CN102830099A - pH fluorescent sensor based on allyl calcein and preparation method thereof - Google Patents
pH fluorescent sensor based on allyl calcein and preparation method thereof Download PDFInfo
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- CN102830099A CN102830099A CN2012102531896A CN201210253189A CN102830099A CN 102830099 A CN102830099 A CN 102830099A CN 2012102531896 A CN2012102531896 A CN 2012102531896A CN 201210253189 A CN201210253189 A CN 201210253189A CN 102830099 A CN102830099 A CN 102830099A
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Abstract
The invention discloses a pH fluorescent sensor based on allyl calcein and a preparation method of the pH fluorescent sensor. The preparation method comprises the following steps: taking a fluorescent carrier in a film substrate monomer as the allyl calcein; dissolving 15mg of the allyl calcein in 0.2ml of N, N-dimethylformamide for sufficient dissolving; sequentially adding 200mg of acrylamide, 0.4ml of hydroxyethyl methylacrylate, 0.15ml of cross-linking agent triethoxy trimethyl acrylate and 0.2ml of triethanolamine; uniformly mixing to drop on a clean Teflon plate; covering a quartz slide after silanization on the Teflon plate; irradiating for 30 minutes by an ultraviolet lamp; washing by water and ethanol; and drying at room temperature. Fluorescent molecules of the pH fluorescent sensor are easy to synthesize, loss of the fluorescent carrier is effectively blocked through a covalence fixing method; and the sensor is simple in structure and expresses good stability, reproducibility and reversibility.
Description
Technical field
The present invention relates to a kind of fluorescent optical sensor, be specifically related to a kind of pH fluorescent optical sensor based on the allyl calcein and preparation method thereof.
Background technology
PH measures with many fields closely bound up.Carry out tending to produce certain difficulty when pH measures with the conventional glass electrode, this is because glass electrode exists impedance height, cracky, can not be used for the mensuration of the pH value of solution of hydrofluoric acid containing, and under high basicity situation, has defectives such as " sodium errors ".Because its volume is big, be not suitable for the online detection of microcell, microenvironment and biological living simultaneously, more difficult the adaptation, used under rugged surroundings.
Optical fiber pH fluorescent optical sensor mainly is a sensitive fluorescent membrane of modifying one deck identification pH through optical fiber end, detects the pH value of solution.Such sensor is measured wide dynamic range because to have equilibration time fast, is easy to demarcate, and measuring-signal is stable, be easy to carry, and the data long-distance transmissions, service life of structure is long, is difficult for characteristics such as impaired and constantly develops.In recent years; Increasing pH sensitive fluorescence carrier such as naphthalimide analog derivative, Coumarins derivant etc. are used to prepare the pH fluorescent optical sensor; This has expanded pH application of sensor space greatly, and the analysis to measure of pH value has also been obtained breakthrough progress under various solution environmentals.Yet quite a few material that can be used to prepare sensitive fluorescent membrane has shortcomings such as complex synthetic route, pH responding range be narrow.Calcein itself is exactly a fluorescent reagent, and is cheap, and its derivant extensively is prepared into the fluorescence probe of metallic ion, and the application in the pH context of detection does not appear in the newspapers about the calcein derivant.
Main fixing means to fluorescence carrier has two kinds: 1, based on the fixing means of embedding.Embedding method is simple, is exactly that fluorescence carrier is easy to run off but a very big shortcoming is arranged.2, fluorescence carrier is bonded on polymeric membrane or the solid matrix through Covalent Immobilization.This method makes fluorescence carrier run off hardly and has advantages such as the auroral poles membrane lifetime is long.In addition, most fluorescent pH sensors mainly is the measurement that the change of length or the stokes displacement of variation, fluorescence lifetime through measuring fluorescence intensity realizes pH value of solution.But the cost of equipment of measuring fluorescence lifetime is higher, and seldom the stokes displacement of fluorescence carrier changes along with the variation of pH simultaneously, and these two kinds of methods are restricted in practical application.
Therefore, develop a kind of preparation easily and fast, long novel pH fluorescent optical sensor of portable, life-span has very important meaning.Constantly the occurring of fluorescence carrier makes the pH sensor get into a new field.And immobilization technology is also perfect constantly, has overcome with glass electrode to carry out all drawbacks that pH measures.As everyone knows, aspects such as the sensitivity of many sensors, stability, selectivity and repeatability face great challenge.Therefore, more function property sensitive material can studied and find out to hope, and the problem that we faced so just might be resolved.
Summary of the invention
The objective of the invention is to overcome the defective of above-mentioned pH fluorescent optical sensor, develop a kind of pH fluorescent optical sensor based on new support and preparation method thereof, is carrier with the allyl calcein; Response time is short; Good stability, long service life, the pH that is mainly used in alkaline solution measures.
For achieving the above object, the present invention provides the pH fluorescent optical sensor based on the allyl calcein, comprises the slide of silanization, and the membrane matrix monomer is attached to the auroral poles film on the slide through photopolymerization, and the fluorescence carrier in the said membrane matrix monomer is the allyl calcein.
Allyl calcein structural formula is following:
Preparation method based on the pH fluorescent optical sensor of allyl calcein comprises the silanization of slide, the preparation of auroral poles film, and the preparation method of said auroral poles film is following:
1), 15mg allyl calcein is dissolved in 0.2ml N, in the dinethylformamide, fully dissolving;
2), in the solution of step 1), add the 200mg acrylic amide successively, the 0.4ml hydroxyethyl methacrylate, 0.15ml crosslinking chemical triethoxy trimethyl acrylic ester, the 0.2ml triethanolamine adds 0.25ml 1-hydroxy-cyclohexyl phenyl ketone at last, mixes;
3), removing step 2) in the drips of solution that obtains on polyfluortetraethylene plate; Using, the slide of silanization covers on it; Irradiation is 30 minutes under uviol lamp, successively water and alcohol flushing then, drying at room temperature; Make the slide that contains auroral poles film thin layer, promptly make pH fluorescent optical sensor based on the allyl calcein.
To be pack into the top of teflon flow cell of auroral poles film that fluorescence carrier is processed with the allyl calcein; Testing sample is with the speed input flow cell of 1.5ml/min; Measure fluorescence intensity at maximum excitation wavelength 505nm and emission wavelength 526nm place, and measure the pH value of unknown appearance according to correction equation.
Beneficial effect: the present invention prepares sensor auroral poles film with allyl calcein Covalent Immobilization on the surface of glass slide of silanization, on Perkin-Elmer Ls-55 phosphorescence spectrophotometer, measures the fluorescence intensity of auroral poles film in the different pH solution then.Exist linear relationship between the fluorescence intensity of auroral poles film and the variation of pH.And utilize this linear relationship just can detect the pH in the actual water of pH scope some between 6.5-12.0.Sensor sheet has revealed excellent selectivity, reappearance, response speed simultaneously.The accuracy of its measurement is also very high, through the test (waste water appearance, river appearance, river appearance) to some actual water sample, finds that the result is the same basically with the glass electrode measurement result, can be used for the mensuration of actual water sample fully.
The fluorescence molecule of the present invention's design is easy to synthesize, and has effectively stoped the loss of fluorescence carrier through the method for Covalent Immobilization; And this sensor construction is simple, has shown good stability, reappearance and reversibility; According to experimental technique, the drawing curve, the pH value has presented good linear relationship in the 6.5-12.0 scope and between the fluorescence intensity; Realized continuous, real-time, online mensuration to pH value in the solution.
Description of drawings
Fig. 1 is the fluorescence spectrum figure of sensitive membrane in the different pH solution, is respectively from top to bottom: (1) 6.5 (2) 7.0 (3) 7.5 (4) 8.0 (5) 9.0 (6) 10.0 (7) 11.0 (8) 11.5 (9) 12.0 (10) 12.5.As can be seen from the figure the fluorescence intensity along with the minimizing auroral poles film of pH also significantly reduces, and the maximum excitation of auroral poles film and emission wavelength do not change, and can measure the pH value of solution through the variation of fluorescence intensity.
Fig. 2 is a fluorescence intensity situation over time when circulation feeds different pH value.Wherein horizontal ordinate is the time, and ordinate is a fluorescence intensity.PH is followed successively by from high to low: (1) 12.0 (2) 10.4 (3) 8.5 (4) 7.5.The result has shown the variation from a kind of solution to another kind of fluorescence intensity of solution, and 5 times repetitive cycling is measured fluorescence intensity, and reappearance and reversibility are all more satisfactory, and release time and change in concentration order are also irrelevant.This sensor sheet has revealed good stability, reappearance and reversibility.
Fig. 3 is the linear relationship between pH value and the fluorescence intensity, and as can be seen from the figure the pH value has presented good linear relationship in the 6.5-12.0 scope and between the fluorescence intensity.Therefore we can measure the value of fluorescence intensity under the different pH and correspond to the true pH value that just can obtain solution on the curve.
Embodiment
Embodiment 1: allyl calcein synthetic
Allyl calcein synthesis step list of references: Wuhan Institute of Chemical Technology's journal, the synthetic route among the 1998:V20 15-17 is synthetic, and concrete steps are as follows:
(1) weighing 500mg calcein is dissolved in the NaOH solution of pH=13, adds lime chloride with 1:1.1 (mol ratio), after the dissolving, regulates pH with NaOH again and makes the pH value greater than 12 fully.Add the 0.4ml bromopropene and add 0.1ml 18-hat-6 ethers simultaneously, on DF-101B heat collecting type constant temperature blender with magnetic force, reacting 3 hours under 60 ℃ the condition.
(2), transfer to acidity with HCl with reacted solution.After solution is yellow, put into the refrigerator cooling.Can obtain the product (MS:657) that needs in 6 hours 70 ℃ of following vacuum drying behind second day suction filtration.
Under alkali condition, the hydrogen ion on 5 carboxyls of allyl calcein is all replaced, along with the increase of pH value, and hydrionic replaced many more of fluorescence, fluorescence intensity descends.Under different pH, its degree of ionization is different, will exist in solution with different shape, so difference has just appearred in its fluorescence intensity.
Embodiment 2:
The preparation and the mensuration of fluorescent optical sensor may further comprise the steps:
1, the silanization of slide: slide (diameter 12.5mm) immerses in the chromic acid lotion and soaked 40 minutes; Putting into 3% hydrofluorite successively soaked 30 minutes; Totally make with distilled water flushing then and fully no longer speckle with 3% hydrofluorite, add again in 10% the oxydol and soaked 30 minutes.Clean with distilled water flushing again.Transfer pipet with dry is measured the 0.2ml propyl methacrylate, 2ml0.2mol l
-1PH be that 3.6 acetic acid-sodium-acetate buffer and 8ml distilled water mix, stirs propyl methacrylate is melted to prepare silanization solution fully.(container that in chromic acid lotion, soaks is the beaker, and other used containers are plastic containers) slide immersed in this solution 2-3 hour, cleaned with distilled water then, and drying at room temperature is subsequent use.
2, the preparation of auroral poles film: 15mg allyl calcein is dissolved in 0.2ml N, in the dinethylformamide, fully dissolving.Add the 200mg acrylic amide successively, 0.4ml hydroxyethyl methacrylate, 0.15ml crosslinking chemical triethoxy trimethyl acrylic ester, 0.2ml triethanolamine and 0.25ml 1-hydroxy-cyclohexyl phenyl ketone.After mixing, (measure above solution and all must use dry transfer pipet) pipette a small amount of dripping on clean polyfluortetraethylene plate; Common slide with silanization is good covers on it; Use uviol lamp (254nm) irradiation 30 minutes again; Take out the good slide of photocuring successively water and alcohol flushing, till the loss that does not observe carrier.
To be pack into the top of teflon flow cell of auroral poles film that fluorescence carrier is processed with the allyl calcein; Testing sample is with the speed input flow cell of 1.5ml/min; Measure fluorescence intensity at maximum excitation wavelength and transmitted wave strong point, and measure unknown appearance pH value according to correction equation.
Fluorescence measurement carries out on the PerkingElmer LS55 phosphorescence spectrophotometer of band computer data processing system.The quartzy slide that will adhere to the auroral poles film is fixed in the teflon flow cell by nut, and face makes the auroral poles film contact with sample solution down.A both arms optical fiber one terminates on the spectrophotometer; The other end inserts flow cell and is close to the slide reverse side; The end of the radiation that excitation source sends through both arms optical fiber is transferred to and shines surface of glass slide in the flow cell; And exciting the fluorescent material in the auroral poles film, emitted fluorescence turns back to detecting device by other end transmission again, measures.Sample solution is by the speed input flow cell of peristaltic pump with 1.5ml/min, and auroral poles film and sample solution just can obtain a stable fluorescence intensity level after reaching balance.Fluorescence chemical sensor of the present invention can be applicable to the mensuration of pH value of solution value.
Embodiment 3: the mensuration of correction equation
The solution of different pH values is the B-R damping fluid, prepares with phosphoric acid, acetate, boric acid, and uses 0.2mol l
-1NaOH solution adjustment pH reach desirable value.
With the allyl calcein is that auroral poles film that fluorescence carrier is processed is packed in the teflon flow cell; B-R buffer solution (pH=6.5,6.8,7.1,7.4,7.7,8.0,8.3,8.6,8.9,9.2,9.5,9.8,10.1,10.4,10.8,11.1,11.4,11.7,12.0) with different pH values; By the speed input flow cell of peristaltic pump with 1.5ml/min; At maximum excitation wavelength 505nm, fluorescence intensity of solution (I) is measured at maximum emission wavelength 526nm place, makes fitting a straight line with Sigmaplot software; As shown in Figure 3, obtain correction equation and be: I=-42.16pH+615.93 (R=0.9920).Therefore we can measure the value of fluorescence intensity under the different pH and correspond to the true pH value that just can obtain solution on the curve.
Embodiment 4:
In order to check the application of this sensor in reality, existing pH value with three kinds of actual water sample of its mensuration:
(1) waste water appearance is taken from the catalyzer washing fluid in chemical plant, Changsha, 3 parts of parallel appearance;
(2) river appearance is taken from the preceding river of graduate school of Hunan University, 3 parts of parallel appearance;
(3) spring appearance is taken from Yue Lu mountain, Changsha white crane spring, 3 parts of parallel appearance.
Mensuration result is as shown in the table:
PH
aThe testing result of fluorescent optical sensor of the present invention
PH
bThe pH meter testing result, the accurate pH meter (Shanghai Precision Scientific Apparatus Co., Ltd) of PHS-3B
Sensor determination result and glass electrode measurement result basically identical shown in as above showing, this sensor can be used for the measurement of these three kinds of actual water sample.
Claims (2)
1. pH fluorescent optical sensor based on the allyl calcein comprises the slide of silanization, and the membrane matrix monomer is attached to the auroral poles film on the slide through photopolymerization, it is characterized in that the fluorescence carrier in the membrane matrix monomer is the allyl calcein.
2. the preparation method based on the pH fluorescent optical sensor of allyl calcein comprises the silanization of slide, the preparation of auroral poles film, it is characterized in that the preparation method of said auroral poles film is following:
1), 15 mg allyl calceins are dissolved in 0.2 ml N, in the dinethylformamide, fully dissolving;
2), in the solution of step 1), add 200 mg acrylic amides successively; 0.4 the ml hydroxyethyl methacrylate, 0.15 ml crosslinking chemical triethoxy trimethyl acrylic ester, 0.2 ml triethanolamine; Add 0.25 ml 1-hydroxy-cyclohexyl phenyl ketone at last, mix;
3), removing step 2) in the drips of solution that obtains on polyfluortetraethylene plate, with the slide lid of silanization on it, irradiation is 30 minutes under uviol lamp, priority water and alcohol flushing then, drying at room temperature makes the slide that contains auroral poles film thin layer.
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Citations (5)
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|---|---|---|---|---|
| US4965087A (en) * | 1982-12-07 | 1990-10-23 | Avl Ag | Method of making a sensor element for fluorescence-optical measurements |
| CN1959381A (en) * | 2006-11-13 | 2007-05-09 | 湖南大学 | Fluorescence PII sensor with inner reference and comparison, and preparation method |
| US20090044744A1 (en) * | 2007-01-30 | 2009-02-19 | Luna Innovations Incorporated | Thermal Sensitive Material |
| CN101430282A (en) * | 2008-12-16 | 2009-05-13 | 焦广旭 | Fluorescent chemical sensor for furadantin detection |
| CN101881732A (en) * | 2010-06-02 | 2010-11-10 | 复旦大学附属中山医院 | Fluorescent optical fiber sensor capable of monitoring pH value of blood in real time and preparation method thereof |
-
2012
- 2012-07-20 CN CN2012102531896A patent/CN102830099A/en active Pending
Patent Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US4965087A (en) * | 1982-12-07 | 1990-10-23 | Avl Ag | Method of making a sensor element for fluorescence-optical measurements |
| CN1959381A (en) * | 2006-11-13 | 2007-05-09 | 湖南大学 | Fluorescence PII sensor with inner reference and comparison, and preparation method |
| US20090044744A1 (en) * | 2007-01-30 | 2009-02-19 | Luna Innovations Incorporated | Thermal Sensitive Material |
| CN101430282A (en) * | 2008-12-16 | 2009-05-13 | 焦广旭 | Fluorescent chemical sensor for furadantin detection |
| CN101881732A (en) * | 2010-06-02 | 2010-11-10 | 复旦大学附属中山医院 | Fluorescent optical fiber sensor capable of monitoring pH value of blood in real time and preparation method thereof |
Non-Patent Citations (2)
| Title |
|---|
| W. D. SLOAN等: "A fibre-optic calcium ion sensor using a calcein derivative", 《LUMINESCENCE》 * |
| 桂小琴: "新型荧光试剂的合成及荧光pH传感器的研制", 《中国优秀硕士学位论文全文数据库信息科技辑》 * |
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Application publication date: 20121219 |