Traditional Chinese medicine compound essential oil and application thereof in preparation of anti-depression product

Abstract

The application provides a traditional Chinese medicine compound essential oil and application thereof in preparation of an antidepressant product, and relates to the technical field of traditional Chinese medicine essential oil pharmaceuticals. The traditional Chinese medicine compound essential oil is composed of benzoin essential oil 3-15 parts, Chinese rose flower essential oil 2-10 parts, immature bitter orange fruit essential oil 2-10 parts, white cardamom essential oil 1-8 parts and fennel essential oil 1-8 parts. The traditional Chinese medicine compound essential oil is administered in the form of sniffing, stimulates olfactory nerves, influences the cerebral cortex margin and neurotransmitters, and realizes improvement of brain depression-like symptoms. After the CUMS depression animal model is treated with the traditional Chinese medicine compound essential oil, the symptoms of loss of pleasure are reversed and improved, the state of reduced movement of the animal is improved, and the behavior state of the depression animal model of despair and giving up struggle is obviously improved. Meanwhile, the levels of DA, 5-HT and NE in the brain tissue of the depression animal model are significantly improved, and the pathological changes in the hippocampus region of the depression animal model are obviously improved. The traditional Chinese medicine compound essential oil can be used for preparing drugs for aromatherapy, has no adverse reactions, has high acceptability, and provides a new way for the treatment of depression.

Description

Technical Field

[0001] This invention relates to the field of pharmaceutical technology of traditional Chinese medicine essential oils, specifically to a compound essential oil of traditional Chinese medicine and its application in the preparation of antidepressant products. Background Technology

[0002] With the continuous development of society, people's life pressure is increasing, and the incidence of depression is gradually rising. As a clinical type of affective disorder, depression is specifically manifested as difficulty sleeping, loss of interest, decreased appetite, and mood instability. In the treatment of depression, most of the antidepressants used clinically are synthetic antidepressants. First-generation drugs of this class have been gradually abandoned due to safety issues, and second-generation synthetic antidepressants have become the norm. However, second-generation drugs still have drawbacks such as slow onset of action and relatively singular target. These factors have prompted scientists to develop third-generation antidepressants that are more targeted, more effective, and safer. However, current third-generation antidepressants still have problems such as insufficient efficacy and numerous adverse reactions in both short-term and long-term treatment. Therefore, the search for natural antidepressants with less toxicity and higher efficacy has become a new research hotspot.

[0003] In traditional Chinese medicine, aromatic herbs are collectively referred to as aromatic drugs. Aromatic drugs constitute a significant proportion of Chinese medicinal materials and play an indispensable role in clinical treatment. Utilizing aromatic drugs as a medium can improve traditional aromatherapy by preparing aromatic substances into inhalers, essential oils, etc., which can be absorbed into the body through respiration or skin to relieve stress, treat diseases, and promote health.

[0004] Volatile oils are a general term for oily substances with volatile aromatic odors, also known as essential oils. Most aromatic traditional Chinese medicines contain volatile oil components. Studies have shown that aromatic traditional Chinese medicines regulate bodily functions by influencing emotions and psychological states. When medicinal plant volatile oils are inhaled into the nasal cavity, some are absorbed into the bloodstream, while others combine with olfactory receptors in the olfactory cells and are transmitted to the cerebral cortex via the olfactory nerve, producing either a soothing or stimulating effect, thus alleviating depressive symptoms. Currently, research on the intervention of medicinal plant volatile oils on depression mainly focuses on the application of volatile oils extracted from single plants, with few reports on compound essential oils in traditional Chinese medicine for antidepressant purposes. Summary of the Invention

[0005] (a) Technical problems to be solved

[0006] To address the shortcomings of existing technologies, this invention provides a traditional Chinese medicine compound essential oil and its application in the preparation of antidepressant products.

[0007] (II) Technical Solution

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

[0009] First, the present invention provides a traditional Chinese medicine compound essential oil, which is composed of the following parts by volume of traditional Chinese medicine plant essential oils: 3-15 parts of benzoin essential oil, 2-10 parts of rose essential oil, 2-10 parts of immature bitter orange essential oil, 1-8 parts of cardamom essential oil, and 1-8 parts of fennel essential oil.

[0010] Preferably, the herbal compound essential oil is composed of the following parts by volume of herbal plant essential oils: 5-10 parts of benzoin essential oil, 4-8 parts of rose essential oil, 4-8 parts of immature bitter orange essential oil, 1-5 parts of cardamom essential oil, and 1-5 parts of fennel essential oil.

[0011] Preferably, the herbal compound essential oil is composed of the following parts by volume of herbal plant essential oils: 6 parts benzoin essential oil, 5 parts rose essential oil, 5 parts immature bitter orange essential oil, 3 parts cardamom essential oil, and 3 parts fennel essential oil.

[0012] The above-mentioned single-herb Chinese herbal essential oils are obtained by distillation or supercritical extraction using the corresponding Chinese herbal plants as raw materials, and the purity of the essential oils is above 99%.

[0013] The essential oils of the aforementioned single Chinese medicinal herbs can also be purchased commercially.

[0014] Secondly, the present invention also provides the application of the above-mentioned compound essential oil of traditional Chinese medicine in the preparation of antidepressant products. The antidepressant products are products used in aromatherapy. Specifically, each single herbal essential oil is weighed according to the above-mentioned volume proportions, mixed evenly, and then medically acceptable excipients are added to prepare a dosage form for administration by inhalation.

[0015] The antidepressant product contains 5-15% effective content of traditional Chinese medicine compound essential oil.

[0016] Benzoin is the dried resin of *Styrax tonkinensis* (Pierre) Craib ex Hart., a plant in the Styraxaceae family. It has a pungent, aromatic, neutral, and non-toxic taste, and is used to clear the mind, promote blood circulation, and relieve pain. It is used for stroke with phlegm syncope, sudden syncope due to qi stagnation, coma due to sudden illness, abdominal pain, postpartum hemorrhage, and infantile convulsions. Benzoin essential oil mainly contains benzyl benzoate, benzyl cinnamate, vanillin, and styraxin, among other components. Modern pharmacological studies have shown that benzoin essential oil has antibacterial, anti-inflammatory, expectorant, skin-repairing, and mood-regulating effects.

[0017] Rose, or Chinese rose, is the dried flower of *Rosa chinensis* Jacq., a plant in the Rosaceae family. It has a sweet taste and warm nature, and enters the liver meridian; it has the effects of promoting blood circulation, regulating menstruation, soothing the liver, and relieving depression. It is commonly used for qi stagnation and blood stasis, irregular menstruation, dysmenorrhea, amenorrhea, and chest and rib pain. Rose essential oil mainly contains terpenoids, aromatic compounds, and aliphatic compounds, and has pharmacological effects such as antibacterial and anti-inflammatory properties, antioxidant effects, immunomodulatory effects, cardiovascular and neuroprotective effects, antitumor effects, and mood-regulating effects.

[0018] Citrus aurantium L., the dried young fruit of the Rutaceae plant Citrus aurantium L., is bitter, pungent, and sour, slightly cold in nature. It enters the spleen and stomach meridians and has the effects of breaking up stagnation, resolving phlegm, and dispersing masses. It is used for symptoms such as internal stagnation, abdominal distension and pain, tenesmus after diarrhea, constipation, phlegm stagnation and qi obstruction, chest pain, chest tightness, and organ prolapse. The essential oil of Citrus aurantium mainly contains monoterpenes, sesquiterpenes, aldehydes, ketones, and esters. Modern pharmacological studies have shown that it has pharmacological effects such as regulating the digestive system, antibacterial and anti-inflammatory properties, antioxidant effects, anti-aging effects, cardiovascular protection, and immune regulation.

[0019] White cardamom is the dried, ripe fruit of *Amomum kravanh Picrre cx Gagnep.*, a plant in the ginger family. It has a pungent and warm nature, and enters the lung, spleen, and stomach meridians. It has the effects of resolving dampness and promoting qi circulation, warming the middle jiao and stopping vomiting, and stimulating appetite and digestion. It is mainly used for symptoms such as dampness obstructing qi, spleen and stomach disharmony, abdominal distension, loss of appetite, early stages of damp-heat syndrome, chest tightness and lack of hunger, stomach cold and vomiting, and indigestion. White cardamom essential oil mainly contains monoterpenes, sesquiterpenes, esters, and ethers. Modern pharmacological studies have shown that it has pharmacological effects such as regulating the digestive system, antibacterial and anti-inflammatory properties, antioxidant effects, anti-aging effects, regulating immune function, and improving the respiratory system.

[0020] Fennel, the dried, ripe fruit of *Foeniculum vdgare* Mill., a plant in the Apiaceae family, has a pungent and warm nature, and enters the liver, kidney, spleen, and stomach meridians. It has the effects of dispelling cold and relieving pain, regulating qi and harmonizing the stomach, and is used for cold-induced abdominal pain, testicular swelling, dysmenorrhea, lower abdominal pain due to cold, abdominal distension and pain, poor appetite, vomiting, and diarrhea. Fennel essential oil mainly contains monoterpenes, sesquiterpenes, aldehydes, and ketones. Modern pharmacological studies have shown that it has pharmacological effects such as regulating the digestive system, antibacterial and anti-inflammatory properties, antioxidant effects, anti-aging effects, improving the respiratory system, regulating immune function, and regulating endocrine function.

[0021] (III) Beneficial Effects

[0022] This invention, based on the principles of TCM syndrome differentiation and treatment, targets the pathogenesis of depression. Following the principles of principal, assistant, adjuvant, and guide herbs, it uses benzoin as the principal herb. Its pungent and bitter properties can soothe the liver, relieve depression, and awaken the mind, directly addressing the core pathogenesis of depression: qi stagnation and phlegm obstructing the orifices. Rose, sweet and warm in nature, has the effects of invigorating blood, regulating menstruation, soothing the liver, and regulating qi, assisting the principal herb in enhancing its qi-regulating and depression-relieving effects, and promoting the smooth flow of qi and blood to relieve liver depression. Immature bitter orange breaks up qi stagnation, eliminates accumulations, and resolves phlegm, enhancing its qi-regulating and phlegm-resolving effects, and eliminating phlegm obstruction. These two herbs serve as assistant herbs. Fennel is added to warm the middle jiao and regulate qi, guiding the medicine to the liver meridian and enhancing its liver-soothing and depression-relieving effects. White cardamom is used as the guide herb to resolve dampness, regulate qi, and harmonize the spleen and stomach, preventing the pungent and aromatic qi-regulating herbs from damaging the spleen and stomach, and harmonizing the liver and spleen through "nourishing the earth and nourishing the wood." The entire formula follows the principle of "soothing the liver, regulating qi, resolving phlegm, and strengthening the spleen." Benzoin relieves stagnation and refreshes the mind, while rose softens the liver, immature bitter orange breaks up stagnation, fennel warms the middle jiao, and cardamom harmonizes the stomach. This aligns with the "qi stagnation first, while also addressing phlegm and dampness" principle in the Six Depressions Theory of Danxi's Heart Method, and also reflects the characteristic treatment of depression in "Clinical Guidelines and Medical Cases," which emphasizes both "regulating qi" and "strengthening the middle jiao." The combination of these herbs works synergistically to soothe the liver, relieve stagnation, strengthen the spleen, and calm the mind, making it suitable for depression caused by qi stagnation, phlegm obstruction, and malnourishment of the mind. Based on the above formula, volatile oil components are extracted from benzoin, rose, immature bitter orange, cardamom, and fennel to create a compound essential oil with the effects of soothing the liver, relieving stagnation, strengthening the spleen, and calming the mind. This compound essential oil is administered via inhalation, stimulating the olfactory nerves to affect the limbic system and neurotransmitters, thereby improving depressive-like symptoms. The compound essential oil of traditional Chinese medicine in this invention can be used to prepare medicines for aromatherapy. It has no adverse reactions, is highly acceptable, and provides a new approach for the treatment of depression.

[0023] Animal experiments showed that after treatment with the herbal compound essential oil of this invention, the anhedonia symptoms in the CUMS animal depression model were significantly reversed and improved. The reduced movement and the despairing, resigned behavior of the depressed animal models were also significantly improved. Simultaneously, the levels of DA, 5-HT, and NE in the brain tissue of the depressed animal models were significantly increased, and the pathological changes in the hippocampus of the depressed animal models were significantly improved. Furthermore, the herbal compound essential oil of this invention showed better therapeutic effects on the depressed animal models than Western medicine. This herbal compound essential oil of this invention holds promise as a new drug for aromatherapy treatment of depression. Attached Figure Description

[0024] Figure 1 HE staining images of hippocampal tissue from rats in each group, HE×400. Detailed Implementation

[0025] To make the objectives, technical solutions, and advantages of the embodiments of the present invention clearer, the technical solutions of the embodiments of the present invention will be clearly and completely described below in conjunction with the embodiments of the present invention. Obviously, the described embodiments are only some embodiments of the present invention, not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0026] Example 1

[0027] A compound essential oil made from traditional Chinese medicine for the purpose of treating depression, wherein the compound essential oil is composed of the following parts by volume of essential oils from traditional Chinese medicinal plants: 3 parts benzoin essential oil, 2 parts rose essential oil, 2 parts immature bitter orange essential oil, 1 part cardamom essential oil, and 1 part fennel essential oil.

[0028] The above-mentioned single-herb Chinese herbal essential oils are obtained by distillation or supercritical extraction using the corresponding Chinese herbal plants as raw materials, and the purity of the essential oils is above 99%.

[0029] Weigh out the essential oils of each single Chinese herbal medicine according to the above volume proportions, mix them evenly, add medically acceptable excipients, and prepare a dosage form for inhalation administration. The effective content of the compound essential oils is 5-15%.

[0030] Example 2

[0031] The difference between this embodiment and Embodiment 1 is that the herbal compound essential oil is composed of the following volume parts of herbal plant essential oils: 13 parts benzoin essential oil, 9 parts rose essential oil, 9 parts immature bitter orange essential oil, 6 parts cardamom essential oil, and 6 parts fennel essential oil.

[0032] Example 3

[0033] The difference between this embodiment and Embodiment 1 is that the compound essential oil of traditional Chinese medicine is composed of the following volume parts of Chinese herbal plant essential oils: 4 parts benzoin essential oil, 3 parts rose essential oil, 3 parts immature bitter orange essential oil, 2 parts white cardamom essential oil, and 1 part fennel essential oil.

[0034] Example 4

[0035] The difference between this embodiment and Embodiment 1 is that the herbal compound essential oil is composed of the following volume parts of herbal plant essential oils: 10 parts benzoin essential oil, 6 parts rose essential oil, 6 parts immature bitter orange essential oil, 4 parts cardamom essential oil, and 3 parts fennel essential oil.

[0036] Example 5

[0037] The difference between this embodiment and Embodiment 1 is that the compound Chinese herbal essential oil is composed of the following volume parts of Chinese herbal plant essential oils: 5 parts benzoin essential oil, 4 parts rose essential oil, 4 parts immature bitter orange essential oil, 1 part cardamom essential oil, and 1 part fennel essential oil.

[0038] Example 6

[0039] The difference between this example and Example 1 is that the traditional Chinese medicine compound essential oil is composed of the following essential oils of traditional Chinese medicine plants by volume: 10 parts of benzoin essential oil, 8 parts of Chinese rose essential oil, 8 parts of bitter orange essential oil, 5 parts of amomum kravanh essential oil, and 5 parts of fennel essential oil.

[0040] Example 7

[0041] The difference between this example and Example 1 is that the traditional Chinese medicine compound essential oil is composed of the following essential oils of traditional Chinese medicine plants by volume: 7 parts of benzoin essential oil, 5 parts of Chinese rose essential oil, 5 parts of bitter orange essential oil, 2 parts of amomum kravanh essential oil, and 3 parts of fennel essential oil.

[0042] Example 8

[0043] The difference between this example and Example 1 is that the traditional Chinese medicine compound essential oil is composed of the following essential oils of traditional Chinese medicine plants by volume: 8 parts of benzoin essential oil, 6 parts of Chinese rose essential oil, 4 parts of bitter orange essential oil, 3 parts of amomum kravanh essential oil, and 2 parts of fennel essential oil.

[0044] Example 9

[0045] The difference between this example and Example 1 is that the traditional Chinese medicine compound essential oil is composed of the following essential oils of traditional Chinese medicine plants by volume: 6 parts of benzoin essential oil, 5 parts of Chinese rose essential oil, 5 parts of bitter orange essential oil, 3 parts of amomum kravanh essential oil, and 3 parts of fennel essential oil.

[0046] Test Example 1

[0047] Effects of traditional Chinese medicine compound essential oils with different components on depressive-like behaviors in mice

[0048] 1 Experimental materials

[0049] 1.1 Experimental animals

[0050] 60 KM mice, all male, weighing 19 - 22 g, SPF grade. Animal certificate number: No.370726211100923115; provided by Jinan Pengyue Experimental Animal Breeding Co., Ltd., Shandong Province, license number SCXK(Shandong)20190003; laboratory use license number: SYXK(Henan)2020 - 0004. Raised in the Animal Experiment Center of Henan University of Traditional Chinese Medicine, unit use license number: SYXK(Henan)2020 - 0004.

[0051] 1.2 Main drugs

[0052] The traditional Chinese medicine compound essential oil of the present invention: 6 mL of benzoin essential oil, 5 mL of Chinese rose essential oil, 5 mL of bitter orange essential oil, 3 mL of amomum kravanh essential oil, 3 mL of fennel essential oil

[0053] Comparison of essential oil blend 1: 6mL benzoin essential oil, 3mL white cardamom essential oil, 3mL fennel essential oil

[0054] Comparison of essential oil blend 2: 5mL rose essential oil, 5mL bitter orange essential oil, 3mL white cardamom essential oil, 3mL fennel essential oil

[0055] Comparison of essential oil compound 3: 6mL benzoin essential oil, 5mL rose essential oil, 5mL bitter orange essential oil

[0056] The essential oils mentioned above were prepared by supercritical carbon dioxide extraction using benzoin, rose, immature bitter orange, white cardamom, and fennel as raw materials. The purity of the essential oils is above 99%. After accurately weighing the ingredients according to the above volume proportions, they are mixed evenly to obtain the final product.

[0057] 2 Experimental Methods

[0058] 2.1 Model Preparation

[0059] Ten male SPF-grade ICR mice were randomly selected as the control group. The remaining 60 mice were used to establish a CUMS (Chronic Unpredictable Mild Stress) depression model using a combination of isolation and chronic unpredictable mild stress. Specifically, ten different stimuli were selected: cage tilt (24 h), damp bedding (24 h), reversed day-night cycle, swimming in ice water (4℃, 5 min), fasting (24 h), water deprivation (24 h), heat stimulation (50℃, 5 min), tail clamping (1 min), electric shock to the soles of the feet (50 mV, once every 30 s, each lasting 10 s, for a total of 15 times), and noise stimulation (100 Hz, 2 h). One stimulus was administered daily in a randomized order, and each stimulus was not used consecutively for two days to ensure the animals could not predict the occurrence of the stimulus. The stress lasted for 6 weeks. The control group received no stimulation.

[0060] 2.2 Experimental grouping and drug administration

[0061] Ten mice were randomly selected as the control group before CUMS stimulation. Six weeks after CUMS treatment, mice with abnormal sucrose preference rates were excluded based on the sucrose preference rate experiment results. Fifty mice with successful model establishment and good condition were then randomly divided into five groups of ten each.

[0062] Model group: No drug treatment was given, only pure water was administered orally;

[0063] Experimental group 1: The traditional Chinese medicine compound essential oil of this invention, dosage is 5μL / 20g;

[0064] Experimental group 2: Control compound essential oil 1, dosage was 5μL / 20g;

[0065] Experimental group 3: Control compound essential oil 2, dosage was 5μL / 20g;

[0066] Experimental group 4: Control compound essential oil 3, dosage was 5μL / 20g;

[0067] All the above compound essential oils were administered via inhalation. For inhalation, mice were first placed in a self-made sealed inhalation chamber (45×30×25cm). Then, the compound essential oils of this invention were mixed with 5mL of distilled water according to the above dosage and placed in a nebulizer cup. The air compressor pump was started to atomize the essential oil solution into fine mist particles. Timing began after the mist had diffused throughout the entire inhalation chamber, and the inhalation administration lasted for 30 minutes. Each treatment group began administration after the completion of CUMS, once daily for 2 weeks, i.e., days 43-57 of the experiment.

[0068] 2.3 Neurobehavioral index detection

[0069] Neurobehavioral studies were conducted on mice in each group before and after the experiment. The specific tests are shown below:

[0070] (1) Sugar water preference test (SPT)

[0071] A continuous sucrose preference test was conducted on mice. Mice were trained to drink sucrose for two days prior to the experiment. After a 24-hour fast (no food or water), the test was performed. The experiment began at 9:00 AM each day. Detailed procedures are shown in the table below.

[0072] Table 2 Experimental Design of Sugar Water Preference Rate

[0073]

[0074]

[0075] (2) Open field test (OFT)

[0076] The OFT rat open field activity experiment system was used to observe the movement distance and movement time of mice in each group within 5 minutes (open field size: 50×50×40cm).

[0077] (3) Tail suspension test (TST)

[0078] A mouse tail suspension video analysis system was used to suspend mice in the tail suspension device for 6 minutes and record the time the mice remained still for the next 4 minutes.

[0079] 2.4 Statistical Methods

[0080] Data analysis was performed using SPSS 25.0 software for statistical processing. Quantitative data were expressed as mean ± standard deviation (x ± s). One-way ANOVA was used for comparisons between groups; Games-Howell's test was used for unequal variances, and LSD test was used for homogeneous variances. Ridit's test was used for ordinal data.

[0081] 3. Results: Effects of the herbal relapse essential oil of this invention on the neurobehavioral characteristics of the CUMS mouse depression model.

[0082] Table 1 shows the results of sucrose preference rates in each group of mice before and after treatment. Before treatment, compared with the control group, the sucrose preference rates in all model groups were significantly reduced (P<0.01), and there was no significant difference in sucrose preference rates among the model groups (P>0.05). After treatment, compared with the control group, the sucrose preference rate in the model group was significantly reduced (P<0.01); compared with the model group, the sucrose preference rates in experimental groups 1-4 were significantly increased (P<0.01), with the increase in sucrose preference rate in experimental group 1 (the compound essential oil of the traditional Chinese medicine in this invention) being significantly higher than that in experimental groups 2-3.

[0083] Table 1. Sugar water preference rate of mice in each group before and after treatment.

[0084] Blank group 81.2±3.6 81.4±2.7 Model group <![CDATA[62.4±3.9 △△ ]]> <![CDATA[65.7±2.6 △△ ]]> Experimental group 1 <![CDATA[62.3±3.1 △△ ]]> <![CDATA[75.1±2.8 ** ]]> Experimental group 2 <![CDATA[62.4±2.8 △△ ]]> <![CDATA[69.8±2.6 ** ]]> Experimental group 3 <![CDATA[62.6±2.8 △△ ]]> <![CDATA[69.1±2.4 ** ]]> Experimental group 4 <![CDATA[62.5±2.9 △△ ]]> <![CDATA[70.2±3.0 ** ]]>

[0085] Note: Compared with the blank group, △△ P<0.01, △ P < 0.05; compared with the model group, **P < 0.01, *P < 0.05

[0086] Table 2 shows the results of open field tests for each group of mice before and after treatment. Before treatment, compared with the control group, the movement distance and movement time of mice in each model group were significantly reduced (P<0.01), and there was no significant difference in movement distance and movement time among the model groups (P>0.05). After treatment, compared with the model group, experimental group 1 (the compound essential oil of the traditional Chinese medicine of this invention) significantly increased the total movement distance of mice (P<0.01), experimental group 4 (control compound essential oil 3) significantly increased the total movement distance of mice (P<0.05), and experimental groups 2 and 3 only showed a trend of increasing the movement distance of mice (P>0.05); experimental group 1 significantly prolonged the movement time of mice (P<0.01), and experimental groups 2 and 4 significantly prolonged the movement time of mice (P<0.05).

[0087] Table 2 Results of open field test in mice before and after treatment.

[0088]

[0089] Note: Compared with the blank group, △△P < 0.01, △ P < 0.05; compared with the model group, **P < 0.01, *P < 0.05

[0090] Table 3 shows the test results of the tail suspension test for mice in each group. Before treatment, compared with the blank group, the immobile time of each model group was significantly prolonged (P < 0.01), and there was no significant difference between the model groups (P > 0.05). Compared with the model group, Test Group 1 could significantly shorten the immobile time of mice (P < 0.01), Test Group 4 could significantly shorten the immobile time of mice (P < 0.05), and Test Groups 2 and 3 only had a tendency to shorten the immobile time of mice (P > 0.05).

[0091] Table 3 shows the test results of the tail suspension test for mice in each group

[0092] Blank group 115.5±22.3 124.8±18.3 Model group <![CDATA[177.5±19.4 △△ ]]> <![CDATA[183.2±16.0 △△ ]]> Experimental group 1 <![CDATA[177.1±19.5 △△ ]]> <![CDATA[142.9±21.7 ** ]]> Experimental group 2 <![CDATA[177.2±18.8 △△ ]]> 161.2±23.1 Experimental group 3 <![CDATA[177.5±19.1 △△ ]]> 165.9±22.5 Experimental group 4 <![CDATA[177.3±19.4 △△ ]]> <![CDATA[154.3±20.9 * ]]>

[0093] Note: Compared with the blank group, △△ P < 0.01, △ P < 0.05; compared with the model group, **P < 0.01, *P < 0.05

[0094] 4 Conclusions

[0095] After the CUMS mouse depression model was treated with the traditional Chinese medicine compound essential oil of the present invention, the symptoms of anhedonia were reversed and improved, and the state of reduced movement in mice and the behavioral states of despair and giving up struggling in the depression model mice could be significantly improved; the improvement effect of the traditional Chinese medicine compound essential oil of the present invention on the depressive-like behavior of mice was significantly better than that of other compound essential oils, and each component of the traditional Chinese medicine compound essential oil of the present invention cooperated with each other and synergistically enhanced the effect. The traditional Chinese medicine compound essential oil of the present invention is expected to become a new drug for the treatment of depression by aromatherapy

[0096] Experimental Example 2

[0097] Effect of the traditional Chinese medicine compound essential oil of the present invention on the CUMS-induced rat depression model

[0098] 1 Experimental materials

[0099] 1.1 Experimental animals

[0100] Male, SD rats, SPF grade, weighing 180 - 200 g, provided by Jinan Pengyue Experimental Animal Breeding Co., Ltd., license number: SCXK(Shandong)20190003; animal certificate number: 1107262011002341; the animals were raised in the Animal Experiment Center of Henan University of Traditional Chinese Medicine, unit use license number: SYXK(Henan)2020 - 0004

[0101] 1.2 Main drugs

[0102] The compound essential oil of traditional Chinese medicine in this invention consists of 6 mL of benzoin essential oil, 5 mL of rose essential oil, 5 mL of immature bitter orange essential oil, 3 mL of cardamom essential oil, and 3 mL of fennel essential oil. The above essential oils are prepared by supercritical carbon dioxide extraction using benzoin, rose, immature bitter orange, cardamom, and fennel as raw materials. The purity of the essential oils is above 99%. After accurately weighing the ingredients according to the above volume proportions, they are mixed evenly to obtain the final product.

[0103] Fluoxetine hydrochloride (manufacturer: Changzhou No. 4 Pharmaceutical Co., Ltd., product batch number: 20201110).

[0104] 2 Experimental Design and Methods

[0105] 2.1 Experimental Design

[0106] After purchasing, all rats were acclimatized for 3 days, then kept in solitary confinement for 2 weeks, followed by 6 weeks of (solitary confinement + CUMS) modeling. Drug intervention lasted for 4 weeks. After sampling, indicators were measured. The sucrose preference rate of the rats was measured every 2 weeks. Before sampling, the sucrose preference rate test, open field test, and forced swimming test were performed in sequence. All behavioral tests started at 9:00 am.

[0107] 2.2 Experimental Methods

[0108] 2.2.1 Model Preparation

[0109] Ten male SD rats (180-200g, SPF grade) were randomly selected as the control group, and the remaining 40 rats were used to establish the CUMS model. Before CUMS, each rat was kept in solitary confinement for 2 weeks. CUMS (continued solitary confinement) began in the third week, with different stimuli given randomly each day. Each stimulus was not to be given consecutively. Ten different stimulation methods were randomly applied over 6 weeks to establish the CUMS depression model. The ten stimuli included: ① fasting (24h), ② water deprivation (24h), ③ reversed day and night cycle, ④ damp bedding, ⑤ heat stress (45℃, 5min), ⑥ swimming in ice water (4℃, 5min), ⑦ tail clamping (1min), ⑧ noise stimulation (100Hz, 2h), ⑨ odor stimulation (appropriate amount of pepper powder, 12h), and ⑩ flashing stimulation (high-frequency flashing of colored lights, 12h).

[0110] 2.2.2 Experimental Grouping and Processing

[0111] The rats that were successfully modeled were divided into 4 groups, with 10 rats in each group, as follows:

[0112] Model group: administered pure water orally;

[0113] Fluoxetine hydrochloride group: dose was 3.33 mg / kg, concentration was 0.333 mg / ml;

[0114] The high-dose group of the compound essential oil of traditional Chinese medicine in this invention (hereinafter referred to as the high-dose group): the dosage is 100 μL / kg;

[0115] The low-dose group of the compound essential oil of traditional Chinese medicine in this invention (hereinafter referred to as the low-dose group): the dosage is 50 μL / kg;

[0116] A separate control group was set up: no CUMS modeling was performed, and the control group was given pure water.

[0117] Dosing began on week 4 and continued for a total of 4 weeks, from days 28 to 56 of the experiment. In the rat experiment, fluoxetine hydrochloride was converted to 10 times the clinical dosage, and the dosage of the herbal compound essential oil in this invention was determined based on the results of preliminary experiments. Rats were weighed weekly, and the dosage was adjusted according to their body weight.

[0118] The herbal compound essential oil of this invention is administered by inhalation. During inhalation, the rat is first placed in a self-made sealed inhalation box (45×30×25cm). Then, the herbal compound essential oil of this invention is mixed with 5mL of distilled water according to the above dosage and placed in the nebulizer cup. The air compressor pump is started to atomize the essential oil solution into fine mist particles. The timing begins after the mist has diffused throughout the entire inhalation box. The inhalation administration lasts for 30 minutes.

[0119] 2.3 Observation Indicators and Detection

[0120] 2.3.1 Behavioral Indicator Testing

[0121] (1) Sugar water preference experiment (SPT)

[0122] The experimental procedure is the same as in Experiment Example 1, and the test time is 1 hour.

[0123] (2) Open Field Experiment (OFT)

[0124] The experimental procedure was the same as in Experiment 1, using a rat open field test system (100×100×40cm) for 5 minutes.

[0125] (3) Forced Swimming Experiment (FET)

[0126] Rats were placed in a forced swimming tub (21×21×50cm, water depth approximately 33cm) at a water temperature of approximately 25℃ for 6 minutes. The duration of immobility was recorded for the last 4 minutes. Behavioral criteria: Swimming—Rats moved around in the water; Climbing—Rats paddled the water surface with their forepaws; Struggling—Rats struggled violently with their limbs / the forepaws remained still while the hind limbs continued to paddle; Immobility—Rats floated on the water surface with their limbs still / occasionally slid their limbs, with only their heads above the water surface to breathe.

[0127] 2.3.2 Biochemical index detection

[0128] The rats were fasted for 12 hours the night before collection, but allowed free access to water. One hour after the last administration of medication, they were weighed. The rats were euthanized by cervical dislocation and dissected. The brains were collected, and a 10% brain tissue homogenate was prepared. Enzyme-linked immunosorbent assay (ELISA) was used to detect the levels of 5-HT, DA, and NE in the brain homogenate.

[0129] 2.3.3 Histopathological examination

[0130] HE staining was used to detect histopathological changes in the hippocampus of rats in each group.

[0131] 2.4 Statistical Analysis

[0132] Data analysis was performed using SPSS 25.0 software for statistical processing. Quantitative data were expressed as mean ± standard deviation (x ± s). One-way ANOVA was used for comparisons between groups; Games-Howell's test was used for unequal variances, and LSD test was used for homogeneous variances. Ridit's test was used for ordinal data.

[0133] 3 Results

[0134] 3.1 Effects of the herbal relapse essential oil of this invention on the behavior of a CUMS-induced rat depression model

[0135] 3.1.1 Effect on body weight of CUMS-induced rat depression model

[0136] The results are shown in Table 4. Compared with the blank group, the body weight of rats in the model group was significantly reduced (P<0.01); compared with the model group, the body weight of rats in the high-dose group was significantly increased (P<0.01), and the low-dose group showed a trend of increasing body weight (P>0.05).

[0137] Table 4. Changes in body weight of rats before and after the experiment in each group.

[0138] Blank group 299.1±5.4 381.4±13.0 Model group <![CDATA[281.2±8.8 △△ ]]> <![CDATA[336.9±21.6 △△ ]]> Fluoxetine hydrochloride group <![CDATA[281.8±6.8 △△ ]]> 343.2±20.8 High-dose group <![CDATA[281.0±7.2 △△ ]]> <![CDATA[360.9±19.9 ** ]]> low-dose group <![CDATA[282.2±7.3 △△ ]]> 352.3±20.3

[0139] Note: Compared with the blank group, △△ P<0.01, △ P < 0.05; compared with the model group, **P < 0.01, *P < 0.05

[0140] 3.1.2 Effect on sucrose preference rate in a CUMS-induced rat model of depression

[0141] For each group of SD rats, the sucrose preference rate was measured every two weeks, and the results are shown in Table 5. At week 2 of intervention, compared with the control group, the sucrose preference rate in the model group and each drug-treated group was significantly reduced (P<0.01). At week 4 of intervention, compared with the control group, the sucrose preference rate in the model group was significantly reduced (P<0.01); compared with the model group, the sucrose preference rate in the fluoxetine hydrochloride group, high-dose group, and low-dose group was significantly increased (P<0.01).

[0142] Table 5 Results of the sugar water preference test in each group of rats

[0143] Blank group 84.79±6.97 85.47±5.43 Model group <![CDATA[63.76±9.30 △△ ]]> <![CDATA[59.65±10.31 △△ ]]> Fluoxetine hydrochloride group <![CDATA[66.61±6.27 △△ ]]> <![CDATA[75.13±8.71 ** ]]> High-dose group <![CDATA[69.33±5.61 △△ ]]> <![CDATA[75.39±6.20 ** ]]> low-dose group <![CDATA[67.84±5.78 △△ ]]> <![CDATA[73.56±6.29 ** ]]>

[0144] 3.1.3 Effects on the open field test of the CUMS-induced rat depression model

[0145] On the mornings of days 52-53 (days 24-25 after drug administration), open field tests were conducted on rats in each group using a 100×100×40cm rat exercise box. The rats' movement was assessed by recording the distance and time of movement over 5 minutes. The results are shown in Table 6. Compared with the control group, the model group showed significantly reduced movement distance and time (P<0.01). Compared with the model group, the high- and low-dose groups significantly prolonged the rat movement distance (P<0.05), and the fluoxetine hydrochloride group significantly prolonged the rat movement distance (P<0.01). Compared with the model group, the high-dose group and the fluoxetine hydrochloride group significantly increased the rat movement time (P<0.01), and the low-dose group significantly increased the rat movement time (P<0.01).

[0146] Table 6 Results of open field test for rats in each group

[0147] Blank group 3190.3±236.6 264.6±14.3 Model group <![CDATA[2501.4±405.9 △△ ]]> <![CDATA[237.6±19.7 △△ ]]> Fluoxetine hydrochloride group <![CDATA[3159.0±347.2 ** ]]> <![CDATA[257.0±13.7 * ]]> High-dose group <![CDATA[2911.9±375.5 * ]]> <![CDATA[261.8±13.9 ** ]]> low-dose group <![CDATA[2897.4±408.8 * ]]> <![CDATA[258.6±13.7 * ]]>

[0148] 3.1.4 Effects on the forced swimming test in a CUMS-induced rat model of depression

[0149] On day 54 of the experiment (26 days after drug administration), rats in each group underwent forced swimming training for 15 minutes each. After a 24-hour rest period (day 27 after drug administration), the rats were formally tested. The results are shown in Table 7. Compared with the control group, the immobility time in the forced swimming test of rats in the model group was significantly prolonged (P<0.01). Compared with the model group, the high-dose group and the fluoxetine hydrochloride group significantly shortened the immobility time of rats (P<0.01), while the low-dose group significantly shortened the immobility time of rats (P<0.05). This suggests that the compound essential oil of traditional Chinese medicine in this invention can improve the desperate struggle state induced by CUMS in rats.

[0150] Table 7 Results of the forced swimming test in each group of rats

[0151] Blank group 62.5±14.4 Model group <![CDATA[139.3±22.2 △△ ]]> Fluoxetine hydrochloride group <![CDATA[107.5±17.4 ** ]]> High-dose group <![CDATA[105.9±24.9 ** ]]> low-dose group <![CDATA[111.4±26.2 * ]]>

[0152] 3.2 Effects of the compound essential oil of traditional Chinese medicine on the biochemical indicators of brain tissue in a CUMS-induced rat depression model

[0153] The results are shown in Table 8. Compared with the control group, the levels of DA, 5-HT, and NE in the brain tissue homogenate of rats in the model group were significantly decreased (P<0.01), indicating a reduction in monoamine neurotransmitters in the brain tissue of rats in the model group. Compared with the model group, the high-dose group, the low-dose group, and the fluoxetine hydrochloride group significantly increased the DA level in rat brain tissue (P<0.01); the high-dose group and the fluoxetine hydrochloride group significantly increased the 5-HT and NE levels in rat brain tissue (P<0.01), and the low-dose group significantly increased the 5-HT and NE levels in rat brain tissue (P<0.05).

[0154] Table 8. Levels of DA, 5-HT, and NE in rat brain tissue homogenates from each group.

[0155] Blank group 1483.8±119.7 131.59±5.62 122.17±6.72 Model group <![CDATA[1310.8±71.9 △△ ]]> <![CDATA[110.55±3.18 △△ ]]> <![CDATA[103.14±7.00 △△ ]]> Fluoxetine hydrochloride group <![CDATA[1486.2±97.0 ** ]]> <![CDATA[124.56±5.76 ** ]]> <![CDATA[118.68±3.90 ** ]]> High-dose group <![CDATA[1480.8±82.4 ** ]]> <![CDATA[126.48±6.08 ** ]]> <![CDATA[118.54±7.99 ** ]]> low-dose group <![CDATA[1428.1±84.7 ** ]]> <![CDATA[117.37±5.81 * ]]> <![CDATA[111.31±7.64 * ]]>

[0156] 3.3 Effects of the herbal relapse essential oil of this invention on pathological changes in the hippocampus of a CUMS-induced rat model of depression

[0157] The arrangement and distribution of neurons in the hippocampus of rats in each group, as well as the degree of cellular changes, were observed. Pathological tissues were graded according to semi-quantitative standards. The results are shown in Table 9. Pathological photographs are shown below. Figure 1 As shown in the figure. Ridit test revealed that, compared to the control group, the model group rats exhibited dispersed nerve cell arrangement, fewer cells, and showed signs of nerve cell indentation, deformation, cell lysis, blurred edges, and cytoplasmic staining in the hippocampus (P<0.01). Compared to the model group, both the high-dose group and the fluoxetine hydrochloride group significantly improved the CUMS-induced morphological and pathological changes in rat hippocampal neurons (P<0.01), while the low-dose group significantly improved the CUMS-induced morphological and pathological changes in rat hippocampal neurons (P<0.05).

[0158] Table 9. Pathological changes in the hippocampus of rats in each group.

[0159] Blank group 10 0 0 0 <0.01 Model group 0 1 5 4 - Fluoxetine hydrochloride group 1 8 1 0 <0.01 High-dose group 2 5 3 0 <0.01 low-dose group 1 4 5 0 <0.05

[0160] 4. Conclusion

[0161] The above animal experimental results show that after treatment with the compound essential oil of traditional Chinese medicine in the present invention, the weight of the CUMS depression model rats can be significantly increased, the symptoms of anhedonia can be reversed and improved, the reduced movement of rats and the despair and giving up behavior of the depression model rats can be significantly improved; at the same time, the levels of DA, 5-HT and NE in the brain tissue of the depression model rats are significantly increased, the pathological changes in the hippocampus of the depression model rats are significantly improved, and the therapeutic effect of the compound essential oil of traditional Chinese medicine in the present invention on the depression model rats is better than that of Western medicine.

[0162] The above embodiments are only used to illustrate the technical solutions of the present invention, and are not intended to limit it. Although the present invention has been described in detail with reference to the foregoing embodiments, those skilled in the art should understand that modifications can still be made to the technical solutions described in the foregoing embodiments, or equivalent substitutions can be made to some of the technical features. Such modifications or substitutions do not cause the essence of the corresponding technical solutions to deviate from the spirit and scope of the technical solutions of the embodiments of the present invention.

Claims

1. A compound essential oil made from traditional Chinese medicine, characterized in that, The herbal compound essential oil is composed of the following parts by volume of herbal plant essential oils: 6 parts benzoin essential oil, 5 parts rose essential oil, 5 parts immature bitter orange essential oil, 3 parts cardamom essential oil, and 3 parts fennel essential oil.

2. The traditional Chinese medicine compound essential oil as described in claim 1, characterized in that, Benzoin essential oil, rose essential oil, immature bitter orange essential oil, white cardamom essential oil, and fennel essential oil are extracted from the corresponding Chinese medicinal plants using distillation or supercritical extraction methods, with a purity of over 99%.

3. The application of the traditional Chinese medicine compound essential oil as described in claim 1 in the preparation of antidepressant products.

4. The application of the traditional Chinese medicine compound essential oil as described in claim 3 in the preparation of antidepressant products, characterized in that, The antidepressant product is an aromatherapy product, specifically made by weighing each herbal essential oil according to the above-mentioned volume proportions, mixing them evenly, adding medically acceptable excipients, and preparing a dosage form for administration by inhalation.

5. The application of the traditional Chinese medicine compound essential oil as described in claim 4 in the preparation of antidepressant products, characterized in that, The antidepressant product contains 5-15% effective content of traditional Chinese medicine compound essential oil.

Images