Optimization of Soybean Protein Extraction with Ammonium Hydroxide (NH₄OH) Using Response Surface Methodology

Plants have been recognized as renewable and sustainable sources of proteins. However, plant protein extraction is challenged by the plant's recalcitrant cell wall. The conventional extraction methods make use of non-reusable strong alkali chemicals in protein-denaturing extraction conditions. In this study, soy protein was extracted using NH₄OH, a weak, recoverable, and reusable alkali. The extraction conditions were optimized using response surface methodology (RSM). A central composite design (CCD) with four independent variables: temperature (25, 40, 55, 70, and 85 °C); NH₄OH concentration (0.5, 1, and 1.5%); extraction time (6, 12, 18, and 24 h) and solvent ratio (1:5, 1:10, 1:15 and 1:20 w/v) were used to study the response variables (protein yield and amine concentration). Amine concentration indicates the extent of protein hydrolysis. The RSM model equation for the independent and response variables was computed and used to create the contour plots. A predicted yield of 64.89% protein and 0.19 mM amine revealed a multiple R-squared value of 0.83 and 0.78, respectively. The optimum conditions to obtain the maximum protein yield (65.66%) with the least amine concentration (0.14 Mm) were obtained with 0.5% NH₄OH concentration, 12 h extraction time, and a 1:10 (w/v) solvent ratio at 52.5 °C. The findings suggest that NH₄OH is suitable to extract soybean protein with little or no impact on protein denaturation.