A compound essential oil for soothing liver, relieving depression and helping sleep and preparation and application thereof

Abstract

The present application relates to a kind of compound essential oil of soothing liver and relieving depression and sleep aid and its preparation and application, including the following weight parts component: bupleurum essential oil 3~10 parts, cyperus rotundus essential oil 2~8 parts, orange peel essential oil 1~5 parts, fried liquorice essential oil 0.5~3 parts.Compared with prior art, the sleep aid compound essential oil composition prepared by the present application can play the synergistic effect of multi-component essential oil through simple essential oil diffusion, effectively treat insomnia symptoms, and has good soothing liver and relieving depression effect.

Description

Technical Field

[0001] This invention belongs to the field of essential oil technology, and relates to a compound essential oil for soothing the liver, relieving depression, and aiding sleep, as well as its preparation and application. Background Technology

[0002] In modern medicine and sleep physiology, insomnia is defined as a highly complex sleep-wake rhythm disorder, primarily characterized by a lack of subjective satisfaction with sleep duration, depth, or quality, accompanied by severe daytime dysfunction and psychosocial impairment. With increasing competitive pressures, rapid lifestyle changes, and information overload in modern society, insomnia is no longer merely an individual sleep disorder but has evolved into a widespread and serious public health crisis. From the perspective of biological evolution and homeostasis, insomnia is essentially an abnormal "warning response" to prolonged psychological stress or environmental discomfort, indicating that the body's internal environmental balance has exceeded the self-repair capabilities of the sleep system. This pathological characteristic is becoming increasingly prevalent across all age groups. Long-term chronic insomnia not only leads to cognitive decline, inattention, and irritability but is also an independent risk factor for depression, anxiety, chronic fatigue syndrome, and various cardiovascular diseases, severely impacting the quality of life for approximately 10% to 30% of the global adult population.

[0003] The pathogenesis of insomnia exhibits strong heterogeneity and network characteristics, being a product of the combined effects of genetic susceptibility, neurophysiological disorders, environmental stress, and psychological cognitive biases. Although neurobiology has made significant progress in sleep induction pathways and homeostasis maintenance, the precise molecular pathological logic of insomnia, especially persistent chronic insomnia, remains incompletely understood. This ambiguity at the mechanistic level directly leads to passive clinical interventions, characterized by limited treatment options, highly variable individual efficacy, and a high likelihood of symptom recurrence. Furthermore, long-term use of chemical drugs often leads to a vicious cycle of "drug resistance-increase-addition," posing a significant challenge to clinical medicine.

[0004] In-depth research reveals that persistent psychological stress triggered by stressful life events is the "driving center" of chronic insomnia. Under this stress, the body produces a series of neuroendocrine cascade responses, the most crucial of which is the overactivation of the hypothalamic-pituitary-adrenal (HPA) axis and hypervigilance of the limbic system. This physiological state of "high arousal" precisely corresponds to the pathological process of "liver qi stagnation, leading to fire" in Traditional Chinese Medicine (TCM). Liver qi stagnation causes qi obstruction, resulting in loss of mental and emotional balance, and consequently, insomnia. Therefore, revealing the complex interactions between the neural regulatory system, the endocrine feedback axis, and sleep structure from a biological perspective is crucial for identifying more broadly applicable and targeted interventions.

[0005] At the microscopic neurotransmitter level, the stability of the sleep-wake cycle depends on the dynamic interplay between two major systems within the brain: on one hand, the "wake-promoting system" centered on orexin, glutamate, and norepinephrine; on the other hand, the "sleep-promoting system" centered on gamma-aminobutyric acid (GABA) and adenosine. The underlying physiological logic of insomnia lies in the imbalance of this dynamic antagonistic relationship between excitation and inhibition, manifested as the pathological overexpression of wake-promoting neurotransmitters or the systemic decline in the function of GABAergic neurons. Although existing benzodiazepines can produce a short-term hypnotic effect by positively regulating GABA receptors, this is merely "passive sleep induction" and fails to fundamentally repair the damaged neurotransmitter regulatory network.

[0006] Meanwhile, brain-derived neurotrophic factor (BDNF) plays a crucial "neuroprotective" role in maintaining synaptic plasticity and sleep homeostasis. BDNF not only supports neuronal survival and repair but is also a key molecule in the precise regulation of circadian rhythms. Studies have confirmed that BDNF levels in the cerebrospinal fluid and serum of insomnia patients are significantly reduced, leading to abnormal metabolic activity in core regulatory areas such as the prefrontal cortex. This lack of neurotrophic support makes it difficult for the brain to generate sufficient "sleep drive," resulting in cognitive impairment and creating a vicious cycle where insomnia worsens and neurological damage reinforce each other.

[0007] Against this pathological backdrop, aromatherapy, as a natural, non-invasive, multi-target intervention strategy, demonstrates unique clinical value. Extensive scientific literature has empirically demonstrated that volatile active molecules (such as monoterpenes and sesquiterpenes) in single and compound essential oils can directly reach the limbic system through the olfactory mucosa, bypassing part of the blood-brain barrier and exerting a "harmonizing" effect on the central nervous system. For example, specific components (linalool, linalyl acetate, etc.) in lavender, bergamot, and bitter orange essential oils have been shown to regulate GABAA receptor subtypes and reduce HPA-axis-mediated stress corticosterone impulses, thereby effectively alleviating hyperarousal states. Compound essential oils offer greater advantages than single-component formulas; through synergistic effects between components, they not only prolong sleep duration but also exhibit multi-target regulatory effects in improving sleep depth and regulating mood complications such as depression and anxiety, demonstrating excellent safety.

[0008] However, current mainstream insomnia intervention programs in the market and clinical practice still mainly rely on chemically synthesized drugs. Although these drugs are fast-acting, they are often accompanied by severe "hangover effects" (residual effects the next day), memory impairment, and disruption of sleep structure (such as inhibiting REM sleep). Most importantly, these methods mostly provide symptom relief and fail to address the underlying imbalance of homeostasis caused by "qi stagnation and liver stagnation" in insomnia.

[0009] Based on the deep integration of traditional Chinese medicine wisdom and modern pharmacology, TCM believes that "the liver governs the free flow of Qi and regulates its movement," and the smooth flow of liver Qi is the cornerstone of sleep homeostasis. Therefore, developing a compound essential oil composition that can "soothe the liver, regulate Qi, relieve depression, and calm the mind," while simultaneously synergistically regulating neurotransmitter balance, activating neurotrophic pathways, and being safe and non-addictive, not only overcomes the limitations of modern synthetic drugs but also provides an innovative scientific solution for the natural treatment of chronic insomnia. This multi-dimensional biological intervention helps patients rebuild a healthy sleep-wake rhythm, possessing extremely high clinical application potential and social benefits.

[0010] Chinese patent application CN120437221A discloses the use of Bupleurum chinense essential oil and Cyperus rotundus essential oil in the preparation of a non-oral drug for treating liver qi stagnation syndrome, as well as an essential oil composition for treating liver qi stagnation syndrome. However, this application only uses Bupleurum chinense and Cyperus rotundus as its core components, with relatively low dosages. It primarily targets liver qi stagnation syndrome, lacking auxiliary herbs to harmonize qi and moderate efficacy. The formulation is relatively simple, limited to regulating liver qi stagnation syndrome itself, without focusing on insomnia and anxiety induced by liver qi stagnation, nor considering the need for qi regulation and stable, sustained drug release. Furthermore, this application does not disclose animal efficacy data related to insomnia and anxiety, lacks experimental support regarding mechanisms of action such as neurotransmitters, the HPA axis, and oxidative stress, and lacks evidence of the actual effects of olfactory administration on improving sleep latency, prolonging sleep duration, and alleviating anxiety and hyperactivity. Summary of the Invention

[0011] The purpose of this invention is to provide a compound essential oil for soothing the liver, relieving depression, and aiding sleep, as well as its preparation and application, providing a new solution for preparing drugs and daily necessities for treating insomnia and anxiety.

[0012] The objective of this invention can be achieved through the following technical solutions: In a first aspect, the present invention provides a compound essential oil for soothing the liver, relieving depression, and aiding sleep, comprising the following components: 3-10 parts of Bupleurum essential oil, 2-8 parts of Cyperus essential oil, 1-5 parts of Citrus aurantium essential oil, and 0.5-3 parts of Glycyrrhiza uralensis essential oil.

[0013] Furthermore, it includes the following raw materials: 4-8 parts Bupleurum essential oil, 3-6 parts Cyperus essential oil, 2-4 parts Citrus aurantium essential oil, and 1-2 parts Glycyrrhiza uralensis essential oil.

[0014] Furthermore, it includes the following raw materials: 5-7 parts Bupleurum essential oil, 4-5 parts Cyperus essential oil, 2-3 parts Citrus aurantium essential oil, and 1-1.5 parts Glycyrrhiza uralensis essential oil.

[0015] Furthermore, it includes the following raw materials: 6 parts Bupleurum essential oil, 4 parts Cyperus essential oil, 2 parts Citrus aurantium essential oil, and 1 part Glycyrrhiza uralensis essential oil.

[0016] Furthermore, it includes the following raw materials: 8 parts Bupleurum essential oil, 3 parts Cyperus essential oil, 3 parts Citrus aurantium essential oil, and 1 part Glycyrrhiza uralensis essential oil.

[0017] Furthermore, it includes the following raw materials: 4 parts Bupleurum essential oil, 3 parts Cyperus essential oil, 3 parts Citrus aurantium essential oil, and 1 part Glycyrrhiza uralensis essential oil.

[0018] In a second aspect, the present invention provides a method for preparing a compound essential oil for soothing the liver, relieving depression, and aiding sleep as described in the first aspect above, comprising the following steps: weighing each component essential oil raw material into a container, stirring, and thus obtaining the product.

[0019] Furthermore, the stirring must be carried out under light-proof and cool conditions.

[0020] Furthermore, the stirring must be carried out under sealed conditions.

[0021] Furthermore, the essential oil raw materials of each component are purchased commercially or obtained by distillation, supercritical CO2 extraction, or solvent extraction.

[0022] In a third aspect, the present invention provides an application of the liver-soothing, mood-lifting, and sleep-aiding compound essential oil as described in the first aspect above in the preparation of pharmaceutical preparations and daily necessities for treating insomnia and anxiety; the sleep-aiding compound essential oil can soothe the liver and relieve mood through aroma diffusion and inhalation.

[0023] Compared with the prior art, the present invention has the following advantages: (1) The sleep-aid compound essential oil prepared in this invention can effectively alleviate insomnia symptoms and has a good sleep-aiding effect by combining the theories of traditional Chinese and Western medicine for treating insomnia and modern aromatherapy, and by using a simple essential oil diffusion method.

[0024] (2) The sleep-aid compound essential oil prepared by this invention uses a variety of natural plant ingredients and has the synergistic pharmacological characteristics of multiple components, multiple targets and multiple mechanisms. It can make up for the shortcomings of traditional single Western medicine and improve the relief effect on insomnia symptoms. In addition, when using compound essential oil, essential oils with similar functions can be blended together to enhance efficacy; essential oils with large differences in function can be blended together to expand the therapeutic effect.

[0025] (3) The Bupleurum essential oil in this invention is rich in saikosauce alcohol and monoterpenoids. It can significantly regulate the feedback mechanism of the hypothalamus-pituitary-adrenal (HPA) axis through the olfactory pathway, downregulate the pathological pulses of adrenocorticotropic hormone (ACTH) and plasma cortisol (CORT), and fundamentally suppress the "over-arousal" state caused by emotional stress. The sesquiterpenoids such as cyperene and cyperone contained in Cyperus essential oil can synergistically enhance the affinity of the central inhibitory neurotransmitter γ-aminobutyric acid (GABA), and dynamically balance the metabolic turnover of 5-HT in the brain, effectively relieve anxiety-like emotions, exert the "regulating qi and relieving depression" sleep-promoting effect, and increase the duration of deep sleep. The high D-limonene and myrcene content in the essential oil of Citrus aurantium can significantly act on the amygdala and anterior cingulate cortex pathways of the limbic system, regulate the rise and fall of Qi, relieve the tension caused by Qi stagnation in the body, thereby shortening the sleep latency and inducing skeletal muscles to enter a relaxed state. The glycyrrhizin derivatives and trace amounts of volatile flavonoids contained in the essential oil of Glycyrrhiza uralensis play a key role in "harmonizing" and "stabilizing" the compound. It can not only reduce oxidative stress indicators in brain tissue (such as MDA content) and protect damaged neurons, but also effectively slow down the release rate of other volatile components in the compound, so that the drug effect is stable and long-lasting, and enhance parasympathetic nerve tone, inducing the body to enter a physiological balance that is conducive to sleep initiation.

[0026] (4) This invention deeply integrates the formula logic of "regulating qi and soothing the liver" in traditional Chinese medicine with the concept of modern aromatherapy. The aromatic essential oil composition acts directly on the limbic system of the brain through the olfactory pathway, which can produce a positive "bidirectional regulation" and "stimulating effect" on the neuroendocrine network. Through olfactory intervention, this invention can help restore the disordered sleep-wake circadian rhythm to a normal homeostasis and correct the abnormal expression of biological clock genes caused by long-term insomnia. In addition, this mechanism of soothing the liver and regulating qi can not only solve the problem of falling asleep, but also regulate the physiological coordination of various tissue systems in the body, thereby achieving a sleep-aiding function from the perspective of overall regulation.

[0027] (5) The sleep-aid compound essential oil prepared by this invention has extremely high bioavailability and clinical application advantages. Due to the excellent lipid solubility of the essential oil components, their active small molecules can quickly penetrate the blood-brain barrier (BBB) ​​and directly act on the central nervous system targets, which is more rapid in onset than oral preparations and avoids the first-pass effect of the liver. At the same time, the compound essential oil form adopted by this invention effectively makes up for the inherent defects of volatile aromatic Chinese medicines such as Bupleurum, Cyperus, and Citrus aurantium in the traditional high-temperature decoction process, which result in the loss of aroma and effective ingredients, and preserves the original efficacy of the medicinal materials to the greatest extent. In addition, the compound essential oil of this invention is stable in nature, extremely convenient to carry and use, and can achieve the therapeutic goals of soothing the liver and relieving depression and regulating insomnia through simple diffusion or inhalation, which significantly improves the patient's treatment compliance. Attached Figure Description

[0028] Figure 1 This is the target of action between the sleep-aiding essential oil of this invention and insomnia symptoms.

[0029] Figure 2 This is a graph showing the target-related PPI information between the sleep-aiding essential oil of this invention and insomnia symptoms.

[0030] Figure 3 This is a computational graph showing the GO functional enrichment of target sites between the sleep-aiding essential oil of this invention and insomnia symptoms.

[0031] Figure 4 This is a computational graph showing the KEGG pathway enrichment of the target sites between the sleep-aiding essential oil of this invention and insomnia symptoms.

[0032] Figure 5 This shows the changes in mouse body weight during the behavioral experiments of this invention.

[0033] Figure 6 This is a graph showing the proportion of time mice spent entering the open arm during the elevated cross maze test of this invention.

[0034] Figure 7 This is a graph showing the proportion of time mice spent entering the closed arm during the elevated cross maze test of this invention.

[0035] Figure 8 and Figure 9 This is a comparative diagram of sleep patterns during the sodium pentobarbital-induced sleep test of this invention.

[0036] Figure 10 and Figure 11 This is a comparison diagram of autonomous activity in the open field test of this invention.

[0037] Figure 12 This is a comparison chart of the 5-HT content in mouse serum as part of the physiological index testing of this invention.

[0038] Figure 13 This is a comparison chart of DA content in mouse serum for physiological index testing according to the present invention.

[0039] Figure 14 This is a comparison chart of GABA content in mouse serum for physiological index testing according to the present invention.

[0040] Figure 15 This is a comparison chart of ACTH levels in mouse serum as part of the physiological index testing of this invention.

[0041] Figure 16 This is a comparison chart of CORT levels in mouse serum for physiological index testing according to the present invention.

[0042] Figure 17 This is a comparison chart of CRH levels in mouse serum used in the physiological index testing of this invention.

[0043] Figure 18 This is a comparison chart of MDA content in mouse serum for physiological index testing according to the present invention. Detailed Implementation

[0044] The present invention will now be described in detail with reference to the accompanying drawings and specific embodiments. These embodiments are based on the technical solution of the present invention and provide detailed implementation methods and specific operating procedures. However, the scope of protection of the present invention is not limited to the following embodiments.

[0045] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this application belongs. The terminology used herein is for the purpose of describing particular embodiments only and is not intended to be limiting of the application.

[0046] As used herein, the terms "and / or," "or / and," and "and / or" encompass any one of two or more of the related listed items, as well as any and all combinations of the related listed items. These arbitrary and all combinations include any two related listed items, any more related listed items, or a combination of all related listed items. It should be noted that when at least three items are connected using at least two conjunctions selected from "and / or," "or / and," and "and / or," it should be understood that, in this application, the technical solution undoubtedly includes solutions connected by "logical AND," and also undoubtedly includes solutions connected by "logical OR."

[0047] In this invention, the technical features described in an open-ended manner include both closed-ended technical solutions composed of the listed features and open-ended technical solutions that include the listed features.

[0048] In this invention, numerical ranges are involved. Unless otherwise specified, the numerical ranges are considered continuous and include the minimum and maximum values ​​of the range, as well as every value between the minimum and maximum values. Furthermore, when the range refers to integers, it includes every integer between the minimum and maximum values ​​of the range. Additionally, when multiple ranges are provided to describe features or characteristics, the ranges may be merged. In other words, unless otherwise specified, all ranges disclosed herein should be understood to include any and all subranges to which they are included.

[0049] This document only specifically discloses some numerical ranges. However, any lower limit can be combined with any upper limit to form an unspecified range; and any lower limit can be combined with other lower limits to form an unspecified range, just as any upper limit can be combined with any other upper limit to form an unspecified range. Furthermore, each individually disclosed point or single value can itself serve as a lower or upper limit and be combined with any other point or single value or with other lower or upper limits to form an unspecified range.

[0050] In this document, the term "suitable" as used in phrases such as "suitable combination," "suitable method," and "any suitable method" refers to the ability to implement the technical solution of this application, solve the technical problem of this application, and achieve the expected technical effect of this application.

[0051] In this invention, terms such as "further," "even more," and "particularly" are used for descriptive purposes and to indicate differences in content, but should not be construed as limiting the scope of protection of this application.

[0052] In this invention, "optionally," "optionally," and "optional" mean that they are optional, that is, they are selected from either "with" or "without." If multiple "options" appear in a technical solution, unless otherwise specified and there are no contradictions or mutual constraints, each "option" is independent.

[0053] In the description of the invention, "multiple" means at least two, such as two, three, etc., unless otherwise explicitly specified.

[0054] Unless otherwise specified, all preparations and tests described herein took place at 25°C.

[0055] The terms “comprising,” “including,” “containing,” “having,” “comprising,” or other variations thereof are intended to cover non-closed inclusion, and no distinction is made between these terms. The term “comprising” means that other steps and ingredients may be added without affecting the final result. The compositions and methods / processes of the present invention comprise, consist of, and substantially consist of the essential elements and limitations described herein, as well as any additional or optional ingredients, components, steps, or limitations described herein. No distinction is made between the terms “efficacy,” “performance,” “effect,” and “potency” herein.

[0056] Unless otherwise specified, all embodiments and optional embodiments of the present invention can be combined with each other to form new technical solutions. Unless otherwise specified, all technical features and optional technical features of the present invention can be combined with each other to form new technical solutions.

[0057] Unless otherwise specified, all steps of the present invention may be performed sequentially or randomly, but are preferably performed sequentially.

[0058] The essential oil raw materials used in the embodiments of this invention were all purchased from Shanxi Hongyi Traditional Chinese Medicine Technology Co., Ltd.

[0059] Example 1: A sleep-aiding compound essential oil is prepared by weighing each single essential oil in the following proportions, placing them in a clean brown glass bottle, and stirring and mixing them in the dark to obtain the sleep-aiding compound essential oil composition: 6 parts Bupleurum essential oil, 4 parts Cyperus essential oil, 2 parts Citrus aurantium essential oil, and 1 part Glycyrrhiza uralensis essential oil.

[0060] Example 2: A sleep-aiding compound essential oil is prepared by weighing each single essential oil in the following proportions, placing them in a clean brown glass bottle, and stirring and mixing them in the dark to obtain the sleep-aiding compound essential oil composition: 8 parts Bupleurum essential oil, 3 parts Cyperus essential oil, 3 parts Citrus aurantium essential oil, and 1 part Glycyrrhiza uralensis essential oil.

[0061] Example 3: A sleep-aiding compound essential oil is prepared by weighing each single essential oil in the following proportions, placing them in a clean brown glass bottle, and stirring and mixing them in the dark to obtain the sleep-aiding essential oil composition: 4 parts Bupleurum essential oil, 3 parts Cyperus essential oil, 3 parts Citrus aurantium essential oil, and 1 part Glycyrrhiza uralensis essential oil.

[0062] Comparative Example 1: It is a single-ingredient essential oil of Bupleurum chinense.

[0063] Comparative Example 2: It is a single-ingredient essential oil of Cyperus rotundus.

[0064] Comparative Example 3: It is a single-ingredient essential oil of Citrus aurantium.

[0065] Comparative Example 4: This is a single-ingredient essential oil made from roasted licorice root.

[0066] Comparative Example 5: A compound essential oil: Weigh each single essential oil according to the following weight proportions, place them in a clean brown glass bottle, and stir and mix them in the dark: 4 parts Bupleurum essential oil, 3 parts Cyperus essential oil, and 3 parts Citrus aurantium essential oil.

[0067] Comparative Example 6: A compound essential oil: Weigh each single essential oil according to the following weight proportions, place them in a clean brown glass bottle, and stir and mix them in the dark: Bupleurum essential oil 1.0 part, Cyperus essential oil 0.75 part, Ligusticum chuanxiong essential oil 0.75 part, Tangerine peel essential oil 0.75 part, Citrus aurantium essential oil 0.75 part, Clove essential oil 0.75 part.

[0068] Comparative Example 7: A compound essential oil: Weigh each single essential oil according to the following weight proportions, place them in a clean brown glass bottle, and stir and mix them in the dark: 12 parts Bupleurum essential oil, 7 parts Cyperus essential oil, 5 parts Citrus aurantium essential oil, and 5 parts Glycyrrhiza uralensis essential oil.

[0069] Experimental methods: Screening for potential pharmacologically active ingredient targets in sleep-aiding essential oils First, the active ingredients of the sleep-aid essential oil were entered into PubChem (https: / / PubChem.ncbi.nlm.nih.gov / ), the world's largest free chemical information database, to obtain corresponding chemical structure data; then, the SwissTargetPrediction (http: / / www.swisstargetprediction.ch / ) database was used to predict the potential targets of the sleep-aid essential oil and to build a relevant target library.

[0070] Acquisition of insomnia-related targets Using "Insomnia" as the keyword, this study utilized the therapeutic target database TTD (https: / / db.idrblab.net / ttd / ), the human gene database GeneCards (http: / / www.genecards.org / ), and the human Mendelian genetics database OMIM (https: / / www.omim.org / ) to mine target information related to insomnia. After integrating the target data from these multiple sources and removing duplicate entries, a target intersection analysis was conducted using the Bioinformatics platform (http: / / www.bioinformatics.com.cn / ) to identify the overlap between the active ingredient targets of sleep-aiding essential oils and insomnia-related targets.

[0071] Construction and visualization of protein-protein interaction (PPI) networks Import the relevant target data into the STRING database (https: / / cn.string-db.org / ) to draw a protein-protein interaction network diagram. Then, import the obtained network file into Cytoscape 3.10.0 software for visualization analysis. The core targets were selected based on the criteria of having betweenness centrality, proximity centrality, and degree values ​​higher than the average value. The selection criteria were: degree > median, betweenness > median, and closeness > median.

[0072] Pathway enrichment analysis Common drug-disease targets were uploaded to the Metascape database (https: / / metascape.org / ), and Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) metabolic pathway enrichment analyses were performed to obtain relevant functional information such as molecular function, biological process, cellular components, and KEGG pathways corresponding to the targets. GO and KEGG diagrams were generated using a bioinformatics platform (http: / / www.bioinformatics.com.cn / ), and related visualization analyses were completed using Cytoscape 3.10.0 software.

[0073] Results and Discussion Disease target construction for insomnia and target prediction of various components of sleep-aiding essential oils Based on GC-MS component data, bioinformatics techniques were used to search and deduplicate data in the PubChem and Swiss TargetPrediction databases, identifying 194 predictive action sites for sleep-aiding essential oils. Furthermore, a mapping analysis was performed between these sites and 6182 pathological targets related to insomnia. Figure 1 As shown in the analysis, the results indicate that there are 76 common targets between the active ingredients of sleep-aiding essential oils and insomnia.

[0074] Protein-protein interactions Protein-protein interaction (PPI) information for the 76 intersecting target sites was obtained from the STRING database and converted to TSV format for network construction in Cytoscape 3.10.0 software. The network was then subjected to topological evaluation and visualization modifications using the CentiScaPe 2.2 plugin. Figure 2 Thus, 27 key targets of sleep-aiding essential oils, represented by AKT1, ESR1, MAOA, MAPK3 and JAK2, were extracted from the complex network.

[0075] Pathway enrichment analysis Enrichment calculations for GO function and KEGG pathway of the aforementioned comorbidity targets were performed using the Metascape database. In the GO enrichment analysis, the top 20 significant entries from biological processes (BP), cellular composition (CC), and molecular function (MF) were extracted according to the p-value from smallest to largest and plotted as follows: Figure 3. The results show that MF mainly involves nuclear receptor activity, metal ion transmembrane transporter activity, kinase binding and the like; BP covers cellular response to hormone stimulus, regulation of secretion by cell, synaptic signaling and the like; CC focuses on presynapse, membrane raft, receptor complex and the like. For KEGG pathways, with P < 0.01 as the screening threshold, the 20 pathways with the highest enrichment degree are selected for visual display as Figure 4 shown. The analysis indicates that the potential pathways by which the sleep-aiding exerts its sleep-aiding effect include Chemical carcinogenesis - receptor activation, AGE-RAGE signaling pathway in diabetic complications, Apoptosis and the like.

[0076] The present invention conducts the following animal behavior tests on the sleep-aiding compound essential oil compositions prepared in the above examples and the single essential oils of the comparative examples: Animal selection and rearing: Healthy SPF-grade ICR mice, male, 6-8 weeks old, with a body weight of 27-33 g, were selected, purchased from Shanghai Jiesijie Laboratory Animal Co., Ltd., and the animal production license number is SCXK (Shanghai) 2023-0004. The animals were reared in an SPF environment under conventional rearing conditions, at a temperature of 22-25°C, a humidity of 40%-70%, and a 12-hour light-dark cycle.

[0077] Establishment of mouse insomnia model (PCPA model) and administration: The animals were acclimatized for 7 days, weighed, and randomly divided into 12 groups, with 5 mice in each group. The groups are respectively the blank group (Control), the model group (Model), and the essential oil group (including each example group and the comparative example group). Mice in the blank group were not given any stimulation. Mice in the model group, the positive control group and the essential oil group received PCPA injection for 3 consecutive days. The specific implementation scheme is as follows: intraperitoneal injection of PCPA solution was continuously performed on the mice at fixed time periods (i.e., 9:00 AM, 10:00 PM) every day, ensuring that the experimental animals move without any discomfort. Body weight was detected every two days, and the changes in body weight of the mice were recorded.

[0078] Mice in the blank control group and model group were treated with distilled water nebulization for 1.5 hours after injection, while mice in the essential oil group were treated with 0.3% concentration of experimental essential oil nebulization for 2 hours.

[0079] Behavioral tests: Elevated cross maze test Before the experiment, each mouse was placed in the experimental area for 5 minutes to acclimatize. After 5 minutes of free exploration, the mice were quickly placed in the center of an elevated cross maze, with their heads facing the open arms. Upon release, four metrics were measured: the number of times the mice entered the open arms (OE), the time taken to enter the open arms (OT), the number of times the mice entered the closed arms (CE), and the time taken to enter the closed arms (CT). The percentage of time taken to enter the open arms (OT%) and the percentage of time taken to enter the closed arms (CT%) were calculated based on OE, OT, CE, and CT. Measurements were performed for 5 minutes per mouse. After each test, the elevated cross maze was cleaned with 75% alcohol to remove fecal matter, urine, and other residues, as well as to eliminate odors, and then dried with a cloth.

[0080] Sodium pentobarbital sleep-inducing test After the last administration intervention, mice in each group were intraperitoneally injected with a suprathreshold dose of sodium pentobarbital solution (50 mg / kg) according to their body weight. They were then placed in a quiet and dimly lit observation box, and their sleep latency (the time from administration to the disappearance of the righting reflex) and sleep duration (the time from the disappearance of the righting reflex to its recovery) were recorded in detail. The criterion for the disappearance of the righting reflex was that the mouse could not turn over voluntarily within 30 seconds when placed in a supine position. The environment was kept quiet and constant throughout the experiment, and the observation box was thoroughly cleaned with 75% alcohol and odor removed after each mouse was tested to eliminate interference factors.

[0081] Open field test Before the experiment, mice were placed in the central square of a 40cm×40cm×40cm black open box for 3 minutes to acclimatize. During the experiment, the mice's activities in the open field were recorded on video over 5 minutes, and the total distance traveled and the time spent in the central area were analyzed. The experimental environment was kept quiet, and after each mouse was tested, the open field test box was wiped with 75% alcohol and excrement was removed before testing another mouse.

[0082] ELISA method for measuring the expression of insomnia-related indicators After the behavioral tests, blood was collected from the orbital cavity of anesthetized mice. After standing for 2 hours, the blood was centrifuged twice, and the supernatant was collected after 20 minutes. The levels of various physicochemical indicators in mouse serum were measured according to the steps listed on the respective ELISA kits for mice.

[0083] Mice were euthanized, and their brains were quickly dissected, rinsed with physiological saline, and rapidly frozen at -80°C. A specific amount of tissue was weighed from each group using a precision balance to prepare a homogenate. The supernatant was collected, and the levels of various physicochemical parameters in the mouse brain tissue were determined according to the steps listed on the mouse ELISA kit.

[0084] Data are expressed as mean ± standard deviation. SPSS 13.0 software was used for statistical analysis, and one-way ANOVA was employed to determine statistical significance. p < 0.05 was considered statistically significant. Origin 2023 was used for graphing.

[0085] The results of the above behavioral tests are as follows: Behavioral experiments on mice Table 1. Changes in mouse body weight before and after modeling. The changes in mouse body weight during the experiment are shown in Table 1 and Figure 5 As shown, the weight of mice in the blank control group steadily increased. Compared with the blank control group, the weight of mice in the model group and the essential oil group generally showed a trend of first decreasing and then increasing during the establishment of the insomnia model, but both were lower than that of the blank control group, indicating that the PCPA insomnia model was successfully established. Nine days after smelling the essential oil (the blank control and model group mice smelled distilled water), the mice in the essential oil groups of Comparative Examples 1-7 and Examples 1-3 rapidly increased in body weight. This was because the mice's insomnia was relieved after smelling the essential oil, their food intake increased, and their physiological indicators gradually returned to normal, resulting in rapid weight gain. Among them, the mice in the essential oil groups of Examples 3 and 2 showed more significant weight increases after smelling the essential oil.

[0086] Table 2. Behavioral results of mice in the elevated cross maze model. The behavioral effects of mice in the elevated cross maze model are shown in Table 2 and Figure 6 , 7As shown, the elevated cross maze and anxiety-like behavior in mice exhibit a stable correlation, making it suitable for rapid screening of anxiety-related phenotypes and evaluation of anti-anxiety drugs or products. Experimental results showed that the proportion of open-arm dwell time in the elevated cross maze test for the model group mice was only 10.41 ± 3.527%, significantly lower than the 43.45 ± 1.245% in the control group (p < 0.01), while the proportion of closed-arm dwell time was significantly increased. The elevated cross maze is a classic experiment for evaluating anxiety responses in animals. The above results indicate that the model group mice showed significantly reduced exploratory behavior towards novel environments, exhibiting significant fear and anxiety. Given the high physiological correlation between anxiety and sleep disorders, this high-anxiety behavior strongly demonstrates the successful establishment of an anxiety-related insomnia model. After intervention with different essential oils, the open arm dwell time of mice in Comparative Groups 1-7 and the essential oil groups in Examples 1, 2, and 3 was significantly increased compared to the model group (ranging from 17.49% to 38.87%), indicating that smelling the essential oils restored the exploratory interest of insomniac mice to varying degrees and improved their insomnia behavior. Data comparison showed that the overall open arm dwell time percentage (30.74% - 38.87%) of mice smelling the compound essential oils in the example groups was higher than that of the comparative group (17.49% - 29.83%), strongly demonstrating that the compound essential oils of this invention are more effective in restoring the exploratory interest of insomniac mice and improving their insomnia behavior. Within the example groups, Example 3 showed the most outstanding performance, with an open arm dwell time percentage of 38.87 ± 1.432%, a greater increase and the best regulatory effect compared to Examples 1 and 2.

[0087] Table 3. Behavioral results in the sodium pentobarbital-induced sleep test in mice. Table 3 summarizes the improvement in sleep in mice. Figure 8 , 9As shown in the figure. The experimental results showed that the sleep latency of the model group mice (610.00 ± 9.61 s) was significantly higher than that of the blank group (324.33 ± 8.04 s), and the sleep duration (26.33 ± 2.42 min) was significantly lower than that of the blank group (54.17 ± 3.06 min) (p < 0.01), indicating that the model group mice had difficulty falling asleep and their sleep time was significantly shortened, and the insomnia model was successfully established. The sleep latency of mice in the control groups 1-7 and the essential oil groups in Examples 1, 2 and 3 was shorter than that of the model group, and the sleep duration was longer than that of the model group, indicating that smelling essential oil can effectively shorten the time to fall asleep and prolong the sleep duration of insomnia mice, and the insomnia behavior is improved. Moreover, the overall sleep improvement of the mice in the example group after smelling the compound essential oil was better than that of the control group (the latency of the example group was shorter and the duration was longer), indicating that the compound essential oil of the present invention can significantly improve the sleep quality of insomnia mice. Compared to Examples 1 and 2, Example 3 showed a greater reduction in sleep latency and a greater extension in sleep duration, with a shorter latency and a longer duration.

[0088] Table 4. Behavioral results of mice in the open field test. The behavioral results of mice in the open field test are shown in Table 4 and Figure 10 , 11 As shown in the figure. The experimental results showed that the total distance traveled (236.42 ± 18.54 cm) and activity time (324.18 ± 16.32 s) in the open field test of the model group mice were significantly higher than those in the control group (171.54 ± 12.45 cm and 312.45 ± 14.85 s), indicating that the model group mice exhibited an abnormally excited state induced by insomnia, and the insomnia model was successfully established. Although the behavioral performance of the mice in each comparative group (1-7) in the open field showed some regression compared with the model group, the differences were relatively small. In contrast, the compound essential oil groups of Examples 1, 2 and 3 showed better improvement, with their total distance traveled and activity time significantly reduced and returning to the level of the control group. Among them, the mice in the compound essential oil group of Example 3 showed the most significant improvement, with their total distance recovered to 148.21 ± 9.85 cm and activity time recovered to 282.45 ± 10.42 s. This indicates that smelling the compound essential oil can effectively inhibit the abnormal excitement of insomnia mice and significantly improve the insomnia and anxiety symptoms of mice.

[0089] Based on the preferred liver-soothing and mood-regulating compound essential oil formula disclosed in the prior art, this invention designed Comparative Examples 5 and 6 as controls to fully demonstrate the advantages of the technical solution of this invention. As the results above show, due to the lack of licorice root essential oil in Comparative Example 5, the mice performed worse than the solution of the embodiment of this invention in all groups of behavioral experiments. This is because licorice root essential oil contains glycyrrhizin derivatives and trace amounts of volatile flavonoids, which can play a key "harmonizing" and "stabilizing" role in the compound. It can not only reduce oxidative stress indicators in brain tissue (such as MDA content) and protect damaged neurons, but also effectively delay the release rate of other volatile components in the compound, making the drug effect stable and lasting, and enhancing parasympathetic nerve tone, inducing the body to enter a physiological balance conducive to sleep initiation. Comparative Example 6 is a preferred compound formulation in the prior art. As shown in the results above, although its performance in various data is better than that of the single essential oil comparative example, it is still weaker than that of the embodiment of the present invention. This is because the main component of clove essential oil in Comparative Example 6 is eugenol, and its aromatic positioning is biased towards warming and relieving nausea and vomiting, which cannot achieve the effect of soothing the liver and relieving depression. Its correlation with the target of soothing the liver and relieving depression is weak, and the direction of compatibility is deviated. Although clove essential oil can be used to warm the spleen and stomach and inhibit bacteria, its effect on improving liver depression is limited. At the same time, its synergistic effect of combining with chuanxiong essential oil is enhanced. As a daily liver-soothing essential oil for long-term use and emotional relief, it is too irritating. Moreover, the combination of bupleurum, chuanxiong and clove has a messy aroma layer, large differences in volatility, uneven aroma retention, and uncoordinated top, middle and base notes, which affects the final effect and cannot achieve a good effect of soothing the liver, relieving depression and promoting sleep. The design of Comparative Example 7 demonstrates that the proportions of the essential oils selected in this invention are not random combinations, but rather the optimal solutions derived from extensive screening and summarization. Too much or too little of any component will affect the final "synergistic" and "harmonious" effects, thereby affecting the therapeutic effect. The sleep-aiding compound essential oil prepared by this invention uses a simple essential oil diffusion method to leverage the synergistic effect of multiple essential oil components, which can effectively alleviate insomnia symptoms and has a good sleep-aiding effect.

[0090] Expression of physiological indicators in mice The compound essential oil group of Example 3, which showed better relief of insomnia symptoms in mice in animal behavior studies, was selected for testing of the expression of various physiological indicators in mice.

[0091] Table 5. Results of neurotransmitter content in mouse brain tissue Based on research on the correlation between the function of the central monoamine neurotransmitter system and the mechanism of insomnia, abnormal levels of monoamine neurotransmitters and imbalances in amino acid neurotransmitter homeostasis are key factors leading to central wakefulness-sleep regulation disorders. Abnormal serotonin (5-HT) function can affect sleep initiation and maintenance, excessive dopamine (DA) activity promotes wakefulness and delays sleep, while insufficient function of γ-aminobutyric acid (GABA), as a major inhibitory neurotransmitter, weakens the central inhibitory process. (See Table 5 and...) Figure 12 , 13 As shown in Figure 14, compared with the control group, the levels of the inhibitory neurotransmitters 5-HT (2.12 ± 0.62 pg / mL) and GABA (2.25 ± 0.37 µmol / L) in the brain tissue of the model group mice were significantly decreased, while the level of DA (136.15 ± 1.53 pg / mL), which promotes wakefulness, was significantly increased (P<0.05). This suggests that in the insomnia model state, the central inhibitory function of mice is weakened and the level of wakefulness is abnormally hyperactive, which is generally manifested as an imbalance in the homeostasis of the neural excitation-inhibition system, which is highly consistent with the behavioral manifestations of insomnia. Compared with the model group, after the intervention in Example 3, the levels of each neurotransmitter in the mouse brain showed a significant recovery. Among them, the 5-HT level rose to 6.90 ± 0.63 pg / mL, the GABA level rose to 6.91 ± 0.36 µmol / L, while the DA level significantly decreased to 51.27 ± 8.35 pg / mL. This result indicates that Example 3, by regulating the release of monoamine neurotransmitters and restoring amino acid metabolic balance, enhanced the inhibitory and rhythmic stability of the nervous system, thereby exerting a significant sedative and sleep-inducing effect.

[0092] Table 6 Results of HPA axis hormone levels in mice Research on the neuroendocrine and central nervous system mechanisms of insomnia indicates that overactivation of the HPA axis is closely related to sleep-wake rhythm disorders. Prolonged stress or sleep disturbances can lead to HPA axis hyperactivity, abnormally elevated secretion of corticotropin-releasing hormone (CRH) and adrenocorticotropic hormone (ACTH), resulting in persistently high levels of terminal glucocorticoids. Excessive CORT not only disrupts the circadian rhythm system and sleep homeostasis but also enhances central nervous system arousal and inhibits normal sleep processes. (See Table 6 and...) Figure 15 , 16As shown in Figure 17, compared with the control group, the serum levels of ACTH (85.56 ± 7.07 pg / mL), CORT (13.08 ± 0.51 ng / mL), and CRH (85.56 ± 4.76 pg / mL) in the model group mice were significantly increased (P<0.05), indicating that the insomnia model successfully induced HPA axis hyperfunction and neuroendocrine rhythm disorder. Compared with the model group, after intervention in Example 3, all three indicators showed significant regression. Specifically, ACTH level decreased significantly to 30.36 ± 7.23 pg / mL, CORT level decreased to 6.50 ± 0.76 ng / mL, and CRH level decreased to 38.45 ± 4.63 pg / mL. The regression of ACTH and CORT was particularly significant, basically returning to the levels of the control group. This result indicates that the intervention can effectively inhibit the overactivation of the HPA axis, reduce the abnormal secretion of stress hormones, help restore neuroendocrine homeostasis and sleep-wake balance, thereby improving insomnia-related physiological and behavioral manifestations, and providing neuroendocrine-level mechanistic support for exerting sedative and sleep-promoting effects.

[0093] Table 7 Results of serum oxidative stress markers in mice As shown in Table 7 and Figure 18 As shown, oxidative stress is closely related to the occurrence and development of insomnia. Under long-term sleep disorders, the body generates more free radicals, leading to elevated levels of lipid peroxidation products such as malondialdehyde (MDA). As the end product of membrane lipid peroxidation, MDA levels directly reflect the degree of oxidative damage. Experimental results showed that compared with the control group (14.38 ± 2.01 ng / mL), the serum MDA level in the model group mice was significantly increased to 42.21 ± 2.89 ng / mL (P < 0.05), indicating that the insomnia model successfully induced oxidative stress in the body, leading to aggravated central and peripheral lipid peroxidation damage. Compared with the model group, after intervention in Example 3, the serum MDA level in mice significantly decreased to 21.33 ± 2.47 ng / mL, showing a significant recovery trend. This suggests that this intervention can effectively alleviate oxidative stress damage and inhibit the lipid peroxidation process. This function helps protect the integrity of sleep-related neural structures and maintain homeostasis, thus providing important antioxidant mechanisms to support improved sleep initiation and maintenance and enhanced sleep quality.

[0094] The above description of the embodiments is provided to enable those skilled in the art to understand and use the invention. It will be apparent to those skilled in the art that various modifications can be made to these embodiments, and the general principles described herein can be applied to other embodiments without inventive effort. Therefore, the present invention is not limited to the above embodiments, and any improvements and modifications made by those skilled in the art based on the disclosure of the present invention without departing from the scope of the invention should be within the protection scope of the present invention.

Claims

1. A compound essential oil for soothing the liver, relieving depression, and aiding sleep, characterized in that: The ingredients include the following components: 3-10 parts Bupleurum essential oil, 2-8 parts Cyperus essential oil, 1-5 parts Citrus aurantium essential oil, and 0.5-3 parts Glycyrrhiza uralensis essential oil.

2. The liver-soothing, mood-lifting, and sleep-aiding compound essential oil according to claim 1, characterized in that, The ingredients include: 4-8 parts Bupleurum essential oil, 3-6 parts Cyperus essential oil, 2-4 parts Citrus aurantium essential oil, and 1-2 parts Glycyrrhiza uralensis essential oil.

3. The liver-soothing, mood-lifting, and sleep-aiding compound essential oil according to claim 1, characterized in that, The ingredients include the following components: 5-7 parts Bupleurum essential oil, 4-5 parts Cyperus essential oil, 2-3 parts Citrus aurantium essential oil, and 1-1.5 parts Glycyrrhiza uralensis essential oil.

4. The liver-soothing, mood-lifting, and sleep-aiding compound essential oil according to claim 1, characterized in that, The ingredients include: 6 parts Bupleurum essential oil, 4 parts Cyperus essential oil, 2 parts Citrus aurantium essential oil, and 1 part Glycyrrhiza uralensis essential oil.

5. The liver-soothing, mood-lifting, and sleep-aiding compound essential oil according to claim 1, characterized in that, The ingredients include: 8 parts Bupleurum essential oil, 3 parts Cyperus essential oil, 3 parts Citrus aurantium essential oil, and 1 part Glycyrrhiza uralensis essential oil.

6. The liver-soothing, mood-lifting, and sleep-aiding compound essential oil according to claim 1, characterized in that, The ingredients include: 4 parts Bupleurum essential oil, 3 parts Cyperus essential oil, 3 parts Citrus aurantium essential oil, and 1 part Glycyrrhiza uralensis essential oil.

7. A method for preparing a liver-soothing, mood-lifting, and sleep-aiding compound essential oil as described in any one of claims 1-6, characterized in that, Includes the following steps: Weigh each component essential oil raw material into a container, stir, and you have the product.

8. The method for preparing a compound essential oil for soothing the liver, relieving depression, and aiding sleep according to claim 7, characterized in that, The stirring is carried out under light-protected conditions.

9. The method for preparing a compound essential oil for soothing the liver, relieving depression, and aiding sleep according to claim 7, characterized in that, The stirring is carried out under closed conditions.

10. The application of the liver-soothing, mood-regulating, and sleep-aiding compound essential oil as described in claim 1 in the preparation of pharmaceutical preparations and daily necessities for treating insomnia and anxiety.

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