Synthesis Of Mannan Oligosaccharide-sialic Acid Conjugates And Their Inhibitory Effects On Aβ Aggregation And Neurotoxicity

The abnormal aggregation of amyloidβ-peptide（Aβ）is considered one of the main pathological changes in the onset of Alzheimer’s disease（AD）and a target for the development of AD treatment drugs.In the previous research,a mannuronic acid oligosaccharide-sialic acid complex（MOS-Sia）synthesized from seaweed-derived mannuronic acid oligosaccharides（MOS）and sialic acid（Sia）was found to disrupt Aβ42 aggregation and inhibit inflammatory responses.Based on this,MOS-Sia was applied to an AD mouse model to investigate its effects on the pathological process of AD.Additionally,this study further utilized locust bean gum-derived mannose oligosaccharides（LBOS）to synthesize a mannose oligosaccharide-sialic acid conjugate and its derivatives（p LBOS-Sia）through sialic acid modification and studied their structure and function to provide possible ideas for the development of AD prevention and treatment drugs.The main research of this paper is as follows:1.Firstly,based on the existing MOS-Sia synthesis process,the structures of MOS prepared by microwave,oxidation,and acid hydrolysis methods and the thiol substitution degree of thiolated MOS（MOS-SH）synthesized were compared.Through infrared spectroscopy and mass spectrometry analysis,it was found that the acid hydrolysis method could prepare MOS with relatively high structural integrity and substitution degree（38.76%）.Then,through the thiol condensation method,MOS-SH reacted with methyl ester acetylated protected Sia to produce MOS-Sia with a yield of 78.86%.Infrared spectroscopy and mass spectrometry analysis showed that MOS-Sia was successfully synthesized.Finally,a double transgenic AD model(FAD^4T)mouse experiment was used for drug intervention using MOS-Sia aqueous solution with a concentration gradient.Behavioral tests were performed using a water maze,histopathological examination was performed using H&E staining and immunohistochemistry,and changes in inflammatory factors in the body were detected by enzyme-linked immunosorbent assay.The results showed that MOS-Sia could slow down the damage to the learning and cognitive memory function of AD mice.A higher concentration(1.5 mg·m L^-1)of MOS-Sia could slow down the loss and apoptosis of hippocampal neurons in AD mice to a certain extent.Compared with the positive group,there was a certain reduction in Aβplaques in the hippocampus of AD mice in the low and high-concentration groups.Among them,the expression of phosphorylated Tau protein（p-Tau）in the low-concentration group was reduced,and the IL-1βlevels in all dosing groups and IL-6 levels in the medium and high-concentration groups of the dosing group were significantly reduced（P<0.05）.2.The synthesis and characterization of p LBOS-Sia were carried out.Using LBG as raw material,LBOS with a degree of polymerization of 3-13 and mainly composed of mannose was obtained by stepwise hydrolysis usingβ-mannanase andα-galactosidase.Then,the activated LBOS was reacted with Sia to synthesize a mannose oligosaccharide derivative（LBOS-Sia）with sialic acid connected at the end and then phosphorylated to obtain p LBOS-Sia.Infrared spectroscopy,mass spectrometry,and nuclear magnetic resonance hydrogen spectrum analysis showed that p LBOS-Sia was successfully synthesized.Soluble protein content determination,biological microscope observation,thioflavin T labeling,and circular dichroism spectroscopy analysis showed that both LBOS-Sia and p LBOS-Sia had the effect of inhibiting Aβ42 aggregation.MTT analysis and enzyme-linked immunosorbent assay showed that LBOS-Sia and p LBOS-Sia had no cytotoxicity to BV-2 cells and could significantly reduce the release of TNF-αinduced by Aβ42 in BV-2 cells（P<0.05）.Therefore,the mannose oligosaccharide-sialic acid derivative p LBOS-Sia synthesized using LBG as raw material has the function of inhibiting Aβ42 aggregation and inflammatory response,which may provide ideas for the development of carbohydrate drugs raw materials or additives for the treatment of AD in the future.