CN106706591B - A kind of reversible nanometer porphyrin fluorescence sensor recognition quantitative chiral amino acid method - Google Patents
A kind of reversible nanometer porphyrin fluorescence sensor recognition quantitative chiral amino acid method Download PDFInfo
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- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
- G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
- G01N21/64—Fluorescence; Phosphorescence
- G01N21/6428—Measuring fluorescence of fluorescent products of reactions or of fluorochrome labelled reactive substances, e.g. measuring quenching effects, using measuring "optrodes"
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
- G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
- G01N21/64—Fluorescence; Phosphorescence
- G01N21/6428—Measuring fluorescence of fluorescent products of reactions or of fluorochrome labelled reactive substances, e.g. measuring quenching effects, using measuring "optrodes"
- G01N2021/6432—Quenching
Abstract
A kind of reversible nanometer porphyrin fluorescence sensor recognition quantitative chiral amino acid method belongs to nano material preparation and chemical analysis detection technique field.The specific recognition used quantifies the reversible nanometer porphyrin fluorescence sensor of proline, lysine and serine chirality, using CdTe quantum as fluorescence probe, the self-assembled nanometer porphyrin that four-(4- pyridyl group) zinc protoporphyrin tetrahydrofuran solutions and cetyl trimethylammonium bromide (CTAB) are prepared is fluorescence quencher, and the specific binding of the two obtains switch nanometer porphyrin fluorescence sensor.Switch nanometer porphyrin fluorescence sensor and chiral proline, lysine and serine act on obtaining reversible (On-Off-On) nanometer porphyrin fluorescence sensor.There are many advantages compared to tradition in the method for chromatography hair method separation identification amino acid chiral.
Description
Technical field
The invention belongs to nano material preparation and chemical analysis detection technique fields, and in particular to a kind of reversible nanometer
The method of the controllable preparation of porphyrin fluorescence sensor and its highly sensitive detection chiral amino acid.
Background technique
Chirality is a kind of critical nature that amino acid, sugar and heterocycle etc. construct living matter primitive molecule.Chiral molecules pair
The difference for reflecting micro-space configuration between isomers frequently can lead to its huge difference that macroscopical physiological and pharmacological acts in vivo
It is different.Therefore, the chiral Recognition research for carrying out chiral material and large biological molecule, to the mechanism of action and rule for disclosing biomolecule
It is of great significance.Proline, lysine and serine play important adjusting in the physiology of biology and pathologic process and make
With.Proline can be used for the supplement of malnutrition, hypoproteinosis, gastrointestinal disease, scald and postoperative protein in medicine
Deng.Lysine is mainly used as feed addictive, food additive and pharmacy.Serine and fat and fatty acid metabolism and
The growth of muscle is related.Studies have shown that serine facilitates the generation of immunoglobulin and antibody, help to maintain siberian crabapple
System.The physiological and pharmacological effect that their L-type and the different structure of D type generate in vivo also has greatest differences.Therefore it studies
The detection technique of effective proline, lysine and serine chiral Recognition and quantitative analysis, in chemistry, biology and medicine
Field is all extremely important.Due to nanometer porphyrin sensors have it is easy to operate, radiationless, it is highly sensitive and high stability excellent
Gesture becomes the ideal chose of quantitative judge proline, lysine and serine.Conventional method has chromatography, electrochemistry side
Method and electron microscope technique etc., these methods have the characteristics that high sensitivity or separating capacity are strong, but simultaneously there is also it is some not
Foot, such as being difficult to avoid that, influence, complicated sample preparation preparation, detection time that derivative reagent identify amino acid chiral are grown.
Therefore a kind of method of quick, high sensitivity and selective amino acid chiral identification and quantitative analysis is studied to amino acid
Biological action has important very important meaning.
The present invention is to overcome the defect of existing method, provides and a kind of new is known based on reversible nanometer porphyrin fluorescence sensor
Other quantitative detection chiral amino acid method.
Summary of the invention
An object of the present invention, which provides, a kind of to be prepared reversible nanometer porphyrin fluorescence simple, that reaction condition is mild and passes
Sensor controllable method for preparing；It is good that the second purpose is to provide a kind of high sensitivity, selectivity, is based on fluorescence On-Off-On mode method
Fast quantification identifies the reversible nanometer porphyrin fluorescence sensor of proline, lysine and serine chirality.
The specific recognition that the present invention uses quantifies the reversible nanometer porphyrin fluorescence of proline, lysine and serine chirality
Sensor, using CdTe quantum as fluorescence probe, four-(4- pyridyl group) zinc protoporphyrin tetrahydrofuran solutions and cetyl three
The self-assembled nanometer porphyrin that methyl bromide ammonium (CTAB) is prepared is fluorescence quencher, and the specific binding of the two is switched
Nanometer porphyrin fluorescence sensor.Switch nanometer porphyrin fluorescence sensor acts on obtaining with chiral proline, lysine and serine
Reversible (On-Off-On) nanometer porphyrin fluorescence sensor.
The present invention solves the problems, such as that the technical solution taken is, quantitative judge chiral proline, lysine and serine are received
The preparation method of rice porphyrin fluorescence sensor, comprising the following steps:
(1) dichloride cadmium and N-acetyl-L-cysteine are dissolved in ultrapure water, are stirred 15 minutes under room temperature, normal pressure
PH value of solution is adjusted to 8.00 with sodium hydroxide solution afterwards, then inflated with nitrogen ice bath stirring 20 minutes；Sodium tellurite, stirring 15 is added
Minute；Sodium borohydride is added, is stirred 15 minutes；Finally this solution is put into reaction kettle, reacts 50 in 200 DEG C of baking oven
Minute, obtain the feux rouges CdTe quantum fluorescence probe that launch wavelength is 641nm；
(2) four-(4- pyridyl group) zinc protoporphyrins are dissolved in tetrahydrofuran solution, to cetyl trimethylammonium bromide
(CTAB) in aqueous solution be added four-(4- pyridyl group) zinc protoporphyrin tetrahydrofuran solutions, room temperature, atmospheric agitation 10 minutes, solution by
Muddiness becomes clarification, and reaction stops, and obtains four-(4- pyridyl group) zinc protoporphyrin self-assembly for nanosphere solution；
(3) four-(4- pyridyl group) zinc protoporphyrin self-assembly for nanosphere solution are added in CdTe quantum fluorescence probe, then
The Tris-HCl buffer solution of pH=4.02 is added, four-(4- pyridyl group) zinc protoporphyrin self-assembly for nanosphere solution are turned by electronics
It moves and fluorescence resonance energy transfer effect, quenching quantum dot fluorescence provides quantum dot by specifically binding obtained compound
The state of one " Turn-off "；
(4) quantum dot fluorescence that step (3) obtain is added in the D type of various concentration range and L-type amino acid suitably to quench
Compound in, quantum dot fluorescence restores, and the phenomenon that D type and L-type amino acid cause quantum dot fluorescence to restore generates apparent poor
It is different, it realizes in the identification of reversible nanometer porphyrin fluorescence sensing modes assistant's acidic amino acid and quantitative；To obtain it is reversible (open-
On/off) nanometer porphyrin fluorescence sensor.
Or directly step (3) and (4) are merged: by synthesis in the D type of various concentration range and L-type amino acid, step (2)
Four-(4- pyridyl group) zinc protoporphyrin self assembly solution, PH=4.02 Tris-HCl buffer solution mixing, stand 5 minutes；Again plus
The CdTe quantum for entering step (1) synthesis carries out fluorescence spectrometry at 540-720nm, measures its spectrum after five minutes.
It is further preferred:
Dichloride cadmium in the present invention, N-acetyl-L-cysteine, sodium tellurite substance amount ratio are as follows: 1.0:(1.2
~1.5): 0.2, general step (1) CdTe quantum fluorescence probe launch wavelength is 620~640nm；
Four-(4- pyridyl group) zinc protoporphyrins and cetyl trimethylammonium bromide are in mixed solution in step (2) of the present invention
The mass ratio of the material is 1:(1.3~1.5)；
Four-(4- pyridyl group) zinc protoporphyrins and CdTe amount in step (3) four-(4- pyridyl group) zinc protoporphyrin nanosphere in the present invention
The mass ratio of the material of son point solution is 40~42:1；
The concentration, CdTe quantum of four-(4- pyridyl group) zinc protoporphyrin nanospheres are dense in step (3) mixed solution in the present invention
Degree is respectively 8.0 × 10-9-1.28×10-7mol/L、5.8×10-9When mol/L, four-(4- pyridyl group) zinc protoporphyrin nanospheres with
The fluorescence intensity of CdTe quantum is at good linear relationship.
Further preferred: reversible nanometer porphyrin fluorescence sensor of the invention is by quantum dot and nanometer porphyrin specificity
In conjunction with obtained compound.Absorbing wavelength is down to 365 by 851 by 641nm red shift to 649nm, fluorescence intensity.
The D type and L-type amino acid of reversible nanometer porphyrin fluorescence sensor quantitative judge of the invention are respectively D/L- dried meat ammonia
Acid, D/L- lysine, D/L- serine.
Reversible nanometer porphyrin fluorescence transducer sensitivity of the invention is high.The fluorescence intensity of CdTe quantum fluorescence probe with
The increase of four-(4- pyridyl group) zinc protoporphyrin self assembly solution gradually weaken, it might even be possible to quenching on earth, as long as the present invention carries out
Part is quenched or is quenched (preferably in linear relation part range) completely, is able to achieve the qualitative and quantitative detection of step (4)；
Four-(4- pyridyl group) zinc protoporphyrin nanosphere concentration (1.46 × 10-9-1.28×10-7Mol/L) with CdTe quantum (2.9 × 10- 8Mol/L fluorescence intensity) is at good linear relationship；The fluorescence of the D type and L-type amino acid that identify in a certain range is strong
Degree has linear relationship.
Reversible nanometer porphyrin fluorescence sensor quantitative judge D-/L- proline, D-/L- lysine and D-/L- of the invention
Serine ability is strong.Concentration is from 1.0 × 10-10Mol/L to 5.0 × 10-6L-PROLINE, L-lysine and the D-Ser of mol/L
Rear fluorescence intensity does not have linear relationship in conjunction with reversible nanometer porphyrin sensors.D-PROLINE (1.0 × 10-9-1.5×10- 7Mol/L), D-Lys (1.0 × 10-9-1.5×10-8) and Serine (1.0 × 10 mol/L-9-5.0×10-9Mol/L) with
Fluorescence intensity of the reversible nanometer porphyrin fluorescence sensor in conjunction with after enhances as the concentration of amino acid increases, and at good line
Sexual intercourse.Linearly dependent coefficient can be 0.9984,0.9952,0.9931 respectively.Due to its binding ability be better than nanometer porphyrin with
Weak electrostatic interaction between quantum dot adds CdTe quantum reaction a period of time, nanometer porphyrin and CdTe quantum
In conjunction with dying down, fluorescence restores.The phenomenon that D type and L-type amino acid cause quantum dot fluorescence to restore generates apparent difference, realizes
In the identification of reversible nanometer porphyrin fluorescence sensing modes assistant's acidic amino acid and quantitative.To obtain reversible (On-Off-On) nanometer
Porphyrin fluorescence sensor；So step (3) and (4) merge and can separately obtain identical effect.
Reversible nanometer porphyrin sensors stability of the invention is good.The reversible nanometer porphyrin fluorescence sensor is 1.0 × 10- 6Mol/L ion (KCl, Na2SO4、CaCl2、ZnCl2), 1 μ g/mL bio-matrix it is (human serum albumins, bovine serum albumin(BSA), thin
Born of the same parents' culture solution, calf thymus DNA) and 0.1 μ g/mL mixing interference in the case where, with proline, lysine and serine act on it is glimmering
The intensity that light restores is almost unchanged.
Reversible nanometer porphyrin fluorescence sensor of the invention is to proline, lysine and serine fast response time.It is reversible
After proline, lysine and serine is added in nanometer porphin fluorescent optical sensor, the fast quick-recovery of fluorescence reaches most stationary value in 5 minutes.
Method of the present invention, compared to tradition with chromatography hair method separation identification amino acid chiral method have it is many excellent
Gesture, including preparation is simple, reaction condition is mild, the chiral Recognition and quantitation capabilities to proline, lysine and serine are strong, clever
Sensitivity height, strong antijamming capability, fast response time, this nanometer of porphyrin fluorescence sensor have in biochemistry, medicine and other fields
Actual application value.
Detailed description of the invention
Fig. 1 is the controllable method for preparing and its Gao Ling of reversible of the present invention (On-Off-On) nanometer porphyrin fluorescence sensor
The method schematic diagram of quick detection chiral amino acid.
Fig. 2 is the ultraviolet of four-(4- pyridyl group) zinc protoporphyrin self assembly solution in the reversible nanometer porphyrin sensors of the present invention
Visible light, abscissa are wavelength, and ordinate is absorbance.
Fig. 3 is the transmission of four-(4- pyridyl group) zinc protoporphyrin self assembly solution in the reversible nanometer porphyrin sensors of the present invention
Formula electron microscope picture, is nanosphere.
Fig. 4 is CdTe quantum and four-(4- pyridyl group) zinc protoporphyrin self assemblies in the reversible nanometer porphyrin sensors of the present invention
Solution specifically binds forward and backward fluorescence spectra, and abscissa is wavelength, and ordinate is fluorescence intensity.
Fig. 5 is the sensitivity of the reversible nanometer porphyrin sensors of the present invention.Four-(4- pyridyl group) zinc protoporphyrin self assembly solution
(1.46×10-9-1.28×10-7Mol/L) and CdTe quantum is glimmering after Tris-HCl buffer solution (PH=4.02) effect
Light spectrogram, abscissa are wavelength, and ordinate is fluorescence intensity.
Fig. 6 is the reversible nanometer porphyrin sensors of the present invention and various concentration D-PROLINE (1.0 × 10-9-1.5×10- 7Mol/L the fluorescence after) acting on restores spectrum, and abscissa is wavelength, and ordinate is fluorescence intensity.
Fig. 7 is the reversible nanometer porphyrin sensors of the present invention and various concentration L-PROLINE (1.0 × 10-10-5.0×10- 6Mol/L the fluorescence after) acting on restores spectrum, and abscissa is wavelength, and ordinate is fluorescence intensity.
Fig. 8 is the reversible nanometer porphyrin sensors of the present invention and various concentration D-Lys (1.0 × 10-9-1.5×10- 8Mol/L the fluorescence after) acting on restores spectrum, and abscissa is wavelength, and ordinate is fluorescence intensity.
Fig. 9 is the reversible nanometer porphyrin sensors of the present invention and various concentration L-lysine (1.0 × 10-10-5.0×10- 6Mol/L the fluorescence after) acting on restores spectrum, and abscissa is wavelength, and ordinate is fluorescence intensity.
Figure 10 is the reversible nanometer porphyrin sensors of the present invention and various concentration Serine (1.0 × 10-9-5.0×10- 9Mol/L the fluorescence after) acting on restores spectrum, and abscissa is wavelength, and ordinate is fluorescence intensity.
Figure 11 is the reversible nanometer porphyrin sensors of the present invention and various concentration D-Ser (1.0 × 10-10-5.0×10- 6Mol/L the fluorescence after) acting on restores spectrum, and abscissa is wavelength, and ordinate is fluorescence intensity.
Figure 12, which is that the reversible nanometer porphyrin sensors of the present invention are linearly related with after the effect of various concentration D-PROLINE, to scheme, horizontal seat
It is designated as the concentration of D-PROLINE, ordinate is fluorescence recovery strength (F2) and CdTe quantum raw florescent intensity (F0) ratio.
Figure 13, which is that the reversible nanometer porphyrin sensors of the present invention are linearly related with after the effect of various concentration D-Lys, to scheme, horizontal seat
It is designated as the concentration of D-Lys, ordinate is the ratio of fluorescence recovery strength and CdTe quantum raw florescent intensity.
Figure 14, which is that the reversible nanometer porphyrin sensors of the present invention are linearly related with after the effect of various concentration Serine, to scheme, horizontal seat
It is designated as the concentration of Serine, ordinate is the ratio of fluorescence recovery strength and CdTe quantum raw florescent intensity.
Figure 15 is the stability of the reversible nanometer porphyrin sensors of the present invention.Reversible nanometer porphyrin sensors and D-PROLINE exist
Ca2+、Zn2+、SO4 2-, human serum albumins (HSA), bovine serum albumin(BSA) (BSA), cell culture fluid (CCF), calf thymus DNA
Stability after being acted in the case where interfering (Mixture) with mixing.Abscissa is the interfering substance being added, and ordinate is vertical
Coordinate is fluorescence recovery strength (F2) and CdTe quantum raw florescent intensity (F0) ratio.
Figure 16 is the stability of the reversible nanometer porphyrin sensors of the present invention.Reversible nanometer porphyrin sensors and D-Lys exist
Ca2+、Zn2+、SO4 2-, human serum albumins (HSA), bovine serum albumin(BSA) (BSA), cell culture fluid (CCF), calf thymus DNA
Stability after being acted in the case where interfering (Mixture) with mixing.Abscissa is the interfering substance being added, and ordinate is vertical
Coordinate is fluorescence recovery strength (F2) and CdTe quantum raw florescent intensity (F0) ratio.
Figure 17 is the stability of the reversible nanometer porphyrin sensors of the present invention.Reversible nanometer porphyrin sensors and Serine exist
Ca2+、Zn2+、SO4 2-, human serum albumins (HSA), bovine serum albumin(BSA) (BSA), cell culture fluid (CCF), calf thymus DNA
Stability after being acted in the case where interfering (Mixture) with mixing.Abscissa is the interfering substance being added, and ordinate is glimmering
Light recovery strength (F2) and CdTe quantum raw florescent intensity (F0) ratio.
Specific embodiment
Applicant will the present invention is described in further detail in conjunction with specific embodiments below, so that the skill of this field
The present invention is more clearly understood in art personnel.But the following contents should not be understood as that claims of the present invention is claimed
The limitation of range.
Embodiment:
Chemical reagent used in embodiment and solvent are that analysis is pure.The stirring uses magnetic stirrer side
Formula.The fluorescence spectrometry condition is launch wavelength 540-720nm, excitation wavelength 380nm, slit width 10-
15nm。
Embodiment 1: identification and quantitative analysis of the reversible nanometer porphyrin fluorescence sensor to Proline, the method are shown
It is intended to such as 1, steps are as follows:
(1) synthesis of CdTe quantum fluorescence probe
It is super that dichloride cadmium (0.1142g, 12.5mM) and N-acetyl-L-cysteine (0.0979g, 15mM) are dissolved in 40mL
In pure water, solution PH is adjusted to 8.00 with sodium hydroxide solution after stirring 15 minutes under room temperature, normal pressure, then inflated with nitrogen ice bath
Stirring 20 minutes.It is added sodium tellurite (0.0216g, 2.5mM), stirs 15 minutes；Add sodium borohydride (0.0113g,
7.5mM), it stirs 15 minutes.Finally this solution is put into reaction kettle, is reacted 50 minutes in 200 DEG C of baking oven.It is cooled to room
Temperature obtains 2.9 × 10-7Mol/LCdTe quantum dot fluorescence probe.
The synthesis of (2) four-(4- pyridyl group) zinc protoporphyrin self assembly solution
Appropriate four-(4- pyridyl group) zinc protoporphyrins are dissolved in tetrahydrofuran solution, obtaining concentration is 1.46 × 10-3mol/L
Four-(4- pyridyl group) zinc protoporphyrin tetrahydrofuran solutions, ultraviolet spectrogram such as Fig. 2.By cetyl trimethyl bromination
(0.0183g) is dissolved in 10mL aqueous solution, and 240 μ L, tetra--(4- pyridyl group) zinc protoporphyrin tetrahydrofuran solution is added, and normal temperature and pressure stirs
It mixes 10 minutes, solution becomes clarification by muddiness, and reaction stops.Obtain 3.42 × 10-5Mol/L tetra--(4- pyridyl group) zinc protoporphyrin is from group
Fill solution, ultraviolet spectrogram such as Fig. 2.Its transmission electron microscope characterizes the nanosphere for being shown as partial size 40nm or so, such as
Fig. 3.
(3) preparation of nanometer porphyrin fluorescence sensor is switched
200 L2.9 × 10 μ are added in 1.5mL cuvette-8The CdTe quantum and 800 μ LPH of mol/L step (1) synthesis
=4.02 Tris-HCl buffer solution carries out fluorescence spectrometry at 540-720nm, and obtaining fluorescence intensity at 641nm is
851 peak, such as Fig. 4.200 L2.9 × 10 μ are added in 1.5mL cuvette-8Mol/L step (1) synthesis CdTe quantum and
70μL3.42×10-6Four-(4- pyridyl group) the zinc protoporphyrin self assembly solution synthesized in mol/L step (2), add 730 μ LPH
=4.02 Tris-HCl buffer solution mixes after five minutes, and fluorescence spectrometry is carried out at 540-720nm, at 649nm
The peak for being 365 to fluorescence intensity, such as Fig. 4.
(4) identification and quantitative analysis of the reversible nanometer porphyrin fluorescence sensor to D-/L- proline
100 μ LD-/L- proline aqueous solutions, 70 L3.42 × 10 μ are added in 1.5mL cuvette-6In mol/L step (2)
The Tris-HCl buffer solution of four-(4- pyridyl group) zinc protoporphyrin self assembly solution and 630 μ LPH=4.02 of synthesis stands 5 points
Clock.Add 200 L2.9 × 10 μ-8The CdTe quantum of mol/L step (1) synthesis, carries out fluorescence spectrum at 540-720nm
Measurement, measures its spectrum after five minutes.D-PROLINE (1.00 × 10-9-1.5×10-7Mol/L it) is sensed with nanometer porphyrin fluorescence
Fluorescence intensity of the device in conjunction with after enhances, such as Fig. 6, linearly dependent coefficient 0.9984 as the concentration of amino acid increases, and such as schemes
12.L-PROLINE (1.00 × 10-10-5.0×10-6Mol/L) rear fluorescence intensity is not closed linearly in conjunction with nanometer porphyrin sensors
System, such as Fig. 7.100 μ LD- proline, 100 μ L interfering substances, 70 L3.42 × 10 μ are added in 1.5mL cuvette-6Mol/L step
Suddenly the Tris-HCl buffer solution of four-(4- pyridyl group) the zinc protoporphyrin self assembly solution and 530 μ LpH=4.02 synthesized in (2),
Stand 5 minutes.Add 200 L2.9 × 10 μ-8The CdTe quantum of mol/L step (1) synthesis, uses fluorescence at 540-720nm
Spectroscopic assay measures its spectrum after five minutes, and fluorescence restores the influence of almost interference-free factor, shows very strong anti-dry
Disturb ability, such as Figure 15.
Embodiment 2: identification and quantitative analysis of the reversible nanometer porphyrin fluorescence sensor to lysine chirality, the method are shown
It is intended to such as 1, steps are as follows:
(1) synthesis of CdTe quantum fluorescence probe
CdTe quantum fluorescence probe is synthesized using the method for step (1) in embodiment 1.
The synthesis of (2) four-(4- pyridyl group) zinc protoporphyrin self assembly solution
Four-(4- pyridyl group) zinc protoporphyrin self assembly solution are synthesized using the method for step (2) in embodiment 1.
(3) preparation of nanometer porphyrin fluorescence sensor is switched
Nanometer porphyrin fluorescence sensor is prepared using the method for step (3) in embodiment 1.
(4) identification and quantitative analysis of the reversible nanometer porphyrin fluorescence sensor to D-/L- lysine
100 μ LD-/L- lysine solutions, 70 L3.42 × 10 μ are added in 1.5mL cuvette-6In mol/L step (2)
The Tris-HCl buffer solution of four-(4- pyridyl group) zinc protoporphyrin self assembly solution and 630 μ LPH=4.02 of synthesis stands 5 points
Clock.Add 200 L2.9 × 10 μ-8The CdTe quantum of mol/L step (1) synthesis, carries out fluorescence spectrum at 540-720nm
Measurement, measures its spectrum after five minutes.D-Lys (1.00 × 10-9-1.50×10-8Mol/L it) is passed with nanometer porphyrin fluorescence
Fluorescence intensity of the sensor in conjunction with after enhances, such as Fig. 8 as the concentration of amino acid increases, and linearly dependent coefficient is 0.9952 as schemed
13.L-lysine (1.00 × 10-10-5.0×10-6Mol/L) rear fluorescence intensity is not closed linearly in conjunction with nanometer porphyrin sensors
System, such as Fig. 9.100 μ LD- lysines, 100 μ L interfering substances, 70 L3.42 × 10 μ are added in 1.5mL cuvette-6Mol/L step
Suddenly the Tris-HCl buffer solution of four-(4- pyridyl group) the zinc protoporphyrin self assembly solution and 530 μ LPH=4.02 synthesized in (2),
Stand 5 minutes.Add 200 L2.9 × 10 μ-8The CdTe quantum of mol/L step (1) synthesis, uses fluorescence at 540-720nm
Spectroscopic assay measures its spectrum after five minutes, and fluorescence restores the influence of almost interference-free factor, shows very strong anti-dry
Disturb ability, such as Figure 16.
Embodiment 3: identification and quantitative analysis of the reversible nanometer porphyrin fluorescence sensor to serine chirality, the method are shown
It is intended to such as 1, steps are as follows:
(1) synthesis of CdTe quantum fluorescence probe
CdTe quantum fluorescence probe is synthesized using the method for step (1) in embodiment 1.
The synthesis of (2) four-(4- pyridyl group) zinc protoporphyrin self assembly solution
Four-(4- pyridyl group) zinc protoporphyrin self assembly solution are synthesized using the method for step (2) in embodiment 1.
(3) preparation of nanometer porphyrin fluorescence sensor is switched
Nanometer porphyrin fluorescence sensor is prepared using the method for step (3) in embodiment 1.
(4) identification and quantitative analysis of the reversible nanometer porphyrin fluorescence sensor to D-/L- serine
100 μ LD-/L- serine aqueous solutions, 70 L3.42 × 10 μ are added in 1.5mL cuvette-6In mol/L step (2)
The Tris-HCl buffer solution of four-(4- pyridyl group) zinc protoporphyrin self assembly solution and 630 μ LPH=4.02 of synthesis stands 5 points
Clock.Add 200 L2.9 × 10 μ-8The CdTe quantum of mol/L step (1) synthesis, carries out fluorescence spectrum at 540-720nm
Measurement, measures its spectrum after five minutes.Serine (1.00 × 10-9-5.00×10-9Mol/L it) is passed with nanometer porphyrin fluorescence
Fluorescence intensity of the sensor in conjunction with after enhances, such as Figure 10, linearly dependent coefficient 0.9931 as the concentration of amino acid increases, such as
Figure 14.D-Ser (1.00 × 10-10-5.0×10-6Mol/L) rear fluorescence intensity is not linear in conjunction with nanometer porphyrin sensors
Relationship, such as Figure 11.100 μ LL- serines, 100 μ L interfering substances, 70 L3.42 × 10 μ are added in 1.5mL cuvette-6mol/L
The Tris-HCl buffering of four-(4- pyridyl group) the zinc protoporphyrin self assembly solution and 530 μ LPH=4.02 that synthesize in step (2) is molten
Liquid stands 5 minutes.Add 200 L2.9 × 10 μ-8The CdTe quantum of mol/L step (1) synthesis, is used at 540-720nm
Fluorescence spectrometry, measures its spectrum after five minutes, and fluorescence restores the influence of almost interference-free factor, shows very strong
Anti-interference ability, such as Figure 17.
Claims (6)
1. a kind of reversible nanometer porphyrin fluorescence sensor recognition quantitative chiral amino acid method, which is characterized in that including following step
It is rapid:
(1) dichloride cadmium and N-acetyl-L-cysteine are dissolved in ultrapure water, are used after being stirred 15 minutes under room temperature, normal pressure
PH value of solution is adjusted to 8.00 by sodium hydroxide solution, then inflated with nitrogen ice bath stirring 20 minutes；Sodium tellurite is added, stirs 15 points
Clock；Sodium borohydride is added, is stirred 15 minutes；Finally this solution is put into reaction kettle, 50 points are reacted in 200 DEG C of baking oven
Clock obtains the feux rouges CdTe quantum fluorescence probe that launch wavelength is 641nm；
(2) four-(4- pyridyl group) zinc protoporphyrins are dissolved in tetrahydrofuran solution, to cetyl trimethylammonium bromide (CTAB) water
Four-(4- pyridyl group) zinc protoporphyrin tetrahydrofuran solutions, room temperature, atmospheric agitation 10 minutes are added in solution, solution becomes clear by muddiness
Clearly, reaction stops, and obtains four-(4- pyridyl group) zinc protoporphyrin self-assembly for nanosphere solution；
(3) four-(4- pyridyl group) zinc protoporphyrin self-assembly for nanosphere solution are added in CdTe quantum fluorescence probe, are added
The Tris-HCl buffer solution of pH=4.02, four-(4- pyridyl group) zinc protoporphyrin self-assembly for nanosphere solution by electronics transfer and
Fluorescence resonance energy transfer effect, quenches quantum dot fluorescence, by specifically binding obtained compound, provides quantum dot one
The state of " Turn-off "；
(4) it is answered what the quantum dot fluorescence that the D type of various concentration range and L-type amino acid addition step (3) obtain suitably quenched
It closes in object, quantum dot fluorescence restores, and the phenomenon that D type and L-type amino acid cause quantum dot fluorescence to restore generates apparent difference, real
Show in the identification of reversible nanometer porphyrin fluorescence sensing modes assistant's acidic amino acid and quantitative；It is received to obtain reversible On-Off-On
Rice porphyrin fluorescence sensor；
Or directly step (3) and (4) are merged: by what is synthesized in the D type of various concentration range and L-type amino acid, step (2)
The Tris-HCl buffer solution mixing of four-(4- pyridyl group) zinc protoporphyrin self assembly solution, pH=4.02, stands 5 minutes；It adds
The CdTe quantum of step (1) synthesis, fluorescence spectrometry is carried out at 540-720nm, measures its spectrum after five minutes.
2. a kind of reversible nanometer porphyrin fluorescence sensor recognition quantitative chiral amino acid method described in accordance with the claim 1,
Be characterized in that, dichloride cadmium, N-acetyl-L-cysteine, sodium tellurite substance amount ratio be 1.0:(1.2~1.5):
0.2。
3. a kind of reversible nanometer porphyrin fluorescence sensor recognition quantitative chiral amino acid method described in accordance with the claim 1,
It is characterized in that, (4- pyridyl group) zinc protoporphyrin and the cetyl trimethylammonium bromide substance in mixed solution four-in step (2)
Amount is than being 1:1.3~1.5.
4. a kind of reversible nanometer porphyrin fluorescence sensor recognition quantitative chiral amino acid method described in accordance with the claim 1,
It is characterized in that, four-(4- pyridyl group) zinc protoporphyrins and CdTe quantum are molten in step (3) four-(4- pyridyl group) zinc protoporphyrin nanosphere
The mass ratio of the material of liquid is 40~42:1.
5. a kind of reversible nanometer porphyrin fluorescence sensor recognition quantitative chiral amino acid method described in accordance with the claim 1,
It is characterized in that, concentration, CdTe quantum the concentration difference of four-(4- pyridyl group) zinc protoporphyrin nanospheres in step (3) mixed solution
It is 8.0 × 10-9-1.28×10-7mol/L、5.8×10-9mol/L。
6. a kind of reversible nanometer porphyrin fluorescence sensor recognition quantitative chiral amino acid method described in accordance with the claim 1,
It is characterized in that, the D type and L-type amino acid of reversible nanometer porphyrin fluorescence sensor quantitative judge are respectively D/L- proline, D/L-
Lysine, D/L- serine.
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