Exogenous abiotic stimulant treatments are a straightforward and effective method for enhancing secondary metabolites in plants. In this study, the response surface optimization method was used to optimize the conditions for enriching flavonoids in short-germinated black soybeans under a slight acid treatment, and the mechanism of flavonoid accumulation during black soybean germination was explored. The results show that the use of a 126.2 mM citric acid–sodium citrate buffer (pH 5.10) as a slight acid treatment resulted in the highest flavonoid content when the black soybeans were germinated for 24 h. Under these conditions, the isoflavonoid (glycitin, daidzein, and genistein) increased significantly, and the flavonoid content reached 2.32 mg/g FW. The microacidified germination treatment significantly increased the activities and relative gene expression levels of key enzymes involved in flavonoid metabolism (4-coumarate-CoA ligase and cinnamic acid 4-hydroxylase, etc.). However, the slight acid treatment inhibited the growth of the black soybeans and caused damage to their cells. This was evidenced by significantly higher levels of malondialdehyde, superoxide anion, and hydrogen peroxide compared to the control group. Furthermore, the antioxidant system in the short-germinated soybeans was activated by the slight acid treatment, leading to a significant increase in the activities and relative gene expression levels of catalase and peroxidase. The results above show that a slight acid treatment was beneficial in inducing the accumulation of flavonoids during the growth of black soybean sprouts. This lays a technical foundation for producing black soybean products that are rich in flavonoids.

Plant hormones play a critical role in the physiological and biochemical mechanisms of plants, with functions such as regulating the metabolic pathways of secondary metabolite production and alleviating external stresses. In this study, the synthesis of flavonoids in black soybean sprouts was induced by slight acid combined with the plant hormones abscisic acid (ABA) and melatonin (MT). The results indicated that the contents of daidzin, genistin, daidzein, and genistein in black soybean sprouts treated with slight acid were increased by 10 μM ABA and 75 μM MT, and the total flavonoid content was significantly enhanced. Compared with the slight acid treatment, the H₂O₂ and malondialdehyde (MDA) contents in black soybean sprouts were increased after ABA treatment, and the black soybean sprouts were further stressed. However, the H₂O₂ and MDA contents in black soybean sprouts were significantly decreased after MT treatment, indicating that the stress of black soybean sprouts can be alleviated by MT. Under slight acid stress, the genes related to flavonoid synthesis in black soybean sprouts were induced by exogenous ABA, promoting the accumulation of flavonoids; under exogenous MT treatment, the activity of phenylpropanoid metabolism enzymes was significantly increased, the genes related to flavonoid synthesis were upregulated, and flavonoid synthesis was induced. These results suggest that the combination of slight acid and plant hormone treatments promotes the accumulation of flavonoid substances during the germination of black soybeans. This research lays the foundation for improving the growth conditions of black soybeans and promoting the enrichment of flavonoid substances in black soybeans.

Broccoli sprouts have high isothiocyanate and selenium accumulation capacity. This study used a combination of methods, including physiological and biochemical, gene transcription and proteomic, to investigate the isothiocyanate and endogenous selenium accumulation mechanisms in broccoli sprouts under exogenous sodium selenite treatment during germination. Compared with the control, the sprouts length of broccoli sprouts under exogenous selenium treatment was significantly lower, and the contents of total phenol and malondialdehyde in 6-day-old broccoli sprouts were substantially higher. The contents of isothiocyanate and sulforaphane in 4-day-old were increased by up-regulating the relative expression of genes of UGT74B1, OX-1, and ST5b. The relative expression of BoSultr1;1, BoSMT, BoHMT1, and BoCOQ5-2 genes regulating selenium metabolism was significantly up-regulated. In addition, 354 proteins in 4-day-old broccoli sprouts showed different relative abundance compared to the control under selenium treatment. These proteins were classified into 14 functional categories. It was discovered that metabolic pathways and biosynthetic pathways of secondary metabolites were significantly enriched. The above results showed that exogenous selenium was beneficial in inducing the accumulation of isothiocyanate and selenium during the growth of broccoli sprouts.