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20191
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Subject: Processing and Food Engineering × Author: Gaibimei, Palmei × Thesis Type: Ph.D ×
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    ThesisItemOpen Access
    Process optimization for anthocyanin extraction from black soyabean (Glycine max (L.) Merr.) using different methods, comparative study and its characterization
    (G.B. Pant University of Agriculture and Technology, Pantnagar - 263145 (Uttarakhand), 2019-07) Gaibimei, Palmei; Singh, Anupama
    Black soyabean hulls are considered as one of the agro-industrial residues which contain abundant anthocyanin in its seed coats. An effective, environment friendly and energy-efficient extraction technique need to be explored to improve the quantity and quality of anthocyanin from black soyabean hulls. In the present investigation four extraction techniques were employed to extract anthocyanin from black soyabean hulls. Design experiments using BBD for aqueous extraction and enzyme assisted extraction while CCD for microwave assisted extraction and ultrasound assisted extraction were employed with Design Expert 10.0.1 software considering independent parameters having varied levels of each. Independent parameters for AE were chosen as solid to solvent ratio, extraction temperature and treatment period having three levels (1:20, 1:40, 1:60 g/ml; 40, 60, 80 °; 40, 60 and 80 min)of each, for EAE, independent parameters chosen were enzyme concentration, extraction temperature and treatment period having three levels (2, 4, 6 %; 35, 40, 45 °; 30, 60 and 90 min) of each. For MAE, independent parameters chosen were solid to solvent ratio, pH, ethanol concentration and treatment period with five levels (1:20, 1:30, 1:40, 1:50, 1:60 g/ml; 1, 2, 3, 4, 5; 50, 60, 70, 80, 90 %; 30, 60, 90, 120, 150 s), for UAE, independent parameters chosen were solid to solvent ratio, pH, ethanol concentration and treatment period with five levels (1:20, 1:30, 1:40, 1:50, 1:60 g/ml; 1, 2, 3, 4, 5; 50, 60, 70, 80, 90 %; 10, 20, 30, 40, 50 min) of each. The responses selected were Total anthocyanin content, Degradation index, Total phenolic content, DPPH radical scavenging activity, Ferric reducing antioxidant power, Total flavonoids content, Percent polymeric color and Colour difference for entire range of experiments. Comparative study based on the optimized responses values obtained from the different extraction techniques was carried out. Scanning electron microscopy (SEM) analysis for each technique was carried to observe the morphological changes in the hulls powder before and after extraction. Also optimized extracts sample obtained from AE, MAE, UAE and EAE were screened for the identification of various bioactive compounds using GC-MS analysis. Result revealed that the optimized conditions obtained for AE were solid to solvent ratio of 1:20 g/ml, extraction temperature of 40° and treatment period of 65 min with the responses having TAC 30.90 mg/g, DI 1.08, TPC 91.14 mg GAE/g, DPPH 35.41 %, FRAP 295.22 mg AAE/g, TFC 1157.80 mg CE/g, PPC 30.68 % and CD 24.90. While for MAE, the optimized conditions were solid to solvent ratio of 1:22 g/ml, pH of 1.01 and ethanol concentration of 75.20 % and treatment period of 91 sec with the responses having TAC 366.24 mg/g, DI 0.91, TPC 117.01 mg GAE/g, DPPH 100.46 %, FRAP 437.11 mg AAE/g, TFC 2704.82 mg CE/g, PPC 18.30 % and CD 1.86. The optimized conditions for UAE were solid to solvent ratio of 1:26 g/ml, pH of 2.93 and ethanol concentration of 90 % and treatment period of 50 min with the values of responses i.e. TAC 559.95 mg/g, DI 1.27, TPC 170.30 mg GAE/g, DPPH 97.57 %, FRAP 603.33 mg AAE/g, TFC 2536.71 mg CE/g, PPC 97.57 % and CD 25.15. The optimized conditions obtained for EAE were ethanol concentration of 2 %, extraction temperature of 37° and treatment period of 90 min having responses of TAC 1125.38 mg/g, DI 1.45, TPC 119.38 mg GAE/g, DPPH 91.22 %, FRAP 1733.36 mg AAE/g, TFC 4125.32 mg CE/g, PPC 66.11 % and CD 16.87. The micrograph of EAE reveals large, porous, flaky and damaged tough cell wall structure and was found to show biggest pore size opening of 54.468 μm enabling them to leach out more pigment as compared to other extraction techniques. The GC-MS analysis result reveals that the bioactive compounds identified for AE, MAE, UAE and EAE were 22, 43, 50 and 40 respectively. EAE was found to be the best technique as it was able to leach 12 antioxidant compounds, followed by UAE (9 compounds), MAE (8 compounds) and AE (1 compound). Hence the studies showed that all the extraction conditions and techniques selected could effectively extract anthocyanin from black soyabean hulls. Among all the techniques, EAE could be considered as best extraction techniques as it gave outstanding yield of TAC i.e. 97.25 % higher than AE, 67.46 % higher than MAE and 50.24 % higher than UAE method. The cell wall of the black soyabean hulls powder treated by EAE causes large, porous, flaky and damaged tough cell wall structure and was found to show biggest pore size opening of 54.468 μm enabling them to leach out more pigment as compared to other extraction techniques. GC-MS analysis for EAE optimized samples also identified 12 antioxidant compounds in the black soyabean hulls extract. Second order model fitted well and the parameters solid to solvent was most effective for AE, MAE, UAE while for EAE, the parameter enzyme concentration was the most effective for anthocyanin extraction.