CN104694119A - Functional fluorescent carbon dot as well as preparation method and application thereof - Google Patents
Functional fluorescent carbon dot as well as preparation method and application thereof Download PDFInfo
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- CN104694119A CN104694119A CN201310662948.9A CN201310662948A CN104694119A CN 104694119 A CN104694119 A CN 104694119A CN 201310662948 A CN201310662948 A CN 201310662948A CN 104694119 A CN104694119 A CN 104694119A
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Abstract
The invention discloses a carbon dot component in cola as well as a property characterization method and a practical application method of the carbon dot. The preparation method comprises the following steps: by taking original-taste (1+2) Nestle coffee, pepsi (330ml) and coca-cola (330ml) as main raw materials, separating and purifying through a dextrane gel G-25 column and performing freeze-drying for 24 hours to collect the carbon dot. The coffee/cola carbon dot is relatively low in cost, low in toxicity, simple in extracting method, good in water solubility, good in biocompatibility and can be widely used as a novel fluorescent carbon nano particle in the field of biomarkers.
Description
Technical field
The present invention relates to fluorescent carbon nano material, specifically at coffee, the fluorescent carbon nano-particles existed in cola.
Background technology
Nano material has good optical property and receives increasing concern at biological technical field.Wherein, in the Application Areas that biological detection, sensing, radiography, cytochemistry, immunohistochemistry etc. are many, the fluorescent labelling techniques of nano material obtains good development in recent years, and becomes the topmost technique means of the one realizing above-mentioned functions.But traditional fluorescence labeling material organic synthesis process is more complicated, and biocompatibility is poor, and the solubleness in water is very low, its application in aqueous phase is made to receive restriction to a certain degree.The correlative study of fluorescent nano particles receives the concern of Chinese scholars just under this background gradually, and is in recent years being developed rapidly.The light stability of fluorescent nano material---semiconductor-quantum-point and water-soluble superior, but more recently by affecting in vivo nanoparticle and the research of biological safety, it is found that except the nanoparticle (as CdSe) containing toxic element has except toxic side effect in vivo, the nano material (as Si, ZnO, SiO2, Au etc.) that other biological is good also shows certain toxicity, and its toxicity depends mainly on the particle diameter of nano material, chemical constitution, charge, wetting ability, agglomeration and pattern etc.A kind of novel fluorescent material---carbon nano-luminescent material is found along with to the further research of nano material.Carbon nanomaterial comprises: the carbon nanotube of the carbon point of zero dimension, soccerballene, one dimension, the Graphene of two dimension.Carbon is widespread in nature, and is extensively be present in organism, is the important element that organism is formed, so carbon nanomaterial can surely can be more extensive relative to the source of other nano materials.Carbon point is exactly a kind of novel carbon material that Clyde's gloomy university research person finds for 2004 in once testing, compare with traditional pigment dyestuff with traditional semiconductor-quantum-point, it is little that newcomer in this Wei Tan family not only maintains carbon material toxicity, biocompatibility is good, the advantages such as particle diameter is less.In addition, carbon nano-particle such as is also easy to realize surface-functionalized, stable luminescence and fluorescence are changed at the advantage, so fluorescent carbon nano particle is considered to a kind of ideal fluorescent mark or test material.Report now that in the synthesis of carbon point, Problems existing comprises following several aspect:
(1) synthetic materials select permeability (document 3:Pin-Che Hsu and Huan-Tsung Chang.ChemComm, 2012,48,3984 – 3986. document 4:Swagatika Sahu, Birendra Behera, Tapas K.ChemComm, 2012,48,8835 – 8837).Carbon point synthesis material mainly contains graphite, plant ash, oil smoke etc., and raw material sources are extensively easy to get and cheap, but quantum yield is lower, brings a lot of problem to the application of later biomaterial.Although natural carbohydrate, EDTA and TRIS quantum yield are very high, synthetic materials is expensive, is unfavorable for scale operation and the application of carbon point.
(2) synthetic method problem.Present synthetic method the most general is hydrothermal method and microwave method (document 1:SwagatikaSahu, Birendra Behera.ChemComm., 2012,48,8835 – 8837. document 2:Xinyun Zhai, PengZhang, Changjun Liu.ChemComm., 2012,48,7955 – 7957.) generated time of hydrothermal method is usually longer, and need to react at relatively high temperatures, but quantum yield is usually lower.Microwave method generated time is shorter, but has radiation in building-up process, to environment, is not a kind of synthetic method of green.These two kinds of usual costs of synthetic method are very high.And the carbon point of these two kinds of method synthesis all needed post to purify usually or dialysis removal of impurities.
The main component of food is carbohydrate, and in its course of processing, and through frying, fermentation, cures, the process such as high-temperature sterilization, carbon point itself to be formed in the course of processing, without the need to carrying out synthesis specially.And food is eaten dozens or even hundreds of year by the mankind, has no side effect.The mankind for the carbon point of food sources safely without the need to doubting.The biocompatibility of the carbon point of food sources is excellent, will bring huge prospect in bio-imaging application aspect.
Summary of the invention
In order to solve the problem, the object of this invention is to provide one at coffee, that extracts in cola is well water-soluble, good biocompatibility, the carbon point that security is high and cheap.
To achieve these goals, technical scheme of the present invention is as follows:
1. coffee carbon point
2g nest original flavor coffee is dissolved in 20ml, in the hot water of 88 DEG C, centrifugation (14*10
3rpm, 15min), remove precipitation, obtain supernatant liquor, filter.By the gel columns of dextran G-25, separating-purifying carbon point.Excited at 300nm by spectrophotofluorometer, collect and have fluorescent carbon point.And the carbon of collection point is carried out vacuum lyophilization.And sample is carried out property representation, cell marking and living imaging.
2. laughable carbon point
Coca-Cola of 100ml and pepsi cola are placed on respectively on magnetic stirring apparatus and stir 4h, stirring velocity is 500rpm/min, makes CO
2air release is gone out.By sample respectively by 0.45 μm, the filter membrane of 0.22 μm, and put it into freeze-drying in lyophilizer.And by the diluted sample of freeze-drying to 20ml.By the pillar separating-purifying of dextran G-25, excited at 300nm by spectrophotofluorometer, collect and have the component of fluorescence and carry out characterization experiments.
Described carbon to select in food extracting directly without the need to synthesis, and carbon is selected composition and all to be originated coffee, and pepsi cola, in Coca-Cola.Described carbon point toxicity is extremely low substantially nontoxic.
Tool of the present invention has the following advantages:
(1) carbon of the present invention is selected at coffee, is formed, need not carry out synthesis specially in the laughable course of processing.The step eliminating synthesis will promote working efficiency greatly.
(2) coffee of the present invention, laughable carbon point there is good biocompatibility and photoluminescent property superior.
(3) carbon point of the present invention has high security.Coffee, cola has century-old history, never occurs safety problem, and like by young people and drink, so the security of laughable source carbon point is indubitable.
Accompanying drawing explanation
Fig. 1 is coffee, the transmission electron microscope photo that laughable carbon is selected: a) coffee, b) Coca-Cola, c) pepsi cola.
Fig. 2 is coffee, the ultraviolet of laughable carbon point, fluorescence spectrum and fluorescence photo: a) coffee, b) Coca-Cola, c) pepsi cola.
Fig. 3 is coffee, the fluorescence lifetime figure that laughable carbon is selected: a) coffee, b) Coca-Cola, c) pepsi cola.
Fig. 4 is coffee, the fluorescence spectrum of the upper conversion that laughable carbon is selected: a) coffee, b) Coca-Cola, c) pepsi cola.
Fig. 5 is coffee, the XPS collection of illustrative plates that laughable carbon is selected: a) coffee, b) Coca-Cola, c) pepsi cola.
Fig. 6 is coffee, the FTIR collection of illustrative plates that laughable carbon is selected: a) coffee, b) Coca-Cola and pepsi cola.
Fig. 7 is coffee, the collection of illustrative plates of the XRD that laughable carbon is selected: a) coffee, b) Coca-Cola, c) pepsi cola.
Fig. 8 is coffee, the ionic strength stability diagram that laughable carbon is selected: a) coffee, b) Coca-Cola, c) pepsi cola.
Embodiment
Adopt coffee, laughable as raw material.
2g nest original flavor coffee is dissolved in 20ml by the making that coffee carbon is selected, in the hot water of 88 DEG C, and centrifugation (14*10
3rpm, 15min), remove precipitation, obtain supernatant liquor, filter.By the gel columns of dextran G-25, separating-purifying carbon point.The stronger carbon point of fluorescence intensity is collected by spectrophotofluorometer.And the carbon of collection point is carried out vacuum lyophilization.
Coca-Cola of 100ml and pepsi cola are placed on magnetic stirring apparatus by making that laughable carbon is selected respectively stirs 4h, and stirring velocity is 500rpm/min, makes CO
2air release is gone out.Sample respectively by 0.45 μm, the filter membrane of 0.22 μm, and put it into freeze-drying in lyophilizer.And by the diluted sample of freeze-drying to 20ml.By the pillar separating-purifying of dextran G-25, excited at 300nm by spectrophotofluorometer, collect the component having fluorescence.And lyophilize carbon point.
Below by embodiment, the invention will be further described.
1. the extraction of carbon point in coffee
The nest's original flavor coffee getting 4g to be dissolved in the pure water of about 90 DEG C and high speed centrifugation (14000rpm, 15min), successively by the filtering membrane of 0.45nm and 0.22nm.Separation and Extraction is carried out with the gel columns of dextran G-25.Each applied sample amount is 1ml, excites measurement fluorescence intensity with spectrophotofluorometer at 300nm, gets the component of fluorescence, carries out lyophilize 24h.
2. the extraction of carbon point in cola
Coca-Cola of 100ml and pepsi cola are placed on magnetic stirring apparatus respectively and stir, until CO
2gas all discharges.Sample respectively by 0.45 μm, the filter membrane of 0.22 μm, and put it into freeze-drying in lyophilizer.And by the diluted sample of freeze-drying to 20ml.By the pillar separating-purifying of dextran G-25, each applied sample amount 1ml, will have fluorescent components collect and sample is carried out lyophilize again.Sample drying completed carries out characterization experiments.
3. coffee, the sign of laughable carbon point character
(1) coffee, the form of laughable carbon point and size dimension
Fig. 1 is coffee, the transmission electron microscope photo of laughable carbon point, result shows that the laughable carbon point form through purification processes is spheroidal, dispersed, size is even, coffee carbon point grain size concentrates on 2-3nm, and Coca-Cola grain size concentrates on 2.0-3.0nm, and Pepsi carbon point grain size concentrates on 2.5-3.5nm.
(2) coffee, fluorescence spectrum, uv-absorbing and the fluorescence photo spectral response curve of laughable carbon point
Fig. 2 is coffee, fluorescence spectrum, uv-absorbing and the fluorescence photo spectrum of laughable carbon point.Coffee in figure, there is obvious red shift along with excitation wavelength increases in the fluorescence spectrum of visible Coca-Cola and pepsi cola carbon point, the maximum excitation wavelength of coffee carbon point appears at 390nm place and laughable carbon point is all appearing at 300nm.But the absorption spectrum of ultraviolet all appears at 280nm stronger absorption peak, it is the charateristic avsorption band of aromatics.
(3) coffee, the fluorescence lifetime of laughable carbon point
Fig. 3 is coffee, the fluorescence lifetime figure of laughable carbon point.Caffeinated water's solution of configuration 150mg/ml, excites under the exciting light of 376nm, and maximum emission peak is that 420nm launches, and records fluorescence lifetime.The experimental technique that laughable carbon is selected is identical with coffee, but maximum emission peak is 365nm.In experiment, find coffee, Coca-Cola and pepsi cola carbon point are three index quenchers, and the fluorescence lifetime of coffee is 4.64ns as calculated, and the fluorescence lifetime of Coca-Cola carbon point is 4.32ns, and the fluorescence lifetime of pepsi cola carbon point is 4.26ns.
(4) coffee, the up-conversion fluorescence character of laughable carbon point
Fig. 4 is coffee, the upper conversion collection of illustrative plates of laughable carbon point.The coffee getting 200mg/ml drinks laughable carbon point, with spectrophotofluorometer at excitation wavelength 750nm---800nm excites, at 375nm---, and 600nm detects intensity and the red shift situation of emmission spectrum.Visible coffee in figure, laughable carbon point has the character of upper conversion but the more weak and maximum emission wavelength of intensity changes along with the change of excitation wavelength, excites lower intensity maximum at 800nm.
(5) coffee, the XPS experiment of laughable carbon point
Fig. 5 is coffee, the collection of illustrative plates of laughable carbon point XPS.In figure, result shows coffee, main containing C and O two kinds of elements and containing the N element seldom measured in laughable carbon point.
(6) coffee, the infrared experiment of laughable carbon point
Fig. 6 is coffee, the infared spectrum of laughable carbon point.Result in figure (a) shows that purified 3326cm-1 contains the stretching vibration peak of O-H and N-H, and 2927cm-1 has the stretching vibration peak of saturated alkane, and 1637cm-1 has the formation vibration peak of N-H, and 1044cm-1 has the stretching vibration peak of C-O.And the stretching vibration peak also having C=O except above-mentioned functional group at 1745cm-1 of not purified coffee.This phenomenon is because the vegetable fat powder contained in coffee in ultracentrifugal process and hydrogenated vegetable oil are removed, so carbonyl disappears.Figure (b) in show two kinds of carbon points with group substantially identical, all contain the group of-OH, this also demonstrates carbon point with negative electricity.
(7) coffee, the XRD experiment of laughable carbon point
Fig. 7 is the collection of illustrative plates of the XRD of laughable carbon point.In figure (a), visible spacing d=0.43nm, proves that carbon point is the structure with decolorizing carbon.The increase of spacing illustrates the increase of oxy radical in material.Figure (b) coca carbon point without obvious peak, proves the structure of agraphitic carbon without obvious peak.The collection of illustrative plates d=5.04nm of figure (c) Pepsi carbon point.Prove that two kinds of carbon points all have the structure of decolorizing carbon.And the increase of Pepsi carbon point spacing illustrates the increase of oxy radical in material.
(8) coffee, the pH stability experiment of laughable carbon point
Fig. 8 is coffee, the stability diagram of the pH of laughable carbon point.Configuration coffee carbon point 100mg/ml and different pH(2-11) B-R buffered soln, added respectively in different pH solution by 250 μ l, survey fluorescence intensity with spectrophotofluorometer, each pH value does three parallel laboratory tests.It is identical with coffee that laughable carbon selects experimental technique.The having good stability of pH of coffee carbon point, increases fluorescence intensity at pH substantially constant in figure below (a).Can find out in figure (b) (c) that two kinds of carbon points are more stable in the first stability of acidic conditions, occur significantly declining at pH=6, continue to occur stability under the condition of pH in alkalescence, just weak than intensity under acidic conditions.
2. coffee, the application experiment of laughable carbon point
Coffee carbon point cell labeling experiment
(1) cell selects SMMC-7721 cell.Select containing 10% foetal calf serum RPMI-1640 substratum and add microbiotic and non-essential amino acid at 37 DEG C, the CO of 5%
2lower cultivation.The inoculum density of cell is 1 × 10
5cells/ml.The concentration of adding coffee carbon point is in the medium 15mg/ml.24h is cultivated at 37 DEG C.The cell phosphate buffer soln of precipitation cleans.Observation of cell mark situation under fluorescent microscope.
(2) the labelling experiment photo of cell
The labelling experiment photo of cell is shown to be compared with control cells and does not add carbon point.B () is the cell that experimental group cell adds 15mg/ml carbon point.Can find out that the cell through mark sends obvious fluorescence in the drawings, and obvious fluorescence can be observed under different excitation wavelength.Under the irradiation of UV-light, mark carbon point cell sends bright blue light, and control group only has more weak blue light; Under the irradiation of blue light, the cell of mark carbon point sends bright green glow, control group green glow and faint; Under the irradiation of green glow, the cell of mark carbon point sends bright ruddiness, and the fluorescence of control group almost redfree.Illustrating that carbon point can enter cell and labeled cell, is the very good material of cell marking, and carbon point can by the optical excitation of each wavelength, for cell marking in the future provides more wide development space.
The cell labeling experiment of laughable carbon point
(1) cell selects Tca-8113 cell.Select containing 10% foetal calf serum RPMI-1640 substratum and add microbiotic and non-essential amino acid at 37 DEG C, the CO of 5%
2lower cultivation.The inoculum density of cell is 1 × 10
5cells/ml.The concentration of adding carbon point is in the medium 15mg/ml.24h is cultivated at 37 DEG C.The cell phosphate buffer soln of precipitation cleans.Observation of cell mark situation under Two Photon Fluorescence.
(2) the labelling experiment photo of cell
The labelling experiment photo of cell shows that two groups of carbon point concentration are 10mg/ml.The cell can finding out through mark in the drawings excites at 405nm and 488nm and all sends obvious blue light and green glow and fluorescence mainly concentrates in tenuigenin and do not enter nucleus down, and the cell fluorescence of control group does not almost have, illustrate that carbon point can enter cell and labeled cell, the very good material of cell marking, for cell marking in the future provides more wide development space.
Claims (4)
1. a functional fluorescent carbon point, is characterized in that: adopt coffee or cola as raw material;
The making that coffee carbon is selected: about 2g coffee is dissolved in 20ml-25ml, in the hot water of 85-90 DEG C, centrifugation (14*10
3rpm, 15min), remove precipitation, obtain supernatant liquor, respectively by the membrane filtration of 0.45 μm, 0.22 μm; Collect the gel columns of filtrate by dextran G-25, separating-purifying carbon point; Excited by spectrophotofluorometer 340nm, collect the carbon point having fluorescence; And the carbon of collection point is carried out vacuum lyophilization;
The making that laughable carbon is selected: Coca-Cola of about 100ml and pepsi cola are placed on respectively on magnetic stirring apparatus and stir 4h, rotating speed is 500rpm/min; By the filter membrane of sample respectively by 0.45 μm, 0.22 μm, collect filtrate and put it into freeze-drying in lyophilizer; And by the diluted sample of freeze-drying to 20ml.By the pillar separating-purifying of dextran G-25, excited by spectrophotofluorometer 340nm, collect the component having fluorescence strong, dry carbon point.
2. a preparation method for functional fluorescent carbon point described in claim 1, is characterized in that: about 2g coffee is dissolved in 20ml-25ml, in the hot water of 85-90 DEG C, and centrifugation (14*10
3rpm, 15min), remove precipitation, obtain supernatant liquor, respectively by the membrane filtration of 0.45 μm, 0.22 μm; Collect the gel columns of filtrate by dextran G-25, separating-purifying carbon point; Excited by spectrophotofluorometer 340nm, collect the carbon point having fluorescence; And the carbon of collection point is carried out vacuum lyophilization;
The making that laughable carbon is selected: Coca-Cola of about 100ml and pepsi cola are placed on respectively on magnetic stirring apparatus and stir 4h, rotating speed is 500rpm/min; By the filter membrane of sample respectively by 0.45 μm, 0.22 μm, collect filtrate and put it into freeze-drying in lyophilizer; And by the diluted sample of freeze-drying to 20ml.By the pillar separating-purifying of dextran G-25, excited by spectrophotofluorometer 340nm, collect the component having fluorescence strong, dry carbon point.
3. the application of a Functional carbon point according to claim 1 in biomarker.
4., according to application according to claim 3, describedly be can be cell by the biology that Functional carbon point marks.
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| CN106829917A (en) * | 2015-12-04 | 2017-06-13 | 中国科学院大连化学物理研究所 | A kind of carbon quantum dot and its application |
| CN107555417A (en) * | 2017-08-29 | 2018-01-09 | 湖南农业大学 | Commercialization beverage prepares the method for fluorescent carbon point and its fluorescent carbon point of preparation |
| CN107641632A (en) * | 2017-10-18 | 2018-01-30 | 福州大学 | A kind of method with the carbon-based point of Microbe synthesis |
| CN109054821A (en) * | 2018-07-13 | 2018-12-21 | 山西大学 | A kind of fluorescent carbon point and its preparation method and application |
| CN114044511A (en) * | 2021-12-02 | 2022-02-15 | 南京财经大学 | Preparation method and application of beer-based nitrogen-phosphorus co-doped biomass carbon dots |
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Cited By (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106829917A (en) * | 2015-12-04 | 2017-06-13 | 中国科学院大连化学物理研究所 | A kind of carbon quantum dot and its application |
| CN105567227A (en) * | 2016-01-16 | 2016-05-11 | 上海大学 | Method for extracting graphene quantum dots from coffee-ground solid waste |
| CN107555417A (en) * | 2017-08-29 | 2018-01-09 | 湖南农业大学 | Commercialization beverage prepares the method for fluorescent carbon point and its fluorescent carbon point of preparation |
| CN107641632A (en) * | 2017-10-18 | 2018-01-30 | 福州大学 | A kind of method with the carbon-based point of Microbe synthesis |
| CN109054821A (en) * | 2018-07-13 | 2018-12-21 | 山西大学 | A kind of fluorescent carbon point and its preparation method and application |
| CN109054821B (en) * | 2018-07-13 | 2021-05-14 | 山西大学 | Fluorescent carbon dot and preparation method and application thereof |
| CN114044511A (en) * | 2021-12-02 | 2022-02-15 | 南京财经大学 | Preparation method and application of beer-based nitrogen-phosphorus co-doped biomass carbon dots |
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Application publication date: 20150610 |