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Numéro de publicationCN103691318 A
Type de publicationDemande
Numéro de demandeCN 201410005331
Date de publication2 avr. 2014
Date de dépôt7 janv. 2014
Date de priorité7 janv. 2014
Autre référence de publicationCN103691318B
Numéro de publication201410005331.4, CN 103691318 A, CN 103691318A, CN 201410005331, CN-A-103691318, CN103691318 A, CN103691318A, CN201410005331, CN201410005331.4
Inventeurs李东浩, 杨翠, 任众, 朴吉寿, 刘翠翠
Déposant延边大学
Exporter la citationBiBTeX, EndNote, RefMan
Liens externes:  SIPO, Espacenet
Micro-scale substance separating method and capillary column transverse eletrochromatography separating device
CN 103691318 A
Résumé
The invention discloses a micro-scale substance separating method and a capillary column transverse eletrochromatography separating device. The separating method comprises the step of introducing a sample into a separating column of a separating device, wherein the separating column is composed of a capillary tube which is used as an external electrode and a conductive material which is used as an internal electrode and penetrates through the capillary tube along the axial direction; the internal electrode and the external electrode are coaxial, and are respectively connected with the two ends of a power supply; the separating column transversely applies electric fields upwards, and electrophoresis effect is utilized for transversely transferring to-be-separated substances, so that contact probability of the to-be-separated substances and a column surface is increased, and separation is realized according to difference of interaction force sizes of the to-be-separated substances and the column surface; an absorption analyzing process of the to-be-separated substances and the column surface is completed by switching direction of the electric fields, so that resolution among the to-be-separated substances is improved. According to the invention, an electrophoretic technique and a chromatographic technique are integrated to realize effective separation of different substances, operation of the separating method and manufacturing of the special-purpose device are simple; and moreover, online detection can be realized.
Revendications(4)  Langue du texte original : Chinois
1.一种微尺度物质的分离方法,包括下列步骤: ①样品由微量注射器引入进样器内; ②打开毛细管柱横向电色谱分离装置的电源开关,使毛细管柱横向电色谱分离装置的分离柱横向上形成电场; ③样品在由运载液的带动下由进样阀进入所述的分离柱,在外加横向电场作用下,样品中待分离物质在分离柱内发生横向迁移,待分离物质迁移到柱表面时,待分离物质会与柱表面发生相互作用; ④电源正负极方向的切换,使待分离物质在分离柱内部横向上来回迁移,并与分离柱内表面发生吸附解吸过程,依据横向力的差异导致物质间分配系数存在差异,从而导致待分离物质沿流动方向有不同的速度,实现不同物质的分离; ⑤分离物质以不同洗脱时间进入检测器内,进行在线检测。 1. A method of separating substances of microscale, comprising the steps of: ① introduced into the sample by a micro-syringe injector; ② open capillary column chromatographic separation means transverse electric power switch, so that the transverse electric capillary column chromatography separation means the transverse electric field is formed; ③ sample in the carrier liquid is driven by the injection valve into the separation column, wherein, under an applied transverse electric field, the sample substances to be separated in the separation column lateral migration, migration of the material to be separated When the surface of the column, the column will be separated from the material surface interact; ④ power of positive and negative switching direction, so that the material to be separated migrate back and forth on the separation column inside the lateral and surface adsorption and desorption processes occur within the separation column, according to the transverse The difference in force leads to differences in the partition coefficient between the material, causing the material to be separated in the flow direction have different speeds, to achieve the separation of different substances; ⑤ separated substances elute at different times into the detector, online detection.
2.根据权利要求1所述的一种微尺度物质的分离方法,其特征在于所述的电场提供方式为脉冲电源或交流电源。 2. A method of separating microscale substance according to claim 1, wherein said electric field provides methods for pulse power or AC power.
3.一种毛细管柱横向电色谱分离装置,包括有提供运载液的泵、进样器、检测器、电源和一端连接进样器,另一端连接检测器的分离柱,其特征在于所述的分离柱包括有: 一个毛细管,该毛细管作为外电极; 一根导电材料,该导电材料作为内电极,沿轴向贯穿毛细管并处于毛细管中心处,即内电极与外电极同轴,内电极与外电极连接电源两端。 3. A transverse electric capillary column chromatography equipment, including providing carrier liquid pump, injector, detector, power supply and connect one end of the injector, and the other end connected to the detector separation column, wherein the separation column comprising: a capillary tube, the capillary tube as the outer electrode; a conductive material, the conductive material as the internal electrode, and axially through the capillary in the capillary at the center, i.e., the inner electrode and the outer electrode coaxial, inner electrode and the outer electrodes are connected across the power supply.
4.根据权利要求3所述的一种毛细管柱横向电色谱分离装置,其特征在于所述的导电材料为钼丝、金丝或银丝。 A capillary column 3, wherein the lateral chromatographic separation apparatus according to claim, wherein said conductive material is molybdenum wire, gold or silver.
Description  Langue du texte original : Chinois

微尺度物质的分离方法及毛细管柱横向电色谱分离装置 Microscale separation material and capillary column chromatographic separation device transverse electric

技术领域 FIELD

[0001] 本发明涉及一种微尺度物质的分离方法及其专用设备,具体是指将电泳技术与色谱技术融合于一体的一种微尺度物质的分离方法及毛细管柱横向电色谱分离装置。 [0001] The present invention relates to a method for the separation of microscale materials and special equipment, specifically refers to the electrophoresis separation and chromatographic techniques, combining to a microscale materials and capillary column chromatographic separation transverse electric devices.

背景技术 BACKGROUND

[0002]目前,微尺度物质如细胞、生物大分子、合成颗粒物、胶体等,其分离和筛选已经成为当代各研究领域中一个优选的主要研究内容。 [0002] Currently, the microscale materials such as cells, biomolecules, synthetic particles, colloids, with its separation and screening has become the field of contemporary research in a preferred main content. 传统的分离技术很难同时分离尺寸范围较宽的样本,而且很难获得单一尺寸的目标物质,这些都将给后续研究带来不便。 The traditional separation technology is difficult to simultaneously separate a wide range of sample sizes, and it is difficult to obtain a single-size target substance, which will follow-up studies inconvenience. 为实现微尺度物质的分离,一些无阻塞、非填充型分离系统被建立,而绝大多数都属于连续型分离技术,此类技术有诸多优势,比如可以实现样品的连续分离与筛选、可实现大量样品的快速分离等。 To achieve the separation of microscale material, some of the non-blocking, non-filled separation system has been established, while the vast majority belong to the continuous separation technology, such technology has many advantages, such as can be achieved continuous separation and screening samples can be realized Rapid separation of large number of samples and the like. 然而此类技术的装置结构和制作过程均比较复杂,并且设备制作精度要求高,对于多个组分的分离,装置制备更为困难。 However, the device structure and fabrication process of such techniques are relatively complex, and requires high precision production equipment, more difficult for the separator, means for preparing a plurality of components. 多出口设计特点很难实现样品的在线检测,故此不适用于未知或复杂样品的分离分析。 Multi-outlet design features difficult to achieve online testing samples, therefore not suitable for the separation and analysis of unknown or complex samples. 另外,大部分微分离技术与现有色谱技术相比,其分离度均较低。 In addition, most of the micro-chromatographic separation technique compared with the prior art, which were lower resolution. 因此,连续型分离技术实际应用过程中必然面临一些挑战。 Therefore, the practical application of continuous separation process is bound to face some challenges. 基于以上局限,为更好的研究微尺度物质,需要开发一种高分辨率、装置制作和实验操作均较简单的技术。 Based on the above limitations, for better research microscale materials needed to develop a high-resolution, device fabrication and experimental operation compared with simple techniques. 更为重要的是,要求技术能够容易实现样品的在线检测。 More importantly, the technology can be easily achieved online request detecting samples.

发明内容 SUMMARY

[0003] 本发明的目的是为了克服非填充型分离系统中微尺度物质分离度低的不足,而提供一种微尺度物质的分离方法及其专用设备毛细管柱横向电色谱分离装置,本发明在分离装置的分离柱横向上引入电场,且柱表面引入色谱作用,微尺度物质进入分离柱后能够在电场作用下发生横向电迁移,通过物质的电迁移行为增加粒子与柱表面的接触几率,进而提高色谱作用的贡献,最终达到提高分离度的目的 [0003] The object of the present invention is to overcome the unfilled material separating separation system microscale low degree insufficient to provide a separation method and its special equipment Microscale lateral capillary column chromatographic separation apparatus, the present invention the introduction of the separation column separation device on the transverse electric field, and the introduction of column chromatography surface effect, microscale material into the separation column can occur transverse electric mobility in the electric field, increasing the chance of contact with the surface of the particles and the column material by electromigration behavior, and thus contribution to improving the chromatographic effects, and ultimately achieve the purpose of improving the degree of separation

为解决上述技术问题,本发明的技术方案如下: To solve the above technical problem, technical solutions of the invention as follows:

一种微尺度物质的分离方法,包括下列步骤: One kind of substance microscale separation method, comprising the steps of:

①样品由微量注射器引入进样器内; ① sample is introduced into the micro-syringe injector;

②打开毛细管柱横向电色谱分离装置的电源开关,使毛细管柱横向电色谱分离装置的分离柱横向上形成电场; ② Open the lateral capillary column chromatographic separation device power switch, so that the lateral capillary column chromatographic separation means the transverse electric field is formed on the separation column;

③样品在由运载液的带动下由进样阀进入所述的分离柱,在外加横向电场作用下,样品中待分离物质在分离柱内发生横向迁移,待分离物质迁移到柱表面时,待分离物质会与柱表面发生相互作用; ③ sample when driven by the carrier liquid into said by injection valve separation column, under an applied transverse electric field, the sample to be separated substances to migrate laterally within the separation column, the column to be separated migrate to the surface of the material to be separation of substances will interact with the column surface occurs;

④电源正负极方向的切换,使待分离物质在分离柱内部横向上来回迁移,并与分离柱内表面发生吸附解吸过程,依据横向力的差异导致物质间分配系数存在差异,从而导致待分离物质沿流动方向有不同的速度,实现不同物质的分离; ④ switching power supply positive and negative direction, so that the material to be separated in the separation column migrate back and forth inside the lateral and surface adsorption and desorption processes occur within the separation column, according to the lateral force distribution coefficient differences lead to differences between the material, causing to be separated material along the flow direction have different speeds, to achieve the separation of different substances;

⑤分离物质以不同洗脱时间进入检测器内,进行在线检测。 ⑤ separation of substances with different elution times into the detector, online testing.

[0004] 所述的电场提供方式为脉冲电源或交流电源。 Field [0004] The manner for providing pulse power or AC power. [0005] 一种毛细管柱横向电色谱分离装置,包括有提供运载液的泵、进样器、检测器、电源和一端连接进样器,另一端连接检测器的分离柱,其中所述的分离柱包括有: [0005] A transverse electric capillary column chromatography equipment, including providing carrier liquid pump, injector, detector, power supply and connect one end of the injector, and the other end connected to the detector separation column, wherein the separation Column comprising:

一个毛细管,该毛细管作为外电极; A capillary, the capillary as an external electrode;

一根导电材料,该导电材料作为内电极,沿轴向贯穿毛细管并处于毛细管中心处,即内电极与与外电极同轴,内电极与与外电极连接电源两端。 An electrically conductive material, the conductive material as the internal electrodes, in the axial direction through the center of the capillary and in the capillary, i.e., the inner electrode and the outer electrode coaxial with the inner electrode and the outer electrode is connected across the power supply.

[0006] 所述的导电材料为钼丝、金丝或银丝。 [0006] The conductive material is molybdenum wire, gold or silver.

[0007] 本发明的分离机理:待分离物质经进样阀引入分离柱后,在分离柱内,待分离物质将受横向电场作用发生横向移动即电泳技术,当待分离物质迁移到柱表面时,待分离物质与柱表面将发生相互作用即色谱技术。 [0007] The separation mechanism of the invention: the material to be separated after the injection valve into a separation column, in the separation column, the material will be separated by the lateral movement of the transverse electric field electrophoresis technique that occur when the material to be separated migrate to the surface of the column to be separated from the surface of the material and the interactions that column chromatographic techniques. 本发明中,待分离物质因自身性质不同,其所受横向作用力将产生差异,进而使待分离物质横向迁移距离产生差异,最终导致待分离物质与柱表面作用力存在差异,即待分离物质在本发明的分离柱内的分配系数存在差异,最终实现待分离物质的分离与分析。 The present invention, the material to be separated due to their different nature, which suffered lateral forces will produce differences, thereby enabling the lateral migration of the material to be separated from a difference, resulting in differences in the material to be separated with the column surface forces, i.e. substances to be separated there is a difference in the separation column of the present invention the distribution coefficient, and ultimately the separation and analysis of substances to be separated. 在本发明系统内,利用电场方向的切换,使待分离物质在分离柱横向上有反复迁移过程,进而使待分离物质与柱表面之间有吸附解吸过程,通过改变电场的频率,提高理论塔板数,进而提高无阻塞、非填充型分离系统的分离度。 In the system of the invention, by the switching electric field direction, so that the material to be separated in the separation column laterally repeated migration, thereby enabling the material to be separated and the cylindrical surface between the adsorption and desorption process, by changing the frequency of the electric field, improving the theoretical column plate number, thus improving separation blocking, non-filled separation system.

[0008] 上述的自身性质包括尺寸大小或分子量、表面电荷以及扩散系数;上述的横向作用力为电泳力和黏滞阻力;上述的待分离物质与柱表面之间的相互作用可以为离子相互作用(静电作用)、非极性相互作用或离子相互作用和非极性相互作用同时存在。 [0008] The properties described above include their size or molecular weight, surface charge and a diffusion coefficient; lateral forces described above for the electrophoretic force and viscous resistance; the interaction between the above-mentioned substances to be separated and the cylindrical surface can be an ionic interaction (electrostatic interaction), non-polar interactions or ionic interactions and non-polar interactions exist.

[0009] 总之,本发明通过将分离柱与检测器的在线联用,克服了当前微分离技术对未知的或复杂的微尺度物质难以在线分离分析的困难。 [0009] In summary, the present invention by the separation column and detector online in conjunction with, overcome the current difficulties microdissection technique unknown substance or complex microscale hard line separation analysis. 另外,本发明有效的将电泳技术和色谱技术融于一体,大大提高了无阻塞、非填充型分离柱的理论塔板数,进而实现样品的高效分离,有望获得单一尺寸的微尺度物质,为微尺度物质的后续研究奠定基础。 In addition, the present invention is effective will electrophoresis and chromatographic techniques into one, greatly improving the number of theoretical plates blocking, unfilled separation column, and thus achieve efficient separation of the sample, is expected to receive a single size of microscale materials for follow-up study microscale material foundation.

附图说明 Brief Description

[0010] 图1为本发明毛细管柱横向电色谱分离装置示意图。 [0010] Figure 1 of the present invention capillary column chromatographic separation Schematic transverse.

[0011] 图2为本发明实施例中不同尺寸聚苯乙烯颗粒分离效果图。 [0011] Figure 2 embodiment of the invention the separation of different sizes of polystyrene particles embodiment FIG.

[0012] 图3为本发明实施例中不同表面电荷聚苯乙烯颗粒分离效果图。 [0012] Figure 3 a different embodiment of the invention the polystyrene particle surface charge separation effect [Fig.

具体实施方式: DETAILED DESCRIPTION:

在图1中,1、运载液,2、六通进样阀,3、样品,4、废液,5、PEEK管,6、导电材料一内电极, In Figure 1, 1, carrier liquid, two, six-way injection valve, 3, sample 4, the waste liquid, 5, PEEK tube, 6, an electrically conductive material within the electrode,

7、毛细管一外电极,8、电源,9、检测器,10、三通阀。 7, a capillary outer electrode, 8, power supply, 9, detector, 10, three-way valve.

[0013] 参考图1,一种毛细管柱横向电色谱分离装置,包括有提供运载液的泵、进样器、检测器、电源和一端连接进样器,另一端连接检测器的分离柱,其中所述的分离柱包括有:一个毛细管7,该毛细管7作为外电极;一根导电材料6,所述的导电材料6可为钼丝、金丝或银丝,该导电材料6作为内电极,沿轴向贯穿毛细管7,为保证分离柱内各处待分离物质所受横向电场力相同,内电极必须置于外电极中心处,即内电极与与外电极同轴,并分别连接电源8两端,进而在分离柱横向上产生电场。 [0013] Referring to FIG. 1. A transverse electric capillary column chromatography equipment, including providing carrier liquid pump, injector, detector, power supply and connect one end of the injector, and the other end connected to the detector separation column, which said separation column comprising: a capillary tube 7, the capillary tube as the outer electrode 7; 6 a conductive material, said conductive material 6 may be a molybdenum wire, gold or silver, the conductive material 6 as an internal electrode, axially through the capillary 7, in order to ensure the separation column separation throughout the substance to be suffered in the same transverse electric field strength, inner electrode must be placed at the center of the outer electrode, that is coaxial with the outer electrode inner electrode, and are connected to the power supply 8 two end, and produce an electric field in the lateral separation column. 本发明无需使用常规液相系统的高压泵,进样器采用六通进样阀2,为将待分离物质引入分离柱,并实现在线检测,分离柱的毛细管7 —端通过固定设置的三通阀10连接六通进样阀2以便引入样品3,另一端通过固定设置的三通阀10连接检测器9以便实分离物质的在线检测,处于毛细管外侧的导电材料6两端由PEEK管5承接。 The present invention eliminates the need for conventional high pressure liquid pump systems, injector with six-way injection valve 2, for the matter to be separated into a separation column, and to achieve online detection, capillary separation column 7-- end by a fixed set of tee six-way valve 10 is connected to the introduction of the injection valve 2 Sample 3, and the other end by a fixed set of three-way valve 10 is connected to the detector in order to achieve the separation line 9 of the test substance, at the outer ends of the capillary conductive material 6 to undertake the PEEK tube 5 . [0014] 利用上述专用的毛细管柱横向电色谱分离装置而公开的一种微尺度物质的分离方法,包括下列步骤: [0014] The use of the dedicated lateral capillary column chromatographic separation apparatus and a separation method disclosed microscale substance, comprising the steps of:

①样品3由微量注射器引入六通进样阀2内; ① Sample 3 introduced by micro-syringe six-way injection valve 2;

②打开电源8的开关,使分离柱横向上形成电场; ② Turn the power switch 8, make the separation column on the transverse electric field is formed;

③样品3在由泵提供的运载液I的带动下由六通进样阀2、三通阀10进入分离柱,在外加横向电场作用下,样品中待分离物质在分离柱内发生横向迁移; ③ Sample 3, driven by a pump provided by the carrier liquid I six-way injection valve 2, the three-way valve 10 into the separation column, under an applied transverse electric field, the sample substances to be separated in the separator string lateral migration;

④电源8的正负极方向切换,使待分离物质在横向上来回迁移,并与柱表面发生吸附解吸过程,横向力的差异使物质间分配系数存在差异,进而导致待分离物质沿流动方向有不同的速度,实现不同物质的分离; ④ power switching positive and negative direction 8, so that the material to be separated migrate back and forth in the transverse direction, and the adsorption process and desorption column surface occurs, so that the difference between the lateral force of the material distribution coefficient differences, leading to material to be separated along the flow direction different speeds, to achieve the separation of different substances;

⑤分离物质以不同洗脱时间由三通阀10进入检测器9内,进行在线检测,同时排出废液4。 ⑤ separation of substances with different elution time from the three-way valve 10 into the detector 9, line detection, while discharging waste 4.

[0015] 在具体分离操作过程中,所述的电源为脉冲电源或交流电源。 [0015] In the process of separating operation in particular, the power to the pulse power or AC power. 所述的横向作用力为电泳力、黏滞阻力、待分离物质与柱表面之间的相互作用力;所述的待分离物质与柱表面之间的相互作用可以为离子相互作用(静电作用)、非极性相互作用或离子相互作用与非极性相互作用同时存在。 The lateral force is electrophoretic force, the viscous resistance, the material to be separated and the interaction force between the surface of the column; according to the material to be separated and the interaction between the surface of the column may be ionic interactions (electrostatic interaction) , non-polar interactions or ionic interactions with the non-polar interactions exist.

[0016] 在毛细管柱横向电色谱系统中,当分离柱固定时,待分离物质与柱表面之间的吸附解析次数直接影响待分离物质间的分离度。 [0016] In the lateral capillary electrochromatography (CEC) system, when the separation column is fixed, the number of adsorption analytical substances to be separated between the cylindrical surface and directly affect the degree of separation between the substance to be separated. 在毛细管柱横向电色谱技术中,通过优化电场大小(电压和频率)、电泳时间、运载液流速等参数可提高待分离物质间的分离度。 In the transverse electric capillary column chromatography technique, by optimizing the electric field magnitude (voltage and frequency), the run time, the carrier liquid flow rate and other parameters can increase the degree of separation between the substance to be separated. 优化结果显示,外加电场频率/和持续时间ί与洗脱时间(6成正相关,电压U对无影响,但影响颗粒的保留量和峰形;另外,电压U和电场持续时间t与半峰宽#1/2成负相关,f与#1/2成正相关;一定范围内,运载液流速影响不大。本发明中在电场作用下,待分离物质将发生横向迁移,因待分离物质横向迁移距离不同使其与柱表面的相互作用力存在差异,即分配系数不同。另外,本发明中采用脉冲频率的变化使待分离物质与柱表面发生吸附解吸过程。基于以上两个原因,最终实现微尺度的不同物质之间的分离。 Optimization results show that the frequency of the applied electric field and / duration ί and elution time (60% positive correlation, no effect on the voltage U, but the impact of particle retention and peak shape; in addition, the voltage U and the electric field and the duration t FWHM # 1/2 negative correlation, f positive correlation with # 1/2 percent; within a certain range, the flow rate of carrier liquid is not affected in the present invention, the electric field, the material will be separated laterally migrate, because the lateral migration of substances to be separated. Different interactions differ from it with stem surface, i.e., the distribution coefficient different. Further, the present invention uses a pulse frequency of changes to be separated so that the surface of the material and the process of adsorption-desorption column occurs. For these two reasons, ultimately micro scale separation between different substances.

[0017] 本发明在实际分离过程中对微尺度物质实现分离,以聚苯乙烯颗粒物为例评价其分辨率。 [0017] The present invention is in the actual separation process to achieve separation of microscale material, polystyrene particles Case evaluated resolution. 毛细管柱横向电色谱技术分尚条件如下,分尚柱长度为20 cm;电压为1.2 V ;频率为0.5 Hz ;电场施加时间为10 min ;脉冲周期分别为0.4和0.6 ;运载液为蒸馏水;流速为1.0mL/h。 Transverse electric capillary chromatography conditions are still divided, sub-column length is still 20 cm; voltage of 1.2 V; frequency of 0.5 Hz; electric field application time is 10 min; pulse period were 0.4 and 0.6; carrier liquid is distilled water; the flow rate of 1.0mL / h. 如图2、图3。 Figures 2, 3. 从图2、3可以得出,毛细管柱横向电色谱分离技术实现了不同微尺度物质(不同尺寸或不同表面电荷)分离的目的。 Figures 2 and 3 can be drawn from, the lateral capillary column chromatographic separation technology to achieve a purpose different microscale materials (different sizes or different surface charge) separation. 本发明公开了毛细管柱横向电色谱技术,首次有效的将电泳技术和色谱技术融于一体,提高待分离物质间的分辨率,获得单一尺寸的微尺度物质,而且该技术可以实现在线检测。 The present invention discloses a transverse electric capillary chromatography, electrophoresis technology will be effective for the first time and chromatographic techniques into one, to improve the resolution between the material to be separated, to obtain a single dimension of microscale materials, and the technology can be implemented on line.

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Classifications
Classification internationaleG01N30/06, G01N1/28, B01D57/02, B01D15/10, G01N1/34
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2 avr. 2014C06Publication
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