Separation and purification device for biomarkers in marine sediments

Abstract

The utility model provides a separation and purification device for biomarkers in marine sediments. The separation and purification device comprises a purification column and an extraction box, the purification column comprises a liquid storage cup and a glass column body, the glass column body comprises a column body and an insertion end, the column body is filled with one kind of filler or two or more kinds of combined filler, a sand core sieve plate is arranged in the column body, and a flow regulating valve is further arranged on the column body; the extraction box is of a side opening type closed box body structure, a plurality of column heads are arranged at the top of the extraction box, insertion ends are inserted into the column heads, the tail parts of the column heads are connected with flow guide pipes, a push-pull plate is lapped in the extraction box, a collecting frame is arranged on the push-pull plate, a plurality of constant-volume collecting pipes are placed on the collecting frame, and the constant-volume collecting pipes are communicated with the flow guide pipes. The separation and purification device disclosed by the utility model is efficient, rapid, economical, safe and environment-friendly, can be used for simultaneously enriching and purifying samples and simultaneously separating and purifying one or more biomarkers, and is suitable for batch operation, so that the analysis speed is greatly improved.

Description

Technical Field

[0001] The utility model belongs to the field of marine bioorganic geochemistry, and in particular relates to a separation and purification device for biomarkers in marine sediments. Background Art

[0002] Biomarkers: After an organism dies and is buried, it undergoes a series of geochemical changes, including oxidation, reduction, and isomerization. The less stable components in its biochemical composition are oxidized and decomposed, and the stable components form fingerprint markers with a certain structure that can identify the source of organic matter.

[0003] In the field of marine bio-organic geochemical testing, when detecting biomarkers such as alkanes, aromatic hydrocarbons and fatty alcohols in marine sediments, due to the complex matrix of marine sediment samples, some sediment samples have a darker color after extraction and concentration. If they are directly tested with a gas chromatography-mass spectrometer, it will not only affect the detection of target compounds, but also contaminate the instrument, so sample pretreatment is required.

[0004] Sample pretreatment is an extremely time-consuming, tedious process that can easily introduce analytical errors, and the method used plays a crucial role in sample analysis. Currently, purification techniques such as silica gel column chromatography and solid-phase extraction are primarily used to remove impurities from the extract, reduce interference, and improve detection sensitivity. However, both methods have certain drawbacks when applied to biomarkers in marine sediments:

[0005] 1. Silica gel column chromatography technology

[0006] In silica gel column chromatography, the columns are secured to a metal stand with test tube clamps, which occupies a large experimental space and makes large-scale processing difficult in typical laboratories. Column chromatography typically requires large amounts of solvent, resulting in lengthy nitrogen flushing times and extended experimental cycles. During the column chromatography process, the operating system is not airtight, allowing for continuous solvent evaporation, which poses a health risk to the experimenter and pollutes the environment.

[0007] 2. Solid Phase Extraction Technology

[0008] Solid-phase extraction (SPE) is a sample pretreatment technology developed in recent years. This technology mainly uses solid-phase extraction (SPE) instruments and SPE cartridges. However, the adsorption of many substances is irreversible. Once adsorbed, they cannot be eluted, which affects the next adsorption and causes the extraction cartridges to be non-reusable. The high cost of SPE instruments and cartridges increases the cost burden of the experiment. The SPE cartridge packing is fixed, but in actual application, it is necessary to select the appropriate column packing for separation and purification based on the specific conditions of the sample, resulting in poor selectivity. The SPE column bed is short and has poor selectivity, making it difficult to extract all target compounds from complex mixtures. It is also susceptible to interference from hybrids, thus affecting the accuracy of the results.

[0009] Therefore, we urgently need a separation and purification device for biomarkers in marine sediments that is efficient, rapid, economical, safe and environmentally friendly to solve the above problems. Utility Model Content

[0010] In order to solve the defects in the prior art, the present invention proposes a separation and purification device for biomarkers in marine sediments, the scheme of which is as follows:

[0011] A device for separating and purifying biomarkers in marine sediments comprises a purification column and an extraction box; the purification column comprises, from top to bottom, a liquid storage cup and a glass cylinder, the glass cylinder comprising a cylinder body and an insertion end, the cylinder body being filled with one filler or a combination of two or more fillers, a sand core sieve plate being provided in the cylinder body, a flow regulating valve being further provided on the cylinder body, the flow regulating valve being located below the sand core sieve plate; the extraction box being a side-opening closed box structure, a plurality of column heads being provided at the top of the extraction box, the insertion end being inserted into the column heads, the tail of the column heads being connected to a guide pipe, a push-pull plate being overlapped in the extraction box, a collection rack being provided on the push-pull plate, a plurality of constant-volume collection pipes being placed on the collection rack, the constant-volume collection pipes being connected to the guide pipe.

[0012] The purification column can simultaneously complete sample enrichment and purification, and can simultaneously separate and purify one or more biomarkers.

[0013] The volume of eluent required varies depending on the type of biomarker. Storage cups of different sizes can be selected to achieve a one-time addition of the eluent to complete the separation and purification of the target component.

[0014] Glass columns of varying lengths and thicknesses can be selected for different samples, and can be filled with a single filler or a combination of two or more fillers, making them suitable for a wider range of applications. Purification columns can be reused after washing, significantly reducing experimental costs.

[0015] The sand core sieve plate is embedded in the glass column to ensure that the filler is stable in the column and prevents the column head from being blocked, thereby controlling the flow rate of the sample and eluent and ensuring the smooth progress of the separation and purification process.

[0016] By setting the flow regulating valve, the flow regulating valve can be turned off when the experimenter needs to go out for something, so as to prevent the glass column from drying up and affecting the separation and purification effect.

[0017] Furthermore, the constant volume collection tube includes a first collection tube and a second collection tube of an integrated structure from bottom to top. The constant volume of the first collection tube is 1 ml, the graduation value is 0.1 ml, and the diameter of the second collection tube is larger than that of the first collection tube.

[0018] The marine sediment organic biomarker sample collected generally needs to be constant volume to 1mL, then carries out gas chromatography-mass spectrometer and tests.Collection tube of prior art is the round bottom collection tube of unified diameter up and down, need to be put on nitrogen blower and blow to 0.5mL and then transfer to chromatogram injection bottle, and wash 3 times collection tubes with solvent and transfer to chromatogram injection bottle in a lump, then after drying with nitrogen blower, add 1mL solvent constant volume in chromatogram injection bottle with pipette gun or pipette, test with gas chromatography-mass spectrometer after constant volume, constant volume error is larger directly in round bottom collection bottle like this, affects the accuracy of test data, and pre-treatment cycle becomes longer, and because some organic components are easily volatilized, nitrogen drying easily causes the problem of low recovery rate.The utility model is set to elongated structure by the first collection tube, can directly collect liquid after nitrogen blow to 1mL in constant volume component collection tube, directly transfer to chromatogram injection bottle and carry out instrument test, not only improves pre-treatment efficiency, can guarantee the accuracy of data again.

[0019] Furthermore, a boss extends from the bottom of the extraction box toward the side wall for overlapping the push-pull plate, and a handle is provided at the front end of the push-pull plate, so that the collection rack can be directly pulled out of the box as a whole, making it convenient to replace the fixed-volume collection tube and clean the collection rack. The purification column does not move due to the removal or placement of the collection rack / tube, thus avoiding the problem of the purification column tipping over or spilling liquid.

[0020] Furthermore, a positioning groove is provided on the top surface of the push-pull plate for positioning the collection rack.

[0021] Furthermore, the push-pull plate is provided with a plurality of slots to facilitate waste liquid to flow to the bottom of the box.

[0022] Furthermore, the collection rack includes three layers of horizontal plates, and adjacent horizontal plates are connected by vertical beams. The upper horizontal plate and the middle horizontal plate are both provided with a first through hole for placing the second collection tube, and the lower horizontal plate is provided with a second through hole for positioning the first collection tube, and the diameter of the second through hole is smaller than the diameter of the first collection tube.

[0023] Furthermore, the liquid storage cup is funnel-shaped, and the bottom of the liquid storage cup is inserted into the top of the glass cylinder. The volume of the liquid storage cup includes 5 ml, 10 ml or 15 ml, etc.

[0024] Furthermore, a waste liquid outlet is provided at the bottom of the extraction box, and a waste liquid discharge valve is provided on the waste liquid outlet to collect waste liquid generated during the pretreatment process to avoid environmental pollution.

[0025] Furthermore, the length of the column body ranges from 15 to 25 cm, and the inner diameter ranges from 6 to 10 mm.

[0026] Compared with the prior art, the advantages of the present invention are as follows:

[0027] The separation and purification device of the utility model is efficient, rapid, economical, safe and environmentally friendly. Through the structure of the purification column and the extraction box, it can simultaneously complete the enrichment and purification of the sample, and can realize the simultaneous separation and purification of one or more biomarkers. It is suitable for batch operation, thereby greatly improving the analysis speed.

[0028] The glass column of the utility model can be filled with not only a single filler but also a combination of two or more fillers, which is suitable for more application scenarios. The volume of eluent required varies depending on the type of biomarker, and storage cups of different volumes can be replaced to achieve a one-time addition of the eluent dosage to complete the separation and purification of the target component.

[0029] The extraction box of the utility model is a closed integral box body, which is made of transparent material resistant to organic solvents, so that the liquid level of the collection tube in the box body can be easily observed; a door panel is provided on the side for easy operation; a waste liquid discharge valve is provided at the bottom of the box to centrally treat the waste liquid generated during the experiment, thereby protecting human health and avoiding environmental pollution. BRIEF DESCRIPTION OF THE DRAWINGS

[0030] Figure 1 This is a schematic structural diagram of the separation and purification device according to an embodiment of the present invention (with the door open);

[0031] Figure 2 This is a schematic diagram of the structure of the separation and purification device according to an embodiment of the utility model (with the door open);

[0032] Figure 3 This is a schematic structural diagram of a purification column according to an embodiment of the present utility model;

[0033] Figure 4 This is a structural diagram of a constant volume collecting pipe according to an embodiment of the present utility model;

[0034] In the above figures: 1. Purification column; 11. Liquid storage cup; 12. Glass cylinder; 121. Column body; 122. Insertion end; 123. Filler; 124. Sand core sieve plate; 125. Flow regulating valve; 2. Extraction box; 21. Column head; 22. Guide tube; 23. Push-pull plate; 231. Positioning groove; 232. Handle; 233. Slot hole; 24. Collection rack; 25. Constant volume collection tube; 251. First collection tube; 252. Second collection tube; 26. Door; 27. Boss; 28. Waste liquid outlet. DETAILED DESCRIPTION

[0035] To facilitate those skilled in the art to understand the present invention, specific implementations of the present invention are described below with reference to the accompanying drawings.

[0036] like Figures 1 to 4As shown, the utility model provides a separation and purification device for biomarkers in marine sediments, comprising a purification column 1 and an extraction box 2.

[0037] Specifically, the purification column 1 includes a liquid storage cup 11 and a glass column 12 from top to bottom.

[0038] Further, if Figure 3 As shown, the glass column 12 includes a column body 121 and an insertion end 122. The column body 121 is filled with a filler 123, or a combination of two or more fillers 123. A sand core sieve plate 124 is provided in the column body 121 and is located at the bottom of the filler 123. A flow regulating valve 125 is also provided on the column body 121, and the flow regulating valve 125 is located at the bottom of the sand core sieve plate 124. Depending on the different components of the sample measured, the amount of eluent used is also different. The liquid storage cup 11 of different sizes can be replaced to add the required amount of solvent at one time, reducing the number of times the laboratory technicians repeatedly add reagents and reducing labor intensity. The glass column 12 can be changed in length, thickness and thickness for different samples, and can be filled with a single filler 123 or a combination of different fillers 123 to meet the needs of more scenarios. In addition, the glass purification column 1 can be reused, saving costs. With the addition of the sand core sieve plate 124, the column head 21 will not be blocked when the filler 123 is filled. The flow regulating valve 125 can be turned off when the experimenter needs to go out to prevent the column from running dry.

[0039] Specifically, the length of the column body 121 ranges from 15 to 25 cm, and the inner diameter ranges from 6 to 10 mm. In this embodiment, the column body 121 is 20 cm long and has an inner diameter of 8 cm, and the insertion end 122 is 1 cm long and has an inner diameter of 2 mm.

[0040] Further, if Figure 3 As shown, to reduce the evaporation rate of the eluent, the main body of the liquid storage cup 11 is circular, but other shapes such as pear-shaped are also possible. A small square opening is provided at the top of the liquid storage cup 11, the opening being sized to facilitate the injection of the eluent. The bottom of the liquid storage cup 11 is inserted into the top of the glass cylinder 12. Depending on the type of biomarker, the volume of eluent required also varies. Liquid storage cups 11 of varying sizes can be selected to achieve separation and purification of the target component with a single addition of the eluent. For example, the volume of the liquid storage cup 11 may be 5ml, 10ml, or 15ml, etc.

[0041] Specifically, the extraction box 2 is a sealed box structure with a side-opening door. Several stigmas 21 are fixedly mounted on the top of the extraction box 2. In this embodiment, the stigmas 21 are arranged in two rows and six columns, facilitating high-throughput sample processing. The insertion end 122 of the glass cylinder 12 is inserted into the stigmas 21. The tail of each stigma 21 is connected to a flow guide tube 22. A sliding plate 23 is connected to the extraction box 2. In this embodiment, a collection rack 24 is mounted on the sliding plate 23. Several fixed-volume collection tubes 25 are placed on the collection rack 24, and the fixed-volume collection tubes 25 are connected to the flow guide tube 22.

[0042] When in use, close the door 26. When the constant volume collecting tube 25 needs to be replaced, open the door 26. Figure 2 As shown, pull out the push-pull plate 23 and replace the constant volume collection tube 25 on the collection rack 24. In this way, the collection rack 24 can be directly pulled out of the box as a whole, which is convenient for replacing the constant volume collection tube 25 and cleaning the collection rack 24. There is no need to move the top cover of the extraction box 2 as in the prior art. The purification column 1 does not move due to the removal or placement of the collection rack 24 / tube, thus avoiding the problem of the purification column 1 tipping over or spilling liquid.

[0043] In this embodiment, Figure 2 As shown, a boss 27 extends from the bottom of the extraction box 2 to the side wall for overlapping the push-pull plate 23; a handle portion 232 is provided at the front end of the push-pull plate 23; a positioning groove 231 is provided on the top surface of the push-pull plate 23 for positioning the collection rack 24; a plurality of slots 233 are also provided on the push-pull plate 23 to facilitate the waste liquid to be retained at the bottom of the box body. In this embodiment, the slots are provided as long holes in three rows and four columns; the side panels of the extraction box 2 can be made of a transparent material resistant to methanol and dichloromethane to facilitate observation of the liquid level in the collection tube.

[0044] Specifically, such as Figure 4 As shown, the constant-volume collection tube 25 comprises, from bottom to top, an integral first collection tube 251 and a second collection tube 252. The first collection tube 251 has a constant volume of 1 ml and a graduation of 0.1 ml. The diameter of the second collection tube 252 is larger than that of the first collection tube 251. The tail of the constant-volume collection tube 25 of the present invention is slender and long (i.e., the first collection tube 251 is configured as a slender structure), allowing for direct nitrogen blowing to maintain constant volume, eliminating intermediate steps and shortening the pre-processing cycle.

[0045] In order to facilitate observation of the volume of the collected eluent, a scale is set on the constant volume collection tube 25.

[0046] In this embodiment, Figure 2 As shown, the collection rack 24 includes three layers of horizontal plates, and adjacent horizontal plates are connected by vertical beams. The upper and middle horizontal plates are both provided with a first through hole for placing the second collection tube 252, and the lower horizontal plate is provided with a second through hole for positioning the first collection tube 251. The diameter of the second through hole is smaller than the diameter of the first collection tube 251.

[0047] A waste liquid outlet 28 is provided at the bottom of the extraction box 2 , and a waste liquid discharge valve is provided on the waste liquid outlet 28 .

[0048] Working process:

[0049] Filling the column - first add about 1.6g of silica gel to the glass column 12, then add 1cm of anhydrous sodium sulfate, and fill it tightly to avoid loose problems.

[0050] Activation—Clean interfering substances and solvent residue from the glass cylinder 12; wet the packing 123 so that the sample solution can flow through the glass cylinder 12. Keep the flow control valve 125 closed, select a reservoir cup 11 of appropriate volume, and add 10 mL of a 5:95 methanol:dichloromethane solution to the glass cylinder 12. Once the solvent completely soaks the packing 123, allow the solution to flow until the liquid surface is barely visible, then close the flow valve. Then, add 10 mL of n-hexane, open the flow valve, allow the solution to flow until the liquid surface is barely visible, and then close the flow valve.

[0051] Sample loading: Allow the sample solution to flow through glass column 12, allowing filler 123 to adsorb the target compound. Transfer the concentrated sample to column head 21 after nitrogen blow-through, allowing the target compound components to be fully adsorbed by filler 123. Open flow control valve 125, allow the sample to flow until the liquid surface is barely visible, and then close flow control valve 125.

[0052] Elution—Select a suitable solvent that will not wash away the target product adsorbed on the filler 123 within the glass column 12, while also removing any interfering substances. Elute with 1 mL of n-hexane. Open the flow control valve 125 and allow the solution to flow until the liquid surface is barely visible. Then close the flow control valve 125.

[0053] Elution—Select an appropriate solvent to elute the target compound adsorbed on the filler 123 within the glass column 12 and collect it in the constant-volume collection tube 25. Add 8 mL of n-hexane all at once, open the flow control valve 125, and elute to obtain the alkane and aromatic components. Close the flow control valve 125, open the door 26, pull out the push-pull plate 23, pull out the collection rack 24, replace the constant-volume collection tube 25, push the push-pull plate 23 into the extraction box 2, close the door 26, and add 12 mL of methanol / dichloromethane (5:95) all at once. Open the flow control valve 125 to elute to obtain the fatty alcohols.

[0054] Waste liquid-The waste liquid generated by activation and elution is discharged through the waste liquid discharge valve.

[0055] The above-described embodiments of the present invention do not limit the scope of protection of the present invention. Any modifications, equivalent replacements, and improvements made within the spirit and principles of the present invention shall be included in the scope of protection of the claims of the present invention.

Claims

1. A device for separating and purifying biomarkers in marine sediments, characterized in that: It includes a purification column (1) and an extraction box (2); The purification column (1) comprises a liquid storage cup (11) and a glass column (12) from top to bottom, the glass column (12) comprises a column body (121) and an insertion end (122), the column body (121) is filled with a filler (123) or a combination of two or more fillers (123), a sand core sieve plate (124) is provided in the column body (121), and a flow regulating valve (125) is further provided on the column body (121), and the flow regulating valve (125) is located at the lower part of the sand core sieve plate (124); The extraction box (2) is a side-opening closed box structure, with a plurality of column heads (21) arranged on the top of the extraction box (2), the insertion end (122) being inserted into the column head (21), the tail of the column head (21) being connected to the guide tube (22), a push-pull plate (23) being overlapped in the extraction box (2), a collection rack (24) being arranged on the push-pull plate (23), a plurality of fixed-volume collection tubes (25) being placed on the collection rack (24), and the fixed-volume collection tubes (25) being connected to the guide tube (22).

2. The device for separating and purifying biomarkers in marine sediments according to claim 1, characterized in that: The constant volume collection tube (25) comprises, from bottom to top, a first collection tube (251) and a second collection tube (252) of an integrated structure; the constant volume of the first collection tube (251) is 1 ml, the graduation value is 0.1 ml, and the diameter of the second collection tube (252) is larger than the diameter of the first collection tube (251).

3. The device for separating and purifying biomarkers in marine sediments according to claim 1, characterized in that: A boss (27) extends from the bottom of the extraction box toward the side wall for overlapping the push-pull plate (23), and a handle portion (232) is provided at the front end of the push-pull plate (23).

4. The device for separating and purifying biomarkers in marine sediments according to claim 3, characterized in that: The top surface of the push-pull plate is also provided with a positioning groove (231) for positioning the collection rack (24).

5. The device for separating and purifying biomarkers in marine sediments according to claim 4, characterized in that: The push-pull plate (23) is also provided with a plurality of slots (233).

6. The device for separating and purifying biomarkers in marine sediments according to claim 2, characterized in that: The collection rack (24) comprises three layers of transverse plates, adjacent transverse plates being connected by vertical beams, the upper transverse plate and the middle transverse plate both being provided with a first through hole for placing the second collection tube (252), the lower transverse plate being provided with a second through hole for positioning the first collection tube (251), the diameter of the second through hole being smaller than the diameter of the first collection tube (251).

7. The device for separating and purifying biomarkers in marine sediments according to claim 1, characterized in that: The main body of the liquid storage cup (11) is round or pear-shaped, and the bottom of the liquid storage cup is inserted into the top of the glass cylinder (12).

8. The device for separating and purifying biomarkers in marine sediments according to claim 1, characterized in that: A waste liquid outlet (28) is provided at the bottom of the extraction box (2), and a waste liquid discharge valve is provided on the waste liquid outlet (28).

9. The device for separating and purifying biomarkers in marine sediments according to claim 1, characterized in that: The length of the column body (121) ranges from 15 to 25 cm, and the inner diameter ranges from 6 to 10 mm.

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