A metal-loaded microalgae-based biomass carbon material and its preparation method and application

Abstract

The present invention relates to a metal-loaded microalgae-based biomass carbon material and a preparation method and application thereof, and belongs to the technical field of biomass carbon materials. The preparation method of the present invention comprises the following steps: (1) culturing microalgae with a culture medium to a logarithmic growth phase, and obtaining microalgae powder by centrifugation, desalting, and freeze drying; (2) mixing a microalgae powder solution and a metal ion solution, and obtaining metal-loaded algae powder by centrifugation and freeze drying; and carbonizing the metal-loaded algae powder under an inert atmosphere to obtain the metal-loaded microalgae-based biomass carbon material. The preparation method of the present invention synthesizes NC@Me by directly carbonizing the metal elements modified on the surface of microalgae, which not only retains the rich nitrogen component, but also successfully realizes the functional modification of the metal elements, constructs an electrochemical sensor for highly sensitive detection of biomolecules such as dopamine, and achieves a low detection limit of 3nM and a wide detection range of 0.01-2000μM, and has good selectivity.

Description

Technical Field

[0001] The present invention belongs to the technical field of biomass carbon materials, and in particular relates to a metal-loaded microalgae-based biomass carbon material and a preparation method and application thereof. Background Art

[0002] Electrochemical biosensors usually use nanomaterials to modify electrodes to achieve high sensitivity and high selectivity detection, which can be roughly divided into carbon nanomaterials, metal and metal oxide nanomaterials, etc. Among them, carbon nanomaterials have excellent chemical and thermal stability and are widely used in many fields. When used alone or modified with other nanomaterials, carbon materials have great advantages in sensing due to their fast electron transfer kinetics, excellent conductivity and extremely high biocompatibility. Currently, the most widely used carbon materials are mostly carbon nanotubes, graphene and biomass carbon. The nanocarbon materials obtained by high-temperature carbonization of biomass in nature have a wide source, low price, simple preparation and good biocompatibility, which has attracted much attention. Biomass carbon can be obtained from the thermochemical decomposition of naturally abundant raw materials. It has excellent physical and chemical properties, sustainability and low cost, and has gradually become a substitute for traditional synthetic carbon nanomaterials to produce a greener and more environmentally friendly electrochemical sensing platform. Microalgae have a wide range of sources and contain a variety of structures. The adjustability of specific surface area, porosity, surface charge and element content makes microalgae-based biomass carbon have excellent performance in different research fields from energy materials to environmental remediation. Highly active, surface-functionalized spherical and porous structures are used as effective pollutant adsorbents. For microalgae-based biomass carbon, different internal microstructures and surface properties also have different applications. In electrochemical sensors, compared with widely studied graphene and carbon nanotubes, microalgae-based biomass carbon can achieve better sensing performance in the field of electrochemical sensing due to its unique properties and structures. At present, microalgae can be used as a precursor of biomass carbon to achieve self-doping with nitrogen elements and directly used to construct dopamine sensors. However, in addition to self-doping with nitrogen elements, microalgae as biomass carbon is also expected to further improve the sensing performance of electrochemical biosensors through functionalization treatment methods such as surface modification of metal elements.

[0003] Currently, microalgae can be used as a precursor of biomass carbon to achieve self-doping with nitrogen elements and can be directly used to construct dopamine sensors. However, for the clinical diagnosis and monitoring of dopamine-related diseases, the dopamine content in the extracellular fluid of human tissues is low, and the development of dopamine sensors with lower detection limits requires further research. Summary of the invention

[0004] In order to solve the above technical problems, the present invention provides a metal-loaded microalgae-based biomass carbon material and its preparation method and application. A simple and rapid method is used to prepare a metal-loaded microalgae-based biomass carbon material for constructing high-performance sensor devices for biomolecules such as dopamine. The metal-loaded microalgae-based biomass carbon material can provide more active sites for biomolecules such as dopamine, promote electron transfer efficiency, and thus achieve efficient detection of biomolecules such as dopamine.

[0005] The first object of the present invention is to provide a method for preparing a metal-loaded microalgae-based biomass carbon material, comprising the following steps:

[0006] (1) culturing microalgae in a culture medium until the logarithmic growth phase, and obtaining microalgae powder by centrifugation, desalting, and freeze-drying;

[0007] (2) mixing the microalgae powder solution and the metal ion solution, centrifuging and freeze-drying to obtain metal-loaded algae powder;

[0008] (3) Carbonizing the metal-loaded algae powder described in step (2) under an inert atmosphere to obtain the metal-loaded microalgae-based biomass carbon material.

[0009] In one embodiment of the present invention, in step (1), the culture medium is f / 2 culture medium.

[0010] In one embodiment of the present invention, in step (1), the accession amount of microalgae in the culture medium is 5%-20%; this accession amount is just suitable for a relatively good growth state. If the accession amount is too low, the state of the microalgae will be poor and the growth will be slow. If the accession amount is too high, the growth will be too fast and will also affect the state of the algae.

[0011] In one embodiment of the present invention, in step (1), the microalgae is one or more of Chlorella vulgaris, Chrysophyte 8701, IMET-1 and Menispermum officinale, and the nitrogen content of the microalgae is relatively high.

[0012] In one embodiment of the present invention, in step (2), the concentration of the microalgae powder before mixing is 0.02 g / mL-0.2 g / mL; the concentration of the metal ions after mixing is 0.05 M-0.2 M. The concentration is limited to prevent the modified metal from being unevenly distributed due to too low a concentration, and the performance improvement is not obvious. Too high a concentration will cause metal agglomeration and even carbon material agglomeration, which will also cause performance degradation.

[0013] In one embodiment of the present invention, in step (2), the metal ions are one or more of iron ions, copper ions, manganese ions, cobalt ions, nickel ions, gold ions and platinum ions.

[0014] In one embodiment of the present invention, in steps (1) and (2), the centrifugal speed is 2000 rpm-6000 rpm. The setting of the centrifugal speed can better collect the material. If the speed is too low, the centrifugation is incomplete and the material is lost. If the speed is too high, the morphology of the microalgae will be destroyed.

[0015] In one embodiment of the present invention, in step (3), the carbonization temperature is 600°C-900°C and the carbonization time is 1.5h-3h. If the carbonization temperature is too low, the carbonization is incomplete, and if the temperature is too high, the carbon material structure will collapse and affect the modification effect of N and metal.

[0016] In one embodiment of the present invention, in step (3), the inert atmosphere is argon.

[0017] The second object of the present invention is to provide a metal-loaded microalgae-based biomass carbon material prepared by the method described above.

[0018] The third object of the present invention is to provide a use of the metal-loaded microalgae-based biomass carbon material as an electrochemical biosensor.

[0019] The technical solution of the present invention has the following advantages over the prior art:

[0020] (1) The preparation method described in the present invention modifies the metal elements on the surface of microalgae by a direct surface method, and synthesizes NC@Me (Me represents metal, including transition metals and precious metals) by direct carbonization. NC@Me not only retains the rich nitrogen components in microalgae, but also successfully incorporates metal elements to construct an electrochemical sensor for highly sensitive detection of biological molecules such as dopamine. The dopamine electrochemical sensor constructed based on NC@Fe achieves a low detection limit of 3nM and a wide detection range of 0.01-2000μM, and has good selectivity.

[0021] (2) The preparation method described in the present invention makes the carbon material tend to be highly graphitized under low temperature conditions by adding metal elements, increases the defects in the carbon material structure, promotes the formation of catalytic active sites, and adjusts the pore structure of the material to increase the specific surface area to achieve rapid charge transfer.

[0022] (3) The preparation method of the metal-loaded microalgae-based biomass carbon material described in the present invention is simple, and can efficiently and quickly realize the functionalization treatment of the microalgae-based biomass carbon material. The sensing enhancement for biological molecules such as dopamine is because the addition of metal inhibits the aggregation of carbon particles and promotes the microalgae-based biomass carbon material to be highly graphitized at a lower temperature. The introduction of metal can form uniformly dispersed metal elements while adjusting the pore structure of the material to provide more catalytic active centers for the specific detection of target biological molecules. The nitrogen-doped porous carbon nano-based material formed by the microalgae's own nitrogen element during the carbonization process can effectively increase the interface electron transfer rate to provide a guarantee for the rapid response of biological molecules such as dopamine. BRIEF DESCRIPTION OF THE DRAWINGS

[0023] In order to make the content of the present invention more clearly understood, the present invention is further described in detail below according to specific embodiments of the present invention in conjunction with the accompanying drawings, wherein:

[0024] Figure 1 Characterization diagrams of different materials in Test Example 1 of the present invention; wherein A is the TEM diagram of NC; B is the TEM diagram of NC@Fe; CE are element distribution diagrams of C, N, and Fe in NC@Fe, respectively;

[0025] Figure 2 This is the electrochemical response test of NC and NC@Fe to dopamine in Test Example 2 of the present invention; wherein A is the CV response of NC@Fe to different concentrations of dopamine; B is the CV response of NC@Fe to different concentrations of dopamine;

[0026] Figure 3 This is the selectivity test of NC@Fe for dopamine in Test Example 2 of the present invention. DETAILED DESCRIPTION

[0027] The present invention is further described below in conjunction with the accompanying drawings and specific embodiments so that those skilled in the art can better understand the present invention and implement it, but the embodiments are not intended to limit the present invention.

[0028] In the present invention, unless otherwise specified, the formula of f / 2 medium is as follows:

[0029] (1) Accurately weigh 7.5 g of NaNO 3 and 0.5 g of NaH 2 PO 4 ·H 2 O, dissolve them with about 90 mL of ultrapure water respectively, wait for them to be completely dissolved and make up to 100 mL, and sterilize them by high pressure at 121℃ for 20 min.

[0030] (2) Preparation of metal salt mixed solution:

[0031] ① Accurately weigh 0.98g of CuSO 4 ·5H 2 O, 0.63 g Na 2 MoO 4 ·2H 2 O, 2.2 g ZnSO 4 7H 2 O, 1.0 g CoCl 2 6H 2 O and 18.0 g of MnCl 2 ·4H 2 O, respectively, dilute to 100 mL with ultrapure water and wait for use.

[0032] ② Weigh 0.315g of FeCl 3 6H 2 O and 0.436 g Na 2 EDTA·2H 2 O was dissolved in 90 mL of ultrapure water, and 1 mL of each of the five reagents prepared in step ① was added to the mixed solution. Finally, the mixed solution was fixed to 100 mL with ultrapure water, sterilized at 121 °C for 20 min, and stored in a dark place.

[0033] (3) Preparation of vitamin mixture:

[0034] ① Accurately weigh 0.01g of biotin and 0.01g of vitamin B12 and dilute to 100mL with ultrapure water for use.

[0035] ② Weigh 0.02g of thiamine hydrochloride and dissolve it in 90mL, and add 1mL of each of the two solutions prepared in step ①. Add ultrapure water to make the volume 100mL, filter and sterilize with a 0.22μm sterile filter membrane, and finally store the solution in a 4℃ refrigerator away from light.

[0036] (4) Prepare 950 mL of filtered natural seawater, sterilize it under high pressure at 110°C for 30 min, and add 1 mL of NaNO prepared in step (1) to it. 3 and NaH 2 PO 4 solution, 1 mL of the mixed metal solution prepared in step (2) and 0.5 mL of the vitamin mixture prepared in step (3), and finally fixed to 1 L with sterilized seawater to obtain an f / 2 culture medium suitable for the growth of microalgae.

[0037] Example 1

[0038] A metal-loaded microalgae-based biomass carbon material and a preparation method thereof, specifically comprising the following steps:

[0039] (1) Cultivation and collection of microalgae: 50 mL of Nannochloropsis algae was inoculated into a 1 L conical flask with 450 mL of f / 2 culture medium according to the inoculation volume of 10%. The inoculated culture medium was placed in a light incubator with the following settings: light intensity 100 μmolphotons / m 2 ·s, the light-dark time ratio is 1:1, the temperature is 25±2℃, and the microalgae will be in the late logarithmic growth stage after growing in the culture medium for about a week, and it will be harvested at this time. The microalgae liquid is concentrated by low-speed centrifugation at 4000rpm, and after removing the seawater salt, the obtained algae mud is freeze-dried to obtain microalgae powder.

[0040] (2) Preparation of algae powder loaded with metal Fe: The algae powder was dissolved in water to prepare an algae solution with a concentration of 0.1 g / mL, and FeCl was added to the solution. 3 The solution was made to have a final concentration of 0.1 M, mixed evenly and stirred at room temperature for 40 min. The algae mud mixed with metal element Fe was collected by low-speed centrifugation at 4000 rpm and freeze-dried to obtain algae powder loaded with metal Fe;

[0041] (2) Preparation of microalgae-based biomass carbon material loaded with metal Fe: The algae powder loaded with metal Fe was carbonized at 600 °C in a tube furnace under an argon atmosphere for 2 h, ground, and washed with 1 M hydrochloric acid to remove excess metal impurities to obtain a microalgae-based biomass carbon material loaded with metal Fe (NC@Fe).

[0042] Example 2

[0043] A metal-loaded microalgae-based biomass carbon material and a preparation method thereof, specifically comprising the following steps:

[0044] (1) Cultivation and collection of microalgae: 50 mL of Nannochloropsis algae was inoculated into a 1 L conical flask with 450 mL of f / 2 culture medium according to the 10% inoculation volume. The inoculated culture medium was placed in a light incubator with the following settings: light intensity 100 μmol photons / m 2 ·s, the light-dark time ratio is 1:1, the temperature is 25±2℃, and the microalgae will be in the late logarithmic growth stage after growing in the culture medium for about a week, and it will be harvested at this time. The microalgae liquid is concentrated by low-speed centrifugation at 4000rpm, and after removing the seawater salt, the obtained algae mud is freeze-dried to obtain microalgae powder.

[0045] (2) Preparation of algae powder loaded with metal Mn: The algae powder was dissolved in water to prepare an algae solution with a concentration of 0.1 g / mL, and MnCl was added to the solution. 2 The solution was made to have a final concentration of 0.1 M, mixed evenly and stirred at room temperature for 40 min. The algae mud mixed with the metal element Mn was collected by low-speed centrifugation at 4000 rpm and freeze-dried to obtain algae powder loaded with metal Mn;

[0046] (2) Preparation of microalgae-based biomass carbon material loaded with metal Mn: The algae powder loaded with metal Mn was carbonized at 600 °C in a tube furnace under an argon atmosphere for 2 h, ground, and washed with 1 M hydrochloric acid to remove excess metal impurities to obtain microalgae-based biomass carbon material loaded with metal Mn (NC@Mn).

[0047] Example 3

[0048] A metal-loaded microalgae-based biomass carbon material and a preparation method thereof, specifically comprising the following steps:

[0049] (1) Cultivation and collection of microalgae: 110 mL of Nannochloropsis spp. was inoculated into a 1 L conical flask with 450 mL of f / 2 medium according to an inoculation volume of about 10%. The inoculated medium was placed in a light incubator with a light intensity of 100 μmol photons / m 2 ·s, the light-dark time ratio is 1:1, the temperature is 25±2℃, and the microalgae will be in the late logarithmic growth stage after growing in the culture medium for about a week, and it will be harvested at this time. The microalgae liquid is concentrated by low-speed centrifugation at 6000rpm, and after removing the seawater salt, the obtained algae mud is freeze-dried to obtain microalgae powder.

[0050] (2) Preparation of algae powder loaded with metal Fe: The algae powder was dissolved in water to prepare an algae solution with a concentration of 0.2 g / mL, and FeCl was added to the solution. 3 The solution was made to have a final concentration of 0.2 M, mixed evenly and stirred at room temperature for 60 min. The algae mud mixed with metal element Fe was collected by low-speed centrifugation at 6000 rpm and freeze-dried to obtain algae powder loaded with metal Fe;

[0051] (2) Preparation of microalgae-based biomass carbon material loaded with metal Fe: The algae powder loaded with metal Fe was carbonized at 900°C in a tube furnace under an argon atmosphere for 3 h, ground, and washed with 2M hydrochloric acid to remove excess metal impurities to obtain a microalgae-based biomass carbon material loaded with metal Fe (NC@Fe).

[0052] Example 4

[0053] A metal-loaded microalgae-based biomass carbon material and a preparation method thereof, specifically comprising the following steps:

[0054] (1) Cultivation and collection of microalgae: 25 mL of microalgae was inoculated into a 1 L conical flask with 450 mL of f / 2 culture medium according to the inoculation volume of about 5%. The inoculated culture medium was placed in a light incubator and the program was set as follows: light intensity 100 μmol photons / m 2·s, the light-dark time ratio is 1:1, the temperature is 25±2℃, and the microalgae will be in the late logarithmic growth stage after growing in the culture medium for about a week, and it is harvested at this time. The microalgae liquid is concentrated by low-speed centrifugation at 2000rpm, and after removing the seawater salt, the obtained algae mud is freeze-dried to obtain microalgae powder.

[0055] (2) Preparation of algae powder loaded with metal Fe: The algae powder was dissolved in water to prepare an algae solution with a concentration of 0.02 g / mL, and FeCl was added to the solution. 3 The solution was made to have a final concentration of 0.05 M, mixed evenly and stirred at room temperature for 20 min. The algae mud mixed with metal element Fe was collected by low-speed centrifugation at 2000 rpm and freeze-dried to obtain algae powder loaded with metal Fe;

[0056] (2) Preparation of microalgae-based biomass carbon material loaded with metal Fe: The algae powder loaded with metal Fe was carbonized at 600°C in a tube furnace under an argon atmosphere for 1.5 h, ground, and washed with 0.2 M hydrochloric acid to remove excess metal impurities to obtain a microalgae-based biomass carbon material loaded with metal Fe (NC@Fe).

[0057] Comparative Example 1

[0058] (1) Cultivation and collection of microalgae: 50 mL of Nannochloropsis algae was inoculated into a 1 L conical flask with 450 mL of f / 2 culture medium according to the 10% inoculation volume. The inoculated culture medium was placed in a light incubator with the following settings: light intensity 100 μmol photons / m 2 ·s, the light-dark time ratio is 1:1, the temperature is 25±2℃, and the microalgae will be in the late logarithmic growth stage after growing in the culture medium for about a week, and it will be harvested at this time. The microalgae liquid is concentrated by low-speed centrifugation at 4000rpm, and after removing the seawater salt, the obtained algae mud is freeze-dried to obtain microalgae powder.

[0059] (2) Preparation of microalgae-based biomass carbon material: The microalgae powder was carbonized at 600 °C in a tube furnace under argon atmosphere for 2 h and ground to obtain microalgae-based biomass carbon material (NC).

[0060] Test Example 1

[0061] Based on Example 1 and Comparative Example 1, the golden algae (NC) without any metal loading and the microalgae-based biomass carbon material (NC@Fe) loaded with metal Fe were characterized. The results are as follows Figure 1 As shown. Figure 1 It can be seen that the control microalgae does not load any metal (NC, Figure 1 A) and carbonized carbon materials after loading 0.1 M metallic Fe (NC@Fe, Figure 1B) shows different microstructures. TEM images show that the Fe element has an important influence on the microstructure of microalgae carbon materials. Figure 1 The TEM image in B clearly shows that Fe atoms are evenly distributed in the microalgae-based biomass carbon material. It also shows that the synthesized NC@Fe completes self-doping with N and combines with externally modified Fe elements to achieve graphitization of biomass at a lower carbonization temperature. At the same time, during the carbonization process, Fe elements will be evenly incorporated into the carbon material to produce more active sites, so that the dual functionalization of Fe and N is completed on the carbon skeleton, promoting rapid response to target molecules. Figure 1 From C-1E, it can be seen that C, N, and Fe elements are evenly distributed on the material.

[0062] Test Example 2

[0063] Based on Example 1 and Comparative Example 1, the electrochemical performance of dopamine sensors constructed by NC and NC@Fe was analyzed. NC and NC@Fe were modified with materials (5 μL 2 mg / mL) on glassy carbon electrodes using a standard three-electrode system to test dopamine biomolecules. The redox reaction at the electrode interface was tested and analyzed using cyclic voltammetry (CV) and differential pulse voltammetry (DPV). The results are shown in Figure 2. Figure 2-3 shown.

[0064] from Figure 2 It can be seen that Figure 2 A shows that NC@Fe obtained by carbonization after loading 0.1M metal Fe on microalgae can detect different concentrations of dopamine. The results show that DPV can detect dopamine as low as 0.01μM ( Figure 2 A), the detection range of the sensor is 0.01-2000μM ( Figure 2 B), it can be calculated that the detection limit of the sensor is as low as 3nM, which is much better than the detection limit of carbon material alone (6nM).

[0065] from Figure 3 It can be seen that by testing different interferents to analyze the selectivity of the NC@Fe modified electrode for detecting dopamine, the sensor has the highest oxidation current response to 100 μM dopamine (DA), a weaker response to the same concentration of uric acid (UA), and almost no response to ascorbic acid (AA) and glucose (Glu), proving that the material has excellent anti-interference ability.

[0066] Obviously, the above embodiments are merely examples for clear explanation and are not intended to limit the implementation methods. For those skilled in the art, other different forms of changes or modifications can be made based on the above description. It is not necessary and impossible to list all the implementation methods here. The obvious changes or modifications derived from these are still within the protection scope of the invention.

Claims

1. Application of a metal-loaded microalgae-based biomass carbon material as an electrochemical biosensor for dopamine detection, characterized in that: The preparation method of the metal-loaded microalgae-based biomass carbon material, The following steps are included: (1) culturing microalgae in a culture medium until the logarithmic growth phase, and obtaining microalgae powder by centrifugation, desalting, and freeze-drying; the microalgae is one or more of Chlorella vulgaris, Chrysophyte 8701, IMET-1, and Crocus ovalis; (2) mixing the microalgae powder solution and the metal ion solution, centrifuging and freeze-drying to obtain metal-loaded algae powder; the metal ions are one or more of iron ions, copper ions, manganese ions, cobalt ions, nickel ions, gold ions and platinum ions; (3) Carbonizing the metal-loaded algae powder described in step (2) under an inert atmosphere to obtain the metal-loaded microalgae-based biomass carbon material.

2. The use according to claim 1, characterized in that: In step (1), the culture medium is f / 2 culture medium.

3. The use according to claim 1, characterized in that: In step (1), the inoculation amount of microalgae in the culture medium is 5%-20%.

4. The use according to claim 1, characterized in that: In step (2), the concentration of the microalgae powder before mixing is 0.02 g / mL-0.2 g / mL; the concentration of the metal ions after mixing is 0.05 M-0.2 M.

5. The use according to claim 1, characterized in that: In steps (1) and (2), the centrifugal rotation speed is 2000 rpm-6000 rpm.

6. The use according to claim 1, characterized in that: In step (3), the carbonization temperature is 600°C-900°C; the carbonization time is 1.5h-3h.

7. The use according to claim 1, characterized in that: In step (3), the inert atmosphere is argon.

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