Nanoemulsion for preventing and treating porcine respiratory diseases and preparation method thereof

Abstract

The invention provides a nano-emulsion for preventing and treating porcine respiratory diseases and a preparation method of the nano-emulsion, and belongs to the field of preparation of nano-emulsion preparations. The nano-emulsion comprises an outer water phase, and a first water-in-oil emulsion core, a second water-in-oil emulsion core, a third water-in-oil emulsion core and a fourth water-in-oil emulsion core which are dispersed in the outer water phase, wherein oil phases of the first water-in-oil emulsion core and the second water-in-oil emulsion core comprise eucalyptus globulus oil and peppermint oil; the D90 particle size of the inner water phase of the first water-in-oil emulsion core is between 100nm and 150nm; the D90 particle size of the inner water phase of the second water-in-oil emulsion core is between 250 nm and 300 nm; oil phases of the third water-in-oil emulsion core and the fourth water-in-oil emulsion core comprise carvacrol and thymol; the D90 particle size of the inner water phase of the third water-in-oil emulsion core is 200-300 nm; and the D90 particle size of the inner water phase of the fourth water-in-oil emulsion core is 500-700nm. The nano-emulsion has long drug effect and long-term stability.

Description

Technical Field

[0001] The present invention relates to the field of manufacturing nanoemulsion preparations, and more particularly, to a nanoemulsion for preventing and treating swine respiratory diseases and a preparation method thereof. Background Art

[0002] The large-scale breeding of pigs has become the trend of the contemporary pig industry. Under the large-scale breeding mode, the density of the pig population is high, and the stress factors increase, which easily cause pigs to get sick or even die. Among them, swine respiratory diseases have become one of the common and serious diseases in pig farms. Antibiotics are an important means to deal with swine respiratory diseases, but the extensive use of antibiotics easily leads to the problems of antibiotic residues and drug resistance in pigs.

[0003] In some antibiotic-free pig farms, attempts have been made to use natural substances such as plant essential oils to prevent or treat swine respiratory diseases and reduce the dosage of antibiotics. In pig houses, the current main utilization method of plant essential oils is to temporarily extract plant essential oils from plants by means of fumigation or steaming and directly apply them. There are many restrictive factors in this application method. In some other attempts, plant essential oils can be prepared into emulsions and applied by spraying or adding them to drinking water; however, the encapsulation rate and stability of these plant essential oil emulsions are relatively poor, and they can only meet the temporary / short-term use requirements. Therefore, there is an urgent need for a preparation with long efficacy and good stability for the prevention and treatment of respiratory diseases during pig breeding.

[0004] It should be noted that the information disclosed in the above background art section is only used to enhance the understanding of the background of the present disclosure, and therefore may include information that does not constitute the prior art known to those of ordinary skill in the art. Summary of the Invention

[0005] The purpose of the present invention is to overcome the above-mentioned deficiencies of the prior art. The present invention provides a nanoemulsion for preventing and treating swine respiratory diseases and a preparation method thereof, so as to improve the long-term efficacy and long-term stability of the nanoemulsion.

[0006] According to the first aspect of the present invention, there is provided a nanoemulsion for preventing and treating swine respiratory diseases, including an outer aqueous phase and a first water-in-oil emulsion core, a second water-in-oil emulsion core, a third water-in-oil emulsion core, and a fourth water-in-oil emulsion core dispersed in the outer aqueous phase;

[0007] Wherein, the oil phase of the first water-in-oil emulsion core and the second water-in-oil emulsion core is a mixture of eucalyptus oil, peppermint oil, and Span 80, and the inner aqueous phase of the first water-in-oil emulsion core and the second water-in-oil emulsion core is water; the D90 particle size of the inner aqueous phase of the first water-in-oil emulsion core is between 100 nm and 150 nm; the D90 particle size of the inner aqueous phase of the second water-in-oil emulsion core is between 250 nm and 300 nm;

[0008] The oil phase of the third water-in-oil emulsion core is a mixture of carvacrol, thymol, span 60, vitamin E, and MCT oil, and the inner water phase of the third water-in-oil emulsion core is an aqueous sodium alginate solution; the D90 particle size of the inner water phase of the third water-in-oil emulsion core is 200 - 300 nm;

[0009] The oil phase of the fourth water-in-oil emulsion core is a mixture of carvacrol, thymol, span 60, vitamin E, ascorbyl palmitate, and MCT oil, and the inner water phase of the fourth water-in-oil emulsion core is an aqueous solution of sodium alginate and hydroxypropyl-β-cyclodextrin; the D90 particle size of the inner water phase of the fourth water-in-oil emulsion core is 500 - 700 nm;

[0010] The outer water phase is an aqueous solution of tween 80, xanthan gum, sodium caseinate, and sodium benzoate.

[0011] According to another aspect of the present invention, there is provided a method for preparing a nanoemulsion for preventing and treating swine respiratory diseases, comprising:

[0012] Dissolve tween 80, xanthan gum, sodium caseinate, and sodium benzoate in water to form a first aqueous solution;

[0013] Prepare a first coarse emulsion by mixing a mixture of eucalyptus oil, peppermint oil, and span 80 with water, and then prepare a first water-in-oil emulsion by high-pressure microfluidic nanoemulsification technology at a pressure of 150 MPa - 200 MPa; after mixing the first water-in-oil emulsion with a part of the first aqueous solution, stir at a speed of 4000 rpm - 6000 rpm for 1 - 3 minutes to form a first water-in-oil-in-water emulsion; the first water-in-oil-in-water emulsion has a first water-in-oil emulsion core, and the D90 particle size of the inner water phase of the first water-in-oil emulsion core is between 100 nm and 150 nm;

[0014] Prepare a second coarse emulsion by mixing a mixture of eucalyptus oil, peppermint oil, and span 80 with water, and then prepare a second water-in-oil emulsion by high-pressure microfluidic nanoemulsification technology at a pressure of 100 MPa - 130 MPa; after mixing the second water-in-oil emulsion with a part of the first aqueous solution, stir at a speed of 4000 rpm - 6000 rpm for 1 - 3 minutes to form a second water-in-oil-in-water emulsion; the second water-in-oil-in-water emulsion has a second water-in-oil emulsion core, and the D90 particle size of the inner water phase of the second water-in-oil emulsion core is between 250 nm and 300 nm;

[0015] Dissolve sodium alginate in water to form a second aqueous solution;

[0016] Dissolve sodium alginate and hydroxypropyl-β-cyclodextrin in water to form a third aqueous solution;

[0017] After preparing a third coarse emulsion from a mixture of carvacrol, thymol, span 60, vitamin E, and MCT oil with a second aqueous solution, a third water-in-oil emulsion is prepared by high-pressure microfluidization nanoemulsification technology at a pressure of 90 MPa to 130 MPa; after mixing the third water-in-oil emulsion with a portion of the first aqueous solution, it is stirred at a speed of 4000 rpm to 6000 rpm for 1 to 3 minutes to form a third water-in-oil-in-water emulsion; the third water-in-oil-in-water emulsion has a third water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the third water-in-oil emulsion core is between 200 nm and 300 nm;

[0018] After preparing a fourth coarse emulsion from a mixture of carvacrol, thymol, span 60, vitamin E, ascorbyl palmitate, and MCT oil with a third aqueous solution, a fourth water-in-oil emulsion is prepared by high-pressure microfluidization nanoemulsification technology at a pressure of 50 MPa to 70 MPa; after mixing the fourth water-in-oil emulsion with a portion of the first aqueous solution, it is stirred at a speed of 4000 rpm to 6000 rpm for 1 to 3 minutes to form a fourth water-in-oil-in-water emulsion; the fourth water-in-oil-in-water emulsion has a fourth water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the fourth water-in-oil emulsion core is between 500 nm and 700 nm;

[0019] After cooling the first water-in-oil-in-water emulsion, the second water-in-oil-in-water emulsion, the third water-in-oil-in-water emulsion, and the fourth water-in-oil-in-water emulsion to 10°C to 15°C, they are mixed and stirred at a speed of 2000 rpm to 3000 rpm for 1 to 3 minutes to form the nanoemulsion.

[0020] The present invention disperses eucalyptus oil and peppermint oil in one oil phase, and carvacrol and thymol are dispersed in another oil phase. Thus, the prepared emulsion has high stability. The particle sizes of the eucalyptus oil / peppermint oil emulsion nuclei are divided into two levels. The particle size of the first water-in-oil emulsion nucleus is smaller and the volatilization rate is fast. The particle size of the second water-in-oil emulsion nucleus is larger and the volatilization rate is slow. When spraying pesticides, the first water-in-oil emulsion nucleus volatilizes quickly and rapidly increases the concentration of eucalyptus oil / peppermint oil in the pigsty, achieving the effect of quickly repelling mosquitoes and flies. The second water-in-oil emulsion nucleus has a certain slow-release effect, so that the concentration of eucalyptus oil / peppermint oil in the pigsty can be maintained for a relatively long time, achieving the purpose of extending the effective duration of deworming and facilitating the reduction of spraying frequency. At the same time, the overall particle sizes of the first water-in-oil emulsion nucleus and the second water-in-oil emulsion nucleus are both nanoscale, and the overall volatilization rate is relatively fast, which is easy to form a local high concentration to disinfect the spraying dead corners, improving the onset time and comprehensiveness of antibacterial disinfection. The particle sizes of the carvacrol / thymol emulsion nuclei are slightly larger, and the release is slightly later and slower, which can achieve a relay in the deworming effect, thus achieving the purpose of long-term repelling of mosquitoes and flies; at the same time, the release rate is slightly slower, which can achieve a long-term disinfection effect. The particle sizes of the carvacrol / thymol emulsion nuclei (i.e., the third water-in-oil emulsion nucleus and the fourth water-in-oil emulsion nucleus) are divided into two levels, achieving a controlled-release effect and further improving the effects of long-term disinfection and long-term repelling of mosquitoes and flies. The particle sizes of the eucalyptus oil / peppermint oil emulsion nuclei are divided into two levels, and the particle sizes of the carvacrol / thymol emulsion nuclei are divided into two levels, which not only realizes the slow release of components to extend the effect, but also improves the stability of the nanoemulsion.

[0021] In addition, vitamin E and ascorbyl palmitate can be added to the oil phase of the carvacrol / thymol emulsion nucleus with a large particle size to improve the antioxidant property, and the oil-phase vitamin E and ascorbyl palmitate have a certain synergistic effect in the antioxidant effect.

[0022] It should be understood that the above general description and the following detailed description are only exemplary and explanatory, and cannot limit the present disclosure. Detailed embodiments

[0023] The present invention provides a nanoemulsion for preventing and treating swine respiratory diseases, comprising an outer aqueous phase and a first water-in-oil emulsion nucleus, a second water-in-oil emulsion nucleus, a third water-in-oil emulsion nucleus, and a fourth water-in-oil emulsion nucleus dispersed in the outer aqueous phase.

[0024] Among them, the oil phase of the first water-in-oil emulsion core and the second water-in-oil emulsion core is a mixture of eucalyptus oil, peppermint oil, and Span 80, and the internal water phase of the first water-in-oil emulsion core and the second water-in-oil emulsion core is water; the D90 particle size (the particle size value corresponding to 90% in the cumulative particle size distribution curve) of the internal water phase of the first water-in-oil emulsion core is between 100 nm and 150 nm; the D90 particle size of the internal water phase of the second water-in-oil emulsion core is between 250 nm and 300 nm;

[0025] The oil phase of the third water-in-oil emulsion core is a mixture of carvacrol, thymol, Span 60, vitamin E, and MCT oil, and the internal water phase of the third water-in-oil emulsion core is an aqueous sodium alginate solution; the D90 particle size of the internal water phase of the third water-in-oil emulsion core is 200 - 300 nm;

[0026] The oil phase of the fourth water-in-oil emulsion core is a mixture of carvacrol, thymol, Span 60, vitamin E, ascorbyl palmitate, and MCT oil, and the internal water phase of the fourth water-in-oil emulsion core is an aqueous solution of sodium alginate and hydroxypropyl-β-cyclodextrin; the D90 particle size of the internal water phase of the fourth water-in-oil emulsion core is 500 - 700 nm;

[0027] The external water phase is an aqueous solution of Tween 80, xanthan gum, sodium caseinate, and sodium benzoate.

[0028] The nanoemulsion provided by the present invention contains four different plant essential oils, namely eucalyptus oil, peppermint oil, carvacrol, and thymol. Eucalyptus oil is a volatile essential oil extracted from the leaves of eucalyptus trees. It not only has the effect of repelling mosquitoes and insects (such as mosquitoes, ants, etc.), but also has a certain inhibitory effect on some viruses and bacteria. Moreover, eucalyptus oil has a certain relieving effect on respiratory symptoms (such as nasal congestion, cough, etc.). The nanoemulsion of the present invention can be applied by spraying. When spraying, the volatiles of eucalyptus oil / peppermint oil can effectively repel mosquitoes and insects in the pigsty, reducing the risk of mosquito-borne diseases. At the same time, when eucalyptus oil / peppermint oil adheres to the facilities in the pigsty or the skin of live pigs, it can have a certain inhibitory and killing effect on bacteria and viruses on the pigsty or the skin. Thus, this nanoemulsion helps to prevent the occurrence of respiratory diseases in live pigs and relieve respiratory symptoms when respiratory diseases occur in live pigs. When a pig respiratory disease breaks out or the symptoms are relatively severe, this nanoemulsion can also be directly added to the drinking water. Both carvacrol and thymol are relatively broad-spectrum natural monoterpene phenol antibacterial compounds, and there is a certain complementarity in their antibacterial spectra. Moreover, both can also relieve inflammation caused by infection. By adding carvacrol and thymol, the nanoemulsion of the present invention can provide a more powerful and more persistent antibacterial and bactericidal effect, and more effectively prevent and treat pig respiratory diseases.

[0029] In the preliminary exploration of the present invention, eucalyptus oil, peppermint oil, carvacrol, and thymol were mixed to prepare an emulsion. However, no matter how the emulsifier and the aqueous phase were adjusted, the emulsions prepared in these preliminary explorations had relatively serious stability problems. It was not only difficult to achieve sufficient emulsification, but also prone to oil-water separation. Moreover, the prepared emulsion also had the problem of rapid color change, possibly because substances such as carvacrol and thymol underwent rapid oxidation.

[0030] To solve this problem, in the further exploration of the present invention, eucalyptus oil and peppermint oil were dispersed in one oil phase, and carvacrol and thymol were dispersed in another oil phase. In this way, the stability of the prepared emulsion was improved. However, it was found in further tests that both the effective time for repelling mosquitoes and the duration of antibacterial action needed to be improved, otherwise there would be a problem of frequent spraying.

[0031] In a further exploration, the technical solution proposed by the present invention was formed. In the present invention, the particle sizes of the eucalyptus oil / peppermint oil emulsion nuclei (the first water-in-oil emulsion nucleus and the second water-in-oil emulsion nucleus) are divided into two levels. The particle size of the first water-in-oil emulsion nucleus is smaller and the volatilization rate is fast, while the particle size of the second water-in-oil emulsion nucleus is larger and the volatilization rate is slow. When spraying pesticides, the first water-in-oil emulsion nucleus volatilizes rapidly and quickly increases the concentration of eucalyptus oil / peppermint oil in the pigsty, achieving the effect of quickly repelling mosquitoes. The second water-in-oil emulsion nucleus has a certain sustained-release effect, so that the concentration of eucalyptus oil / peppermint oil in the pigsty can be maintained for a relatively long time, achieving the purpose of extending the effective duration of deworming and facilitating the reduction of the spraying frequency. At the same time, the overall particle sizes of the first water-in-oil emulsion nucleus and the second water-in-oil emulsion nucleus are all in the nanometer range, and the overall volatilization rate is relatively fast, which is easy to form a local high concentration to disinfect the spraying dead corners, improving the onset time and comprehensiveness of antibacterial disinfection. In the present invention, the particle size of the carvacrol / thymol emulsion nucleus is slightly larger, the release is slightly later and slower, which can achieve a relay in the deworming effect, and thus achieve the purpose of long-term mosquito repellent; at the same time, the release speed is slightly slower, which can achieve a long-term disinfection effect. The particle sizes of the carvacrol / thymol emulsion nuclei (i.e., the third water-in-oil emulsion nucleus and the fourth water-in-oil emulsion nucleus) are divided into two levels to achieve a controlled-release effect, further improving the effects of long-term disinfection and long-term mosquito repellent. More unexpectedly, the particle sizes of the eucalyptus oil / peppermint oil emulsion nuclei are divided into two levels, and the particle sizes of the carvacrol / thymol emulsion nuclei are divided into two levels, which not only realizes the sustained release of the components to extend the effect, but also improves the stability of the nanoemulsion. This may be because the small particle size emulsion nuclei are embedded in the gaps between the large particle size emulsion nuclei, reducing the migration of each emulsion nucleus and thus reducing the ripening process.

[0032] In addition, during the research of the present invention, it was found that in the oil phase of the carvacrol / thymol emulsion core with large particle size, the antioxidant property can be improved by adding vitamin E and ascorbyl palmitate. Simply increasing the dosage of vitamin E without adding ascorbyl palmitate will lead to limited improvement in the antioxidant effect, and the antioxidant effect is also relatively ordinary when only ascorbyl palmitate is used. Therefore, it is speculated that vitamin E and ascorbyl palmitate in the oil phase of the carvacrol / thymol emulsion core with large particle size have a certain synergistic effect on the antioxidant effect.

[0033] In addition, during the research of the present invention, it was found that when hydroxypropyl-β-cyclodextrin is added to the internal aqueous phase of the carvacrol / thymol emulsion core with large particle size, it may be possible to achieve a longer sustained release and may help reduce the oxidation rate of carvacrol / thymol.

[0034] In one embodiment of the present invention, the mass ratio of the first water-in-oil emulsion core to the second water-in-oil emulsion core is in the range of 7:3 to 3:7; the mass ratio of the third water-in-oil emulsion core to the fourth water-in-oil emulsion core is in the range of 7:3 to 3:7.

[0035] In one example, the mass ratio of the first water-in-oil emulsion core to the second water-in-oil emulsion core is 2:1; the mass ratio of the third water-in-oil emulsion core to the fourth water-in-oil emulsion core is 6:5. In this way, a balance can be achieved among long-lasting mosquito repellent, long-lasting bacteriostasis, and ensuring the long-term stability of the emulsion.

[0036] In one example, the mass of eucalyptus oil in the first water-in-oil emulsion core is 2.8% to 4.5% of the total mass of the nanoemulsion. The mass of eucalyptus oil in the second water-in-oil emulsion core is 2.5% to 4.1% of the total mass of the nanoemulsion. The mass of peppermint oil in the first water-in-oil emulsion core is 1.2% to 2.1% of the total mass of the nanoemulsion. The mass of peppermint oil in the second water-in-oil emulsion core is 0.9% to 1.8% of the total mass of the nanoemulsion.

[0037] In one example, the mass of carvacrol in the third water-in-oil emulsion core is 1.5% to 2.1% of the total mass of the nanoemulsion. The mass of thymol in the third water-in-oil emulsion core is 1.5% to 2.1% of the total mass of the nanoemulsion. The mass of carvacrol in the fourth water-in-oil emulsion core is 0.9% to 1.8% of the total mass of the nanoemulsion. The mass of thymol in the fourth water-in-oil emulsion core is 0.9% to 1.8% of the total mass of the nanoemulsion.

[0038] In one example, the D90 particle size of the internal aqueous phase of the first water-in-oil emulsion core is between 115 nm and 125 nm; the D90 particle size of the internal aqueous phase of the second water-in-oil emulsion core is between 280 nm and 290 nm; the D90 particle size of the internal aqueous phase of the third water-in-oil emulsion core is between 250 nm and 260 nm; the D90 particle size of the internal aqueous phase of the fourth water-in-oil emulsion core is between 500 nm and 550 nm.

[0039] In one example, in the third water-in-oil emulsion core, the mass of vitamin E is 3% - 7% of the total mass of carvacrol and thymol; in the fourth water-in-oil emulsion core, the mass of vitamin E is 3% - 7% of the total mass of carvacrol and thymol, and the mass of ascorbyl palmitate is 3% - 5% of the total mass of carvacrol and thymol. Thus, the oxidation of carvacrol and thymol can be reduced, which is beneficial to verifying the antibacterial duration.

[0040] In one example, in the fourth water-in-oil emulsion core, the mass content of hydroxypropyl-β-cyclodextrin in the internal aqueous phase is 0.1% - 0.3%. This is beneficial to improving the antibacterial duration of the cross-linked emulsion.

[0041] In one embodiment of the present invention, in the nanoemulsion, the mass content of eucalyptus oil is 6.3% - 7.5%, the mass content of peppermint oil is 2.7% - 3.5%, the mass content of carvacrol is 2.5% - 3.6%, and the mass content of thymol is 2.5% - 3.6%.

[0042] In one example, the mass of eucalyptus oil in the first water-in-oil emulsion core is 4.5% of the total mass of the nanoemulsion. The mass of eucalyptus oil in the second water-in-oil emulsion core is 2.5% of the total mass of the nanoemulsion. The mass of peppermint oil in the first water-in-oil emulsion core is 2.0% of the total mass of the nanoemulsion. The mass of peppermint oil in the second water-in-oil emulsion core is 1.0% of the total mass of the nanoemulsion. The mass of carvacrol in the third water-in-oil emulsion core is 1.8% of the total mass of the nanoemulsion. The mass of thymol in the third water-in-oil emulsion core is 1.8% of the total mass of the nanoemulsion. The mass of carvacrol in the fourth water-in-oil emulsion core is 1.2% of the total mass of the nanoemulsion. The mass of thymol in the fourth water-in-oil emulsion core is 1.2% of the total mass of the nanoemulsion. In one example, the D90 particle size of the internal aqueous phase of the first water-in-oil emulsion core is 120 nm; the D90 particle size of the internal aqueous phase of the second water-in-oil emulsion core is 285 nm; the D90 particle size of the internal aqueous phase of the third water-in-oil emulsion core is 255 nm; the D90 particle size of the internal aqueous phase of the fourth water-in-oil emulsion core is 520 nm.

[0043] The present invention also provides a method for using a nanoemulsion for preventing and treating swine respiratory diseases. The nanoemulsion can be diluted with water and used by spraying, or added to the drinking water of live pigs. For example, for prevention during the high-incidence season, the nanoemulsion can be diluted 200 times with water and used for environmental spraying, or added to the drinking water (diluted 3000 - 4000 times). When early symptoms appear in live pigs, the nanoemulsion is added to the drinking water (diluted 3000 - 4000 times). Of course, if the symptoms of the live pigs worsen, the nanoemulsion can also assist other treatment means, such as assisting in treatment in combination with antibiotics (for example, adding the nanoemulsion to the drinking water, diluted 3000 times).

[0044] The present invention also provides a method for preparing the nanoemulsion, including:

[0045] Dissolve Tween 80, xanthan gum, sodium caseinate, and sodium benzoate in water to form a first aqueous solution;

[0046] Prepare a first coarse emulsion from a mixture of eucalyptus oil, peppermint oil, and Span 80 and water, and then prepare a first water-in-oil emulsion under a pressure of 150 MPa - 200 MPa by high-pressure microfluidic nanoemulsification technology; mix the first water-in-oil emulsion with a part of the first aqueous solution and stir at a rotation speed of 4000 rpm - 6000 rpm for 1 - 3 minutes to form a first water-in-oil-in-water emulsion; the first water-in-oil-in-water emulsion has a first water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the first water-in-oil emulsion core is between 100 nm and 150 nm;

[0047] Prepare a second coarse emulsion from a mixture of eucalyptus oil, peppermint oil, and Span 80 and water, and then prepare a second water-in-oil emulsion under a pressure of 100 MPa - 130 MPa by high-pressure microfluidic nanoemulsification technology; mix the second water-in-oil emulsion with a part of the first aqueous solution and stir at a rotation speed of 4000 rpm - 6000 rpm for 1 - 3 minutes to form a second water-in-oil-in-water emulsion; the second water-in-oil-in-water emulsion has a second water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the second water-in-oil emulsion core is between 250 nm and 300 nm;

[0048] Dissolve sodium alginate in water to form a second aqueous solution;

[0049] Dissolve sodium alginate and hydroxypropyl-β-cyclodextrin in water to form a third aqueous solution;

[0050] After preparing a mixture of carvacrol, thymol, span 60, vitamin E, and MCT oil with a second aqueous solution into a third coarse emulsion, a third water-in-oil emulsion is prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 90 MPa to 130 MPa; after mixing the third water-in-oil emulsion with a portion of the first aqueous solution, it is stirred at a speed of 4000 rpm to 6000 rpm for 1 to 3 minutes to form a third water-in-oil-in-water emulsion; the third water-in-oil-in-water emulsion has a third water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the third water-in-oil emulsion core is between 200 nm and 300 nm;

[0051] After preparing a mixture of carvacrol, thymol, span 60, vitamin E, ascorbyl palmitate, and MCT oil with a third aqueous solution into a fourth coarse emulsion, a fourth water-in-oil emulsion is prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 50 MPa to 70 MPa; after mixing the fourth water-in-oil emulsion with a portion of the first aqueous solution, it is stirred at a speed of 4000 rpm to 6000 rpm for 1 to 3 minutes to form a fourth water-in-oil-in-water emulsion; the fourth water-in-oil-in-water emulsion has a fourth water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the fourth water-in-oil emulsion core is between 500 nm and 700 nm;

[0052] After cooling the first water-in-oil-in-water emulsion, the second water-in-oil-in-water emulsion, the third water-in-oil-in-water emulsion, and the fourth water-in-oil-in-water emulsion to 10°C to 15°C, they are mixed and stirred at a speed of 2000 rpm to 3000 rpm for 1 to 3 minutes to form the nanoemulsion.

[0053] As follows, taking some of the examples / experimental examples in the R & D stage of the present invention as examples, the preparation method and effects of the nanoemulsion provided by the present invention are introduced.

[0054] During the preparation process of the nanoemulsion in the embodiment of the present invention, unless otherwise stated, the first aqueous solution, the second aqueous solution, and the third aqueous solution are prepared according to the following method. Tween 80 (4 g), xanthan gum (0.5 g), sodium caseinate (1 g), and sodium benzoate (0.2 g) are dissolved in water (200 g) to form the first aqueous solution. Sodium alginate (5 g) is dissolved in water (200 g) to form the second aqueous solution. Sodium alginate (5 g) and hydroxypropyl-β-cyclodextrin (0.5 g) are dissolved in water (200 g) to form the third aqueous solution.

[0055] In the preparation process of the nanoemulsion of the embodiment of the present invention, unless otherwise specified, the following method is adopted: first, four water-in-oil-in-water emulsions (the first water-in-oil-in-water emulsion, the second water-in-oil-in-water emulsion, the third water-in-oil-in-water emulsion, and the fourth water-in-oil-in-water emulsion) are prepared, and then the four water-in-oil-in-water emulsions are mixed to form a nanoemulsion. The mixing conditions are as follows: after the four emulsions are cooled to 10°C to 15°C, they are mixed and stirred at a rotation speed of 2500 rpm for 2 minutes to form a nanoemulsion.

[0056] The following method is adopted in the present invention to evaluate the mosquito repellent duration of the nanoemulsion.

[0057] A net cover is obtained. The net cover has five side gauzes (polyester mosquito net cloth, non-cotton cloth), and the size of each side is 50 cm × 50 cm. The opening of the net cover is used to buckle on the test bench. One of the side gauzes is detachable and is the side gauze for applying the medicine. The side gauze for applying the medicine is removed. The nanoemulsion is diluted 200 times with water, and then sprayed on the side gauze by spray application. The spraying direction is directly towards the side gauze for applying the medicine. Spray once at each of the four corners and the center of the side gauze for applying the medicine, and the distance between the spray head and the gauze during each spraying is 30 cm. After installing the medicated gauze back on the net cover, the net cover is inverted and 20 Culex quinquefasciatus are placed in it, and this moment is recorded as the zero moment. Subsequently, every ten minutes, each non-medicated gauze of the net cover is shaken to make the mosquitoes take off. Observe the staying situation of the mosquitoes on the medicated gauze. When the staying time of the mosquitoes on the medicated gauze exceeds ten seconds, it is recorded as one mosquito stay. The moment when the fourth mosquito stay occurs is recorded as the mosquito repellent duration of the nanoemulsion.

[0058] The present invention uses the following method to detect the antibacterial / bactericidal ability of the nanoemulsion. The bacterium solution of Actinobacillus pleuropneumoniae suis is mixed evenly with the nutrient solution (containing agar solution), and then diluted 3 times to form a low-viscosity and nutrient-containing bacterium solution. The diluted bacterium solution is evenly sprayed onto a plurality of culture dishes by spraying. Then, the culture dishes are left standing at room temperature for 15 minutes to allow the sprayed bacterium solution to volatilize. Taking this as the zero time, a plurality of culture dishes are taken for bacterial culture, and the number of colonies is recorded after the culture; the average value of the number of colonies of these culture dishes at the zero time is the bacterial number reference value N0. At the zero time, the nanoemulsion is sprayed onto the remaining plurality of culture dishes. The specific method of spraying the drug is as follows: The nanoemulsion is diluted 200 times with water, and then sprayed from the side rear of the culture dish. The spraying direction is horizontal and towards the culture dish. The height of the spray head is 40 cm from the height of the culture dish, and the distance of the spray head from the culture dish in the horizontal direction is 30 cm. Spray twice. In this way, the process of the drug droplets naturally falling onto the surface of the pigsty facilities during the spraying of the nanoemulsion is simulated. 6 hours after the spraying of the drug, some of the culture dishes are taken for bacterial culture and the number of colonies N6 formed is recorded. The 6-hour bacterial growth rate of the nanoemulsion is (N6 - N0) / N0. 20 hours after the spraying of the drug, the remaining culture dishes are taken for bacterial culture and the number of colonies N20 formed is recorded. The 20-hour bacterial growth rate of the nanoemulsion is (N20 - N0) / N0.

[0059] The present invention uses the following method to test the stability of the nanoemulsion. The nanoemulsion is filled into a plurality of test tubes and sealed, and then stored at a constant temperature of 45°C. In the first ten days, observation is carried out once a day and oil separation rating is carried out according to the observation results; after ten days, observation is carried out once every two days and oil separation rating is carried out according to the observation results. When oil droplets appear but no oil film appears, it is defined as initial oil separation. When an oil film appears but the thickness of the oil film is not greater than 0.3 mm, it is defined as medium oil separation. When an oil film appears and the thickness of the oil film exceeds 0.3 mm, it is defined as severe oil separation. Record the initial oil separation time, medium oil separation time and severe oil separation time. Monitor up to fifty days.

[0060] Example 1

[0061] After preparing a first crude emulsion from a mixture of eucalyptus oil (7 g), peppermint oil (3 g), span 80 (1 g) and water (3 g), a first water-in-oil emulsion is prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 170 MPa; after mixing the first water-in-oil emulsion with a first aqueous solution (36 g) and stirring at a speed of 5000 rpm for 2 minutes, a first water-in-oil-in-water emulsion is formed; the first water-in-oil-in-water emulsion has a first water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the first water-in-oil emulsion core is 120 nm.

[0062] After preparing a second crude emulsion from a mixture of eucalyptus oil (7 g), peppermint oil (3 g), span 80 (1 g) and water (3 g), a second water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 120 MPa; the second water-in-oil emulsion was mixed with the first aqueous solution (36 g) and stirred at a speed of 5000 rpm for 2 minutes to form a second water-in-oil-in-water emulsion; the second water-in-oil-in-water emulsion had a second water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the second water-in-oil emulsion core was 285 nm.

[0063] After preparing a third crude emulsion from a mixture of carvacrol (3 g), thymol (3 g), span 60 (0.9 g), vitamin E (0.3 g), MCT oil (3 g) and the second aqueous solution (2.7 g), a third water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 105 MPa; the third water-in-oil emulsion was mixed with the first aqueous solution (37.1 g) and stirred at a speed of 5000 rpm for 2 minutes to form a third water-in-oil-in-water emulsion; the third water-in-oil-in-water emulsion had a third water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the third water-in-oil emulsion core was 255 nm.

[0064] After preparing a fourth crude emulsion from a mixture of carvacrol (3 g), thymol (3 g), span 60 (0.9 g), vitamin E (0.3 g), ascorbyl palmitate (0.24 g), MCT oil (3 g) and the third aqueous solution (3.6 g), a fourth water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 50 MPa; the fourth water-in-oil emulsion was mixed with the first aqueous solution (35.96 g) and stirred at a speed of 5000 rpm for 2 minutes to form a fourth water-in-oil-in-water emulsion; the fourth water-in-oil-in-water emulsion had a fourth water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the fourth water-in-oil emulsion core was 520 nm.

[0065] The four water-in-oil-in-water emulsions were cooled and mixed to form nanoemulsion 1.

[0066] The zeta potential of nanoemulsion 1 was -38 V; the mosquito repellent duration was 6.2 h; the colony growth rate at 6 h was -75%; the colony growth rate at 20 h was 275%; the initial oil separation time was 12 days; the medium oil separation time was 36 days; no severe oil separation was observed.

[0067] Example 2

[0068] After preparing a first crude emulsion from a mixture of eucalyptus oil (5.6 g), peppermint oil (2.4 g), and span 80 (0.8 g) with water (2.4 g), a first water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 170 MPa; after mixing the first water-in-oil emulsion with a first aqueous solution (28.8 g) and stirring at a speed of 5000 rpm for 2 minutes, a first water-in-oil-in-water emulsion was formed; the first water-in-oil-in-water emulsion has a first water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the first water-in-oil emulsion core is 120 nm.

[0069] After preparing a second crude emulsion from a mixture of eucalyptus oil (8.4 g), peppermint oil (3.6 g), and span 80 (1.2 g) with water (3.6 g), a second water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 120 MPa; after mixing the second water-in-oil emulsion with a first aqueous solution (43.2 g) and stirring at a speed of 5000 rpm for 2 minutes, a second water-in-oil-in-water emulsion was formed; the second water-in-oil-in-water emulsion has a second water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the second water-in-oil emulsion core is 285 nm.

[0070] After preparing a third crude emulsion from a mixture of carvacrol (3 g), thymol (3 g), span 60 (0.9 g), vitamin E (0.3 g), and MCT oil (3 g) with a second aqueous solution (2.7 g), a third water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 105 MPa; after mixing the third water-in-oil emulsion with a first aqueous solution (37.1 g) and stirring at a speed of 5000 rpm for 2 minutes, a third water-in-oil-in-water emulsion was formed; the third water-in-oil-in-water emulsion has a third water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the third water-in-oil emulsion core is 255 nm.

[0071] After preparing a fourth crude emulsion from a mixture of carvacrol (3 g), thymol (3 g), span 60 (0.9 g), vitamin E (0.3 g), ascorbyl palmitate (0.24 g), and MCT oil (3 g) with a third aqueous solution (3.6 g), a fourth water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 50 MPa; after mixing the fourth water-in-oil emulsion with a first aqueous solution (35.96 g) and stirring at a speed of 5000 rpm for 2 minutes, a fourth water-in-oil-in-water emulsion was formed; the fourth water-in-oil-in-water emulsion has a fourth water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the fourth water-in-oil emulsion core is 520 nm.

[0072] Cool and mix the four water-in-oil-in-water emulsions to form nanoemulsion 2.

[0073] The zeta potential of the nanoemulsion 2 was measured to be -39 V; the mosquito repellent duration was 5.1 h; the colony growth rate at 6 h was -73%; the colony growth rate at 20 h was 302%; the initial oil separation time was 12 days; the medium oil separation time was 38 days; and no severe oil separation was observed.

[0074] Example 3

[0075] After preparing a first coarse emulsion from a mixture of eucalyptus oil (4.2 g), peppermint oil (1.8 g), and span 80 (0.6 g) with water (1.8 g), a first water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 170 MPa; after mixing the first water-in-oil emulsion with a first aqueous solution (21.6 g), it was stirred at a speed of 5000 rpm for 2 minutes to form a first water-in-oil-in-water emulsion; the first water-in-oil-in-water emulsion had a first water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the first water-in-oil emulsion core was 120 nm.

[0076] After preparing a second coarse emulsion from a mixture of eucalyptus oil (9.8 g), peppermint oil (4.2 g), and span 80 (1.4 g) with water (4.2 g), a second water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 120 MPa; after mixing the second water-in-oil emulsion with a first aqueous solution (50.4 g), it was stirred at a speed of 5000 rpm for 2 minutes to form a second water-in-oil-in-water emulsion; the second water-in-oil-in-water emulsion had a second water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the second water-in-oil emulsion core was 285 nm.

[0077] After preparing a third coarse emulsion from a mixture of carvacrol (3 g), thymol (3 g), span 60 (0.9 g), vitamin E (0.3 g), and MCT oil (3 g) with a second aqueous solution (2.7 g), a third water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 105 MPa; after mixing the third water-in-oil emulsion with a first aqueous solution (37.1 g), it was stirred at a speed of 5000 rpm for 2 minutes to form a third water-in-oil-in-water emulsion; the third water-in-oil-in-water emulsion had a third water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the third water-in-oil emulsion core was 255 nm.

[0078] After preparing a mixture of carvacrol (3 g), thymol (3 g), span 60 (0.9 g), vitamin E (0.3 g), ascorbyl palmitate (0.24 g), and MCT oil (3 g) with the third aqueous solution (3.6 g) into a fourth crude emulsion, a fourth water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 50 MPa; after mixing the fourth water-in-oil emulsion with the first aqueous solution (35.96 g), it was stirred at a speed of 5000 rpm for 2 minutes to form a fourth water-in-oil-in-water emulsion; the fourth water-in-oil-in-water emulsion has a fourth water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the fourth water-in-oil emulsion core is 520 nm.

[0079] Cool and mix the four water-in-oil-in-water emulsions to form nanoemulsion 3.

[0080] Measure the zeta potential of nanoemulsion 3 to be -38 V; the mosquito repellent duration is 3.8 h; the colony growth rate at 6 h is -73%; the colony growth rate at 20 h is 280%; the initial oil separation time is 10 days; the medium oil separation time is 32 days; no severe oil separation is observed.

[0081] Example 4

[0082] After preparing a mixture of eucalyptus oil (8.4 g), peppermint oil (3.6 g), and span 80 (1.2 g) with water (3.6 g) into a first crude emulsion, a first water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 170 MPa; after mixing the first water-in-oil emulsion with the first aqueous solution (43.2 g), it was stirred at a speed of 5000 rpm for 2 minutes to form a first water-in-oil-in-water emulsion; the first water-in-oil-in-water emulsion has a first water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the first water-in-oil emulsion core is 120 nm.

[0083] After preparing a mixture of eucalyptus oil (5.6 g), peppermint oil (2.4 g), and span 80 (0.8 g) with water (2.4 g) into a second crude emulsion, a second water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 120 MPa; after mixing the second water-in-oil emulsion with the first aqueous solution (28.8 g), it was stirred at a speed of 5000 rpm for 2 minutes to form a second water-in-oil-in-water emulsion; the second water-in-oil-in-water emulsion has a second water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the second water-in-oil emulsion core is 285 nm.

[0084] After preparing a third coarse emulsion from a mixture of carvacrol (3 g), thymol (3 g), span 60 (0.9 g), vitamin E (0.3 g), MCT oil (3 g) and a second aqueous solution (2.7 g), a third water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 105 MPa; the third water-in-oil emulsion was mixed with a first aqueous solution (37.1 g) and stirred at a speed of 5000 rpm for 2 minutes to form a third water-in-oil-in-water emulsion; the third water-in-oil-in-water emulsion has a third water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the third water-in-oil emulsion core is 255 nm.

[0085] After preparing a fourth coarse emulsion from a mixture of carvacrol (3 g), thymol (3 g), span 60 (0.9 g), vitamin E (0.3 g), ascorbyl palmitate (0.24 g), MCT oil (3 g) and a third aqueous solution (3.6 g), a fourth water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 50 MPa; the fourth water-in-oil emulsion was mixed with a first aqueous solution (35.96 g) and stirred at a speed of 5000 rpm for 2 minutes to form a fourth water-in-oil-in-water emulsion; the fourth water-in-oil-in-water emulsion has a fourth water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the fourth water-in-oil emulsion core is 520 nm.

[0086] The four water-in-oil-in-water emulsions were cooled and mixed to form nanoemulsion 4.

[0087] The zeta potential of nanoemulsion 4 was measured to be -37 V; the mosquito repellent duration was 8.9 h; the colony growth rate at 6 h was -75%; the colony growth rate at 20 h was 295%; the initial oil separation time was 12 days; the medium oil separation time was 36 days; no severe oil separation was observed.

[0088] Example 5

[0089] After preparing a first coarse emulsion from a mixture of eucalyptus oil (9.8 g), peppermint oil (4.2 g), span 80 (1.4 g) and water (4.2 g), a first water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 170 MPa; the first water-in-oil emulsion was mixed with a first aqueous solution (50.4 g) and stirred at a speed of 5000 rpm for 2 minutes to form a first water-in-oil-in-water emulsion; the first water-in-oil-in-water emulsion has a first water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the first water-in-oil emulsion core is 120 nm.

[0090] After preparing a second crude emulsion from a mixture of eucalyptus oil (4.2 g), peppermint oil (1.8 g), span 80 (0.6 g) and water (1.8 g), a second water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 120 MPa; the second water-in-oil emulsion was mixed with the first aqueous solution (21.6 g) and stirred at a speed of 5000 rpm for 2 minutes to form a second water-in-oil-in-water emulsion; the second water-in-oil-in-water emulsion had a second water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the second water-in-oil emulsion core was 285 nm.

[0091] After preparing a third crude emulsion from a mixture of carvacrol (3 g), thymol (3 g), span 60 (0.9 g), vitamin E (0.3 g), MCT oil (3 g) and the second aqueous solution (2.7 g), a third water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 105 MPa; the third water-in-oil emulsion was mixed with the first aqueous solution (37.1 g) and stirred at a speed of 5000 rpm for 2 minutes to form a third water-in-oil-in-water emulsion; the third water-in-oil-in-water emulsion had a third water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the third water-in-oil emulsion core was 255 nm.

[0092] After preparing a fourth crude emulsion from a mixture of carvacrol (3 g), thymol (3 g), span 60 (0.9 g), vitamin E (0.3 g), ascorbyl palmitate (0.24 g), MCT oil (3 g) and the third aqueous solution (3.6 g), a fourth water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 50 MPa; the fourth water-in-oil emulsion was mixed with the first aqueous solution (35.96 g) and stirred at a speed of 5000 rpm for 2 minutes to form a fourth water-in-oil-in-water emulsion; the fourth water-in-oil-in-water emulsion had a fourth water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the fourth water-in-oil emulsion core was 520 nm.

[0093] The four water-in-oil-in-water emulsions were cooled and mixed to form nanoemulsion 5.

[0094] The zeta potential of nanoemulsion 5 was measured to be -38 V; the mosquito repellent duration was 9.5 h; the colony growth rate at 6 h was -69%; the colony growth rate at 20 h was 315%; the initial oil separation time was 14 days; the medium oil separation time was 42 days; no severe oil separation was observed.

[0095] Example 6

[0096] After preparing a first coarse emulsion from a mixture of eucalyptus oil (8.4 g), peppermint oil (3.6 g), and Span 80 (1.2 g) with water (3.6 g), a first water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 170 MPa; after mixing the first water-in-oil emulsion with a first aqueous solution (43.2 g), it was stirred at a speed of 5000 rpm for 2 minutes to form a first water-in-oil-in-water emulsion; the first water-in-oil-in-water emulsion has a first water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the first water-in-oil emulsion core is 120 nm.

[0097] After preparing a second coarse emulsion from a mixture of eucalyptus oil (5.6 g), peppermint oil (2.4 g), and Span 80 (0.8 g) with water (2.4 g), a second water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 120 MPa; after mixing the second water-in-oil emulsion with a first aqueous solution (28.8 g), it was stirred at a speed of 5000 rpm for 2 minutes to form a second water-in-oil-in-water emulsion; the second water-in-oil-in-water emulsion has a second water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the second water-in-oil emulsion core is 285 nm.

[0098] After preparing a third coarse emulsion from a mixture of carvacrol (2.4 g), thymol (2.4 g), Span 60 (0.72 g), vitamin E (0.24 g), and MCT oil (2.4 g) with a second aqueous solution (2.16 g), a third water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 105 MPa; after mixing the third water-in-oil emulsion with a first aqueous solution (29.68 g), it was stirred at a speed of 5000 rpm for 2 minutes to form a third water-in-oil-in-water emulsion; the third water-in-oil-in-water emulsion has a third water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the third water-in-oil emulsion core is 255 nm.

[0099] After preparing a fourth coarse emulsion from a mixture of carvacrol (3.6 g), thymol (3.6 g), Span 60 (1.08 g), vitamin E (0.36 g), ascorbyl palmitate (0.288 g), and MCT oil (3.6 g) with a third aqueous solution (4.32 g), a fourth water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 50 MPa; after mixing the fourth water-in-oil emulsion with a first aqueous solution (43.152 g), it was stirred at a speed of 5000 rpm for 2 minutes to form a fourth water-in-oil-in-water emulsion; the fourth water-in-oil-in-water emulsion has a fourth water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the fourth water-in-oil emulsion core is 520 nm.

[0100] The four water-in-oil-in-water emulsions were cooled and mixed to form nanoemulsion 6.

[0101] The zeta potential of nanoemulsion 6 was measured to be -37 V; the mosquito repellent duration was 9.1 h; the colony growth rate at 6 h was -65%; the colony growth rate at 20 h was 105%; the initial oil separation time was 12 days; the medium oil separation time was 38 days; no severe oil separation was observed.

[0102] Example 7

[0103] After preparing a first coarse emulsion from a mixture of eucalyptus oil (8.4 g), peppermint oil (3.6 g), and span 80 (1.2 g) with water (3.6 g), a first water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 170 MPa; after mixing the first water-in-oil emulsion with a first aqueous solution (43.2 g) and stirring at a speed of 5000 rpm for 2 minutes, a first water-in-oil-in-water emulsion was formed; the first water-in-oil-in-water emulsion has a first water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the first water-in-oil emulsion core is 120 nm.

[0104] After preparing a second coarse emulsion from a mixture of eucalyptus oil (5.6 g), peppermint oil (2.4 g), and span 80 (0.8 g) with water (2.4 g), a second water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 120 MPa; after mixing the second water-in-oil emulsion with a first aqueous solution (28.8 g) and stirring at a speed of 5000 rpm for 2 minutes, a second water-in-oil-in-water emulsion was formed; the second water-in-oil-in-water emulsion has a second water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the second water-in-oil emulsion core is 285 nm.

[0105] After preparing a third coarse emulsion from a mixture of carvacrol (1.8 g), thymol (1.8 g), span 60 (0.54 g), vitamin E (0.18 g), and MCT oil (1.8 g) with a second aqueous solution (1.62 g), a third water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 105 MPa; after mixing the third water-in-oil emulsion with a first aqueous solution (22.26 g) and stirring at a speed of 5000 rpm for 2 minutes, a third water-in-oil-in-water emulsion was formed; the third water-in-oil-in-water emulsion has a third water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the third water-in-oil emulsion core is 255 nm.

[0106] After preparing a mixture of carvacrol (4.2 g), thymol (4.2 g), span 60 (1.26 g), vitamin E (0.42 g), ascorbyl palmitate (0.336 g), and MCT oil (4.2 g) with the third aqueous solution (5.04 g) into a fourth crude emulsion, a fourth water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 50 MPa; after mixing the fourth water-in-oil emulsion with the first aqueous solution (50.344 g), it was stirred at a speed of 5000 rpm for 2 minutes to form a fourth water-in-oil-in-water emulsion; the fourth water-in-oil-in-water emulsion has a fourth water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the fourth water-in-oil emulsion core is 520 nm.

[0107] Cool and mix the four water-in-oil-in-water emulsions to form nanoemulsion 7.

[0108] Measure the zeta potential of nanoemulsion 7 to be -38 V; the mosquito repellent duration is 8.9 h; the colony growth rate at 6 h is -53%; the colony growth rate at 20 h is 79%; the initial oil separation time is 9 days; the medium oil separation time is 26 days; no severe oil separation is observed.

[0109] Example 8

[0110] After preparing a mixture of eucalyptus oil (8.4 g), peppermint oil (3.6 g), and span 80 (1.2 g) with water (3.6 g) into a first crude emulsion, a first water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 170 MPa; after mixing the first water-in-oil emulsion with the first aqueous solution (43.2 g), it was stirred at a speed of 5000 rpm for 2 minutes to form a first water-in-oil-in-water emulsion; the first water-in-oil-in-water emulsion has a first water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the first water-in-oil emulsion core is 120 nm.

[0111] After preparing a mixture of eucalyptus oil (5.6 g), peppermint oil (2.4 g), and span 80 (0.8 g) with water (2.4 g) into a second crude emulsion, a second water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 120 MPa; after mixing the second water-in-oil emulsion with the first aqueous solution (28.8 g), it was stirred at a speed of 5000 rpm for 2 minutes to form a second water-in-oil-in-water emulsion; the second water-in-oil-in-water emulsion has a second water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the second water-in-oil emulsion core is 285 nm.

[0112] After preparing a third crude emulsion from a mixture of carvacrol (3.6 g), thymol (3.6 g), span 60 (1.08 g), vitamin E (0.36 g), and MCT oil (3.6 g) and a second aqueous solution (3.24 g), a third water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 105 MPa; the third water-in-oil emulsion was mixed with a first aqueous solution (44.52 g) and stirred at a speed of 5000 rpm for 2 minutes to form a third water-in-oil-in-water emulsion; the third water-in-oil-in-water emulsion has a third water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the third water-in-oil emulsion core is 255 nm.

[0113] After preparing a fourth crude emulsion from a mixture of carvacrol (2.4 g), thymol (2.4 g), span 60 (0.72 g), vitamin E (0.24 g), ascorbyl palmitate (0.192 g), and MCT oil (2.4 g) and a third aqueous solution (2.88 g), a fourth water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 50 MPa; the fourth water-in-oil emulsion was mixed with a first aqueous solution (28.768 g) and stirred at a speed of 5000 rpm for 2 minutes to form a fourth water-in-oil-in-water emulsion; the fourth water-in-oil-in-water emulsion has a fourth water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the fourth water-in-oil emulsion core is 520 nm.

[0114] The four water-in-oil-in-water emulsions were cooled and mixed to form nanoemulsion 8.

[0115] The zeta potential of nanoemulsion 8 was measured to be -37 V; the mosquito repellent duration was 8.9 h; the colony growth rate at 6 h was -78%; the colony growth rate at 20 h was 425%; the initial oil separation time was 14 days; the medium oil separation time was 34 days; no severe oil separation was observed.

[0116] Example 9

[0117] After preparing a first crude emulsion from a mixture of eucalyptus oil (8.4 g), peppermint oil (3.6 g), span 80 (1.2 g), and water (3.6 g), a first water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 170 MPa; the first water-in-oil emulsion was mixed with a first aqueous solution (43.2 g) and stirred at a speed of 5000 rpm for 2 minutes to form a first water-in-oil-in-water emulsion; the first water-in-oil-in-water emulsion has a first water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the first water-in-oil emulsion core is 120 nm.

[0118] After preparing a second crude emulsion from a mixture of eucalyptus oil (5.6 g), peppermint oil (2.4 g), span 80 (0.8 g) and water (2.4 g), a second water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 120 MPa; the second water-in-oil emulsion was mixed with the first aqueous solution (28.8 g) and stirred at a speed of 5000 rpm for 2 minutes to form a second water-in-oil-in-water emulsion; the second water-in-oil-in-water emulsion has a second water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the second water-in-oil emulsion core is 285 nm.

[0119] After preparing a third crude emulsion from a mixture of carvacrol (4.2 g), thymol (4.2 g), span 60 (1.26 g), vitamin E (0.42 g), and MCT oil (4.2 g) and the second aqueous solution (3.78 g), a third water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 105 MPa; the third water-in-oil emulsion was mixed with the first aqueous solution (51.94 g) and stirred at a speed of 5000 rpm for 2 minutes to form a third water-in-oil-in-water emulsion; the third water-in-oil-in-water emulsion has a third water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the third water-in-oil emulsion core is 255 nm.

[0120] After preparing a fourth crude emulsion from a mixture of carvacrol (1.8 g), thymol (1.8 g), span 60 (0.54 g), vitamin E (0.18 g), ascorbyl palmitate (0.144 g), and MCT oil (1.8 g) and the third aqueous solution (2.16 g), a fourth water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 50 MPa; the fourth water-in-oil emulsion was mixed with the first aqueous solution (21.576 g) and stirred at a speed of 5000 rpm for 2 minutes to form a fourth water-in-oil-in-water emulsion; the fourth water-in-oil-in-water emulsion has a fourth water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the fourth water-in-oil emulsion core is 520 nm.

[0121] The four water-in-oil-in-water emulsions were cooled and mixed to form nanoemulsion 9.

[0122] The zeta potential of nanoemulsion 9 was measured to be -38 V; the mosquito repellent duration was 9 h; the colony growth rate at 6 h was -85%; the colony growth rate at 20 h was 481%; the initial oil separation time was 14 days; the medium oil separation time was 38 days; no severe oil separation was observed.

[0123] Example 10

[0124] After preparing a first crude emulsion from a mixture of eucalyptus oil (8.4 g), peppermint oil (3.6 g), and span 80 (1.2 g) with water (3.6 g), a first water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 170 MPa; after mixing the first water-in-oil emulsion with a first aqueous solution (43.2 g), it was stirred at a speed of 5000 rpm for 2 minutes to form a first water-in-oil-in-water emulsion; the first water-in-oil-in-water emulsion has a first water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the first water-in-oil emulsion core is 120 nm.

[0125] After preparing a second crude emulsion from a mixture of eucalyptus oil (4.2 g), peppermint oil (1.8 g), and span 80 (0.6 g) with water (1.8 g), a second water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 120 MPa; after mixing the second water-in-oil emulsion with a first aqueous solution (21.6 g), it was stirred at a speed of 5000 rpm for 2 minutes to form a second water-in-oil-in-water emulsion; the second water-in-oil-in-water emulsion has a second water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the second water-in-oil emulsion core is 285 nm.

[0126] After preparing a third crude emulsion from a mixture of carvacrol (3 g), thymol (3 g), span 60 (0.9 g), vitamin E (0.3 g), and MCT oil (3 g) with a second aqueous solution (2.7 g), a third water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 105 MPa; after mixing the third water-in-oil emulsion with a first aqueous solution (37.1 g), it was stirred at a speed of 5000 rpm for 2 minutes to form a third water-in-oil-in-water emulsion; the third water-in-oil-in-water emulsion has a third water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the third water-in-oil emulsion core is 255 nm.

[0127] After preparing a fourth crude emulsion from a mixture of carvacrol (3.6 g), thymol (3.6 g), span 60 (1.08 g), vitamin E (0.36 g), ascorbyl palmitate (0.288 g), and MCT oil (3.6 g) with a third aqueous solution (4.32 g), a fourth water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 50 MPa; after mixing the fourth water-in-oil emulsion with a first aqueous solution (43.152 g), it was stirred at a speed of 5000 rpm for 2 minutes to form a fourth water-in-oil-in-water emulsion; the fourth water-in-oil-in-water emulsion has a fourth water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the fourth water-in-oil emulsion core is 520 nm.

[0128] The four water-in-oil-in-water emulsions were cooled and mixed to form nanoemulsion 10.

[0129] The zeta potential of the nanoemulsion 10 was measured to be -38 V; the mosquito repellent duration was 8.5 h; the colony growth rate at 6 h was -79%; the colony growth rate at 20 h was 51%; the initial oil separation time was 12 days; the medium oil separation time was 34 days; and no severe oil separation was observed.

[0130] Example 11

[0131] After preparing a first coarse emulsion from a mixture of eucalyptus oil (8.4 g), peppermint oil (3.6 g), and span 80 (1.2 g) with water (3.6 g), a first water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 170 MPa; after mixing the first water-in-oil emulsion with a first aqueous solution (43.2 g) and stirring at a speed of 5000 rpm for 2 minutes, a first water-in-oil-in-water emulsion was formed; the first water-in-oil-in-water emulsion had a first water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the first water-in-oil emulsion core was 120 nm.

[0132] After preparing a second coarse emulsion from a mixture of eucalyptus oil (2.8 g), peppermint oil (1.2 g), and span 80 (0.4 g) with water (1.2 g), a second water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 120 MPa; after mixing the second water-in-oil emulsion with a first aqueous solution (14.4 g) and stirring at a speed of 5000 rpm for 2 minutes, a second water-in-oil-in-water emulsion was formed; the second water-in-oil-in-water emulsion had a second water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the second water-in-oil emulsion core was 285 nm.

[0133] After preparing a third coarse emulsion from a mixture of carvacrol (3.6 g), thymol (3.6 g), span 60 (1.08 g), vitamin E (0.36 g), and MCT oil (3.6 g) with a second aqueous solution (3.24 g), a third water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 105 MPa; after mixing the third water-in-oil emulsion with a first aqueous solution (44.52 g) and stirring at a speed of 5000 rpm for 2 minutes, a third water-in-oil-in-water emulsion was formed; the third water-in-oil-in-water emulsion had a third water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the third water-in-oil emulsion core was 255 nm.

[0134] After preparing a mixture of carvacrol (3.6 g), thymol (3.6 g), span 60 (1.08 g), vitamin E (0.36 g), ascorbyl palmitate (0.288 g), and MCT oil (3.6 g) with a third aqueous solution (4.32 g) into a fourth crude emulsion, a fourth water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 50 MPa; after mixing the fourth water-in-oil emulsion with a first aqueous solution (43.152 g), it was stirred at a speed of 5000 rpm for 2 minutes to form a fourth water-in-oil-in-water emulsion; the fourth water-in-oil-in-water emulsion has a fourth water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the fourth water-in-oil emulsion core is 520 nm.

[0135] Cool and mix four water-in-oil-in-water emulsions to form nanoemulsion 11.

[0136] Measure the zeta potential of nanoemulsion 11 to be -38 V; the mosquito repellent duration is 7.2 h; the colony growth rate at 6 h is -88%; the colony growth rate at 20 h is 37%; the initial oil separation time is 14 days; the medium oil separation time is 40 days; no severe oil separation is observed.

[0137] Example 12

[0138] After preparing a mixture of eucalyptus oil (8.4 g), peppermint oil (3.6 g), and span 80 (1.2 g) with water (3.6 g) into a first crude emulsion, a first water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 170 MPa; after mixing the first water-in-oil emulsion with a first aqueous solution (43.2 g), it was stirred at a speed of 5000 rpm for 2 minutes to form a first water-in-oil-in-water emulsion; the first water-in-oil-in-water emulsion has a first water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the first water-in-oil emulsion core is 120 nm.

[0139] After preparing a mixture of eucalyptus oil (4.2 g), peppermint oil (1.8 g), and span 80 (0.6 g) with water (1.8 g) into a second crude emulsion, a second water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 120 MPa; after mixing the second water-in-oil emulsion with a first aqueous solution (21.6 g), it was stirred at a speed of 5000 rpm for 2 minutes to form a second water-in-oil-in-water emulsion; the second water-in-oil-in-water emulsion has a second water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the second water-in-oil emulsion core is 285 nm.

[0140] After preparing a third crude emulsion from a mixture of carvacrol (3.6 g), thymol (3.6 g), span 60 (1.08 g), vitamin E (0.36 g), MCT oil (3.6 g) and a second aqueous solution (3.24 g), a third water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 105 MPa; the third water-in-oil emulsion was mixed with a first aqueous solution (44.52 g) and stirred at a speed of 5000 rpm for 2 minutes to form a third water-in-oil-in-water emulsion; the third water-in-oil-in-water emulsion had a third water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the third water-in-oil emulsion core was 255 nm.

[0141] After preparing a fourth crude emulsion from a mixture of carvacrol (3.6 g), thymol (3.6 g), span 60 (1.08 g), vitamin E (0.36 g), ascorbyl palmitate (0.288 g), MCT oil (3.6 g) and a third aqueous solution (4.32 g), a fourth water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 50 MPa; the fourth water-in-oil emulsion was mixed with a first aqueous solution (33.152 g) and stirred at a speed of 5000 rpm for 2 minutes to form a fourth water-in-oil-in-water emulsion; the fourth water-in-oil-in-water emulsion had a fourth water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the fourth water-in-oil emulsion core was 520 nm.

[0142] The four water-in-oil-in-water emulsions were cooled and mixed to form nanoemulsion 12.

[0143] The zeta potential of nanoemulsion 12 was measured to be -39 V; the mosquito repellent duration was 8.4 h; the colony growth rate at 6 h was -85%; the colony growth rate at 20 h was 59%; the initial oil separation time was 12 days; the medium oil separation time was 36 days; no severe oil separation was observed.

[0144] Example 13

[0145] After preparing a first crude emulsion from a mixture of eucalyptus oil (9 g), peppermint oil (4 g), span 80 (1.3 g) and water (3.9 g), a first water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 170 MPa; the first water-in-oil emulsion was mixed with a first aqueous solution (41.8 g) and stirred at a speed of 5000 rpm for 2 minutes to form a first water-in-oil-in-water emulsion; the first water-in-oil-in-water emulsion had a first water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the first water-in-oil emulsion core was 120 nm.

[0146] After preparing a second crude emulsion from a mixture of eucalyptus oil (5 g), peppermint oil (2 g), and span 80 (0.7 g) with water (2.1 g), a second water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 120 MPa. After mixing the second water-in-oil emulsion with the first aqueous solution (20.2 g), it was stirred at a speed of 5000 rpm for 2 minutes to form a second water-in-oil-in-water emulsion. The second water-in-oil-in-water emulsion has a second water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the second water-in-oil emulsion core is 285 nm.

[0147] After preparing a third crude emulsion from a mixture of carvacrol (3.6 g), thymol (3.6 g), span 60 (1.08 g), vitamin E (0.36 g), and MCT oil (3.6 g) with the second aqueous solution (3.24 g), a third water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 105 MPa. After mixing the third water-in-oil emulsion with the first aqueous solution (44.52 g), it was stirred at a speed of 5000 rpm for 2 minutes to form a third water-in-oil-in-water emulsion. The third water-in-oil-in-water emulsion has a third water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the third water-in-oil emulsion core is 255 nm.

[0148] After preparing a fourth crude emulsion from a mixture of carvacrol (2.4 g), thymol (2.4 g), span 60 (0.72 g), vitamin E (0.24 g), ascorbyl palmitate (0.192 g), and MCT oil (2.4 g) with the third aqueous solution (2.88 g), a fourth water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 50 MPa. After mixing the fourth water-in-oil emulsion with the first aqueous solution (38.768 g), it was stirred at a speed of 5000 rpm for 2 minutes to form a fourth water-in-oil-in-water emulsion. The fourth water-in-oil-in-water emulsion has a fourth water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the fourth water-in-oil emulsion core is 520 nm.

[0149] The four water-in-oil-in-water emulsions were cooled and mixed to form nanoemulsion 13.

[0150] The zeta potential of nanoemulsion 13 was measured to be -38 V; the mosquito repellent duration was 8.5 h; the colony growth rate at 6 h was -83%; the colony growth rate at 20 h was 46%; the initial oil separation time was 14 days; the medium oil separation time was 42 days; no severe oil separation was observed.

[0151] Example 14

[0152] After preparing a first coarse emulsion from a mixture of eucalyptus oil (9 g), peppermint oil (3 g), and Span 80 (1.2 g) with water (3.6 g), a first water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 170 MPa; after mixing the first water-in-oil emulsion with a first aqueous solution (43.2 g) and stirring at a speed of 5000 rpm for 2 minutes, a first water-in-oil-in-water emulsion was formed; the first water-in-oil-in-water emulsion has a first water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the first water-in-oil emulsion core is 120 nm.

[0153] After preparing a second coarse emulsion from a mixture of eucalyptus oil (5 g), peppermint oil (3 g), and Span 80 (0.8 g) with water (2.4 g), a second water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 120 MPa; after mixing the second water-in-oil emulsion with a first aqueous solution (18.8 g) and stirring at a speed of 5000 rpm for 2 minutes, a second water-in-oil-in-water emulsion was formed; the second water-in-oil-in-water emulsion has a second water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the second water-in-oil emulsion core is 285 nm.

[0154] After preparing a third coarse emulsion from a mixture of carvacrol (4 g), thymol (3 g), Span 60 (1.05 g), vitamin E (0.35 g), and MCT oil (3.5 g) with a second aqueous solution (3.15 g), a third water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 105 MPa; after mixing the third water-in-oil emulsion with a first aqueous solution (44.95 g) and stirring at a speed of 5000 rpm for 2 minutes, a third water-in-oil-in-water emulsion was formed; the third water-in-oil-in-water emulsion has a third water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the third water-in-oil emulsion core is 255 nm.

[0155] After preparing a fourth coarse emulsion from a mixture of carvacrol (3 g), thymol (2 g), Span 60 (0.75 g), vitamin E (0.25 g), ascorbyl palmitate (0.2 g), and MCT oil (2.5 g) with a third aqueous solution (3 g), a fourth water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 50 MPa; after mixing the fourth water-in-oil emulsion with a first aqueous solution (38.3 g) and stirring at a speed of 5000 rpm for 2 minutes, a fourth water-in-oil-in-water emulsion was formed; the fourth water-in-oil-in-water emulsion has a fourth water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the fourth water-in-oil emulsion core is 520 nm.

[0156] The four water-in-oil-in-water emulsions were cooled and mixed to form nanoemulsion 14.

[0157] The zeta potential of the nanoemulsion 14 was measured to be -38 V; the mosquito repellent duration was 8.5 h; the colony growth rate at 6 h was -85%; the colony growth rate at 20 h was 49%; the initial oil separation time was 14 days; the medium oil separation time was 38 days; and no severe oil separation was observed.

[0158] Example 15

[0159] After preparing a first coarse emulsion from a mixture of eucalyptus oil (8 g), peppermint oil (4 g), and span 80 (1.2 g) with water (3.6 g), a first water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 170 MPa; after mixing the first water-in-oil emulsion with a first aqueous solution (43.2 g) and stirring at a speed of 5000 rpm for 2 minutes, a first water-in-oil-in-water emulsion was formed; the first water-in-oil-in-water emulsion has a first water-in-oil emulsion core, and the D90 particle size of the internal aqueous phase of the first water-in-oil emulsion core is 120 nm.

[0160] After preparing a second coarse emulsion from a mixture of eucalyptus oil (5 g), peppermint oil (3 g), and span 80 (0.8 g) with water (2.4 g), a second water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 120 MPa; after mixing the second water-in-oil emulsion with a first aqueous solution (18.8 g) and stirring at a speed of 5000 rpm for 2 minutes, a second water-in-oil-in-water emulsion was formed; the second water-in-oil-in-water emulsion has a second water-in-oil emulsion core, and the D90 particle size of the internal aqueous phase of the second water-in-oil emulsion core is 285 nm.

[0161] After preparing a third coarse emulsion from a mixture of carvacrol (4 g), thymol (3 g), span 60 (1.05 g), vitamin E (0.35 g), and MCT oil (3.5 g) with a second aqueous solution (3.15 g), a third water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 105 MPa; after mixing the third water-in-oil emulsion with a first aqueous solution (44.95 g) and stirring at a speed of 5000 rpm for 2 minutes, a third water-in-oil-in-water emulsion was formed; the third water-in-oil-in-water emulsion has a third water-in-oil emulsion core, and the D90 particle size of the internal aqueous phase of the third water-in-oil emulsion core is 255 nm.

[0162] After preparing a mixture of carvacrol (3 g), thymol (3 g), span 60 (0.9 g), vitamin E (0.3 g), ascorbyl palmitate (0.24 g), and MCT oil (3 g) with a third aqueous solution (3.6 g) into a fourth crude emulsion, a fourth water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 50 MPa; after mixing the fourth water-in-oil emulsion with a first aqueous solution (35.96 g), it was stirred at a speed of 5000 rpm for 2 minutes to form a fourth water-in-oil-in-water emulsion; the fourth water-in-oil-in-water emulsion has a fourth water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the fourth water-in-oil emulsion core is 520 nm.

[0163] Cool and mix the four water-in-oil-in-water emulsions to form nanoemulsion 15.

[0164] Measure the zeta potential of nanoemulsion 15 to be -40 V; the mosquito repellent duration is 8.4 h; the colony growth rate at 6 h is -84%; the colony growth rate at 20 h is 48%; the initial oil separation time is 12 days; the medium oil separation time is 36 days; no severe oil separation is observed.

[0165] Example 16

[0166] After preparing a mixture of eucalyptus oil (8 g), peppermint oil (3 g), and span 80 (1.1 g) with water (3.3 g) into a first crude emulsion, a first water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 170 MPa; after mixing the first water-in-oil emulsion with a first aqueous solution (44.6 g), it was stirred at a speed of 5000 rpm for 2 minutes to form a first water-in-oil-in-water emulsion; the first water-in-oil-in-water emulsion has a first water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the first water-in-oil emulsion core is 120 nm.

[0167] After preparing a mixture of eucalyptus oil (5 g), peppermint oil (2 g), and span 80 (0.7 g) with water (2.1 g) into a second crude emulsion, a second water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 120 MPa; after mixing the second water-in-oil emulsion with a first aqueous solution (20.2 g), it was stirred at a speed of 5000 rpm for 2 minutes to form a second water-in-oil-in-water emulsion; the second water-in-oil-in-water emulsion has a second water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the second water-in-oil emulsion core is 285 nm.

[0168] After preparing a third crude emulsion from a mixture of carvacrol (3 g), thymol (4 g), span 60 (1.05 g), vitamin E (0.35 g), and MCT oil (3.5 g) and a second aqueous solution (3.15 g), a third water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 105 MPa. After mixing the third water-in-oil emulsion with a first aqueous solution (44.95 g), it was stirred at a speed of 5000 rpm for 2 minutes to form a third water-in-oil-in-water emulsion. The third water-in-oil-in-water emulsion has a third water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the third water-in-oil emulsion core is 255 nm.

[0169] After preparing a fourth crude emulsion from a mixture of carvacrol (3 g), thymol (3 g), span 60 (0.9 g), vitamin E (0.3 g), ascorbyl palmitate (0.24 g), and MCT oil (3 g) and a third aqueous solution (3.6 g), a fourth water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 50 MPa. After mixing the fourth water-in-oil emulsion with a first aqueous solution (35.96 g), it was stirred at a speed of 5000 rpm for 2 minutes to form a fourth water-in-oil-in-water emulsion. The fourth water-in-oil-in-water emulsion has a fourth water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the fourth water-in-oil emulsion core is 520 nm.

[0170] The four water-in-oil-in-water emulsions were cooled and mixed to form nanoemulsion 16.

[0171] The zeta potential of nanoemulsion 16 was measured to be -38 V; the mosquito repellent duration was 8.6 h; the colony growth rate at 6 h was -83%; the colony growth rate at 20 h was 59%; the initial oil separation time was 14 days; the medium oil separation time was 40 days; and no severe oil separation was observed.

[0172] Example 17

[0173] After preparing a first crude emulsion from a mixture of eucalyptus oil (9 g), peppermint oil (4 g), span 80 (1.3 g), and water (3.9 g), a first water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 150 MPa. After mixing the first water-in-oil emulsion with a first aqueous solution (41.8 g), it was stirred at a speed of 5000 rpm for 2 minutes to form a first water-in-oil-in-water emulsion. The first water-in-oil-in-water emulsion has a first water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the first water-in-oil emulsion core is 100 nm.

[0174] After preparing a second coarse emulsion from a mixture of eucalyptus oil (5 g), peppermint oil (2 g), and span 80 (0.7 g) with water (2.1 g), a second water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 120 MPa. After mixing the second water-in-oil emulsion with the first aqueous solution (20.2 g), it was stirred at a speed of 5000 rpm for 2 minutes to form a second water-in-oil-in-water emulsion. The second water-in-oil-in-water emulsion has a second water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the second water-in-oil emulsion core is 285 nm.

[0175] After preparing a third coarse emulsion from a mixture of carvacrol (3.6 g), thymol (3.6 g), span 60 (1.08 g), vitamin E (0.36 g), and MCT oil (3.6 g) with the second aqueous solution (3.24 g), a third water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 105 MPa. After mixing the third water-in-oil emulsion with the first aqueous solution (44.52 g), it was stirred at a speed of 5000 rpm for 2 minutes to form a third water-in-oil-in-water emulsion. The third water-in-oil-in-water emulsion has a third water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the third water-in-oil emulsion core is 255 nm.

[0176] After preparing a fourth coarse emulsion from a mixture of carvacrol (2.4 g), thymol (2.4 g), span 60 (0.72 g), vitamin E (0.24 g), ascorbyl palmitate (0.192 g), and MCT oil (2.4 g) with the third aqueous solution (2.88 g), a fourth water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 50 MPa. After mixing the fourth water-in-oil emulsion with the first aqueous solution (38.768 g), it was stirred at a speed of 5000 rpm for 2 minutes to form a fourth water-in-oil-in-water emulsion. The fourth water-in-oil-in-water emulsion has a fourth water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the fourth water-in-oil emulsion core is 520 nm.

[0177] The four water-in-oil-in-water emulsions were cooled and mixed to form nanoemulsion 17.

[0178] The zeta potential of nanoemulsion 17 was measured to be -38 V; the mosquito repellent duration was 8.2 h; the colony growth rate at 6 h was -85%; the colony growth rate at 20 h was 72%; the initial oil separation time was 14 days; the medium oil separation time was 42 days; and no severe oil separation was observed.

[0179] Example 18

[0180] After preparing a first crude emulsion from a mixture of eucalyptus oil (9 g), peppermint oil (4 g), and Span 80 (1.3 g) with water (3.9 g), a first water-in-oil emulsion is prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 200 MPa; after mixing the first water-in-oil emulsion with a first aqueous solution (41.8 g) and stirring at a speed of 5000 rpm for 2 minutes, a first water-in-oil-in-water emulsion is formed; the first water-in-oil-in-water emulsion has a first water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the first water-in-oil emulsion core is 150 nm.

[0181] After preparing a second crude emulsion from a mixture of eucalyptus oil (5 g), peppermint oil (2 g), and Span 80 (0.7 g) with water (2.1 g), a second water-in-oil emulsion is prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 120 MPa; after mixing the second water-in-oil emulsion with a first aqueous solution (20.2 g) and stirring at a speed of 5000 rpm for 2 minutes, a second water-in-oil-in-water emulsion is formed; the second water-in-oil-in-water emulsion has a second water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the second water-in-oil emulsion core is 285 nm.

[0182] After preparing a third crude emulsion from a mixture of carvacrol (3.6 g), thymol (3.6 g), Span 60 (1.08 g), vitamin E (0.36 g), and MCT oil (3.6 g) with a second aqueous solution (3.24 g), a third water-in-oil emulsion is prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 105 MPa; after mixing the third water-in-oil emulsion with a first aqueous solution (44.52 g) and stirring at a speed of 5000 rpm for 2 minutes, a third water-in-oil-in-water emulsion is formed; the third water-in-oil-in-water emulsion has a third water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the third water-in-oil emulsion core is 255 nm.

[0183] After preparing a fourth crude emulsion from a mixture of carvacrol (2.4 g), thymol (2.4 g), Span 60 (0.72 g), vitamin E (0.24 g), ascorbyl palmitate (0.192 g), and MCT oil (2.4 g) with a third aqueous solution (2.88 g), a fourth water-in-oil emulsion is prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 50 MPa; after mixing the fourth water-in-oil emulsion with a first aqueous solution (38.768 g) and stirring at a speed of 5000 rpm for 2 minutes, a fourth water-in-oil-in-water emulsion is formed; the fourth water-in-oil-in-water emulsion has a fourth water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the fourth water-in-oil emulsion core is 520 nm.

[0184] Cool and mix the four water-in-oil-in-water emulsions to form nanoemulsion 18.

[0185] The zeta potential of the nanoemulsion 18 was measured to be -37 V; the mosquito repellent duration was 8.6 h; the colony growth rate at 6 h was -85%; the colony growth rate at 20 h was 59%; the initial oil separation time was 12 days; the medium oil separation time was 38 days; no severe oil separation was observed.

[0186] Example 19

[0187] After preparing a first coarse emulsion from a mixture of eucalyptus oil (9 g), peppermint oil (4 g), and span 80 (1.3 g) with water (3.9 g), a first water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 150 MPa; after mixing the first water-in-oil emulsion with a first aqueous solution (41.8 g) and stirring at a speed of 5000 rpm for 2 minutes, a first water-in-oil-in-water emulsion was formed; the first water-in-oil-in-water emulsion had a first water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the first water-in-oil emulsion core was 100 nm.

[0188] After preparing a second coarse emulsion from a mixture of eucalyptus oil (5 g), peppermint oil (2 g), and span 80 (0.7 g) with water (2.1 g), a second water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 100 MPa; after mixing the second water-in-oil emulsion with a first aqueous solution (20.2 g) and stirring at a speed of 5000 rpm for 2 minutes, a second water-in-oil-in-water emulsion was formed; the second water-in-oil-in-water emulsion had a second water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the second water-in-oil emulsion core was 250 nm.

[0189] After preparing a third coarse emulsion from a mixture of carvacrol (3.6 g), thymol (3.6 g), span 60 (1.08 g), vitamin E (0.36 g), and MCT oil (3.6 g) with a second aqueous solution (3.24 g), a third water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 105 MPa; after mixing the third water-in-oil emulsion with a first aqueous solution (44.52 g) and stirring at a speed of 5000 rpm for 2 minutes, a third water-in-oil-in-water emulsion was formed; the third water-in-oil-in-water emulsion had a third water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the third water-in-oil emulsion core was 255 nm.

[0190] After preparing a mixture of carvacrol (2.4 g), thymol (2.4 g), span 60 (0.72 g), vitamin E (0.24 g), ascorbyl palmitate (0.192 g), and MCT oil (2.4 g) with the third aqueous solution (2.88 g) into a fourth crude emulsion, a fourth water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 50 MPa; after mixing the fourth water-in-oil emulsion with the first aqueous solution (38.768 g), it was stirred at a speed of 5000 rpm for 2 minutes to form a fourth water-in-oil-in-water emulsion; the fourth water-in-oil-in-water emulsion has a fourth water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the fourth water-in-oil emulsion core is 520 nm.

[0191] Cool and mix the four water-in-oil-in-water emulsions to form nanoemulsion 19.

[0192] Measure the zeta potential of nanoemulsion 19 to be -38 V; the mosquito repellent duration is 8.2 h; the colony growth rate at 6 h is -84%; the colony growth rate at 20 h is 53%; the initial oil separation time is 14 days; the medium oil separation time is 32 days; no severe oil separation is observed.

[0193] Example 20

[0194] After preparing a mixture of eucalyptus oil (9 g), peppermint oil (4 g), and span 80 (1.3 g) with water (3.9 g) into a first crude emulsion, a first water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 170 MPa; after mixing the first water-in-oil emulsion with the first aqueous solution (41.8 g), it was stirred at a speed of 5000 rpm for 2 minutes to form a first water-in-oil-in-water emulsion; the first water-in-oil-in-water emulsion has a first water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the first water-in-oil emulsion core is 120 nm.

[0195] After preparing a mixture of eucalyptus oil (5 g), peppermint oil (2 g), and span 80 (0.7 g) with water (2.1 g) into a second crude emulsion, a second water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 120 MPa; after mixing the second water-in-oil emulsion with the first aqueous solution (20.2 g), it was stirred at a speed of 5000 rpm for 2 minutes to form a second water-in-oil-in-water emulsion; the second water-in-oil-in-water emulsion has a second water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the second water-in-oil emulsion core is 285 nm.

[0196] After preparing a third crude emulsion from a mixture of carvacrol (3.6 g), thymol (3.6 g), span 60 (1.08 g), vitamin E (0.36 g), and MCT oil (3.6 g) and a second aqueous solution (3.24 g), a third water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 90 MPa; the third water-in-oil emulsion was mixed with a first aqueous solution (44.52 g) and stirred at a speed of 5000 rpm for 2 minutes to form a third water-in-oil-in-water emulsion; the third water-in-oil-in-water emulsion has a third water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the third water-in-oil emulsion core is 210 nm.

[0197] After preparing a fourth crude emulsion from a mixture of carvacrol (2.4 g), thymol (2.4 g), span 60 (0.72 g), vitamin E (0.24 g), ascorbyl palmitate (0.192 g), and MCT oil (2.4 g) and a third aqueous solution (2.88 g), a fourth water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 50 MPa; the fourth water-in-oil emulsion was mixed with a first aqueous solution (38.768 g) and stirred at a speed of 5000 rpm for 2 minutes to form a fourth water-in-oil-in-water emulsion; the fourth water-in-oil-in-water emulsion has a fourth water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the fourth water-in-oil emulsion core is 520 nm.

[0198] The four water-in-oil-in-water emulsions were cooled and mixed to form nanoemulsion 20.

[0199] The zeta potential of nanoemulsion 20 was measured to be -38 V; the mosquito repellent duration was 8.1 h; the colony growth rate at 6 h was -86%; the colony growth rate at 20 h was 68%; the initial oil separation time was 12 days; the medium oil separation time was 36 days; no severe oil separation was observed.

[0200] Example 21

[0201] After preparing a first crude emulsion from a mixture of eucalyptus oil (9 g), peppermint oil (4 g), span 80 (1.3 g), and water (3.9 g), a first water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 170 MPa; the first water-in-oil emulsion was mixed with a first aqueous solution (41.8 g) and stirred at a speed of 5000 rpm for 2 minutes to form a first water-in-oil-in-water emulsion; the first water-in-oil-in-water emulsion has a first water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the first water-in-oil emulsion core is 120 nm.

[0202] After preparing a second coarse emulsion from a mixture of eucalyptus oil (5 g), peppermint oil (2 g), and span 80 (0.7 g) with water (2.1 g), a second water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 120 MPa. After mixing the second water-in-oil emulsion with the first aqueous solution (20.2 g), it was stirred at a speed of 5000 rpm for 2 minutes to form a second water-in-oil-in-water emulsion. The second water-in-oil-in-water emulsion has a second water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the second water-in-oil emulsion core is 285 nm.

[0203] After preparing a third coarse emulsion from a mixture of carvacrol (3.6 g), thymol (3.6 g), span 60 (1.08 g), vitamin E (0.36 g), and MCT oil (3.6 g) with the second aqueous solution (3.24 g), a third water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 90 MPa. After mixing the third water-in-oil emulsion with the first aqueous solution (44.52 g), it was stirred at a speed of 5000 rpm for 2 minutes to form a third water-in-oil-in-water emulsion. The third water-in-oil-in-water emulsion has a third water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the third water-in-oil emulsion core is 210 nm.

[0204] After preparing a fourth coarse emulsion from a mixture of carvacrol (2.4 g), thymol (2.4 g), span 60 (0.72 g), vitamin E (0.24 g), ascorbyl palmitate (0.192 g), and MCT oil (2.4 g) with the third aqueous solution (2.88 g), a fourth water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 70 MPa. After mixing the fourth water-in-oil emulsion with the first aqueous solution (38.768 g), it was stirred at a speed of 5000 rpm for 2 minutes to form a fourth water-in-oil-in-water emulsion. The fourth water-in-oil-in-water emulsion has a fourth water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the fourth water-in-oil emulsion core is 700 nm.

[0205] The four water-in-oil-in-water emulsions were cooled and mixed to form nanoemulsion 21.

[0206] The zeta potential of nanoemulsion 21 was measured to be -38 V; the mosquito repellent duration was 8.3 h; the colony growth rate at 6 h was -84%; the colony growth rate at 20 h was 51%; the initial oil separation time was 9 days; the medium oil separation time was 28 days; the severe oil separation time was 48 days.

[0207] Example 22

[0208] After preparing a first crude emulsion from a mixture of eucalyptus oil (9 g), peppermint oil (4 g), and span 80 (1.3 g) with water (3.9 g), a first water-in-oil emulsion is prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 170 MPa; after mixing the first water-in-oil emulsion with a first aqueous solution (41.8 g) and stirring at a speed of 5000 rpm for 2 minutes, a first water-in-oil-in-water emulsion is formed; the first water-in-oil-in-water emulsion has a first water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the first water-in-oil emulsion core is 120 nm.

[0209] After preparing a second crude emulsion from a mixture of eucalyptus oil (5 g), peppermint oil (2 g), and span 80 (0.7 g) with water (2.1 g), a second water-in-oil emulsion is prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 120 MPa; after mixing the second water-in-oil emulsion with a first aqueous solution (20.2 g) and stirring at a speed of 5000 rpm for 2 minutes, a second water-in-oil-in-water emulsion is formed; the second water-in-oil-in-water emulsion has a second water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the second water-in-oil emulsion core is 285 nm.

[0210] After preparing a third crude emulsion from a mixture of carvacrol (3.6 g), thymol (3.6 g), span 60 (1.08 g), vitamin E (0.36 g), and MCT oil (3.6 g) with a second aqueous solution (3.24 g), a third water-in-oil emulsion is prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 130 MPa; after mixing the third water-in-oil emulsion with a first aqueous solution (44.52 g) and stirring at a speed of 5000 rpm for 2 minutes, a third water-in-oil-in-water emulsion is formed; the third water-in-oil-in-water emulsion has a third water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the third water-in-oil emulsion core is 300 nm.

[0211] After preparing a fourth crude emulsion from a mixture of carvacrol (2.4 g), thymol (2.4 g), span 60 (0.72 g), vitamin E (0.24 g), ascorbyl palmitate (0.192 g), and MCT oil (2.4 g) with a third aqueous solution (2.88 g), a fourth water-in-oil emulsion is prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 65 MPa; after mixing the fourth water-in-oil emulsion with a first aqueous solution (38.768 g) and stirring at a speed of 5000 rpm for 2 minutes, a fourth water-in-oil-in-water emulsion is formed; the fourth water-in-oil-in-water emulsion has a fourth water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the fourth water-in-oil emulsion core is 640 nm.

[0212] Cool and mix the four water-in-oil-in-water emulsions to form nanoemulsion 22.

[0213] The zeta potential of the nanoemulsion 22 was measured to be -38 V; the mosquito repellent duration was 8.4 h; the colony growth rate at 6 h was -83%; the colony growth rate at 20 h was 63%; the initial oil separation time was 8 days; the medium oil separation time was 28 days; the severe oil separation time was 42 days.

[0214] Experimental Example 23

[0215] After preparing a first crude emulsion from a mixture of eucalyptus oil (14 g), peppermint oil (6 g), span 80 (2 g) and water (6 g), a first water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 170 MPa; after mixing the first water-in-oil emulsion with a first aqueous solution (72 g), it was stirred at a speed of 5000 rpm for 2 minutes to form a first water-in-oil-in-water emulsion; the first water-in-oil-in-water emulsion had a first water-in-oil emulsification core, and the D90 particle size of the inner aqueous phase of the first water-in-oil emulsification core was 120 nm.

[0216] After preparing a second crude emulsion from a mixture of eucalyptus oil (0 g), peppermint oil (0 g), span 80 (0 g) and water (0 g), a second water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at MPa pressure; after mixing the second water-in-oil emulsion with a first aqueous solution (0 g), it was stirred at a speed of 5000 rpm for 2 minutes to form a second water-in-oil-in-water emulsion; the second water-in-oil-in-water emulsion had a second water-in-oil emulsification core, and the D90 particle size of the inner aqueous phase of the second water-in-oil emulsification core was between nm.

[0217] After preparing a third crude emulsion from a mixture of carvacrol (6 g), thymol (6 g), span 60 (1.8 g), vitamin E (0.6 g), MCT oil (6 g) and a second aqueous solution (5.4 g), a third water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 105 MPa; after mixing the third water-in-oil emulsion with a first aqueous solution (74.2 g), it was stirred at a speed of 5000 rpm for 2 minutes to form a third water-in-oil-in-water emulsion; the third water-in-oil-in-water emulsion had a third water-in-oil emulsification core, and the D90 particle size of the inner aqueous phase of the third water-in-oil emulsification core was between 255 nm.

[0218] After preparing a mixture of carvacrol (0 g), thymol (0 g), span 60 (0 g), vitamin E (0 g), ascorbyl palmitate (0 g), and MCT oil (0 g) with a third aqueous solution (0 g) into a fourth crude emulsion, a fourth water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of MPa; after mixing the fourth water-in-oil emulsion with a first aqueous solution (0 g), it was stirred at a speed of 5000 rpm for 2 minutes to form a fourth water-in-oil-in-water emulsion; the fourth water-in-oil-in-water emulsion has a fourth water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the fourth water-in-oil emulsion core is between nm.

[0219] Cool and mix the four water-in-oil-in-water emulsions to form nanoemulsion 23.

[0220] Measure the zeta potential of nanoemulsion 23 to be -38 V; the mosquito repellent duration is 4.5 h; the colony growth rate at 6 h is -82%; the colony growth rate at 15 h is 782%; the initial oil separation time is 12 days; the medium oil separation time is 46 days; no severe emulsification is observed.

[0221] Experimental Example 24

[0222] After preparing a mixture of eucalyptus oil (14 g), peppermint oil (6 g), and span 80 (2 g) with water (6 g) into a first crude emulsion, a first water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 200 MPa; after mixing the first water-in-oil emulsion with a first aqueous solution (72 g), it was stirred at a speed of 5000 rpm for 2 minutes to form a first water-in-oil-in-water emulsion; the first water-in-oil-in-water emulsion has a first water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the first water-in-oil emulsion core is 150 nm.

[0223] After preparing a mixture of carvacrol (6 g), thymol (8 g), span 60 (2.1 g), vitamin E (0.7 g), and MCT oil (7 g) with a second aqueous solution (6.3 g) into a third crude emulsion, a third water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 130 MPa; after mixing the third water-in-oil emulsion with a first aqueous solution (69.9 g), it was stirred at a speed of 5000 rpm for 2 minutes to form a third water-in-oil-in-water emulsion; the third water-in-oil-in-water emulsion has a third water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the third water-in-oil emulsion core is 300 nm.

[0224] Cool and mix the above two water-in-oil-in-water emulsions to form nanoemulsion 24.

[0225] The zeta potential of nanoemulsion 24 was measured to be -38 V; the mosquito repellent duration was 5.2 h; the colony growth rate at 6 h was -83%; the colony growth rate at 20 h was 904%; the initial oil separation time was 18 days; the medium oil separation time was 42 days; no severe emulsification was observed.

[0226] Experimental Example 25

[0227] After preparing a first crude emulsion from a mixture of eucalyptus oil (12 g), peppermint oil (8 g), span 80 (2 g) and water (6 g), a first water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 200 MPa; after mixing the first water-in-oil emulsion with a first aqueous solution (72 g), it was stirred at a speed of 5000 rpm for 2 minutes to form a first water-in-oil-in-water emulsion; the first water-in-oil-in-water emulsion had a first water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the first water-in-oil emulsion core was 150 nm.

[0228] After preparing a fourth crude emulsion from a mixture of carvacrol (8 g), thymol (6 g), span 60 (2.1 g), vitamin E (0.7 g), ascorbyl palmitate (0.56 g), MCT oil (7 g) and a third aqueous solution (8.4 g), a fourth water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 50 MPa; after mixing the fourth water-in-oil emulsion with a first aqueous solution (67.24 g), it was stirred at a speed of 5000 rpm for 2 minutes to form a fourth water-in-oil-in-water emulsion; the fourth water-in-oil-in-water emulsion had a fourth water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the fourth water-in-oil emulsion core was between 5 and 20 nm.

[0229] After cooling the first water-in-oil-in-water emulsion, the second water-in-oil-in-water emulsion, the third water-in-oil-in-water emulsion and the fourth water-in-oil-in-water emulsion to 10 °C to 15 °C, they were mixed and stirred at a speed of 2500 rpm for 2 minutes to form nanoemulsion 36.

[0230] The zeta potential of nanoemulsion 25 was measured to be -37 V; the mosquito repellent duration was 5.3 h; the colony growth rate at 6 h was -74%; the colony growth rate at 20 h was 139%; the initial oil separation time was 8 days; the medium oil separation time was 24 days; the severe oil separation time was 36 days.

[0231] Experimental Example 26

[0232] After preparing a first crude emulsion from a mixture of eucalyptus oil (14 g), peppermint oil (8 g), and span 80 (2.2 g) with water (6.6 g), a first water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 200 MPa; after mixing the first water-in-oil emulsion with a first aqueous solution (69.2 g), it was stirred at a speed of 5000 rpm for 2 minutes to form a first water-in-oil-in-water emulsion; the first water-in-oil-in-water emulsion has a first water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the first water-in-oil emulsion core is 150 nm.

[0233] After preparing a fourth crude emulsion from a mixture of carvacrol (8 g), thymol (8 g), span 60 (2.4 g), vitamin E (0.8 g), ascorbyl palmitate (0.64 g), and MCT oil (8 g) with a third aqueous solution (9.6 g), a fourth water-in-oil emulsion was prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 70 MPa; after mixing the fourth water-in-oil emulsion with a first aqueous solution (62.56 g), it was stirred at a speed of 5000 rpm for 2 minutes to form a fourth water-in-oil-in-water emulsion; the fourth water-in-oil-in-water emulsion has a fourth water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the fourth water-in-oil emulsion core is between 700 nm.

[0234] The above two water-in-oil-in-water emulsions were cooled and mixed to form nanoemulsion 26.

[0235] The zeta potential of nanoemulsion 26 was measured to be -38 V; the mosquito repellent duration was 4.8 h; the colony growth rate at 6 h was -76%; the colony growth rate at 20 h was 157%; the initial oil separation time was 6 days; the medium oil separation time was 20 days; the severe oil separation time was 32 days.

[0236] Experimental Example 27

[0237] The preparation process of Example 27 was basically the same as that of Example 1, except that in Example 27, the oil phase of the fourth water-in-oil emulsion core did not contain ascorbyl palmitate. The nanoemulsion prepared in Example 27 was labeled as nanoemulsion 27.

[0238] The zeta potential of nanoemulsion 27 was measured to be -37 V; the mosquito repellent duration was 8.6 h; the colony growth rate at 6 h was -62%; the colony growth rate at 20 h was 635%; the initial oil separation time was 14 days; the medium oil separation time was 44 days; no severe emulsification was observed.

[0239] Experimental Example 28

[0240] The preparation process of Example 28 was basically the same as that of Example 1, except that in Example 28, the inner aqueous phase of the fourth water-in-oil emulsion core did not contain hydroxypropyl-β-cyclodextrin. The nanoemulsion prepared in Example 28 was labeled as nanoemulsion 28.

[0241] The zeta potential of the nanoemulsion 28 was measured to be -38 V; the mosquito repellent duration was 8.3 h; the colony growth rate at 6 h was -90%; the colony growth rate at 20 h was 791%; the initial oil separation time was 12 days; the medium oil separation time was 38 days; no severe emulsification was observed.

[0242] The test results of each nanoemulsion are summarized in Table 1 below.

[0243] Table 1: Test Results of Nanoemulsions

[0244]

[0245]

[0246] Among them, "-" indicates that no severe emulsification was observed until the 50th day.

[0247] Nanoemulsions 1 to 22 are applicable nanoemulsions provided by the embodiments of the present invention, and most of them can have a long mosquito repellent duration, a long bacteriostatic and bactericidal duration, and high stability.

[0248] It can be seen from nanoemulsions 1 to 22 that the adjustment of the ratio of small-sized emulsification nuclei to large-sized emulsification nuclei will affect the duration of the effect of the nanoemulsion. For example, in nanoemulsions 2 and 3, the proportion of the first water-in-oil emulsification nuclei is lower than that of other nanoemulsions, and the proportion of the second water-in-oil emulsification nuclei is higher than that of other nanoemulsions. The mosquito repellent duration of these nanoemulsions 2 and 3 is insufficient. This may be related to the fact that nanoemulsions 2 and 3 cannot quickly form a high-concentration volatile essential oil to quickly repel mosquitoes at the initial stage of spray application, resulting in insufficient mosquito repellent rate in a short time after application (individual mosquitoes stay on the treated gauze). For another example, in nanoemulsion 7, the proportion of the third water-in-oil emulsification nuclei is lower than that of other nanoemulsions, and the proportion of the fourth water-in-oil emulsification nuclei is higher than that of other nanoemulsions. It can be seen from the test results that the bacteriostatic / bactericidal effect of nanoemulsion 7 within 6 hours is not prominent, but the bacteriostatic / bactericidal effect within 18 hours is relatively prominent, showing an obvious long-acting bacteriostatic characteristic.

[0249] It can be seen from nanoemulsions 1 to 22 that the adjustment of the ratio of small-sized emulsification nuclei to large-sized emulsification nuclei, as well as the adjustment of the particle size of each emulsification nucleus, has a certain impact on the stability of the nanoemulsion. Generally, the more small-sized emulsification nuclei and the smaller the small-sized emulsification nuclei, the higher the stability of the nanoemulsion. For example, in nanoemulsions 21 and 22, the particle size of the emulsification nuclei is relatively large, which leads to a decrease in their stability.

[0250] Experimental Examples 23 to 26 are exploratory experimental examples of the present invention with only two emulsifying nuclei set in the R & D stage. It can be seen from Experimental Examples 23 to 26 that the long-lasting mosquito repellent effects of these nanoemulsions are not ideal, and there is an obvious conflict between the goals of long-lasting bactericidal / bacteriostatic and long-term stability.

[0251] Experimental Example 27 is a typical example among multiple experimental examples of the present invention where ascorbyl palmitate is not added to the oil phase of the fourth water-in-oil emulsion in the R & D stage. It can be seen from Table 1 that when ascorbyl palmitate is not added to the oil phase, the colony growth at 6 hours is -62%, and the colony growth rate reaches 635% at 20 hours. This indicates that the bactericidal and disinfecting effect of nanoemulsion 27 is initially weak, and the continuous bactericidal time and continuous inhibition of bacteria are also weak. This may be because ascorbyl palmitate and vitamin E can cooperate more effectively to provide antioxidant protection for carvacrol and thymol, reducing the oxidative loss of carvacrol and thymol during the long-term slow release process.

[0252] Experimental Example 28 is a typical example among multiple experimental examples of the present invention where hydroxypropyl-β-cyclodextrin is not added to the inner aqueous phase of the fourth water-in-oil emulsion in the R & D stage. It can be seen from Table 1 that when hydroxypropyl-β-cyclodextrin is not added to the inner aqueous phase, the colony growth rate at 6 hours is as low as -90%, indicating that about 90% of the bacteria have lost their activity, showing a very good rapid bactericidal and disinfecting effect. However, the colony growth rate reaches 791% at 20 hours, indicating that this nanoemulsion does not inhibit bacteria for a long time. And hydroxypropyl-β-cyclodextrin is contained in the aqueous phase of the fourth water-in-oil emulsifying nucleus of nanoemulsions 1 to 22, and their colony growth rates at 20 hours are all lower than that of Experimental Example 28. This may be because if hydroxypropyl-β-cyclodextrin is added to the inner aqueous phase, carvacrol and thymol can be partially combined with hydroxypropyl-β-cyclodextrin and protected by it; after the water phase in the nanoemulsion droplets volatilizes, carvacrol and thymol in the oil phase play a role in rapidly inhibiting and killing bacteria. The carvacrol and thymol combined with hydroxypropyl-β-cyclodextrin are slowly released under the wrapping of sodium alginate, so that a low concentration of carvacrol and thymol can be continuously maintained around the bacteria, and this low concentration of carvacrol and thymol may have a certain inhibitory or interfering effect on the proliferation of bacteria, thereby slowing down the rapid rebound of bacteria after the bactericidal and disinfecting effect fails.

[0253] Those skilled in the art will readily think of other embodiments of the present invention after considering the specification and practicing the invention herein. This application is intended to cover any variations, uses, or adaptations of the present invention, which follow the general principles of the present invention and include the common general knowledge or conventional technical means in the technical field not invented by the present invention. The specification and examples are only regarded as exemplary, and the true scope and spirit of the present invention are pointed out by the appended claims.

Claims

1. A nanoemulsion for preventing and treating porcine respiratory diseases, characterized in that, It includes an external aqueous phase and the first water-in-oil emulsion core, the second water-in-oil emulsion core, the third water-in-oil emulsion core, and the fourth water-in-oil emulsion core dispersed in the external aqueous phase; Among them, the oil phase of the first water-in-oil emulsion core and the second water-in-oil emulsion core is a mixture of eucalyptus oil, peppermint oil, and span 80, and the internal aqueous phase of the first water-in-oil emulsion core and the second water-in-oil emulsion core is water; the D90 particle size of the internal aqueous phase of the first water-in-oil emulsion core is between 100 nm and 150 nm; the D90 particle size of the internal aqueous phase of the second water-in-oil emulsion core is between 250 nm and 300 nm; The oil phase of the third water-in-oil emulsion core is a mixture of carvacrol, thymol, span 60, vitamin E, and MCT oil, and the internal aqueous phase of the third water-in-oil emulsion core is an aqueous sodium alginate solution; the D90 particle size of the internal aqueous phase of the third water-in-oil emulsion core is 200 - 300 nm; The oil phase of the fourth water-in-oil emulsion core is a mixture of carvacrol, thymol, span 60, vitamin E, ascorbyl palmitate, and MCT oil, and the internal aqueous phase of the fourth water-in-oil emulsion core is an aqueous solution of sodium alginate and hydroxypropyl-β-cyclodextrin; the D90 particle size of the internal aqueous phase of the fourth water-in-oil emulsion core is 500 - 700 nm; The external aqueous phase is an aqueous solution of tween 80, xanthan gum, sodium caseinate, and sodium benzoate.

2. The nanoemulsion for preventing and treating swine respiratory diseases according to claim 1, characterized in that, The mass ratio of the first water-in-oil emulsion core to the second water-in-oil emulsion core is in the range of 7:3 to 3:7; The mass ratio of the third water-in-oil emulsion core to the fourth water-in-oil emulsion core is in the range of 7:3 to 3:

7.

3. The nanoemulsion for preventing and treating porcine respiratory diseases according to claim 1, wherein The mass ratio of the first water-in-oil emulsion core to the second water-in-oil emulsion core is 2:1; The mass ratio of the third water-in-oil emulsion core to the fourth water-in-oil emulsion core is 6:

5.

4. The nanoemulsion for preventing and treating porcine respiratory diseases according to claim 1, wherein The mass of eucalyptus oil in the first water-in-oil emulsion core is 2.8% - 4.5% of the total mass of the nanoemulsion; The mass of eucalyptus oil in the second water-in-oil emulsion core is 2.5% - 4.1% of the total mass of the nanoemulsion; The mass of peppermint oil in the first water-in-oil emulsion core is 1.2% - 2.1% of the total mass of the nanoemulsion; The mass of peppermint oil in the second water-in-oil emulsion core is 0.9% - 1.8% of the total mass of the nanoemulsion.

5. The nanoemulsion for preventing and treating swine respiratory diseases according to claim 1, wherein The mass of carvacrol in the third water-in-oil emulsion core is 1.5% - 2.1% of the total mass of the nanoemulsion; The mass of thymol in the third water-in-oil emulsion core is 1.5% - 2.1% of the total mass of the nanoemulsion; The mass of carvacrol in the fourth water-in-oil emulsion core is 0.9% - 1.8% of the total mass of the nanoemulsion; The mass of thymol in the fourth water-in-oil emulsion core is 0.9% - 1.8% of the total mass of the nanoemulsion.

6. The nanoemulsion for preventing and treating porcine respiratory diseases according to any one of claims 1 to 5, characterized in that, The D90 particle size of the internal aqueous phase of the first water-in-oil emulsion core is between 115 nm and 125 nm; The D90 particle size of the internal aqueous phase of the second water-in-oil emulsion core is between 280 nm and 290 nm; The D90 particle size of the internal aqueous phase of the third water-in-oil emulsion core is between 250 nm and 260 nm; The D90 particle size of the internal aqueous phase of the fourth water-in-oil emulsion core is between 500 nm and 550 nm.

7. The nanoemulsion for preventing and treating swine respiratory diseases according to any one of claims 1 to 5, characterized in that, In the third water-in-oil emulsion core, the mass of vitamin E is 3% - 7% of the total mass of carvacrol and thymol; In the fourth water-in-oil emulsion core, the mass of vitamin E is 3% - 7% of the total mass of carvacrol and thymol, and the mass of ascorbyl palmitate is 3% - 5% of the total mass of carvacrol and thymol.

8. The nanoemulsion for preventing and treating porcine respiratory diseases according to any one of claims 1 to 5, characterized in that, In the fourth water-in-oil emulsion core, the mass content of hydroxypropyl-β-cyclodextrin in the internal aqueous phase is 0.1% - 0.3%.

9. The nanoemulsion for preventing and treating porcine respiratory diseases according to any one of claims 1 to 5, characterized in that, In the nanoemulsion, the mass content of eucalyptus oil is 6.3% - 7.5%, the mass content of peppermint oil is 2.7% - 3.5%, the mass content of carvacrol is 2.5% - 3.6%, and the mass content of thymol is 2.5% - 3.6%.

10. The preparation method of the nanoemulsion for preventing and treating porcine respiratory diseases according to any one of claims 1 to 9, characterized in that, Comprising: Dissolve Tween 80, xanthan gum, sodium caseinate, and sodium benzoate in water to form a first aqueous solution; After preparing a first coarse emulsion from a mixture of eucalyptus oil, peppermint oil, and Span 80 and water, prepare a first water-in-oil emulsion by high-pressure microfluidic nanoemulsification technology at a pressure of 150 MPa - 200 MPa; after mixing the first water-in-oil emulsion with a part of the first aqueous solution, stir at a speed of 4000 rpm - 6000 rpm for 1 - 3 minutes to form a first water-in-oil-in-water emulsion; the first water-in-oil-in-water emulsion has a first water-in-oil emulsion core, and the D90 particle size of the internal aqueous phase of the first water-in-oil emulsion core is between 100 nm and 150 nm; After preparing a second coarse emulsion from a mixture of eucalyptus oil, peppermint oil, and Span 80 and water, prepare a second water-in-oil emulsion by high-pressure microfluidic nanoemulsification technology at a pressure of 100 MPa - 130 MPa; after mixing the second water-in-oil emulsion with a part of the first aqueous solution, stir at a speed of 4000 rpm - 6000 rpm for 1 - 3 minutes to form a second water-in-oil-in-water emulsion; the second water-in-oil-in-water emulsion has a second water-in-oil emulsion core, and the D90 particle size of the internal aqueous phase of the second water-in-oil emulsion core is between 250 nm and 300 nm; Dissolve sodium alginate in water to form a second aqueous solution; Dissolve sodium alginate and hydroxypropyl-β-cyclodextrin in water to form a third aqueous solution; After preparing a third coarse emulsion from a mixture of carvacrol, thymol, Span 60, vitamin E, and MCT oil and the second aqueous solution, prepare a third water-in-oil emulsion by high-pressure microfluidic nanoemulsification technology at a pressure of 90 MPa - 130 MPa; after mixing the third water-in-oil emulsion with a part of the first aqueous solution, stir at a speed of 4000 rpm - 6000 rpm for 1 - 3 minutes to form a third water-in-oil-in-water emulsion; the third water-in-oil-in-water emulsion has a third water-in-oil emulsion core, and the D90 particle size of the internal aqueous phase of the third water-in-oil emulsion core is between 200 nm and 300 nm; After preparing a mixture of carvacrol, thymol, span 60, vitamin E, ascorbyl palmitate, and MCT oil with a third aqueous solution into a fourth crude emulsion, a fourth water-in-oil emulsion is prepared by high-pressure microfluidic nanoemulsification technology at a pressure of 50 MPa to 70 MPa; after mixing the fourth water-in-oil emulsion with a part of the first aqueous solution, it is stirred at a speed of 4000 rpm to 6000 rpm for 1 to 3 minutes to form a fourth water-in-oil-in-water emulsion; the fourth water-in-oil-in-water emulsion has a fourth water-in-oil emulsion core, and the D90 particle size of the inner aqueous phase of the fourth water-in-oil emulsion core is between 500 nm and 700 nm; After cooling the first water-in-oil-in-water emulsion, the second water-in-oil-in-water emulsion, the third water-in-oil-in-water emulsion, and the fourth water-in-oil-in-water emulsion to 10 °C to 15 °C, they are mixed and stirred at a speed of 2000 rpm to 3000 rpm for 1 to 3 minutes to form the nanoemulsion.

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