Composition containing magnolia and phellodendron

Abstract

To provide a composition for relieving anxiety. [Solution] This is a composition for anxiety relief administered to mammals. It contains whey protein, magnolia extract, and phellodendron extract, and induces anxiety relief without sedation.

Description

Related Application

[0001] The present application claims the priority of U.S. Provisional Patent Application No. 62 / 212,080 filed on August 31, 2015, and incorporates the content of this provisional patent application by reference. Technical Field

[0002] The present invention generally relates to an anxiolytic composition containing Magnolia, Phellodendron, theanine, whey protein and / or S-adenosylmethionine. Background Art

[0003] Anxiety, fear and stress play important roles, and in a sense contribute to behavioral disorders in a large number of humans and non-human animals. For example, in dogs and cats, storm phobia, noise phobia, social avoidance, fear-related aggression, threat disorders and submissive urination are clear anxiety factors, and even problems such as urine marking, territorial aggression and resource guarding are amplified by fear and anxiety. In the United States, it is estimated that 23 million dogs suffer from fear disorders and anxiety disorders, and the costs for treatment and property damage reach 1 billion US dollars. Of the dogs transferred to rescue facilities, more than approximately 40% are abandoned due to behavioral problems, and 14% of cats are abandoned due to behavioral problems. For cats kept as pets, behavioral problems remain the most common cause of euthanasia. According to a recent survey of pet owners, 41% of dog owners report that their dogs occasionally show anxiety, and 29% of currently kept dogs are reported to have anxiety problems. The most common anxiety reported by these owners is noise phobia (17%), separation anxiety (13%) and generalized anxiety (5%).

[0004] Anxiety in horses is a common problem in the horse industry, contributing to behavioral problems, training issues, and performance decline. Depending on the case, elevated anxiety levels can lead to health problems such as stomach ulcers, and some studies have shown that this affects a large number of horses. In horses, anxiety can manifest not only as stomach ulcers but also as behavioral symptoms such as sudden galloping, jumping, panic behavior, difficulty getting into horse transport vehicles, lateral gait, refusal to enter or leave stalls, scratching the ground with hooves, and biting. In dogs, the most common risk factor for anxiety disorders is factors acquired in animal shelters and multiple foster homes, with less than 68.3% of dogs from shelters estimated to have some form of anxiety disorder. One previous study on behavioral diagnosis suggests that breed predictions may be possible in Dalmatians, English Springer Spaniels, German Shepherds, and mixed breeds. On the other hand, other studies suggest that breeds such as Cocker Spaniels, Schnauzers, and Dachshunds are at risk of developing separation anxiety. The incidence of anxiety disorders increases with age, largely due to cognitive impairments, with a prevalence of 22.5% in dogs under three years old and reaching 36.5% in dogs over eight years old.

[0005] Fear is an emotional response to the presence of a specific stimulus (such as an object, noise, or a person) that a pet perceives as a threat or danger. In contrast, anxiety is a response to concern or discomfort regarding an anticipated danger or threat. In other words, anxiety occurs even without an identifiable stimulus, whereas with fear, the stimulus is usually identifiable. While anxiety and the associated psychological stress do not become pathological in the short term, chronic stress and anxiety can impair a pet's health, comfort, and lifespan. Therefore, to ensure optimal health and comfort for all pets, veterinarians consider anxiety to be another possible pathological condition in pets. In extreme cases, phobias are intense, excessive, and abnormal fear responses that occur even without a real threat or regardless of the need to deal with an actual threat. Fear is a normal adaptive response, while phobias are abnormal, maladaptive, and generally interfere with normal functioning. Anxiety is a moderate response, often overlooked or misunderstood, but can have a significant impact on a pet's daily health.

[0006] In dogs, clinical signs of fear and anxiety include excessive vigilance, urination, destruction, excessive vocalization, excessive salivation, panting, hiding behavior, trembling, and escape behavior. In cats, chronic anxiety and fear can lead to secondary behavioral problems such as excessive grooming, spraying, and aggression among cats, predisposing them to health problems due to the sacrifice of the cat's immune system. Many of the clinical signs of fear and anxiety are harmful and distressing to both pets and owners, so it should be easy to understand why owners seek advice about these disorders. No coincidence, the most common risk factors for abandoning animals to shelters, and the most common risk factors for euthanasia of both dogs and cats, are house-soiling (improper urination), destruction, aggression, and excessively abnormal behavior, all of which are potential clinical signs of anxiety.

[0007] Anxiety disorders rarely occur in isolation; they often manifest in combination. While thunderstorm phobia and noise phobia do not necessarily occur simultaneously, thunderstorm phobia, noise phobia, and separation anxiety are more frequently observed together than in isolation. This evidence suggests that the precise cause of pet anxiety is difficult to determine, and that multiple conditions may be present in the same pet.

[0008] In humans, while the triggers and manifestations of behavioral disorders differ, anxiety, fear, and stress are predisposing factors for these disorders. Attempts to alleviate behavioral disorders in humans and non-human animals have led to the development of certain synthetic drugs. Examples include clomipramine hydrochloride, fluoxetine hydrochloride, benzodiazepines, and acepromazine maleate, all of which are drug compositions used in attempts to alleviate anxiety in humans and / or non-human animals. While these drugs offer some relief from anxiety, a major side effect is sedation, causing drowsiness in humans and non-human animals. Furthermore, many of these drugs are synthetic compositions, making them undesirable for humans to ingest and also raising concerns about administering them to pets.

[0009] The biochemical processes of anxiety are extremely complex, and much remains unknown. Research suggests that almost every type of neurotransmitter and hormone, from serotonin, gamma-aminobutyric acid (GABA), glutamate, and dopamine to cortisol, adrenaline, and even thyroid hormones, tends to play some role in anxiety. In many ways, anxiety is simply a biochemical response to brain stress. Often, when something disrupts the delicate chemical balance in the brain, anxiety results.

[0010] Therefore, ideal anxiety management must be multimodal, meaning that one or more mechanisms of action should target and correct the underlying chemical imbalance. However, FDA-approved drugs such as serotonin selective reuptake inhibitors (SSRIs), tricyclic antidepressants (TCAs), and benzodiazepines generally act through a single mechanism of action.

[0011] Because it is difficult to track adverse drug reactions in combination with other medications, the FDA is not in favor of approving drugs with multiple active ingredients. Therefore, a multimodal approach using only drugs is not feasible. Nutritional and botanical supplements, which are regulated by the FDA as foods rather than drugs, can be configured as novel multi-component administration methods and offer greater opportunities for the multimodal management of anxiety. For this reason, and for other reasons, there is an increasing demand for the use of natural products in humans and non-human animals, as consumers now seek drug alternatives. Currently, some of these natural products are already incorporated into dietary supplements and various foods.

[0012] As mentioned above, there is a demand for natural product compositions with anxiety-relieving properties. These compositions need to help support normal behavior and promote sedative effects without inducing extreme drowsiness. Furthermore, such compositions must have synergistic effects to manage and control the clinical signs or symptoms of anxiety. [Overview of the project]

[0013] In accordance with the objectives and effects described herein, one aspect of the present invention provides an anxiety-relieving composition comprising L-theanine and whey protein in a synergistic combination. In the embodiment, the whey protein is α-lactalbumin. The anxiety-relieving composition of the present invention introduces an anxiety-relieving alteration in the release pattern of one or more neurotransmitters, including gamma-aminobutyric acid (GABA) and serotonin. The composition of the present invention can be formulated for oral administration to mammals, including humans, pet animals, and equids.

[0014] In embodiments of the present invention, the synergistic composition further comprises an extract of magnolia and an extract of phellodendron. In embodiments of the present invention, the synergistic composition further comprises RELORA.

[0015] Another aspect of the present invention relates to a method for suppressing, alleviating, or treating symptoms of anxiety in mammals by administering a synergistic composition having L-theanine and whey protein. In embodiments of the present invention, the synergistic composition further comprises magnolia extract and phellodendron extract. In embodiments of the present invention, the whey protein is provided as a whey protein concentrate having α-lactalbumin. In the method of the present invention, the synergistic composition is orally administered in an amount sufficient to induce an anxiety-relieving change in the release pattern of one or more neurotransmitters, including gamma-aminobutyric acid (GABA) and serotonin. The target animals may be humans, canids, felines, or equids.

[0016] In yet another embodiment, the anxiety-relieving composition comprises a composition exhibiting a synergistic effect of L-theanine and S-adenosylmethionine (SAMe). This synergistic composition introduces anxiety-relieving changes into the release patterns of one or more neurotransmitters, including glutamic acid and gamma-aminobutyric acid (GABA). This composition can be formulated for oral administration to mammals. In embodiments of the present invention, SAMe comprises SAMe phytate.

[0017] In yet another embodiment, the anxiety relief composition comprises a composition exhibiting a synergistic effect of L-theanine, whey protein, and RELORA. In embodiments of the present invention, the whey protein comprises α-lactalbumin. In embodiments of the present invention, the whey protein is a whey protein concentrate.

[0018] In one embodiment of the present invention, the composition comprises at least 2.0 mg of L-theanine, at least 0.5 mg of whey protein, and at least 0.5 mg of RELORA. In another embodiment of the present invention, the composition comprises at least 2.0 mg of L-theanine, at least 0.5 mg of whey protein, and at least 3.0 mg of RELORA.

[0019] The following describes some preferred embodiments of the anxiety-relieving compositions and corresponding methods of the present invention. As those skilled in the art will understand, the compositions and methods of the present invention can be implemented in other different embodiments, and some details can be modified in various obvious embodiments without departing from the compositions and methods described in the claims. Accordingly, the accompanying drawings and description are illustrative and not intended to be limiting. [Brief explanation of the drawing]

[0020] Several embodiments of the anxiety-relieving compositions of the present invention are shown in the accompanying drawings, which are attached to and constitute part of this specification. These drawings, along with a detailed description of the invention, illustrate the principles of the present invention. [Figure 1]It is a figure showing basal serotonin (5-HT) output after administration of the composition of the present invention. [Figure 2] It is a figure showing basal GABA output after administration of the composition of the present invention. [Figure 3] It is a figure showing basal glutamate output after administration of the composition of the present invention. [Figure 4] It is a figure showing a comparison of GABA output after administration of one embodiment of the composition of the present invention. [Figure 5] It is a figure showing a comparison of glutamate output after administration of the embodiment shown in Figure 4. [Figure 6A] It is a figure showing the results of an elevated plus maze (EPM) test after administration of the composition of the present invention. [Figure 6B] It is a figure showing a comparison of EPM test results after administration of one embodiment of the composition of the present invention. [Figure 6C] It is a figure showing a comparison of EPM test results after administration of another embodiment of the composition of the present invention. [Figure 7A] It is a figure showing the results of an open field (OF) test after administration of the composition of the present invention. [Figure 7B] It is a figure showing a comparison of OF test results after administration of one embodiment of the composition of the present invention. [Figure 7C] It is a figure showing a comparison of OF test results after administration of another embodiment of the composition of the present invention.

[0021] Hereinafter, embodiments of the anxiolytic composition of the present invention will be described in detail. Examples of the present invention are shown in the accompanying drawings. DETAILED DESCRIPTION OF THE INVENTION

[0022] This invention relates to a composition of one or more substances, including magnolia, phellodendron amurense, theanine, S-adenosylmethionine (SAMe), and / or whey protein, which can be applied to humans or non-human animals. In one embodiment, the composition of the present invention contains Magnolia officinalis, Phellodendron amurense, L-theanine, and α-lactalbumin. When these substances are used together, a number of different biochemical processes are involved. That is, the composition of the present invention enables a variety of approaches to anxiety relief. While not bound by any particular theory, one potential mode is to act on multiple neurotransmitters (such as glutamate and GABA), to increase the production of neurotransmitters such as serotonin, or to alter electroencephalogram activity.

[0023] To further explain the background of this invention, Magnolia officinalis is a species of magnolia native to China. The bark and / or extracts of Magnolia officinalis contain honokiol and magnolol, which enhance the activity of both synaptic GABA receptors and extrasynaptic GABA receptors in the brain. GABA is the chief inhibitory neurotransmitter in the brain that regulates the function of neurons that have become extremely excited by excessive stimulation of fear and anxiety. The action of these compounds is thought to be due to their selective binding to specific GABA receptors, which explains why honokiol and magnolol exhibit anxiety-relieving effects without showing sedative effects.

[0024] Phellodendron amurense is a species of tree commonly known as Phellodendron amurense. The fruit, bark, and / or extracts of Phellodendron amurense are rich in the compound berberine. When Phellodendron amurense extract is used in combination with magnolia, a synergistic effect is achieved, resulting in a stronger stress and anxiety-reducing effect than when used individually. Generally, a synergistic effect refers to the effect where the combined use of two or more components yields a stronger result than the sum of the effects of using each component individually. In a preferred embodiment of this invention, the synergistic effect is stronger than the additive effect. The synergistic effect observed when Phellodendron amurense extract is used in combination with magnolia can be attributed to the fact that berberine inhibits the release of glutamate into the synaptic cleft by presynaptic neurons. Such a combination allows for the modulation of both the excitatory neurotransmitter glutamate and the inhibitory neurotransmitter GABA at the synaptic level. In laboratory models, the combined use of magnolia and Phellodendron amurense can alleviate anxiety in beagle dogs in placebo-type clinical trials of noise-induced anxiety. In some embodiments of the present invention, a commercially available formulation (RELORA, InterHealth Nutraceuticals, Inc., Benicia, Canada) can be administered as a combination drug of magnolia and phellodendron.

[0025] The neurotransmitters glutamate and GABA act antagonistically, regulating interactions between neurons in the brain. Glutamate is the primary excitatory neurotransmitter, increasing the sensitivity of neurons to stimuli and electrical impulses. Glutamate plays a crucial role in fear and anxiety, which are often the result of excessive stimulation. GABA is the primary inhibitory neurotransmitter in the nervous system, playing an equally important role in controlling anxiety by sedating and reversing the effects of glutamate.

[0026] In one embodiment, magnolia extract is used in combination with Phellodendron amurense extract, and this combination exhibits a synergistic effect, more effectively controlling stress and anxiety than when the compounds are used alone. This synergistic effect is due to the effects these compounds exert on both glutamate and GABA at the synaptic level. Magnolia officinalis extract, specifically its components honokiol and magnolol, enhances the activity of both GABA receptors at synapses and extrasynaptic GABA receptors in the brain. Phellodendron amurense extract contains berberine. Berberine inhibits the release of glutamate into the synaptic cleft by presynaptic neurons. In other words, magnolia extract enhances the GABA-stabilizing effect, and berberine from Phellodendron amurense extract inhibits the release of excitatory glutamate.

[0027] Berberine is a bright yellow ammonium salt found in Phellodendron amurense, and is also present in plants such as Oregon grapes, barley, buttercups, coptis, and turmeric. Berberine is typically found in the roots, rhizomes, trunks, and bark of these plants.

[0028] L-theanine is a structural analogue of the amino acid glutamate and is one of the most important excitatory neurotransmitters in the nervous system. Naturally present in various tea plants, theanine is thought to exert its neuroprotective effects by binding to and blocking glutamate receptors, thereby suppressing excitatory impulses and reducing the stimulating effects of glutamate. Theanine increases the stability levels of neurotransmitters such as serotonin, dopamine, and GABA in the brain. Furthermore, theanine directly stimulates the generation of alpha brain waves, creating states of deep relaxation, alertness, and mental alertness.

[0029] Alpha-lactalbumin is a component of certain whey protein compositions and a high-quality protein source that supplies amino acids to animals. Milk has long been considered a beverage with postprandial sedative properties, particularly in infants and young animals. The first human studies on the anxiety-relieving effects of milk began in the 1930s, confirming that certain proteins in milk have sedative effects. Alpha-lactalbumin exerts neuroprotective properties by providing amino acid precursors to the antioxidant glutathione (cysteine) and the mood-enhancing neurotransmitter serotonin (tryptophan).

[0030] The brain consumes nearly 20% of the body's oxygen, resulting in extremely high rates of reactive oxygen species (ROS) production, making brain cells particularly susceptible to oxidative damage. When fully developed neurons in the brain are lost, this cannot be compensated for by the generation of new neurons. In other words, an imbalance between ROS production and antioxidants leads to several types of neurological disorders. Glutathione is the dominant antioxidant present in the nervous system. The glutathione content of brain cells strongly depends on the extent to which they can utilize glutathione precursors. Alpha-lactalbumin supplements cysteine, the amino acid precursor of glutathione.

[0031] Since brain serotonin levels rise under stress, neurotransmitters are important substances that regulate emotional states and mood. When chronic stress and anxiety are present, the available concentrations of serotonin and its precursor, tryptophan, become insufficient, and serotonin may not reach the functionally necessary levels. Alpha-lactalbumin contains a mixture of tryptophan and other amino acids derived from natural food protein sources, and studies suggest that dietary supplementation with alpha-lactalbumin enhances brain tryptophan and serotonin activity, thereby improving cognitive abilities in stress-sensitive subjects. In one embodiment of the present invention, alpha-lactalbumin synergistically enhances the serotonin effects of other components of the composition, thus adding another mode of operation.

[0032] S-adenosylmethionine (SAMe) is a naturally occurring compound found in tissues throughout the body. At the molecular level, SAMe is involved in various metabolic pathways, including the transmethylation, transsulfation, and aminopropylation pathways. In the body, SAMe is synthesized from amino acids, methionine, and triphosphate nucleotides, and then participates in the biosynthesis of numerous biological molecules, including hormones and neurotransmitters.

[0033] SAMe is known to exert various health-promoting effects when administered to subjects. SAMe regulates gene expression and helps prevent gene mutations. It also maintains mitochondrial function, participates in phospholipid synthesis, maintains cell membrane integrity, and regulates neurotransmitters such as serotonin, dopamine, and epinephrine (adrenaline), as well as hormones such as estrogen and melatonin. Furthermore, SAMe is known to suppress post-ischemic neuronal death, improve glucose utilization in the brain, suppress hydrocephalus, normalize EEG and induced potential findings, and improve motor function, including motor function impaired by seizures. For example, in meta-analyses of numerous drug studies, SAMe enhances emotional health and shows comparable efficacy to many prescription drugs, but with significantly fewer side effects. SAMe is also used to treat anxiety, chronic pain, arthritis, rheumatic fibromyalgia, chronic fatigue syndrome, cognitive impairment associated with Alzheimer's disease, neurovascular disorders, and neurological disorders associated with AIDS. SAMe can treat not only central and peripheral nervous system disorders, but also improve joint diseases, cardiovascular diseases, and liver diseases.

[0034] Initially, SAMe was considered impractical due to the instability of its ions during manufacturing, transport, and storage. However, stable salts of SAMe were eventually developed. Examples include SAMe tosylate disulfate, SAMe butanedisulfonate salt, SAMe di-p-toluenesulfonate disulfate salt, SAMe tri-p-toluenesulfonic acid salt, SAMe butanedisulfonate salt, and SAMe disulfate p-toluensulfonate salt. In exemplary embodiments of the present invention, SAMe is administered as a commercially available SAMe salt of phytic acid.

[0035] Those skilled in the art will understand that the composition of Magnolia officinalis, Phellodendron amurense, L-theanine, and whey protein can contain each component in any amount. Each component may also be in the form of a concentrate, such as a whey protein concentrate. Furthermore, this composition can be used on human subjects and non-human animal subjects (mammals, birds, fish, reptiles, etc.).

[0036] In one embodiment, a composition intended for non-human animals may contain 0.5 to 3 mg of Magnolia officinalis, 0.03 to 0.2 mg of Phellodendron amurense, 17.0 to 450 mg of L-theanine, and 12.0 to 100 mg of α-lactalbumin. Alternatively, in another embodiment, a composition intended for non-human animals may contain 0.01 to 10 g of Magnolia officinalis, 0.01 to 10 g of Phellodendron amurense, 0.01 mg to 10 g of L-theanine, and 0.01 mg to 10 g of α-lactalbumin.

[0037] In one embodiment, the composition of the present invention may contain at least 0.5 mg of RELORA, at least 2.0 mg of L-theanine, at least 0.5 mg of whey protein, and at least 1.5 mg of SAMe.

[0038] In one embodiment, the composition of the present invention, intended for non-human animals such as canids, may contain at least 6.0 mg of RELORA, at least 3.0 mg of L-theanine, at least 1.0 mg of whey protein, and at least 3.0 mg of SAMe.

[0039] In one embodiment, the present invention composition for non-human animals such as felines may contain at least 3.0 mg of RELORA, at least 2.0 mg of L-theanine, at least 0.5 mg of whey protein, and at least 1.0 mg of SAMe.

[0040] In one embodiment, the present invention composition for non-human animals of the equid family may contain at least 20.0 mg of RELORA, at least 15.0 mg of L-theanine, at least 5.0 mg of whey protein, and at least 10.0 mg of SAMe.

[0041] In one embodiment, the composition of the present invention intended for human subjects may contain at least 20.0 mg of RELORA, at least 15.0 mg of L-theanine, at least 5.0 mg of whey protein, and at least 10.0 mg of SAMe.

[0042] In one embodiment targeting pet animals such as dogs and cats, the composition of the present invention may contain 37 mg of RELORA, 17 mg of L-theanine, and 12 mg of whey protein as a single dose.

[0043] In one embodiment targeting pet animals such as dogs and cats, the composition of the present invention may contain 75 mg of RELORA, 35 mg of L-theanine, and 25 mg of whey protein as a single dose.

[0044] In one embodiment targeting pet animals such as dogs and cats, the composition of the present invention may contain 450 mg of RELORA, 205 mg of L-theanine, and 100 mg of whey protein as a single dose.

[0045] In one embodiment targeting pet animals such as dogs and cats, the composition of the present invention may contain 0.5 mg of Magnolia officinalis extract, 0.03 mg of Phellodendron amurense extract, 17 mg of L-theanine, and 12 mg of α-lactalbumin as a single dose.

[0046] In one embodiment targeting equid animals, the composition of the present invention may contain 750 mg of RELORA, 150 mg of L-theanine, and 1000 mg of whey protein.

[0047] In one embodiment targeting equid animals, the composition of the present invention may contain 1500 mg of RELORA, 300 mg of L-theanine, and 1000 mg of whey protein.

[0048] Those skilled in the art will understand that each component, magnolia, Phellodendron amurense, theanine, and / or whey protein (or α-lactalbumin), can be arbitrarily combined and bound together. For example, one embodiment may consist of magnolia extract and α-lactalbumin, without using Phellodendron amurense or theanine. Alternatively, the composition of the present invention may consist of Phellodendron amurense extract and α-lactalbumin, without using magnolia or theanine. Furthermore, we can provide a single component from magnolia, Phellodendron amurense, theanine, and / or whey protein (or α-lactalbumin) that gives the desired reaction, or a combination having these single components. The formulation may also contain other flavonoids, ω-3 fatty acids, eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), valerian, SAMe, or other milk proteins (such as α-S1 trypsin casein), or milk protein-containing products, and / or Sceletium tortuosum.

[0049] The compositions of the present invention can be combined in any way and can be provided to humans or non-human animals in any combination. In one embodiment of the present invention, the compositions of the present invention have a unit dosage / administration form, but are not limited to this form. Examples of preferred methods include oral administration, rectal administration, intravenous injection, transdermal administration, intramuscular injection, transdermal administration, transmucosal administration, and local administration.

[0050] The compositions of the present invention have an orally administered dosage form. Examples of orally administered dosage forms, though not limited to them, include tablets, capsules, powders that can be dispersed in liquid or sprinkled on food, solutions, liquids such as suspensions and emulsions, soft gels / chewing capsules, chewing bars, and other known dosage forms. In some embodiments of the present invention, the compositions of the present invention include tablets, capsules, or soft chewing forms. The orally administered dosage forms may be immediate-release, delayed-release, or delayed-release. The compositions of the present invention may or may not be coated. Example 1: Evaluation of neurotransmitter release patterns in hippocampal dialysis samples from C57B1 / 6 mice treated with an anxiety-relieving composition for 7 days.

[0051] The release patterns of neurotransmitters in the brain were measured by a procedure in which a microdialysis probe was inserted into the hypothalamic region of experimental animals and the probe was flushed by dialysis using artificial cerebrospinal fluid (CSF). This is an accepted common method for obtaining and measuring in vivo brain levels of neurotransmitters and other small proteins.

[0052] The recovered fluid represents the extracellular and interstitial fluid present in the hippocampus region of the brain. This region is generally preferred and common for microdialysis because any changes measured in this area of ​​the brain are thought to represent approximately 80% of the total brain tissue. The levels of neurotransmitters in the fluid represent not only the complex balance between the release of these neurotransmitters by presynaptic neurons and their reuptake into neurons, but also the interactions between neurotransmitters, if any. Therefore, these results indicate how compounds and compound compositions affect the neurochemistry of the brain.

[0053] Female C57B1 mice were divided into groups of 6 and administered orally with various compounds or a placebo for 7 days for evaluation. On day 6, each mouse was anesthetized, and a microdialysis probe was inserted into the hippocampus of the brain. Each probe was connected to a microperfusion pump, and microdialysis sampling was started the day after surgery while perfusing with artificial CSF. On day 7, samples were taken in 30-minute aliquots 90 minutes before administration of each compound. Next, each aliquot was analyzed, and GABA, glutamate, and serotonin (5-HT) in each sample were quantified. Neurotransmitter analysis was performed in two stages to confirm the chronic and acute effects of each compound.

[0054] In the results described below, compound A is L-theanine, compound B is a composition containing Magnolia officinalis extract and Phellodendron amurense extract, compound C is a whey protein containing α-lactalbumin, compound D is SAMe, and compound E is glutathione. Compound B used in the embodiment is a commercially available formulation (RELORA, InterHealth Nutraceuticals, Inc. Benicia, CA). Compound C used in the embodiment was standardized to 80% α-lactalbumin. Compound D used in the embodiment was commercially available SAMe phytate. The dosage of compound A was 6.66 mg / kg, compound B was 12.5 mg / kg, compound C was 2.36 mg / kg, compound D was 6.02 mg / kg, and compound E was 13.6 mg / kg. Equivalent dosages for interspecies extrapolation can be calculated by a person skilled in the art using a standard dosage conversion method. Basic neurotransmitter output

[0055] First, the baseline output levels of each neurotransmitter were measured to determine the long-term chronic effects of each compound on neurotransmitters. The baseline level represents the lowest level of each neurotransmitter in a 24-hour cycle. These levels were determined by averaging the levels of each neurotransmitter in three 30-minute aliquots prior to the 7th day of administration (Sample -90 min to -60 min, Sample -60 min to -30 min, and Sample -30 min to 0 min). The baseline level was measured to represent the chronic effect of the compound on neurotransmitter levels; that is, it measured the minimum effect observed before initial administration. Next, to determine the acute effects of these compounds, the levels of each neurotransmitter were analyzed in aliquots administered up to 3.5 hours (210 minutes) after the 7th day of administration for each compound / composition.

[0056] Basal levels of serotonin (5-HT), GABA, and glutamate were measured not only for each individual component of the above composition, but also after administration of compositions containing each component for 6 days. Next, the mean concentrations of each neurotransmitter were compared with the mean basal levels measured for the placebo group using a student test to determine if there was a significant difference between the two sets of data.

[0057] Regarding serotonin, none of the evaluated compounds showed a statistically significant effect (p-value < 0.05) on basal serotonin output (see Figure 1). Specifically, L-theanine, RELORA, and whey protein all showed no effect on basal serotonin levels. On the other hand, the composition A+B+C (theanine + magnolia / Phellodendron amurense + whey protein) showed a statistically significant effect (p-value < 0.05) on basal serotonin output. Compositions containing L-theanine and whey protein (A+C) also resulted in significantly lower basal serotonin levels. These results explain the unexpected synergistic effect of the compositions on brain serotonin levels.

[0058] The compositions of theanine / magnolia / Phellodendron amurense (A+B) and L-theanine and whey protein (A+C) showed unexpected synergistic improvements in GABA levels (see Figure 2). Similarly, the composition of theanine and SAMe (A+B) also significantly improved basal GABA output.

[0059] Regarding basal glutamate output (see Figure 3), an increase was observed with the theanine / magnolia / phellodendron (A+B) composition. Similarly, a significant improvement in basal glutamate output was also observed with the theanine and SAMe (A+B) composition. Acute neurotransmitter output

[0060] After the seventh dose of each compound and / or composition, CSF aliquots were collected in 30-minute increments up to 210 minutes post-administration. The neurotransmitter concentrations in each aliquot show acute, short-term changes not reflected in the baseline output results.

[0061] The results were plotted on a graph. The x-axis represents the time after administration on day 7, and the y-axis represents the percentage increase or decrease in basal output observed in the first half of the experiment. This measurement is used because the actual concentrations of neurotransmitters vary significantly among the animal subjects. This method allows for data standardization and essentially allows each animal to account for its own individual control. Next, analysis of variance (ANOVA) was used to analyze the results and detect any statistically significant therapeutic effects.

[0062] First, examining the acute effects of these compounds on GABA, analysis by ANOVA (p<0.05) showed that, compared to placebo during the study period, no statistically significant therapeutic effect was observed for individual components A, B, or C. However, as shown in Figure 4, ANOVA analysis revealed a statistically significant difference in the therapeutic effect of the (A+B+C) composition compared to placebo. Therefore, these results explain that the composition of L-theanine, magnolia / Phellodendron amurense, and whey protein (A+B+C) shows a significant synergistic effect on GABA, but each component itself does not show an effect.

[0063] Regarding glutamate, neither the individual components (A, B, C) nor the composition (A+B+C) showed a direct therapeutic effect on glutamate levels. However, these results explain the tendency for the composition (A+B+C) to exert its therapeutic effect 120-210 minutes after administration (see Figure 5). Those skilled in the art will understand that changes in glutamate levels are one of the more difficult neurotransmitter changes to induce. Example 2: Evaluation of the anxiety-relieving properties of the composition of the present invention by behavioral testing.

[0064] Wistar rats (16 rats / group) were administered individual components (L-theanine, magnolia / Phellodendron, and whey protein), as well as several compositions of these three components, including a three-way composition (theanine + magnolia / Phellodendron + whey protein). The magnolia / Phellodendron composition used was a commercially available product (RELORA, InterHealth Nutraceuticals, Inc., Benicia, Canada). The administered whey protein was standardized to 80% α-lactalbumin. The rats were administered daily for 14 days, followed by a series of behavioral assessments to detect whether the rats experienced anxiety-relieving or sedative effects. The treatment groups were as follows: 1: L-theanine + Magnolia / Phellodendron 2: L-theanine + whey protein 3: Magnolia / Phellodendron + Whey Protein 4: L-theanine + Magnolia / Phellodendron amurense + Whey protein 5: L-theanine 6: Magnolia / Yellow Amur Cork Tree 7: Whey protein The rats were divided into the following groups. TIFF2026139799000002.tif72163 Elevated Cross Maze Test

[0065] The elevated cross maze (EPM) test utilizes the natural aversion (characteristic anxiety) of rodents in a brightly lit, open elevated area. EPMs have extremely high predictive power and are often used to elucidate the potential anxiety-relieving activity of compounds. During this test, rats are videotaped after being placed in the elevated maze. The video footage is analyzed to determine the percentage of time the rats spend on the open elevated section of the maze. Rats that spend longer on the open arms are considered to be experiencing less anxiety. Table 1 below shows the average time each rat spent on the open arms of the maze. Table 1 TIFF2026139799000003.tif51105

[0066] In this experiment, rats administered with the composition containing the components of Treatment 4 stayed on the open arm of the maze for an average of 3 minutes longer than rats administered with a placebo (see Figure 6A), demonstrating the anxiety-relieving effect of the compound. Similarly, rats administered with the composition containing L-theanine / whey protein (Treatment 2) stayed on the open arm of the maze for an average of 5 minutes longer than rats administered with a placebo (see Figure 6), demonstrating the anxiety-relieving effect of the compound. The L-theanine / magnolia / Phellodendron composition (Treatment 1) and the magnolia / Phellodendron / whey protein composition (Treatment 3) showed statistically significant effects in post-hoc comparisons (both analyzed at p<0.05; see Figures 6B and 6C, respectively). Open field activity as a measure of sedation

[0067] Walking activity in open-field tests is an excellent measure of a compound's sedative properties. For example, a significant side effect of many anxiety-relieving compounds, such as benzodiazepines, is a marked sedative effect associated with their administration. While these substances are highly effective anxiety relievers, they often possess undesirable sedative effects.

[0068] This test involves placing rats in a brightly lit enclosure with free movement. The rats are videotaped, and the total distance they walk during the test is recorded as an indicator of walking activity. Sedated rats walk shorter distances, while active rats walk longer distances. Table 2 TIFF2026139799000004.tif51105a: One rat was excluded from the analysis due to inaccurate video tracking.

[0069] The results, shown in Table 2 and illustrated in Figure 7A, indicate that none of the administered compounds / compositions had a sedative effect on the walking motor activity of the rats. All rats were more active than those in the placebo group and showed a voracious interest in exploration and detection. In fact, post-hoc comparisons showed a tendency toward increased walking motor activity. This was particularly true for treatment 1 [t(30)=3.58, p<0.01; t-test] and treatment 3 [t(30)=1.99, p<0.06; t-test] (see Figures 7B and 7C). Compounds that exhibit anxiety-relieving effects without showing sedative effects are considered highly desirable candidates for anxiety treatments.

[0070] From these results, it should be understood that we can provide a composition that is suitable for oral administration and is effective, and that it can affect neurochemistry, as demonstrated by the fact that it can introduce changes in the release patterns of various neurotransmitters in various embodiments. Furthermore, various behavioral evaluations demonstrate the anxiety-relieving effects of the various compositions of the present invention. Although the individual compounds of the compositions of the present invention do not show effects on neurochemistry or behavior, surprisingly, a synergistic effect was observed in the compositions.

[0071] The above description is for illustrative purposes only and is not exhaustive; in other words, it is not limited to the exact forms disclosed. Obvious modifications are possible in light of the above disclosure. The above embodiments have been selected to allow those skilled in the art to implement the invention in various forms and to provide the best possible implementable modes to address the specific intended uses. These modifications are included within the claims when interpreted in a fair, legal, and equitable manner.

Claims

[Claim 1] An anxiety-relieving composition for administration to mammals in need thereof, comprising whey protein, magnolia extract, and phellodendron extract, which induces anxiety-relieving changes without sedative effects.

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