Chicken feather: Chicken feather was obtained from the animal processing unit at Landmark University Commercial Farm, Omu-Aran, Kwara State, Nigeria.
Ginger: Ginger roots was purchased from Omu-Aran market and transported fresh in polythene bag to Biochemistry Laboratory.
Chicken feather and keratin extraction: Chicken feather was obtained from the animal processing unit at Landmark University Commercial Farms. The white coloured feather wastes were free of blood and grease by washing severally with detergent and then rinsed several times with tap water to remove every trace of the detergent. The washed feathers were spread on a clean dry white board to remove any extraneous tissues such as skin, intestine, beak, etc. Thereafter, the feathers were disinfected by soaking in 0.5% hypochlorite solution, sodium thiosulphate was added to neutralize the hypochlorite before being rinsed severally with distilled water before being dried to constant weight at 60 oC in a forced-air oven. The dried feathers were cut into small pieces and milled into powder, using a mechanical grinder. One thousand gram (1000 g) of the powdered feather was soaked in acetone (Sigma-Aldrich, UK) overnight, filtered and allowed to air-dry at 27 oC. Keratin was extracted from the powdered feather following a combine methods of Akpor et al.  and Tesfaye et al. . Fifty gram (50 g) feather powder was dissolved in 150 mL of 0.1 M sodium hydroxide solution, mixed thoroughly and then placed in an orbital shaker at 27 oC for 8 h. The resulting solution was filtered through a muslin cloth. The pH of the dissolved feather solution was adjusted to neutral, using 10% trichloroacetic acid and the precipitated keratin filtered through a muslin cloth, washed severally in distilled water and then freeze-dried. The dried keratin was pulverized using a mechanical grinder, sieved through a 50 µm sieve and then stored at 4 oC until required for further analysis.
Starch was extracted from ginger rhizome according to the method of Afolayan et al. . Freshly purchased ginger roots were neatly peeled, washed with copious water, sliced into small pieces and soaked in 1% sodium metabisulphite solution to prevent oxidative browning. Afterward, the chopped ginger roots were milled into a slurry using an electric blender. The starch slurry was then dispersed in a large volume of 1% sodium metabisulphite solution before been filtered using muslin cloth. The obtained filtrate was centrifuged for 15 min at 3000 rpm after which the supernatant was carefully decanted and the mucilage scraped off. The process was repeated trice until the starch was obtained in a pure form. The extracted starch was oven-dried to constant weight at 30 oC, weighed and then stored at 4 oC, until required for further analysis.
Preparation of the keratin-starch bio-composite film
In the preparation of the keratin-starch bio-composite film, the method described by Tesfaye et al.  was followed with little modifications. Briefly, a 5% starch solution was prepared and gelatinized by heating on an electric heater at 70 oC under constant stirring. Keratin solution was made by adding 5 g keratin powder in 100 mL of 0.1 M NaOH and heated in an electric heater at 70°C for 20 minutes with constant stirring. Different blends of starch and keratin solutions were then prepared as follow: 20:0, 20:1, 20:3 and 20:5 (starch:keratin v/v). To each starch-keratin blend 1 mL of glycerol was added as plasticizer, after which the mixture was heated at 70 oC on an electric heater for 10 min, with vigorous stirring before being poured into a glass petri dish and oven-dried at 50 oC for 24 h. The resulting film was carefully peeled and kept in a cool dried carton at 25 oC before further analysis.
Physicochemical characterization of ginger starch
Swelling power determination
The swelling power of the starch sample was determined using the method of Afolayan et al. . Ginger starch (0.1 g) was added to 10 ml of distilled water and heated at 50 oC for 30 min in a water bath with continuous shaking. The resulting slurry was centrifuged at 2000 rpm for 15 min after which the supernatant was carefully decanted and the resulting starch paste weighed. Thereafter, the swelling power was estimated using the formula:
The swelling power of the starch sample was estimated over a temperature range of 50 – 100 oC.
Solubility index determination
The swelling power of the starch sample was determined using the method of Afolayan et al. . A starch solution comprising of 0.5 g dry starch sample dissolved in 10 mL distilled water and heated at 50 oC over a water bath for 30 min with vigorous shaking was prepared. The starch solution was then centrifuged at 2000 rpm for 10 min and the supernatant carefully decanted. The resulting starch paste was weighed and the solubility index was estimated, using the formula:
The solubility index of the starch sample was evaluated over a temperature range of 50 – 100 oC.
The pH was determined using a British Standard Institution (BSI 757) as described by Taheri et al. . A10% (w/v) starch dispersion was prepared with continuous shaking in a water bath at 50 oC for 5 minutes. Having allowed the starch gel to cool to about 25 oC, the pH determined using a pH meter with a glass electrode after standardizing with 4 and 7 pH buffers.
Gelatinization temperature determination
The gelatinization temperature was determined following the method described by Nwokocha et al. . One gram (1 g) starch sample was dispersed in 10 mL distilled water and stirred vigorously. The starch solution was placed on a hot plate and heated with continuous stirring. With the aid of a suspended thermometer, the gelatinization temperature was then estimated.
Physicochemical characterization of the keratin-starch bio-composite film
The bio-composite film thickness was estimated with the aid of a digital MDC-MX 923 micrometer (Mitutoyo, Japan). For each bio-composite film, 10 random points were sampled and measurements were taken and the mean values calculated.
The degree of transparency or opaqueness of the bio-composite film with the aid of a UV-visible spectrophotometer (Jenway 7305, UK) at 560 nm. The measurement was done following Santacruz et al.  method.
Moisture content determination
The moisture content of the bio-composite fil was determined by drying the bio-composite film to constant weight in a forced air oven at 100 °C. The moisture content was then calculated using the equation:
Where W1, Initial weight of the biocomposte film, W2, weight of the bio-composite film after drying.
Tensile strength analysis
The tensile strength of the bio-composite film was determined according to the method of Rahman and Jamalulail , with the aid of a Universal Testing Machine (Hounsfield Series S, UK) according to ASTM standard D882. In measuring the tensile strength, a 1.0 kN load was applied at 10 mm/mim constant at 27 oC. From the data obtained, tensile strength and elongation at break were calculated.
The solubility of the keratin-starch bio-composite film in water was determined according to the method of Pavin et al. . A weighed amount of the keratin-starch bio-composite films was completely immersed in water for a period of 12 da and at every three-day interval, the bio-composite film was exhumed and weighed.
The degradability test of the keratin-starch bio-composite films in soil was studied, following the method of Pavin et al. . In this study, a weighed amount of the keratin-starch bio-composite films was vertically buried at 3 – 5 cm depth inside the soil to ensure aerobic degradation conditions at 25 ± 2 oC and 35 – 50% relative humidity for a period of 12 days and at every three-day interval, the bio-composite film was exhumed, washed with water, dried to constant weight and weighed.
Morphological analysis of the bio-composite films
The morphological analysis of keratin, ginger starch, and keratin-starch bio-composite film was carried out with a field emission gun scanning electron microscope (SEM) (Zeiss, Germany). The sample was placed on a gold coated stub with the aid of a diode sputter coater. The crystalline phase evaluation was carried out with the aid of a benchtop XRD analyzer (ARL EQUINOX 1000, Thermo Fischer, UK), equipped with a diffracted beam monochromator and a copper target X-ray tube operated at 40 kV and 30 mA.
The experimental set up was carried out in triplicate and the results expressed as mean ± SD. The results were statistically analyzed using two-way analysis of variance (ANOVA) followed by Turkey multiple range test. Charts were drawn using GraphPad prism 8.0 software (GraphPad Software Inc., San Diego, California). Significant level was set at P < 0.05.