基于在线HPLC-自由基清除检测-ESI-IT-TOF-MSn技术的蓝刺头抗氧化成分筛选及其构效关系研究

doi:10.5246/jcps.2021.04.022

中国药学(英文版) ›› 2021, Vol. 30 ›› Issue (4): 267-279.DOI: 10.5246/jcps.2021.04.022

• 【研究论文】 • 下一篇

基于在线HPLC-自由基清除检测-ESI-IT-TOF-MSn技术的蓝刺头抗氧化成分筛选及其构效关系研究

董馨¹, 王弘², 马飞祥¹, 高建萍¹, 陈世忠²^,^*(), 薛培凤¹^,^*()

1. 内蒙古医科大学药学院, 内蒙古呼和浩特 010110
2. 北京大学药学院天然药物学系, 北京 100191

收稿日期:2020-03-29 修回日期:2020-04-15 接受日期:2020-06-10 出版日期:2021-04-30 发布日期:2021-04-30
通讯作者: 陈世忠, 薛培凤
作者简介:
+ Tel.: +86-10-82802723, E-mail: chenshizhong66@163.com
+ Tel.: +86-471-6653146, E-mail: Xpfdc153@163.com
基金资助:
National Natural Science Foundation of China (Grant No. 81860756, 81960758), Natural Science Foundation of Inner Mongolia Autonomous Region (Grant No. 2017MS08122, 2019MS08111 and 2019MS08119), Inner Mongolia Science and Technology Innovation Guide Project (Grant No. 02039001), Rolling Support Plan for Grassland Talents Project in Inner Mongolia Autonomous Region, Inner Mongolia Autonomous Region Higher Education Science Research Project (Grant No. NJZY19099).

A strategy for structure-activity relationship study on antioxidants in Echinops latifolius Tausch extracts by online HPLC-radical scavenging detection coupled with ESI-IT-TOF-MSn

Xin Dong¹, Hong Wang², Feixiang Ma¹, Jianping Gao¹, Shizhong Chen²^,^*(), Peifeng Xue¹^,^*()

1 Department of Pharmacy, Inner Mongolia Medical University, Jinshan Development Zone, Hohhot 010110, China
2 Department of Natural Medicines, School of Pharmaceutical Sciences, Peking University Health Science Center, Beijing 100191, China

Received:2020-03-29 Revised:2020-04-15 Accepted:2020-06-10 Online:2021-04-30 Published:2021-04-30
Contact: Shizhong Chen, Peifeng Xue

摘要/Abstract

摘要：

蓝刺头是治疗骨质疏松的传统蒙药, 其所含的有效成分不明确是限制其现代化、国际化发展的关键因素。传统活性物质筛选策略费时费力, 本研究采用在线HPLC-自由基清除检测-ESI-IT-TOF-MSn技术对蓝刺头中有效成分进行筛选, 实现分离、鉴定、活性筛选一体化。此外, 该技术还有助于阐明活性成分间的构效关系。由于骨质疏松发病过程与氧化应激有关, 因此本研究主要针对蓝刺头中抗氧化成分进行筛选, 通过DPPH等效法确定其中抗氧化活性最高的成分。该策略不仅能够筛选、鉴定出蓝刺头中抗氧化活性成分, 还能够评价这些成分的构效关系。最终, 蓝刺头中20个成分 (包括芹菜素葡萄糖苷、咖啡酸、双咖啡酰奎尼酸、双咖啡酰奎尼酸甲酯等)被准确鉴定, 其中18种成分具有抗氧化活性。通过研究14种蓝刺头中分离所得的化合物发现各物质氧化活性与以下结构特点有关: (1) 有酚羟基的成分具有抗氧化活性; (2) 酚羟基的数目并不能影响抗氧化活性的强弱, 但咖啡酰基的数目会影响抗氧化剂的活性; (3) 咖啡酰基在奎宁酸上的取代位置对DPPH活性有较大影响; (4) 甲氧基可降低其抗氧化活性。综上所述, 该研究以生化视角将蓝刺头中的活性化合物与其抗骨质疏松作用联系起来, 为进一步阐释蓝刺头如何在骨质疏松症中发挥作用提供依据。

关键词: 蓝刺头, 抗氧化剂, 构效关系, 在线HPLC-自由基清除检测, ESI-IT-TOF-MSn

Abstract:

Echinops latifolius Tausch (ELT) is the traditional Mongolian medicine for the treatment of osteoporosis, and the ambiguous composition of active ingredients is an important factor in restricting the modernization and globalization of this herb. Considering the traditional activity screening strategy is time-consuming and labor intensive, online HPLC active ingredient detection coupled with ESI-IT-TOF-MSⁿ strategy was employed in this study to isolate, identify and screen active compounds from the herbal medicines at the same time. The structure-activity relationship of these compounds was elucidated as well. Owing to the association of osteoporosis progression and oxidative stress, the antioxidants screening from ELT could be a good interpretive of the active substance in this herb. Meanwhile, DPPH equivalent method was an indicative of the most powerful antioxidant in ELT. Consequently, the screening and identification of the antioxidants in ELT was performed by using on-line HPLC-radical scavenging detection coupled with ESI-IT-TOF-MSⁿ strategy, and the structure-activity relationship was investigated based on DPPH equivalent method. Finally, 20 constituents (including apigenin glucosides, caffeic acid, biscaffeoylquinic acids, biscaffeoylquinic acid methyl esters, ect.) were characterized in ELT extracts, and 18 components showed appreciable radical scavenging capacity. In addition, the structure-activity relationship study was carried out based on 14 compounds isolated from our laboratory, and the structural requirements of the compounds on antioxidant activity were obtained: (1) compounds with phenolic hydroxyl groups could have antioxidant activity; (2) the antioxidant activity could not be facilitated by the number of hydroxyl groups, but affected by the number of caffeoyl groups; (3) the substitution position of caffeoyl on quinic acid had a greater influence on DPPH activity; (4) methoxy groups could reduce the antioxidant activity. Collectively, this work provided the biochemical perspective to link active compounds and anti-osteoporosis action of ELT, and further explained how ELT worked in osteoporosis patients with bone loss.

Key words: Echinops latifolius Tausch, Antioxidants, Structure-activity relationship, Online HPLC-radical scavenging detection, ESI-IT-TOF-MSn

Supporting:

董馨, 王弘, 马飞祥, 高建萍, 陈世忠, 薛培凤. 基于在线HPLC-自由基清除检测-ESI-IT-TOF-MSn技术的蓝刺头抗氧化成分筛选及其构效关系研究[J]. 中国药学(英文版), 2021, 30(4): 267-279.

Xin Dong, Hong Wang, Feixiang Ma, Jianping Gao, Shizhong Chen, Peifeng Xue. A strategy for structure-activity relationship study on antioxidants in Echinops latifolius Tausch extracts by online HPLC-radical scavenging detection coupled with ESI-IT-TOF-MSn[J]. Journal of Chinese Pharmaceutical Sciences, 2021, 30(4): 267-279.

图/表 6

Table 1. Identified the compounds in ELT extracts (negative ion mode).

Figure 1. The chemical structures of major compounds in ELT extracts.

Figure 2. The chromatograms of ELT extracts by HPLC-ESI-IT-TOF-MSn-PDA after treated with methanol.

Figure 3. The chromatograms of ELT extracts by HPLC-ESI-IT-TOF-MSn-PDA after treated with DPPH solutions.

Table 2. The free radical scavenging ability of 14 compounds isolated from our laboratory in ELT extract.

Figure 4. Structure-activity relationships of compounds from ELT extracts.

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基于在线HPLC-自由基清除检测-ESI-IT-TOF-MSn技术的蓝刺头抗氧化成分筛选及其构效关系研究

A strategy for structure-activity relationship study on antioxidants in Echinops latifolius Tausch extracts by online HPLC-radical scavenging detection coupled with ESI-IT-TOF-MSn

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