CN107043097A - Method and its carbon point and the application of carbon point are prepared by mushroom - Google Patents
Method and its carbon point and the application of carbon point are prepared by mushroom Download PDFInfo
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Abstract
It is by 0.08g ~ 0.1g by mushroom powder and water the invention discloses a kind of method that carbon point is prepared by mushroom：After 1mL ratio mixing, 180 200 DEG C of reactions, 57 hours obtained carbon points.The protophyte resource mushroom that the present invention is widely present using nature is cooked carbon source, carbon point is prepared by simple hydro-thermal method, environmental protection, gained carbon point has good fluorescence property, also have the advantages that good light stability, toxicity be low, good biocompatibility, had a good application prospect in terms of cell imaging.
Description
Technical field
The invention belongs to the preparing technical field of luminescent material, it is related to a kind of method that carbon point is prepared by mushroom, further relates to
Carbon point obtained by this method, and the carbon point application.
Background technology
In recent years, as an emerging important member in carbon nanomaterial family, carbon point is received more and more attention.
Except with the excellent optical property similar to traditional semiconductor-quantum-point, carbon point also have good light stability, toxicity it is low,
Traditional quantum dot such as good biocompatibility and environment friendly, preparation condition are gentle, step is simple, abundant raw material is cheap without
Analogous advantage.At present, the research emphasis of carbon point concentrates on the preparation method for finding simplicity and answering in fluorescence analysis
With.
The synthesis of carbon point is generally divided into method from top to bottom（Such as arc discharge method, laser ablation, electrochemical synthesis and acid
Oxidizing process etc.）Method from bottom to top（Such as microwave method, hydro-thermal method and ultrasonic method）Two major classes.From top to bottom method by physics or
By the carbon material of bulk from large to small, size is until nanoscale for the method for chemistry.Method is that have with the carbon containing of small size from bottom to top
Machine thing is carbon source, is changed from small to big by orderly or unordered being assembled of small molecule, so as to reach certain size to prepare
Carbon point.The carbon point that this two classes method is prepared all shows the excellent characteristics of luminescence.Prepare carbon source used in carbon point basic
On can be divided into inorganic carbon source and the major class of organic carbon source two.Inorganic carbon source mainly has candle ash, graphite powder, activated carbon, carbon nanometer
Pipe and carbon fiber etc..It is main using method from top to bottom when preparing carbon point with inorganic carbon source.Organic carbon source has glucose, amino
The nature of the environmental protection such as the carbon containing organic chemical reagent such as acid and grass, pericarp, sweet potato, coffee grounds, capsicum, apple and sugarcane
Product.When preparing carbon point with organic carbon source mainly using method from bottom to top, most common method has hydro-thermal method and microwave method.But
So far, there is not yet preparing the method report of carbon point by carbon source of mushroom class.
The content of the invention
In view of this, carbon source is done it is an object of the invention to the protophyte resource mushroom being widely present using nature,
Carbon point, environmental protection are prepared by simple hydro-thermal method.
Through research, the present invention provides following technical scheme：
1. the method for carbon point is prepared by mushroom, it is characterised in that mushroom powder and water are pressed into 0.08g ~ 0.1g：1mL ratio mixing
Afterwards, 5-7 hours obtained carbon points of 180-200 DEG C of reaction.
It is furthermore preferred that mushroom powder and water are pressed into 0.1g：After 1mL ratio mixing, 200 DEG C of reactions, 6 hours obtained carbon points.
It is preferred that, reaction solution is cooled down, centrifuged, supernatant is collected, with the filtering with microporous membrane that aperture is 0.22 μm, filtrate
After being dialysed with molecular cut off for 100 bag filter, dry, obtain carbon point powder.
It is furthermore preferred that reaction solution is cooled down, 10000rpm is centrifuged 15 minutes, collects supernatant, is 0.22 μm with aperture
Filtering with microporous membrane, after filtrate molecular cut off is dialysed 24 hours for 100 bag filter, freeze-drying obtains carbon point powder.
It is preferred that, the mushroom powder be by new fresh mushroom in after 60 DEG C ~ 80 DEG C dryings 24 ~ 48 hours, with mortar grinder into
Powder.
Carbon point made from the above method is used.
Application of the carbon point in terms of cell imaging.
The beneficial effects of the present invention are：The invention provides a kind of method that carbon point is prepared by mushroom, nature is utilized
The protophyte resource mushroom being widely present is cooked carbon source, and carbon point, environmental protection, gained carbon point tool are prepared by simple hydro-thermal method
There is good fluorescence property, also have the advantages that good light stability, toxicity be low, good biocompatibility, in terms of cell imaging
Have a good application prospect.
Brief description of the drawings
Fig. 1 is prepared the method schematic diagram of carbon point by mushroom.
The sign of Fig. 2 carbon point：（A）The fluorescence excitation and emission spectrum of carbon point, wherein illustration are carbon point dispersion liquid visible
Light（I）And ultraviolet light（II）Under photo；（B）The high resolution TEM figure of carbon point；（C）The infrared spectrum of carbon point；（D）Carbon
The X-ray energy spectrum of point.
Emission spectrum of Fig. 3 carbon point under different excitation wavelengths.
The study on the stability of Fig. 4 carbon point：（A）Saline-alkaline tolerance is investigated；（B）Study on the stability under different pH condition；（C）
Stabilization time is investigated；（D）Study on the stability in different organic reagents.
Toxicity test of the carbon point of Fig. 5 various concentrations to CKK-8 cells.
Fig. 6 is prepared the condition optimizing of carbon point by mushroom：（A）Reaction time optimizes；（B）Reaction temperature optimizes.
Fig. 7（A）NaCl under uviol lamp（It is left）With carbon point（It is right）；（B）NaCl under fluorescent lamp（It is left）With carbon point（It is right）.
Fig. 8（A）The absorbent cotton of carbon point dispersion liquid is not smeared under uviol lamp（It is left）With the absorbent cotton for smearing carbon point dispersion liquid
（It is right）；（B）The filter paper of carbon point dispersion liquid is not smeared under uviol lamp（It is left）With the filter paper for smearing carbon point dispersion liquid（It is right）.
Fluoroscopic image of Fig. 9 carbon point in human A549 cell lines：（A）For cell photograph via bright field；（B）Marked for carbon point
Cell fluorescence is imaged（Excitation wavelength is 488nm）；（C）For light field and fluorescence picture superposition photo.
Embodiment
In order that the object, technical solutions and advantages of the present invention are clearer, below in conjunction with accompanying drawing to the excellent of the present invention
Embodiment is selected to be described in detail.
The main agents used in preferred embodiment are as follows：Mushroom is purchased from It In Beibei, Chongqing Yong Hui supermarkets；Methanol
（Methanol）, ethanol（Ethanol）, acetone（Acetone）, acetic acid（Acetic Acid）, acetonitrile（Acetonitrile）、
Dimethyl sulfoxide (DMSO)（DimethylSulfoxide）, tetrahydrofuran（Tetrahydrofuran ）And trichloroethanes
（Trichloroethane）Purchased from Chengdu Ke Long chemical reagents factory；Disodium hydrogen phosphate（Na2HPO4）, sodium dihydrogen phosphate
（NaH2PO4）, glacial acetic acid（HAC）, phosphoric acid（H3PO4）, boric acid（H3BO3）, sodium hydroxide（NaOH）And sodium chloride（NaCl）It is purchased from
Beijing Ding Guo bio tech ltd；Experimental water is ultra-pure water（Millipore, 18.25 M Ω）.
The key instrument used in preferred embodiment is as follows：FA2004A electronic balances（The limited public affairs of the upper sub- instrument of Nereid's atmospheric electricity
Department）, 25 ml polytetrafluoroethylene (PTFE) hydrothermal reaction kettles（Jinan China Heng Hua Science and Technology Ltd.s）, F-7000 type fluorescence spectrophotometries
Meter（HIT）, Tecnai G2 F20 S-TWIN type transmission electron microscopes（FEI Co. of the U.S.）, ESCALAB
250 model x-ray photoelectron spectroscopy（Thermo companies of the U.S.）, Prestige-21 Fourier infrared spectrographs（Japanese Shimadzu
Company）, PiloFD8-4.3V type freeze driers（Xi Meng international corporations of the U.S.）, DHG-9140A electric heating constant-temperature blowing drying boxes
（Shanghai Qi Xin scientific instrument Co., Ltd）, UPT laboratories Superpure water machine（The Chinese excellent general experimental instruments and equipment limited in Xi'an）,
TG16-W high speed centrifugal machine for minim.
Embodiment 1, carbon point is prepared by carbon source of mushroom
1st, the preparation of carbon point
As shown in figure 1, by new fresh mushroom after 80 DEG C are dried 24 hours, with mortar grinder into powder；The g of mushroom powder 0.6 is taken, plus
Enter the mL of ultra-pure water 6, mix, put in 25 mL polytetrafluoroethylene (PTFE) stainless steel cauldrons, 200 DEG C are reacted 6 hours；Gained reaction solution is cold
But to room temperature, 10000 rpm are centrifuged 15 minutes, collect supernatant, and with the filtering with microporous membrane that aperture is 0.22 μm, filtrate, which is used, cuts
Stay molecular weight（MWCO）For 100 bag filter dialyse 24 hours after, freeze-drying obtain carbon point powder；By carbon point powder with ultrapure
Moisture dissipates, and obtains 1 μ g/mL carbon point dispersion liquid, 4 DEG C of preservations.
2nd, the sign of carbon point
A length of 370 nm of optimum excitation wave of carbon point, optimal launch wavelength is 455 nm（Fig. 2A）, it is in micro- Huang under visible light
Color（Illustration I in Fig. 2A）, hanced cyan fluorescent is sent under 365 nm ultraviolet lights（Illustration II in Fig. 2A）.
The shape characteristic of carbon point is observed using high resolution TEM（Fig. 2 B）, it is seen that carbon point has good disperse
Property, average diameter is 2.3 ± 1.3 nm.
The surface texture of carbon point is analyzed using FTIR spectrum（Fig. 2 C）, 3438 cm-1The peak at place is attributed to O-H and stretched
Contracting vibration, 1678 cm-1The peak at place is attributed to C=O stretching vibrations, 2964 cm-1The peak at place is attributed to N-H stretching vibrations, display
There is the characteristic groups such as carboxyl, hydroxyl, amino in the surface of carbon point.
Surface composition and the elementary analysis of carbon point are carried out using X-ray electron spectrum（Fig. 2 D）, it is seen that mainly deposited on spectrogram
At the peak of C, N and O three types（284.6,399.8 and 531.4 eV are respectively from C 1s, N 1s and O 1s）, illustrate carbon
Point doping N.
The wavelength of fluorescence of carbon point has obvious Red Shift Phenomena with the increase of excitation wavelength, when excitation wavelength increases from 370 nm
450 nm are added to, the launch wavelength of carbon point increases to 450 nm from 370 nm（Fig. 3）.
3rd, the study on the stability of carbon point
（1）Saline-alkaline tolerance is investigated：Carbon point dispersion liquid is taken, different final concentrations are added（0、0.1、0.2、0.3、0.4、0.5、0.6、
0.7、0.8、0.9、1.0 mol/L）Sodium chloride, be incubated, the nm of wavelength 455 at measure fluorescence intensity.As a result such as Fig. 4 A institutes
Show, substantially change does not occur in the case that sodium chloride concentration gradually increases in system for the fluorescence intensity of carbon point, shows that it has
Higher saline-alkaline tolerance.
（2）Study on the stability under different pH condition：Carbon point dispersion liquid is taken, in different pH value（2、3、4、5、6、7、8、
9、10、11、12）Under the conditions of be incubated after, the nm of wavelength 455 at measure fluorescence intensity, as a result as shown in Figure 4 B, the fluorescence of carbon point
Intensity is slightly changed under different pH condition, overall relatively stable, and the optimal pH value that is incubated is 7.
（3）Stabilization time is investigated：Carbon point dispersion liquid is taken, different time is incubated（1、2、3、4、5、6、7、8、9、10、11、12
Hour）Afterwards, fluorescence intensity is determined at the nm of wavelength 455.As a result as shown in Figure 4 C, the fluorescence intensity of carbon point is steady in 12 hours
It is fixed, do not change with time and change.
（4）Study on the stability in different organic reagents：Carbon point dispersion liquid is taken, in different organic reagents（Methanol, ethanol,
Acetone, acetic acid, acetonitrile, dimethyl sulfoxide (DMSO), tetrahydrofuran, trichloroethanes）After middle incubation, fluorescence is determined at the nm of wavelength 455 strong
Degree.As a result as shown in Figure 4 D, carbon is selected keeps stable in most of organic reagents.
4th, the Toxicity test of carbon point
Take 10 mg/mL carbon point dispersion liquids of filtration sterilization, be diluted to fresh 1640 culture medium final concentration be respectively 2.5,
2.0、1.5、1.0、0.5 mg/mL.The CKK-8 cells of growth of taking the logarithm are inoculated in 60 mm Tissue Culture Dish, and totally 6 ware, is put into
Cell culture incubator, which is incubated 24 h, makes cell enter exponential phase, absorbs nutrient solution in each culture dish, is separately added into foregoing difference
The mL of carbon point dilution 3 of concentration, while setting solvent control group（Add 1640 trainings that carbon point dispersion liquid is substituted with physiological saline
The mL of nutrient solution 3）, it is put into cell culture incubator culture 24 hours.As a result as shown in figure 5, even in high dose（2.5mg/mL）With it is long when
Between be incubated（24 hours）Under the conditions of, carbon point also hardly produces toxicity to cell, shows good biocompatibility.
5th, the fluorescence of carbon point
The carbon point powder that freeze-drying is obtained is taken, respectively in uviol lamp with being compared under fluorescent lamp with NaCl.As a result such as Fig. 6 institutes
Show, carbon point powder shows obvious fluorescence under uviol lamp（Fig. 6 A）, there is also obvious color with NaCl under fluorescent light
Difference（Fig. 6 B）.
Carbon point dispersion liquid is applied on absorbent cotton and filter paper respectively, with not smearing the de- of carbon point dispersion liquid under uviol lamp
Fat cotton and filter paper are compared.As a result as shown in fig. 7, under uviol lamp, smear the absorbent cotton and filter paper of carbon point dispersion liquid with not
Smearing the absorbent cotton and filter paper of carbon point dispersion liquid has obvious fluorescent differences.
Embodiment 2, the condition optimizing for preparing by carbon source of mushroom carbon point
1st, the reaction time optimizes
By the methods described of embodiment 1, in 200 DEG C respectively reaction 4, prepare carbon point within 5,6,7,8,9 hours.As a result as shown in Figure 8 A,
The fluorescent effect of 5-7 hours obtained carbon points of reaction is preferable, most preferably 6 hours reaction time.
2nd, reaction temperature optimizes
By the methods described of embodiment 1, respectively at 120,140,160,180,200 DEG C of reactions prepare within 6 hours carbon point.As a result such as Fig. 8 B
Shown, the fluorescent effect of the obtained carbon point of 180-200 DEG C of reaction is preferable, most preferably 200 DEG C of reaction temperature.
Embodiment 3, using the mushroom application of carbon point in terms of cell imaging made from carbon source
By human lung adenocarcinoma cell（A549 cells）6 holes in 96 microwell plates are inoculated in, is added per hole after confining liquid, is put into wet box
In, place into 37 DEG C, 5%CO2It is incubated 30 minutes in incubator, 96 microwell plates is taken out, first with 0.01mol/L phosphate buffer
Confining liquid is washed away, then is blotted orifice plate surrounding liquid with filter paper, by A549 in carbon point dispersion liquid made from embodiment 1 and 6 holes
Cell is in 37 DEG C, 5%CO2Incubator is incubated 1 hour, is washed with 0.01mol/L phosphate buffer, is then put fluorescence microscope
The mark situation of lower observation cell.
Carbon point mark A549 cell results are as shown in Figure 9, it can be seen that A549 cells show bright under fluorescence microscope
Aobvious green fluorescence, illustrates that carbon point produced by the present invention can be used for cell imaging as fluorescent dye.
Above-mentioned result of the test explanation, the present invention has good fluorescence property using mushroom carbon point made from carbon source, also had
Have the advantages that good light stability, toxicity be low, good biocompatibility, had a good application prospect in terms of cell imaging.
Finally illustrate, the above embodiments are merely illustrative of the technical solutions of the present invention and it is unrestricted, although pass through ginseng
According to the preferred embodiments of the present invention, invention has been described, it should be appreciated by those of ordinary skill in the art that can
So that various changes are made to it in the form and details, the present invention limited without departing from appended claims
Spirit and scope.
Claims (7)
1. the method for carbon point is prepared by mushroom, it is characterised in that mushroom powder and water are pressed into 0.08g ~ 0.1g：1mL ratio mixing
Afterwards, 5-7 hours obtained carbon points of 180-200 DEG C of reaction.
2. the method as claimed in claim 1 that carbon point is prepared by mushroom, it is characterised in that mushroom powder and water are pressed into 0.1g：1mL
Ratio mixing after, 200 DEG C reaction 6 hours obtained carbon points.
3. the method as claimed in claim 1 that carbon point is prepared by mushroom, it is characterised in that cool down reaction solution, is centrifuged, collect
Supernatant, with the filtering with microporous membrane that aperture is 0.22 μm, after filtrate molecular cut off is 100 bag filter dialysis, is dried,
Obtain carbon point powder.
4. the method as claimed in claim 3 that carbon point is prepared by mushroom, it is characterised in that cool down reaction solution, 10000rpm
Centrifugation 15 minutes, collects supernatant, is 0.22 μm of filtering with microporous membrane with aperture, filtrate molecular cut off for 100 it is saturating
After analysing bag dialysis 24 hours, freeze-drying obtains carbon point powder.
5. the method that carbon point is prepared by mushroom as described in any one of Claims 1-4, it is characterised in that the mushroom powder is
By new fresh mushroom in after 60 DEG C ~ 80 DEG C dryings 24 ~ 48 hours, with mortar grinder into powder.
6. using carbon point made from any one of claim 1 to 5 methods described.
7. application of the carbon point in terms of cell imaging described in claim 6.
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Cited By (10)
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| CN108069414A (en) * | 2017-12-19 | 2018-05-25 | 重庆文理学院 | A kind of method for preparing carbon nanomaterial using discarded pine tree tower |
| CN108101022A (en) * | 2017-12-27 | 2018-06-01 | 重庆文理学院 | A kind of method that carbon quantum dot is prepared using fennel seeds as raw material |
| CN109593523A (en) * | 2018-11-13 | 2019-04-09 | 东北林业大学 | A kind of preparation method and application of hypha,hyphae nitrogen sulphur auto-dope carbon dots |
| CN109734073A (en) * | 2018-12-13 | 2019-05-10 | 南京师范大学 | It is a kind of using wormwood leaf as carbon quantum dot of carbon source and the preparation method and application thereof |
| CN109821525A (en) * | 2019-03-28 | 2019-05-31 | 东北林业大学 | A kind of preparation method loading biomass carbon point defect titanium dioxide optical catalyst |
| CN110194953A (en) * | 2019-06-27 | 2019-09-03 | 上海大学 | Biomass-based fluorescent carbon quantum dot, preparation method and application |
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| CN108069414A (en) * | 2017-12-19 | 2018-05-25 | 重庆文理学院 | A kind of method for preparing carbon nanomaterial using discarded pine tree tower |
| CN108101022A (en) * | 2017-12-27 | 2018-06-01 | 重庆文理学院 | A kind of method that carbon quantum dot is prepared using fennel seeds as raw material |
| CN109593523A (en) * | 2018-11-13 | 2019-04-09 | 东北林业大学 | A kind of preparation method and application of hypha,hyphae nitrogen sulphur auto-dope carbon dots |
| CN109734073A (en) * | 2018-12-13 | 2019-05-10 | 南京师范大学 | It is a kind of using wormwood leaf as carbon quantum dot of carbon source and the preparation method and application thereof |
| CN109821525A (en) * | 2019-03-28 | 2019-05-31 | 东北林业大学 | A kind of preparation method loading biomass carbon point defect titanium dioxide optical catalyst |
| CN110194953A (en) * | 2019-06-27 | 2019-09-03 | 上海大学 | Biomass-based fluorescent carbon quantum dot, preparation method and application |
| CN110320196A (en) * | 2019-08-23 | 2019-10-11 | 云南健牛生物科技有限公司 | A kind of highly sensitive method for detecting inorganic mercury and organic mercury |
| WO2021066767A1 (en) * | 2019-09-30 | 2021-04-08 | Selçuk Üni̇versi̇tesi̇ Rektörlüğü | A method for obtaining a fluorophore dye from the edible wild plants |
| CN113789173A (en) * | 2021-09-16 | 2021-12-14 | 青岛农业大学 | Preparation method and application of edible fungus-based fluorescent carbon quantum dots for indicating freshness of food |
| CN114958359A (en) * | 2022-05-12 | 2022-08-30 | 青岛农业大学 | Mimic enzyme material copper-doped carbon quantum dot using mushroom as carbon source and preparation method and application thereof |
| CN114958359B (en) * | 2022-05-12 | 2023-10-13 | 青岛农业大学 | Simulated enzyme material copper-doped carbon quantum dot with mushrooms as carbon sources, and preparation method and application thereof |
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