CN112730358A - Optical sensing film for monitoring pH two-dimensional dynamic distribution in sediment - Google Patents
Optical sensing film for monitoring pH two-dimensional dynamic distribution in sediment Download PDFInfo
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Abstract
An optical sensing film for monitoring two-dimensional dynamic distribution of pH in sediments is prepared by the following steps: firstly, dissolving 5-fluoroescein in ethanol to prepare a pH sensitive dye stock solution, dissolving DPA in ethanol to obtain a reference dye stock solution, and mixing the two stock solutions in equal volume to obtain a pH fluorescent dye stock solution; adding HydroMed D4 into a mixed solution of pH fluorescent dye stock solution and water, stirring and dissolving to prepare pH sensitive layer casting solution, and then adding HydroMed D4 into a mixed solution of ethanol and water, stirring and dissolving to prepare protective film casting solution; finally, uniformly coating the pH sensitive layer casting solution on the surface of a transparent polyester film, and after the pH sensitive layer is completely dried, uniformly coating a protective film stock solution on the surface of the sensitive layer to obtain an optical sensing film; the preparation method of the optical sensing film is simple, convenient and efficient, does not need a complex chemical synthesis method and experimental steps, and can quickly obtain the two-dimensional spatial distribution of pH in sediments in a short time.
Description
[ technical field ] A method for producing a semiconductor device
The invention belongs to the technical field of monitoring of pH in sediments, and particularly relates to an optical sensing film for monitoring two-dimensional dynamic distribution of pH in sediments.
[ background of the invention ]
Sediments act as "sinks" and "sources" of pollutants, playing a vital role in the circulation of biogeochemical elements; deposits also have a high degree of heterogeneity and the resolution of monitoring techniques determines the degree of heterogeneity in the deposits monitored, and therefore high resolution deposit monitoring techniques have received much attention in recent years.
The pH is one of important parameters for representing the biogeochemical characteristics of the water environment, and the change of the pH in the sediment reflects various biogeochemical reactions and transportation processes, such as degradation of organic matters in the sediment, oxidation of ammonia and the like; therefore, monitoring of pH in the sediment is critical; microelectrode technology and optical fiber sensor are used as two important means for monitoring pH, and the two technologies play more and more important roles in sediment pH monitoring due to high resolution and fast reaction time; however, microelectrode and fiber sensor technologies can only achieve single-point measurement, and are expensive, cumbersome to operate, and unable to obtain two-dimensional spatial distribution of pH in a deposit.
The development of planar optical sensors provides a new approach for monitoring the two-dimensional spatial distribution of pH in sediments; the planar optical sensor is mainly characterized in that a photosensitive fluorescent dye is uniformly fixed on the surface of a planar substrate by a physical or chemical method to form a planar optical sensing film, after the sensing film is contacted with the pH in a deposit, the optical properties (luminous intensity and luminous life) of the fluorescent dye are changed, then a camera or other photosensitive elements are used for capturing a fluorescent image of the sensing film under the irradiation of an excitation light source, and the optical information of each pixel point on the image is converted into the pH according to a calibration equation, so that the two-dimensional spatial distribution information of the pH in the deposit is obtained; currently, the commonly used optical sensing film using HPTS as a main raw material has a complicated preparation process: one is that the lipid HPTS is prepared by chemically modifying water-soluble HPTS dye, and then the dye is fixed in polymer material through physical embedding place to prepare optical sensing film; another method is to prevent leakage of the dye by directly immobilizing HPTS on a polymer having a functional group by a covalent bonding method, both of which are too complicated.
Therefore, the problem to be solved in the field is to provide an optical sensing film which is simple, convenient and efficient and can ensure the performance of the sensing film.
[ summary of the invention ]
In order to solve the problems, the invention provides an optical sensing film for monitoring pH two-dimensional dynamic distribution in sediments, and a preparation method of the optical sensing film comprises the following steps:
the method comprises the following steps: dissolving 5-fluoroescein in ethanol to prepare a pH sensitive dye stock solution, dissolving DPA in ethanol to obtain a reference dye stock solution, and mixing the two stock solutions in equal volume to obtain a pH fluorescent dye stock solution;
step two: adding HydroMed D4 into a mixed solution of pH fluorescent dye stock solution and water, stirring and dissolving to prepare pH sensitive layer casting solution, and then adding HydroMed D4 into a mixed solution of ethanol and water, stirring and dissolving to prepare protective film casting solution;
step three: uniformly coating the pH sensitive layer casting solution on the surface of a transparent polyester film, and drying for 12 h; and after the pH sensitive layer is completely dried, uniformly coating the protective film stock solution on the surface of the sensitive layer, and drying for 12 hours to obtain the optical sensing film.
Further, the concentration of the pH sensitive dye stock solution is 2-4 mg/ml, and the concentration of the reference dye stock solution is 1-3 mg/ml.
Further, the concentration of D4 in the pH sensitive layer casting solution and the protective film stock solution is 0.05-0.1 g/ml.
Further, the calibration method comprises the following steps:
s1: preparing phosphate buffer solutions with different pH values by using disodium hydrogen phosphate and sodium citrate, attaching the optical sensing film on the inner wall of a cuvette, respectively adding the phosphate buffer solutions with different pH values, and obtaining fluorescent images of the optical sensing film under different pH conditions by using an imaging system;
and S2, analyzing the fluorescence image obtained in S1, extracting RGB three channels of the image by using Matlab software to obtain two fluorescence intensity images of G and B channels corresponding to the two dyes 5-fluoroscein and DPA respectively, calculating the fluorescence intensity ratio r of the two channels, performing curve fitting on the fluorescence intensity ratio r and the corresponding pH value to obtain a pH response curve of the optical sensing membrane, and performing fitting by using a Boltzmann equation.
Compared with the prior art, the invention has the following beneficial effects:
1. the preparation method of the optical sensing film is simple, convenient and efficient, and does not need a complex chemical synthesis method and experiment steps;
2. the optical sensing film can reduce sediment background interference by using a fluorescence ratio imaging method;
3. the optical sensing film has a monitoring range of 6.0-8.0 and is suitable for monitoring the pH in a common fresh water environment.
4. The optical sensing film has high space-time resolution, the space resolution is about 80 microns, the time resolution is about 6 seconds, and the two-dimensional space distribution of pH in sediments can be rapidly acquired in a short time.
[ description of the drawings ]
FIG. 1 shows fluorescence spectra of the present invention in buffer solutions of different pH values.
FIG. 2 is a graph showing the response of the fluorescence ratio of the optical sensing film of the present invention in buffer solutions with different pH values.
Fig. 3 is a graph showing the uniformity of the optical sensing film in a buffer solution with pH 7 according to an embodiment of the present invention.
FIG. 4 is a graph showing response times of optical sensing films according to embodiments of the present invention.
FIG. 5 is a graph illustrating the response of an optical sensing film to ionic strength in an embodiment of the present invention.
FIG. 6 is a schematic structural diagram of an optical sensing film for detecting pH in sediment according to an embodiment of the present invention.
FIG. 7 is a two-dimensional spatial distribution of pH at the sediment-water interface obtained by the optical sensing film of the embodiment of the present invention.
FIG. 8 is a two-dimensional spatial distribution diagram of the rhizosphere pH of the tape grass obtained by the optical sensing film according to the embodiment of the invention.
[ detailed description ] embodiments
The directional terms of the present invention, such as "up", "down", "front", "back", "left", "right", "inner", "outer", "side", etc., are only directions in the drawings, and are only used to explain and illustrate the present invention, but not to limit the scope of the present invention.
The present invention is further illustrated by the following figures and specific examples, which are to be understood as illustrative only and not as limiting the scope of the invention, which is to be given the full breadth of the appended claims and any and all equivalent modifications thereof which may occur to those skilled in the art upon reading the present specification.
Example 1: the preparation of the optical sensing film comprises the following steps
The method comprises the following steps: firstly, dissolving 5-fluoroescein in ethanol to prepare a pH sensitive dye stock solution; then dissolving DPA in ethanol to obtain a reference dye stock solution; mixing the two stock solutions in equal volume to obtain a pH fluorescent dye stock solution; the concentration of the pH sensitive dye stock solution is 2-4 mg/ml, and the concentration of the reference dye stock solution is 1-3 mg/ml.
Step two: adding HydroMed D4 into a mixed solution of pH fluorescent dye stock solution and water with the volume ratio of 9:1, stirring and dissolving to prepare pH sensitive layer casting solution; adding HydroMed D4 into a mixed solution of ethanol and water with the volume ratio of 9:1, stirring and dissolving to prepare a protective film casting solution; the concentration of D4 in the pH sensitive layer casting solution and the protective film stock solution is 0.05-0.1 g/ml.
Step three: uniformly coating the pH sensitive layer casting solution on the surface of a transparent polyester film, and drying for 12h to form a pH sensitive layer; and after the pH sensitive layer is completely dried, uniformly coating the protective film stock solution on the surface of the sensitive layer, and drying for 12 hours to obtain the optical sensing film.
Step four: placing the optical sensing film in a quartz cup, and measuring fluorescence spectra (figure 1) of the optical sensing film in buffer solutions with different pH values by a photometer, wherein 5-fluoroscein emits at 525nm, and the fluorescence intensity is increased along with the increase of the pH value, so that the optical sensing film can be used as a pH sensitive fluorescent dye; DPA emits at 440nm, and the fluorescence intensity does not change obviously with the increase of pH, which indicates that DPA can be used as a reference dye.
Example 2: drawing a standard curve
S1, preparing phosphate buffer solutions with different pH values by using 0.2M disodium hydrogen phosphate and 0.1M sodium citrate, attaching the optical sensing film on the inner wall of a cuvette, respectively adding the phosphate buffer solutions with different pH values, and obtaining fluorescence images of the optical sensing film under different pH conditions by using the imaging system shown in figure 6.
S2, analyzing the fluorescence image obtained in S1, extracting RGB three channels of the image by using Matlab software to obtain two fluorescence intensity images of G and B channels corresponding to two dyes 5-fluoroscein and DPA respectively, calculating the fluorescence intensity ratio r of the two channels, performing curve fitting on the fluorescence intensity ratio r and the corresponding pH value to obtain a pH response curve of the optical sensing membrane, and performing fitting by using a Boltzmann equation:
wherein r is the intensity ratio (G/B) of the green channel and the blue channel of the optical sensing film under different pH values; a, b and dx represent empirical parameters and the width of the curve, respectively, and pKa is the coefficient at the center of the measurement range.
The response curve of the optical sensing film to pH is shown in FIG. 2, and a calibration curve is finally obtained:
the monitoring range of the optical sensing film is 6.0-8.0.
Example 3: testing of optical sensing films for uniformity, response time, and interference of ionic strength
(1) Uniformity test
The optical sensing film is attached to the inner wall of a cuvette, phosphoric acid buffer solutions with different pH values of 7 are added, after stabilization, an imaging system shown in figure 6 is used for obtaining a fluorescence image of the optical sensing film, Matlab software is used for extracting G and B channels of the image and taking the channel as a ratio, and the RSD (RSD) of the optical sensing film within the range of 5cm x 4cm is obtained to be less than 5% (figure 3), and the optical sensing film has good uniformity.
(2) Response time testing
The optical sensing film is attached to the inner wall of the cuvette, a buffer solution with pH 6 is added, a picture is taken every 1 second, when the time reaches 20 seconds, the optical sensing film is immediately taken out and put into the buffer solution with pH 8, and a picture is taken every 1 second. Matlab software was used to perform the ratio of the G and B channels of the extracted image, and the ratio was used as the ordinate and the photographing time was used as the abscissa to obtain a response time graph (FIG. 4) of the optical sensor film, and it was found that the response time was <6s and the response time was fast.
(3) Effect of ionic strength on optical sensing Membrane response
The ionic strength of the pH buffer solution is adjusted by using sodium chloride to obtain the pH buffer solution with different ionic strengths, then the optical sensing membrane is pasted on the inner wall of a cuvette and is respectively added with the pH buffer solution with different ionic strengths, after stabilization, an imaging system shown in figure 6 is used for obtaining a fluorescence image of the optical sensing membrane, Matlab software is used for carrying out G and B channels for extracting the image and carrying out ratio, the ratio is used as a vertical coordinate, the pH value is used as a horizontal coordinate, the response of the optical sensing membrane under different ionic strengths is obtained (figure 5), and the result shows that the ionic strength has certain interference on the optical sensing membrane, so that the optical sensing membrane is corrected by using synthetic seawater particularly in marine sediments.
Example 4: application of optical sensing film in sediment
Collecting a sediment columnar sample and a water sample of the Taihu lake, cutting the sediment columnar sample layer by layer, mixing uniformly, sieving, removing benthos, large particles and the like, adding the layered sediment sample into a root box with a detachable side, adding lake water for culturing, after the sediment is stabilized for 2 weeks, taking out the upper cover water, detaching the detachable side of the root box, attaching the prepared optical sensing film on a detachable organic glass plate, then re-installing the organic glass plate, after stabilizing for a period of time, moving the root box into a dark room to obtain a fluorescence image, and taking a picture by using an imaging system shown in figure 6; after the fluorescence image of the sediment-water interface pH is obtained, inserting a submerged plant tape grass near the optical sensing film, culturing the submerged plant tape grass with a root box inclined at 45 degrees so that the plant can grow on the optical sensing film, and taking a picture after culturing for one week.
According to the method, the two-dimensional spatial distribution of pH in the sediment can be obtained after calculation and mapping. The results show that the optical sensing film can acquire the gradient distribution of pH at the sediment-water interface (figure 7) and the influence of the root system of the aquatic plant on the pH in the sediment (figure 8); the pH is distributed in a gradient way at a sediment-water interface, and the pH of the root system is reduced due to the acidification phenomenon of the plant root system; the result shows that the optical sensing film can meet the dynamic detection of pH in the sediment micro-interface environment.
The above-mentioned embodiments are intended to illustrate the objects, technical solutions and advantages of the present invention in further detail, and it should be understood that the above-mentioned embodiments are only exemplary embodiments of the present invention, and are not intended to limit the present invention, and any modifications, equivalents, improvements and the like made within the spirit and principle of the present invention should be included in the protection scope of the present invention.
Furthermore, it should be understood that although the present description refers to embodiments, not every embodiment may contain only a single embodiment, and such description is for clarity only, and those skilled in the art should integrate the description, and the embodiments may be combined as appropriate to form other embodiments understood by those skilled in the art.
Claims (4)
1. An optical sensing film for monitoring two-dimensional dynamic distribution of pH in sediment, which is characterized in that the preparation method of the optical sensing film comprises the following steps:
the method comprises the following steps: dissolving 5-fluoroescein in ethanol to prepare a pH sensitive dye stock solution, dissolving DPA in ethanol to obtain a reference dye stock solution, and mixing the two stock solutions in equal volume to obtain a pH fluorescent dye stock solution;
step two: adding HydroMed D4 into a mixed solution of pH fluorescent dye stock solution and water, stirring and dissolving to prepare pH sensitive layer casting solution, and then adding HydroMed D4 into a mixed solution of ethanol and water, stirring and dissolving to prepare protective film casting solution;
step three: uniformly coating the pH sensitive layer casting solution on the surface of a transparent polyester film, and drying for 12 h; and after the pH sensitive layer is completely dried, uniformly coating the protective film stock solution on the surface of the sensitive layer, and drying to obtain the optical sensing film.
2. The optical sensing film for monitoring pH two-dimensional dynamic distribution in sediments according to claim 1, wherein the concentration of the pH sensitive dye stock solution is 2-4 mg/ml, and the concentration of the reference dye stock solution is 1-3 mg/ml.
3. The optical sensing film for monitoring the two-dimensional dynamic distribution of the pH in the sediment as claimed in claim 1, wherein the concentration of D4 in the pH sensitive layer casting solution and the protective film stock solution is 0.05-0.1 g/ml.
4. A calibration method applied to the optical sensing film of claim 1, wherein the calibration method comprises the steps of:
s1: preparing phosphate buffer solutions with different pH values by using disodium hydrogen phosphate and sodium citrate, attaching the optical sensing film on the inner wall of a cuvette, respectively adding the phosphate buffer solutions with different pH values, and obtaining fluorescent images of the optical sensing film under different pH conditions by using an imaging system;
and S2, analyzing the fluorescence image obtained in S1, extracting RGB three channels of the image by using Matlab software to obtain two fluorescence intensity images of G and B channels corresponding to the two dyes 5-fluoroscein and DPA respectively, calculating the fluorescence intensity ratio r of the two channels, performing curve fitting on the fluorescence intensity ratio r and the corresponding pH value to obtain a pH response curve of the optical sensing membrane, and performing fitting by using a Boltzmann equation.
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Cited By (2)
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| CN113624728A (en) * | 2021-07-02 | 2021-11-09 | 湖北文理学院 | Ratiometric fluorescent probe system for detecting hydrazine and method for detecting hydrazine concentration |
| CN115353659A (en) * | 2022-08-08 | 2022-11-18 | 中国科学院南京地理与湖泊研究所 | Preparation and application of gel film for in-situ and high-resolution detection of pH in sediment |
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