CN105524079B - Ratio-type pH fluorescence probes, preparation method, application and test method for water-soluble positioning lysosome - Google Patents
Ratio-type pH fluorescence probes, preparation method, application and test method for water-soluble positioning lysosome Download PDFInfo
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Abstract
It is a kind of to be shown below for the Ratio-type pH fluorescence probes of water-soluble positioning lysosome, preparation method, application and test method, chemical structural formula：The probe of the present invention can detect the fluorescence intensity change of two launch wavelengths simultaneously, and ectocine is eliminated by self calibration, and accuracy in detection is high, rapid to pH variation responses, have excellent water solubility, have good anti-interference ability in the presence of various metals ion.In pH, fluorescence intensity change is notable in range from 5.8 to 6.8, adapts to pH under solutions of weak acidity and detects.Experiments have shown that the probe can selectively dye the lysosome in cell, the good light stability in cell, and to cytotoxic, can be used for the pH detections of living cells.The probe of the present invention has important application value in cell imaging and detection lysosome in terms of pH value variation.
Description
One, technical field
The present invention relates to a kind of Ratio-type pH fluorescence probes, preparation method, application and test method, especially a kind of use
In Ratio-type pH fluorescence probes, preparation method, application and the test method of water-soluble positioning lysosome.
Two, background technology
Intracellular ph value plays a significant role in many cell behaviors, such as cell growth, Apoptosis, signal transduction and
Cell Proliferation etc..As an important indicator of cell health, the minor change of internal pH can cause cell abnormal, even
It can cause cell dysfunction and serious disease, such as cancer and Alzheimer's disease.Intracellular pH value be not it is identical,
Cytoplasm is weakly alkaline.And lysosome internal pH-values are about 4.0-6.0, it is weakly acidic.Weak acid environment in lysosome can
With the function of the internal hydrolase of activation, the decomposing protein in cell metabolism.Therefore, it is used for the ratio of water-soluble positioning lysosome
Type pH fluorescence probes, preparation method, application and test method are that the pH value of lysosome is quantitatively detected to cell analysis or examined
Disconnected method detects the method master of intracellular ph value in existing Ratio-type pH fluorescence probes and preparation method and application
There are nuclear magnetic resonance, Absorption and fluorescence spectrum etc., detection technique of fluorescence is easy to operate due to high sensitivity, can be real-time
Monitoring and other advantages and be widely applied, but most of fluorescence probe is all the variation for detecting single wavelength fluorescent intensity,
Such probe output signal can be influenced by temperature, solvent polarity, concentration and probe concentration, can influence the accuracy of testing result.
Three, invention content
The object of the present invention one is a kind of Ratio-type pH fluorescence probes for water-soluble positioning lysosome,
The object of the present invention is the second is prepared by its of a kind of Ratio-type pH fluorescence probes for water-soluble positioning lysosome
Method,
The object of the present invention the third is a kind of Ratio-type pH fluorescence probes for water-soluble positioning lysosome application,
The object of the present invention is the fourth is a kind of test side for the water-soluble Ratio-type pH fluorescence probes for positioning lysosome
Method.
In order to overcome the technical drawbacks described above, the object of the present invention is to provide a kind of ratios for water-soluble positioning lysosome
Type pH fluorescence probes, preparation method, application and test method, therefore the fluorescence intensity for detecting two launch wavelengths simultaneously becomes
Change, ectocine is eliminated by self calibration, improves detection accuracy.
In order to achieve the above objectives, the technical solution adopted by the present invention is that：
A kind of Ratio-type pH fluorescence probes for water-soluble positioning lysosome, chemical structural formula are shown below：
。
Due to devising Ratio-type pH fluorescence probes, pass through the Ratio-type pH fluorescence probes for water-soluble positioning lysosome
Fluorescence resonance energy transfer FRET mechanism show therefore simultaneously detect two launch wavelengths fluorescence intensity change, by from
Ectocine is eliminated in calibration, improves detection accuracy.
The present invention devises, and a kind of Ratio-type pH fluorescence probes for water-soluble positioning lysosome have：
Nuclear magnetic resonance hydrogen spectruming determining result：1HNMR (DMSO-d 6 , 300 MHz), δ (ppm)=8.71-8.75 (m,
1H), 8.67 (d, J = 7.5 Hz, 1H), 8.14 (s, 1H), 7.80 (t, J = 4.5 Hz, 1H), 7.51
(t, J = 4.5 Hz, 2H), 7.39 (dd, J = 1.5 Hz, J = 7.5 Hz, 1H), 7.00 (t, J = 7.5
Hz, 1H), 6.30-6.43 (m, 6H), 4.03 (s, 3H), 3.39-3.48 (m, 4H), 3.18-3.32 (m,
10H), 1.56-1.64 (m, 2H), 1.39 (q, J = 7.5 Hz, 2H), 0.97-1.09 (m, 12H), 0.91
(t, J=7.5 Hz, 3H),
Magnetic resonance carbon composes measurement result：13C NMR (DMSO-d 6 , 75 MHz), δ (ppm) = 12.31 (4C),
13.48 (2C), 21.66 (2C), 25.12, 28.57, 29.96, 43.51 (3C), 64.31, 64.31, 97.27
(2C), 104.74 (2C), 108.07 (2C), 110.93 (2C), 116.01, 119.24, 121.51, 122.22,
123.60, 124.20, 128.25, 128.60, 130.65 (2C), 131.45, 132.68, 138.47 (2C),
152.51 (2C), 153.87,163.76,167.69,
High resolution mass spectrum measurement result： Calcd. [M-I]-: 733.3633; found value [M-I]-:
733.3623
Compound.
The present invention devises, a kind of preparation method of Ratio-type pH fluorescence probes for water-soluble positioning lysosome,
Its step：
Intermediate compound III, HOBT（I-hydroxybenzotriazole）, EDC [1- (3- dimethylamino-propyls) -3- ethyls carbon two
Inferior amine salt hydrochlorate], triethylamine be dissolved in DMF and obtain in intermediate solution, intermediate compound V is added in intermediate solution, instead
Final compound RMPM should be obtained,
Wherein：III chemical structural formula of intermediate compound is shown below：
；
V chemical structural formula of intermediate compound is shown below：
。
The present invention devises, a kind of preparation method of Ratio-type pH fluorescence probes for water-soluble positioning lysosome,
Its step：Its step：
III 1.0-1.4 mmol of intermediate compound, HOBT 1.0-1.4 mmol, EDC 1.0-1.4 mmol, triethylamine
1.8-2.2 mmol are dissolved in DMF18-22mL, and 0.8-1.2h is stirred at room temperature under nitrogen protection, obtain intermediate solution, then
V 0.8-1.2 mmol of intermediate compound are added in intermediate solution, under nitrogen protection, 18-22h, silica gel plate are stirred at room temperature
Tracking determines reaction end, reaction solution reduction vaporization is concentrated to give thick terminal compound, by thick terminal compound dichloromethane
It is extracted in water, collects organic phase, dry, then concentration is purified, obtained faint yellow solid i.e. terminal chemical combination by column chromatography
Object RMPM, yield 45.2%, fusing point:197-201℃.
The present invention devises, a kind of preparation method of Ratio-type pH fluorescence probes for water-soluble positioning lysosome,
Its step：Its step：
3- butyl -1- chlorine imidazo (1,5- α) pyridine-7-carboxylic acids ethyl ester, that is, intermediate compound I 8-12mmol and NaOH18-
22mmol is dissolved in the mixed solution of ethyl alcohol 18-22ml and water 13-17ml, after back flow reaction 1.8-2.2 h, by acquired solution
It pours into 180-220 ml water, neutrality is neutralized to dilute hydrochloric acid 1.8-2.2M, have yellow solid that filtering is precipitated, it is washed with water 2 ×
5mL in an oven after 50 DEG C of dryings, obtains yellow solid i.e. intermediate compound II, yield 90.8%,
By intermediate compound II 8-12mmol and CH3I 48-52mmol are dissolved in tetrahydrofuran 28-32ml solution,
Heating reflux reaction 7.8-8.2h under nitrogen protection is cooled to room temperature, and yellow solid is precipitated, and passes through vacuum filtration, filter cake second
Acetoacetic ester 8-12ml is washed 3 times, obtains the solid i.e. intermediate compound III of yellow, yield 73.0%,
Intermediate compound V can by being synthesized by the intermediate compound IV and reacting ethylenediamine of method in the literature,
Wherein：IV chemical structural formula of intermediate compound is shown below：
。
The present invention devises, a kind of preparation method of Ratio-type pH fluorescence probes for water-soluble positioning lysosome
Synthetic route is as follows:
The present invention devises, and a kind of Ratio-type pH fluorescence probes for water-soluble positioning lysosome are glimmering as Ratio-type pH
The application of light probe.
The present invention devises, and a kind of Ratio-type pH fluorescence probes for water-soluble positioning lysosome are as pH is measured
The application of 5.8 to 6.8 Ratio-type pH fluorescence probes.
The present invention devises, and a kind of Ratio-type pH fluorescence probes for water-soluble positioning lysosome are used as in Hela cells
Ratio-type pH fluorescence probes application.
The present invention devises, a kind of test method of Ratio-type pH fluorescence probes for water-soluble positioning lysosome,
Step：The Ratio-type pH fluorescence probes of above-mentioned water-soluble positioning lysosome are prepared, solution at various ph values (ethyl alcohol/
Britton--Robinson buffer solutions, 40mM acetic acid, phosphoric acid, boric acid, 1:4, volume ratio), surveyed by fluorescence spectrophotometry
Try its responding ability to different pH value；
It is quantitatively adding AgNO3, Al (NO3) 3, Ba respectively into the solution of the above-mentioned fluorescence probe containing pH in acid condition
(NO3) 2, Cd (NO3) 2, Co (NO3) 2, Cu (NO3) 2, Fe (NO3) 3, HgCl2, Ni (NO3) 2, Pb (NO3) 2, Zn
(NO3) 2 aqueous solution tests its anti-interference ability；
Using the above-mentioned pH fluorescence probes culture Hela cells of various concentration, fluorescence imaging is carried out respectively, and observation is intracellular
Fluorescence intensity change,
Using above-mentioned pH fluorescence probes and the lysosome probe Lysosensor Green DND-189 of commercialization to work
Hela cells dye altogether, and dyeing positioning is carried out by fluorescence imaging,
Using the above-mentioned pH fluorescence probes culture Hela cells of a certain concentration difference pH value, observes fluorescence intensity and red is glimmering
The variation of light/blue-fluorescence ratio,
Using certain density above-mentioned pH fluorescence probes culture Hela cells, under the continuous agitation of laser, observation blue
Photostability of the probe in Hela cells is tested in the variation of the ratio of fluorescence/red fluorescence,
After certain density above-mentioned 6 h of pH fluorescence probes culture Hela cells, detection probe is to Hela cell survivals
The influence of rate.
The technical effects of the invention are that：The preferred Hela cells of living cells.The pH fluorescence probes for positioning lysosome are rung
Should be rapid, there is excellent water solubility, there is good anti-interference ability to various metals ion.In pH from 5.8 to 6.8 in range
It is changed significantly, adapts to pH under solutions of weak acidity and detect.The probe can select, by the lysosome dyeing in Hela cells, to can be used for
The detection of pH in living cells.Above-mentioned probe has rhodamine-imidazo [1,5- α] pyridine structure, when pH value increases from 5.50
To 7.00, the blue-fluorescence of imidazo [1,5- α] pyridine groups is almost constant.Meanwhile the red fluorescence of rhodamine group
Decline.And the ratio of red fluorescence/blue-fluorescence is as pH value increases to 7.00 from 5.50 and increases.Two hairs can be detected simultaneously
The fluorescence intensity change of ejected wave length eliminates ectocine by self calibration, and accuracy in detection is high, rapid to pH variation responses, tool
There is excellent water solubility, to metal ion strong antijamming capability.In pH, fluorescence intensity change is notable in range from 5.8 to 6.8, fits
PH under solutions of weak acidity is answered to detect.Experiments have shown that the probe can selectively dye the lysosome in cell, in cell
Good light stability, to cytotoxic.The probe of the present invention has weight in cell imaging and detection lysosome in terms of pH value variation
The application value wanted.
In the technical scheme, column chromatography purifying is silica gel, methylene chloride/methanol=10：1, v/v.
In the technical scheme, the chemical structural formula of Ratio-type pH fluorescence probes is important technical characteristic, for water-soluble
Property the positioning Ratio-type pH fluorescence probes of lysosome, preparation method, application and test method technical field in, there is novelty
Property, creativeness and practicability, term in the technical scheme is all that can be solved with patent document in the art
It releases and understands.
Four, it illustrates
In order to more clearly explain the embodiment of the invention or the technical proposal in the existing technology, to embodiment or will show below
There is attached drawing needed in technology description to be briefly described, it should be apparent that, the accompanying drawings in the following description is only this
Some embodiments of invention for those of ordinary skill in the art without creative efforts, can be with
Obtain other attached drawings according to these attached drawings.
Fig. 1 is the chemical structural formula of intermediate compound III：
The fluorescence spectrum (a) and probe of Fig. 2 RMPM (10 μm) in the B-R buffer solutions of different pH value (5.6-7.2)
Fluorescence intensity ratio under corresponding pH value（I580/I426）(b), (λex=340 nm, slit:10 nm / 5 nm),n =
3：
Fig. 3 pH value is 5.6（a）When with 7.2 (b), RMPM (10 μ in the B-R buffer solutions containing different metal ions
M fluorescence intensity ratio I)580/I426, the concentration of all metal cations is 100 μM, (λex=340 nm, slit:10 nm
/ 5 nm) ：
Fig. 4 RMPM (10 μM) fluorescence intensity ratios I580 / I426In the B-R buffer solutions of pH 5.6 and pH 7.2
Reaction time：
Fig. 5 Hela cells and 5 μM of probe handle the fluorescence microscopy images after 2 h at 37 DEG C.Ctr:Do not visit
Needle.The first row:The fluorescent image (400-555 nm) of blue channel, the second row:The fluorescent image (560-of red channel
700 nm), the third line:Bright field image, fourth line:The first, the second and the third line covering image, λex = 405 nm。(*p
< 0.05;n = 3)：
Positioning of Fig. 6 probes in Hela Cytolysosomes.(a):RMPM probes (5 μM), (b): LysoSensors
Green DND-189 (1 μM), (c):(a) and the coverage diagram of (b), (d):Bright field image, (e):RMPM and
The common location coefficient figure of LysoSensors Green DND-189：
Fig. 7 is buffered after 2 μM of probes handle 2 h, then in the PBS containing 10 μM of nigericins of known pH value
After cultivating 0.5 h in liquid, the fluorescence microscopy images (a) of Hela cells.The pH value of PBS buffer solution is respectively pH5.5 (a-d),
PH6.0 (e-h), pH6.5 (i-l) and pH7.00 (m-p).First row:The fluorescent image (400-555 nm) of blue channel；
Secondary series:The fluorescent image (560-700 nm) of red channel；Third arranges:Bright field image；4th row:First, second and
The image of three row superpositions.λ is analyzed using image jex=405 nm fluorescence intensity ratios (b) as a result use ± SE to indicate (* p<
0.05 and * * p< 0.01；n = 3)：
The light resistance of Fig. 8 probes.After 2 μM of probes handle 2 h, in λexIt is surveyed under the continuous agitation of=405 nm laser
Try the light resistance of Hela cells：
After Fig. 9 Hela cells and 6 h of probe culture of known concentration, influence of the probe to Hela cell survival rates.(p >
0.05; n = 3)：
Figure 10 is the chemistry knot for Ratio-type pH fluorescence probes, that is, terminal compound RMPM of water-soluble positioning lysosome
Structure formula.
Five, specific implementation mode
With reference to embodiment, the present invention is further described, following embodiment is intended to illustrate invention rather than to this
Invention further limits.Its step is：
A kind of one embodiment of the preparation method of Ratio-type pH fluorescence probes for water-soluble positioning lysosome,
Its step：
3- butyl -1- chlorine imidazo (1,5- α) pyridine-7-carboxylic acids ethyl ester, that is, intermediate compound I 8mmol and NaOH18mmol
It is dissolved in the mixed solution of ethyl alcohol 18ml and water 13ml, after 1.8 h of back flow reaction, acquired solution is poured into 180 ml water,
It is neutralized to neutrality with dilute hydrochloric acid 1.8M, has yellow solid that filtering is precipitated, 2 × 5mL is washed with water, in an oven after 50 DEG C of dryings,
Obtain yellow solid i.e. intermediate compound II, yield 90.8%,
By intermediate compound II 8mmol and CH3I 48mmol are dissolved in tetrahydrofuran 28ml solution, under nitrogen protection
Heating reflux reaction 7.8h is cooled to room temperature, and yellow solid is precipitated, and by vacuum filtration, filter cake washs 3 with ethyl acetate 8ml
It is secondary, obtain the solid i.e. intermediate compound III of yellow, yield 73.0%,
Intermediate compound V can by being synthesized by the intermediate compound IV and reacting ethylenediamine of method in the literature,
III 1.0 1.0 1.0 mmol of mmol, EDC of mmol, HOBT of intermediate compound, 1.8 mmol of triethylamine are dissolved in
In DMF18mL, 0.8h is stirred at room temperature under nitrogen protection, obtains intermediate solution, then adds V 0.8 mmol of intermediate compound
Enter into intermediate solution, under nitrogen protection, 18h is stirred at room temperature, silica gel plate tracking determines reaction end, reaction solution is depressurized and is steamed
Hair is concentrated to give thick terminal compound, will be extracted in thick terminal compound dichloromethane and water, collects organic phase, dry, concentration,
Then it is purified by column chromatography, obtained faint yellow solid i.e. terminal compound RMPM, yield 45.2%, fusing point:197-201
℃。
Wherein：III chemical structural formula of intermediate compound is shown below：
；
Wherein：IV chemical structural formula of intermediate compound is shown below：
V chemical structural formula of intermediate compound is shown below：
。
A kind of synthetic route of the preparation method of Ratio-type pH fluorescence probes for water-soluble positioning lysosome is as follows:
The fluorescence resonance energy transfer FRET mechanism of Ratio-type pH fluorescence probes for water-soluble positioning lysosome：
A kind of second embodiment of the preparation method of Ratio-type pH fluorescence probes for water-soluble positioning lysosome,
Its step：
3- butyl -1- chlorine imidazo (1,5- α) pyridine-7-carboxylic acids ethyl ester, that is, intermediate compound I 8-12mmol and NaOH18-
22mmol is dissolved in the mixed solution of ethyl alcohol 18-22ml and water 13-17ml, after back flow reaction 1.8-2.2 h, by acquired solution
It pours into 180-220 ml water, neutrality is neutralized to dilute hydrochloric acid 1.8-2.2M, have yellow solid that filtering is precipitated, it is washed with water 2 ×
5mL in an oven after 50 DEG C of dryings, obtains yellow solid i.e. intermediate compound II, yield 90.8%,
By intermediate compound II 8-12mmol and CH3I 48-52mmol are dissolved in tetrahydrofuran 28-32ml solution,
Heating reflux reaction 7.8-8.2h under nitrogen protection is cooled to room temperature, and yellow solid is precipitated, and passes through vacuum filtration, filter cake second
Acetoacetic ester 8-12ml is washed 3 times, obtains the solid i.e. intermediate compound III of yellow, yield 73.0%,
Intermediate compound V can by being synthesized by the intermediate compound IV and reacting ethylenediamine of method in the literature,
III 1.0-1.4 mmol of intermediate compound, HOBT 1.0-1.4 mmol, EDC 1.0-1.4 mmol, triethylamine
1.8-2.2 mmol are dissolved in DMF18-22mL, and 0.8-1.2h is stirred at room temperature under nitrogen protection, obtain intermediate solution, then
V 0.8-1.2 mmol of intermediate compound are added in intermediate solution, under nitrogen protection, 18-22h, silica gel plate are stirred at room temperature
Tracking determines reaction end, reaction solution reduction vaporization is concentrated to give thick terminal compound, by thick terminal compound dichloromethane
It is extracted in water, collects organic phase, dry, then concentration is purified, obtained faint yellow solid i.e. terminal chemical combination by column chromatography
Object RMPM, yield 45.2%, fusing point:197-201℃.
A kind of third embodiment of the preparation method of Ratio-type pH fluorescence probes for water-soluble positioning lysosome,
Its step：
3- butyl -1- chlorine imidazo (1,5- α) pyridine-7-carboxylic acids ethyl ester, that is, I 12mmol of intermediate compound and
NaOH22mmol is dissolved in the mixed solution of ethyl alcohol 22ml and water 17ml, and after 2.2 h of back flow reaction, acquired solution is poured into
In 220 ml water, it is neutralized to neutrality with dilute hydrochloric acid 2.2M, has yellow solid that filtering is precipitated, 2 × 5mL is washed with water, in an oven
After 50 DEG C of dryings, yellow solid i.e. intermediate compound II is obtained, yield 90.8%,
By intermediate compound II 12mmol and CH3I 52mmol are dissolved in tetrahydrofuran 32ml solution, in nitrogen protection
Lower heating reflux reaction 8.2h is cooled to room temperature, and yellow solid is precipitated, and by vacuum filtration, filter cake is washed with ethyl acetate 12ml
It washs 3 times, obtains the solid i.e. intermediate compound III of yellow, yield 73.0%,
Intermediate compound V can by being synthesized by the intermediate compound IV and reacting ethylenediamine of method in the literature,
III 1.4 1.4 mmol of mmol, HOBT of intermediate compound, EDC1.4 mmol, 2.2 mmol of triethylamine are dissolved in
In DMF22mL, 1.2h is stirred at room temperature under nitrogen protection, obtains intermediate solution, then adds V 1.2 mmol of intermediate compound
Enter into intermediate solution, under nitrogen protection, 22h is stirred at room temperature, silica gel plate tracking determines reaction end, reaction solution is depressurized and is steamed
Hair is concentrated to give thick terminal compound, will be extracted in thick terminal compound dichloromethane and water, collects organic phase, dry, concentration,
Then it is purified by column chromatography, obtained faint yellow solid i.e. terminal compound RMPM, yield 45.2%, fusing point:197-201
℃。
A kind of 4th embodiment of the preparation method of Ratio-type pH fluorescence probes for water-soluble positioning lysosome,
Its step：
3- butyl -1- chlorine imidazo (1,5- α) pyridine-7-carboxylic acids ethyl ester, that is, I 10mmol of intermediate compound and
NaOH20mmol is dissolved in the mixed solution of ethyl alcohol 20ml and water 15ml, and after 2.0 h of back flow reaction, acquired solution is poured into
In 200 ml water, it is neutralized to neutrality with dilute hydrochloric acid 2.0M, has yellow solid that filtering is precipitated, 2 × 5mL is washed with water, in an oven
After 50 DEG C of dryings, yellow solid i.e. intermediate compound II is obtained, yield 90.8%,
By intermediate compound II 10mmol and CH3I50mmol is dissolved in tetrahydrofuran 30ml solution, under nitrogen protection
Heating reflux reaction 8.0h is cooled to room temperature, and yellow solid is precipitated, and by vacuum filtration, filter cake washs 3 with ethyl acetate 10ml
It is secondary, obtain the solid i.e. intermediate compound III of yellow, yield 73.0%,
Intermediate compound V can by being synthesized by the intermediate compound IV and reacting ethylenediamine of method in the literature,
III 1.2 1.2 mmol of mmol, HOBT of intermediate compound, EDC 1.2mmol, 2.0 mmol of triethylamine are dissolved in
In DMF20mL, 1.0h is stirred at room temperature under nitrogen protection, obtains intermediate solution, then adds V 1.0 mmol of intermediate compound
Enter into intermediate solution, under nitrogen protection, 20h is stirred at room temperature, silica gel plate tracking determines reaction end, reaction solution is depressurized and is steamed
Hair is concentrated to give thick terminal compound, will be extracted in thick terminal compound dichloromethane and water, collects organic phase, dry, concentration,
Then it is purified by column chromatography, obtained faint yellow solid i.e. terminal compound RMPM, yield 45.2%, fusing point:197-201
℃。
A kind of Ratio-type pH fluorescence probes, that is, terminal compound RMPM for water-soluble positioning lysosome, chemical constitution
Formula is shown below：
The spectrogram of Ratio-type pH fluorescence probes, that is, terminal compound RMPM for water-soluble positioning lysosome is measured：
Nuclear magnetic resonance hydrogen spectruming determining：1HNMR (DMSO-d 6 , 300 MHz), δ (ppm)=8.71-8.75 (m, 1H),
8.67 (d, J = 7.5 Hz, 1H), 8.14 (s, 1H), 7.80 (t, J = 4.5 Hz, 1H), 7.51 (t, J
= 4.5 Hz, 2H), 7.39 (dd, J = 1.5 Hz, J = 7.5 Hz, 1H), 7.00 (t, J = 7.5 Hz,
1H), 6.30-6.43 (m, 6H), 4.03 (s, 3H), 3.39-3.48 (m, 4H), 3.18-3.32 (m, 10H),
1.56-1.64 (m, 2H), 1.39 (q, J = 7.5 Hz, 2H), 0.97-1.09 (m, 12H), 0.91 (t, J =
7.5 Hz, 3H)。
Magnetic resonance carbon spectrum measures：13C NMR (DMSO-d 6 , 75 MHz), δ (ppm) = 12.31 (4C), 13.48
(2C), 21.66 (2C), 25.12, 28.57, 29.96, 43.51 (3C), 64.31, 64.31, 97.27 (2C),
104.74 (2C), 108.07 (2C), 110.93 (2C), 116.01, 119.24, 121.51, 122.22,
123.60, 124.20, 128.25, 128.60, 130.65 (2C), 131.45, 132.68, 138.47 (2C),
152.51 (2C), 153.87, 163.76, 167.69。
High resolution mass spectrum measures： Calcd. [M-I]-: 733.3633; found value [M-I]-: 733.3623。
A kind of test method of Ratio-type pH fluorescence probes for water-soluble positioning lysosome, step：It prepares above-mentioned
The Ratio-type pH fluorescence probes of water solubility positioning lysosome, solution (ethyl alcohol/Britton--Robinson at various ph values
Buffer solution, 40mM acetic acid, phosphoric acid, boric acid, 1:4, volume ratio), it is tested to different pH value by fluorescence spectrophotometry
Responding ability；
It is quantitatively adding AgNO3, Al (NO3) 3, Ba respectively into the solution of the above-mentioned fluorescence probe containing pH in acid condition
(NO3) 2, Cd (NO3) 2, Co (NO3) 2, Cu (NO3) 2, Fe (NO3) 3, HgCl2, Ni (NO3) 2, Pb (NO3) 2, Zn
(NO3) 2 aqueous solution tests its anti-interference ability；
Using the above-mentioned pH fluorescence probes culture Hela cells of various concentration, fluorescence imaging is carried out respectively, and observation is intracellular
Fluorescence intensity change,
Using above-mentioned pH fluorescence probes and the lysosome probe Lysosensor Green DND-189 of commercialization to work
Hela cells dye altogether, and dyeing positioning is carried out by fluorescence imaging,
Using the above-mentioned pH fluorescence probes culture Hela cells of a certain concentration difference pH value, observes fluorescence intensity and red is glimmering
The variation of light/blue-fluorescence ratio,
Using certain density above-mentioned pH fluorescence probes culture Hela cells, under the continuous agitation of laser, observation blue
Photostability of the probe in Hela cells is tested in the variation of the ratio of fluorescence/red fluorescence,
After certain density above-mentioned 6 h of pH fluorescence probes culture Hela cells, detection probe is to Hela cell survivals
The influence of rate.
The measurement embodiment 1 of Ratio-type pH fluorescence probes, that is, terminal compound RMPM for water-soluble positioning lysosome：
Prepare the solution (ethyl alcohol/Britton-- of the above-mentioned fluorescence probe RMPM of 20mL (10 μM) at various ph values
Robinson buffer solutions, 40mM acetic acid, phosphoric acid, boric acid, 1:4, volume ratio), fluorescence light is tested by fluorescence spectrophotometry
Spectrum.The fluorescence intensity ratio of Fig. 2 (a) and probe in the region pH value (5.6-7.2)（I580/I426）Fig. 2 (b).
As a result it shows：It is in pH for the water-soluble Ratio-type pH fluorescence probes, that is, terminal compound RMPM for positioning lysosome
Fluorescence intensity change is notable in 5.8 and 6.8 solution, fluorescent emission ratio I580/I426Reached peak and at room temperature at 4 minutes
It keeps stablizing, this shows that RMPM can change pH value quick response, and accuracy in detection is high.(Fig. 2,4)
The measurement embodiment 2 of Ratio-type pH fluorescence probes, that is, terminal compound RMPM for water-soluble positioning lysosome：
PH value be 5.6 and 7.2 under the conditions of to the above-mentioned RMPM of fluorescence probe containing pH (10 μM) solution (ethyl alcohol/
Britton-Robinson buffer solutions, 40mM acetic acid, phosphoric acid, boric acid, 1:4, volume ratio) in be separately added into AgNO3, Al
(NO3) 3, Ba (NO3) 2, Cd (NO3) 2, Co (NO3) 2, Cu (NO3) 2, Fe (NO3) 3, HgCl2, Ni (NO3) 2, Pb
(NO3) concentration of the aqueous solution of 2, Zn (NO3) 2, all metal cations is 100 μM, and test is containing different metal ions
Solution in fluorescence intensity ratio I580/I426(λex=340 nm, Fig. 3).
As a result it shows：For water-soluble positioning lysosome Ratio-type pH fluorescence probes, that is, terminal compound RMPM pH=
5.6 and pH=7.2 under the conditions of to above-mentioned metal ion almost without response, it was demonstrated that the probe is to H+With very high selectivity.
Strong interference immunities of the probe RMPM to common metal ion.
The measurement embodiment 4 of Ratio-type pH fluorescence probes, that is, terminal compound RMPM for water-soluble positioning lysosome：
(1,2 or 5 μM of Hela cells and probe RMPM）2 h are cultivated under the conditions of 37 DEG C.Under laser confocal microscope
Observation.Fixed excitation wavelength is 405nm, collects the fluorescent image (400-555 nm) that emission band is respectively blue channel,
The fluorescent image (560-700 nm) of red channel.In Hela cells blue and red fluorescence intensity all with probe it is dense
The increase of degree and increase, this shows that the Ratio-type pH fluorescence probes i.e. terminal compound RMPM for water-soluble positioning lysosome has
The ability (Fig. 5) that living cells is dyed.
The measurement embodiment 5 of Ratio-type pH fluorescence probes, that is, terminal compound RMPM for water-soluble positioning lysosome：
Hela cells and probe RMPM (5 μM) cultivate 2h at 37 DEG C, then with containing Lysosensor Green DND-
The fresh culture of 189 (1 μM) further cultivates 1 h.Respectively obtain probe RMPM blue fluorescence photograph (Fig. 6 a) and
Two photos are superimposed by the green fluorescence photo (Fig. 6 b) of Lysosensor Green DND-189, blue and green overlapping
Region shows cyan（Fig. 6 c）.The result shows that two photo pigmented sections can overlap well, common location coefficient is 0.931,
The Ratio-type pH fluorescence probes, that is, terminal compound RMPM for being used for water-soluble positioning lysosome can selectively mark lysosome
Note（Fig. 6）.
The measurement embodiment 6 of Ratio-type pH fluorescence probes, that is, terminal compound RMPM for water-soluble positioning lysosome：
By Hela cells respectively in different pH value（5.50,6.00,6.50,7.00）RMPM（4.00 μM, 5.00 μM,
6.00 μM and 7.00 μM）2 h of middle culture.Then processed Hela cells are buffered in the PBS containing 10 μM of nigericins
0.5 h is cultivated in liquid.From the image that Laser Scanning Confocal Microscope is shot (Fig. 7 a), when pH value increases to 7.00 from 5.50, miaow
Azoles simultaneously [1,5- α] pyridine groups（First row）Blue-fluorescence be almost constant, while rhodamine group（Secondary series）It is red
Color fluorescence declines.And the ratio of red fluorescence/blue-fluorescence is as pH value increases to 7.00 from 5.50 and increases (Fig. 7 b).
The measurement embodiment 7 of Ratio-type pH fluorescence probes, that is, terminal compound RMPM for water-soluble positioning lysosome：
After the method processing same as above of Hela cells, in λexThe laser of=405 nm is in different time（30S,60S,
90S,120S,180 S）Under continuous agitation, from the image that Laser Scanning Confocal Microscope is shot, blue-fluorescence/red fluorescence
Ratio is substantially constant（Fig. 8）, show the Ratio-type pH fluorescence probes i.e. terminal compound for water-soluble positioning lysosome
The photostability that RMPM has had in Hela cells.
The measurement embodiment 8 of Ratio-type pH fluorescence probes, that is, terminal compound RMPM for water-soluble positioning lysosome：
After Hela cells cultivate 6h with probe RMPM (5 μM) at 37 DEG C, Hela cell survival rates, knot are detected with mtt assay
Fruit shows the Ratio-type pH fluorescence probes i.e. terminal compound RMPM for being used for water-soluble positioning lysosome under experimental conditions to thin
Born of the same parents do not have toxicity (Fig. 9).
The present invention has the following characteristics：
1, due to devising Ratio-type pH fluorescence probes, the fluorescence intensity change of two launch wavelengths can be detected simultaneously, led to
It crosses self calibration and eliminates ectocine, improve detection accuracy..
2, due to devising Ratio-type pH fluorescence probes, there is excellent water solubility, to metal ion strong antijamming capability.
3, due to devising Ratio-type pH fluorescence probes, the energy transmission of FRET donors is given to the receptor in ground state, this
A process can reduce the error of fluoroscopic examination.
4, the compound structure new due to devising Ratio-type pH fluorescence probes, experiments have shown that the compound structure feature takes
Obtained good technique effect.
5, due to devising the technical characteristic of the present invention, in the effect of the independent and mutual set of technical characteristic,
By experiments have shown that, property indices of the invention are that existing property indices want high, are had by assessment good
Market value.
Technical characteristic as also other and Ratio-type pH fluorescence probes chemical structural formulas are same or similar is all this
One of embodiment of invention, and each technical characteristic of embodiment described above can be combined arbitrarily, to meet patent
The requirement of method, patent regulation and guidelines for examination, no longer to all possible group of each technical characteristic in above-described embodiment
The embodiment of conjunction is all described.
Above-described embodiment be Ratio-type pH fluorescence probes provided by the present invention for water-soluble positioning lysosome, its
A kind of way of realization of preparation method, application and test method, according to scheme provided by the present invention other deformation, increase or
Person reduces composition therein or step, or the present invention is belonged to this for other technical fields close with the present invention
The protection domain of invention.
Claims (5)
1. a kind of Ratio-type pH fluorescence probes for water-soluble positioning lysosome, it is characterized in that：Its chemical structural formula such as following formula
It is shown：
。
2. the preparation method of the Ratio-type pH fluorescence probes according to claim 1 for water-soluble positioning lysosome,
It is characterized in：Its step：
Intermediate compound III, HOBT（I-hydroxybenzotriazole）, EDC, triethylamine be dissolved in DMF and obtain intermediate solution, will in
Between compound V be added to intermediate solution, final compound is obtained by the reaction,
Wherein：III chemical structural formula of intermediate compound is shown below：
；
V chemical structural formula of intermediate compound is shown below：
。
3. the preparation method of the Ratio-type pH fluorescence probes according to claim 2 for water-soluble positioning lysosome,
It is characterized in：Its step：
III 1.0-1.4 mmol of intermediate compound, HOBT 1.0-1.4 mmol, EDC 1.0-1.4 mmol, triethylamine 1.8-2.2
Mmol is dissolved in DMF18-22mL, and 0.8-1.2h is stirred at room temperature under nitrogen protection, obtains intermediate solution, then by centreization
It closes V 0.8-1.2 mmol of object to be added in intermediate solution, under nitrogen protection, 18-22h is stirred at room temperature, silica gel plate tracking determines
Reaction solution reduction vaporization is concentrated to give thick terminal compound, thick terminal compound is extracted in dichloromethane and water by reaction end
It takes, collects organic phase, dry, then concentration is purified, obtained faint yellow solid i.e. terminal compound, yield by column chromatography
45.2%, fusing point:197-201℃.
4. the preparation method of the Ratio-type pH fluorescence probes according to claim 2 for water-soluble positioning lysosome,
It is characterized in：Its step：
3- butyl -1- chlorine imidazo (1,5- α) pyridine-7-carboxylic acids ethyl ester, that is, intermediate compound I 8-12mmol and NaOH18-
22mmol is dissolved in the mixed solution of ethyl alcohol 18-22ml and water 13-17ml, after back flow reaction 1.8-2.2 h, by acquired solution
It pours into 180-220 ml water, neutrality is neutralized to dilute hydrochloric acid 1.8-2.2M, have yellow solid that filtering is precipitated, it is washed with water 2 ×
5mL in an oven after 50 DEG C of dryings, obtains yellow solid i.e. intermediate compound II, yield 90.8%,
By intermediate compound II 8-12mmol and CH3I 48-52mmol are dissolved in tetrahydrofuran 28-32ml solution, are protected in nitrogen
The lower heating reflux reaction 7.8-8.2h of shield, is cooled to room temperature, yellow solid is precipitated, pass through vacuum filtration, filter cake ethyl acetate
8-12ml is washed 3 times, obtains the solid i.e. intermediate compound III of yellow, yield 73.0%,
Intermediate compound V can by being synthesized by the intermediate compound IV and reacting ethylenediamine of method in the literature,
Wherein：IV chemical structural formula of intermediate compound is shown below：
。
5. the preparation method of the Ratio-type pH fluorescence probes according to claim 2 for water-soluble positioning lysosome,
It is characterized in：The synthetic route of Ratio-type pH fluorescence probe preparation methods for water-soluble positioning lysosome is as follows:
。
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN114262333B (en) * | 2020-09-16 | 2023-01-17 | 中国科学院大连化学物理研究所 | Near-infrared fluorescent dye for lysosome super-resolution imaging and preparation method and application thereof |
| CN114656476A (en) * | 2022-02-18 | 2022-06-24 | 晋中学院 | Lysosome targeting rhodamine B hydrazide fluorescent probe as well as preparation method and application thereof |
Citations (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102219789A (en) * | 2011-04-08 | 2011-10-19 | 山东大学 | Chromone acylhydrazone derivative of rhodamine B, and preparation method and application thereof |
| CN102321093A (en) * | 2011-07-06 | 2012-01-18 | 山东大学 | A kind of is the chloro chromone acyl hydrazone derivative of parent and as the application of fluorescent probe with the rhodamine B |
| CN102746313A (en) * | 2012-06-20 | 2012-10-24 | 山东大学 | Rhodamine-B hydrazide derivative containing 1,2,4-triazole structural unit, preparation method and application thereof |
| CN102942919A (en) * | 2012-11-13 | 2013-02-27 | 山东大学 | Application of rhodamine B thio-bishydrazide derivative as Hg<2+> fluorescent probe |
| CN103012418A (en) * | 2012-12-04 | 2013-04-03 | 山东大学 | 1,3,4-oxadizaole structure unit-containing Rhodamine B pH fluorescence probe and applications thereof |
| CN104017567A (en) * | 2014-04-29 | 2014-09-03 | 苏州科技学院 | High-molecular pH probe containing rhodamine lactam group and synthetic method thereof |
| CN104277826A (en) * | 2014-10-16 | 2015-01-14 | 天津工业大学 | Fluorescent probe for intracellular Hg<2+> detection by using oxygen atoms as combination sites |
-
2016
- 2016-02-04 CN CN201610078944.XA patent/CN105524079B/en active Active
Patent Citations (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102219789A (en) * | 2011-04-08 | 2011-10-19 | 山东大学 | Chromone acylhydrazone derivative of rhodamine B, and preparation method and application thereof |
| CN102321093A (en) * | 2011-07-06 | 2012-01-18 | 山东大学 | A kind of is the chloro chromone acyl hydrazone derivative of parent and as the application of fluorescent probe with the rhodamine B |
| CN102746313A (en) * | 2012-06-20 | 2012-10-24 | 山东大学 | Rhodamine-B hydrazide derivative containing 1,2,4-triazole structural unit, preparation method and application thereof |
| CN102942919A (en) * | 2012-11-13 | 2013-02-27 | 山东大学 | Application of rhodamine B thio-bishydrazide derivative as Hg<2+> fluorescent probe |
| CN103012418A (en) * | 2012-12-04 | 2013-04-03 | 山东大学 | 1,3,4-oxadizaole structure unit-containing Rhodamine B pH fluorescence probe and applications thereof |
| CN104017567A (en) * | 2014-04-29 | 2014-09-03 | 苏州科技学院 | High-molecular pH probe containing rhodamine lactam group and synthetic method thereof |
| CN104277826A (en) * | 2014-10-16 | 2015-01-14 | 天津工业大学 | Fluorescent probe for intracellular Hg<2+> detection by using oxygen atoms as combination sites |
Non-Patent Citations (3)
| Title |
|---|
| A ratiometric lysosomal pH probe based on the coumarin–rhodamine FRET system;Xiao-Fan Zhang et al.;《RSC Advances》;20150528;第5卷;第49115-49121页，摘要，第49116页Scheme 1，右栏最后1段至第49119页右栏第1段，FIG.1-8 * |
| A ratiometric lysosomal pH probe based on the naphthalimide– rhodamine system;Xiao-Fan Zhang et al.;《Journal of Materials Chemistry B》;20150304;第3卷;第3260-3266页，摘要，第3261页Scheme 1，右栏第2段至第3264页右栏第1段 * |
| A selective turn-on fluorescent chemosensor based on rhodamine for Hg2+ and its application in live cell imaging;Fanyong Yan et al.;《Sensors and Actuators B》;20111229;第162卷;第313-320页 * |
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