南极磷虾中不同形态虾青素的分离制备、结构鉴定及含量分析
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摘要
以南极磷虾为研究对象,采用丙酮提取南极磷虾中的虾青素。优化硅胶柱层析技术分离虾青素双酯、单酯及游离虾青素;筛选合适的展开剂,利用薄层层析法对得到的样品进行鉴定;通过Chiralpak IC柱鉴别其立体结构;运用C_(30)-HPLC法测定不同形态虾青素的含量。结果表明,硅胶柱层析以不同比例的丙酮-石油醚溶液梯度淋洗,可以分离制备虾青素双酯、单酯及游离虾青素; V(正己烷)∶V(丙酮)∶V(乙酸)=8∶2∶0. 2为薄层层析最佳展开剂。南极磷虾中不同形态虾青素均由3S,3'S、3S,3'R、3R,3'R 3种光学异构体组成,虾青素双酯和单酯中3R,3'R所占比例> 70%,3S,3'S的比例最低。南极磷虾中总虾青素含量为18. 9 mg/kg,其中虾青素双酯中虾青素含量为12. 0 mg/kg,虾青素单酯中虾青素含量为6. 1 mg/kg,游离虾青素含量为0. 8 mg/kg。
In this study,acetone was used to extract the astaxanthin from Antarctic krill. The silica gel column chromatography was optimised for separating astaxanthin diester,astaxanthin monoester,and free astaxanthin. A suitable developing solvent was selected. Samples were identified by a thin layer chromatography and the three-dimensional structures were confirmed by Chiralpak IC column. The contents of astaxanthin in different forms were quantified using C_(30)-HPLC. The results showed that a gradient elution through silica gel column chromatography with different ratios of acetone-petroleum ether could completely separate free astaxanthin,astaxanthin monoester and diester. V( nhexane) ∶ V( acetone) ∶ V( acetic acid) = 8∶ 2∶ 0. 2 was the optimum developing solvent for thin layer chromatography.Astaxanthin from Antarctic krill included three optical isomers: 3 S,3' S,3 S,3' R,and 3 R,3' R. The proportions of3 R,3'R in astaxanthin diesters and monoesters were more than 70%,and the proportion of 3 S,3' S was the lowest.The total astaxanthin content in Antarctic krill was 18. 9 mg/kg,of which the astaxanthin contents of astaxanthin diester,astaxanthin monoester,and free astaxanthin were 12. 0,6. 1 and 0. 8 mg/kg,respectively.
In this study,acetone was used to extract the astaxanthin from Antarctic krill. The silica gel column chromatography was optimised for separating astaxanthin diester,astaxanthin monoester,and free astaxanthin. A suitable developing solvent was selected. Samples were identified by a thin layer chromatography and the three-dimensional structures were confirmed by Chiralpak IC column. The contents of astaxanthin in different forms were quantified using C_(30)-HPLC. The results showed that a gradient elution through silica gel column chromatography with different ratios of acetone-petroleum ether could completely separate free astaxanthin,astaxanthin monoester and diester. V( nhexane) ∶ V( acetone) ∶ V( acetic acid) = 8∶ 2∶ 0. 2 was the optimum developing solvent for thin layer chromatography.Astaxanthin from Antarctic krill included three optical isomers: 3 S,3' S,3 S,3' R,and 3 R,3' R. The proportions of3 R,3'R in astaxanthin diesters and monoesters were more than 70%,and the proportion of 3 S,3' S was the lowest.The total astaxanthin content in Antarctic krill was 18. 9 mg/kg,of which the astaxanthin contents of astaxanthin diester,astaxanthin monoester,and free astaxanthin were 12. 0,6. 1 and 0. 8 mg/kg,respectively.
引文
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[7]张瑞莲.左旋、右旋及外消旋虾青素抗脂质过氧化活性差异的研究[D].广州:华南农业大学,2016:1-2.
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[12]张晓娜,惠伯棣,裴凌鹏,等.功能因子虾青素研究概况[J].中国食品添加剂,2017(8):208-214.
[13] MIAO Feng-ping,GENG Ya-hong,LU Da-yan,et al.Stability and changes in astaxanthin ester composition from Haematococcus pluvialis during storage[J]. Chinese Journal of Oceanology and Limnology,2013,31(6):1 181-1 189.
[14]REZANKA T,NEDBALOVAL,KOLOUCHOVAI,et al. LC-MS/APCI identification of glucoside esters and diesters of astaxanthin from the snow alga Chlamydomonas nivalis including their optical stereoisomers[J]. Phytochemistry,2013,88(1):34-42.
[15]杨楠,惠伯棣,宫平,等.雨生红球藻萃取物中虾青素及其衍生物的含量测定[J].中国食品添加剂,2016(11):195-203.
[16] JIANG Xi,YAN Qi-zhi,LIU Fu-he,et al. Chronic trans-astaxanthin treatment exerts antihyperalgesic effect and corrects co-morbid depressive like behaviors in mice with chronic pain[J]. Neuroscience Letters,2018,662(1):36-43.
[17]张沙沙.南极磷虾虾青素及虾青素酯分离研究[D].济南:山东师范大学,2015:13-30.
[18] MAOKA T,MASAAKI K,NOBUYUKI K,et al. Stereochemical Investigation of Carotenoids in the Antarctic krill Euphausia superba[J]. Bulletin of the Japanese Society of Scientific Fisheries,1985,51(10):1 671-1 673.
[19] YAMAGUCHI K,MIKI W,TORIU N,et al. The composition of carotenoid pigments in the antarctic krill euphausia superba[J]. Nippon Suisan Gakkaishi,1983,49(9):1 411-1 415.
[20]孙伟红,邢丽红,冷凯良,等.高效液相色谱法测定南极磷虾及其制品中虾青素的含量[J].食品安全质量检测学报,2017,8(4):1 248-1 253.
[21]杨澍,张婷,徐杰,等.高效液相色谱手性拆分法分析生物体内虾青素光学异构体[J].食品科学,2015,36(8):139-144.
[22]孙晓昀.南极磷虾虾青素酯的纯化与分析[D].济南:山东师范大学,2014:20-33.
[23] MORETTI V M,MENTASTI T,BELLAGAMBA F,et al. Determination of astaxanthin stereoisomers and colour attributes in flesh of rainbow trout(Oncorhynchus mykiss)as a tool to distinguish the dietary pigmentation source[J]. Food Additives and Contaminants,2006,23(11):1 056-1 063.
[2]刘丽,刘承初,赵勇,等.南极磷虾的营养保健功效以及食用安全性评价[J].食品科学,2010,31(17):443-447.
[3] CICERO A F G,COLLETTI A. Krill oil:Evidence of a new source of polyunsaturated fatty acids with high bioavailability[J]. Clinical Lipidology,2015,10(1):1-4.
[4] ULVEN S M,HOLVEN K B. Comparison of bioavailability of krill oil versus fish oil and health effect[J]. Vascular Health and Risk Management,2015,11(8):511-524.
[5]麦康森,魏玉婷,王嘉,等.南极磷虾的主要营养组成及其在水产饲料中的应用[J].中国海洋大学学报(自然科学版),2016,46(11):1-15.
[6]彭永健,吕红萍,王胜南,等.天然虾青素的研究进展[J].中国食品添加剂,2017(4):193-197.
[7]张瑞莲.左旋、右旋及外消旋虾青素抗脂质过氧化活性差异的研究[D].广州:华南农业大学,2016:1-2.
[8]周庆新,刘婷婷,杨鲁.虾青素的来源、生物功效及吸收代谢研究进展[J].食品研究与开发,2017,38(16):214-219.
[9]徐健.雨生红球藻中虾青素的提取及虾青素对皮肤细胞损伤的保护作用[D].杭州:浙江大学,2016:42-55.
[10]吕亭亭,刘海丽,冯晓慧,等.虾青素来源及生物活性的研究进展[J].中国食物与营养,2016,22(7):67-70.
[11] CAMPOIO T R,OLIVEIRA F A,OTTON R. Oxidative stress in human lymphocytes treated with fatty acid mixture:Role of carotenoid astaxanthin[J]. Toxicology in Vitro,2011,25(7):1 448-1 456.
[12]张晓娜,惠伯棣,裴凌鹏,等.功能因子虾青素研究概况[J].中国食品添加剂,2017(8):208-214.
[13] MIAO Feng-ping,GENG Ya-hong,LU Da-yan,et al.Stability and changes in astaxanthin ester composition from Haematococcus pluvialis during storage[J]. Chinese Journal of Oceanology and Limnology,2013,31(6):1 181-1 189.
[14]REZANKA T,NEDBALOVAL,KOLOUCHOVAI,et al. LC-MS/APCI identification of glucoside esters and diesters of astaxanthin from the snow alga Chlamydomonas nivalis including their optical stereoisomers[J]. Phytochemistry,2013,88(1):34-42.
[15]杨楠,惠伯棣,宫平,等.雨生红球藻萃取物中虾青素及其衍生物的含量测定[J].中国食品添加剂,2016(11):195-203.
[16] JIANG Xi,YAN Qi-zhi,LIU Fu-he,et al. Chronic trans-astaxanthin treatment exerts antihyperalgesic effect and corrects co-morbid depressive like behaviors in mice with chronic pain[J]. Neuroscience Letters,2018,662(1):36-43.
[17]张沙沙.南极磷虾虾青素及虾青素酯分离研究[D].济南:山东师范大学,2015:13-30.
[18] MAOKA T,MASAAKI K,NOBUYUKI K,et al. Stereochemical Investigation of Carotenoids in the Antarctic krill Euphausia superba[J]. Bulletin of the Japanese Society of Scientific Fisheries,1985,51(10):1 671-1 673.
[19] YAMAGUCHI K,MIKI W,TORIU N,et al. The composition of carotenoid pigments in the antarctic krill euphausia superba[J]. Nippon Suisan Gakkaishi,1983,49(9):1 411-1 415.
[20]孙伟红,邢丽红,冷凯良,等.高效液相色谱法测定南极磷虾及其制品中虾青素的含量[J].食品安全质量检测学报,2017,8(4):1 248-1 253.
[21]杨澍,张婷,徐杰,等.高效液相色谱手性拆分法分析生物体内虾青素光学异构体[J].食品科学,2015,36(8):139-144.
[22]孙晓昀.南极磷虾虾青素酯的纯化与分析[D].济南:山东师范大学,2014:20-33.
[23] MORETTI V M,MENTASTI T,BELLAGAMBA F,et al. Determination of astaxanthin stereoisomers and colour attributes in flesh of rainbow trout(Oncorhynchus mykiss)as a tool to distinguish the dietary pigmentation source[J]. Food Additives and Contaminants,2006,23(11):1 056-1 063.
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