关键词: 虾青素；全反式虾青素；顺式虾青素；胰腺α-淀粉酶；抑制机制

Abstract: This study prepared high-purity Z-astaxanthin (Z-AST) by successive iodine-induced isomerization, ethanol extraction, and column chromatography. The interaction between AST and α-amylase as a key regulatory factor for blood sugar reduction was taken as a starting point to investigated the differences in the inhibitory activity of astaxanthin isomers against porcine pancreatic α-amylase (PPAA) and the underlying mechanism by enzyme inhibition assays, spectroscopic techniques and molecular docking. The results demonstrated that high-purity 9Z-AST (95.07%) and 13Z-AST (90.87%) were successfully prepared and identified. Z-AST exhibited a significantly stronger inhibitory effect on PPAA activity compared with all-E-AST (P < 0.05). Interaction studies revealed that the binding of astaxanthin isomers to PPAA caused only minor changes in the microenvironment of tryptophan (Trp) residues without significant alteration in protein conformation. Molecular docking analysis showed that the binding sites of astaxanthin isomers were located within the catalytic pocket of PPAA, coinciding with the binding site of the competitive inhibitor acarbose, and interacted with key amino acid residues at the enzyme’s catalytic center, namely Asp197, Glu233, and Asp300. This indicated that the inhibition of PPAA by astaxanthin isomers was competitive in nature. The fluorescence of PPAA was statically quenched after binding with astaxanthin isomers. Furthermore, the major interaction forces between PPAA and astaxanthin isomers were identified as van der Waals forces, hydrogen bonds, and hydrophobic interactions, and both isomers formed hydrogen bonds with the Glu233 residue. Notably, Z-AST exhibited shorter hydrogen bonds, indicating a stronger hydrogen bonding interaction. Additionally, Z-AST demonstrated higher binding affinity, interacted with more amino acid residues, and exhibited lower thermodynamic parameters compared with all-E-AST. This suggests that Z-AST possesses a stronger binding activity to PPAA, resulting in a more inhibition effect on the enzyme activity compared with all-E-AST. The findings contribute to the understanding of the differences in hypoglycemic activity between astaxanthin isomers and the potential mechanism, holding significant implications for the development of novel functional foods and hypoglycemic agents.

Key words: astaxanthin; all-E-astaxanthin; Z-astaxanthin; pancreatic α-amylase; inhibition mechanism