红松子油中皮诺敛酸的纯化及降脂活性评价
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摘要
以红松子油为原料,采用尿素包合法纯化皮诺敛酸,通过脂肪酸与尿素的比例、尿素与乙醇的比例、包合温度及包合时间等因素对皮诺敛酸含量及产率的影响,优化了尿素包合法的工艺参数,最终确定了皮诺敛酸的最佳纯化条件为:脂肪酸与尿素的比例为1:6. 5(w/w),尿素与90%乙醇的比例为1:4(w/v),包合温度为-10℃,包合时间12 h。在上述条件下,经GC-MS检测获得了纯度为58. 3%的皮诺敛酸,回收率为51. 8%,经二次尿素包合纯化后可获得纯度为72. 5%的皮诺敛酸,回收率为64. 7%。为了评估皮诺敛酸的降脂活性,以油酸诱导HepG2细胞成脂,最终确定40μmol·L~(-1)的皮诺敛酸能显著的降低HepG2细胞内甘油三酯及总胆固醇的水平。
With Korean pine seed oil we enriched and purifiedpinoleic acid by urea inclusion. Through the influence of the ratio of fatty acid to urea,the ratio of urea to ethanol,we optimized the inclusion temperature and the inclusion time on the content of pinoleic acid and recovery of pinoleic acid,the process parameters of urea bagging. Finally,the optimal purification condition of pinoleic acid was determined as follows: the ratio of fatty acid to urea was 1:6. 5( w/w); the ratio of urea to 90% ethanol was 1:4( w/v); Inclusion temperature was -10℃; Inclusion time was 12 h. Under the above conditions,a purity of 58. 3% pinoleic acid could be obtained,and the recovery rate was 51. 8%. After purification by secondary urea inclusion,a purity of 72. 5%of pinoleic acid could be obtained,and the recovery rate was 64. 7%. To verify the lipid-lowering activity of pinoleic acid,oleic acid was used to induce lipogenesis in HepG2 cells. The results showed 40 μmol · L~(-1) pinoleic acid could significantly reduce the level of triglycerides and total cholesterol in HepG2 cells.
With Korean pine seed oil we enriched and purifiedpinoleic acid by urea inclusion. Through the influence of the ratio of fatty acid to urea,the ratio of urea to ethanol,we optimized the inclusion temperature and the inclusion time on the content of pinoleic acid and recovery of pinoleic acid,the process parameters of urea bagging. Finally,the optimal purification condition of pinoleic acid was determined as follows: the ratio of fatty acid to urea was 1:6. 5( w/w); the ratio of urea to 90% ethanol was 1:4( w/v); Inclusion temperature was -10℃; Inclusion time was 12 h. Under the above conditions,a purity of 58. 3% pinoleic acid could be obtained,and the recovery rate was 51. 8%. After purification by secondary urea inclusion,a purity of 72. 5%of pinoleic acid could be obtained,and the recovery rate was 64. 7%. To verify the lipid-lowering activity of pinoleic acid,oleic acid was used to induce lipogenesis in HepG2 cells. The results showed 40 μmol · L~(-1) pinoleic acid could significantly reduce the level of triglycerides and total cholesterol in HepG2 cells.
引文
1.苏晓雨,王振宇.红松子种皮提取物活性成分及抗氧化作用研究[J].林产化学与工业,2010,30(4):99-102.Su X Y,Wang Z Y. Study on the active component and antioxidant activities of extract from Pinus koraiensis Sieb. et Zucc. seed shell[J]. Chemistry and Industry of Forest Products,2010,30(4):99-102.
2 .杨明非,苏雯,寇萍,等.高速匀质—微波辅助提取红松籽油工艺及其品质评价[J].植物研究,2017,37(5):789-796.Yang M F,Su W,Kou P,et al. Ultra turrax-microwave assisted extraction of oil from Pinus koraiensis Sieb. Seed and its quality evaluation[J]. Bulletin of Botanical Research,2017,37(5):789-796.
3 .杨明非,张晶,苏雯,等.皮诺敛酸/左旋肉碱降低HepG2细胞脂质的最佳浓度配比研究[J].植物研究,2017,37(6):941-946.Yang M F,Zhang J,Su W,et al. The optimized proportion of combination pinoleic acid/l-carnitinedecrease lipid in HepG2[J]. Bulletin of Botanical Research,2017,37(6):941-946.
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6 .戚登斐,张润光,韩海涛,等.核桃油中亚油酸分离纯化技术研究及其降血脂功能评价[J].中国油脂,2019,44(2):104-108.Qi D F,Zhang R G,Han H T,et al. Separation and purification of linoleic acid from walnut oil and its hypolipidemic function evaluation[J]. China Oils and Fats,2019,44(2):104-108.
7 .刘志鑫,刘志彬,张根生,等.尿素包合法富集压榨松籽油中皮诺敛酸工艺优化[J].食品与机械,2018,34(8):183-188.Liu Z X,Liu Z B,Zhang G S,et al. Optimization of the process for the enrichment of pinolenic acid from pressed pine seed oil by urea inclusion method[J]. Food&Machinery,2018,34(8):183-188.
8 .徐水祥,周淡宜,徐飞鹤,等.甲鱼油中多不饱和脂肪酸的提取与纯化研究[J].食品科学,2004,25(11):118-120.Xu S Y,Zhou D Y,Xu F H,et al. Study on extracting and purification PUFA from turtle oil[J]. Food science,2004,25(11):118-120.
9 .郑晓吉,罗鹏,陈计峦,等.尿素包合法纯化棉籽油中亚油酸的研究[J].食品研究与开发,2010,31(5):51-53.Zheng X J,Luo P,Chen J L,et al. Purification of linoleic acid from cotton seed oil by urea adduction fractionation[J]. Food Research and Development,2010,31(5):51-53.
10 .程亚鹏,毕艳兰,刘伟,等.尿素包合法制备高纯度油酸乙酯的工艺条件优化[J].中国油脂,2018,43(9):1-7.Cheng Y P,Bi Y L,Liu W,et al. Optimization of preparation of high purity ethyl oleate by urea inclusion method[J]. China Oils and Fats,2018,43(9):1-7.
11 .董秀婷,杨国恩,王秋敏,等.尿素包合法分离油用牡丹籽油的不饱和脂肪酸工艺优化研究[J].中国油脂,2018,43(8):39-41,46.Dong X T,Yang G E,Wang Q M,et al. Optimization of urea inclusion process for separation of unsaturated fatty acids from peony seed oil[J]. China Oils and Fats,2018,43(8):39-41,46.
12 .王培燕,黄元波.尿素包合法分离混合脂肪酸研究进展[J].绿色科技,2017,(20):186-189.Wang Y P,Huang Y B. Development of urea-inclusion separation of mixed fatty acids[J]. Journal of Green Science and Technology,2017,(20):186-189.
13 .陶川东,鲁厚芳,梁斌.尿素包合法分离麻疯树籽油脂肪酸甲酯的研究[J].中国油脂,2013,38(5):64-68.Tao C D,Lu H F,Liang B. Separation of fatty acid methyl esters from Jatropha curcus L. seed oil using urea adduction method[J]. China Oils and Fats,2013,38(5):64-68.
14 . Hays S M,Mcginnis C. Nonalcoholic Fatty Liver Disease in Children:Beyond Metabolic Syndrome[J]. The Journal for Nurse Practitioners,2018,14(10):725-731.
15 .徐杰,李伟,杨辉.脂代谢紊乱对2型糖尿病患者心血管疾病发生和病死率的影响[J].岭南心血管病杂志,2017,23(06):759-763.Xu J,Li W,Yang H. Effect of dyslipidemia on cardiovascular disease and mortality in patients with type 2 diabetes[J]. South China Journal of Cardiovascular Diseases,2017,23(06):759-763.
16 .刘锐,何嘉,刘秀芳,等.多烯磷脂酰胆碱干预大鼠非酒精性脂肪肝细胞模型基因芯片组学分析[J].中国组织工程研究,2018,22(36):5833-5839.Liu R,He J,Liu X F,et al. Gene chipset analysis in rats with non-alcoholic fatty liver disease intervened by polyene phosphatidyl choline[J]. Chinese Journal of Tissue Engineering Research,2018,22(36):5833-5839.
17 . Cui W,Chen S L,Hu K Q. Quantification and mechanisms of oleic acid-induced steatosis in HepG2 cells[J]. American Journal of Translational Research,2010,2(1):95-104.
18 . Elshaghabee F M F,Rokana N,Panwar H,et al. Probiotics as a dietary intervention for reducing the risk of nonalcoholic fatty liver disease[M].//Arora D,Sharma C,Jaglan S,et al. Pharmaceuticals from microbes:impact on drug discovery. Cham:Springer,2019:207-223.
19 . Vidyashankar S,Varma R S,Patki P S. Quercetin ameliorate insulin resistance and up-regulates cellular antioxidants during oleic acid induced hepatic steatosis in HepG2cells[J]. Toxicology in Vitro,2013,27(2):945-953.
20 . Rafiei H,Omidian K,Bandy B. Dietary polyphenols protect against oleic acid-induced steatosis in an in vitro model of NAFLD by modulating lipid metabolism and improving mitochondrial function[J]. Nutrients,2019,11(3):541.
2 .杨明非,苏雯,寇萍,等.高速匀质—微波辅助提取红松籽油工艺及其品质评价[J].植物研究,2017,37(5):789-796.Yang M F,Su W,Kou P,et al. Ultra turrax-microwave assisted extraction of oil from Pinus koraiensis Sieb. Seed and its quality evaluation[J]. Bulletin of Botanical Research,2017,37(5):789-796.
3 .杨明非,张晶,苏雯,等.皮诺敛酸/左旋肉碱降低HepG2细胞脂质的最佳浓度配比研究[J].植物研究,2017,37(6):941-946.Yang M F,Zhang J,Su W,et al. The optimized proportion of combination pinoleic acid/l-carnitinedecrease lipid in HepG2[J]. Bulletin of Botanical Research,2017,37(6):941-946.
4 . Xie K Y,Miles E A,Calder P C. A review of the potential health benefits of pine nut oil and its characteristic fatty acid pinolenicacid[J]. Journal of Functional Foods,2016,23:464-473.
5 . Christiansen E,Watterson K R,Stocker C J,et al. Activity of dietary fatty acids on FFA1 and FFA4 and characterisation of pinolenic acid as a dual FFA1/FFA4 agonist with potential effect against metabolic diseases[J]. British Journal of Nutrition,2015,113(11):1677-1688.
6 .戚登斐,张润光,韩海涛,等.核桃油中亚油酸分离纯化技术研究及其降血脂功能评价[J].中国油脂,2019,44(2):104-108.Qi D F,Zhang R G,Han H T,et al. Separation and purification of linoleic acid from walnut oil and its hypolipidemic function evaluation[J]. China Oils and Fats,2019,44(2):104-108.
7 .刘志鑫,刘志彬,张根生,等.尿素包合法富集压榨松籽油中皮诺敛酸工艺优化[J].食品与机械,2018,34(8):183-188.Liu Z X,Liu Z B,Zhang G S,et al. Optimization of the process for the enrichment of pinolenic acid from pressed pine seed oil by urea inclusion method[J]. Food&Machinery,2018,34(8):183-188.
8 .徐水祥,周淡宜,徐飞鹤,等.甲鱼油中多不饱和脂肪酸的提取与纯化研究[J].食品科学,2004,25(11):118-120.Xu S Y,Zhou D Y,Xu F H,et al. Study on extracting and purification PUFA from turtle oil[J]. Food science,2004,25(11):118-120.
9 .郑晓吉,罗鹏,陈计峦,等.尿素包合法纯化棉籽油中亚油酸的研究[J].食品研究与开发,2010,31(5):51-53.Zheng X J,Luo P,Chen J L,et al. Purification of linoleic acid from cotton seed oil by urea adduction fractionation[J]. Food Research and Development,2010,31(5):51-53.
10 .程亚鹏,毕艳兰,刘伟,等.尿素包合法制备高纯度油酸乙酯的工艺条件优化[J].中国油脂,2018,43(9):1-7.Cheng Y P,Bi Y L,Liu W,et al. Optimization of preparation of high purity ethyl oleate by urea inclusion method[J]. China Oils and Fats,2018,43(9):1-7.
11 .董秀婷,杨国恩,王秋敏,等.尿素包合法分离油用牡丹籽油的不饱和脂肪酸工艺优化研究[J].中国油脂,2018,43(8):39-41,46.Dong X T,Yang G E,Wang Q M,et al. Optimization of urea inclusion process for separation of unsaturated fatty acids from peony seed oil[J]. China Oils and Fats,2018,43(8):39-41,46.
12 .王培燕,黄元波.尿素包合法分离混合脂肪酸研究进展[J].绿色科技,2017,(20):186-189.Wang Y P,Huang Y B. Development of urea-inclusion separation of mixed fatty acids[J]. Journal of Green Science and Technology,2017,(20):186-189.
13 .陶川东,鲁厚芳,梁斌.尿素包合法分离麻疯树籽油脂肪酸甲酯的研究[J].中国油脂,2013,38(5):64-68.Tao C D,Lu H F,Liang B. Separation of fatty acid methyl esters from Jatropha curcus L. seed oil using urea adduction method[J]. China Oils and Fats,2013,38(5):64-68.
14 . Hays S M,Mcginnis C. Nonalcoholic Fatty Liver Disease in Children:Beyond Metabolic Syndrome[J]. The Journal for Nurse Practitioners,2018,14(10):725-731.
15 .徐杰,李伟,杨辉.脂代谢紊乱对2型糖尿病患者心血管疾病发生和病死率的影响[J].岭南心血管病杂志,2017,23(06):759-763.Xu J,Li W,Yang H. Effect of dyslipidemia on cardiovascular disease and mortality in patients with type 2 diabetes[J]. South China Journal of Cardiovascular Diseases,2017,23(06):759-763.
16 .刘锐,何嘉,刘秀芳,等.多烯磷脂酰胆碱干预大鼠非酒精性脂肪肝细胞模型基因芯片组学分析[J].中国组织工程研究,2018,22(36):5833-5839.Liu R,He J,Liu X F,et al. Gene chipset analysis in rats with non-alcoholic fatty liver disease intervened by polyene phosphatidyl choline[J]. Chinese Journal of Tissue Engineering Research,2018,22(36):5833-5839.
17 . Cui W,Chen S L,Hu K Q. Quantification and mechanisms of oleic acid-induced steatosis in HepG2 cells[J]. American Journal of Translational Research,2010,2(1):95-104.
18 . Elshaghabee F M F,Rokana N,Panwar H,et al. Probiotics as a dietary intervention for reducing the risk of nonalcoholic fatty liver disease[M].//Arora D,Sharma C,Jaglan S,et al. Pharmaceuticals from microbes:impact on drug discovery. Cham:Springer,2019:207-223.
19 . Vidyashankar S,Varma R S,Patki P S. Quercetin ameliorate insulin resistance and up-regulates cellular antioxidants during oleic acid induced hepatic steatosis in HepG2cells[J]. Toxicology in Vitro,2013,27(2):945-953.
20 . Rafiei H,Omidian K,Bandy B. Dietary polyphenols protect against oleic acid-induced steatosis in an in vitro model of NAFLD by modulating lipid metabolism and improving mitochondrial function[J]. Nutrients,2019,11(3):541.