A two level full factorial is performed with a model equation designed such that the variance of Y is constant for all points equidistant from the centre of the design. Minitab (14.0) statistical software package was used in the experimental design and data analysis. Response surface graphs were obtained to know the effect of the variables, individually and in combination, and to determine their optimum levels for maximum melanin production. All trials were performed in duplicate, and the average melanin yield was used as response Y. As per the adapted method, spectra of learn more melanin samples were collected
over the spectral range 400–280 nm with 1 nm data point resolution on a UV-visible UV-3200 double beam spectrophotometer (LABINDIA analytical Instruments Pvt Ltd India). The SPF values of melanin from the purchased strain and microbial isolate were determined using Mansur STA-9090 purchase mathematical Eq. (1). equation(1) SPF=CF×∑290320EE(λ)×I(λ)×Abs(λ)where, SPF=Sunscreen
protection factor; EE(λ) = erythremal effect spectrum, I(λ) = solar intensity spectrum; Abs(λ) = absorbance of sunscreen product; CF = correction factor (36.2 for SNOW LOTUS® SPF 15 and 30) taken from Huang et al. [20] The radical scavenging activity by melanin pigment was investigated by the modified method of Ju et al. [21]. Primarily, 1 mg/mL of microbial melanin/standard (Ascorbic acid) in different dilutions was added to 2 mL of DPPH in ethanol; so that the total strength of the melanin used was varied from 15–100 μg/mL in each solution. After keeping
for 30 min at 37 °C the absorbance at 516 nm was measured using UV‐spectrophotometer with reference blank samples. The experiment was performed in duplicate. The absorbance of DPPH as control was measured at 516 nm. The lower absorbance of the reaction mixture indicated higher radical scavenging activity. The scavenging effect was measured using Eq. (2). equation(2) DPPHinhibition(%)=[(Controlabsorbance−testabsorbance)]×100DPPHinhibition(%)=[(Controlabsorbance−testabsorbance)]×100 Branched chain aminotransferase The chelation of ferrous ions by the melanin pigment was estimated by the method of Huang et al. [20]. Different concentrations of melanin were mixed with a solution of 2 mM FeCl2 (0.05 mL). The reaction was initiated by the addition of 5 mM ferrozine (0.2 mL) and the mixture was shaken vigorously and left standing at room temperature for 10 min. Absorbance of the solution was then measured spectrophotometrically at 562 nm. All the tests and analysis were performed in duplicate and averaged. The inhibition of metal chelating activity by the melanin pigment in percentage (%) was calculated using Eq. (3) equation(3) Metalchelatingeffect(%)=[A0−A1A0]×100%where A0 is the absorbance of control reaction and A1 is the absorbance in the presence of the sample of the melanin pigment or standards. The control contains FeCl2 and ferrozine. XRD analysis was performed using the CuKα radiation (λ = 1.
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