Optimization of germination temperature and CaCl2 concentration to improve γ-amino butyric acid content in germination millet

Abstract: Millet is an ancient crop and used as a staple food grain in many semi-arid and tropical areas of the world due to its strong vitality in drought prone areas and very poor soils where other crops fail to grow. Because of its varieties essential nutrients for the human body, and suitable ratio of nutrients, millet has been considered extensively an ideal source of nutrition. γ-amino butyric acid, a non-protein amino acid with four-carbon components, has showed many health benefits. Germination is a biochemical process that can improve effectively the nutrients in whole grains. Especially, germination can increase the content of-amino butyric acid of cereals. The germinated millet is a potential source of functional food. To improve the content of the γ-amino butyric acid in germinated millet, the optimal technological conditions was performed through the single factor experiment, Plackett-Burman experiment, steepest ascent path experiment and response surface methodology. Through the Single factor experiment, the optimum level of each factor was soaking time of 12 h, soaking temperature of 35℃, CaCl2 concentration of 3 mmol/L, germination time of 48 h and germination temperature of 35℃. On the base of the single factor experiment, soaking time, CaCl2 concentration and germination temperature which affected significantly the content of the γ-amino butyric acid were selected through the Plackett-Burman experiment. By use of Minitab software, a first-order model was fitted to the date obtained from the experiment. The effects of the five factors: soaking time (A), soaking temperature (B), CaCl2 concentration (C), germination time (D) and germination temperature (E) were calculated to be GABA = 175.8 - 7.11A - 0.680B +16.85C - 0.365D + 3.894E. At the same time, the first-order linear model of all factors and the content of γ- amino butyric acid were formulated according to the results. Based on the formula and the three important effect factors above, the steepest ascent path experiment was carried out to determine the levels that affected significantly (P<0.05) the content of γ- amino butyric acid at the central point of response surface experiment. It was obtained that the optimal processing for the accumulation of γ-amino butyric acid in germinated millet. The millet was soaked in 3.5 mmol/L CaCl2 solution at 35℃ for 13 h and germinated at 31℃ for 48 h. The content of γ-amino butyric acid was obtained 251.46 mg/100 g in germinated millet under optimized condition, which was 2.90 times higher than the initial γ-amino butyric acid 86.72 mg/100 g. The maximum predicted value 254.19 mg/100 g of GABA content in germinated millet obtained by Response surface methodology was close to the actual value 249.47 mg/100 g, indicating that the formula designed by Response surface methodology was close to the actual situation and the suitability of the formula to be used in optimizing the conditions of germination for the millet. It can be seen that the Single factor experiment, Plackett-Burman experiment, Steepest ascent path experiment and Response surface methodology can effectively screen and optimize the significant factors affecting the GABA content in the germinated millet, and greatly increase the GABA content.