improved biodegradability and thermo-mechanical properties, and to extract keratin from the feathers for inclusion in biomaterials for potential consumer applications. The first step in the application of chicken feathers involved thorough cleaning and disinfection since plucked chicken feathers impose severe microbiological hazards. Therefore, the design of a proper purification method in respect to the final application was necessary. Different surfactants including anionic, non-ionic, and cationic; bleach such as ozone and chlorine dioxide; ethanol extraction; and a combined method comprising surfactant–bleach–ethanol extraction were applied to chicken feathers and their bactericidal performance was investigated via a) Standard Plate Count, b) the enumeration of Escherichia coli, Pseudomonas species, coagulase positive Staphylococcus, aerobic and anaerobic spore-formers and c) Salmonella and Campylobacter detection tests. Among all practices, only the ethanol extraction and combined method eliminated Salmonella from the feathers. Although ethanol-extraction showed superior bactericidal decontamination compared with the combined method, the feathers purified with the latter method showed better morphological and mechanical properties. Scanning electron microscopy-energy dispersive spectroscopy confirmed the presence of sodium lauryl sulphate remnants in the feathers after applying the combined method. Fourier-transform infrared spectroscopy was adopted for the qualitative characterisation of the feathers before and after purification. Chicken feather characterisation including a-helix conformation in the feather wool, and pleated sheet in barbs and rachis, are presented herein. The pH, visual observation, optical microscopy under visible and ultraviolet lights, scanning electron microscopy, micro X-ray diffraction, wide-angle X-ray scattering, infrared spectroscopy, vibrational spectroscopy and thermogravimetry were used to characterise the feathers before and after purification and residues after extraction. The next consideration was to find a use for waste feathers. Two polyurethane based polymers were combined with chicken feather fibres, to form bio-composites. Thermoplastic polyether–polyurethane was used via solvent–casting–evaporation–compression moulding method at 10, 20, 30, 40, 50, 60 and 70 %·w/w of chicken feather fibres; and thermoplastic polysiloxane–polyurethane was used via solvent–casting–evaporation–compression moulding, and solvent–precipitation–evaporation–compression moulding methods to create new bio-composites incorporating 10 and 20 %·w/w of chicken feather fibres into the polyurethane. Compatibility of polyurethanes with the feather fibres and the thermo-mechanical properties of the resulting bio-composites were determined and using thermogravimetry, dynamic mechanical analysis and stress–strain measurements with hysteresis loops. The uniformity of the feather fibres dispersion in the polyurethane matrix was investigated via macro-photography. Scanning electron microscopy of fractured surfaces of the bio-composites was used to verify that the adhesion between fibre and polymer was effective. Molecular modelling visualisation predicted the existence of hydrogen bonding between fibres and polyurethane molecules and this result was supported by Fourier-transform infrared analysis of the composite. The addition of chicken feather fibres to the polyurethane matrixes was found to decrease the glass transition temperature, recovery strain and thermal mass loss of the composites, but increase the elastic modulus (hardness), storage modulus and char level on thermal decomposition. The thermo-mechanical properties of these polymers were enhanced by addition of keratin feather fibres. The utilization of ecofriendly, bio-based composites has been reported in many areas including, but not limited to, the packaging, insulation, automotive, building and roofing industries, as well as for separation membranes for water treatment. The applications of the produced bio-composites are steps towards more environmentally-friendly and more cost effective products. Keratin was then extracted from different segments of disposable chicken feathers including whole feathers, calamus and rachis (composed mainly of beta-pleated sheet structures), barbs and barbules (composed mainly of alpha-helix), using sodium sulfide or L‑cysteine. The extraction process involved dissolving the chicken feathers by reducing its disulfide links, then separating the protein from the medium by centrifugation. Once the feathers were dissolved, the pH of solution was adjusted to the isoelectric point using hydrochloric acid, to precipitate the proteins, and the yield of extracted keratin with sodium sulfide (88 ± 3 %) was higher than with L-cysteine (66 ± 4 %). The precipitated keratin was washed three times with distilled water. The presence of protein obtained from different methods was confirmed using the biuret test, and the Bradford assay enabled the concentration of keratin to be determined. The precipitated keratin was characterised using gel electrophoresis, which confirmed soluble protein of molar mass 11 kg/mol and estimated its purity to be over 95 %. Liquid chromatography-mass spectrometry verified the molar mass of the extracted material matched that of chicken keratin. Vibrational and nuclear magnetic resonance spectroscopy confirmed the structure of keratin was retained following extraction. Thermogravimetry of original purified chicken feather and keratin extracted via sodium sulfide treatment showed virtually identical decomposition behaviour, proving the purity of the keratin. In contrast, thermogravimetry of keratin extracted with L-cysteine indicated it may contain residual L-cysteine. The structure of keratins extracted from different segments of waste chicken feathers via sodium sulfide and L‑cysteine, have been subjected to further nuclear magnetic resonance spectroscopy and analysed for their antibacterial properties on Staphylococcus aureus and Escherichia coli as Gram-positive and Gram-negative species, respectively. The goal of this section was to produce an extract and to characterise several aspects of its behaviour that may have implications for its use as a biomaterial. Hence, the keratin extracted using sodium sulfide was incorporated into hair conditioner and cream, and used in hair and leather treatments to determine their interactions with animal tissues. These experiments confirmed and expanded earlier findings that keratin demonstrated excellent compatibility in biological systems, as the highest keratin concentration experimental cream and conditioner, had the best outcomes. Finally, this study presents suggestions for future fundamental studies and proposals for the development of keratin-based materials for biomedical and consumer product applications.
Degree Doctor of Philosophy (PhD)
Institution RMIT University
School, Department or Centre Science
Subjects Theory and Design of Materials
Keyword(s) Keratin
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Created: Fri, 17 Nov 2017, 09:54:26 EST by Adam Rivett
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