Kim Sang Min, Kang Suk Woo, Jeon Je-Seung, Jung Yu-Jin, Kim Chul Young, Pan Cheol Ho, Um Byung-Hun
Functional Food Center, KIST Gangneung Institute, Gangwon 210-340, Republic of Korea.
Biomed Chromatogr. 2012 Feb;26(2):199-207. doi: 10.1002/bmc.1646. Epub 2011 Jun 8.
Gradient HPLC coupled to Diode Array Detector (DAD), MS/MS and NMR was applied to the rapid structure determination of major compounds of methanol extracts from leaves and roots of Petasites japonicus. The relative antioxidant capacities of the compounds were evaluated by an HPLC system with post-column on-line antioxidant detection based on 2,2'-azinobis-3-ethylbenzothiazoline-6-sulfonic acid radical scavenging. Six compounds were successfully separated on a reverse-phase C(18) column and were identified as 5-caffeoylquinic acid (5-CQA), fukinolic acid (FA), 3,5-di-O-caffeoylquinic acid (3,5-DCQA), quercetin-3-O-(6″-acetyl)-β-glucopyranoside (QAG), 4,5-di-O-caffeoylquinic acid (4,5-DCQA) and kaempferol-3-O-(6″-acetyl)-β-glucopyranoside (KAG) by MS/MS and (1)H NMR data. Among these compounds, those containing a caffeoyl moiety (5-CQA, FA, 3,5- and 4,5-DCQA) showed relatively strong radical scavenging capacity, with 3,5-DCQA having the greatest radical scavenging capacity in leaf (23.09% of total antioxidant capacity) and root (26.47%) extracts. The relative radical scavenging portion of QAG was only 3.41% in the leaves and KAG did not show any radical scavenging activity. These results demonstrate that the hyphenated HPLC techniques can be successfully applied to rapidly identify structures and evaluate antioxidant activities without prior purification of compounds from plant tissues of P. japonicus.
采用梯度高效液相色谱法结合二极管阵列检测器（DAD）、串联质谱（MS/MS）和核磁共振（NMR）对蜂斗菜叶片和根部甲醇提取物中的主要化合物进行快速结构鉴定。基于2,2'-偶氮二（3-乙基苯并噻唑啉-6-磺酸）自由基清除能力，通过柱后在线抗氧化检测的高效液相色谱系统评估了这些化合物的相对抗氧化能力。六种化合物在反相C(18)柱上成功分离，并通过MS/MS和(1)H NMR数据鉴定为5-咖啡酰奎尼酸（5-CQA）、蜂斗菜酸（FA）、3,5-二-O-咖啡酰奎尼酸（3,5-DCQA）、槲皮素-3-O-(6″-乙酰基)-β-葡萄糖苷（QAG）、4,5-二-O-咖啡酰奎尼酸（4,5-DCQA）和山奈酚-3-O-(6″-乙酰基)-β-葡萄糖苷（KAG）。在这些化合物中，含有咖啡酰部分的化合物（5-CQA、FA、3,5-和4,5-DCQA）表现出相对较强的自由基清除能力，其中3,5-DCQA在叶片提取物（占总抗氧化能力的23.09%）和根部提取物（占26.47%）中具有最大的自由基清除能力。QAG在叶片中的相对自由基清除比例仅为3.41%，而KAG未表现出任何自由基清除活性。这些结果表明，联用的高效液相色谱技术可成功应用于快速鉴定结构并评估抗氧化活性，而无需事先从蜂斗菜植物组织中纯化化合物。
