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CN-122003282-A - Method and device for extracting substances from adsorption layers of chromatographic plates

CN122003282ACN 122003282 ACN122003282 ACN 122003282ACN-122003282-A

Abstract

The subject of the invention is a method and a device for extracting substances from the adsorption layer of a chromatography plate. According to the invention, the method comprises forming a micro-container with a side wall of a sleeve vertically penetrating the adsorption layer and pressing against the chromatographic carrier plate and a bottom of the sleeve wall of the micro-container being the chromatographic carrier plate. To the microcontainers formed in this way, one or more parts of several tens to several hundreds of microliters of extract are added. The extract is obtained and, after extraction from the cartridge, is subjected to qualitative and/or quantitative analysis using instrumental techniques. The method is performed using a mechanism that presses the sleeve vertically against the chromatographic plate. The sleeve has a specially contoured lower edge which helps achieve a tight connection between the lower portion of the sleeve and the chromatographic carrier plate. The device uses a replaceable sleeve made of a chemically resistant material, preferably plastic, such as polypropylene.

Inventors

  • T. H. Guido
  • M. Sharek
  • M. Soldier

Assignees

  • SFPE有限责任公司

Dates

Publication Date
20260508
Application Date
20240919
Priority Date
20230920

Claims (20)

  1. 1. A method of extracting a substance from an adsorption layer of a chromatography plate/from a specific location in the adsorption layer of a chromatography plate, characterized by comprising the steps of: Forming a micro-container between the support of the chromatography plate and the sleeve pressed thereon, wherein the sleeve is pressed at right angles against the support of the adsorption layer, such that the bottom is the support of the adsorption layer and the wall of the micro-container is the wall of the sleeve, wherein the height of the micro-container is preferably up to 50 mm, Introducing a predetermined small volume of extraction liquid into the resulting micro-container through the upper opening of the cartridge, the volume being greater than 20 μl, preferably between 20 μl and 500 μl, Maintaining the extract in contact with the adsorbent layer in a micro-vessel for a prescribed short period of time, in the range 1 min to 10 min, Extract (i.e. the extract with the extracted material) is withdrawn from the micro-container. Preferably, a syringe or pipette is used to remove the liquid from the micro-containers.
  2. 2. The extraction process according to claim 1, characterized in that in a subsequent stage the extract is sent to an instrumental analysis or the solid particles of the adsorption layer are separated from the extract before being sent to an instrumental analysis.
  3. 3. The extraction method according to claim 2, characterized in that it comprises a step of removing solid particles of the adsorption layer, wherein the solid particles of the adsorption layer are separated from the extract and removed by filtration or centrifugation before the extract is placed in a vial or by subjecting the solid particles to a sedimentation process after the placement in a vial.
  4. 4. A method according to claims 1-3, characterized in that the micro-containers have a circular inner opening, preferably with an inner diameter of 2mm to 10 mm, or an oval inner opening, preferably with a short axis of 2mm to 6 mm and a long axis of 3 mm to 10 mm.
  5. 5. Extraction method according to claims 1-4, characterized in that a part of the introduced extraction liquid (11) is withdrawn from the micro-container and immediately reintroduced into the micro-container, and the operations of withdrawing and introducing the part of the extraction liquid (11) are repeated 2 to 10 times.
  6. 6. The extraction method according to claims 1-4, characterized in that the extraction process is repeated several times, preferably with a new portion of the extract (11), and the obtained extraction solutions are combined and the number of extraction repetitions is experimentally determined.
  7. 7. Extraction process according to claims 1-6, characterized in that the extraction liquid (11) is mixed in a micro-container, preferably using a micro-stirrer or by shaking or sonication.
  8. 8. The method of extracting material from a chromatography plate of claims 1-6, wherein the time of one extraction process is up to 10 min.
  9. 9. Method for extracting substances from chromatography plates according to claims 1-8, characterized in that during the extraction run the upper opening of the sleeve is preferably closed with a plug/cork (7) with an opening.
  10. 10. An apparatus for extracting a substance from a chromatography plate, comprising a body and a table for holding the chromatography plate, characterized in that it comprises: -an element for fixing at least one sleeve (1) in the form of a bracket (19) and/or a fastening beam (30, 36) and connected to the body (17, 32, 44); -a pressing mechanism (22, 23, 24, 26, 31, 39) connected to the device body (17, 32, 44) ensuring that the at least one sleeve (1) is pressed vertically against the chromatography plate (8) and that the at least one sleeve (1) can be removed from the chromatography plate (1); -elastic elements (20, 35, 40) ensuring the fitting of the edges of the sleeve (1) to the chromatographic plate (8).
  11. 11. Device for extracting substances from chromatography plates according to claim 10, characterized in that it has at least one exchangeable sleeve (1) made of chemically inert material, preferably plastic, which sleeve is sharp over the entire circumference of the lower edge (3) and which sharp end is formed by the connection of the two surfaces of the sleeve, i.e. the outer surface of the sleeve (1) and the surface of the chamfer (4) inside the sleeve, or the edge (3) is formed by two surfaces, one being the surface constituting the inner wall surface of the sleeve and the other being the surface constituting the outer chamfer (4 ') of the bottom of the sleeve, or the edge is formed by the surface of the outer chamfer (4 ") and the inner chamfer (4'").
  12. 12. Device for extracting substances from chromatography plates according to claim 11, characterized in that the angle between the surfaces constituting the sharp ends of the edge (3) is an acute angle, ranging between 15 ° and 70 °.
  13. 13. Device for extracting substances from chromatography plates according to claims 10-12, characterized in that the edge formed at the junction of the outer surface of the sleeve (1) and the chamfer (3) surface of the inner sleeve (1) has a passivation treatment (6) over the whole circumference and the width of the passivation (6) surface is preferably 0.1 to 0.5 mm.
  14. 14. Device for extracting substances from chromatography plates according to claims 10-13, characterized in that the sleeve (1) has at least one hole (5) in its wall near the end opposite the end with sharp edge (3), preferably with a diameter in the range of 0.3 to 1.5 mm.
  15. 15. Device for extracting substances from chromatography plates according to claims 10-14, characterized in that the sleeve (1) has at least one longitudinal groove on the inner side wall, starting from the end opposite to the end of the sleeve with sharp edges (3), up to 10mm in length, 0.3-1.0 mm in width and 0.3-0.6 mm in depth.
  16. 16. Device for extracting substances from chromatography plates according to claims 10-15, characterized in that the inner cross section of the sleeve (1) is circular, preferably with a diameter of 2 to 10mm, or the inner cross section of the sleeve (1) is elliptical, preferably with a short axis in the range of 2 to 6mm, preferably with a long axis in the range of 3 to 10mm, or the inner cross section of the sleeve has a shape close to circular or elliptical.
  17. 17. Device for extracting substances from chromatography plates according to claims 10-16, characterized in that the length of the sleeve (1) can reach 50 mm, preferably between 5mm and 50 mm, even more preferably between 10 mm and 50 mm.
  18. 18. Device for extracting substances from chromatography plates according to claims 10-17, characterized in that the sleeve (1) is provided with one projection/flange or two projections/flanges (2) over the entire circumference of its outer surface or with one recess/groove or two recesses/recesses over its entire circumference.
  19. 19. Device for extracting substances from chromatography plates according to claims 10-18, characterized in that the sleeve (1) is preferably provided with a plug (7) at the opening opposite the sleeve opening with sharp edges (3), and that the plug (7) is provided with a hole having a diameter of up to 2 mm.
  20. 20. Device for extracting substances from chromatography plates according to claims 10-19, characterized in that the holder (19) is equipped with a snap-on mechanism securing the sleeve (1), wherein the holder (19) is connected to the pressing mechanism by means of an elastic element (20).

Description

Method and device for extracting substances from adsorption layers of chromatographic plates The subject of the present invention is a method and a device for extracting a substance from a substance in a specific zone/spot in an adsorption layer on a chromatographic plate, which zone/spot is obtained after development of the chromatogram, which zone contains one or more components originating from the separation mixture. The obtained extract can be used for qualitative and/or quantitative analysis using instrumental techniques. In laboratory practice, the simplest method of extracting a substance from the adsorption layer of a chromatographic plate is to scrape off a specific area containing the substance or mixture of substances, transfer it to a filter paper filter layer placed on a funnel, and elute the substance with a suitable solvent. Depending on the application, for example for quantitative or qualitative analysis or for preparation purposes, the extract obtained is subjected to further treatment. The method is described in thin layer chromatography, fourth revised and expanded edition （Thin-Layer Chromatography, Fourth Edition, Revised and Expanded, B. Fried, J. Sherma, 1999, Marcel Dekker, Inc.） Is described in chapter 10 and chapter 12. In publication J.chromatogr.A.103 (1975), 279-288, B.Falk and K.Krummen, a method and apparatus for eluting substances from spots on chromatography plates is described. The essence of the method and apparatus is that an annular groove is formed in the adsorption layer around the spot, a teflon head with a seal is pressed onto the groove, and an elution solvent is supplied to one opening in the head using a syringe pump and the effluent is collected through the other opening. A suitably selected solvent is flowed through the region of the adsorbent layer defined by the aforementioned seals, eluting the material therefrom. With this device, it is possible to elute substances from six substance spots in the adsorption layer of the chromatography plate simultaneously. The disadvantage of this device is that a recess needs to be made manually in the adsorption layer for the head seal and a flushing device is required before the next elution process. Furthermore, a syringe pump must be used, which additionally increases the cost of the device. One known device for eluting substances from an adsorption layer on a chromatography plate is described in the publication Anal. Bioanal. Chem. (2004) 378:964-968, DOI 10.1007/s00216-003-2293-3 and in patent specification DE10036293A1 of H Luftmann. The device is in the form of a steel piston with a sharp edge on the entire circumference of its lower surface and two capillary channels, one for supplying the extraction liquid and the other for discharging the extraction liquid. The supply of the extract is carried out by means of a pressure pump. In order to elute the substance from the adsorption layer, the piston is pressed against the layer and the eluting solution is pressed in by a pressure pump via the supply channel, the adsorption layer and the discharge channel. From the latter channel, the liquid containing the eluted substance is directed to a vial or directly into the instrument analysis device. The disadvantage of the presented solution is that the device channels are often blocked by adsorbent particles, which need to be thoroughly cleaned before the next elution process from the next position of the adsorption layer of the chromatography plate, and the use of pressure pumps, which are often expensive equipment. This protocol consumes a large amount of eluent, and in addition, the obtained sample is diluted, sometimes requiring concentration for subsequent steps. From U.S. patent No. 5208458, 5 and 4, 1993, a device is known that connects various types of planar electropherograms, including gel electropherograms, to a mass spectrometer. The connection means comprises a pump for delivering solvent to the surface of the electrophoresed pattern at a prescribed speed, a system for disrupting/pulverizing the gel and releasing molecules of the substance contained therein so that they can be absorbed by the solvent, and a capillary for transferring the solvent containing the eluting substance to the mass spectrometer. The limitation of this device is that it is intended for use in electropherograms from which material is eluted into a mass spectrometer. Furthermore, the elution solvent must be supplied by a separate pump that is connected to the sample injector of the mass spectrometer, which makes the device complex, relatively expensive and the connection channels prone to clogging. Another solution is described in patent application US 2011/0269166 A1, 11/3/2011. The sample in the adsorption layer (acting as a filter layer) is placed on a flexible hydrophobic substrate. The sample may comprise biological or chemical materials and absorbing/filtering materials/layers. The filter layer comprises an absorbent material embedd