Traditional Chinese medicine cacumen biotae essential oil, preparation method thereof and application of antidepressant drug

Abstract

The invention provides traditional Chinese medicine cacumen biotae essential oil, a preparation method thereof and application of an antidepressant drug, and relates to the technical field of medicine. The traditional Chinese medicine cacumen biotae essential oil disclosed by the invention is prepared from the following components: alpha-pinene, gamma-terpinene, beta-terpilenol, alpha-terpilenol, gamma-terpilenol, alpha-cedrene, beta-cedrene, cis-thujopsene, curcumene, cupreene, Alaskenene, delta-junifolene and cedrol. The traditional Chinese medicine cacumen biotae essential oil with 13 specific components has a remarkable anti-depression effect, depression indexes can be improved from gene and protein levels, and depression-like behaviors can be improved. Meanwhile, the traditional Chinese medicine cacumen biotae essential oil is good in safety. Therefore, the traditional Chinese medicine cacumen biotae essential oil prepared by the invention can be used for preparing medicines for preventing and / or treating depression, and has a good application prospect.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of medicine, in particular to a traditional Chinese medicine Cacumen Platycladi essential oil, a preparation method thereof and an application of an antidepressant. BACKGROUND

[0002] At present, the commonly used antidepressants in clinic mainly include selective serotonin reuptake inhibitors (SSRIs), tricyclic antidepressants (TCAs) and the like, but these drugs generally have problems such as slow effect, large side effects and poor compliance. Therefore, developing new antidepressants with safety, effectiveness and small side effects has become a research hotspot at present.

[0003] The olfactory pathway directly acts on the brain limbic system (including amygdala, hippocampus and hypothalamus, etc.), and its potential to regulate emotions, stress response and sleep rhythm has been confirmed in aroma intervention studies. The olfactory intervention of essential oil has become an important direction in the international emotional health and complementary medicine research in recent years due to its fast effect, low dose and good safety. In the field of aromatherapy, Cacumen Platycladi essential oil is considered to be a dangerous essential oil, because the raw material of this Cacumen Platycladi essential oil is selected from Thuja occidentalis belonging to the genus of Thuja of Cupressaceae. The essential oil is composed of aromatic molecules such as monoterpenone, monoterpene, ester and the like, and the main components include thujaketone, thujaketene, camphor, isobornyl acetate and the like, including up to 60% of α-thujaketone and about 10% of β-thujaketone. Thujaketone is a component that can cause epilepsy, damage to the liver and central nervous system, and even a small amount of oral administration can cause vomiting, hallucinations and coordination disorders.

[0004] In addition to the above Thuja occidentalis, there is a Cacumen Platycladi with the Latin name of PLatycladus orientalis (L.) Franco, which is a commonly used Cacumen Platycladi in traditional Chinese medicine. The traditional Chinese medicine research and modern pharmacological research are mainly focused on its hemostatic, anti-inflammatory, antioxidant, antibacterial, anti-hair loss and the like, including the treatment of hematochezia, epistaxis, metrorrhagia and the like, and the application in dermatology and hair management. Modern research in recent years also suggests that traditional Chinese medicine Cacumen Platycladi contains flavonoids, triterpenes and a small amount of volatile components, which have been proved to have anti-inflammatory, antioxidant and hair follicle growth promoting activities. The volatile components (essential oil part) of traditional Chinese medicine Cacumen Platycladi have not been systematically studied and developed, and the olfactory inhalation of traditional Chinese medicine Cacumen Platycladi essential oil has almost no mature medical evidence or application mode in the regulation of emotions, anxiety and depression intervention. At present, there is no systematic research or application report on the olfactory inhalation of traditional Chinese medicine Cacumen Platycladi essential oil for the intervention of depression or emotional abnormalities. SUMMARY

[0005] The application aims to provide a traditional Chinese medicine Platycladus orientalis essential oil, a preparation method thereof and an antidepressant use.

[0006] The application provides a traditional Chinese medicine Platycladus orientalis essential oil, which is composed of the following components in mass percentage: α-pinene 21.02%, γ-terpinene 11.61%, β-terpineol 1.89%, α-terpineol 13.71%, γ-terpineol 5.02%, α-cuparene 15.98%, β-cuparene 6.32%, cis-thujaplicin 6.86%, zingiberene 0.95%, thujaplicin 0.96%, alaskene 1.28%, δ-cadinene 1.03%, cedrol 13.38%; the Platycladus orientalis in the traditional Chinese medicine Platycladus orientalis essential oil is Platycladus orientalis (L.) Franco.

[0007] The traditional Chinese medicine Platycladus orientalis essential oil is detected by gas chromatography-mass spectrometry to confirm that it contains 13 specific components, the mass percentage of each component is determined by the percentage of peak area sum, the result is kept to two decimal places, and the data processing leads to a deviation of ±0.5%.

[0008] The application further provides a preparation method of the aforementioned traditional Chinese medicine Platycladus orientalis essential oil, which comprises the following steps: (1) picking traditional Chinese medicine Platycladus orientalis branches and leaves, washing and putting into a distiller containing distilled water to immerse them completely; (2) heating and distilling, the heating and distilling temperature is 90-96°C, the condenser temperature is 15-25°C during distillation, and the distillation time is 60-90 min; after the distillate is left to stand, the upper essential oil is separated, and impurities are removed by filtration, to obtain the traditional Chinese medicine Platycladus orientalis essential oil.

[0009] The application further provides a use of the aforementioned traditional Chinese medicine Platycladus orientalis essential oil in preparing a medicine for preventing and / or treating depression.

[0010] Further, the medicine is a medicine for improving depression-like behavior.

[0011] Further, the medicine is a medicine for improving cell morphology of the hippocampal tissue CA1 area, CA3 area and DG area and / or increasing the number of nerve cells in the hippocampal tissue CA1 area, CA3 area and DG area.

[0012] Further, the medicine is a medicine for reducing the content of CORT in serum, increasing the contents of 5-HT and BDNF in serum; and / or, the medicine is a medicine for increasing the expression level of BDNF gene and 5-HT gene in the hippocampal tissue; and / or, the medicine is a medicine for increasing the expression level of BDNF protein in the hippocampal tissue.

[0013] Further, the drug is a nasal administration drug.

[0014] Further, the drug is an olfactory administration drug.

[0015] The "Chinese medicine Platycladus orientalis leaf essential oil" referred to in the present application corresponds to the leaves of Platycladus orientalis (L.) Franco, not the leaves of Thuja occidentalis commonly named "Platycladus orientalis leaf essential oil" in the field of aromatherapy. The raw material in the present application is commonly used in the field of traditional Chinese medicine, and therefore "Chinese medicine Platycladus orientalis leaf essential oil" is used in the present application to distinguish it.

[0016] In the background that neither traditional efficacy nor modern research involves the mood regulation effect of "Chinese medicine Platycladus orientalis leaf essential oil", the present application develops an anti-depression essential oil based on Chinese medicine Platycladus orientalis leaf essential oil, and clearly defines its preparation method and mechanism of action. The essential oil can be added with a medically acceptable adjuvant to form a dosage form for olfactory administration for the regulation of depressive mood. The present application has technical innovation significance, important research value and application prospect.

[0017] Compared with the prior art, the present application has the following beneficial effects: The present application found that the Chinese medicine Platycladus orientalis leaf essential oil prepared from Platycladus orientalis with Latin name PLatycladus orientalis (L.) Franco and containing 13 specific components has a significant anti-depression effect, can improve depression indicators and depressive behavior from the genetic and protein levels. At the same time, the Chinese medicine Platycladus orientalis leaf essential oil in the present application has good safety. Therefore, the Chinese medicine Platycladus orientalis leaf essential oil prepared in the present application can be used for preparing a drug for preventing and / or treating depression, and has good application prospect.

[0018] Obviously, according to the above content of the present application, according to the ordinary technical knowledge and common means in the art, without departing from the above technical idea of the present application, other various forms of modification, replacement or change can also be made.

[0019] The above content of the present application will be further described in detail through the specific embodiments below. However, it should not be understood that the scope of the above subject matter of the present application is limited to the following examples. Any technology realized based on the above content of the present application belongs to the scope of the present application. BRIEF DESCRIPTION OF DRAWINGS

[0020] Figure 1 The total ion chromatogram of the Chinese medicine Platycladus orientalis leaf essential oil detected by GC-MS.

[0021] Figure 2 The body weight change result graph of each group of mice.

[0022] Figure 3 Activity trajectory and heat map of each group of mice in the open field test.

[0023] Figure 4 Statistical graph of detection results of each group of mice in the open field test: A is the statistical graph of the total moving distance in the closed area; B is the frequency graph of entering the central area; C is the time graph of staying in the central area.

[0024] Figure 5 Activity trajectory and heat map of each group of mice in the elevated plus maze test.

[0025] Figure 6 Statistical graph of detection results of each group of mice in the elevated plus maze test: A is the statistical graph of the total distance of movement in the open arm; B is the frequency graph of entering the open arm; C is the time graph of staying in the open arm.

[0026] Figure 7 H&E staining results of the hippocampus of each group of mice.

[0027] Figure 8 Nissl staining results of the hippocampus of each group of mice.

[0028] Figure 9 Statistical graph of CORT, 5-HT and BDNF content in the serum of each group of mice: A is the statistical graph of CORT content in the serum; B is the statistical graph of 5-HT content in the serum; C is the statistical graph of BDNF content in the serum.

[0029] Figure 10 Statistical graph of the relative expression amount of BDNF gene and 5-HT gene in the hippocampus of each group of mice: A is the statistical graph of the relative expression amount of BDNF gene; B is the statistical graph of the relative expression amount of 5-HT gene.

[0030] Figure 11 Immunohistochemical detection results of BDNF protein in the hippocampus of each group of mice.

[0031] Figure 12 Western Blot detection results of BDNF protein in the hippocampus of each group of mice: A is the Western Blot graph; B is the BDNF protein expression statistical graph. DETAILED DESCRIPTION

[0032] The raw materials and equipment used in the specific embodiments of the present application are known products, which are obtained by purchasing commercially available products.

[0033] Example 1, Preparation of Chinese Medicine Cypresses Leaf Essential Oil The Latin name of the traditional Chinese medicine Platycladus orientalis (L.) Franco used in this embodiment is PLatycladus orientalis (L.) Franco, belonging to Cupressaceae (Cupressaceae) and Platycladus (Platycladus).

[0034] The preparation method of the traditional Chinese medicine Platycladus orientalis leaf essential oil is as follows: In summer and autumn, choose fresh, dry, and uncontaminated traditional Chinese medicine Platycladus orientalis (L.) Franco leaves and branches during the growing season, and wash them. Disassemble the distillation apparatus and clean all parts to ensure that there is no oil stain and impurity. Add 3 / 4 of the distillation water to the distillation pot, place it on the heating plate, and put the traditional Chinese medicine Platycladus orientalis leaf branches and leaves into the distillation pot to ensure that they are completely immersed in water, and the water level reaches the upper limit. Connect the distillation apparatus with a cooling tube, and put the other end of the tube into the collection bottle. The heating temperature is 90-96°C, the condenser temperature is 15-25°C, the cooking time is 60-90 min, and the steam will be condensed through the cooling tube and flow into the collection bottle. Wait for 24 hours, the essential oil will float on the upper layer of water, and use a dropper or a separation funnel to separate it from the water. After collecting the essential oil, filter it with filter paper or filter cloth to remove particles or other impurities suspended in the essential oil, and obtain the traditional Chinese medicine Platycladus orientalis leaf essential oil.

[0035] Example 2, composition study of traditional Chinese medicine Platycladus orientalis leaf essential oil The composition of the traditional Chinese medicine Platycladus orientalis leaf essential oil obtained in Example 1 was studied, and the traditional Chinese medicine Platycladus orientalis leaf essential oil obtained in Example 1 was sent to Shanghai University of Chinese Medicine for gas chromatography-mass spectrometry detection.

[0036] Detection instrument: Agilent gas chromatography-quadrupole mass spectrometry Preparation of test sample: Mix the essential oil to be tested, transfer 0.5 mL to a 1.5 mL centrifuge tube, and centrifuge for 1 min; take 50 μL of supernatant, weigh, add ethyl acetate, and prepare an essential oil solution with a concentration of 100 mg / mL; add anhydrous sodium sulfate powder according to a mass ratio of 10:1 of anhydrous sodium sulfate powder to essential oil solution, mix well, stand, and centrifuge; take 10 μL of supernatant, dilute 100 times with ethyl acetate to obtain a test solution with a concentration of 1 mg / mL; centrifuge the test solution for 1 min, take the supernatant, and perform gas chromatography-mass spectrometry detection.

[0037] Detection method: the chromatographic column is Agilent HP-5 MS UI (30 m x 0.25 mm x 0.25 μm); the carrier gas is high-purity helium; the flow rate is 1.0 mL / min; the injection port temperature is 250℃; the split ratio is 10:1; the injection amount is 1 μL; the temperature rising program is 50℃, which is raised to 180℃ at a rate of 3℃ / min, then raised to 300℃ at a rate of 20℃ / min, and then kept for 5 min; the ion source is EI+; the ion source temperature is 230℃; the scan mode is full ion scan; the scan range is m / z 40-600; the solvent delay is 4 min.

[0038] Data analysis: the original data of GC-MS detection is analyzed by Agilent MassHunter Qualitative Analysis (V B.07.00) software to obtain the information of each component peak (retention time, integrated peak area, peak area percentage, and peak height); the structure of each component peak is attributed by matching the mass spectrum database (NIST 2020 V2.4) and the retention index library (NIST ChemistryWebBook); the result of peak area sum is kept to two decimal places, and the data processing results have a deviation of ±0.5%.

[0039] Table 1. Component peak information table Table 2. Component peak attribution table The total ion chromatogram obtained by detection is shown in Figure 1 , the component peak information table is shown in Table 1, and the component peak attribution table is shown in Table 2. It can be seen that the components of the traditional Chinese medicine cypress leaf essential oil of the embodiment include: α-pinene, the mass percentage is 21.02%; γ-terpinene, the mass percentage is 11.61%; β-terpineol, the mass percentage is 1.89%; α-terpineol, the mass percentage is 13.71%; γ-terpineol, the mass percentage is 5.02%; α-cuparene, the mass percentage is 15.98%; β-cuparene, the mass percentage is 6.32%; cis-thujaplicin, the mass percentage is 6.86%; zingiberene, the mass percentage is 0.95%; flavesene, the mass percentage is 0.96%; alaskane, the mass percentage is 1.28%; δ-cadinene, the mass percentage is 1.03%; cedrol, the mass percentage is 13.38%.

[0040] The beneficial effects of the present application are demonstrated by the following specific test examples.

[0041] Test Example 1, Study on the Antidepressant Effect of Traditional Chinese Medicine Cypress Leaf Essential Oil 1. Experimental materials 1.1 Experimental animals: C57BL / 6J mice, male, 7 weeks old, grade: SPF, purchased from Beijing Vito Lihua, license number: SCXK (Shanghai) 2021-0006.

[0042] 1.2 Main reagents: Chinese medicine cypress leaf essential oil prepared in Example 1, isoflurane, HE staining kit, Nissl staining kit, ELISA kit (CORT, 5-HT and BDNF), RT-PCR reverse transcription kit, RT-PCR amplification kit, immunohistochemical kit, RIPA lysis buffer, BDNF antibody, β-tubulin antibody, BCA protein concentration determination kit, BDNF antibody, Tween 80.

[0043] 1.3 Main instruments and equipment: Small animal aerosol administration system, mouse elevated plus maze, open field reaction box, multifunctional enzyme label instrument, pathological section scanner, real-time fluorescence quantitative PCR instrument, chemiluminescence imaging system.

[0044] 2. Experimental method 2.1 Animal grouping: randomly divided into four groups, 10 mice in each group, control group (Ctr.), chronic restraint group (CRS), chronic restraint + 2.5% Chinese medicine cypress leaf essential oil group (2.5%), chronic restraint + 5% Chinese medicine cypress leaf essential oil group (5%).

[0045] 2.2 Preparation of chronic restraint (CRS) induced depression model: except for the control group, the mice in the other groups were put into the restraint device (to ensure that the mice could not move), restrained for 3 hours (9:00-12:00) every day, and lasted for 4 weeks.

[0046] 2.3 Essential oil intervention: after adding essential oil to deionized water, add appropriate amount of Tween 80, stir to dissolve the essential oil in water, get essential oil solution with concentrations of 2.5% and 5% respectively. Continue modeling for 4 weeks, from the 3rd week, except for the control group and the chronic restraint group, the other two groups of mice were put into the sniffing box of the small animal aerosol administration system, the prepared essential oil solution of each concentration was put into the aerosol cup, the gas flow was set to 3L / min, after starting, the essential oil solution was atomized into fine mist particles and sprayed into the sniffing box, each sniffing lasted for 15 min, once a day, for 2 weeks.

[0047] 2.4 Behavioral test 2.4.1 Elevated plus maze (EPM) The experiment lasted for 4 weeks, and the elevated plus maze test was immediately performed after 4 weeks. The mice were placed in a cross-shaped maze composed of two open arms and two closed arms, and the frequency of mice entering the open arms, the residence time and the running path were observed.

[0048] 2.4.2 Open field test (OFT) The experiment lasted for 4 weeks, and immediately after the open field test was performed at the end of the 4 weeks. The mice were placed in an open field box, and the time and frequency of their activity in the central and peripheral regions were analyzed, as well as the total running path.

[0049] 2.5 Sampling: After the behavioral test, blood was collected from the abdominal aorta under isoflurane anesthesia, and after standing, the serum was collected by centrifugation for use; the brain was removed by opening the skull cavity through heart perfusion with PBS, and the hippocampus was separated, part of which was collected for gene and protein detection; part was embedded with paraffin for morphological observation.

[0050] 2.6 H&E staining of mouse hippocampus: deparaffinization to water - hematoxylin 15 min - tap water wash 2 min - 1% hydrochloric acid alcohol 15 s - tap water wash 2 min - Li2CO3 3 s - tap water wash 10 min - eosin 3 min - dehydration - xylene transparency 2 min - neutral resin mounting - photography.

[0051] 2.7 Nissl staining of mouse hippocampus: deparaffinization to water - Nissl staining kit A liquid immersion 15 min - distilled water wash 5 s 2 times - Nissl staining kit B liquid differentiation - distilled water wash 5 s 3 times - xylene transparency 2 min - neutral resin mounting - photography.

[0052] 2.8 ELISA detection of CORT, 5-HT and BDNF content in serum: according to the ELISA kit instruction.

[0053] 2.9 RT-PCR detection of BDNF gene and 5-HA gene expression in hippocampal tissue: take the hippocampal tissue, extract the RNA, and follow the reverse transcription and amplification kit operation, take 18S as the internal reference gene, and use 2 -△△Ct Relative quantitative analysis method for target gene calculation.

[0054] 2.10 Immunohistochemical detection of BDNF protein expression in hippocampal tissue: paraffin section deparaffinization and hydration - 3% H2O2 treatment for 10 min - heating antigen repair - blocking room temperature for 1 h - primary antibody 4℃ overnight - secondary antibody room temperature 1 h - SP reaction - DAB color development - hematoxylin restain - sequential dehydration - transparency - mounting - photography.

[0055] 2.11 Western Blot detection of BDNF protein expression in hippocampal tissue: take the hippocampal tissue, extract the protein, and use the BCA protein concentration determination kit to determine the sample protein concentration, and then electrophoresis and transfer membrane, add primary antibody BDNF (1:1000), β-tubulin (1:1000) 4℃ shaking bed overnight, secondary antibody incubation color development and protein gray scale analysis.

[0056] 2.12 Statistical method: SPSS 24.0 statistical software was used for analysis. The data were measurement data, expressed as (±s), and one-way ANOVA was used for comparison of multiple groups of data. P P < 0.05 was considered statistically significant.

[0057] 3. Experimental results 3.1 Effects of C. sinica essential oil on body weight of CRS-induced mice During the 4-week CRS modeling period, the body weight changes of mice in each group were observed Figure 2 The body weight of the control group gradually increased with the extension of feeding time, while the body weight of the mice decreased significantly compared with the control group within 4 weeks after CRS stimulation.

[0058] 3.2 Effects of C. sinica essential oil on behavior of CRS-induced mice The results of open field test are shown in Figure 3 and Figure 4 The activity trajectory graph and heat map of mice in the open field test are shown in Figure 3 A is the total moving distance of mice in the enclosed area; Figure 4 B is the frequency of mice entering the central area; Figure 4 C is the time of mice staying in the central area; Figure 4 P < 0.01 compared with the control group (Ctrl.); P P < 0.001 compared with the control group (Ctrl.); P P < 0.05 compared with the chronic restraint group (CRS); P P < 0.001 compared with the chronic restraint group (CRS).The results of open field test showed that compared with the control group, the total moving distance of mice decreased (P < 0.05), the frequency of entering the central area and the time of staying in the central area decreased significantly (P < 0.01) after CRS stimulation. After smelling C. sinica essential oil, the frequency of entering the central area and the time of staying in the central area increased significantly (P < 0.05), and the total moving distance also improved (P < 0.05). P P P P P ​​​​​​​​​

[0059] The results of the elevated cross maze experiment are as follows Figure 5 and Figure 6 As shown, Figure 5 The activity trajectory diagram and heat map of mice in the elevated cross maze experiment; Figure 6 In the graph, A represents the total distance the mouse traveled in the open arm. Figure 6 B in the graph represents the frequency statistics of mice entering the open arm; Figure 6 C in the figure represents the statistical graph of the time mice spend in the open arm; in the figure, Indicates comparison with the control group (Ctrl.) P <0.01; Indicates comparison with the control group (Ctrl.) P <0.001; Indicates comparison with the chronic restraint group (CRS) P <0.001. The results of the elevated cross maze experiment showed that, compared with the control group, the frequency of mice entering the open arm and the time spent in the open arm were significantly reduced after CRS stimulation. P <0.01, P <0.001, as the total distance the open arm moves decreases ( P <0.01); Inhaling the essential oil of the Chinese herbal medicine Platycladus orientalis leaves can significantly increase the frequency of mice entering the open arm and the time spent in the open arm. P <0.001, and the distance the open arm moves also increases accordingly ( P <0.001).

[0060] The above experimental results demonstrate that the Chinese herbal medicine Platycladus orientalis leaf essential oil in Example 1 can improve depressive-like behavior in mice.

[0061] 3.3 Effects of Platycladus orientalis leaf essential oil on the pathological morphology of CRS-induced hippocampal tissue in mice H&E staining results are shown in […]. Figure 7 In the control group, the neurons in the hippocampus of mice were tightly arranged. Compared with the control group, after 4 weeks of CRS stimulation, the cells in the CA1, CA3 and DG areas of the hippocampus were loosely arranged and fewer in number. Inhalation of the essential oil of Platycladus orientalis leaves can significantly improve the morphological changes induced by CRS.

[0062] 3.4 Effects of Platycladus orientalis leaf essential oil on the number of neural cells in the hippocampus of mice induced by CRS The number of hippocampal neurons was detected by Niemeyer staining, and the results are shown below. Figure 8Compared with the control group, the nerve cells in the CA1 area, CA3 area and DG area of the hippocampus tissue were sparse, and the number of neurons was significantly reduced after CRS stimulation for 4 weeks; the number of nerve cells in the CA1 area, CA3 area and DG area could be significantly increased after smelling the traditional Chinese medicine cypress leaf essential oil.

[0063] 3.5 Effect of traditional Chinese medicine cypress leaf essential oil on the contents of CORT, 5-HT and BDNF in serum of CRS-induced mice CORT (corticosterone), 5-HT (5-hydroxytryptamine) and BDNF (brain-derived neurotrophic factor) play key roles in the occurrence and treatment of depression, and there is a close interaction among them, which together constitute the neurobiological basis of depressive disorders. The ELISA detection results are shown in Figure 9 , Figure 9 A in the figure is a statistical graph of the CORT content in the serum of mice in each group; Figure 9 B in the figure is a statistical graph of the 5-HT content in the serum of mice in each group; Figure 9 C in the figure is a statistical graph of the BDNF content in the serum of mice in each group; in the figure, indicates that compared with the control group (Ctrl.), P <0.01, indicates that compared with the control group (Ctrl.), P <0.001; indicates that compared with the chronic restraint group (CRS), P <0.05, indicates that compared with the chronic restraint group (CRS), P <0.01. Compared with the control group, the CORT in the serum of mice was significantly increased after CRS stimulation ( P <0.001), the content of BDNF was significantly reduced ( P <0.01), and the neurotransmitter 5-HT had a decreasing trend; smelling the traditional Chinese medicine cypress leaf essential oil could significantly reduce the content of CORT in the serum ( P <0.05), and promote the increase of BDNF content, among which the 5% concentration of traditional Chinese medicine cypress leaf essential oil could significantly increase the expression of BDNF compared with the CRS group alone ( P <0.01), and smelling the essential oil had a trend of promoting the expression of 5-HT. It is shown that the traditional Chinese medicine cypress leaf essential oil in Example 1 has an antidepressant effect.

[0064] 3.6 Effect of traditional Chinese medicine cypress leaf essential oil on the expression amounts of 5-HT gene and BDNF gene in hippocampus tissue of CRS-induced mice The hippocampus tissue was taken to detect the change of 5-HT gene and BDNF gene expression by real-time fluorescent quantitative PCR, and the results are shown in Figure 10 , Figure 10 A in FIG. is a statistical chart of relative expression of BDNF gene; Figure 10 B in FIG. is a statistical chart of relative expression of 5-HT gene; in the chart, indicates that compared with the control group (Ctrl.), P<0.05, P <0.05, <0.05, indicates that compared with the control group (Ctrl.), P<0.01; P <0.01; indicates that compared with the chronic restraint group (CRS), P<0.05, P <0.05, <0.05, indicates that compared with the chronic restraint group (CRS), P<0.01. Compared with the control group, the expression levels of BDNF gene and 5-HT gene in hippocampus tissue of mice after 4 weeks of CRS stimulation were significantly reduced (P<0.05, P <0.01), and the decrease of expression levels of BDNF gene and 5-HT gene caused by CRS could be reversed after the mice were given Chinese medicine cypress leaf essential oil sniffing, and the 5% concentration of Chinese medicine cypress leaf essential oil could significantly increase the expression levels of BDNF gene and 5-HT gene (P<0.01). P <0.01, P <0.01. P <0.01, <0.01,

[0065] 3.7 Effect of Chinese medicine cypress leaf essential oil on BDNF protein expression in hippocampus tissue of mice induced by CRS The expression level of BDNF protein in hippocampus tissue was detected by immunohistochemistry and Western Blot, and the results are shown in Figure 11 and Figure 12 . Figure 11 FIG. is a chart of immunohistochemical detection of BDNF protein in hippocampus tissue of mice in each group. Figure 12 A in FIG. is a chart of Western Blot; Figure 12 B in FIG. is a chart of BDNF protein expression; in the chart, <0.01, <0.01, P <0.01, <0.01, indicates that compared with the chronic restraint group (CRS), P<0.05, P<0.01. Immunohistochemistry results showed that the positive expression of BDNF protein in the hippocampus of mice was significantly reduced after 4 weeks of CRS stimulation compared with the control group; after 2 weeks of administration of Chinese medicine cypress leaf essential oil, the positive expression of BDNF protein in the hippocampus of mice was significantly improved, and the positive expression of BDNF protein in the hippocampus of mice in the group of 5% concentration of Chinese medicine cypress leaf essential oil was the most obvious. The results of Western Blot were consistent with the results of immunohistochemistry. Compared with the control group, the expression of BDNF protein in the hippocampus of mice was significantly reduced after 4 weeks of CRS stimulation (P < 0.01), and the administration of Chinese medicine cypress leaf essential oil could reverse the reduction of BDNF protein level induced by CRS (P < 0.01). P <0.01), and the administration of Chinese medicine cypress leaf essential oil could reverse the reduction of BDNF protein level induced by CRS (P < 0.01). P <0.01).

[0066] The above experimental results show that the Chinese medicine cypress leaf essential oil prepared in Example 1 can significantly improve the depressive behavior of mice; can improve the morphological changes in the CA1 area, CA3 area and DG area of the hippocampus and increase the number of nerve cells; at the same time, can reduce the content of CORT in serum, increase the content of 5-HT and BDNF, increase the expression level of 5-HT and BDNF genes and the expression level of BDNF protein in the hippocampus. Therefore, the Chinese medicine cypress leaf essential oil can be used for preparing a medicine for preventing and / or treating depression.

[0067] The above disclosure is only a specific embodiment of the present application, but the present application is not limited thereto, and any changes that can be thought of by those skilled in the art shall fall within the protection scope of the present application.

Claims

1. A traditional Chinese medicine of cypress leaf essential oil, characterized in that: The traditional Chinese medicine cypress leaf essential oil is composed of the following components in mass percentage The traditional Chinese medicine cypress leaf essential oil is composed of the following components in mass percentage Alpha-pinene 21.02%, gamma-terpinene 11.61%, beta-terpineol 1.89%, alpha-terpineol 13.71%, gamma-terpineol 5.02%, alpha-cuparene 15.98%, beta-cuparene 6.32%, cis-thujaplicin 6.86%, zingiberene 0.95%, floracuparene 0.96%, alaskene 1.28%, delta-cadinene 1.03%, cedrol 13.38%; the cypress in the traditional Chinese medicine cypress leaf essential oil is cypress with Latin name PLatycladus orientalis (L.) Franco.

2. The preparation method of the traditional Chinese medicine cypress leaf essential oil according to claim 1, characterized in that: The method comprises the following steps: (1) picking traditional Chinese medicine cypress leaf branches and leaves, washing and putting into a distiller containing distilled water to immerse all; (2) heating and distilling, the heating and distilling temperature is 90-96°C, the condenser temperature is 15-25°C during distillation, and the distillation time is 60-90 min; after the distillation is left to stand, the upper essential oil is separated, and impurities are removed by filtration to obtain the traditional Chinese medicine cypress leaf essential oil.

3. The use of the traditional Chinese medicine cypress leaf essential oil in claim 1 in the preparation of a medicine for preventing and / or treating depression.

4. Use according to claim 3, characterized in that: The medicine is a medicine for improving depression-like behavior.

5. Use according to claim 3, characterized in that: The medicine is a medicine for improving cell morphology of the hippocampal tissue CA1 region, CA3 region and DG region and / or increasing the number of nerve cells in the hippocampal tissue CA1 region, CA3 region and DG region.

6. Use according to claim 3, characterized in that: The medicine is a medicine for reducing the content of CORT in serum, increasing the content of 5-HT and BDNF in serum; and / or, the medicine is a medicine for increasing the expression level of BDNF gene and 5-HT gene in the hippocampal tissue; and / or, the medicine is a medicine for increasing the expression level of BDNF protein in the hippocampal tissue.

7. Use according to claim 3, characterized in that: The medicine is a medicine for nasal administration.

Images