A positively charged biomass carbon dot and a preparation method and application thereof

Abstract

The application discloses a kind of positive charge biomass carbon dots and preparation method and application thereof, belong to biomass carbon dot technical field.The application provides a kind of method for preparing nanoscale biomass carbon dots with positive charge using agricultural waste-tomato seedling, and the preparation method is simple and easy to operate, and the equipment requirement is low, material source is extensive, cheap and easy to obtain, no toxic chemical reagent is used, and it is environment-friendly, and the preparation cost is low;A new way for high-value utilization of tomato seedling is provided;The biomass carbon dots prepared are approximately spherical in shape, uniform in size, well dispersed, and have no aggregation phenomenon, with an average diameter of 10 nm and an average potential of +42.03 mV;Good hydrophilicity, strong stability;With blue, green, red and other fluorescent properties.

Description

Technical Field

[0001] This invention relates to the field of biomass carbon dot technology, and in particular to a positively charged biomass carbon dot, its preparation method, and its application. Background Technology

[0002] Positively charged carbon nanodots, due to the positive charge on their surface, can efficiently bind to negatively charged cell membranes, showing broad application prospects in fields such as biomedicine and agriculture. Their main known applications can be summarized as follows: 1) Precise antibacterial action and wound healing. This is one of the most researched areas. Positively charged carbon dots can adhere firmly to the surface of negatively charged bacteria through electrostatic interactions, disrupting their cell membrane integrity and thus effectively killing bacteria, including drug-resistant bacteria. Utilizing this property, positively charged carbon dots can be made into wound dressings or hydrogels to accelerate the healing of infected wounds.

[0003] 2) An ideal carrier for gene therapy. The key to gene therapy lies in the safe and efficient delivery of therapeutic genes into cells. The positive charge on the surface of positively charged carbon dots can tightly "grab" negatively charged DNA / RNA, forming a stable complex. It can not only efficiently carry genes across the cell membrane, but also has low toxicity and good biocompatibility.

[0004] 3) High-precision bioimaging and sensing. Thanks to their bright fluorescence and strong ability to penetrate biological membranes, positively charged carbon dots are excellent fluorescent probes. They can penetrate cell membranes and even tissue barriers to enter living cells or organisms, enabling specific labeling and real-time dynamic imaging of key substances such as DNA and RNA.

[0005] 4) Applications in Innovative Agriculture. Research shows that positively charged carbon dots have a higher affinity for plant cells (protoplasts) than negatively charged carbon dots. They can interact with specific components in the plant cell wall (such as pectin), causing changes in cell membrane morphology, thereby allowing them to enter plant cells in a specific manner. This discovery offers new possibilities for using carbon dots to deliver pesticides or genes to plants in the future, achieving precision agriculture.

[0006] However, biomass carbon dots prepared using existing technologies are negatively charged and exhibit limited fluorescence properties, primarily absorbing ultraviolet light and emitting blue light. Furthermore, modifying biomass carbon dots is quite difficult. Therefore, positively charged nanomaterials can only be synthesized using chemical materials. Summary of the Invention

[0007] The purpose of this invention is to provide a positively charged biomass carbon dot, its preparation method, and its application, in order to solve the problem that most biomass carbon dots prepared from straw are negatively charged and have limited fluorescence properties.

[0008] To achieve the above objectives, the present invention provides a method for preparing positively charged biomass carbon dots, comprising the following steps: S1. After removing mud and impurities from the tomato vines, dry them, crush and sieve them to obtain tomato straw powder; S2. Mix tomato straw powder, polyethyleneimine and deionized water thoroughly, sonicate, and then place in a reaction vessel to react; S3. Centrifuge the mixture after reaction S2, take the supernatant and filter it through a 0.22μm filter membrane. Take the liquid that has passed through the membrane and concentrate it to obtain an aqueous solution of positively charged biomass carbon dots. After freeze-drying, obtain solid positively charged biomass carbon dots.

[0009] Preferably, in step S1, the drying process involves air drying, sun drying, or drying in an oven at 50-70°C; the sieving process involves passing the material through a 40-60 mesh sieve.

[0010] Preferably, the mass-to-volume ratio of tomato straw powder, polyethyleneimine, and deionized water in S2 is 1 g: 100 mL.

[0011] Preferably, the ultrasound in S2 is 150W, 40kHz / Hz ultrasound for 20-40 minutes.

[0012] Preferably, the reaction conditions in the reactor during S2 are heating at 180°C for 6-10 hours.

[0013] Positively charged biomass carbon dots prepared by the method described above.

[0014] The application of the positively charged biomass carbon dots as described above in the preparation of foliar fertilizer, wherein the foliar fertilizer uses the positively charged biomass carbon dots as an active ingredient.

[0015] The application of positively charged biomass carbon dots as described above in the preparation of imaging materials, wherein the imaging materials use the positively charged biomass carbon dots as an effective component.

[0016] The application of positively charged biomass carbon dots in plant gene genetic transformation, as described above.

[0017] The application of positively charged biomass carbon dots as described above in the preparation of gene therapy vectors.

[0018] Therefore, the positively charged biomass carbon dots, their preparation method, and applications provided by this invention have the following specific technical effects: (1) This invention provides a method for preparing positively charged, nanoscale biomass carbon dots using agricultural waste—tomato seedlings. The preparation method is simple and easy to operate, requires low equipment, and the materials are widely available, inexpensive and readily available. It does not use toxic chemical reagents, is environmentally friendly, and has low preparation costs; thus, it provides a new way for the high-value utilization of tomato seedlings. (2) The biomass carbon dots prepared by the preparation method provided in this invention are approximately spherical in shape, uniform in size, well dispersed, and without agglomeration. The average diameter is 10 nm and the average potential is +42.03 mV. They have good hydrophilicity and strong stability. They also have multiple fluorescent properties such as blue, green and red.

[0019] The technical solution of the present invention will be further described in detail below with reference to the accompanying drawings and embodiments. Attached Figure Description

[0020] To more clearly illustrate the technical solutions of the embodiments of the present invention, the drawings used in the description of the embodiments of the present invention will be briefly introduced below. Obviously, the drawings described below are only some embodiments of the present invention. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.

[0021] Figure 1 These are TEM images and particle size distribution maps of PEI-CDs in Test Example 1 of this invention; where A is the TEM image and B is the particle size distribution map. Figure 2 These are the Fourier transform infrared spectroscopy results of PEI-CDs in Test Example 1 of this invention; Figure 3 This is the X-ray diffraction pattern of PEI-CDs in Test Example 1 of this invention; Figure 4 This is the Zeta potential analysis result of PEI-CDs in Test Example 1 of this invention; where CD is the biomass carbon dots prepared in Comparative Example 1; Figure 5 This is the fluorescence visible absorption spectrum of PEI-CDs in Test Example 2 of this invention; Figure 6 This is the UV-Vis absorption spectrum of PEI-CDs in Test Example 2 of this invention; Figure 7 These are photographs of plant leaves sprayed with PEI-CDs under ultraviolet light in Test Example 2 of this invention; where A and B are two replicates. Figure 8 These are photographs of melon plants sprayed with PEI-CDs in Test Example 3 of this invention; where A represents the phenotype of plants treated with 5 mg / L; and B represents the phenotype of plants treated with 10 mg / L. Detailed Implementation

[0022] The technical solution of the present invention will be further described below with reference to the accompanying drawings and embodiments.

[0023] To make the objectives, technical solutions, and advantages of this application clearer, more thorough, and more complete, the technical solutions of the present invention will be clearly and completely described below with reference to the accompanying drawings and embodiments. The following detailed descriptions are all illustrations of embodiments, intended to provide further detailed explanation of the present invention. Unless otherwise specified, all technical terms used in this invention have the same meaning as commonly understood by one of ordinary skill in the art to which this application pertains.

[0024] The instruments, equipment, reagents and materials used in the embodiments were all obtained through commercial means; the methods and steps not described in detail in the embodiments are all conventional techniques in the art.

[0025] Example 1 The specific steps for preparing a positively charged biomass carbon dot are as follows: (1) Take disease-free tomato seedlings (including straw and leaves, but excluding underground parts) that have been pulled up after the growing season. After rinsing to remove mud and sand, sun-dry (or oven-dry or air-dry, pay attention to ventilation to prevent mold and rot when air-drying) and then crush them and pass them through a 60-mesh sieve to obtain tomato straw powder.

[0026] (2) Accurately weigh 1g of tomato straw powder and 1g of polyethyleneimine (the mass ratio of tomato straw powder to polyethyleneimine is 1:1), add 100mL of deionized water (the mass ratio of tomato straw powder to deionized water is 1:100), stir and mix well, and then continue to sonicate (150W, 40kHz) for 30min; transfer the ultrasonically mixed liquid to the reaction vessel, and then place it in a high-temperature drying oven to heat to 180℃ within 20min, and keep it at 180℃ for 8h.

[0027] (3) Centrifuge the mixture obtained in step (2) at 4000 r / min for 20 min, collect the supernatant and filter it through a 0.22 μm filter membrane under reduced pressure (-0.1 MPa to -0.08 MPa) to obtain an orange-yellow liquid product.

[0028] (4) The liquid product obtained in step (3) is frozen at low temperature, and after freezing, it is placed in a freeze dryer and freeze-dried into PEI-CD powder. The powder is collected and stored at 4°C for later use.

[0029] Example 2 The specific steps for preparing a positively charged biomass carbon dot are as follows: (1) Take disease-free tomato seedlings (including straw and leaves, but excluding underground parts) that have been pulled up after the growing season. After rinsing to remove mud and sand, sun-dry (or oven-dry or air-dry, pay attention to ventilation to prevent mold and rot when air-drying) and then crush them and pass them through a 60-mesh sieve to obtain tomato straw powder.

[0030] (2) Accurately weigh 1g of tomato straw powder and 0.5g of polyethyleneimine (the mass ratio of tomato straw powder to polyethyleneimine is 1:0.5), add 100mL of deionized water (the mass ratio of tomato straw powder to deionized water is 1:100), stir and mix well, and then treat with ultrasound (150W, 40kHz) for 30min; transfer the suspension to the reaction vessel, and then place it in a high-temperature drying oven and heat it to 180℃ within 20min, and keep it at 180℃ for 8h.

[0031] (3) Centrifuge the mixture obtained in step (2) at 4000 r / min for 20 min, collect the supernatant and filter it through a 0.22 μm filter membrane under reduced pressure (-0.1 MPa to -0.08 MPa) to obtain an orange-yellow liquid product.

[0032] (4) The liquid product obtained in step (3) is frozen at low temperature, and after freezing, it is placed in a freeze dryer and freeze-dried into PEI-CD powder. The powder is collected and stored at 4°C.

[0033] Example 3 The specific steps for preparing a positively charged biomass carbon dot are as follows: (1) Take disease-free tomato seedlings (including straw and leaves, but excluding underground parts) that have been pulled up after the growing season. After rinsing to remove mud and sand, sun-dry (or oven-dry or air-dry, pay attention to ventilation to prevent mold and rot when air-drying) and then crush them and pass them through a 60-mesh sieve to obtain tomato straw powder.

[0034] (2) Accurately weigh 1g of tomato straw powder and 2g of polyethyleneimine (the mass ratio of tomato straw powder to polyethyleneimine is 1:2), add 100ml of deionized water (the mass ratio of tomato straw powder to deionized water is 1:100), stir and mix well, and then continue to sonicate (150W, 40kHz) for 30min; transfer the suspension to the reaction vessel, and then place it in a high temperature drying oven and heat it to 180℃ within 20min, and keep it at 180℃ for 8h.

[0035] (3) Centrifuge the mixture obtained in step (2) at 4000 r / min for 20 min, collect the supernatant and filter it through a 0.22 μm filter membrane under reduced pressure (-0.1 MPa to -0.08 MPa) to obtain an orange-yellow liquid product.

[0036] (4) The liquid product obtained in step (3) is frozen at low temperature, and after freezing, it is placed in a freeze dryer and freeze-dried into PEI-CD powder. The powder is collected and stored at 4°C for later use.

[0037] Comparative Example 1 The conventional method for preparing biomass carbon dots from tomato straw (application publication number CN119430153A) involves the following steps: (1) Wash and dry the tomato vine stalks, then crush them into powder using a pulverizer and sieve them to obtain tomato stalk powder.

[0038] (2) Weigh 3g of tomato straw powder and 300mL of double-distilled water and add them to a beaker (1:100). Treat the mixture with ultrasound (150W power, 40kHz frequency) for 30min. Transfer the suspension to a 500mL polyvinyl fluoride reactor and heat it at 200℃ for 6h. Allow the reactor to cool to room temperature.

[0039] (3) Centrifuge the resulting mixture at 8000 r / min for 15 min. After centrifugation, collect the supernatant and filter it under reduced pressure through a 0.22 μm filter membrane to obtain an orange-yellow liquid product.

[0040] (4) Transfer the obtained product into a dialysis bag with a molecular weight cutoff of 3500 μm, place it in a large beaker containing deionized water, dialyze for 3 days, change the water every 24 hours, until the liquid is basically colorless.

[0041] (5) The solution in the dialysis bag is tomato straw carbon dots (TS-CDs). After freezing, it is placed in a freeze dryer and freeze-dried into carbon dot powder. The powder is collected and stored at 4°C for later use.

[0042] Test Example 1 The positively charged biomass carbon dots (PEI-CDs) prepared in Example 1 were characterized as follows: (1) The structure, size and dispersion of PEI-CDs were observed using TEM, and the results are as follows: Figure 1 As shown, part A is a TEM image, which shows that the prepared PEI-CDs are approximately spherical in shape, uniform in size, well dispersed, and without agglomeration. Part B is a particle size distribution map after analyzing the TEM image using ImageJ, which shows that the average diameter of TS-CDs is 10 nm, which meets the particle size requirements of nanomaterials.

[0043] (2) Fourier transform infrared spectroscopy was performed on the prepared PEI-CDs. Because the vibrations of different chemical bonds in PEI-CDs have specific absorption frequencies in the infrared region, the structure of PEI-CDs can be determined by observing the position and intensity of each peak in the spectral image. The results are as follows: Figure 2 As shown, the prepared PEI-CDs were at 3451.64 cm⁻¹ -1 and 2868.15cm -1 1644.52cm -1 The points are the stretching vibration regions of OH, CH, and C=O, respectively; 1315.87 cm -1 This is the OH bending stretching vibration zone; 1121.07cm -1The surface contains stretching vibrations of CO and CN and single-bond skeletal vibrations of C and C. These abundant surface functional groups can enhance hydrophilicity and stability in aqueous systems, promoting further functionalization and application of PEI-CDs.

[0044] (3) X-ray diffraction (XRD) analysis was performed on the prepared PEI-CDs. Generally, the XRD spectrum of amorphous materials is a straight line. The diffuse peaks that appear at the low 2θ angle are usually composed of liquid solids and gas solids. The results are as follows: Figure 3 As shown, the characteristic peaks of carbon quantum dots typically appear in the 2θ angle range of 20-30°. Among them, the strongest peak is usually located around 25°. A very broad diffraction pattern appears near the peak value of 25°, thus proving that the prepared PEI-CDs are carbon materials with an amorphous structure, consistent with the properties of carbon dots.

[0045] (4) The zeta potential of the prepared PEI-CDs was analyzed, and the results are as follows: Figure 4 As shown, the average potential of PEI-CDs is +42.03mV.

[0046] Test Example 2 The fluorescence properties of the PEI-CDs prepared in Example 1 were tested, as follows: (1) The luminescence characteristics of the prepared PEI-CDs under different excitation wavelengths were investigated, and the results are as follows: Figure 5 As shown, the prepared PEI-CDs absorb ultraviolet light at 370 nm and emit blue light at 465 nm, absorb ultraviolet light at 430 nm and emit green light at 520 nm, and have a weak red light at 640 nm. Due to the change in functional groups, the fluorescence of PEI-CDs is diverse.

[0047] (2) The UV-Vis absorption spectra of the prepared PEI-CDs were examined, and the results are as follows: Figure 6 As shown, the absorbance of the prepared PEI-CDs gradually decreases with increasing excitation wavelength in the ultraviolet wavelength range of 200-800 nm. It has strong absorption capacity in the ultraviolet region of 200-470 nm and has three relatively broad diffraction peaks, corresponding to blue, green and red light in the visible fluorescence absorption spectrum, thus exhibiting superior optical performance.

[0048] The prepared PEI-CD was prepared into a 20 mg / L solution and sprayed evenly on the surface of melon seedlings using a 50 mL spray bottle. After 48 hours, it showed strong blue-green fluorescence after being irradiated with ultraviolet light, indicating that the prepared PEI-CDs have strong fluorescence and stable fluorescence characteristics, and can remain on the plant surface for a long time. Figure 7).

[0049] Test Example 3 The fluorescence properties of the PEI-CDs prepared in Example 1 were tested, as follows: 'Yangjiaomi' melon seeds with plump kernels were selected and soaked in warm water to promote germination. Then, seeds with uniform white sprouts were selected and sown into a substrate (a mixture of peat moss, vermiculite, and perlite in a volume ratio of 6:2:2, with water added and stirred until it can be formed into a clump by hand but crumbles upon falling, with an organic matter content of 45% and a pH of 5.8-6.5). The melons were grown under conditions of 60% relative humidity, 26℃, a photoperiod of 16h / 8h (light / dark), and a light intensity of 22000Lx. When the first true leaf unfolded, the melons were sprayed with 5 mg / L and 10 mg / L solutions (5 mg and 10 mg of PEI-CD powder obtained in step 4 of Example 1 were dissolved in 1L of deionized water) when the fifth true leaf appeared (or when the desired phenotype appeared). The spraying was repeated every 5 days for 30 consecutive days. Pure water was used as a blank control group (Control), and TS-CDs prepared in Comparative Example 1 were used as a control group (CD group). Each treatment was replicated three times. Observe and statistically analyze the plant height and growth of each treatment group. Results are as follows: Figure 8 As shown, compared with the blank control group, the plants treated with 5 mg / L and 10 mg / L TS-CDs and PEI-CDs showed better growth after treatment with 10 mg / L. Furthermore, it was found that at the same concentration, the melon plants treated with PEI-CDs grew significantly better than those treated with TS-CDs, and to a certain extent, the seedling vigor rate of melons was enhanced.

[0050] Therefore, this invention provides a method for preparing positively charged, nanoscale biomass carbon dots using agricultural waste—tomato straw. The preparation method is simple and easy to operate, requires minimal equipment, uses widely available and inexpensive materials, does not use toxic chemical reagents, is environmentally friendly, and has low preparation costs. It provides a new approach for the high-value utilization of tomato seedlings. The prepared biomass carbon dots are approximately spherical in shape, uniform in size, well-dispersed, and do not agglomerate, with an average diameter of 10 nm and an average potential of +42.03 mV. They exhibit good hydrophilicity and strong stability, and possess multiple fluorescent properties, including blue, green, and red.

[0051] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention and not to limit them. Although the present invention has been described in detail with reference to preferred embodiments, those skilled in the art should understand that modifications or equivalent substitutions can still be made to the technical solutions of the present invention, and these modifications or equivalent substitutions cannot cause the modified technical solutions to deviate from the spirit and scope of the technical solutions of the present invention.

Claims

1. A method for preparing positively charged biomass carbon dots, characterized in that, The steps are as follows: S1. After removing mud and impurities from the tomato vines, dry them, crush and sieve them to obtain tomato straw powder; S2. Mix tomato straw powder, polyethyleneimine and deionized water thoroughly, sonicate, and then place in a reaction vessel to react; S3. Centrifuge the mixture after reaction S2, take the supernatant and filter it through a 0.22μm filter membrane. Take the liquid that has passed through the membrane and concentrate it to obtain an aqueous solution of positively charged biomass carbon dots. After freeze-drying, obtain solid positively charged biomass carbon dots.

2. The method for preparing positively charged biomass carbon dots according to claim 1, characterized in that: In S1, drying is done by air drying, sun drying, or drying in an oven at 50-70℃; sieving is done by passing through a 40-60 mesh sieve.

3. The method for preparing positively charged biomass carbon dots according to claim 1, characterized in that: The mass-to-volume ratio of tomato straw powder, polyethyleneimine, and deionized water in S2 is 1 g: 100 mL.

4. The method for preparing positively charged biomass carbon dots according to claim 1, characterized in that: The ultrasound in S2 is 150W, 40kHz ultrasound for 20-40 minutes.

5. The method for preparing positively charged biomass carbon dots according to claim 1, characterized in that: The reaction conditions in the S2 reactor are heating at 180°C for 6-10 hours.

6. Positively charged biomass carbon dots prepared by the preparation method according to any one of claims 1-5.

7. The application of the positively charged biomass carbon dots as described in claim 6 in the preparation of foliar fertilizer, characterized in that: The foliar fertilizer uses the positively charged biomass carbon dots as its active ingredient.

8. The application of the positively charged biomass carbon dots as described in claim 6 in the preparation of imaging materials, characterized in that: The imaging material uses the positively charged biomass carbon dots as its active ingredient.

9. The application of the positively charged biomass carbon dots as described in claim 6 in plant gene genetic transformation.

10. The application of the positively charged biomass carbon dots as described in claim 6 in the preparation of gene therapy vectors.

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