A volatile amine fluorescent probe and application of a double-channel indicating card prepared by the same in fish freshness detection

Abstract

An application of a volatile amine fluorescent probe and its prepared dual-channel indicator card in the detection of fish freshness. The fluorescent probe has the following structure: The dual-channel indicator card for fish freshness, prepared with the fluorescent probe, is shaped like a fish and consists of six square colorimetric cards of equal area and a circular sensor tag. In the upper half of the colorimetric cards, from left to right, are the natural light colorimetric areas for freshness, acceptable quality, and spoilage; in the lower half, from left to right, are the ultraviolet light colorimetric areas for freshness, acceptable quality, and spoilage. The advantages are: the probe can provide both colorimetric and fluorescence responses to various volatile amines in aqueous systems, exhibiting high sensitivity; and by preparing the probe into a sensor tag, it can be used for non-destructive, rapid, and real-time detection of fish freshness, yielding accurate and reliable results.

Description

Technical Field

[0001] This invention relates to the application of a volatile amine fluorescent probe and its prepared dual-channel indicator card in the detection of fish freshness. Background Technology

[0002] As people place increasing emphasis on healthy eating, consumers are choosing safe, healthy, and nutritious foods, and fish perfectly meets this demand, leading to a significant increase in the sales volume of fish products in my country. However, the freshness of fish directly affects its quality. Spoiled fish not only causes huge economic losses but also has harmful effects on human health. Therefore, how to conveniently, quickly, and cost-effectively assess the quality and freshness of fish has become an increasingly important issue. Currently, deep-sea fish are mostly sold in sealed plastic bags or vacuum-packed containers to ensure their quality. However, traditional methods for testing fish freshness often require damaging the outer packaging, are time-consuming in pre-processing, and are complex and cumbersome. Although many analytical methods have been developed in recent years for detecting fish freshness, they usually require expensive instruments, cannot display fish freshness in real time, and cannot meet the needs of retailers or consumers for visual assessment. Developing a method for evaluating fish freshness that does not damage the outer packaging, is low-cost, easy to operate, and provides accurate and reliable results remains a key focus for researchers.

[0003] In recent years, smart packaging has shown promising application prospects in monitoring fish quality. Existing smart packaging designs include chemical or biological indicators and time-temperature indicators. Among these different methods, monitoring fish freshness by designing smart packaging to analyze volatile metabolites produced during fish spoilage has been widely reported. However, some freshness indicator packaging is affected by the poor hygroscopicity of the carrier or the limited humidity environment inside the packaging, directly impacting the sensitivity and effectiveness of the sensor tags. For example, although pH-sensitive sensor tags for monitoring fish freshness were prepared in the literature *Food Science and Biotechnology, 2017, 26(1): 37-42; Carbohydrate Polymers 255 (2021) 117488*, the color changes in colorimetric recognition were not vivid enough, easily leading to identification errors during application. Therefore, developing sensor tags with both colorimetric and fluorescence responses, and comprehensively judging the freshness level of fish through dual-channel color development, can reduce misjudgments. In addition, it is important to develop near-infrared fluorescent probes that are sensitive to volatile amines, fast in speed, and have emission wavelengths in the range of 600 nm to 800 nm. Such probes are less affected by background fluorescence interference, exhibit more significant fluorescence changes, and have greater application prospects. Summary of the Invention

[0004] The technical problem to be solved by this invention is to provide a volatile amine fluorescent probe and its prepared dual-channel indicator card for the detection of fish freshness. This probe can exhibit both colorimetric and fluorescence responses to various amine compounds in a DMF / H2O = 1:1 (v / v) system, with high sensitivity, fast response time, and near-infrared emission. By preparing the probe into a sensor tag, qualitative analysis of fish freshness within packaging boxes can be achieved without destroying the sample or requiring complex pretreatment. The results are accurate and reliable, and it can be used for non-destructive, rapid, and real-time detection of fish freshness.

[0005] An application of a volatile amine fluorescent probe and its prepared dual-channel indicator card in the detection of fish freshness. The structural formula of the fluorescent probe is as follows:

[0006] Furthermore, the specific synthesis steps of this fluorescent probe are as follows:

[0007] Using ethanol as a solvent, a benzopyran derivative and 2-thiobarbituric acid were fed at a molar ratio of 1:(1-1.5), wherein the benzopyran derivative was 6-(diethylamino)-1,2-dihydrocyclopentane[b]benzopyran-3-carboxaldehyde. Then, piperidine was added at a mass ratio of benzopyran derivative to piperidine of 100:1, and the mixture was heated under reflux with stirring for 2 to 4 hours. After cooling to room temperature, the solid was collected by filtration and washed three times with ethanol to obtain the fluorescent probe DCA-Tba. .

[0008] A dual-channel indicator card for fish freshness prepared by the aforementioned fluorescent probe is provided. The indicator card is shaped like a fish and consists of six square colorimetric cards of equal area and a circular sensor label. The six squares are respectively a fresh colorimetric card, a qualified colorimetric card, and a spoilage colorimetric card. The special feature is that the colorimetric card is divided into a natural light colorimetric area and an ultraviolet light colorimetric area. The upper half of the colorimetric card is respectively the fresh natural light colorimetric area, the qualified natural light colorimetric area, and the spoilage natural light colorimetric area, and the lower half of the colorimetric card is respectively the fresh ultraviolet light colorimetric area, the qualified ultraviolet light colorimetric area, and the spoilage ultraviolet light colorimetric area.

[0009] The specific preparation process of the sensor tag is as follows:

[0010] Weigh 3.95 mg of the fluorescent probe DCA-Tba and dissolve it in 10 mL of DMSO. Place a 2 cm diameter circular filter paper into the solution and soak it for 24 hours. Then remove and dry it to obtain a sensor tag loaded with the fluorescent probe. The sensor tag is green when viewed with the naked eye and shows no fluorescence under ultraviolet light.

[0011] Furthermore, the hue corresponding to the natural light colorimetric area of ​​fresh products is green, and the hue corresponding to the ultraviolet light colorimetric area of ​​fresh products is non-fluorescent; the hue corresponding to the natural light colorimetric area of ​​qualified products is light pink, and the hue corresponding to the ultraviolet light colorimetric area of ​​qualified products is pinkish-orange fluorescent; the hue corresponding to the natural light colorimetric area of ​​unqualified products is dark pink, and the hue corresponding to the ultraviolet light colorimetric area of ​​unqualified products is orange fluorescent.

[0012] Application of a dual-channel indicator card for fish freshness in determining fish freshness.

[0013] An application of a dual-channel indicator card for fish freshness in determining fish freshness involves placing the indicator card and fish sample inside the packaging box, ensuring that the sensor label does not directly contact the food. The freshness of the fish is monitored in real time and evaluated based on the color change of the sensor label compared with a standard colorimetric card.

[0014] Under visible light, a green sensor tag indicates freshness, a light pink sensor tag indicates compliance, and a dark pink sensor tag indicates non-compliance. Under 365nm ultraviolet light, a sensor tag with no fluorescence indicates freshness, a pinkish-orange fluorescent sensor tag indicates compliance, and an orange fluorescent sensor tag indicates non-compliance.

[0015] The beneficial effects of this invention are:

[0016] (1) The fluorescent probe has the characteristics of near-infrared emission, with a maximum emission wavelength of 720 nm. It is less affected by scattering and fluorescence background interference, has high quantum efficiency, and stable fluorescence signal. It can respond to a variety of amine compounds in the DMF:H2O= 1:1 (v / v) system.

[0017] (2) The sensor tag prepared by the fluorescent probe can detect the freshness of fish meat through both colorimetric and fluorescence channels. The sensor tag, together with the standard colorimetric card, can monitor the freshness of fish meat in real time.

[0018] In summary, the fluorescent probe designed and synthesized in this invention can not only detect a variety of amine compounds, but can also be prepared into a sensor tag to identify a variety of volatile amines. The sensor tag preparation process is simple, and it can detect fish freshness through both colorimetric and fluorescence channels, resulting in more accurate and reliable qualitative analysis. This sensor tag allows for non-contact and non-destructive real-time monitoring of fish freshness, providing timely and effective freshness information for producers, retailers, and consumers, and has significant practical application value. Attached Figure Description

[0019] Figure 1 It is the fluorescent probe DCA-Tba of this invention. 1 H NMR spectrum;

[0020] Figure 2 It is the fluorescent probe DCA-Tba of this invention. 13 C NMR spectrum;

[0021] Figure 3 This is the high-resolution mass spectrum of the fluorescent probe DCA-Tba of this invention;

[0022] Figure 4 These are the fluorescence emission spectra of the fluorescent probe DCA-Tba of this invention before and after interaction with amine compounds;

[0023] Figure 5 These are the UV absorption spectra of the fluorescent probe DCA-Tba of this invention before and after its interaction with amine compounds;

[0024] Figure 6 These are the fluorescence emission spectra of the fluorescent probe DCA-Tba of this invention after reacting with different concentrations of spermine;

[0025] Figure 7 These are the ultraviolet absorption spectra of the fluorescent probe DCA-Tba of this invention before and after interaction with different concentrations of spermine;

[0026] Figure 8 This describes the linear relationship between the fluorescence intensity of the fluorescent probe DCA-Tba and the concentration of spermine in this invention.

[0027] Figure 9 This relates to the linear relationship between the absorption intensity of the fluorescent probe DCA-Tba and the concentration of spermine in this invention.

[0028] Figure 10 This is a graph showing the change in fluorescence intensity of the fluorescent probe DCA-Tba at different pH values ​​before and after the addition of spermine.

[0029] Figure 11 This describes the change in fluorescence intensity at 600 nm over time after adding spermine to the fluorescent probe DCA-Tba of this invention.

[0030] Figure 12 These are photographs of the changes in sunlight color (top) and fluorescence color (bottom) before and after the sensor tag prepared in this invention identifies volatile amines;

[0031] Figure 13 These are photographs showing the TVB-N content of salmon flesh at 4 ℃ and the corresponding color of the sensor tag under natural light and ultraviolet light as the storage time increases.

[0032] Figure 14It is a standard colorimetric card made based on the correspondence between the TVB-N value of fish meat and the color of the sensor label; the upper layer of the standard colorimetric card, from left to right, is the fresh natural light colorimetric area, the qualified natural light colorimetric area, and the spoiled natural light colorimetric area; the lower layer of the standard colorimetric card, from left to right, is the fresh ultraviolet light colorimetric area, the qualified ultraviolet light colorimetric area, and the spoiled ultraviolet light colorimetric area.

[0033] Figure 15 The freshness of salmon meat is determined by the color change of the label under sunlight and ultraviolet light, combined with a standard colorimetric card, when the sensor label prepared by this invention is stored with salmon samples at 4 ℃. The salmon meat is considered fresh on day 0 of storage at 4 ℃, qualified on day 4, and unqualified on day 8.

[0034] Figure 16 It is an integrated label that combines a standard colorimetric card with a sensor label, enabling rapid colorimetric and fluorescence determination of fish freshness.

[0035] Figure 17 The integrated label prepared by this invention is used to store salmon samples at room temperature (25 ℃). The freshness of salmon meat is quickly determined by the color of sunlight and ultraviolet light. When stored at room temperature for 0 h, the salmon meat is in the fresh grade (upper). When stored for 18 h, the salmon meat is in the qualified grade (middle). When stored for 32 h, the salmon meat is in the unqualified grade (lower). Detailed Implementation

[0036] The technical solution of the present invention will be further described in detail below with reference to specific embodiments.

[0037] Example 1

[0038] The specific synthesis steps of the fluorescent probe DCA-Tba are as follows:

[0039]

[0040] Compound DCA (538 mg, 2 mmol) and 2-thiobarbituric acid (288 mg, 2 mmol) dissolved in 10 mL of ethanol were added, followed by the addition of 5.4 mg of piperidine to obtain a reaction mixture. The reaction mixture was heated under reflux and stirred for 2 hours, cooled to room temperature, and the crude product, a dark green solid, was collected by filtration and washed with ethanol to obtain the fluorescent probe DCA-Tba, with a yield of 85%. 1 H NMR spectrum, 13 C NMR spectrum and mass spectrum as follows Figure 1-3 As shown.

[0041] 1 H NMR (400 MHz, DMSO- d 6) δ 11.65 (s, 2H), 8.15 (s, 1H), 7.59 (s, 1H),7.46 (d, J = 9.0 Hz, 1H), 6.86 (d, J = 9.4 Hz, 1H), 6.81 (s, 1H), 3.50 (dd, J = 15.9, 8.9 Hz, 4H), 3.12 (s, 1H), 2.84 (s, 1H), 1.15 (t, J = 6.9 Hz, 3H), 1.05 (t, J = 7.0 Hz, 3H).

[0042] 13 C NMR (101 MHz, DMSO- d 6 ) δ 176.81, 151.18, 139.57, 133.57, 131.96,129.75, 118.91, 112.92, 111.80, 97.10, 55.14, 44.51, 28.74, 24.95, 18.79,12.64.

[0043] HRMS (ESI - ) for C 21 H 21 N3O3S [M+H] + calcd: 396.1376, found: 396.1385.

[0044] Example 2

[0045] Compound DCA (13.467 g, 50 mmol) and p-aminobenzoic acid (9.37 g, 65 mmol) dissolved in 50 mL of ethanol were added, followed by the addition of 135 mg piperidine. The reaction mixture was heated under reflux and stirred for 3 hours, cooled to room temperature, filtered, and the crude product was collected and washed with ethanol to obtain the fluorescent probe DCA-Tba, with a yield of 77%. In this example, the fluorescent probe DCA-Tba... 1 H NMR spectrum as shown Figure 1 , 13 The C NMR spectrum is as follows Figure 2 High-resolution mass spectrometry, such as Figure 3 As shown.

[0046] Example 3

[0047] Compound DCA (2.6934 g, 10 mmol) and 2-thiobarbituric acid (2.16 g, 15 mmol) dissolved in 20 mL of ethanol were added, followed by the addition of 27 mg piperidine. The reaction mixture was heated under reflux and stirred for 4 hours, cooled to room temperature, and the crude product was collected by filtration and washed with ethanol to obtain the fluorescent probe DCA-Tba, with a yield of 65%. In this example, the fluorescent probe DCA-Tba... 1 H NMR spectrum as shown Figure 1 , 13 The C NMR spectrum is as follows Figure 2 High-resolution mass spectrometry, such as Figure 3 As shown.

[0048] I. Selectivity of fluorescent probe DCA-Tba for amine compounds

[0049] In a 2 mL DMF / H₂O = 1:1 (v / v) system, a 10 μmol / L solution of the fluorescent probe DCA-Tba was added. Then, 20 μL of each of the following solutions (50 mmol / L): cyclohexanediamine, diethylamine, n-propylamine, isopropylamine, triethylamine, ethylamine, spermine, cadaverine, putrescine, 2-phenylethylamine, tyramine, tryptamine, and aniline were added, and the changes in fluorescence intensity were observed. Figure 4 As shown, except for aniline, the solutions of the other 12 amine compounds all caused an increase in fluorescence intensity around 600 nm. Under 365 nm ultraviolet light irradiation, the fluorescence changed from no fluorescence to pink or orange fluorescence. The tested ultraviolet-visible spectra are shown below. Figure 5 As shown, after the addition of various amine compounds, except for aniline, the maximum absorption wavelengths of the other 12 amine compounds showed a significant blue shift, and the color changes were obvious. These results indicate that the fluorescent probe DCA-Tba can achieve colorimetric and fluorescence dual-channel recognition of various amine compounds in a DMF / H2O = 1:1 (v / v) solution, exhibiting excellent selectivity.

[0050] II. Recognition performance of fluorescent probe DCA-Tba on spermine, a representative amine compound

[0051] First, the fluorescence and UV titration of DCA-Tpa with amine compounds were tested. Taking spermine as an example, the fluorescence intensity and UV absorption intensity of the 10 μmol / L fluorescent probe DCA-Tpa in DMF / H2O = 1:1 (v / v) were tested in relation to the concentration of the spermine solution. Figure 6 As shown, as the concentration of spermine solution (0–40 times) gradually increased, the fluorescence intensity of the fluorescent probe DCA-Tba gradually decreased. When 50 μmol / L spermine solution was added, the fluorescence intensity no longer changed, indicating that saturation had been reached. Figure 7As shown, when the concentration of spermine added gradually increases from 0 to 20 times, the maximum UV absorption intensity of the fluorescent probe DCA-Tba gradually decreases. When 200 μmol / L spermine solution is added, the UV absorption intensity no longer changes, indicating that saturation has been reached. Other amines besides aniline exhibit similar effects.

[0052] Secondly, the detection limit for DCA-Tba in recognizing spermine was calculated. Fluorescence intensity was measured in at least 11 parallel samples using a 10 μmol / L DCA-Tba solution, according to the formula: ∑(X i -X) 2 = (X1-X) 2 + (X2-X) 2 +……+ (X n -X) 2 Find the sum of the differences of squares (X) i For each measurement of the receptor's fluorescence intensity, X is the average fluorescence intensity, and n is the number of tests (n≥11). Then, according to the formula: S=[∑(X) i -X) 2 / (n-1)] 0.5 Calculate the sensitivity S, and then use the detection limit formula: Detection limit = 3S / K, where K is the slope of the selected linear segment (Note: the linear segment is based on a titration plot, with the x-axis representing ion concentration and the y-axis representing fluorescence intensity). The calculated detection limit is 1.091 × 10⁻⁶. -5 mol / L (see Figure 8 Based on the UV titration test results, plot a linear graph to calculate Y = AX + B (where A is the slope of the selected linear portion and B is the intercept). When Y = 0, calculate the instantaneous detection limit. According to the formula, the detection limit = 10. -A / B (The horizontal axis represents the logarithmic concentration of spermine, and the vertical axis represents the ratio of (maximum absorption intensity - absorption intensity) to (maximum absorption intensity - absorption intensity value)). The detection limit for ultraviolet light is calculated to be 3.528 × 10⁻⁶. -6 mol / L (see Figure 9 This indicates that the probe can detect low concentrations of spermine in aqueous solution, exhibiting high sensitivity and good potential for practical application.

[0053] Furthermore, the effect of pH on the detection of amine compounds was investigated. Taking spermine as an example, the fluorescence intensity of DCA-Tba under different pH conditions was as follows: Figure 10As shown (Note: the ordinate represents the ratio of fluorescence intensity at 600 and 700 nm), the probe DCA-Tba exhibits good fluorescence intensity within a pH range of 5-11, indicating good stability in weakly acidic, neutral, and weakly alkaline environments. The addition of spermine solution to the probe consistently increased the ratio, indicating a gradual increase in fluorescence intensity at 600 nm. Considering the significant changes in fluorescence before and after recognition, we can conclude that the fluorescent probe DCA-Tba demonstrates a significant recognition effect on spermine within a pH range of 5-11, indicating that this probe has a wide applicable pH range.

[0054] Finally, the response time of the fluorescent probe in recognizing amine compounds was investigated, such as... Figure 11 As shown, after adding spermine solution to the solution containing the fluorescent probe, the fluorescence intensity of the test solution at 600 nm was monitored over time using a fluorescence instrument. A significant increase in fluorescence signal was observed within 2 hours until it plateaued, laying the foundation for the subsequent preparation of a fish freshness sensing tag. Furthermore, we hypothesize the following mechanism by which the fluorescent probe DCA-Tba recognizes volatile amines:

[0055]

[0056] Different volatile amines underwent nucleophilic substitution-elimination reactions with the probe DCA-Tba. Compared with the probe DCA-Tba, the conjugation length of the resulting product was shortened, the push-pull electron system was disrupted, causing the fluorescence emission to shift from long-wavelength emission to short-wavelength emission, and resulting in a decrease in absorption intensity.

[0057] III. The effectiveness of sensor tags in identifying volatile amines in a simulated environment (cylindrical glass bottle, 65 mm high and 18 mm in diameter).

[0058] Preparation process of the sensor tag: Weigh 3.95 mg of DCA-Tba and dissolve it in 10 mL of DMSO. Soak a circular filter paper with a diameter of 2 cm in the above solution overnight, then take it out and dry it to obtain the fluorescent probe DCA-Tba sensor tag. The sensor tag is green when viewed with the naked eye and shows no fluorescence under ultraviolet light.

[0059] During the spoilage process of fish, some volatile metabolites are produced, such as triethylamine, n-propylamine, and diethylamine. Detecting these volatile metabolites can monitor the freshness of the fish. To verify whether the prepared sensor tag is selective for volatile amines, we first placed an aqueous solution containing 0.5% volatile amines in a small bottle, and then placed the sensor tag on top of the bottle to simulate whether the sensor tag responds to volatile amines under spoilage conditions.

[0060] First, 2 mL of a 50 mmol / L volatile amine was added to 3 mL of distilled water to prepare a 20 mmol / L volatile amine solution. Twelve volatile amines, including ethylamine, diethylamine, triethylamine, and n-propylamine, were selected as experimental groups, with pure water serving as the blank control group. The sensor tags in both the 12 experimental groups and the blank control group appeared green under natural light but showed no fluorescence under ultraviolet light. After 1 hour of fumigation with the volatile amines, the color of the sensor tags in all 12 experimental groups changed differently under natural light, and under ultraviolet light, they all exhibited different intensities of pink or orange (see...). Figure 12 The blank control group showed no change. This indicates that the sensor tag has good colorimetric and fluorescence dual responses to 12 volatile amines, including ethylamine, diethylamine, triethylamine, and n-propylamine, suggesting that the sensor tag has the potential to detect the freshness of fish.

[0061] IV. Preparation of Standard Colorimetric Card for Sensor Tags

[0062] To determine the relationship between the color change of the sensor tag and the actual freshness level of the fish, we stored the tag and fish together. The TVB-N content was determined using methods specified in the national standards of the People's Republic of China, and a standard colorimetric card was prepared to identify the freshness level of the fish. We selected salmon meat as the experimental subject. The salmon meat was skinned and cut into multiple 5-6 g pieces, which were placed in disposable round transparent plastic petri dishes. A sensor tag was placed in the opening at the top of the petri dish lid, the lid was sealed, and the dishes were stored at 4 ℃. The TVB-N content of the salmon meat was measured every 24 hours, and natural light and fluorescence photographs of the sensor tags were collected. The obtained data are as follows: Figure 13 As shown, the TVB-N content of fresh salmon meat showed an overall upward trend from an initial 6.53±2.13 mg / 100 g, reaching 15.46±1.40 mg / 100 g on day 4, and then rapidly increasing to 30.18±0.53 mg / 100 g on day 8. The acceptable limit for TVB-N is 30 mg / 100 g. Therefore, day 8 of storage at 4℃ can be considered the threshold for storage spoilage in salmon meat. Under natural light, the sensor tag color changed from initial green to light pink and finally to dark pink; under 365nm ultraviolet light, the sensor tag changed from initially no fluorescence to pinkish-orange fluorescence, and finally to orange fluorescence.

[0063] According to the regulations in the national standard of the People's Republic of China GB / T 18108-2019 (General Rules for Fresh Seawater Fish), the maximum limit of TVB-N content in seawater fish is 30 mg / 100 g. When the TVB-N value ≤ 15 mg / 100 g, the sample is of superior grade; when 15 mg / 100 g < TVB-N value ≤ 30 mg / 100 g, the sample is a qualified product; when the TVB-N value > 30 mg / 100 g, the sample is an unqualified product.

[0064] Based on the measured TVB-N values and the corresponding natural light colors and ultraviolet light color photos of the sensing labels, we selected the sensing labels corresponding to TVB-N values of 6.53 mg / 100 g and 14.74 mg / 100 g, which respectively showed green and light green under natural light, and no fluorescence and light pink fluorescence under ultraviolet light irradiation. These were used as the reference standards for the fresh part in the standard color comparison card. When the sensing label showed one of the above color states, it could be judged that this sample was a fresh product (see Figure 14 the fresh area in Figure 14 ). We selected the sensing labels corresponding to TVB-N values of 15.46 mg / 100 g and 27.46 mg / 100 g, which respectively showed light pink under natural light and medium-strength pink-orange fluorescence under ultraviolet light irradiation. These were used as the reference standards for the qualified part in the standard color comparison card. When the sensing label showed one of the above color states, it could be judged that this sample was a qualified product (see Figure 14 the qualified area in

[0065] V. Practical Application of the Sensing Label

[0066] The sensing label was used for the actual monitoring application of the freshness of salmon. Figure 15 shows the color change of the sensing label of salmon during storage at 4 °C with the increase of storage time. The sensing label was stored together with newly purchased salmon. It showed green under natural light and no fluorescence under ultraviolet light, corresponding to the fresh part in the standard color comparison card; when the storage time reached the 4th day, the natural light color of the sensing label was light pink and it showed medium-strength pink-orange fluorescence under ultraviolet light, corresponding to the qualified part in the standard color comparison card, indicating that the fish was still a qualified product at this time. Further increasing the storage time to the 8th day, the natural light color of the sensing label was bright pink and it showed orange fluorescence under ultraviolet light, corresponding to the spoiled part in the standard color comparison card, indicating that the salmon fish had spoiled and was inedible at this time.

[0067] To improve the convenience of sensor tags, we integrated them with standard color charts to create commercially available integrated tags. For example... Figure 16 As shown, the integrated label is shaped like a fish and consists of six identical square color charts and one circular sensor tag. The six squares represent the fresh, acceptable, and spoiled color charts, respectively. Its unique feature is that the color charts are divided into natural light and ultraviolet (UV) light color matching areas. The upper half of the color charts represents the fresh, acceptable, and spoiled natural light color matching areas, while the lower half represents the fresh, acceptable, and spoiled UV light color matching areas. The circular area is the sensor tag. By observing the color change of the sensor tag in the circular area of ​​the integrated label as the storage time increases, and comparing it with the color of the color matching areas, the freshness of the fish can be determined.

[0068] Taking salmon as an example, integrated tags are used to monitor its freshness. Salmon purchased from the supermarket is placed in a food container and sealed with plastic film. Round holes are punched in the film, and the sensor tag of the integrated tag is aligned with the holes and affixed to the film. The color change of the tag at room temperature over storage time is recorded. Figure 17 As can be seen, when the salmon meat was first stored, the color of the sensor label corresponded to the color of the fresh portion on the color chart. The TVB-N value at this point was 6.27 mg / 100 g, proving that the salmon meat was indeed fresh. After 18 hours at room temperature, the color of the sensor label corresponded to the color of the acceptable area on the color chart, and the TVB-N value was 16.8 mg / 100 g, proving that the salmon meat was within the acceptable range. After 32 hours at room temperature, the color of the sensor label corresponded to the color of the spoiled portion on the color chart, and the TVB-N value was 31.36 mg / 100 g, indicating that the salmon meat at this point exceeded the national standard and was no longer safe to eat.

[0069] In summary, whether in simulated environments or real fish applications, the sensor tag of this invention can detect fish freshness non-contactly and non-destructively using both colorimetric and fluorescence dual-channel detection. The qualitative analysis is more accurate and reliable, providing producers, retailers, and consumers with real-time and effective freshness information, and has good practical application value.

Claims

1. A volatile amine fluorescent probe, characterized in that: The probe structure is as follows:

2. A dual-channel indicator card prepared according to the volatile amine fluorescent probe as described in claim 1, wherein the indicator card is mainly fish-shaped, comprising six square colorimetric cards of equal area and a circular sensor label, wherein the six squares are respectively a fresh colorimetric card, a qualified colorimetric card, and a spoiled colorimetric card, characterized in that: The colorimetric card is divided into a natural light colorimetric area and an ultraviolet light colorimetric area. The upper half of the colorimetric card represents the fresh natural light colorimetric area, the qualified natural light colorimetric area, and the spoiled natural light colorimetric area, respectively. The lower half of the colorimetric card represents the fresh ultraviolet light colorimetric area, the qualified ultraviolet light colorimetric area, and the spoiled ultraviolet light colorimetric area, respectively. The specific preparation process of the sensor tag is as follows: 10 mL of dimethyl sulfoxide (DMSO) solution is added to 10 mg of fluorescent probe. Then, the cut circular filter paper is soaked in the probe solution. After overnight, it is taken out and dried to obtain the sensor tag for monitoring the freshness of fish meat.

3. The dual-channel indicator card prepared by the volatile amine fluorescent probe according to claim 2, characterized in that: The hue of fresh products in the natural light colorimetric area is green, and the hue of fresh products in the ultraviolet light colorimetric area is non-fluorescent; the hue of qualified products in the natural light colorimetric area is light pink, and the hue of qualified products in the ultraviolet light colorimetric area is fluorescent orange-pink; the hue of unqualified products in the natural light colorimetric area is dark pink, and the hue of unqualified products in the ultraviolet light colorimetric area is orange fluorescent.

4. The application of the dual-channel indicator card prepared by the volatile amine fluorescent probe according to claim 2 in determining the freshness of fish meat.

5. The application of the dual-channel indicator card prepared by the volatile amine fluorescent probe according to claim 4 in determining the freshness of fish meat, characterized in that: Align the sensor label in the indicator card with the opening in the packaging, without direct contact with the fish meat. Compare the color change of the sensor label with the standard color chart to monitor the freshness of the fish meat in real time and determine its grade. Under visible light, a green sensor tag indicates the fish is fresh, a light pink sensor tag indicates it is acceptable, and a dark pink sensor tag indicates it is spoiled. Under 365nm ultraviolet light, a sensor tag with no fluorescence indicates the fish is fresh, a sensor tag with medium-intensity orange-pink fluorescence indicates it is acceptable, and a sensor tag with orange fluorescence indicates it is spoiled.

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