CN113403060A

CN113403060A - Ratiometric fluorescent filter membrane based on fluorescein and carbon quantum dots, preparation method and application

Info

Publication number: CN113403060A
Application number: CN202110599790.XA
Authority: CN
Inventors: 王丽; 刘英男; 肖亚庆; 尚明慧
Original assignee: Northwest A&F University
Current assignee: Northwest A&F University
Priority date: 2021-05-31
Filing date: 2021-05-31
Publication date: 2021-09-17

Abstract

The invention discloses a ratio fluorescence filter membrane based on fluorescein and carbon quantum dots, a preparation method and application thereof. The invention takes the ratio fluorescence filter membrane based on the fluorescein and the carbon quantum dot as the intelligent label for detection, the detection result is basically consistent with the result of the standard volatile basic nitrogen detection method, the color change is easy to be identified by naked eyes, and the detection process is economic, efficient, rapid and convenient.

Description

Ratiometric fluorescent filter membrane based on fluorescein and carbon quantum dots, preparation method and application

Technical Field

The invention relates to the field of food detection, in particular to a ratio fluorescence filter membrane based on fluorescein and carbon quantum dots, a preparation method and application.

Background

Seafood has been popular with consumers because of its abundant protein content. With the continuous increase in seafood demand in the global market, more and more fresh seafood is being transported around the world. However, improper transport and storage conditions can result in microorganisms enzymatically producing biogenic amines, and these accumulated biogenic amines can serve as a direct indicator of food spoilage. Therefore, detecting the biogenic amine content in food products is crucial for monitoring seafood freshness and assessing seafood quality.

Currently, a variety of analytical methods for detecting biogenic amines have been developed, including gas chromatography, liquid chromatography, electronic nose, electronic tongue, capillary electrophoresis, and the like. However, most of these methods rely on expensive and complex instrumentation, cumbersome sample preparation, and trained technicians. Therefore, they are not suitable for real-time, in situ detection of biogenic amines, nor are they amenable to detection by the general population. It is imperative to develop a simple, fast and cost effective method to monitor seafood freshness in real time in situ. Although the visual colorimetric detection method can also sensitively detect the biogenic amine, the detection result is easily interfered by environmental factors and the color of the sample, and errors are easily caused.

Disclosure of Invention

Aiming at the defects and shortcomings in the prior art, the invention provides a ratio fluorescence filter membrane based on fluorescein and carbon quantum dots, a preparation method and application. The ratio fluorescence filter membrane as a label has the advantages of low preparation cost, simple preparation process and easy naked eye identification of color change, and solves the problems that the prior biogenic amine detection depends on large-scale instruments and equipment, the pretreatment process is complicated, the reaction time is long and the like.

In order to achieve the purpose, the technical scheme adopted by the invention comprises the following steps:

a ratio fluorescence filter membrane based on fluorescein and carbon quantum dots comprises an organic phase filter membrane as a base material, the fluorescein as an indicator and the carbon quantum dots as an internal reference, wherein the fluorescein and the carbon quantum dots are deposited on the organic phase filter membrane to form the ratio fluorescence filter membrane, the fluorescence indicator is green fluorescence fluorescein, the carbon quantum dots emit red fluorescence in an aggregated state, and the carbon quantum dots are prepared from melamine and dithiosalicylic acid through a solvothermal method.

Specifically, the preparation method of the ratiometric fluorescent filter membrane comprises the steps of mixing an ethanol solution of the carbon quantum dots with an ethanol solution of the fluorescein, immersing the organic phase filter membrane in the mixed solution, taking out and drying to obtain the ratiometric fluorescent filter membrane.

Specifically, the preparation method of the carbon quantum dot comprises the following steps:

the preparation method comprises the steps of dispersing melamine and dithiosalicylic acid in an acetic acid solution through ultrasound, transferring the obtained solution to a polytetrafluoroethylene reaction kettle, heating, cooling, washing, vacuum filtering and purifying sequentially to obtain the melamine dithiosalicylic acid.

The concentration ratio of the ethanol solution of the carbon quantum dots to the ethanol solution of the fluorescein is 4:0.5, and the volume ratio of the ethanol solution of the carbon quantum dots to the ethanol solution of the fluorescein is 4 (0.1-1.5).

Specifically, the mass fraction of the melamine is 0.4-0.6%, the mass fraction of the dithiosalicylic acid is 1-1.5%, the reaction temperature is 175-185 ℃, and the reaction time is 8-12 h.

A preparation method of a ratio fluorescence filter membrane based on fluorescein and carbon quantum dots comprises the steps of mixing an ethanol solution of the carbon quantum dots with an ethanol solution of the fluorescein, immersing an organic phase filter membrane in the mixed solution, taking out and drying to obtain the ratio fluorescence filter membrane.

Furthermore, the concentration ratio of the ethanol solution of the carbon quantum dots to the ethanol solution of the fluorescein is 4:0.5, and the volume ratio is 4 (0.1-1.5).

Specifically, the volume ratio of the ethanol solution of the carbon quantum dots to the ethanol solution of the fluorescein is 4: 1.

The invention relates to application of a ratio fluorescence filter membrane based on fluorescein and carbon quantum dots in detecting food freshness.

The ratiometric fluorescent filter membrane prepared by the preparation method based on the fluorescein and carbon quantum dots is applied to detecting food freshness.

Compared with the prior art, the invention has the following technical effects:

(1) the invention takes the ratio fluorescence filter membrane based on the fluorescein and the carbon quantum dot as the intelligent label for detection, the detection result is basically consistent with the result of the standard volatile basic nitrogen detection method, the color change is easy to be identified by naked eyes, and the detection process is economic, efficient, rapid and convenient.

(2) The detection effects under the two ultraviolet lights can be verified mutually, so that the detection accuracy is further improved.

(3) The R-F filters (RQDs and FL co-modified filters) can be used as smart labels to simply and accurately monitor shrimp and crab freshness. The smart tag can be easily applied to the seafood supply chain to monitor freshness in real time and visually. It is also expected that a smartphone will be used to capture the tag and assess freshness through more accurate color analysis.

Drawings

FIG. 1 is a photograph of (a) blank filters, RQDs modified filters, FL modified filters and R-F modified filters under ambient light, 254nm UV light and 365nm UV light.

FIG. 2 is a photograph of RQDs, FL and R-F filters exposed to 25mg/mL ammonia gas for 7min at 254nm and 365nm violet light.

FIG. 3 shows the fluorescence response of filters modified with mixed solutions of RQDs and FL at different ratios to ammonia. (ammonia water concentration of 2.5 mg/mL. RQDs solution concentration of 4mg/mL, FL solution concentration of 0.5 mg/mL. I: 4mL RQDs solution was mixed with 0.1mL FL solution, II: 4mL RQDs solution was mixed with 0.2mL FL solution, III: 4mL RQDs solution was mixed with 0.6mL FL solution, IV: 4mL RQDs solution was mixed with 1mL FL solution, V: 4mL RQDs solution was mixed with 1.5mL FL solution)

FIG. 4 is a photograph of R-F filters exposed to 2.5mg/mL ammonia for various periods of time under UV light at 254nm and 365 nm.

FIG. 5 is a graph showing the fluorescence response of R-F filters to different concentrations of ammonia at two excitation wavelengths.

FIG. 6 is a photograph of the R-F filter in a repeat use under UV light at 254nm and 365nm for 2.5mg/mL ammonia detection.

FIG. 7 is an R-F filter under 254nm and 365nm UV light to monitor shrimp freshness under different storage conditions.

FIG. 8 is a photograph showing the R-F filter membrane stored in a container containing wet wipes at different temperatures for 1 to 5 days.

FIG. 9 is a graph showing the TVB-N content of shrimps stored at 25 deg.C, 4 deg.C and-20 deg.C for different days.

The following describes embodiments of the present invention in further detail with reference to the accompanying drawings.

Detailed Description

The present invention uses green fluorescent fluorescein as the fluorescence indicator, abbreviated in english as FL, and red fluorescent carbon quantum dots with aggregation-induced emission properties as the internal reference, abbreviated in english as RQDs. The two are modified on the surface of the organic phase filter membrane by a physical deposition method. When the biogenic amine molecule exists, FL molecule conformation is changed, green fluorescence is enhanced, while RQDs do not respond to biogenic amine, and fluorescence is not interfered. The change of the intensity of the two fluorescence signals is closely related to the concentration of the biogenic amine and can be recognized by the naked eye, so that the ratiometric fluorescent film can be used as a smart label for visualization and real-time monitoring of the biogenic amine.

The solvothermal method is developed on the basis of a hydrothermal method, and refers to a synthetic method in which an original mixture is reacted in a closed system such as an autoclave by using an organic or non-aqueous solvent as a solvent at a certain temperature and under the autogenous pressure of the solution.

The experimental reagents used in the present invention include dithiosalicylic acid, fluorescein, and melamine, supplied by alatin chemical co. Ammonia (25%) and acetic acid were obtained from guangdong photowarfare technologies, inc.

Organic phase filters (0.22 μm) were purchased from Shanghai New sub-clean plants.

Detection instruments and equipment are common instruments, and the detection methods for determining total volatile basic nitrogen (TVB-N) in the shrimp meat sample by using an automatic Kjeldahl azotometer are conventional methods. The total nitrogen content (TVB-N) distilled out from the water extract of meat food under alkaline condition together with water vapor is used for showing the freshness of the food.

Biogenic Amines (BAs) are a generic name for a class of biologically active nitrogen-containing low molecular weight organic compounds. It is considered that the ammonia is produced by substituting 1 to 3 hydrogen atoms in the ammonia molecule with an alkyl group or an aryl group, and is a low molecular weight organic base of aliphatic, alicyclic or heterocyclic group, and is often present in the bodies of animals and plants and in foods.

The R-F filter membrane shows sensitive fluorescent response to biogenic amine, reflects the freshness of food, and proves the application reliability of the R-F filter membrane in the aspect of biogenic amine detection through mutual verification of the fluorescent detection and a common TVB-N detection method.

RGB refers to the three primary optical colors: r is Red (Red), G is Green (Green), and B is Blue (Blue). The value of RGB refers to its luminance, the larger the value, the larger the luminance. The RGB values in the examples of the present invention are the mean and standard deviation of 3 parallel values.

Example 1:

according to the technical scheme, the ratiometric fluorescent filter membrane based on the fluorescein and the carbon quantum dots and the preparation method are provided, the ratiometric fluorescent filter membrane is formed by depositing the fluorescein and the carbon quantum dots on the organic phase filter membrane by taking the organic phase filter membrane as a base material, the fluorescein as an indicator and the carbon quantum dots as internal references, the fluorescent indicator is green fluorescent fluorescein, the carbon quantum dots emit red fluorescence in an aggregation state, and the carbon quantum dots are prepared from melamine and dithiosalicylic acid through a solvothermal method.

The preparation method of the ratio fluorescence filter membrane comprises the steps of mixing an ethanol solution of the carbon quantum dots with an ethanol solution of the fluorescein, immersing the organic phase filter membrane in the mixed solution, taking out and drying to obtain the ratio fluorescence filter membrane.

The preparation method of the carbon quantum dot comprises the following steps:

The mass fraction of the melamine is 0.4-0.6%, the mass fraction of the dithio-salicylic acid is 1-1.5%, the reaction temperature is 175-185 ℃, and the reaction time is 8-12 h.

As a preferred embodiment, the method for preparing the ratiometric fluorescent filter comprises the steps of,

first, a (red quat dots, RQDs) RQDs (4mg/mL) ethanol solution and a green fluorescent fluorescein FL (0.5mg/mL) ethanol solution were prepared, then 4mLRQDs solution was mixed with 1mL FL solution uniformly, and the organic phase filter was completely submerged in the solution. After 100s, the organic phase filter was removed and allowed to dry naturally at room temperature. Obtaining the ratio fluorescence filter membrane, called R-F filter membrane for short. The concentration ratio of the ethanol solution of the carbon quantum dots to the ethanol solution of the fluorescein is 4:0.5, and the volume ratio of the ethanol solution of the carbon quantum dots to the ethanol solution of the fluorescein is 4 (0.1-1.5).

Specifically, in this embodiment, the following may be used: (1) mixing 4mL of RQDs solution with 0.1mL of FL solution; (2) mixing 4mL of RQDs solution with 0.2mL of FL solution; (3) mixing 4mL of RQDs solution with 0.6mL of FL solution; (4) mixing 4mL of RQDs solution with 1mL of FL solution; (5)4mL of RQDs solution was mixed with 1.5mL of FL solution.

The volume ratio of the ethanol solution of the carbon quantum dots to the ethanol solution of the fluorescein is 4:1, which is the optimal ratio.

Among them, FL can cause fluorescence enhancement by conformational change in the presence of amine molecules, thereby serving as an indicator. RQDs are hydrophobic and the fluorescence intensity is not interfered by amine molecules. Therefore, RQDs serve as internal references.

In this embodiment, the preparation method of the carbon quantum dot specifically includes:

melamine (MA) and dithiosalicylic acid (DTSA) were dispersed in 60mL of an acetic acid solution at mass fractions of 0.5% and 1.36% by sonication, and the resulting solution was transferred to a 100mL polytetrafluoroethylene reaction vessel and heated at 180 ℃ for 10 hours. After the solution was cooled to room temperature, it was added to 1L of boiling water to wash out residual materials and solvents. The solid quantum dots were obtained by vacuum filtration and then dried at 60 ℃. To further purify the prepared RQDs, the RQDs were redispersed in ethanol solution and insoluble material was removed by centrifugation. The supernatant was again vacuum dried to give purified RQDs.

The prepared filter membrane is shown in FIG. 1. Under ambient light, both filters modified by RQDs and R-F (RQDs and FL) showed pink color, while the FL-modified filter was green. This series of color changes means that two fluorescent materials RQDs and FL can be deposited on the filter.

The fluorescence properties of the three fluorescent filters were investigated. It can be seen that the unmodified filter showed no fluorescence under UV light at 254nm and 365nm, while the FL-modified filter showed green fluorescence. Both RQDs and R-F filters showed bright pink fluorescence. Again, the RQDs and FL were successfully deposited on the filters.

Table 1 shows the RGB values for different filters under Daylight sunlight, 254nm and 365nm UV light conditions. Wherein blank represents the blank membrane, RQDs film represents the RQDs modified filter membrane, FL film represents the FL modified filter membrane, and R-F film represents the RQDs and FL modified filter membrane.

It can be seen that under all three light conditions, the modified filters exhibited RGB values different from those of the blank membranes, confirming the successful modification of RQDs and FL on the filters.

TABLE 1 RGB values of RQDs and FL modified filters

Example 2:

this example uses ammonia as the model biogenic amine to study the ability of the prepared ratiometric fluorescence filters to detect biogenic amine vapors. Namely, discloses the application of a ratio fluorescence filter membrane based on fluorescein and carbon quantum dots to the detection of biogenic amine.

First, a series of ammonia solutions of known concentration were prepared by diluting commercially available aqueous ammonia with deionized water. Thereafter, the ammonia solution was added to a 24-well plate (1 mL per well), and the prepared filter was immediately placed on top of the 24-well plate. The color change of the fluorescence was recorded with a camera.

As shown in FIG. 2, the fluorescence color remained unchanged when RQDs filters were exposed to ammonia gas at a concentration of 25mg/mL, both under UV light at 254nm and 365 nm. This indicates that RQDs do not respond to ammonia and can be used as an internal reference. However, under the same conditions, the green fluorescence of the FL filter was significantly enhanced. This is mainly due to the fact that FL protons are deprived after exposure to ammonia gas, resulting in a change in molecular structure. FL can therefore be used as an indicator of ammonia. For the R-F filter, the fluorescence changed from pink to green under UV light. The corresponding RGB values in Table 2 are consistent with the color change described above, collectively indicating that the prepared R-F filter can be used as a dual signal ratio fluorescent label for visual detection of ammonia.

In Table 2, RQDs-before represent RQDs filters that were not exposed to ammonia, RQDs-after represent RQDs filters that were exposed to ammonia, and the responses were measured; FL-before represents the FL filter membrane not exposed to ammonia gas, FL-after represents the FL filter membrane after exposure to ammonia gas environment and detection response; R-F-before represents the RQDs and FL modified filters that were not exposed to ammonia, and R-F-after represents the RQDs and FL modified filters that were exposed to ammonia and tested after response.

TABLE 2 RGB values of the response of different filters to ammonia

The ratio of the two fluorescent materials, as well as the exposure time in an ammonia environment, is optimized for optimal detection performance. The method comprises the following specific steps: the concentration of the RQDs solution is 4mg/mL, and the concentration of the FL solution is 0.5 mg/mL.

Mixing 4mL of RQDs solution with 0.1mL of FL solution;

(II) mixing 4mL of RQD solution with 0.2mL of FL solution;

(III) mixing 4mL of RQD solution with 0.6mL of FL solution;

(IV) mixing 4mL of RQD solution with 1mL of FL solution;

(V) mixing 4mL of RQD solution with 1.5mL of FL solution;

as shown in FIG. 3, the FL addition tested had little effect on the fluorescence color of the R-F filter prior to reaction with ammonia, whether under UV light at 254nm or 365 nm. The fluorescence of the R-F filters prepared with low concentrations of FL did not change significantly when exposed to an ammonia environment and remained pink. The R-F filters gradually fluoresce green with increasing FL concentration.

A mixture (group IV) of 4mL of RQDs solution (4mg/mL) and 1mL of FL solution (0.5mg/mL) was selected for filter preparation, taking into account the fluorescence color change before and after reaction of the R-F filter under two different UV lights and the RGB value change in Table 3. In Table 3, the R-F filters were prepared from two fluorescent materials at the five ratios I-V, and the RGB values before and after reaction with ammonia were recorded, and control indicated blank control.

TABLE 3 RGB values of R-F filters prepared according to the proportions of two different fluorescent materials

Reaction time is also an important factor affecting assay performance. We recorded the change in fluorescence of the R-F filter to ammonia over 20 min.

As shown in FIG. 4, the fluorescence of the R-F filter did not change significantly in the UV light at 254nm and 365nm within 1 to 20min, indicating that the response of the prepared fluorescent membrane to ammonia reached equilibrium within 1 min. Therefore, after comprehensively considering the RGB value changes in table 4, 1min is set as the optimal response time.

TABLE 4 RGB values of R-F filters at different response times

Under the optimal experimental conditions discussed above, the fluorescent response of the R-F ratio phosphor film to different concentrations of ammonia at two excitation wavelengths was explored.

As shown in FIG. 5, under a 254nm UV lamp, FL green fluorescence becomes dominant with increasing ammonia concentration. The observer clearly identified a distinct fluorescent color change from pink to green without any training, indicating that ratiometric fluorescent films have the potential to visually detect ammonia. The trend of the change of the fluorescence color of the R-F ratio fluorescent film under 365nm ultraviolet light is basically the same as that under 254 nm. And the RGB values in table 5 correspond to the above-described fluorescent color change. Table 5 shows R-F filters for different NH groups₃RGB values before and after fluorescence response at concentrations (0 to 25000 ppm).

Together, the above results indicate that the R-F ratiometric fluorescent films have a sensitive and rapid color response to ammonia and that the color change can be observed by the naked eye without the aid of sophisticated instruments. The prepared fluorescent film has great potential and can be developed as a fluorescent label for monitoring BAs.

TABLE 5 RGB values of fluorescence response of R-F filters to different concentrations of ammonia

In this example, the reaction of the R-F filter with ammonia is reversible. The R-F ratio phosphor film is removed from NH₃Taking out in the atmosphereAfter the film was left in the air for 1min, the fluorescence of FL was quenched, and the color of the film was restored to the original state. Therefore, the fluorescent film can be reused.

Specific test results as shown in fig. 6 and table 6, the R-F ratio fluorescent film showed no significant fluorescence intensity decay after repeating five cycles (1, 2, 3, 4, 5 for 5 cycles) for 2.5mg/mL ammonia gas detection, demonstrating its excellent stability, ammonia reactivity and reversibility. Therefore, the R-F ratio phosphor film is suitable for practical use in sensing systems. This ratiometric fluorescence strategy can avoid the effects of environmental factors and thus achieve more reliable results.

TABLE 6R-F ratio RGB values after different cycles of fluorescent film inspection

Example 3:

this example shows the effect of fluorescence detection of biogenic amines produced by R-F filters during shrimp storage. The detection of the biogenic amine is carried out by taking commercial fresh shrimps as real samples. The R-F filter membrane is prepared by the preparation method of the embodiment 1, and the two fluorescent materials adopt the optimal proportion.

First, the shrimp were placed in a sterile petri dish, then the R-F filter was placed in the petri dish, taking care to avoid direct contact of the R-F filter with the shrimp, and then all dishes were sealed and placed at different temperatures (-20 ℃, 4 ℃ and 25 ℃) for 1-5 days. Meanwhile, in order to eliminate the interference of the environmental humidity on the fluorescence of the membrane, the R-F filter membrane is placed in a container containing wet tissues and is placed at different temperatures (-20 ℃, 4 ℃ and 25 ℃) for 1-5 days as a control test group. During this time, the fluorescence change of the R-F filter under UV light was recorded by a camera to analyze the freshness of the sample. In addition, total volatile basic nitrogen (TVB-N) in the shrimp meat samples was determined using an automatic kjeldahl azotometer, all measurements were repeated three times, and the results are shown in table 7.

In Table 7, 25 ℃ to 0day means that the shrimps were stored at 25 ℃ for 0day, i.e., the sample in the initial state, 25 ℃ to 1day means that the shrimps were stored at 25 ℃ for 1day, and the rest is analogized.

As seen in FIG. 7, under UV light at an excitation wavelength of 254nm, the pink color gradually faded to off-white, indicating a process of slow deterioration to severe deterioration of the shrimp from fresh. It can be seen that the original pink fluorescence of the R-F filters became pale grey after 1day of storage at 25 ℃ and 3 days at 4 ℃ indicating deterioration of the shrimp. However, when the shrimp were stored at-20 ℃ for 5 days, there was little significant change in the fluorescence color of the R-F filter, indicating that the shrimp were still fresh.

Under 365nm ultraviolet light of excitation wavelength, pink represents freshness, yellow-green represents deterioration, and green represents severe deterioration. It can be seen that the R-F filters still showed a sensitive fluorescent response to biogenic amines, and that the original pink fluorescence of the R-F filters turned yellow after 1day of storage at 25 ℃ and 3 days of storage at 4 ℃ indicating deterioration of the shrimp. Under the ultraviolet rays with the excitation wavelengths of 254nm and 365nm, the process of fluorescence change of the R-F filter membrane can embody the process from freshness to deterioration of the shrimps, and the result is reliable.

In Table 8, 25 ℃ to 0day indicates that the R-F filter was stored at 25 ℃ for 0day, i.e., the R-F filter in the initial state, 25 ℃ to 1day indicates that the R-F filter was stored at 25 ℃ for 1day, and so on. The results in FIG. 8 show that the fluorescence intensity of the R-F filters remained almost constant for 5 days under 254nm or 365nm UV light.

Referring to FIG. 7, Table 7, FIG. 8 and Table 8, the control test group in which the R-F filter was placed in the container with the wet towel demonstrated that the fluorescence of the R-F filter was hardly affected by the ambient humidity, thereby demonstrating that the change in fluorescence in FIG. 7 and Table 7 is mainly attributed to the fluorescence response of the R-F filter to biogenic amines, and thus, the R-F filter can be used to detect the freshness of shrimp.

TABLE 7 RGB values of R-F filters for detection of biogenic amines

TABLE 8 RGB values of fluorescence intensity of R-F filters on different days

The detection result of the R-F ratio fluorescent film is compared with the detection result of the standard TVB-N method to verify the detection accuracy. According to the standard, when the TVB-N content is less than 120mg/kg, the raw shrimp are fresh. When the TVB-N content is in the range of 120-250 mg/kg, the shrimp are slightly decomposed but still edible. When the TVB-N content exceeds 250mg/kg, the shrimp deteriorate and are inedible.

As shown in FIG. 9, the TVB-N level of fresh shrimp was 53 mg/kg. After storage at 25 ℃ for 1day, this increased to 1040mg/kg, indicating that the shrimps were completely spoiled. Meanwhile, the fluorescence of the R-F filter membrane changes from pink to grey under 254nm ultraviolet light, and changes from pink to yellow and green under 365nm ultraviolet light.

After the shrimp was allowed to stand at 4 ℃ for 3 days, the TVB-N value was 350 mg/kg. The fluorescent film becomes light and grey-white under 254nm ultraviolet rays, and becomes yellow under 365nm ultraviolet rays, so that the deterioration of the shrimps is reflected.

After 5 days at-20 ℃ this value increased to 86mg/kg, but the fluorescence of the R-F filter membrane hardly changed, which means that the shrimp were still fresh and edible.

The above experimental results show that the fluorescence color change of the R-F filter membrane is consistent with the results of the standard TVB-N method. Thus, the R-F filter membrane can be used as a smart label to simply and accurately monitor the freshness of shrimp and crabs. With this functionality, the smart tag can be easily applied to the seafood supply chain to monitor freshness in real time and visually. In the future, it is also expected to use smartphones to capture tags and assess freshness through more accurate color analysis.

Claims

1. A ratio fluorescence filter membrane based on fluorescein and carbon quantum dots is characterized in that an organic phase filter membrane is used as a base material, the fluorescein is used as an indicator, the carbon quantum dots are used as internal references, the fluorescein and the carbon quantum dots are deposited on the organic phase filter membrane to form the ratio fluorescence filter membrane, the fluorescence indicator is green fluorescence fluorescein, the carbon quantum dots emit red fluorescence, and the carbon quantum dots are prepared from melamine and dithiosalicylic acid through a solvothermal method.

2. The ratiometric fluorescent filter membrane based on fluorescein and carbon quantum dots of claim 1, wherein the preparation method of the ratiometric fluorescent filter membrane comprises mixing an ethanol solution of the carbon quantum dots and an ethanol solution of the fluorescein, immersing the organic phase filter membrane in the mixed solution, taking out and drying.

3. The ratio fluorescence filter membrane based on fluorescein and carbon quantum dots of claim 1, wherein the preparation method of the carbon quantum dots comprises:

4. The fluorescence filter membrane based on the ratio of the fluorescein and the carbon quantum dot as claimed in claim 2, wherein the concentration ratio of the ethanol solution of the carbon quantum dot to the ethanol solution of the fluorescein is 4:0.5, and the volume ratio is 4 (0.1-1.5).

5. The fluorescence filter membrane based on the ratio of the fluorescein and the carbon quantum dots as claimed in claim 3, wherein the mass fraction of the melamine is 0.4-0.6%, the mass fraction of the dithiosalicylic acid is 1-1.5%, the reaction temperature is 175-185 ℃, and the reaction time is 8-12 h.

6. A preparation method of a ratio fluorescence filter membrane based on fluorescein and carbon quantum dots is characterized by comprising the steps of mixing an ethanol solution of the carbon quantum dots with an ethanol solution of the fluorescein, immersing an organic phase filter membrane in the mixed solution, taking out and drying to obtain the ratio fluorescence filter membrane.

7. The method for preparing a fluorescence filter membrane based on the ratio of fluorescein and carbon quantum dot as claimed in claim 6, wherein the concentration ratio of the ethanol solution of carbon quantum dot to the ethanol solution of fluorescein is 4:0.5, and the volume ratio is 4 (0.1-1.5).

8. The method of claim 7, wherein the volume ratio of the carbon quantum dot in the ethanol solution to the fluorescein in the ethanol solution is 4: 1.

9. Use of a ratio fluorescence filter based on fluorescein and carbon quantum dots as claimed in any one of claims 1 to 5 for detecting freshness of a food product.

10. Use of a ratiometric fluorescent filter membrane prepared according to the method for preparing a ratiometric fluorescent filter membrane based on fluorescein and carbon quantum dots as defined in any one of claims 6 to 8 for detecting the freshness of a food product.