| Plant sterols (phytosterols) and stanols (phytostanols) are generally extracted from thedeodorizer distillates. Both of them are regarded as―the key of life‖due to their manyimportant beneficial properties, such as cholesterol-lowering effects, anti-cancer,anti-inflammatory property and so on. Plant stanols (PS) are the hydrogenated counterparts ofplant sterols. However, the unique chemical structure of plant sterols and stanols determinestheir insolubility in water and poor solubility in oil, which greatly limits their application infood, medical, cosmetic and other industries. The present study aimed to synthesize plantstanol derivatives via chemical modification to improve their solubility in oil or water. On thebasis of modification, their physiological functions and the potential molecular mechanismwere investigated.The reaction products were firstly analyzed and separated. The HPLC mobile phases forplant stanyl laurate (PSL), hemisuccinate (PSH) and sorbitol succinate (PSS) analysis weremethanol/isopropanol/n-hexane (8/1/1, v/v/v), methanol/formic acid (1000/1, v/v) andmethanol/formic acid (1000/1, v/v), respectively. And the silica gel column chromatographiceluents for plant stanyl laurate, hemisuccinate and sorbitol succinate were cyclohexane/ethylacetate (8/2, v/v), petroleum ether/ethyl acetate/formic acid (10/10/0.02, v/v/v) and ethylacetate/methanol/formic acid (18/2/0.15, v/v/v), respectively. The purified products wereanalyzed by FT-IR, MS and NMR, and confirmed to be the goal products.The chemical modification of phytostanols can be divided into two aspects. On the onehand, plant stanol fatty acid esters (PSE) were successfully synthesized via chemicalmodification to improve their solubility in vegetable oil. The highest conversion of80%wasobtained under the selected conditions: lauric acid (LA) as acyl donor,25μmol/mL plantstanols,100μmol/mL lauric acid,80mg/mL3molecular sieves and40mg/mL Novozym435at150r/min and55oC for96h in10mL of n-hexane. On the other hand, hydrophilicplant stanol derivatives-plant stanol sorbitol succinate was prepared via chemo-enzymaticways to improve their hydrophilic property. The highest yield (>51%) of plant stanyl sorbitolsuccinate was obtained under the below conditions:50μmol/mL plant stanyl hemisuccinate,1:3molar ratio of plant stanyl hemisuccinate to D-sorbitol,80mg/mL3molecular sieves and100mg/mL Lipozyme RM IM in tert-butanol,150r/min and55oC.The physiological activities of plant sterols (270mg/(kg·d)), stanols (270mg/(kg·d)),phytosteryl laurate (386mg/(kg·d)) and phytostanyl laurate (386mg/(kg·d)) were investigatedin vivo using KM mice as animal model. Results showed that the free plant sterols (-11.5%)and stanols (-14.7%) could markedly lower the serum cholesterol in mice, and plant steryllaurate (-13.2%) and stanyl laurate (-17.6%) still retained the cholesterol-lowering property of the free forms. In detail, plant steryl and stanyl laurate could effectively reduce the serum totalcholesterol (TC) and low density lipoprotein cholesterol (LDL-C) levels in mice, and have noeffect on the serum high density lipoprotein cholesterol (HDL-C) and triglyceride (TAG)levels. There were no significant differences in reducing cholesterol between equimolarphytosterols and phytostanols, or between phytosteryl laurate and phytostanyl laurate. Also,plant sterols, stanols, and their laurate could remarkably decrease liver cholesterol content,and effectively promote the cholesterol excretion via feces.The physiological activities and the influence on the gene expression related tocholesterol metabolism of intermediate products–hemisuccinate (1×:167mg/(kg·d);2×:335mg/(kg·d);5×:839mg/(kg·d)) and hydrophilic plant stanol derivatives–sorbitol succinate(1×:221mg/(kg·d);2×:442mg/(kg·d);5×:1106mg/(kg·d)) were also investigated. Plantstanyl hemisuccinate (2×,-13.3%;5×,-14.6%) and plant stanyl sorbitol succinate (2×,-12.9%;5×,-15.0%) still retained the cholesterol-lowering effect, and had no effect on the serumblood glucose, alanine aminotransferase (ALT) and aspertate aminotransferase (AST) level inmice. Both of them could promote the liver alpha liver X receptor (LXR-α) and cholesterol7alpha-hydroxylase (CYP7A1) gene expression, but had no influence on the liver a3-hydroxy-3-methylglutaryl-coenzyme-a reductase (HMGCR), low density lipoproteinreceptor (LDLR) and sterol regulatory element binding protein (SREBP-2) mRNA level inmice. The cholesterol-lowering effect of plant stanol derivatives may be through activatingthe potential LXRα-CYP7A1-bile acid-fecal excretion pathway.The solubility in oil or water, crystallization and melting behavior, thermal stability ofplant stanols and their derivatives were explored. The solubility of plant stanol fatty acidesters in soybean oil was higher than plant stanols. The oil solubility of plant stanol saturatedfatty acid esters decreased with the increase of the carbon chain in fatty acids, and the oilsolubility of plant stanol unsaturated fatty acid esters increased with the number increase ofunsaturated double bond. Plant stanol sorbitol succinate improved the hydrophilic property ofplant stanols to a certain extent. Plant stanyl fatty acid esters had lower melting temperatureand crystallization temperature than plant stanols, and the melting and crystallizationtemperature increased with the increase of the carbon chain. Plant stanyl hemisuccinate andsorbitol succinate had higher melting and crystallization temperature. The crystallizationprofiles and sensory characteristics of the soybean oil were not affected when the amount ofplant sterols, stanol and laurate was not more than2%, or oleate and linoleate not exceed than5%. The modified products-both plant stanol fatty acid esters and plant stanyl sorbitolsuccinate had good thermal stability. |
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