Traditional Chinese medicine compound essential oil microemulsion gel as well as preparation method and application thereof

Abstract

The invention provides traditional Chinese medicine compound essential oil microemulsion gel as well as a preparation method and application thereof, and relates to the technical field of traditional Chinese medicine compound essential oil preparations. The traditional Chinese medicine compound essential oil microemulsion gel is prepared from traditional Chinese medicine compound essential oil consisting of radix bupleuri essential oil, pericarpium citri reticulatae essential oil, ligusticum wallichii essential oil, murraya paniculata essential oil and rhizoma nardostachyos essential oil according to the theory of traditional Chinese medicine and by taking dialectical treatment as a principle, and has the effects of soothing the liver, promoting the circulation of qi, promoting blood circulation and relieving pain. On the basis of a single factor test, a response surface analysis method is applied to optimize a traditional Chinese medicine compound essential oil micro-emulsion gel prescription, and finally the traditional Chinese medicine compound essential oil micro-emulsion gel which is uniform, good in stability and good in ductility is obtained. The traditional Chinese medicine compound essential oil microemulsion gel can effectively improve depression-like behaviors of chronic unpredictable stress mice, can reduce the inflammatory factor level of a CUMS mouse model, and can effectively improve the bioavailability of active ingredients in the plant essential oil. Reference is provided for development of new dosage forms for preventing and treating depression and application of traditional Chinese medicine essential oil.

Description

Technical Field

[0001] The invention relates to the technical field of traditional Chinese medicine composite essential oil preparations, and in particular to a traditional Chinese medicine composite essential oil microemulsion gel and a preparation method and application thereof. Background Art

[0002] Depression is a common mental disorder of the central nervous system. Currently, about 17% of the world's population suffers from depression. With the continuous development of society, people's life pressure is increasing, and the incidence of depression is gradually increasing. The main manifestations of depression are loss of pleasure and interest, accompanied by other psychological and physical symptoms of varying degrees such as anxiety and sleep disorders. A report from the World Health Organization pointed out that more than 700,000 people commit suicide every year due to depression, which is also the main reason for the increase in global mortality.

[0003] At present, clinical antidepressant treatment mainly includes drug therapy, psychotherapy and physical therapy. Among them, drug therapy accounts for the majority. Most of the antidepressants used in clinical practice are synthetic antidepressants. The first generation of such drugs has been gradually abandoned due to safety issues and transitioned to the second generation of synthetic antidepressants. However, the second generation of drugs still have shortcomings such as slow onset and relatively single target. These factors have prompted scientists to develop new third-generation antidepressants with higher targeting, higher efficiency and higher safety. However, the current third-generation antidepressants still have problems such as insufficient efficacy and many adverse reactions in the long-term and short-term treatment of patients, and their safety and tolerability have always been controversial. Therefore, finding natural antidepressants with less toxicity and higher efficacy has become a new development hotspot.

[0004] In the theory of traditional Chinese medicine, Chinese medicines with aromatic smell are collectively referred to as aromatic drugs. Aromatic drugs account for a large proportion of Chinese medicinal materials and play an indispensable role in clinical treatment. Using aromatic drugs as enzymatic hydrolysis can improve the traditional "aromatherapy", and aromatic substances can be made into inhalants, essential oils, etc., which enter the body through breathing or skin to relieve mental stress, treat diseases, and promote human health. In recent years, more and more studies have revealed that plant essential oils can enter the body through olfactory absorption and skin penetration. They are often used as an auxiliary means of antidepressant treatment in clinical practice, and have a strong effect on people with depression or patients in the early stages of depression. However, due to the clearance of the nasal mucosa, the essential oil stays in the nasal mucosa for a short time, resulting in low bioavailability. However, with the development of drug dosage forms, it has been found that microemulsion gel drug-loaded dosage forms can prolong the retention time of drugs in the nasal mucosa and improve bioavailability. Summary of the invention

[0005] 1. Technical issues to be resolved

[0006] In view of the deficiencies in the prior art, the present invention provides a traditional Chinese medicine composite essential oil microemulsion gel with significant antidepressant effect and a preparation method thereof.

[0007] (II) Technical solution

[0008] To achieve the above objectives, the present invention is implemented through the following technical solutions:

[0009] The invention provides a traditional Chinese medicine compound essential oil microemulsion gel. The microemulsion gel is composed of the following components in weight percentage per 100 parts by weight: 6-9% of the traditional Chinese medicine compound essential oil, 25-35% of a surfactant, 12-16% of a cosurfactant, 1-5% of a moisturizer, 1-2% of a gel matrix, and the balance is purified water.

[0010] Specifically, every 100 parts by weight of the microemulsion gel is composed of the following components in percentage by weight: 7.98% of traditional Chinese medicine compound essential oil, 29.82% of surfactant, 14.91% of cosurfactant, 3% of moisturizer, 1.5% of gel matrix, and the balance is purified water.

[0011] Specifically, the Chinese medicine composite essential oil is composed of bupleurum essential oil, tangerine peel essential oil, chuanxiong essential oil, osmanthus fragrans essential oil and nardus odoratus essential oil in a mass ratio of 5-7:3-5:3-5:1-3:1-3; preferably, the Chinese medicine composite essential oil is composed of bupleurum essential oil, tangerine peel essential oil, chuanxiong essential oil, osmanthus fragrans essential oil and nardus odoratus essential oil in a mass ratio of 6:4.5:4.5:1.5:1.5.

[0012] Specifically, the surfactant is one of polyoxyethylene 40 hydrogenated castor oil (RH40), Tween-80 (TW-80), Tween-20 (TW-20), Span-80, and caprylic / capric macrogol glyceride (Labrasol); preferably, the surfactant is TW-80.

[0013] Specifically, the co-surfactant is one of Transcutol P, polyethylene glycol 400 (PEG400), 1,2-propylene glycol, anhydrous ethanol, and n-butanol; preferably, the co-surfactant is PEG400.

[0014] Specifically, the moisturizing agent is one of glycerin, trehalose, hyaluronic acid, sorbitol, and urea.

[0015] Specifically, the gel matrix is ​​one of Carbomer 934 or Carbomer 940. Preferably, the gel matrix is ​​Carbomer 940.

[0016] The present invention also provides a method for preparing a traditional Chinese medicine composite essential oil microemulsion gel, wherein the microemulsion gel is prepared according to the following steps:

[0017] (1) Preparation of Chinese medicine composite essential oil: 500 g each of bupleurum, dried orange peel, chuanxiong, osmanthus fragrans, and nard were finely ground and passed through a No. 2 sieve to obtain uniform powder; then, each powder was mixed with distilled water 9 times its volume, and each was placed in an electric heating mantle and soaked for 1 hour to fully infiltrate; then, glass beads were added and connected to a volatile oil analyzer, and a reflux condenser was installed at the upper end, and the water was heated to gradually boil and maintain a slightly boiling state; after 6 hours of continuous extraction, when the volume of the volatile oil no longer increased, Stop heating and allow the system to cool naturally for 30 minutes; then, record the volume of the volatile oil and collect it; to ensure purity, use anhydrous sodium sulfate to dry the volatile oil; the dried volatile oil is a yellow oily liquid, which is stored in a brown bottle and placed in a refrigerator at 4°C for subsequent use; Bupleurum essential oil, tangerine peel essential oil, Chuanxiong essential oil, Murraya osmanthus essential oil, and Nardostachys grandiflora essential oil are mixed evenly according to a mass ratio of 5-7:3-5:3-5:1-3:1-3 to obtain a traditional Chinese medicine composite essential oil;

[0018] (2) uniformly mixing the Chinese medicine composite essential oil, surfactant and co-surfactant according to the above mass percentages, slowly adding water under magnetic stirring, and continuously stirring for more than 40 minutes to obtain a microemulsion solution for standby use;

[0019] (3) adding a gel matrix to the microemulsion solution obtained in step (2), mixing evenly, allowing it to fully swell overnight, adjusting the pH to 6-7 with triethanolamine, then adding a moisturizer, and mixing evenly to obtain the traditional Chinese medicine composite essential oil microemulsion gel.

[0020] (III) Beneficial effects

[0021] The liver governs the flow of qi and likes to be smooth. Its meridians are distributed in the ribs and lower abdomen. If the emotions are not satisfied and the wood fails to flow smoothly, it will lead to liver qi stagnation and poor meridian qi, so there will be pain in the ribs, chest tightness, and abdominal distension. If the liver fails to flow, the emotions will be depressed, irritable, and prone to sighing. The stringy pulse is a sign of liver depression and discomfort. According to the principle of "relieving wood depression" in the "Inner Canon of Medicine", the treatment should be to soothe the liver and regulate qi. The invention follows the theory of traditional Chinese medicine, takes dialectical treatment as the principle, and takes Chaihu Shugan San as the basic formula to obtain a Chinese medicine composite essential oil microemulsion gel prepared from Chinese medicine composite essential oils composed of Chaihu essential oil, Chenpi essential oil, Chuanxiong essential oil, Murraya odorata essential oil, and Nardostachys raphis essential oil. In the formula, Chaihu is good at soothing the liver and relieving depression, and is used as the main drug; Chenpi regulates qi, soothes the liver, and relieves stagnation, and is the minister drug; Chuanxiong promotes blood circulation and qi to relieve pain, and the two drugs are combined to help Chaihu to relieve the stagnation of the liver meridian, and increase the effects of promoting qi, promoting blood circulation, and relieving pain, and are the minister drugs. Murraya odorata promotes qi and blood circulation, relieves blood stasis, and relieves pain, and Nardostachys raphis regulates qi and relieves pain, and relieves depression and invigorates the spleen, and the two drugs are the adjuvant drugs; all the drugs are combined to play the functions of soothing the liver, promoting qi, and promoting blood circulation to relieve pain.

[0022] The invention optimizes the prescription of the traditional Chinese medicine composite essential oil microemulsion gel by using the response surface analysis method on the basis of a single factor experiment, and finally obtains the optimal prescription of the traditional Chinese medicine composite essential oil microemulsion gel, which is specifically: 7.98% of traditional Chinese medicine composite essential oil, 29.82% of surface active agent, 14.91% of cosurfactant, 3% of moisturizer, 1.5% of gel matrix, and the balance is purified water; wherein the surfactant is TW-80, the cosurfactant is PEG 400, the moisturizer is glycerin, and the gel matrix is ​​carbomer 940; and the traditional Chinese medicine composite essential oil microemulsion gel with uniformity, good stability and good ductility is prepared.

[0023] The Chinese medicine composite essential oil microemulsion gel of the present invention can effectively improve the depressive-like behavior of chronic unpredictable stress mice, and can reduce the level of inflammatory factors in the CUMS mouse model, and the effect is significantly improved compared with the direct use of Chinese medicine composite essential oil, and can effectively improve the bioavailability of active ingredients in plant essential oils. The addition of Murraya osmanthus essential oil and Nardostachys chinensis essential oil in the present invention greatly improves the antidepressant effect of the Chinese medicine composite essential oil microemulsion gel, and each component in the Chinese medicine composite essential oil microemulsion gel of the present invention synergizes and enhances the effect, and none of them can be missing. The present invention provides reference and reference for the development of new dosage forms for the prevention and treatment of depression and the application of Chinese medicine essential oils. BRIEF DESCRIPTION OF THE DRAWINGS

[0024] Figure 1 The state of microemulsion solution prepared by different surfactants and traditional Chinese medicine composite essential oil when km=2.

[0025] Figure 2 The state of microemulsion solution prepared with different co-surfactants and traditional Chinese medicine composite essential oil when km=2.

[0026] Figure 3 Pseudo-ternary phase diagram for surfactant screening; A: oil phase-RH40 / Transcutol P (2:1)-water; B: oil phase-TW-80 / Transcutol P (2:1)-water; C: oil phase-[RH40 / TW-80 (2:1)] / Transcutol P (2:1)-water.

[0027] Figure 4 Pseudo-ternary phase diagram for co-surfactant screening; A: oil phase-TW-80 / 1,2-propylene glycol (2:1)-water; B: oil phase-TW-80 / PEG 400 (2:1)-water; C: oil phase-TW-80 / Transcutol P (2:1)-water.

[0028] Figure 5Pseudo-ternary phase diagram for screening km values; A: oil phase-TW-80 / PEG 400 (1:1)-water; B: oil phase-TW-80 / PEG 400 (2:1)-water; C: oil phase-TW-80 / PEG 400 (3:1)-water.

[0029] Figure 6 for Figure 6 Three-dimensional effect surface diagram; A: three-dimensional effect surface diagram (particle size); B: contour diagram (particle size); C: three-dimensional effect surface diagram (PDI); D: contour diagram (PDI).

[0030] Figure 7 It is the appearance, properties and particle size distribution of Chinese medicine compound essential oil microemulsion. DETAILED DESCRIPTION

[0031] In order to make the purpose, technical solution and advantages of the embodiments of the present invention clearer, the technical solution in the embodiments of the present invention will be clearly and completely described below in combination with the embodiments of the present invention. Obviously, the described embodiments are part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0032] Example 1

[0033] Preparation of Chinese herbal compound essential oil:

[0034] 500 g each of Bupleurum, Tangerine Peel, Chuanxiong, Murraya Osmanthus, and Nardostachys grandiflora were finely ground and passed through a No. 2 sieve to obtain uniform powders; then, each powder was mixed with distilled water 9 times its volume and immersed in an electric heating jacket for 1 hour to fully infiltrate; then, glass beads were added and connected to a volatile oil analyzer, and a reflux condenser was installed at the upper end, and the water was heated to gradually boil and maintain a slightly boiling state; after 6 hours of continuous extraction, when the volume of the volatile oil no longer increased, the heating was stopped. Allow the system to cool naturally for 30 minutes; then, record the volume of the volatile oil and collect it; to ensure the purity, use anhydrous sodium sulfate to dry the volatile oil; the dried volatile oil is a yellow oily liquid, which is stored in a brown bottle and placed in a refrigerator at 4°C for subsequent use; Bupleurum essential oil, tangerine peel essential oil, Chuanxiong essential oil, Osmanthus fragrans essential oil, and Nardostachys grandiflora essential oil are evenly mixed in a mass ratio of 6:4.5:4.5:1.5:1.5 to obtain a Chinese medicine composite essential oil.

[0035] Example 2

[0036] The difference between this embodiment and embodiment 1 is that the Chinese medicine composite essential oil is obtained by uniformly mixing bupleurum essential oil, tangerine peel essential oil, chuanxiong essential oil, osmanthus fragrans essential oil, and nard essential oil in a mass ratio of 5:3:3:1:1.

[0037] Example 3

[0038] The difference between this embodiment and embodiment 1 is that the Chinese medicine composite essential oil is obtained by uniformly mixing bupleurum essential oil, tangerine peel essential oil, chuanxiong essential oil, osmanthus fragrans essential oil, and nard essential oil in a mass ratio of 7:5:5:3:3.

[0039] Example 4

[0040] The difference between this embodiment and embodiment 1 is that the Chinese medicine composite essential oil is obtained by uniformly mixing bupleurum essential oil, tangerine peel essential oil, chuanxiong essential oil, osmanthus fragrans essential oil, and nard essential oil in a mass ratio of 5.8:4:4.2:2:2.

[0041] Example 5

[0042] The present invention provides a method for preparing a traditional Chinese medicine composite essential oil microemulsion gel, the preparation method comprising the following steps:

[0043] (1) Preparation of Chinese herbal compound essential oil: 500 g each of Bupleurum, Tangerine Peel, Chuanxiong, Murraya Fragrant, and Nardostachys grandiflora were finely ground and passed through a No. 2 sieve to obtain uniform powder; then, each powder was mixed with distilled water 9 times its volume and placed in an electric heating mantle for 1 hour to fully infiltrate; then, glass beads were added and connected to a volatile oil analyzer, and a reflux condenser was installed at the upper end, and the water was heated to gradually boil and maintain a slightly boiling state; after 6 hours of continuous extraction, when the volume of the volatile oil no longer increased, , stop heating, and let the system cool naturally for 30 minutes; then, record the volume of volatile oil and collect it; to ensure purity, use anhydrous sodium sulfate to dry the volatile oil; the dried volatile oil is a yellow oily liquid, which is stored in a brown bottle and placed in a refrigerator at 4°C for subsequent use; Bupleurum essential oil, tangerine peel essential oil, Chuanxiong essential oil, Murraya osmanthus essential oil, and Nardostachys grandiflora essential oil are evenly mixed in a mass ratio of 6:4.5:4.5:1.5:1.5 to obtain a traditional Chinese medicine composite essential oil;

[0044] (2) uniformly mixing the Chinese medicine composite essential oil, surfactant and co-surfactant according to the above mass percentages, slowly adding water under magnetic stirring, and continuously stirring for more than 40 minutes to obtain a microemulsion solution for standby use;

[0045] (3) adding a gel matrix to the microemulsion solution obtained in step (2), mixing evenly, allowing it to fully swell overnight, adjusting the pH to 6-7 with triethanolamine, then adding a moisturizer, and mixing evenly to obtain the traditional Chinese medicine composite essential oil microemulsion gel.

[0046] The present invention optimizes the prescription of the traditional Chinese medicine composite essential oil microemulsion gel by using response surface analysis method on the basis of single factor experiment, which is described in detail below.

[0047] 1 Experimental methods and results

[0048] 1.1 Choice of surfactant

[0049] The co-surfactant was Transcutol P, the oil phase was a traditional Chinese medicine compound essential oil, and different surfactants (RH40, TW-80, TW-20, Labrasol, Span-80) were investigated.

[0050] According to km=2, ​​the co-surfactant was prepared with different surfactants to form a mixed emulsifier, the mixed emulsifier and the traditional Chinese medicine compound essential oil were mixed in a suitable ratio, and water was added and titrated evenly and slowly under magnetic stirring. When the surfactant was RH40 or TW-80, the prepared microemulsion solution was uniform, clear and transparent (see Figure 1 ), so RH40 and TW-80 were initially selected as surfactants for the prescription.

[0051] 1.2 Selection of co-surfactant

[0052] The oil phase was traditional Chinese medicine compound essential oil, the surfactant was RH40, and different co-surfactants (Transcutol P, PEG 400, 1,2-propylene glycol, anhydrous ethanol, and n-butanol) were investigated.

[0053] According to km=2, ​​the surfactants were respectively prepared with different co-surfactants to form mixed emulsifiers, the mixed emulsifiers were mixed with the traditional Chinese medicine compound essential oil in a suitable ratio, water was slowly titrated under magnetic stirring, and the prepared microemulsion was uniform, clear and transparent (see Figure 2 ), but considering the biosafety index, Transcutol P, PEG 400, and 1,2-propylene glycol were initially selected as cosurfactants.

[0054] 1.3 Single factor study of prescription

[0055] 1.3.1 Screening of surfactants

[0056] The auxiliary surfactant is Transcutol P, the oil phase is traditional Chinese medicine compound essential oil, and the surfactants are RH40 and TW-80.

[0057] According to km=2, ​​Transcutol P was respectively mixed with RH40, TW-80, and RH40 / TW-80 (2:1) to prepare mixed emulsifiers. The oil phase and the mixed emulsifier were mixed in the ratios of 0.5:9.5, 1:9, 1.5:8.5, 2:8, 2.5:7.5, 3:7, 3.5:6.5, and 4:6. Water was slowly added and titrated until the critical point of clear to turbid and then to clear was reached. The data was recorded and a pseudo-ternary phase diagram was drawn. The results are shown in Tables 1 and Figure 3It shows that when TW-80 is used as the surfactant, the microemulsion area is the largest, so TW-80 is selected as the surfactant.

[0058] Table 1 Screening of surfactants

[0059] serial number prescription area A Oil phase-RH40 / Transcutol P (2:1)-water 0.0164 B Oil phase-TW-80 / Transcutol P (2:1)-water 0.0207 C Oil phase - [RH40 / TW-80 (2:1)] / Transcutol P (2:1) - Water 0.0150

[0060] 1.3.2 Screening of co-surfactants

[0061] TW-80 was used as a surfactant, Chinese herbal compound essential oil was used as the oil phase, and the co-surfactants were Transcutol P, PEG 400, and 1,2-propylene glycol.

[0062] According to km=2, ​​TW-80 was mixed with Transcutol P, PEG 400, and 1,2-propylene glycol to prepare mixed emulsifiers. The Chinese medicine compound essential oil and the mixed emulsifier were mixed in the ratio of 0.5:9.5, 1:9, 1.5:8.5, 2:8, 2.5:7.5, 3:7, 3.5:6.5, and 4:6. Water was slowly added and titrated under magnetic stirring. When the critical point of clear to turbid and then to clear was reached, the data was recorded and the pseudo-ternary phase diagram was drawn using Origin2019 64Bit. The results are shown in Table 2 and Figure 4 When TW-80 is the surfactant and PEG400 is the co-surfactant, the microemulsion area prepared when the compound essential oil is the oil phase is the largest, so PEG400 is selected as the co-surfactant.

[0063] Table 2 Screening of co-surfactants

[0064] serial number prescription area A Oil phase-TW-80 / 1,2-propylene glycol (2:1)-water 0.0278 B Oil phase-TW-80 / PEG 400(2:1)-water 0.0323 C Oil phase-TW-80 / Transcutol P (2:1)-water 0.0207

[0065] 1.3.3 km value screening

[0066] Different mixed emulsifiers were prepared with TW-80 and PEG 400 according to different km values ​​(km = 1:1, 2:2, 3:1, 4:1), and the Chinese medicine compound essential oil and the mixed emulsifier were mixed in the ratio of 0.5:9.5, 1:9, 1.5:8.5, 2:8, 2.5:7.5, 3:7, 3.5:6.5, 4:6. Water was slowly added and titrated under magnetic stirring. When the critical point of clear to turbid and then to clear was reached, the data was recorded and the pseudo-ternary phase diagram was drawn using Origin2019 64Bit. The results are shown in Table 3 and Figure 5 The microemulsion area is the largest when the ratio of surfactant (TW-80) to co-surfactant (PEG 400) is km=2:1. Therefore, km=2 is selected.

[0067] Table 3 Screening of km values

[0068] serial number prescription area A Oil phase-TW-80 / PEG 400(1:1)-water 0.0163 B Oil phase-TW-80 / PEG 400(2:1)-water 0.0323 C Oil phase-TW-80 / PEG 400(3:1)-water 0.0080

[0069] 1.4 Central-site design-response surface methodology to optimize prescription

[0070] 1.4.1 Optimization of the prescription of Chinese medicine compound essential oil microemulsion

[0071] The oil phase ratio (A%) of the traditional Chinese medicine compound essential oil and the mixed phase ratio (B%) of the mixed emulsifier [TW-80 / PEG 400 (km=2:1)] were used as factors, and the microemulsion particle size Y1 (nm) and Y2 (PDI) were used as indicators to optimize the formulation design. The experimental factor levels are shown in Table 4, and the microemulsion star point design table and results are shown in Table 5.

[0072] Table 4 Microemulsion factor levels

[0073]

[0074] Table 5 Microemulsion star point design table and results

[0075]

[0076] 1.4.2 Data results and processing

[0077] The results of ANOVA and binomial equation fitting are shown in Tables 6 and 7.

[0078] Table 6 Particle size-variance-analysis results

[0079] Table 7 PDI-variance-analysis results

[0080]

[0081] 1.4.3 Effect Surface

[0082] When km=2, ​​when the mixed phase ratio is constant, the particle size and PDI gradually increase with the increase of the oil phase ratio; when the oil phase ratio is constant, the particle size and PDI both increase with the increase of the mixed phase ratio. Figure 6 The final optimized prescription composition is Chinese herbal compound essential oil (7.98%): Tween-80 (29.82%): PEG400 (14.91%): water (47.29%).

[0083] 1.3.4 Verification test

[0084] Three parallel verifications were carried out according to the optimized prescription process, and the obtained microemulsion results were all clear and transparent, with an average particle size of 20.14 (±0.23nm), an average PDI value of 0.23 (±0.01), and an average Zeta potential of -18.36 (±4.2), which were in line with the expected results.

[0085] 1.5 Quality evaluation of traditional Chinese medicine compound microemulsion

[0086] 1.5.1 Appearance and physical and chemical properties

[0087] The traditional Chinese medicine compound essential oil microemulsion prepared according to the prescription is yellow, uniform, transparent, clear and has good fluidity (see Figure 7 ). Three groups of microemulsion preparations were prepared in parallel according to the optimized prescription. The average particle size was 20.14 (±0.23nm), the average PDI value was 0.23 (±0.01), the average Zeta potential was -18.36 (±4.2) mV, the average pH was 6.32 (±0.14), the average refractive index was 1.425 (26.3℃), and the average conductivity was 290μS / cm; indicating that the essential oil microemulsion prepared by this prescription has good physical and chemical properties.

[0088] 1.5.2 Stability study

[0089] 1.5.2.1 Centrifugal stability

[0090] Three groups of volatile oil microemulsions were prepared in parallel at 13000 r·min -1 The mixture was centrifuged for 20 min under the conditions of 40 °C and 10 °C, and its appearance and particle size were observed. The results are shown in Table 8. After centrifugation, it was clear and transparent, and the particle size and PDI did not change significantly, indicating that its centrifugal stability was good.

[0091] Table 8 Centrifugal stability ( n=3)

[0092] serial number Appearance Particle size PDI 1 Clear, transparent, uniform 20.22±0.18 0.21±0.01 2 Clear, transparent, uniform 19.96±0.17 0.20±0.01 3 Clear, transparent, uniform 20.64±0.21 0.23±0.02

[0093] 1.5.2.2 Temperature stability

[0094] The microemulsion solution was placed at 4, 25, and 50°C for 24 hours. The results are shown in Table 9. The microemulsion remained clear and transparent at different temperatures, and there was no obvious change in particle size and PDI. There was no stratification or demulsification.

[0095] Table 9 Temperature stability ( n=3)

[0096] serial number Appearance Particle size PDI 1 Clear, transparent, uniform 21.42±0.21 0.23±0.01 2 Clear, transparent, uniform 22.64±0.18 0.21±0.01 3 Clear, transparent, uniform 21.94±0.26 0.24±0.02

[0097] 1.5.2.3 Dilution stability

[0098] The microemulsion was diluted with double distilled water at a ratio of 1:100, 1:500, and 1:1000, respectively. The results are shown in Table 10. The microemulsion did not stratify, and the particle size and PDI did not change significantly, indicating good dilution stability.

[0099] Table 10 Dilution stability ( n=3)

[0100] serial number Appearance Particle size PDI 1 Clear, transparent, uniform 22.48±0.24 0.16±0.01 2 Clear, transparent, uniform 21.73±0.22 0.19±0.01 3 Clear, transparent, uniform 20.15±0.21 0.18±0.01

[0101] 1.6 Preparation of microemulsion gel

[0102] 1.6.1 Screening of gel matrix dosage

[0103] Carbomer 934 and Carbomer 940 were weighed at 1.0%, 1.5%, and 2.0% by mass, and added to 6.0 g of the traditional Chinese medicine compound essential oil microemulsion solution to allow it to fully swell overnight. The pH was adjusted to 6-7 with triethanolamine, and the mixture was heated at 9000 r·min. -1 After centrifugation for 10 minutes, the properties, viscosity, fluidity, stability and ductility of the microemulsion gel were evaluated. The results are shown in Table 11.

[0104] Table 11 Screening of gel matrix

[0105] Matrix type content Characteristics Viscosity Liquidity stability Ductility Carbomer 934 1.00％ Yellow fluid, agglomerates weak have Uneven, layered Difficult to apply Carbomer 934 1.50％ Yellow gel powerful none Uniform, stable, no stratification Easy to apply Carbomer 934 2.00％ Yellow fluid, agglomerates weak have Uneven, layered Difficult to apply Carbomer 940 1.00％ Yellow fluid weak have Uniform, stable, no stratification Easy to apply Carbomer 940 1.50％ Yellow fluid weak have Uniform, stable, no stratification Easy to apply Carbomer 940 2.00％ Yellow fluid weak have Uniform, stable, no stratification Easy to apply

[0106] 1.6.2 Moisturizer content screening

[0107] Moisturizers with glycerol contents of 1%, 3%, and 5% were added to the traditional Chinese medicine composite essential oil microemulsion gel for screening. The results showed that when the amount of glycerol was 1%, the gel appearance did not change significantly, and there was no greasy feeling; when the amount of glycerol was 5%, the gel felt slightly greasy and the ductility also decreased; when the amount of glycerol was 3%, the gel was clear and bright, with moderate greasy feeling and good ductility. Therefore, the amount of glycerol was selected to be 3%.

[0108] 2 Conclusion

[0109] The invention optimizes the prescription of the traditional Chinese medicine compound essential oil microemulsion gel by using the response surface analysis method on the basis of a single factor experiment, and finally obtains the optimal prescription of the traditional Chinese medicine compound essential oil microemulsion gel, which specifically comprises 7.98% of the traditional Chinese medicine compound essential oil, 29.82% of the surface active agent, 14.91% of the cosurfactant, 3% of the humectant, 1.5% of the gel matrix, and the balance is purified water; wherein the surfactant is TW-80, the cosurfactant is PEG 400, the humectant is glycerin, and the gel matrix is ​​carbomer 940.

[0110] Comparative Example 1

[0111] A microemulsion gel was prepared according to the method described in Example 5. The microemulsion gel had a prescription of: 7.98% of traditional Chinese medicine compound essential oil, 29.82% of TW-80, 14.91% of PEG 400, 3% of glycerol, 1.5% of carbomer 940, and the remainder was purified water; wherein the traditional Chinese medicine compound essential oil was obtained by uniformly mixing bupleurum essential oil, tangerine peel essential oil, and chuanxiong essential oil in a mass ratio of 6:4.5:4.5.

[0112] Comparative Example 2

[0113] The difference between this comparative example and comparative example 1 is that the traditional Chinese medicine composite essential oil is obtained by uniformly mixing tangerine peel essential oil, chuanxiong essential oil and cyperus rotundus essential oil in a mass ratio of 4.5:4.5:3.

[0114] Test example

[0115] 1. Preparation and grouping of depression animal models

[0116] SPF female Kunming mice were selected, with a body weight of (20±2) g. After 3 days of adaptive feeding, the mice were randomly divided into a blank group, a fluoxetine hydrochloride group (0.0025 mg / g), a model group, a Chinese medicine compound essential oil (Essential oil, EO) group (the Chinese medicine compound essential oil prepared in Example 1, 0.25 μl / g), a Chinese medicine compound essential oil microemulsion gel (Essential oilmicroemulsion Gel, EOMG) group (the Chinese medicine compound essential oil microemulsion gel of the present invention prepared in Example 5, 0.25 μl / g), a control group 1 (the microemulsion gel prepared in Comparative Example 1, 0.25 μl / g), and a control group 2 (the microemulsion gel prepared in Comparative Example 2, 0.25 μl / g), 12 mice in each group, and the drug was administered by nasal drops. The mice in the blank group were housed together without the modeling procedure, and the mice in the other groups were housed in single cages and subjected to the CUMS procedure for modeling: cold water swimming (4°C, 5min), empty cage, tail clamp (3min), light-dark reversal (24h), food and water deprivation for 12h, wet bedding (12h), odor stimulation (3h), restraint (2h), crowding (12h), no bedding (24h), cage tilted 45° (24h), noise environment (60Hz, 1h), 45°C heat stimulation (5min), cage vibration for 10min. The above stimulation factors were randomly assigned, and unpredictable stimulation methods were adopted during the stress cycle, that is, one of the above stress stimulation methods was randomly selected every day, and it was ensured that the method was not repeated within a week. Drug administration and modeling were carried out simultaneously, and the modeling procedure was not stopped during drug administration, and the modeling and drug administration were continued for 28 days.

[0117] 2Indicator detection

[0118] 2.1 Weight measurement

[0119] The weight of mice was monitored weekly, and the differences in weight gain among the groups were compared to determine the effects of chronic stress on the feeding and growth of mice and the effects of the traditional Chinese medicine composite essential oil microemulsion gel of the present invention on body weight.

[0120] 2.2 Sugar water preference rate

[0121] Before the sugar water preference experiment, each group of mice was given 1 bottle of plain water and 1 bottle of 1% sucrose water to drink, so as to adapt to the situation of having plain water on one side and sucrose water on the other. After 12 hours of adaptation, all mice were housed in single cages and fasted from food and water for 12 hours. Each mouse was given 1 bottle of plain water and 1 bottle of 1% sucrose water, and the masses of the two water bottles were weighed. After the mice drank plain water or sucrose water for 12 hours, the water bottles were removed and the masses of the water bottles were weighed again. The difference in the masses of the two water bottles was recorded as the consumption of plain water or sucrose water by the mice, and the sugar water preference rate of the mice was calculated. The basal value of the sugar water preference rate of the mice was measured before modeling, and the sugar water preference rate of the mice was measured after the modeling was completed.

[0122] Sugar water preference rate = sucrose water consumption / (white water consumption + sucrose water consumption) × 100%

[0123] 2.3 Tail suspension test

[0124] The mouse was fixed in a horizontal position with tape 2 cm behind its tail, so that it was hanging upside down with its head 5 to 6 cm above the lower horizontal plane. The cumulative immobility time of the animal during the last 4 minutes within 6 minutes was recorded.

[0125] 2.4 Splash test

[0126] 10% sucrose water was sprayed on the back of the mouse near the tail for three times in a row. Then the mouse was placed in the cage where it originally lived and the grooming time of the mouse was recorded within 5 minutes (the time the mouse groomed the location of the sugar water spray on its back was recorded as the grooming time). The measurement was performed one week before sampling.

[0127] 2.5ELISA detection of serum inflammatory factor levels

[0128] 2 hours after the last administration, blood was collected from the mouse eyeballs, left to stand at room temperature for 30 minutes, and centrifuged at 3500 r / min for 10 minutes (centrifugal radius 7.5 cm) to separate the serum. The levels of tumor necrosis factor-α (TNF-α), interleukin (IL-18), IL-1β, and IL-6 in the serum were determined according to the instructions of the kit.

[0129] 2.6 Statistical analysis

[0130] Excel was used to organize behavioral measurement data and expressed as mean ± standard deviation (xˉ±s). Statistical analysis was performed using the prism software package, and one-way ANOVA was used for comparison among multiple groups. P<0.05 indicated a statistically significant difference, and P<0.01 indicated a statistically significant difference.

[0131] 3 Results

[0132] 3.1 Effect of the Traditional Chinese Medicine Composite Essential Oil Microemulsion Gel of the Present Invention on Body Weight of CUMS Mouse Model

[0133] The comparison of body weight is shown in Table 12. Before modeling, the weight difference between the mice in each group was not large, and the results were not significantly different (P>0.05). After 28 days of CUMS stimulation, the body weight of the model group decreased significantly compared with the blank group, with a significant difference (P<0.01); compared with the model group, the body weight of the EOMG group increased significantly (P<0.01); the body weight of the fluoxetine hydrochloride group, EO group, control group 1 and 2 also showed an increasing trend, but there was no significant difference compared with the model group (P<0.01).

[0134] Table 12 Results of weight changes of mice in each group ( n=12)

[0135]

[0136]

[0137] Note: Compared with the model group, ## P<0.01, # P<0.05.

[0138] 3.2 Effect of the Traditional Chinese Medicine Composite Essential Oil Microemulsion Gel of the Present Invention on Sugar Preference Rate of CUMS Mouse Model

[0139] As shown in Table 13, before modeling, there was no significant difference in the sugar preference rate of mice in each group (P>0.05); after modeling, compared with the blank group, the sugar preference rate of mice in the model group was significantly reduced (P<0.01), and the sugar preference rates of mice in the other groups also decreased to varying degrees, proving that the modeling was successful. After modeling and administration, compared with the model group, the sugar preference rate of mice in the hydrochloride group and the EOMG group was significantly increased (P<0.01); compared with the model group, the sugar preference rate of mice in the EO group was significantly increased (P<0.05); compared with the model group, the sugar preference rate of mice in the control group 1 and 2 was increased, but the difference was not significant (P>0.05).

[0140] Table 13 Results of sugar water preference rate of mice in each group ( n=12)

[0141] Group Dosage (mg / kg) Sugar water preference rate before modeling (%) Sugar water preference rate after modeling (%) Blank Group - 83.79％±2.13 <![CDATA[90.37%±1.71 ## ]]> Model Group - 80.35％±7.09 50.36％±8.11 Fluoxetine Hydrochloride 0.0025mg / g 85.08％±1.75 <![CDATA[81.01%±4.67 ## ]]> EO group 0.25μl / g 84.62％±2.95 <![CDATA[63.12%±4.93 # ]]> Control group 1 0.25μl / g 83.33±3.06 60.64±5.47 Control group 2 0.25μl / g 82.26±2.57 58.23±4.03 EOMG group 0.25μl / g 82.94％±2.32 <![CDATA[73.40%±1.12 ## ]]>

[0142] Note: Compared with the model group, ## P<0.01, # P<0.05.

[0143] 3.3 Effect of the Traditional Chinese Medicine Composite Essential Oil Microemulsion Gel of the Present Invention on the Tail Suspension Immobility Time of the CUMS Mouse Model

[0144] As shown in Table 14, compared with the blank group, the immobility time of the tail suspension in the model group was significantly increased (P<0.01). Compared with the model group, the immobility time of the tail suspension in the cetin hydrochloride group and the EOMG group was significantly reduced (P<0.01); compared with the model group, the immobility time of the tail suspension in the EO group was significantly reduced (P<0.05); compared with the model group, the immobility time of the tail suspension in the control group 1 and 2 mice was reduced, but the difference was not significant (P>0.05).

[0145] Table 14 Results of tail suspension immobility time of mice in each group ( n=12)

[0146]

[0147]

[0148] Note: Compared with the model group, ## P<0.01, # P<0.05.

[0149] 3.4 Effect of the Traditional Chinese Medicine Composite Essential Oil Microemulsion Gel of the Present Invention on the Grooming Time of the CUMS Mouse Model

[0150] As shown in Table 15, compared with the blank group, the grooming time of mice in the model group was significantly reduced (P<0.01). Compared with the model group, the grooming time of mice in the cetin hydrochloride group and EOMG group was significantly increased (P<0.01); compared with the model group, the grooming time of mice in the EO group was significantly increased (P<0.05); compared with the model group, the grooming time of mice in the control group 1 and 2 increased, but the difference was not significant (P>0.05).

[0151] Table 15 Results of grooming time of mice in each group ( n=12)

[0152] Group dose Number of animals (n) Grooming time / (S) Blank Group - 12 <![CDATA[89.92±28.80 ## ]]> Model Group - 12 36.33±28.91 Fluoxetine Hydrochloride 0.0025mg / g 12 <![CDATA[79.83±18.32 ## ]]> EO group 0.25μl / g 12 <![CDATA[59.08±18.48 # ]]> Control group 1 0.25μl / g 12 52.17±10.38 Control group 2 0.25μl / g 12 50.64±9.27 EOMG group 0.25μl / g 12 <![CDATA[62.67±15.87 ## ]]>

[0153] Note: Compared with the model group, ## P<0.01, # P<0.05.

[0154] 3.5 Effect of the Traditional Chinese Medicine Composite Essential Oil Microemulsion Gel of the Present Invention on Inflammatory Factors in the CUMS Mouse Model

[0155] As shown in Table 16, compared with the blank group, the IL-18, IL-6, IL-1β and TNF-α of the mice in the model group were significantly increased (P < 0.01); compared with the model group, the level of serum inflammatory factor IL-18 of the mice in the EO group and EOMG group was significantly decreased (P < 0.01), the level of serum inflammatory factor IL-6 of the mice in the EOMG group was significantly decreased (P < 0.01), and the level of serum inflammatory factor IL-6 of the mice in the EO group was significantly decreased (P < 0.05); the level of serum inflammatory factor IL-18 of the mice in the fluoxetine hydrochloride group, EO group and EOMG group was significantly decreased (P < 0.01). The level of serum inflammatory factor IL-1β was significantly decreased (P<0.01); the level of serum inflammatory factor TNF-α in mice in fluoxetine hydrochloride and EOMG groups was significantly decreased (P<0.01), and the level of serum inflammatory factor TNF-α in EO mice was significantly decreased (P<0.05); compared with the model group, the level of inflammatory factor IL-18 in mice in control group 1 and control group 2 was significantly decreased (P<0.05), and the levels of other inflammatory factors TNF-α, IL-6, and IL-1β were decreased, but there was no significant difference in the content (P>0.05).

[0156] Table 16 Results of inflammatory factors in each group of mice ( n=12)

[0157] Group IL-18 (pg / ml) IL-6 (pg / ml) IL-1β (pg / ml) TNF-α(pg / ml) Blank Group <![CDATA[61.9±26.24 ## ]]> <![CDATA[42.59±11.2 ## ]]> <![CDATA[39.54±3.37 ## ]]> <![CDATA[329.64±58.14 ## ]]> Model Group 230.8±23.14 68.22±8.16 98.08±9.57 689.18±115.62 Fluoxetine Hydrochloride 207.37±25.7 62.54±10.02 <![CDATA[69.19±12.34 ## ]]> <![CDATA[502.01±95.92 ## ]]> EO group <![CDATA[131.58±32.23 ## ]]> <![CDATA[52.59±10.26 # ]]> <![CDATA[63.66±11.91 ## ]]> <![CDATA[551.35±42.39 # ]]> Control group 1 <![CDATA[155.39±19.44 # ]]> 60.31±11.75 <![CDATA[72.46±12.57 # ]]> 597.93±47.42 Control group 2 <![CDATA[164.59±21.60 # ]]> 61.24±13.68 <![CDATA[74.83±11.68 # ]]> 608.29±63.64 EOMG group <![CDATA[105.21±22.38 ## ]]> <![CDATA[46.69±9.96 ## ]]> <![CDATA[48.45±16.08 ## ]]> <![CDATA[405.46±82.26 ## ]]>

[0158] Note: Compared with the model group, ## P<0.01, # P<0.05.

[0159] 4 Conclusion

[0160] The invention follows the theory of traditional Chinese medicine, takes dialectical treatment as the principle, and takes Chaihu Shugan San as the basic formula to obtain a Chinese medicine composite essential oil microemulsion gel prepared from Chinese medicine composite essential oils composed of Chaihu essential oil, Chenpi essential oil, Chuanxiong essential oil, Murraya odorata essential oil, and Nardostachys raphis essential oil. In the formula, Chaihu is good at soothing the liver and relieving depression, and is used as the main drug; Chenpi regulates qi, soothes the liver, and relieves stagnation, and is the minister drug; Chuanxiong promotes blood circulation and qi to relieve pain, and the two drugs are combined to help Chaihu to relieve the stagnation of the liver meridian, and increase the effects of promoting qi, promoting blood circulation, and relieving pain, and are the minister drugs. Murraya odorata promotes qi and blood circulation, relieves blood stasis, and relieves pain, and Nardostachys raphis regulates qi and relieves pain, and relieves depression and invigorates the spleen, and the two drugs are the adjuvant drugs; all the drugs are combined to play the functions of soothing the liver, promoting qi, and promoting blood circulation to relieve pain.

[0161] According to the above test results, the Chinese medicine composite essential oil microemulsion gel of the present invention can effectively improve the depressive-like behavior of chronic unpredictable stress mice, and can reduce the level of inflammatory factors in the CUMS mouse model, and the effect is improved compared with the direct use of Chinese medicine composite essential oil, and can effectively improve the bioavailability of active ingredients in plant essential oils. At the same time, the antidepressant effect of the Chinese medicine composite essential oil microemulsion gel of the present invention is significantly better than that of Comparative Example 1, Comparative Example 2 and Western medicine fluoxetine hydrochloride. The addition of Murraya odorifera essential oil and Nardostachys raphanus essential oil in the present invention greatly improves the antidepressant effect of the Chinese medicine composite essential oil microemulsion gel, and the various components in the Chinese medicine composite essential oil microemulsion gel of the present invention are synergistic and indispensable. The present invention provides reference and reference for the development of new dosage forms for the prevention and treatment of depression and the application of Chinese medicine essential oils.

[0162] The above embodiments are only used to illustrate the technical solutions of the present invention, rather than to limit the same. Although the present invention has been described in detail with reference to the aforementioned embodiments, those skilled in the art should understand that the technical solutions described in the aforementioned embodiments may still be modified, or some of the technical features may be replaced by equivalents. However, these modifications or replacements do not deviate the essence of the corresponding technical solutions from the spirit and scope of the technical solutions of the embodiments of the present invention.

Claims

1. A Chinese medicine composite essential oil microemulsion gel, characterized in that: The microemulsion gel is composed of the following components in weight percentage per 100 parts by weight: 6-9% of traditional Chinese medicine compound essential oil, 25-35% of surfactant, 12-16% of co-surfactant, 1-5% of moisturizer, 1-2% of gel matrix, and the balance is purified water.

2. The Chinese medicine composite essential oil microemulsion gel according to claim 1, characterized in that: The microemulsion gel is composed of the following components in weight percentage per 100 parts by weight: 7.98% of traditional Chinese medicine compound essential oil, 29.82% of surface active agent, 14.91% of cosurfactant, 3% of moisturizing agent, 1.5% of gel matrix, and the balance is purified water.

3. A Chinese medicine composite essential oil microemulsion gel according to claim 1 or claim 2, characterized in that: The traditional Chinese medicine composite essential oil consists of bupleurum essential oil, tangerine peel essential oil, chuanxiong essential oil, osmanthus fragrans essential oil and nardus odoratus essential oil in a mass ratio of 5-7:3-5:3-5:1-3:1-3.

4. A Chinese medicine composite essential oil microemulsion gel according to claim 1 or claim 2, characterized in that: The traditional Chinese medicine composite essential oil consists of bupleurum essential oil, tangerine peel essential oil, chuanxiong essential oil, osmanthus fragrans essential oil and nardus odoratus essential oil in a mass ratio of 6:4.5:4.5:1.5:1.

5.

5. A Chinese medicine composite essential oil microemulsion gel according to claim 1 or claim 2, characterized in that: The surfactant is one of polyoxyethylene 40 hydrogenated castor oil (RH40), Tween-80 (TW-80), Tween-20 (TW-20), Span-80, and caprylic / capric macrogol glyceride (Labrasol); preferably, the surfactant is TW-80.

6. The co-surfactant is one of Transcutol P, polyethylene glycol 400 (PEG400), 1,2-propylene glycol, anhydrous ethanol, and n-butanol; preferably, the co-surfactant is PEG400.

7. The moisturizing agent is one of glycerin, trehalose, hyaluronic acid, sorbitol and urea.

8. The gel matrix is ​​one of Carbomer 934 and Carbomer 940. Preferably, the gel matrix is ​​Carbomer 940.

9. The method for preparing a traditional Chinese medicine composite essential oil microemulsion gel according to claim 1, characterized in that: The microemulsion gel is prepared according to the following steps: (1) Preparation of Chinese herbal composite essential oil: 500 g each of bupleurum, dried orange peel, chuanxiong, osmanthus fragrans, and nard were finely ground and passed through a No. 2 sieve to obtain uniform powder; then, each powder was mixed with distilled water 9 times its volume, and each powder was placed in an electric heating mantle and soaked for 1 h to fully infiltrate; then, glass beads were added and connected to a volatile oil analyzer, and a reflux condenser was installed at the upper end, and the water was heated to gradually boil and maintain a slightly boiling state; after 6 h of continuous extraction, when the volume of the volatile oil no longer increased, the heating was stopped and the system was allowed to cool naturally for 30 min; then, the volume of the volatile oil was recorded and collected; To ensure purity, the volatile oil is dried using anhydrous sodium sulfate; the dried volatile oil is a yellow oily liquid, which is stored in a brown bottle and placed in a refrigerator at 4°C for subsequent use; Bupleurum essential oil, tangerine peel essential oil, Chuanxiong essential oil, Murraya osmanthus essential oil, and Nardostachys grandiflora essential oil are uniformly mixed according to a mass ratio of 5-7:3-5:3-5:1-3:1-3 to obtain a traditional Chinese medicine composite essential oil; (2) uniformly mixing the Chinese medicine composite essential oil, surfactant and co-surfactant according to the above mass percentages, slowly adding water under magnetic stirring, and continuously stirring for more than 40 minutes to obtain a microemulsion solution for standby use; (3) adding a gel matrix to the microemulsion solution obtained in step (2), mixing evenly, allowing it to fully swell overnight, adjusting the pH to 6-7 with triethanolamine, then adding a moisturizer, and mixing evenly to obtain the traditional Chinese medicine composite essential oil microemulsion gel.

10. Use of the traditional Chinese medicine composite essential oil microemulsion gel as claimed in claim 1 in preparing a preparation for improving depression.

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