Preparation Of Different Molecular Weight Fucoidans And Their Effect On Prevention And Treatment Of Hepatocarcinoma

Fucoidan is a sulfated polysaccharide mainly derived from the cell walls of brown algae and marine invertebrates.It has rich biological activities such as antioxidant,antitumor,and immune regulation,and is currently a hot research topic.Due to different types of raw materials,harvesting seasons,geographical locations,and extraction methods,the structure and molecular weight of fucoidan are also different during the preparation process.Due to the complexity of the structure and high molecular weight of fucoidan,its low bioavailability limits the research on its biological activity.The research on the preparation of different molecular weight fucoidan gums and clarifying the relationship between biological activity and molecular weight is of great significance.Therefore,this article aims to study the extraction process of kelp derived fucoidan and prepare different molecular weight fucoidan through molecular weight interception and classification,and conduct preliminary characterization and activity research on it.Explore the preventive and therapeutic effects of different molecular weight fucoidan on mouse liver cancer.The main research content is as follows:We use biological enzymatic hydrolysis to effectively release the active components of kelp,and obtain crude fucoidan through graded alcohol precipitation.Then,we use molecular weight interception grading to prepare different molecular weight components F-1(<3.5 k Da),F-2(3.5-10 k Da),F-3(10-100 k Da),F-4(100-300 k Da),and F-5(>300k Da)for activity research.The yield rates were 26.8%,19.4%,10.7%,33.6%,and 6.3%,respectively,with a total recovery rate of 96.8%.Detect the chemical composition of each component,and the total sugar content is 56.28%,59.66%,68.1%,62.14%,and 77.56%,respectively;The sulfate content is 22.8%,16%,21.03%,15.61%,and 19.8% respectively;The content of fucose is 48.98%,33.67%,31.55%,24.78% and 38.14% respectively.The monosaccharide composition of F-1,F-2,F-3,F-4 and F-5 was determined by high performance liquid chromatography(HPLC).It was found that F-1,F-2,F-3,F-4 and F-5 were polysaccharides composed of mannose,fucose,xylose,rhamnose and galactose.F1 had the highest content of fucose,48.98%.By measuring the functional groups of each component through infrared spectroscopy,it was found that all five components contain characteristic absorption peaks of polysaccharides and sulfate ions.Through GC-MS analysis,it was proven that the main chains are all α(1 → 3)Linked L-neneneba fucose is composed of fucose branched chains with partial 1 → 4 links.Sulfate is mainly linked to fucose C2 and C4 positions,and also contains other substituents,such as galactose and uronic acid.By verifying the cell survival rate and colony forming ability of human liver cancer Hep G2 cells with different molecular weight fucoidan,as well as the effect on the transformation ability of mouse BALB/3T3 embryonic fibroblasts,we aim to explore its in vitro anti-tumor activity.The MTT results showed that the IC50 values of the five components of fucoidan were 184.7 μg/m L,274.3 μg/m L,416.2 μg/m L,790.1 μg/m L,and 1556 μg/m L,respectively.The results showed that F-1 had the best in vitro anti-tumor activity.From the perspective of the colony forming ability of Hep G2 cells and the transformation ability of BALB/3T3 cells,the F-1 component has significant in vitro antitumor activity.By comparing the effects of fucoidan on the chemoprevention ability of tumors in H22 mice,the results of the tumor formation rate of fucoidan showed that fucoidan can significantly inhibit tumor generation through immune regulation.The results of mouse blood immunological indicators showed that the number and percentage of white blood cells,neutrophils,neutrophil/lymphocyte ratio,and platelet/lymphocyte ratio were significantly increased,Its thymus index and spleen index were significantly increased compared to the model group(* * p<0.01)and could increase IL-1β in mouse serum 、IL-6 and tumor necrosis factor TNF-α The level and inhibition of vascular endothelial growth factor VEGF expression slow down the generation of blood vessels in tumor tissue,achieving the goal of inhibiting tumors.Through the H22 tumor bearing mouse model,it was shown that fucoidan has good in vivo anti-tumor activity,with a tumor inhibition rate of 58%.Tumor pathological sections showed that the tumor cells treated with fucoidan had apoptosis characteristics such as Karyorrhexis,nuclear concentration,etc.The cell size was reduced,the cytoplasm was dense,eosinophilic staining was enhanced,and the cells were scattered.The tumor cells in the model group were relatively compact.Analyzing the inflammatory factor IL-1β in serum 、 IL-6 and tumor necrosis factor TNF-α The level of fucoidan and the changes in thymus and spleen indices suggest that fucoidan may have anti-tumor effects by activating the body’s immune regulation and enhancing its immune function.We measured the level of VEGF(vascular endothelial growth factor)using ELISA and the expression level of its downstream signaling molecule PI3K/AKT/m TOR using q PCR.The results showed that fucoidan significantly reduced the expression of VEGF and further reduced the expression of its downstream gene PI3K/AKT/m TOR,indicating that it may exert anti-tumor effects through the VEGF pathway.