Abstract:Alzheimer's disease (AD) is a neurodegenerative disease characterized primarily by progressive memory decline, cognitive impairment, and disturbances in language and psychomotor functions. With the aging of the global population and the deterioration of the living environment, the number of patients is also increasing. The World Alzheimer's Disease Report estimates that the global population of Alzheimer's patients is expected to increase to 115 million by 2050, which could notably affect the world economy and society. The cholinergic hypothesis is a widely accepted theory describing AD pathology, which considers acetylcholine to be an important neurotransmitter involved in learning and memory. The lack of acetylcholine results in insufficient cholinergic signal transmission, consequently contributing to the development of AD. Acetylcholinesterase (AChE) inhibitors can reduce the decomposition of acetylcholine by inhibiting the activity of AChE, making them one of the primary drugs for the treatment of AD. The AChE inhibitor drugs approved by the FDA include donepezil and galantamine. However, the micro-molecule drugs have strong side effects; therefore, finding new safe and efficient AChE inhibitors is necessary. Pathological protein aggregation, oxidative stress, and metal ion homeostasis imbalance exacerbate disease progression. The correlation between different pathogenic factors has shifted drug research from single to multiple targets. Bioactive peptides are peptide compounds with biological activity, which have the advantages of high selectivity, high specificity, multi-target, high safety, and low immunogenicity. In particular, bioactive peptides from marine sources have many specific structures and functions owing to their unique growth environment. Researchers have prepared and isolated peptides from marine organisms with antioxidants, anti-inflammatory, high blood pressure, uric acid, immune regulation, and other effects. Trachinotus ovatus has the advantages of high yield, fast growth, strong disease resistance, high nutritional value, high protein and essential amino acid content, and is a good raw material for preparing bioactive peptides. Enzymatic hydrolysis is the most commonly used method for preparing peptides owing to its mildness, controllability and low cost. In this study, the AChE inhibitory peptide was prepared by enzymolysis of T. ovatus. Enzymolysis was performed under five proteases (neutral protease, papain, alkaline protease, protamines, and trypsin), and different enzymolysis times (1, 2, 4, 6, and 8 h), and the optimal enzymolysis time were screened based on AChE inhibition activity and antioxidant capacity. The hydrolysis degree, molecular weight distribution, amino acid composition and chelating ability of metal ions were determined, and the structure of the products was determined by ultraviolet and Fourier transform infrared. The results showed that the 4 h hydrolysate of papain had the highest AChE inhibition rate (18.02±0.78)%, which was significantly higher than other protease and positive control cerebrolin (P＜0.05). The ABTS radical scavenging rate of the enzymolysis product was (52.54±0.89)%, the degree of hydrolysis was 14.86%, and the proportion of components with molecular weight ＜3 kDa was 96.87%. Correlation analysis showed that AChE inhibition activity was significantly positively correlated with these three indices (P＜0.05). In addition, the amino acids with the highest content in the 4 h papain hydrolysate were glutamic acid, aspartic acid, and lysine, all of which were positively or negatively charged. The proportion of hydrophobic amino acids was 34.92%, which contributed to the interaction between the hydrolysate products and AChE and free radicals, improving the inhibition rate of AChE and antioxidant activity. The binding ability of metal ions was determined. The enzymatic hydrolysis product could bind Ca2+ and Fe2+; the binding rates were (26.28±1.20)% and (14.25±0.85)%, respectively, and the AChE inhibition activity was improved after binding. Ultraviolet analysis showed that the hydrolysates interacted with calcium and iron to form new compounds. Fourier transform infrared spectroscopy showed that amino and carboxyl groups participated in the formation of the complexes. Consequently, the screened hydrolysate has good potential for treating AD. In the future, the AChE inhibition effect of AChE inhibitory peptide in vivo will be further verified, and the primary role of peptide composition will be explored, including the mechanism of action. This provides theoretical support and scientific basis for marine bioactive peptides in the improvement and treatment of AD disease.
