| The isolation, purification and structure characterization of polysaccharides of Opuntia monacantha cladode (POMC), as well as their inhibitory effects on the four factors in resulting diabetic chronic complication and the control of blood glucose were investigated in this study. The main contents and results are as follows:Aiming at optimization of the extraction of POMC, response surface methodology was adopted. The results showed that the optimum conditions of Opuntia polysaccharides extraction were as follows: ratio of water to material of 5.5:1, extraction temperature of 75℃, extraction time of 2.2h. Under these conditions the extraction yield could be up to 0.81% by one-time extraction.The purification and concentration of POMC were carried out via membrane ultrafitration using the membranes with different molecular weight cut-off. The optimum conditions for the ultrafiltration were finally determined. It was found that, for the membrane with a molecular mass cut-off of 10000u, when the pressure, temperature, purification time, flow rate and concentration multiple were 0.20 MPa, 25℃, 35 min, 0.04-0.05 L/min and 5~7, respectively, a POMC yield of 0.95% was reached. Moreover, as compared with the traditional vacuum concentration, membrane ultrafiltration was easier to obtain high-quality crude POMC with a higher content of polysaccharide, which was helpful for the further extraction and purification of polysaccharides in the following process.After concentration by membrane ultrafiltration, OP1 was obtained by precipitation extracts with 55% ethanol solution. The supernatant was added ethanol to final concentration of 80% to precipitate OP2. OP3 was obtained by precipitation extracts with 80% of ethanol solution directly. Then, OPM1,OPM2,OPM3 were gained after purification.In vitro trials of inhibitory effects on the four indices reflecting the resulting diabetic chronic complication were evaluated. The results showed: 1) polysaccharide OPM1,OPM2,OPM3 had significant free-radical scavenging activity. Amongst, OPM2 had the highest free-radical scavenging activity. OPM2 and OPM3 had inhibitory effects on Malondialdehyde to different extent; 2) OP2 showed stronger inhibition effect on AGE than aminoguanidine with the same dose at fourth week; 3) OPM2 also showed the strongest inhibition effect on aldose reductase (AR) activity, but lower than epalrestat, the positive control; 4) OPM1 showed the strongest inhibition effect on protein kinase C (PKC) activity in the extraction from cell membrane. OPM2 which had a good performance on above four indices was chosen for hypoglycemic analysis. The resutls showed that it had significant hypoglycemic effect, but lower than aminiguanidine. The body weight loss and polydipsia of diabetic rats were decreased. The high density lipoprotein level and total cholesteral level were increased and decreased, respectively.Then OPM2 was chromatographed on a DEAE Sepharose CL-6B anion exchange column to yield a major fraction (OPMH1). OPMH1 was subjected to further purification on a Sephadex G-100 gel filtration column. Two major fractions, OPMH1 I和OPMH1 II, were collected. By analyses of gel permeation chromatography (GPC), meta-hydroxydiphenyl method and gas chromatography (GC), OPMH1 I, which had a average molecular weight of 42.87 kDa, comprised mainly of rhamnose, arabinose, glucose and glucuronic acid in the molar ratio of 9.15: 1.00: 6.84: 0.51. While OPMH1 II, which had an average molecular weight of 20.67 kDa, comprised mainly of rhamnose, mannose, glucose and glucuronic acid in the molar ratio of 8.72: 1.00: 6.19: 1.06.The results of periodate oxidation and Smith degradation showed that the backbone of OPMH1 I was 1→2 rhamnose and 1→3 rhamnose. Their molar ratio was 2.06: 1. The side chains mainly comprised of branched (1→3)-linked glucose and (1→3)-linked arabinose. The backbone of OPMH1 II was 1→2 rhamnose and 1→3 rhamnose. Their molar ratio was 1.56: 1. The side chains mainly comprised of branched (1→3)-linked glucose and (1→3)-linked mannose.
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