In-situ mass spectrometry device for nano-enzyme reaction
By using Fe3O4 nanoparticle spray needle and conductive wire to construct a microreaction cell in the nanoenzyme reaction, and using electrospray ionization technology to conduct in-situ mass spectrometry monitoring, the simplicity of mass spectrometry monitoring and needle blockage problems in nanoenzyme reactions are solved, and fast and simple high-efficiency mass spectrometry detection is achieved.
Patent Information
- Application Number
- CN202422137870.5
- Authority / Receiving Office
- CN · China
- Patent Type
- Utility models(China)
- Current Assignee / Owner
- Filing Date
- 2024-09-02
- Publication Date
- 2025-07-04
- Estimated Expiration
- 2034-09-02
AI Technical Summary
The prior art is difficult to conduct mass spectrometry monitoring in nanoenzyme reactions in real-time, simple and low-cost, and nanoparticle aggregation leads to the problem of needle blockage.
A Fe3O4 nanoparticle plated needle is used to combine conductive wire and high-voltage power supply to build a micro-reaction cell, and an in-situ mass spectrometry monitoring is used to avoid nanoparticles aggregation.
It realizes simplified analysis of nanoenzyme reactions, and can quickly and easily perform mass spectrometry detection at room temperature and pressure, avoid needle blockage, and has high resolution and multi-stage mass spectrometry detection capabilities.
Smart Images
Figure CN223065231U_ABST
Abstract
Description
Technical Field
[0001] The utility model belongs to the technical field of mass spectrometry analysis, and particularly relates to an in-situ mass spectrometry analysis device for nanozyme reaction. Background Art
[0002] Natural enzymes have high catalytic activity and specific selectivity and can be used to catalyze a variety of biochemical reactions. However, the inherent defects of natural enzymes limit their applications, such as harsh storage conditions, instability, low tolerance, difficult separation, and high cost. Since the discovery of Fe3O4 nanoparticles with enzyme activity similar to horseradish peroxidase (HPR) in 2007, with the major progress of nanotechnology, biotechnology, and nanomaterial science, great progress has also been made in the field of nanozymes, and artificial synthetic enzymes based on various nanomaterials have emerged as the times require. Nanozymes have the catalytic activity of natural enzymes, and nanozymes are used as catalysts in various reactions. For example, in the field of environmental protection, the Fenton reaction in the most studied advanced oxidation technologies (AOPs) uses a metal oxide nanomaterial with enzyme catalytic behavior as the catalyst, which has the advantages of good stability, simple preparation process, and low cost.
[0003] Reaction monitoring is a key aspect in a series of chemical environments, and real-time monitoring of reactions is crucial for many pharmaceutical and synthetic organic chemists. Whether it is traditional chemical, biological production, or (bio) catalysis, reaction monitoring can usually provide important information about the chemical reaction mechanism. Currently, the spectral technologies used for reaction monitoring mainly include Fourier transform infrared spectroscopy (FT-IR), nuclear magnetic resonance (NMR), and infrared spectroscopy (IR). These technologies usually have rich information and can reveal information about chemical composition and physical form. However, these monitoring technologies usually require pretreatment of the reaction solution. For example, IR requires purification and dehydration of the sample; NMR requires good solubility of the sample in a certain deuterated solvent, etc.
[0004] Mass spectrometry is one of the widely used analytical technologies at present. It can accurately determine the elemental composition and structural information of organic molecules, and has the advantages of good accuracy, high sensitivity, and fast signal response speed, and is widely used in various fields. Atmospheric pressure mass spectrometry technology has unique advantages in chemical reaction monitoring. By real-time online monitoring of the substrates, intermediates, and products of the reaction system, short-lived intermediates can be captured and the reaction process can be monitored. However, nanoparticles have a strong aggregation tendency in catalytic reactions, and directly detecting the aggregated nanoparticles by mass spectrometry may clog the injection needle. Therefore, there is an urgent need to explore a device that is applicable, simple to operate, low in cost, and can monitor the reaction process in real time for monitoring the nanozyme reaction process. Content of the Utility Model
[0005] The object of the present utility model is to provide an in-situ mass spectrometry analysis device for nanozyme reaction, and to provide a spray needle coated with nanoparticles, and use electrospray ionization (ESI) technology to monitor the reaction in real time, so as to solve the problems existing in the above-mentioned background technology.
[0006] To achieve the above object, the present application is realized through the following technical solutions:
[0007] An in-situ mass spectrometry analysis device for nanozyme reaction, comprising a spray needle coated with Fe3O4 nanoparticles and a mass spectrometer used in cooperation with the spray needle coated with Fe3O4 nanoparticles; after adding the reaction solution, the spray needle coated with Fe3O4 nanoparticles is placed at the entrance of the mass spectrometer, the tail of the spray needle coated with Fe3O4 nanoparticles is inserted with a conductive iron wire, one end of the conductive iron wire is in contact with the reaction solution in the spray needle coated with Fe3O4 nanoparticles, and the other end of the conductive iron wire is connected to a power supply.
[0008] Further, the tip of the spray needle coated with Fe3O4 nanoparticles is a capillary spray needle with a diameter of 2 μm.
[0009] Further, the distance between the front end of the conductive iron wire and the tip of the spray needle coated with Fe3O4 nanoparticles is 1.5 - 2 cm; the distance between the tip and the sample inlet of the mass spectrometer is 5 - 10 mm.
[0010] Further, a Fe thin layer is covered on the outer surface of the spray needle coated with Fe3O4 nanoparticles and the inner wall of the tip.
[0011] Further, the Fe thin layer deposited on the inner wall of the tip of the spray needle coated with Fe3O4 nanoparticles is annealed to form a nanozyme thin layer of Fe3O4.
[0012] Further, the conductive iron wire is connected to the power supply through an insulating high-voltage wire with an alligator clip.
[0013] Further, the power supply is a high-voltage power supply, and the voltage range is 0.8 - 1.5 kV.
[0014] Further, the power supply is the high-voltage source of an LTQ-Orbitrap Velos linear ion trap - electrostatic field orbitrap combined mass spectrometer.
[0015] The beneficial effects of the present utility model are:
[0016] Compared with the prior art, the beneficial effects of the present utility model are:
[0017] 1. This technical solution is an in-situ mass spectrometry monitoring device for nanozyme reactions. By depositing a thin layer of Fe3O4 nanozyme on the inner wall of the tip of a capillary spray needle, a "tip" micro-reaction cell is constructed, integrating reaction and process monitoring, simplifying the analysis steps of nanozyme reactions, making the analysis process simple and fast. Moreover, the experimental conditions are not stringent and are relatively easy to achieve, enabling immediate mass spectrometry detection.
[0018] 2. This technical solution is an in-situ mass spectrometry monitoring device for nanozyme reactions. Under normal temperature and pressure conditions, it can directly perform mass spectrometry analysis on target analytes in complex samples, and has the advantages of small sample consumption, fast analysis speed, simplicity, high efficiency, environmental protection, etc. The mass spectrometer used is an LTQ-Orbitrap Velos linear ion trap - electrostatic field orbitrap combined mass spectrometer, and its unique high-resolution ability and multi-stage mass spectrometry detection ability can achieve precise analysis of the reaction process.
[0019] 3. This technical solution is an in-situ mass spectrometry monitoring device for nanozyme reactions. By fixing nanoparticles at the tip of the spray needle, it avoids the agglomeration of nanoparticles during the catalytic reaction, causing blockage of the tip of the spray needle. Description of the Drawings
[0020] Figure 1 It is a schematic diagram of the in-situ mass spectrometry monitoring device for nanozyme reactions of the present utility model.
[0021] Figure 2 It is a mass spectrometry result diagram of nicotine degradation analysis using the device of the present utility model.
[0022] Figure 3 It is a diagram of the nicotine degradation reaction process deduced from the mass spectrometry results of this embodiment.
[0023] Description of the Reference Numerals:
[0024] 1. Conductive iron wire; 2. Insulated high-voltage power supply line; 3. Spray needle coated with Fe3O4 nanoparticles; 4. Mass spectrometer. Detailed Description of the Embodiment
[0025] The following elaborates on the technical solution of the present utility model in conjunction with the drawings. The following embodiments are merely exemplary and can only be used to explain and illustrate the technical solution of the present utility model, rather than being construed as a limitation of the technical solution of the present utility model.
[0026] As Figure 1As shown in the figure, the present application provides an in-situ mass spectrometry analysis device for nanozyme reactions, including a capillary needle coated with Fe3O4 nanoparticles and a mass spectrometer used in conjunction with the capillary needle coated with Fe3O4 nanoparticles; after adding the reaction solution, the capillary needle coated with Fe3O4 nanoparticles is placed at the entrance of the mass spectrometer, and the tail of the capillary needle coated with Fe3O4 nanoparticles is inserted into a conductive iron wire. One end of the conductive iron wire is in contact with the reaction solution inside the capillary needle coated with Fe3O4 nanoparticles, and the other end of the conductive iron wire is connected to the high-voltage power supply of the mass spectrometer. In the technical solution of the present application, the mass spectrometer is a conventional device and can be directly obtained through commercial channels. Therefore, in the technical solution of the present application, the structure of the mass spectrometer is not described or explained, and those skilled in the art are well aware of the performance and use of the mass spectrometer.
[0027] In the present application, the capillary needle coated with Fe3O4 nanoparticles is a capillary needle with a tip diameter of 2 μm.
[0028] In the present application, a thin layer of Fe is covered on the outer surface and the inner wall of the tip of the capillary needle coated with Fe3O4 nanoparticles.
[0029] In the present application, the thin layer of Fe deposited on the inner wall of the tip of the capillary needle coated with Fe3O4 nanoparticles is annealed to form a nanozyme thin layer of Fe3O4. The Fe3O4 nanoparticles in this example have enzyme activity similar to that of horseradish peroxidase. The reaction system in this example is a Fenton-like system, and hydroxyl radicals (•OH) with strong oxidation ability are generated in the system in the presence of H2O2 and Fe2+.
[0030] In the present application, the conductive iron wire is connected to the power supply through an insulated high-voltage wire with an alligator clip.
[0031] In the present application, the power supply is a high-voltage power supply, and the voltage range is 0.8 - 1.5 kV.
[0032] In the present application, the power supply is the high-voltage source of an LTQ-Orbitrap Velos linear ion trap - electrostatic field orbitrap combined mass spectrometer.
[0033] The present application also relates to the preparation of the capillary needle coated with Fe3O4 nanoparticles:
[0034] Pulling of the blank needle: A borosilicate capillary is pulled using a P-2000 programmable laser microelectrode puller at certain parameters to obtain a capillary needle with a tip diameter of about 2 μm.
[0035] Specifically, each glass capillary has undergone a comprehensive cleaning process using a plasma cleaner to eliminate the organic impurities present on its surface.
[0036] Specifically, the parameters of the P-2000 programmable laser microelectrode pulling instrument are: Line 1: HEAT=270, FIL=2, VEL=24, DEL=128, PUL=45; Line 2: HEAT=270, FIL=2, VEL=24, DEL=128, PUL=55.
[0037] Fe sputtering: The blank needle drawn in step a) was fixed on a homemade sample stage, with its orifice facing the sputtering source at a certain angle. The rotation speed was set during the coating process to ensure uniform coating thickness at all locations within the needle tip. The magnetron sputtering coater was used in DC mode to first sputter Cr at 50 W for 30 s to form an adhesion layer, and then sputter Fe at 100 W for 3 min to cover the outer surface of the needle and the inside of the needle tip with a thin layer of Fe.
[0038] Annealing treatment of Fe layer: The Fe-coated microtubes were placed in a set pot and then placed in a muffle furnace for treatment. The treatment conditions were as follows: heated from room temperature to 400 °C -450 °C, maintained for a period of time, and then naturally cooled to room temperature.
[0039] Specifically, the angle of the blank needle orifice facing the sputtering source is about 45°; the rotation speed of the coating is 10 r / min.
[0040] During monitoring, the reaction solution was prepared. According to the mechanism of electron transfer and heterogeneous Fenton reaction, the volume of the reaction solution was 1 mL, the concentration ratio of the reaction substrate nicotine and H2O2 was 1:3, and the solvent was distilled water.
[0041] Specifically, in the present technical solution, the concentration of nicotine is 10 μM / L, and the concentration of H2O2 is 30 μM / L.
[0042] Working principle:
[0043] The reaction solution prepared in step b) is injected into the Fe3O4 nanoparticle-coated needle from the tail using a micro-sampler, and then placed in front of the mass spectrometer inlet; the Fe3O4 nanoparticle-coated needle is connected to a high-voltage power supply using a conductive iron wire, and electrospray ionization is triggered by the application of high voltage to achieve mass spectrometry detection.
[0044] Specifically, electrospray ionization is achieved by inserting a conductive iron wire 1 into the tail of a Fe3O4 nanoparticle-coated spray needle 3, and clamping the tail of the conductive iron wire by an insulated high-voltage power line 2 with an alligator clamp at one end; the other end of the high-voltage line is connected to a high-voltage source of a LTQ-Orbitrap Velos linear ion trap-electrostatic field orbital trap combined mass spectrometer 4 for applying a spray voltage to achieve in-situ mass spectrometry monitoring and analysis.
[0045] The distance between the front end of the conductive iron wire and the tip of the Fe3O4 nanoparticle-coated injection needle is 1.5 - 2 cm; the distance between the tip of the Fe3O4 nanoparticle-coated injection needle and the inlet of the mass spectrometer is 5 - 10 mm. The method parameters of the mass spectrometer are as follows: positive ion detection mode; spray voltage: in the range of 0.8 - 1.5 kV; capillary temperature: 350 °C; maximum injection time: 50 ms; collision gas: helium (purity ≥ 99.999%).
[0046] Spectrum results:
[0047] The spectrum is as Figure 2 shown, and its reaction process is as Figure 3 shown. From the data in the mass spectrum, it can be obtained that the nicotine molecule ([M + H] + with m / z 163.1223) first obtains [M] + with m / z 161.1073 through electron transfer, that is, m / z 161.1070 in the mass spectrum. Then, under the oxidative attack of the hydroxyl radical (•OH), the double bond is broken and oxidized to obtain a molecule with m / z 179.1179, that is, m / z 179.1176 in the mass spectrum; m / z 179.1179 then obtains m / z 177.1022 through electron transfer, that is, m / z 177.1017 in the mass spectrum.
[0048] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, and are not intended to limit them; although the present invention has been described in detail with reference to the foregoing embodiments, those of ordinary skill in the art should understand that they can still modify the technical solutions recorded in the foregoing embodiments, or perform equivalent replacements for some of the technologies. However, these modifications or replacements do not make the essence of the corresponding technical solutions deviate from the spirit and scope of the technical solutions of the embodiments of the present invention.
Claims
1. An in-situ mass spectrometry analysis device for nanozyme reactions, characterized in that, It includes a spray needle coated with Fe3O4 nanoparticles and a mass spectrometer used in conjunction with the spray needle coated with Fe3O4 nanoparticles; after adding the reaction solution, the spray needle coated with Fe3O4 nanoparticles is placed at the entrance of the mass spectrometer. The tail of the spray needle coated with Fe3O4 nanoparticles is inserted with a conductive iron wire. One end of the conductive iron wire contacts the reaction solution inside the spray needle coated with Fe3O4 nanoparticles, and the other end of the conductive iron wire is connected to a power supply.
2. The in-situ mass spectrometry analysis device for nanozyme reaction according to claim 1, wherein The tip of the spray needle coated with Fe3O4 nanoparticles is a capillary spray needle with a diameter of 2 μm.
3. The in-situ mass spectrometry analysis device for nanozyme reaction according to claim 1, wherein The distance between the front end of the conductive iron wire and the tip of the spray needle coated with Fe3O4 nanoparticles is 1.5 - 2 cm; the distance between the tip and the sample inlet of the mass spectrometer is 5 - 10 mm.
4. The in-situ mass spectrometry analysis device for nanozyme reaction according to claim 1, wherein, A thin Fe layer is covered on the outer surface of the spray needle coated with Fe3O4 nanoparticles and the inner wall of the tip of the needle.
5. The in-situ mass spectrometry analysis device for nanozyme reaction according to claim 4, wherein The thin Fe layer deposited on the inner wall of the tip of the spray needle coated with Fe3O4 nanoparticles is annealed to form a thin layer of Fe3O4 nanozyme.
6. The in-situ mass spectrometry analysis device for nanozyme reaction according to claim 1, characterized in that The conductive iron wire is connected to the power supply through an insulated high-voltage wire with an alligator clip.
7. The in-situ mass spectrometry analysis device for nanozyme reaction according to claim 1, characterized in that The power supply is a high-voltage power supply, and the voltage range is 0.8 - 1.5 kV.
8. The in-situ mass spectrometry analysis device for nanozyme reaction according to claim 1, wherein The power supply is the high-voltage source of an LTQ-Orbitrap Velos linear ion trap - electrostatic field orbitrap combined mass spectrometer.