CN112305139A - 一种增强氨基柱稳定性的方法 - Google Patents

一种增强氨基柱稳定性的方法 Download PDF

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CN112305139A
CN112305139A CN201910687458.1A CN201910687458A CN112305139A CN 112305139 A CN112305139 A CN 112305139A CN 201910687458 A CN201910687458 A CN 201910687458A CN 112305139 A CN112305139 A CN 112305139A
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梁鑫淼
夏东海
郭志谋
金高娃
闫竞宇
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Dalian Institute of Chemical Physics of CAS
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    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
    • G01N30/02—Column chromatography
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    • G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
    • G01N30/02—Column chromatography
    • G01N30/26—Conditioning of the fluid carrier; Flow patterns
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    • G01N30/26—Conditioning of the fluid carrier; Flow patterns
    • G01N30/28—Control of physical parameters of the fluid carrier
    • G01N30/32—Control of physical parameters of the fluid carrier of pressure or speed
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    • G01N30/88—Integrated analysis systems specially adapted therefor, not covered by a single one of the groups G01N30/04 - G01N30/86
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Abstract

本发明提供了一种增强氨基柱稳定性的方法，能够明显增强氨基柱的使用稳定性和重复性。本发明中先用磷酸盐/甲醇溶液作为流动相冲洗氨基柱，再用水/甲醇溶液作为流动相冲洗氨基柱，使柱表面得以稳定，从而获得良好的使用稳定性。本发明增强氨基柱稳定性的方法可使基于氨基柱的分离分析获得良好的重现性、降低使用蒸发光散射检测器时的基线噪音并提供更好的分离选择性，对氨基柱的应用具有十分重要的价值，同时也是色谱研究的一个重要方向。

Description

一种增强氨基柱稳定性的方法
技术领域
本发明属于分析化学领域，涉及一种增强氨基柱稳定性的方法。
技术背景
氨基柱被广泛用于色谱分离分析当中，如糖和某些极性药物、药物辅料等。但是氨基柱存在稳定性和重现性差、寿命短等问题。虽然氨基柱存在上述弊端，而且新型HILIC固定相不断推陈出新，但氨基柱因其独特的选择性而无法被完全替代。因此，增强氨基柱的稳定性对其应用具有十分重要的价值，同时也是色谱研究的一个重要方向。
目前，增强氨基柱稳定性的方法主要从两方面入手。其一是在填料制备工艺上进行改进，如TOSOH公司的TSKgel NH2-100和Welch公司的Ultimate XB-NH2，但是这两种氨基柱在对山梨醇和甘露醇的ELSD分析检测中仍具有较高的基线噪音和较差的分离选择性，而这种差的分离分析结果则源自于氨基柱的不稳定性。其二是采用合适的流动相对色谱柱进行冲洗，但是采用欧洲药典中异丙醇冲洗的方法不仅耗时(低流速下连续冲洗10h)，而且在对山梨醇和甘露醇的ELSD分析检测中仍无法获得满意的结果。因此，为加速和拓展氨基柱的应用，促进其色谱研究及相关领域的发展，需要开发一种能够有效增强氨基柱稳定性的方法。
发明内容
本发明的目的在于提供一种增强氨基柱稳定性的方法。
本发明的目的通过以下技术方案实现：首先，配制一定物质的量浓度的磷酸盐溶液；然后，将该磷酸盐溶液与甲醇以一定的体积比混合作为流动相冲洗氨基柱；最后，采用一定体积比的水/甲醇溶液作为流动相冲洗氨基柱。
所述氨基柱是以硅胶为基质，键合有氨基官能团的色谱固定相。
所述磷酸盐为磷酸钠、磷酸氢二钠、磷酸二氢钠、磷酸钾、磷酸氢二钾、磷酸二氢钾中的一种或二种以上。
所述磷酸盐溶液的物质的量的浓度为0.01～100mM。
所述磷酸盐/甲醇溶液为磷酸盐溶液与甲醇按照体积比为1/99～99/1配制而成。
所述磷酸盐/甲醇溶液作为流动相冲洗氨基柱的流速为0.01～100mL/min，冲洗1～100个柱体积。
所述水/甲醇溶液为水和甲醇按照体积比为1/99～99/1配制而成。
所述水/甲醇溶液作为流动相冲洗氨基柱的流速为0.01～100mL/min，冲洗1～100个柱体积。
实验证明，本发明能够快速、有效增强氨基柱的使用稳定性，可使基于氨基柱的分离分析获得良好的重现性、降低使用蒸发光散射检测器时的基线噪音并提供更好的分离选择性，对氨基柱的应用及其在色谱领域的发展将起到良好的促进作用。
附图说明
图1为本发明实施例1所述的采用本发明前后R5-NH2柱(4.6×150mm)的重现性对比图。
图2为本发明实施例2所述的采用本发明前后R5-NH2柱(4.6×150mm)的分离分析效果对比图。
图3为本发明实施例3所述的采用本发明前后其他不同品牌氨基柱(4.6×150mm)的分离分析效果对比图。
具体实施方式
以下结合具体实施例进一步说明本发明，应理解，具体实施方式仅用于说明本发明，而非限制本发明。
实施例1
首先，配制20mM磷酸盐溶液(包括磷酸二氢钠10mM、磷酸氢二钠10mM)；然后，将该磷酸盐溶液与甲醇等体积混合作为流动相冲洗R5-NH2柱，流速0.5mL/min冲洗18个柱体积；最后，采用水/甲醇溶液以水与甲醇体积比1/1作为流动相冲洗R5-NH2柱，流速0.5mL/min冲洗6个柱体积。
以尿嘧啶、尿苷为分析对象，采用本发明方法对R5-NH2柱进行冲洗前后R5-NH2柱(4.6×150mm)分析尿嘧啶、尿苷重现性对比如图1所示。
由图1可知，未采用本发明的方法冲洗R5-NH2柱之前，R5-NH2柱连续进样5次重现性很差；但采用本发明冲洗后，连续进样5次重现性很好。上述结果表明，本发明能够快速、有效增强氨基柱的使用稳定性，可使基于氨基柱的分离分析获得良好的重现性。
实施例2
与实施例1不同之处在于，使用磷酸盐/甲醇溶液冲洗48个柱体积；最后以水/甲醇溶液冲洗12个柱体积。
以山梨醇、甘露醇为分析对象，采用本发明方法对R5-NH2柱进行冲洗前后R5-NH2柱(4.6×150mm)对山梨醇、甘露醇分离分析效果对比如图2所示。
由图2可知，R5-NH2柱冲洗前基线噪音强且分离选择性差；但采用本发明方法冲洗后，基线噪音明显减弱且分离选择性明显改善。上述结果表明，本发明在快速、有效增强氨基柱使用稳定性的同时，能够降低使用蒸发光散射检测器时的基线噪音并提供更好的分离选择性。
实施例3
与实施例2不同之处在于，采用其他不同品牌氨基柱(4.6×150mm)，对比了采用本发明方法冲洗前后其他不同品牌氨基柱(4.6×150mm)对山梨醇、甘露醇的分离分析效果，对比图如图3所示。
由图3可知，本发明增强氨基柱稳定性的方法具有普适性，即本发明不仅只适用于某一种氨基柱，对其他品牌氨基柱同样适用。

Claims (8)

1.一种增强氨基柱稳定性的方法，其特征在于包括以下步骤：
(1)配制磷酸盐溶液；
(2)将(1)中配制的磷酸盐溶液与甲醇配制成磷酸盐/甲醇溶液，将其作为流动相冲洗氨基柱；
(3)配制水/甲醇溶液作为流动相冲洗氨基柱。
2.如权利要求1所述的增强氨基柱稳定性的方法，其特征在于，所述氨基柱是以硅胶为基质，键合有氨基官能团的色谱固定相。
3.如权利要求1所述的增强氨基柱稳定性的方法，其特征在于，所述磷酸盐为磷酸钠、磷酸氢二钠、磷酸二氢钠、磷酸钾、磷酸氢二钾、磷酸二氢钾中的一种或二种以上。
4.如权利要求1所述的增强氨基柱稳定性的方法，其特征在于，所述磷酸盐溶液的物质的量的浓度为0.01～100mM。
5.如权利要求1所述的增强氨基柱稳定性的方法，其特征在于，所述磷酸盐/甲醇溶液为磷酸盐溶液与甲醇按照体积比为1/99～99/1配制而成。
6.如权利要求1所述的增强氨基柱稳定性的方法，其特征在于，所述磷酸盐/甲醇溶液作为流动相冲洗氨基柱的流速为0.01～100mL/min，冲洗1～100个柱体积。
7.如权利要求1所述的增强氨基柱稳定性的方法，其特征在于，所述水/甲醇溶液为水和甲醇按照体积比为1/99～99/1配制而成。
8.如权利要求1所述的增强氨基柱稳定性的方法，其特征在于，所述水/甲醇溶液作为流动相冲洗氨基柱的流速为0.01～100mL/min，冲洗1～100个柱体积。
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* Cited by examiner, † Cited by third party
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WO2012125493A1 (en) * 2011-03-11 2012-09-20 Dionex Corporation Electrostatically bound hyperbranched anion exchange surface coating prepared via condensation polymerization using tertiary amine linkers for improved divalent anion selectivity
CN104931603A (zh) * 2015-05-13 2015-09-23 中国医药集团总公司四川抗菌素工业研究所 一种采用氨基柱测定福多司坦有关物质的方法
CN105510460A (zh) * 2015-12-07 2016-04-20 江南大学 一种定量检测葡萄糖系列酸及内酯的方法

Patent Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2012125493A1 (en) * 2011-03-11 2012-09-20 Dionex Corporation Electrostatically bound hyperbranched anion exchange surface coating prepared via condensation polymerization using tertiary amine linkers for improved divalent anion selectivity
CN104931603A (zh) * 2015-05-13 2015-09-23 中国医药集团总公司四川抗菌素工业研究所 一种采用氨基柱测定福多司坦有关物质的方法
CN105510460A (zh) * 2015-12-07 2016-04-20 江南大学 一种定量检测葡萄糖系列酸及内酯的方法

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