Two-dimensional countercurrent chromatography x high performance liquid chromatography for preparative isolation of toad venom
Two-dimensional countercurrent chromatography x high performance liquid chromatography for preparative isolation of toad venom
复制标题
二维逆流色谱×高效液相色谱制备蟾蜍毒液
DOI：
10.1016/j.chroma.2014.01.029
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发表时间：
2014-02-28
影响因子：
4.1
通讯作者：
Zhao, Yu-Fen
中科院分区：
文献类型：
--
作者：
Qiu, Ying-Kun;Yan, Xia;Zhao, Yu-Fen
在这项工作中，开发了一种新型在线二维色谱联用程序逆流色谱和高效液相色谱（2D CCC x HPLC），用于复杂天然产物的制备分离。 CCC 柱用作第一维分离，以反相 (RP) 模式操作的制备型 ODS 柱作为第二维。 CCC 在受控流速下运行，以确保每个级分在一小时内洗脱，对应于二维制备型 HPLC 的分离时间。来自第一维 CCC 的洗脱液使用补充泵稀释并捕获到保持柱上，然后洗脱并转移到第二维 HPLC。从柱尺寸、稀释比、径高比以及系统压力等方面对保留柱的性能进行了评估，以解决 CCC 柱低压洗脱液在线捕集的问题。使用市售的 15 mm x 30 mm i.d. 可以获得令人满意的捕获效率和可承受的 CCC 压力。 ODS 预柱。该集成系统成功应用于从中华大蟾蜍毒液甲醇粗提取物中一步制备分离 12 种化合物。化合物1-12的分离总收率为1.0%、0.8%、2.0%、13%、2.0%、1.5%、1.9%、3.6%、6.1%、4.8%、3.5%和4.1%，HPLC纯度为99.9%、99.7%、90.6%、99.9%，分别为 77.0%、99.9%、90.4%、99.9%、52.0%、99.9%、99.3% 和 85.0%。所有结果表明，流程编程 CCC x HPLC 方法是一种高效、便捷的蟾蜍毒液化合物分离方法，也可用于分离其他复杂的多组分天然产物。 (C) 2014 Elsevier B.V. 保留所有权利。
In this work, a new on-line two-dimensional chromatography coupling of flow programming countercurrent chromatography and high-performance liquid chromatography (2D CCC x HPLC) was developed for preparative separation of complicated natural products. The CCC column was used as the first dimensional isolation and a preparative ODS column operated in reversed-phase (RP) mode as the second dimension. The CCC was operated at a controlled flow rate to ensure that each fraction eluted within one hour, corresponding to the isolation time of the 2nd dimensional preparative HPLC. The eluent from the 1st dimensional CCC was diluted using a makeup pump and trapped onto holding column, before been eluted and transferred to the 2nd dimensional HPLC. The performance of the holding column was evaluated, in terms of column size, dilution ratio and diameter-height ratio, as well as system pressure, for the solution to the issue of online trapping of low pressure eluent from a CCC column. Satisfactory trapping efficiency and tolerable CCC pressure can be achieved using a commercially available 15 mm x 30 mm i.d. ODS pre-column. The present integrated system was successfully applied in a one-step preparative separation of 12 compounds, from the crude methanol extract of venom of Bufo bufo gargarizans. Compounds 1-12 were isolated in overall yield of 1.0%, 0.8%, 2.0%, 13%, 2.0%, 1.5%, 1.9%, 3.6%, 6.1%, 4.8%, 3.5% and 4.1%, with HPLC purity of 99.9%, 99.7%, 90.6%, 99.9%, 77.0%, 99.9%, 90.4%, 99.9%, 52.0%, 99.9%, 99.3%, and 85.0%, respectively. All the results demonstrate that the flow programming CCC x HPLC method is an efficient and convenient way for the separation of compounds from toad venom and it can also be applied to isolate other complex multi-component natural products. (C) 2014 Elsevier B.V. All rights reserved.