Enrichment of γ-aminobutyric acid in mulberry leaves and the inhibitory effects of the water extract on ACE and α-glucosidase activity

To improve the functional properties of mulberry leaves, different methods were used to treat harvested mulberry leaves to accumulate γ-aminobutyric acid (GABA). The results showed that soaking, cold shock, and anoxic treatments all significantly increased GABA content in mulberry leaves, which were all closely related to the increase of glutamate decarboxylase (GAD) activity and Ca2+ content. Further studies showed that the water extract of the treated mulberry leaves had a higher angiotensin I-converting enzyme (ACE) inhibitory activity than that of the untreated sample, which was close to the effects of using the same amount of GABA alone. The result of molecular docking showed that GABA was stretched into the hydrophobic pocket of ACE to form a stable hydrophobic binding with Ala-354 and Val-380 residues, and formed the hydrogen bond interaction with the residue Glu-384. Importantly, GABA could directly chelated with the zinc ion (bond lengths: 2.9 and 3.1 Å), which was the main interaction between the GABA and ACE. However, the inhibition of mulberry leaf extract on α-glucosidase was not closely related to the GABA content. The results showed that the α-glucosidase inhibitory activity was significantly increased by treating with 6 g/L sodium glutamate for 15 h (IC50 =0.29 mg/mL), and further LC/MS analysis demonstrated that some active compounds especially neochlorogenic acid, chlorogenic acid, cryptochlorogenic acid and rutin were significantly increased in the extract. These data indicated that GABA enrichment was an effective postharvest treatment of mulberry leaves, which can improve its prospects as functional raw materials for industrial production.