方格星虫低聚肽的分离、鉴定及其促骨发育作用评价

刘珍 唐晓宁 吴科锋 林芊杏 刘欢 李茜 张茜 梅思 戚怡

刘珍,唐晓宁,吴科锋,等. 方格星虫低聚肽的分离、鉴定及其促骨发育作用评价[J]. 食品工业科技,2021,42(20):334−341. doi:  10.13386/j.issn1002-0306.2020120263
引用本文: 刘珍,唐晓宁,吴科锋,等. 方格星虫低聚肽的分离、鉴定及其促骨发育作用评价[J]. 食品工业科技,2021,42(20):334−341. doi:  10.13386/j.issn1002-0306.2020120263
LIU Zhen, TANG Xiaoning, WU Kefeng, et al. Isolation and Identification of Oligopeptides from Squares Siphon Worm and Its Effect on Promoting Bone Growth[J]. Science and Technology of Food Industry, 2021, 42(20): 334−341. (in Chinese with English abstract). doi:  10.13386/j.issn1002-0306.2020120263
Citation: LIU Zhen, TANG Xiaoning, WU Kefeng, et al. Isolation and Identification of Oligopeptides from Squares Siphon Worm and Its Effect on Promoting Bone Growth[J]. Science and Technology of Food Industry, 2021, 42(20): 334−341. (in Chinese with English abstract). doi:  10.13386/j.issn1002-0306.2020120263

方格星虫低聚肽的分离、鉴定及其促骨发育作用评价

doi: 10.13386/j.issn1002-0306.2020120263
基金项目: 广东省自然科学基金资助项目(2018A030307001);湛江市财政资金科技专项竞争性分配项目(2019A01005,2019A03023);南海海洋生物医药资源研发公共服务平台开放课题资助项目(XM-202012-01A)
详细信息
    作者简介:

    刘珍(1995−),女,硕士研究生,研究方向:药理学,Email:958487385@qq.com

    通讯作者:

    戚怡(1975−),女,博士,助理研究员,研究方向:药理学,Email: 173312453@qq.com

  • 中图分类号: TS201.4

Isolation and Identification of Oligopeptides from Squares Siphon Worm and Its Effect on Promoting Bone Growth

  • 摘要: 目的:从方格星虫中分离和筛选可促进骨形成的生物活性低聚肽。方法:通过酶解法和膜分离法制备寡肽,并通过HPLC-MS/MS鉴定肽段序列。通过在线数据库预测药物形成,并通过斑马鱼模型评估促进骨形成的作用。结果:共鉴定出10种低分子量肽,通过斑马鱼骨发育模型获得了两种潜在的候选肽,其氨基酸序列分别为GFAGDDAPR和GLGGLSPEK;分别命名为GSP1和GSP2;其分子量分别为905.04和857.06 Da;与空白对照组相比,1~50 μg/mL GSP1溶液浓度处理的AB斑马鱼幼鱼颅骨积分光密度和绿色荧光的面积均有所增加,GSP1浓度在5 μg/mL时促进骨发育的效果最好(P<0.05)。1~50 μg/mL GSP2溶液浓度处理的AB型斑马鱼幼鱼颅骨积分光密度和绿色荧光的面积均有所增加,且随着浓度的增加而增大,GSP2浓度在50 μg/mL时促进骨发育的效果最好(P<0.05)。结论:小于1000 Da的寡肽有望成为促进骨形成的候选药物。
  • 图  1  LC-MS/MS分析方格星虫短肽中总离子流色谱图

    Figure  1.  LC-MS/MS analysis of total ion current chromatogram in short peptides of square siphon worm

    图  2  斑马鱼的钙黄绿素荧光染料染色结果和统计分析(10×)

    Figure  2.  Results of zebrafish stained with calcein fluorescent dye and statistical analysis(10×)

    图  3  斑马鱼的钙黄绿素荧光染料染色结果和统计分析(10×)

    Figure  3.  GSP2 concentration group of zebrafish stained with calcein fluorescent dye and statistic analysis(10×)

    图  4  方格星虫提取物多肽GSP1的高效液相色谱图和结构式

    Figure  4.  High performance liquid chromatography and structural formula of square siphon worm polypeptide GSP1

    图  5  方格星虫提取物多肽GSP2的高效液相色谱图和结构式

    Figure  5.  High performance liquid chromatography and structural formula of square siphon worm polypeptide GSP2

    图  6  GSP1质谱图

    Figure  6.  Mass spectrum of GSP1

    图  7  GSP2质谱图

    Figure  7.  Mass spectrum of GSP2

    表  1  方格星虫酶解短肽的LC-MS/MS相对定量结果

    Table  1.   LC-MS/MS relative quantitative results of enzymatic hydrolysis of short peptides hydrolyzed by squares siphon worm

    数字序列长度质量蛋白序号强度
    1DALDELYLEIMME131583.7099R7TSN252954000
    2FPSLVGRP8871.49159R7TR8912021000
    3GFAGDDAPR9904.40389A0A0K1R048; R7TPD2; Q4AC20; R7TMG3; D3YN2324078000
    4GLGGLSPEK9856.46543R7URV642424000
    5LTEAPLNPK9981.5495A0A0K1R048; R7TPD2;Q4AC20;R7TMG3; D3YN23; D3UA35; R7VCX4; R7U3X61666200000
    6PAVKKPAP8806.50142R7VFK06592900
    7SILSPSPR8855.48142R7UJQ710448000
    8TEAPLNPK8868.46543A0A0K1R048; R7TPD2; Q4AC20; R7TMG3; D3YN23; D3UA35;
    R7VCX4; R7U3X6
    0
    9VAPEEHPV8876.43413A0A0K1R048; R7TPD2; Q4AC20; R7TMG3; D3YN23; D3UA35; R7VCX4251970000
    10VVTTPKSG8787.44397R7U0B47325900
    下载: 导出CSV

    表  2  方格星虫酶解短肽筛选结果

    Table  2.   Screening results of short peptides hydrolyzed by squares siphon worm

    序号序列活性水溶性
    1DALDELYLEIMME0.531
    2FPSLVGRP0.777121
    3GFAGDDAPR0.598852
    4GLGGLSPEK0.244678
    5LTEAPLNPK0.20058
    6PAVKKPAP0.278084
    7SILSPSPR0.431578
    8TEAPLNPK0.220992
    9VAPEEHPV0.12208
    10VVTTPKSG0.0732613
    下载: 导出CSV
  • [1] 李娇, 程海涛, 张秀霞, 等. 光裸方格星虫的研究进展[J]. 河北渔业,2020(6):46−51. [Li Jiao, Cheng Haitao, Zhang Xiuxia, et al. Progress in the study of the Saccharomyces glabrifolia[J]. Hebei Fisheries,2020(6):46−51. doi:  10.3969/j.issn.1004-6755.2020.06.013
    [2] 陈伟耀, 李金兰, 陈伟寿, 等. 光祼星虫营养成分和药理作用的研究进展[J]. 安徽农业科学,2018,46(12):20−22. [Chen Weiyao, Li Jinlan, Chen Weishou, et al. Research progress on nutritional components and pharmacological effects of naked starworm[J]. Journal of Anhui Agricultural Sciences,2018,46(12):20−22. doi:  10.3969/j.issn.0517-6611.2018.12.007
    [3] 李映新, 李肖肖, 黄媛恒, 等. 光裸方格星虫纤溶酶SNFE体外溶栓作用及安全性初评[J]. 天然产物研究与开发,2016,Magi 28(11):1789−1792. [Li Yingxin, Li Xiaoxiao, Huang Yuanheng, et al. Preliminary evaluation of thrombolytic effect and safety of fibrinolytic enzyme SNFE in vitro[J]. Research and Development of Natural Products,2016,Magi 28(11):1789−1792.
    [4] 徐艳, 王慧芹, 邹杰, 等. 方格星虫生物活性成分的研究进展[J]. 广西科学院学报,2015,31(4):273−280. [Xu Yan, Wang Huiqin, Zou Jie, et al. Research progress on bioactive components of Alternaria alternata[J]. Journal of Guangxi Academy of Sciences,2015,31(4):273−280. doi:  10.3969/j.issn.1002-7378.2015.04.009
    [5] 陈文, 王湘君, 王玉杰, 等. 方格星虫氨基酸含量测定及其营养价值评定[J]. 保鲜与加工,2018,18(4):108−113. [Chen Wen, Wang Xiangjun, Wang Yujie, et al. Determination of amino acid content and evaluation of its nutritional value[J]. Fresh-keeping and Processing,2018,18(4):108−113. doi:  10.3969/j.issn.1009-6221.2018.04.020
    [6] Zhang Q, Dong L, Tong T, et al. Polysaccharides in Sipunculus nudus: Extraction condition optimization and antioxidant activities[J]. Journal of Ocean University of China,2017,16(1):74−80. doi:  10.1007/s11802-017-2969-5
    [7] 马伏宁, 谭琳, 郭刚, 等. 海南方格星虫纤溶酶分离纯化及部分基因克隆[J]. 基因组学与应用生物学,2018,37(2):698−703. [Ma Funing, Tan Lin, Guo Gang, et al. Isolation, purification and partial gene cloning of fibrinolytic enzyme from S. hainanensis[J]. Genomics and Applied Biology,2018,37(2):698−703.
    [8] 徐艳, 谢一兴, 邹杰, 等. 方格星虫提取物的抗氧化活性及成分分析[J]. 食品工业科技,2017,38(21):7−11. [Xu Yan, Xie Yixing, Zou Jie, et al. Antioxidant activity and composition analysis of the extract of Artemisia punctata[J]. Food Industry Science and Technology,2017,38(21):7−11.
    [9] Ge Y H, Chen Y Y, Zhou G S, et al. A novel antithrombotic protease from marine worm Sipunculus nudus[J]. International Journal of Molecular Sciences,2018,19(10):3023. doi:  10.3390/ijms19103023
    [10] 高春丽, 石特, 张俊玲, 等. 方格星虫多糖对人肠上皮细胞辐射损伤的保护作用[J]. 辐射研究与辐射工艺学报,2019,37(3):24−32. [Gao Chunli, Shi Te, Zhang Junling, et al. Protective effects of polysaccharides from Alternaria alternata on radiation injury of human intestinal epithelial cells[J]. Journal of Radiation Research and radiation Technology,2019,37(3):24−32.
    [11] 陈永祥, 吕萍. 方格星虫药理学活性的研究概况[J]. 中西医结合心血管病电子杂志,2017,5(2):58−59. [Chen Yongxiang, Lv Ping. A survey of studies on the pharmacological activity of Chrysoptera[J]. Electronic Journal of Cardiovascular Diseases with Integrated Traditional Chinese and Western Medicine,2017,5(2):58−59. doi:  10.3969/j.issn.2095-6681.2017.02.042
    [12] 何颖, 沈先荣, 莫琳芳, 等. 方格星虫多糖对低剂量中子辐射损伤大鼠的保护作用[J]. 中国海洋药物,2014,33(3):49−55. [He Ying, Shen Xianrong, Mo Linfang, et al. Protective effects of polysaccharides from Alternaria alternata on rats injured by low dose neutron radiation[J]. China Marine Pharmaceuticals,2014,33(3):49−55.
    [13] 郑志鸿, 章超桦, 林海生, 等. 方格星虫水提物促小鼠皮肤创伤愈合及作用机制的研究[J]. 生物医学工程学杂志,2020,37(3):460−468. [Zheng Zhihong, Zhang Chaohua, Lin Haisheng, et al. Study on the effect and mechanism of aqueous extract of Chrysophora tenuiformis on skin wound healing in mice[J]. Journal of Biomedical Engineering,2020,37(3):460−468.
    [14] 董兰芳, 张琴, 童潼, 等. 方格星虫多糖抗菌和抗氧化活性研究[J]. 广西科学,2013,20(4):289−293. [Dong Lanfang, Zhang Qin, Tong Tong, et al. Study on antibacterial and antioxidant activity of polysaccharides from Alternaria alternata[J]. Guangxi Science,2013,20(4):289−293.
    [15] 梁伟乔, 钟诚, 李宇明. 骨质疏松症的中医病因病机认识与治疗进展[J]. 中国骨质疏松杂志,2020,26(1):135−139. [Liang Weiqiao, Zhong Cheng, Li Yuming. Progress in TCM etiology, pathogenesis and treatment of osteoporosis[J]. Chinese Journal of Osteoporosis,2020,26(1):135−139. doi:  10.3969/j.issn.1006-7108.2020.01.028
    [16] 李云贺, 顾昊, 孙君雷, 等. 上海市莘庄社区中老年人体质量指数与常见慢性病关系研究[J]. 蚌埠医学院学报,2015,40(4):503−506. [Li Yunhe, Gu Hao, Sun Junlei, et al. Study on the relationship between body mass index and common chronic diseases in middle-aged and elderly in Xinzhuang community of Shanghai[J]. Journal of Bengbu Medical College,2015,40(4):503−506.
    [17] Yongkui W, Jian C, Jing C, et al. Daphnetin ameliorates glucocorticoid-induced osteoporosis via activation of Wnt/GSK-3β/β-catenin signaling[J]. Toxicology and Applied Pharmacology,2020,409:115333.
    [18] 吴忠, 易迟. 肾骨颗粒联合复方骨肽注射液治疗骨质疏松的临床疗效及其对腰椎功能的影响[J]. 临床合理用药杂志,2020,r13(33):104−106. [Wu Zhong, Yi Chi. Clinical efficacy of Shengu granule combined with compound osteopeptide injection in the treatment of osteoporosis and its effect on lumbar function[J]. Journal of Rational Clinical Drug Use,2020,r13(33):104−106.
    [19] Li X, Ettore B, Pan W, et al. Separating effects of bone-quality changes at multiple scales in steroid-induced osteoporosis: Combining multiscale experimental and modelling approaches[J]. Mechanics of Materials,2021:157.
    [20] 沈先荣, 蒋定文, 贾福星, 等. 方格星虫延缓衰老作用研究[J]. 中国海洋药物,2004(1):30−32. [Shen Xianrong, Jiang Dingwen, Jia Fuxing, et al. Study on the anti-aging effect of chestnut[J]. Chinese Marine Medicine,2004(1):30−32. doi:  10.3969/j.issn.1002-3461.2004.01.008
    [21] Carnovali M, Luzi L, Terruzzi I, et al. Metabolic and bone effects of high-fat diet in adult zebrafish[J]. Endocrine,2018,61(2):317−326. doi:  10.1007/s12020-017-1494-z
    [22] 许冰洁, 张立将, 李春启, 等. 斑马鱼胚胎评价5种药物的发育毒性与模型验证[J]. 中国药理学通报,2016,32(1):74−79. [Xu Bingjie, Zhang Lijiang, Li Chunqi, et al. Evaluation of developmental toxicity of 5 drugs by zebrafish embryos and model verification[J]. China Pharmacology Bulletin,2016,32(1):74−79. doi:  10.3969/j.issn.1001-1978.2016.01.016
    [23] 邸亚男, 朱丽英, 钱雯, 等. 斑马鱼作为模式动物在人类眼睛疾病研究中的应用[J]. 实验动物与比较医学,2020,40(5):440−448. [Di Yanan, Zhu Liying, Qian Wen, et al. Application of zebrafish as a model animal in the study of human eye diseases[J]. Laboratory Animals and Comparative Medicine,2020,40(5):440−448. doi:  10.3969/j.issn.1674-5817.2020.05.014
    [24] 刘艺, 王瑞昕, 游龙泰, 等. 芦荟大黄素诱导斑马鱼肝毒性及作用机制研究[J]. 环球中医药,2020,13(1):18−22. [Liu Yi, Wang Ruixin, You Longtai, et al. Study on zebrafish hepatotoxicity induced by aloe-emodin and its mechanism[J]. Global Traditional Chinese Medicine,2020,13(1):18−22. doi:  10.3969/j.issn.1674-1749.2020.01.004
    [25] Luo S, Chen J, Zhong Z, et al. Salvianolic acid B stimulates osteogenesis in dexamethasone-treated zebrafish larvae[J]. Acta Pharmacologica Sinica,2016,37(1):1370−1380.
    [26] Son Y, Lee E M, Lee D Y, et al. Longan fruit increase bone mineral density in zebrafish and ovariectomized rat by suppressing RANKL-induced osteoclast differentiation[J]. Phytomedicine,2019:59.
    [27] 白景文, 杨海贤. 人皮质骨矿化基质中骨盐框架结构[J]. 中华骨科杂志,1997(2):53−55. [Bai Jingwen, Yang Haixian. Bone salt frame structure in human cortical bone mineralized matrix[J]. Chinese Journal of Orthopaedics,1997(2):53−55.
    [28] Xi Y, Chen D, Sun L, et al. Characterization of zebrafish mutants with defects in bone calcification during development[J]. Biochemical & Biophysical Research Communications,2013,440(1):132−136.
    [29] Yuehua Y, Yuan S, Wei-wei M, et al. Oxidative stress induces downregulation of TP53INP2 and suppresses osteogenic differentiation of BMSCs during osteoporosis through the autophagy degradation pathway[J]. Free Radical Biology and Medicine,2021:166.
    [30] Qi H, Junzheng Y, Gangyu Z, et al. Sanhuang Jiangtang tablet protects type 2 diabetes osteoporosis via AKT-GSK3β-NFATc1 signaling pathway by integrating bioinformatics analysis and experimental validation[J]. Journal of Ethnopharmacology,2021:273.
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  • 收稿日期:  2020-12-30
  • 网络出版日期:  2021-08-30
  • 刊出日期:  2021-10-11

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