| Phytosterol glycolipids(PG),a membrane-bound sterol molecule derivatives,is mainly composed of sterol glycosides(SG)and acyl sterol glycosides(ASG),with the reported functions of lowering cholesterol,enhancing immunity,coagulation,liver targeting,analgesia,anti-malarial,and lowering blood sugar,the application value of PG is very high.Because of 5%-10% PG content,soybean lecithin is regarded as an important raw material of PG preparation.Therefore,studys on isolation,component analysis and application characteristics of soybean PG,not only provide basis for the design and development of PG goods,but also have great significance of high-end phospholipid production process.In this paper,soybean lecithin powder was used as material of PG preparation,after optimized the isolation parameters,soybean PG were detected for its exact molecular composition,then the adsorption behavior and mechanism mentioned in soybean PG separation process were explored,properties of PG liposome were examined,finally,effects of the combination of PG and PC on nerve cell were discussed.The main researching aspects and results showed below:With 4 indicators containing phospholipid yield(Y1),ASG recovery(Y2),SG recovery(Y3)and PG purity(Y4)as the dependent variables,the optimized parameters including silicagel dosage 3:1,lecithin solution concentration 50 mg/mL,and adsorption temperature 26 oC were determined for the best PG separation efficiency,where Y1-Y4 respectively reached 78.36% ± 4.11%,97.21% ± 3.48%,94.60% ± 2.91% and 59.99% ± 2.76%.The exact molecular compositions of soybean PG were analyzed,after a separation and purification,ASG fraction and SG fraction with a purity of 85.17% ± 1.83 % and 81.60% ± 2.24 % were obtained at a yield of 35.20% and 15.09% respectively.Based on a detection of fat acid and sterol in PG,?-sitosterol(49%-57%),?-stigmasterol(18%-20%)and ?-campesterol(24%-29%)were identified as the main sterol skeleton of PG,while the acyl compositions of ASG were mainly composed of linoleic acid(44.78% ± 2.23%),palmitic acid(27.77% ± 0.82%),stearic acid(5.52% ± 0.21%)and oleic acid(5.27% ± 0.32%).Finally,the specific molecular compositions of the purified ASG and SG fraction were determined by UPLC-MS,with linoleoyl sterol glycoside(53.37%),palmityl sterol glycoside(20.05%)and linolenoyl sterol glycoside(5.83%)as ASG main compositions,SG contained sitosteryl glycoside,campesteryl glycoside and stigmasteryl glycoside.The mechanism involved in PG isolation process was investigated,based on a fitting degree of 0.9807 and 0.9548,the pseudo-first-order kinetic model was best suitable to ASG adsorption at 309 K,while the pseudo-second-order kinetic model to SG adsorption.With an increase of adsorption temperature,the theoretical maximized ASG adsorption capacity decreased from 18.5332 mg/g(299 K)to 14.15138 mg/g(319 K),and its multi-molecular layer adsorption was turned into the monomolecular layer adsorption.Because of the negative ?G,?H,and ?S in ASG adsorption process,ASG adsorption process was considered as a spontaneouse exothermic physical adsorption process driven by changes of enthalpy.While in SG adsorption,its theoretical maximized adsorption amount increased by 2.0822 mg/g,and monolayer adsorption was present in all the isothermal adsorption experiment.Owing to the thermodynamic parameters containing ?G < 0,0 < ?H < 25 KJ/mol and ?S < 0.1 KJ/(mol*K),SG adsorption was regarded as a spontaneous endothermic process,mainly controlled by changes of entropy.Effects of PG on the property of pure PC liposome were studied,with an average particle size of Pure PC liposome at 200 nm,ASG benefited the hydration of liposome membrane,it was natural of ASG liposome to have a lower particle size and turbidity,which was opposite to SG.When stored at 10 oC for 33 days,changes of liposomes diameter could be reduced by PG effectively,meaning the enhanced storage stability.At the mantime,PG liposome has a lower TBARS rate than cholesterol liposome,confirming the better anti-peroxidation ability of PG liposome.With PG addition less than 20%,SG not only reduced the distorted conformation,but also increased the longitudinal order of the pure PC liposome,indicating the enhanced stability of pure PC liposome by PG.Application properties of PG liposome were explored,when carboplatin was encapsulated by PG liposome,its leakage at 35 oC for 120 h was reduced,where carboplatin leakage behavior in SG liposome was a diffusion behavior of the skeleton into the medium,the concentration difference between the liposome and solvent might be the main driving force of carboplatin leakage.Finally,HepG2 was brought as the cell model to investigate effects of PG on the cells safety,cell uptake,and the carboplatin activity of liposome,it was easy of PG liposome to enrich in HepG2 cells,no side-effects were discovered after HepG2 cells were cultured in 600 ?g/mL PG liposome.With IC50 of carboplatin in ASG and SG liposomes occupying 69.84% and 62.13% of free carboplatin,the anticancer activity of carboplatin was enhanced by PG liposome.To reduce the side effects of PG liposome infusion,effects of combination of PG and PC on nerve cells-human neuroblastoma cell SH-SY5 Y were investigated.After SH-SY5 Y cells cultured in mediums containing 50 ?g/mL ASG or 25 ?g/mL SG for 48 h,the cell viabilities respectively reduced to 63.74% ± 2.29% or 67.85% ± 3.51% of the blank group,which could be transfored by PC,when ASG or SG mixed PC at a ratio of 1/8 or 1/12,effects of PG on SHSY5 Y cell were minimized.Compared with PG group,the cell viability and mitochondrial potential of ASG-PC group increased by 29.72% and 30.18%,while its apoptosis rate decreased by 14.71%;While in SG and PC synergistic group,the decreased the cell viability and mitochondrial potential were 29.06% and 27.15%,coupled with a declined apoptosis rate of 13.56%;as the increased expression of phosphorylated AKT in the synergetic group could be blocked by LY294002,the apoptosis of SH-SY5 Y cell.could be mediated through PI3K/AKT mitochondrial pathway by the combination of PG and PC. |
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