Nile blue dye and preparation method thereof, preparation method of packaging film based on prepared dye and application of dye and packaging film

Abstract

The invention discloses a Nile blue dye and a preparation method thereof, a preparation method of a packaging film based on the prepared dye and application of the dye and the packaging film, and belongs to the technical field of food monitoring and preservation. Based on a Nile blue parent structure, different heavy atoms are introduced into a meso-position, and alkyl chains with different chain lengths are introduced into an amino position, so that various photosensitive dyes with pH response and photodynamic therapy capability are constructed. By comparing various heavy atom substitution structures, the dye I and the dye II have certain sensitivity to the pH value of a solution, and the sulfo-dye packaging film and the seleno-dye packaging film both show excellent antibacterial ability to escherichia coli. The dye I packaging film and the dye II packaging film have excellent bacteriostatic activity on escherichia coli. Visual detection of the rotting process of fresh shrimps and preservation of bananas can be realized.

Description

Technical Field

[0001] The present invention belongs to the technical field of food monitoring and preservation, and specifically relates to a Nile blue dye and a preparation method thereof, a preparation method of a packaging film based on the prepared dye, and an application of the dye and the packaging film in the field of food dynamic visualization detection and antibacterial preservation. Background Art

[0002] With economic development and the improvement of living standards, consumers are beginning to pursue higher-quality food with higher cleanliness and freshness. The research and development of food packaging science and technology to meet consumer needs is very urgent. Traditional packaging methods usually only provide static information such as production date and shelf life. Since this static information cannot convey the real-time status and dynamic changes of food freshness, it has limited reference value for consumers to judge the freshness of food.

[0003] Smart packaging can effectively solve the above problems by monitoring the freshness of packaged food in real time during transportation, storage and sales. The application of smart packaging not only improves the quality and safety of food, but also meets consumers' higher demand for food safety. It is an important innovation area for the packaging industry in the future.

[0004] During the food spoilage process, the chemical environment inside the food changes, affecting its pH value. In addition, food is susceptible to microbial contamination at all stages, and the protein in meat and seafood decomposes into alkaline volatile nitrogen compounds (such as ammonia, trimethylamine and dimethylamine, etc.), causing further changes in pH value. Therefore, real-time monitoring of changes in food pH throughout the supply chain can effectively track the freshness and spoilage of packaged foods.

[0005] At present, various methods and techniques for pH sensing and imaging have been reported, including potentiometric titration, microelectrode sensing, and absorption spectroscopy. However, these techniques require relatively professional technical knowledge and are usually operated by technicians in the laboratory, which cannot meet the needs of real-time detection of the freshness of fish and shrimp foods in daily life. As an alternative to traditional detection methods, fluorescent pH-responsive sensors have been used for pH detection and visualization in the fields of optics and biomedicine. Natural pH pigments such as anthocyanins, curcumin, and alizarin have been widely reported as smart films for food packaging due to their visible color changes and non-toxic characteristics. Among them, the most reported is the use of anthocyanins from different sources as food freshness indicators, such as purple sweet potatoes, butterfly pea flowers, purple cabbage, red cabbage, etc. However, anthocyanins are easily degraded and discolored under the influence of environmental factors such as light, temperature, and metal ions, affecting their pH indication ability. In addition, anthocyanins have poor stability, which greatly limits their application in smart food packaging.

[0006] The PCT patent application with publication number WO2017007552A1 discloses a "pH-sensitive nanoparticle for detecting and preventing food spoilage", which involves the technical problems of food pH sensitivity detection and prevention of food spoilage. However, it has the following major defects: First, its pH-sensitive nanoparticles have a complex structure, are difficult to synthesize and purify, and have poor repeatability. It needs to use exogenous hydrophobic active agents as indicators to indicate or prevent food spoilage. Second, the nanoparticles use Nile red dye as an exogenous hydrophobic active agent, and use its fluorescence signal to indicate changes in environmental pH, thereby realizing monitoring of food spoilage. It needs to use additional large instruments such as fluorescence spectrometers for operation, and the process is complicated. Secondly, it is impossible to achieve real-time dynamic visual monitoring, and the purpose of real-time visual monitoring cannot be achieved by the naked eye. Third, the nanoparticles use Nile red dye as an exogenous hydrophobic active agent. According to the changes in environmental pH, the amount of Nile red released changes relatively, and the food spoilage is indirectly indicated according to the changes in the depth of red in the solution. The color recognition is limited and it is easy to cause confusion. Summary of the invention

[0007] The technical problem to be solved by the present invention is to provide a Nile blue dye and a preparation method, a preparation method of a packaging film based on the prepared dye, and uses of the dye and the packaging film, and to explore and develop a dye with high sensitivity, good stability and good biocompatibility, which can be used to monitor pH changes during food storage on the one hand, and to extend the shelf life of food on the other hand.

[0008] The present invention adopts the following technical solutions: A Nile blue dye, the Nile blue dye structural formula is as follows: .

[0009] The preparation method of Nile blue dye comprises the following steps: (1) 1-naphthylamine, K 2 CO 3 and iodoethane are added to ethanol, heated under reflux and stirred; the solvent is removed, and the crude product is eluted and purified to obtain compound B; (2) 3-iodoaniline and K 2 CO 3 Dissolve in acetonitrile, add ethyl iodide, and use N 2 Protective heating and reflux stirring are performed to react; after the reaction is completed, the reaction is cooled, the solvent is removed, and the crude product is eluted and purified to obtain compound C; (3) Compound C, selenium powder, dimethyl sulfoxide, copper oxide and potassium hydroxide are placed in a container and heated with N 2 Protective heating and reflux reaction; extract after filtration, dry and remove the solvent; the crude product is eluted and purified to obtain compound D; (4) In an ice bath, add hydrochloric acid to compound D while stirring, and then add NaNO 2 , and then stirred in an ice bath; extracting, drying, removing the solvent, and eluting and purifying the crude product to obtain compound E; (5) Compound E and compound B prepared in step (1) were dissolved in trifluoroethanol, heated and stirred, and N 2 The method comprises the steps of: protecting the heating reflux reaction; cooling the solution after the reaction stops, adding hydrochloric acid to adjust the pH value of the solution to acidity; extracting, drying, purifying, and rotary evaporating the solution into powder to obtain Nile blue dye for dynamic visualization detection of food and antibacterial preservation.

[0010] Furthermore, in the step (1), 1-naphthylamine: K 2 CO 3 The molar ratio of iodoethane is 1: (1-1.5): (1-1.5); the heating reflux condition is 80-90 °C for 2-6 hours; in the step (2), 3-iodoaniline: K 2 CO 3 The molar ratio of compound C to selenium powder is 1:(2-3):(2-3); the reaction conditions for heating and refluxing are 80-85°C for 14-18 hours; in the step (3), the molar ratio of compound C, selenium powder, copper oxide and potassium hydroxide is 1:(1.5-2.5):(0.4-0.6):(6-8); the reaction conditions for heating and refluxing are 95-105°C for 10-14 hours; the reaction time for stirring in an ice bath in the step (4) is 0.3-4 hours; in the step (5), the molar ratio of compound E to compound B is 1:(2-4); the reaction conditions for heating and refluxing are 85-95°C for 1-3 hours; and the acidification conditions are pH 2-3.

[0011] The method for preparing the packaging film based on the Nile blue dye comprises the following steps: The Nile blue dye is dispersed in a dimethyl sulfoxide solution, and ultrasonic treatment is performed to form a uniform dispersion to obtain a mother solution; sodium alginate is dissolved in water and heated to dissolve; the mother solution is dissolved in a sodium alginate solution, stirred, allowed to stand, poured into a mold, and dried to obtain a packaging film of the Nile blue dye for dynamic visual detection of food and antibacterial preservation.

[0012] Furthermore, the mass ratio of Nile blue dye to sodium alginate is 1:(450-550); the heating dissolution conditions are 45-55°C, 3-6 h; and the film forming temperature is 45-55°C.

[0013] The method for preparing a packaging film of Nile blue dye prepared by the method for preparing Nile blue dye comprises the following steps: The Nile blue dye is dispersed in a dimethyl sulfoxide solution, and ultrasonic treatment is performed to form a uniform dispersion to obtain a mother solution; sodium alginate is dissolved in water and heated to dissolve; the mother solution is dissolved in a sodium alginate solution, stirred, allowed to stand, poured into a mold, and dried to obtain a packaging film of the Nile blue dye for dynamic visual detection of food and antibacterial preservation.

[0014] Furthermore, the mass ratio of Nile blue dye to sodium alginate is 1:(450-550); the heating dissolution conditions are 45-55°C, 3-6 h; and the film forming temperature is 45-55°C.

[0015] The Nile blue dye is used for dynamic visual detection of food and antibacterial preservation.

[0016] The Nile blue dye prepared by the preparation method of the Nile blue dye is applied to dynamic visual detection of food and antibacterial preservation.

[0017] The packaging film prepared by the packaging film preparation method is applied to food dynamic visualization detection and antibacterial preservation.

[0018] Compared with the prior art, the present invention has the following beneficial effects: Nile blue is a benzophenoxazine dye, which is usually used for in vitro tissue staining imaging of RNA, lysosomes or lipids and photodynamic therapy. Due to its excellent photophysical properties, stability and good biosafety, it has become a research hotspot in recent years. Photodynamic antimicrobial therapy (PDAT) with Nile blue dye as the core has been widely reported. The mechanism of PDAT is that under the excitation of a light source of a specific wavelength, the photosensitizer changes from the ground state (S 0 ) transitions to the singlet excited state (S 1 ), S 1 Through intersystem crossing (ISC) to the triplet excited state (T 1 ), which directly or indirectly transfers energy to oxygen, thereby generating reactive oxygen species (ROS). The generated ROS will interact with cells, causing the cytoplasm to leak out, and eventually leading to the death of microorganisms. Therefore, by regulating the relationship between the structure and performance of Nile blue dyes, applying them to intelligent food monitoring, and realizing food preservation and dynamic visualization detection is an effective technical path with broad prospects.

[0019] Based on the parent structure of Nile blue, the present invention constructs a variety of photosensitizing dyes with pH response and photodynamic therapy capabilities by introducing different heavy atoms at the middle position and introducing alkyl chains of different chain lengths at the amino position. By comparing various heavy atom substitution structures, it is found that both thio dyes (dye I of the present invention) and seleno dyes (dye II of the present invention) have a certain sensitivity to the pH of the solution, and both thio dye packaging films and seleno dye packaging films show excellent antibacterial ability against Escherichia coli. Furthermore, seleno dyes are more sensitive to the pH of the solution and begin to change color at a pH of 8; compared with the dye I packaging film of the present invention, the dye II packaging film has stronger antibacterial activity against Escherichia coli.

[0020] The present invention dopes dye I and dye II with sodium alginate as a substrate, respectively, and prepares dye I packaging film and dye II packaging film by using a casting method. Dye I packaging film and dye II packaging film are used to perform visual detection tests on fresh shrimps as the storage time increases. It is found that the dye I packaging film has obvious color changes starting from the third day, and the dye II packaging film has obvious color changes on the second day. Both the dye I packaging film and the dye II packaging film have banana preservation effects. Among them, the dye II packaging film has a longer banana preservation time compared with the dye I packaging film. The Nile blue dye and packaging film of the present invention can achieve more efficient and rapid pH-sensitive detection and light-inhibitory antiseptic. BRIEF DESCRIPTION OF THE DRAWINGS

[0021] Figure 1 The graphs are the color change and ultraviolet absorption spectrum of the Nile blue photosensitizer I obtained in Example 5 of the present invention in different pH solutions.

[0022] Figure 2 The graphs are the color change and ultraviolet absorption spectrum of the Nile blue photosensitizer II obtained in Example 5 of the present invention in solutions with different pH values.

[0023] Figure 3 This is a diagram showing the color changes of Nile blue type photosensitizing dye I packaging film and Nile blue type photosensitizing dye II packaging film obtained in Example 6 of the present invention in different pH solutions. In the figure, film I represents a packaging film based on dye I, and film II represents a packaging film based on dye II.

[0024] Figure 4 This is a diagram showing the color changes of the Nile blue type photosensitizing dye I packaging film and the Nile blue type photosensitizing dye II packaging film obtained in Example 7 of the present invention in various volatile basic nitrogen solutions. In the diagram, film I represents a packaging film based on dye I, and film II represents a packaging film based on dye II.

[0025] Figure 5 This is a graph showing the antibacterial performance test of Nile blue photosensitizer I and Nile blue photosensitizer II obtained in Example 8 of the present invention on Staphylococcus aureus.

[0026] Figure 6 This is a graph showing the antibacterial performance test of the Nile blue type photosensitizing dye I packaging film and the Nile blue type photosensitizing dye II packaging film against Escherichia coli obtained in Example 9 of the present invention. In the graph, film I represents a packaging film based on dye I, and film II represents a packaging film based on dye II.

[0027] Figure 7 This is a diagram showing the visual detection of fresh shrimp decay during the process of the Nile blue type photosensitizing dye I packaging film and the Nile blue type photosensitizing dye II packaging film obtained in Example 10 of the present invention. In the figure, film I represents the packaging film based on dye I, and film II represents the packaging film based on dye II.

[0028] Figure 8 This is a diagram showing the effects of the Nile blue type photosensitizing dye I packaging film and the Nile blue type photosensitizing dye II packaging film obtained in Example 11 of the present invention on the preservation of bananas. In the figure, film I represents a packaging film based on dye I, and film II represents a packaging film based on dye II. DETAILED DESCRIPTION

[0029] The specific implementation modes of the present invention are further described below in conjunction with examples. It should be noted that the specific implementation modes described herein are only for illustrating and explaining the present invention and are not intended to limit the protection scope of the present invention.

[0030] Example 1 (as a comparative example) Preparation example of Nile blue type photosensitizing dye I (abbreviated as dye I).

[0031] (1) In a reaction vessel, N,N -Diethyl-p-phenylenediamine (1.64 g, 10 mmol) was added to 20 mL of an aqueous solution of aluminum sulfate (4.11 g, 12 mmol) and stirred continuously. Sodium thiosulfate (4.42 g, 28 mmol) and zinc chloride (1.74 g, 12.78 mmol) were added in sequence. The reaction vessel was placed in an ice bath and about 8 mL of an aqueous solution of potassium dichromate (0.98 g, 3.36 mmol) was added within 20 min. After continuous stirring in an ice bath for 2 h, the obtained product was washed with acetone and filtered. The obtained crude product was refluxed in 24 mL of methanol, filtered to obtain a dark gray solid, and dried to obtain compound A having the following structure: .

[0032] (2) 1-naphthylamine (0.89 g, 6.25 mmol), potassium carbonate (1.05 g, 7.58 mmol) and iodoethane (1.04 g, 6.61 mmol) were added to 12 mL of ethanol and refluxed at 85 °C for 4 h. The solvent was removed by vacuum distillation, and the crude product was eluted with petroleum ether and ethyl acetate as eluents and purified by silica gel column chromatography to obtain a yellow oil with the following structure Compound B: .

[0033] (3) Compound A (298 mg, 1 mmol) and compound B (308 mg, 1.8 mmol) were added to 25 mL of methanol and refluxed at 70 °C with stirring. Then, silver carbonate (606 mg, 2.2 mmol) was slowly added to the reflux reaction. The reaction mixture was heated for another 30 min until it turned dark blue and cooled to room temperature. The solvent was removed by vacuum distillation to obtain a dark blue crude product. The crude product was dissolved in 25 mL of dichloromethane and extracted three times with saturated aqueous sodium carbonate solution (3×100). The mixed organic extracts were dried over anhydrous sodium sulfate, filtered, acidified with concentrated hydrochloric acid (0.4 mL), and concentrated. The product was purified by silica gel column chromatography using methanol and dichloromethane as eluents to obtain dye I having the following structural formula: .

[0034] Example 2 Preparation of Nile blue type photosensitizing dye II (abbreviated as dye II).

[0035] (1) 3-iodoaniline (1 g, 5 mmol), K 2 CO 3 (1.6 g, 11.5 mmol) was dissolved in 30 mL of MeCN, and iodoethane (1.8 g, 11.5 mmol) was added dropwise. 2 The mixture was protected and placed in an oil bath at 82 °C for 16 h under reflux stirring. After the reaction was completed, the mixture was cooled to room temperature and the reaction progress was monitored by thin layer chromatography (TLC). Silica gel powder was added and rotary evaporated in a vacuum rotary evaporator to form a powder. Finally, column chromatography (PE: EtOAc = 20:1) was used for purification to obtain an orange-yellow oily compound C having the following structural formula: .

[0036] (2) Compound C (1.1 g, 4 mmol), selenium powder (0.632 g, 8 mmol), dimethyl sulfoxide (15 mL), copper oxide (0.152 g, 2 mmol) and potassium hydroxide (1.692 g, 28.8 mmol) were placed in a container and heated to 40 ℃. 2 Protect and place the container in an oil bath, and reflux the reaction at 100 ° C for 12 hours. Place filter paper in a Buchner funnel to vacuum filter the solution. The filtered solution is grass green, and then extracted multiple times with an EtOAc / saturated saline system. Take the upper organic phase to obtain an orange-red solution. The resulting solution is monitored by TLC, and the target product is observed to be generated, and the product contains residual raw materials. The solvent is removed by a vacuum rotary evaporator, and silica gel powder is added to the rotary evaporator to form a powder. After column chromatography (PE:DCM=2:1), compound D with the following structural formula is obtained: .

[0037] (3) Compound D (0.36 g, 0.8 mmol) was added to a 25 mL container, 1 mol / L HCl was added, a magnet was added to the container and the container was placed in a magnetic stirrer with an ice bath, and 1.2 mL of NaNO was added dropwise while stirring. 2 After the addition was completed, the mixture was stirred in an ice bath for 0.5 h. The solution was observed to be dark red. The mixture was extracted several times with an EtOAc / saturated saline solution system. The upper organic phase was taken for TLC detection reaction and a column chromatography mobile phase suitable for the target product was obtained. 2 SO 4 After drying overnight, the solvent was removed by vacuum rotary evaporator, silica gel powder was added to rotary evaporate it into powder, and after purification by column chromatography (MeOH:DCM=1:20), a red powder compound E with the following structural formula was obtained: .

[0038] (4) Compound E (102.8 mg, 0.2 mmol) and compound B prepared in Example 1 (321.6 g, 0.6 mmol) were dissolved in 5 mL of trifluoroethanol in a container. A magnetic stirrer was added and the container was placed in an oil bath. 2Protect and heat to reflux at 90 °C for 2 h. After the reaction stops, the solution is observed to be black-red. After cooling naturally, 1 mL of 1M HCl is added for acidification, and the pH value is 2-3 as detected by pH paper. Extract multiple times with DCM / saturated saline solution system, take the lower organic phase and dry it overnight with anhydrous sodium sulfate. Add silica gel powder and evaporate it into powder. Purify it by column chromatography (methanol: dichloromethane = 1:10) to obtain a blue powder product dye II with the following structural formula: .

[0039] Example 3 (as a comparative example) is based on the preparation example of the packaging film of dye I.

[0040] The dye I (3.581 mg) prepared in Example 1 was dispersed in 3 mL of dimethyl sulfoxide solution and ultrasonically treated for 1 min to form a uniform dispersion to obtain mother solution I. 0.5 g of sodium alginate was dissolved in 50 mL of deionized water and heated at 50 °C for 4 h. After it was completely dissolved, 833.3 μL of mother solution I was dissolved in 50 mL of sodium alginate solution (10 mg / mL), stirred for 30 min, allowed to stand overnight, poured into a mold to form a film, and the mold was placed in an oven and heated at 50 °C for 48 h to obtain a packaging film based on dye I.

[0041] Example 4 is an example of preparing a packaging film based on dye II.

[0042] Dye II (3.684 mg) prepared in Example 2 was dispersed in 3 mL of dimethyl sulfoxide solution and ultrasonically treated for 1 min to form a uniform dispersion to obtain mother solution II. 0.5 g of sodium alginate was dissolved in 50 mL of deionized water and heated at 50°C for 4 h. After it was completely dissolved, 833.3 μL of mother solution II was dissolved in 50 mL of sodium alginate solution (10 mg / mL), stirred for 30 min, allowed to stand overnight, poured into a mold to form a film, and the mold was placed in an oven and heated at 50°C for 48 h to obtain a packaging film based on dye II.

[0043] Example 5 Characterization example of color change of dye I solution and dye II solution in different pH solutions.

[0044] The color and ultraviolet absorption spectra of the dye I prepared in Example 1 and the dye II prepared in Example 2 were analyzed in different pH solutions, and the results were as follows: Figure 1 and Figure 2 Color and UV absorption spectrum shown.

[0045] Depend on Figure 1It can be seen that the dye I solution is blue at pH 2-9, transitions from blue to purple at pH 10, and finally turns pink at pH 11, and the pink color gradually deepens with the increase of pH. Figure 2 It can be seen that the dye II solution is blue at a pH value of 2-8, at a pH value of 9, the solution color transitions from blue to light purple, and at a pH value of 10-12, the pink color gradually deepens.

[0046] Conclusion: As the pH increases, the solution color of both dye I and dye II changes from blue to purple and finally to pink, and the pink color gradually deepens as the pH increases. Furthermore, dye II is more sensitive to pH than dye I, and the color changes at pH 9.

[0047] Example 6 Characterization of the color changes of dye I packaging film and dye II packaging film in solutions with different pH values.

[0048] The color changes of the dye I packaging film prepared in Example 3 and the dye II packaging film prepared in Example 4 in different pH solutions were tested. The dye I packaging film and the dye II packaging film were immersed in a buffer solution with a pH range of 4 to 13 for 5 minutes. The color changes of the packaging film were captured using a mobile phone camera. Figure 3 Schematic diagram of the color change of dye I packaging film and dye II packaging film at different pH values.

[0049] Depend on Figure 3 It can be seen that when the pH value of the solution is between 4-9, the color of the dye I packaging film is blue. When the pH value of the solution is 10-11, the color of the dye I packaging film transitions from gray-blue to purple, and finally turns pink at pH 12, and the pink color gradually deepens with the increase of pH. When the pH value of the solution is between 4-8, the color of the dye II packaging film is blue. When the pH value of the solution is 9-10, the color of the dye II packaging film transitions from gray-blue to light purple, and finally turns pink at pH 11, and the pink color gradually deepens with the increase of pH.

[0050] Conclusion: With the increase of pH, the solution color of both Dye I packaging film and Dye II packaging film changes from blue to gray-blue and purple, and finally turns pink, and the pink color gradually deepens with the increase of pH. Furthermore, Dye II packaging film is more sensitive to pH and has produced color changes at pH 9.

[0051] Example 7 Characterization of the color changes of Dye I packaging film and Dye II packaging film in various volatile basic nitrogen.

[0052] The dye I packaging film prepared in Example 3 and the dye II packaging film prepared in Example 4 were placed in various volatile basic nitrogen environments, and the color changes of the films were detected. Specifically, the dye I packaging film and the dye II packaging film were placed in 10 mL of ammonia, dimethylamine, and trimethylamine (2 mol / L) solution, and the color changes were recorded every 2 minutes using a mobile phone camera for a total of 10 minutes. Figure 4 The graphs shown are the color changes of Nile blue type photosensitizer dye I packaging film and Nile blue type photosensitizer dye II packaging film in various volatile basic nitrogen solutions.

[0053] like Figure 4 In ammonia solution, the color of Dye I packaging film changed from blue to purple within 6 minutes, and then to pink in 8 minutes; in dimethylamine solution, the color of Dye I packaging film changed from blue to purple within 4 minutes, and then to pink in the 6th minute; in trimethylamine solution, the color of Dye I packaging film immediately turned pink after two minutes. In the three volatile basic nitrogen solutions, the color of Dye II packaging film quickly changed from blue to pink within 2 minutes, which shows that both Dye I packaging film and Dye II packaging film have sensitivity and universality. However, Dye II packaging film has better sensitivity and universality.

[0054] Example 8 Characterization example of the antibacterial test of dye I and dye II for Staphylococcus aureus.

[0055] First, a single Staphylococcus aureus colony was picked and transferred to a conical flask containing LB medium (20 mL), and cultured on a shaker for 12 h (speed: 150 rpm / min, temperature: 37°C); then, the obtained fresh bacterial suspension was centrifuged at 5000 rpm / min for 5 min, the supernatant was discarded, and the precipitated bacteria were redispersed in sterile PBS solution, and centrifuged and washed twice at the same speed; finally, the bacteria were directly diluted to 10 with PBS solution. 3 cfu / mL for future use.

[0056] Add 3 mL of the reserved bacterial solution (10 3 cfu / mL), PBS, dye I, and dye II were added to the culture medium to make the concentration 50 μM. The PBS-treated culture medium was set as the control group, the PBS-treated culture medium was set as the illumination group, the dye I and dye II-treated culture medium were set as the dye I group and dye II group, respectively, and the dye I and dye II-treated culture medium were set as the dye I + illumination group and dye II + illumination group, respectively. Among them, the illumination group was incubated in a 37°C shaker after the dye was added, and 2 hours later, it was irradiated with a 650 nm xenon lamp for 10 minutes (light dose of 50 mW / cm 2), and then incubated in a 37℃ shaker for 12 hours. 0.1 mL of bacterial solution from each group was spread on an agar plate and incubated in a 37℃ incubator for 24 hours to obtain Figure 5 The figure shows the test results of the antibacterial performance of Nile blue photosensitizer I and Nile blue photosensitizer II on Staphylococcus aureus.

[0057] Depend on Figure 5 It can be seen that the PBS treatment group was used as the control group, the light-only group had no obvious antibacterial activity, and the dye I solution and dye II solution did not show obvious antibacterial activity after dark treatment. The light-treated groups of dye I solution and dye II solution had an inhibition efficiency of 74.8% and 96.3% on Staphylococcus aureus after light exposure, respectively. The above experiments show that both dye I and dye II show significant antibacterial ability against Staphylococcus aureus. Furthermore, compared with dye I, dye II has stronger antibacterial activity against Staphylococcus aureus.

[0058] Example 9 Characterization example of using dye I packaging film and dye II packaging film for Escherichia coli antibacterial test.

[0059] First, a single E. coli colony was picked and transferred to a conical flask containing LB medium (20 mL), and cultured on a shaker for 12 h (speed: 150 rpm / min, temperature: 37°C); then, the fresh bacterial suspension was centrifuged at 5000 rpm / min for 5 min, the supernatant was discarded, and the precipitated bacteria were redispersed in sterile PBS solution, and centrifuged and washed twice at the same speed; finally, the bacteria were directly diluted to 10 with PBS solution. 6 cfu / mL for future use.

[0060] 0.1 mL (10 6 cfu / mL) of bacterial solution was applied on the agar plate, and the packaging films of dye I and dye II were cut into squares with a side length of 1.0 cm, respectively, and attached to the middle of the agar plate. The plate was incubated in a 37°C incubator for 2 hours, and then irradiated with a 650 nm xenon lamp for 10 minutes (light dose of 50 mW / cm 2 ) and incubated in a 37°C incubator for 24 hours. The diameter of the inhibition zone was measured and obtained. Figure 6 The figure shows the test results of the antibacterial performance of Nile blue type photosensitizer dye I packaging film and Nile blue type photosensitizer dye II packaging film on Escherichia coli.

[0061] Depend on Figure 6It can be seen that the antibacterial activity of the light-exposed group without any treatment is negligible, and the antibacterial activity of the dark-treated dye I packaging film and dye II packaging film is negligible. The inhibition efficiency of dye I packaging film and dye II packaging film against Escherichia coli under light conditions is 50.0% and 52.4%, respectively. The above experiments show that both dye I packaging film and dye II packaging film show obvious antibacterial ability against Escherichia coli. However, compared with dye I packaging film, dye II packaging film has stronger antibacterial activity against Escherichia coli.

[0062] Example 10 Characterization example of using dye I packaging film and dye II packaging film for visual detection of freshness of fresh shrimp.

[0063] The dye I packaging film prepared in Example 3 and the dye II packaging film prepared in Example 4 were used for visual detection of the freshness of fresh shrimp. First, the fresh shrimp sample (10±1.0 g) was placed in a transparent culture dish (Φ 90 mm), and the inside of the culture dish cover was affixed with a rectangular dye I packaging film and a dye II packaging film (length: 1.5 cm, width: 0.8 cm). Then, the fresh shrimp sample was stored at 25±1 ℃, and the color change of the indicator film was recorded regularly using the camera function of a smartphone to obtain Figure 7 Visual detection chart of freshness of fresh shrimp.

[0064] Depend on Figure 7 It can be seen that as the fresh shrimp deteriorates, the sodium alginate film does not change color, while the dye I packaging film and the dye II packaging film both change from blue to pink. Among them, the dye II packaging film changes from blue-green to purple on the second day, and the dye I packaging film changes from blue to light purple on the third day, which shows that the dye I packaging film and the dye II packaging film both respond to pH. Among them, the dye II packaging film responds more sensitively.

[0065] Example 11 Characterization of the effects of dye I packaging film and dye II packaging film on the preservation of bananas.

[0066] The dye I packaging film prepared in Example 3 and the dye II packaging film prepared in Example 4 were used to preserve bananas. Bananas were wrapped with sodium alginate film, dye I packaging film and dye II packaging film, respectively, and the sodium alginate film, dye I packaging film were dark-treated, dye II packaging film was dark-treated, dye I packaging film was light-treated, and dye II packaging film was light-treated. The control group was placed in a natural environment to obtain Figure 8 The diagram shows the banana preservation process.

[0067] Depend on Figure 8It can be seen that compared with the natural environment, the packaging film effectively extends the shelf life of bananas. The appearance of the Dye I packaging film after light exposure can still remain intact on the 4th day, and it begins to rot on the 6th day; the appearance of the Dye II packaging film after light exposure can still remain intact on the 6th day, and it begins to rot on the 8th day. These phenomena show that the light exposure groups of Dye I packaging film and Dye II packaging film both show obvious preservation effects. Furthermore, the Dye II packaging film has a longer shelf life.

[0068] The reagents used in the examples of the present invention, except for the reagents for which the preparation methods are provided, are all commercially available.

Claims

1. A Nile blue dye, characterized in that The Nile blue dye structural formula is as follows: 。 2. The method for preparing Nile blue dye according to claim 1, characterized in that The following steps are involved: (1) 1-naphthylamine, K2CO3 and iodoethane are added to ethanol, heated under reflux and stirred; the solvent is removed, and the crude product is eluted and purified to obtain compound B; (2) Dissolve 3-iodoaniline and K2CO3 in ethyl acetate, add iodoethane, and heat with N2 protection and reflux with stirring to react; after the reaction is completed, cool and remove the solvent, and the crude product is eluted and purified to obtain compound C; (3) Compound C, selenium powder, dimethyl sulfoxide, copper oxide and potassium hydroxide are placed in a container, heated and refluxed under N2 protection; filtered and extracted, dried and the solvent was removed; the crude product was eluted and purified to obtain compound D; (4) Add hydrochloric acid to compound D while stirring in an ice bath, then add NaNO2 while stirring, and stir in an ice bath again; extract, dry, remove the solvent, and elute and purify the crude product to obtain compound E; (5) Compound E and compound B prepared in step (1) are dissolved in trifluoroethanol, heated and stirred, and refluxed under N2 protection; after the reaction stops, the solution is cooled and hydrochloric acid is added to adjust the pH of the solution to acidic; extraction, drying, purification, and rotary evaporation are performed to obtain Nile blue dye for dynamic visualization detection of food and antibacterial preservation.

3. The method for preparing Nile blue dye as claimed in claim 2, wherein: In the step (1), the molar ratio of 1-naphthylamine: K2CO3: iodine ethane is 1: (1-1.5): (1-1.5); the heating reflux condition is 80-90°C for 2-6 hours; in the step (2), the molar ratio of 3-iodoaniline: K2CO3: iodine ethane is 1: (2-3): (2-3); the heating and reflux reaction conditions are 80-85°C for 14-18 hours; in the step (3), the molar ratio of compound C, selenium powder, copper oxide and potassium hydroxide is 1: (1.5-2.5): (0.4-0.6): (6-8); the heating and reflux conditions are 95-105°C for 10-14 hours; in the step (4), the reaction time of ice bath stirring is 0.3-4 hours; in the step (5), the molar ratio of compound E to compound B is 1: (2-4); the heating reflux reaction conditions are 85-95 ℃ reaction for 1-3 hours; the acidification condition is pH 2-3.

4. The method for preparing the packaging film of Nile blue dye according to claim 1, characterized in that The following steps are involved: The Nile blue dye is dispersed in a dimethyl sulfoxide solution, and ultrasonic treatment is performed to form a uniform dispersion to obtain a mother solution; sodium alginate is dissolved in water and heated to dissolve; the mother solution is dissolved in a sodium alginate solution, stirred, allowed to stand, poured into a mold, and dried to obtain a packaging film of the Nile blue dye for dynamic visual detection of food and antibacterial preservation.

5. The method for preparing the packaging film according to claim 4, characterized in that: The mass ratio of Nile blue dye to sodium alginate is 1:(450-550); the heating dissolution conditions are 45-55 ℃, 3-6 h; and the film forming temperature is 45-55 ℃.

6. A method for preparing a packaging film of Nile blue dye prepared by the method for preparing Nile blue dye according to claim 2 or 3, characterized in that The following steps are involved: The Nile blue dye is dispersed in a dimethyl sulfoxide solution, and ultrasonic treatment is performed to form a uniform dispersion to obtain a mother solution; sodium alginate is dissolved in water and heated to dissolve; the mother solution is dissolved in a sodium alginate solution, stirred, allowed to stand, poured into a mold, and dried to obtain a packaging film of the Nile blue dye for dynamic visual detection of food and antibacterial preservation.

7. The method for preparing a packaging film according to claim 6, characterized in that: The mass ratio of Nile blue dye to sodium alginate is 1:(450-550); the heating dissolution conditions are 45-55 ℃, 3-6 h; and the film forming temperature is 45-55 ℃.

8. The Nile blue dye according to claim 1 is used for dynamic visual detection and antibacterial preservation of food.

9. The Nile blue dye prepared by the preparation method of the Nile blue dye according to claim 2 or 3 is used for dynamic visualization detection and antibacterial preservation of food.

10. The packaging film prepared by the method for preparing the packaging film according to claim 4, 5, 6 or 7 is used for dynamic visualization detection and antibacterial preservation of food.

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