A probiotic supplement for improving depression, anxiety and sleep disorders

Abstract

The present invention belongs to the technical field of probiotic food, and specifically relates to a probiotic supplement for improving depression, anxiety and sleep disorders. The probiotic supplement includes mixed probiotics, and the mixed probiotics are composed of Lactobacillus plantarum, Bifidobacterium breve, Bifidobacterium longum subspecies longum, Weizmannia coagulans and Lactobacillus rhamnosus, and the mass ratio is 5-20:5-40:5-30:5-40:1-10. The probiotic supplement of the present invention can make the effect of improving depression, anxiety and sleep disorders more accurate, more significant and more lasting than a single strain, and the probiotic supplement is highly safe and not easy to produce drug resistance by enhancing the body's own neurotransmitter production ability.

Description

Technical Field

[0001] The invention belongs to the technical field of probiotic foods, and in particular relates to a probiotic supplement for improving depression, anxiety and sleep disorders. Background Art

[0002] With the acceleration of the aging process of the population, the health problems of the elderly have become an important topic in medical and sociological research. The deterioration of gastrointestinal function and the imbalance of intestinal flora constitute important challenges to the physical health of the elderly. With the increase of age, the physical and physiological functions of the human gastrointestinal tract gradually decline, which is manifested by reduced gastric acid secretion, reduced digestive enzyme activity and slower intestinal motility. These changes not only lead to decreased digestive efficiency and poor nutrient absorption, but also have a negative impact on the intestinal microenvironment, thereby causing an imbalance of intestinal flora.

[0003] However, an imbalance in the intestinal flora can affect the function of the nervous system through multiple pathways such as metabolites and immune responses, thereby increasing the risk of anxiety, depression and sleep disorders in the elderly. The intestinal flora directly acts on the central nervous system by releasing metabolites, while affecting the intestinal mucosal immune system and intestinal barrier function, and regulating the synthesis and release of neurotransmitters. Various neurotransmitters such as 5-hydroxytryptamine (5-HT) and γ-aminobutyric acid (GABA) are produced in the intestine and transmit information to the brain through the vagus nerve, affecting emotions and behavioral responses. Short-chain fatty acids (SCFAs) such as acetate, butyrate and propionate metabolites of the intestinal flora have an important regulatory effect on the central nervous system. They can enter the brain through the blood-brain barrier, regulate neuroinflammation and the activity of nerve cells, and thus affect emotions and behavior.

[0004] Therefore, the intestinal flora plays an important role in regulating the sleep process. Normal intestinal flora can produce metabolites that affect the secretion of melatonin, thereby regulating the circadian rhythm. On the contrary, an imbalance in the intestinal flora will disrupt the normal production of these metabolites, thereby disrupting the secretion of melatonin and the circadian rhythm. In addition, sleep disorders themselves will in turn affect the composition of the intestinal flora, aggravating the imbalance, forming a vicious circle, and further aggravating the sleep disorders and mental health problems of the elderly.

[0005] At present, probiotics can play a potential role in protecting mental health by improving intestinal barrier function, regulating immune response and promoting the production of neurotransmitters. The lactic acid produced by probiotics can enhance the effect of the neurotransmitter γ-aminobutyric acid (GABA) and relieve anxiety and depression symptoms. Acetic acid in short-chain fatty acids can regulate the release of neurotransmitters by entering the blood circulation and affecting the hypothalamus-pituitary-adrenal (HPA) axis, which has a positive effect on emotional balance and sleep quality. At the same time, probiotics can regulate the production of neurotransmitters by regulating the precursors of neurotransmitters related to mood and sleep, such as tryptophan, glutamate, glutamine, etc. Prebiotics mainly improve the balance of intestinal flora indirectly by selectively promoting the growth of beneficial bacteria. Many studies have shown that probiotics can effectively relieve anxiety and depression symptoms and promote the synthesis of neurotransmitters. They are considered to have a positive effect on alleviating depressive symptoms and improving intestinal flora.

[0006] For example, Chinese patent CN 118575952 A discloses a probiotic composition, preparation method and application thereof for improving the balance of intestinal flora. The probiotic composition comprises the following raw materials in parts by weight: 1-20 parts of composite probiotic powder, 20-80 parts of inulin, 10-30 parts of whole milk powder, and 0.1-5 parts of vitamin C. The composite probiotic powder comprises one or more of lactobacillus, bifidobacterium, streptococcus, lactococcus and pediococcus. The probiotic composition can effectively relieve gastrointestinal discomfort symptoms, reduce the impact of intestinal flora imbalance and help the intestinal flora return to normal. However, it only focuses on regulating intestinal flora and has not studied mood disorders (anxiety and depression). Another example is the Chinese patent CN 117327631A, which only focuses on mood disorders and does not conduct specific research on intestinal flora. Moreover, the strain selection of this patent is relatively single and lacks a diverse strain combination, which may limit its wide application in targeting specific psychological problems such as anxiety and depression. It ignores the potential benefits of probiotic diversity to different individual patients and cannot fully cover different types of anxiety and depression manifestations.

[0007] Another example is Chinese patent CN 111728111A, which discloses a probiotic composition for relieving anxiety or depression and its application. The probiotic composition comprises prebiotics, a K-KLJ probiotic combination and a functional substance, wherein the K-KLJ probiotic combination contains at least one of plant lactobacillus, Lactobacillus reuteri, Lactobacillus casei, and Lactobacillus helveticus, or also includes at least one of Lactobacillus rhamnosus, Lactobacillus acidophilus, Bifidobacterium animalis, and Bifidobacterium longum. The probiotic composition invented in this patent can significantly increase the number of beneficial intestinal flora, improve intestinal discomfort, reduce chronic inflammation, thereby reducing bad emotions, and preventing and treating emotional diseases such as depression and anxiety. Although the patent mentions both improving intestinal discomfort and preventing and treating depression and anxiety, only emotional research was conducted subsequently, and no research on the effect of improving intestinal flora was conducted.

[0008] In summary, there is an urgent need to develop a probiotic supplement that can regulate multiple mechanisms, combine the production of intestinal flora neurotransmitters and short-chain fatty acids, and be more precise in its effectiveness to improve depression, anxiety, and sleep disorders. Summary of the invention

[0009] Based on the deficiencies of the prior art, the present invention aims to provide a probiotic supplement and its application that has multiple mechanisms of regulation, combines the production of neurotransmitters and short-chain fatty acids, and has more precise effects. The probiotic preparation includes Lactobacillus plantarum, Bifidobacterium breve, Bifidobacterium longum subspecies longum, Weizmannella coagulans, and Lactobacillus rhamnosus. Finally, experiments have shown that the probiotic can improve anxiety and depression symptoms, prolong total sleep time, improve sleep disorders and intestinal flora, has high safety, and is not easy to develop drug resistance.

[0010] In order to achieve the above object, the present invention adopts the following technical solutions:

[0011] In a first aspect, the present invention provides a probiotic supplement for improving depression, anxiety, and sleep disorders, characterized in that the probiotic supplement comprises mixed probiotics, and the mixed probiotics are composed of Lactobacillus plantarum, Bifidobacterium breve, Bifidobacterium longum subspecies longum, Weizmannella coagulans, and Lactobacillus rhamnosus, and the mass ratio thereof is 5-20:5-40:5-30:5-40:1-10.

[0012] Preferably, the mass ratio of Lactobacillus plantarum, Bifidobacterium breve, Bifidobacterium longum subsp. longum, Weizmannella coagulans and Lactobacillus rhamnosus is 7-20:10-40:5-30:5-20:1-5.

[0013] Preferably, the Lactobacillus plantarum accounts for 20-70% of the total number of live bacteria; the Bifidobacterium breve accounts for 5-40% of the total number of live bacteria; the Bifidobacterium longum subspecies longum accounts for 5-30% of the total number of live bacteria; the Weizmannella coagulans accounts for 5-40% of the total number of live bacteria; and the Lactobacillus rhamnosus accounts for 5-40% of the total number of live bacteria.

[0014] Among them, the total number of viable bacteria in probiotic supplements should not be less than 0.5×10 10 CFU / g.

[0015] The probiotic supplement also includes a functional additive.

[0016] Preferably, the functional additive is selected from any one or more combinations of fructooligosaccharides, galacto-oligosaccharides, inulin, pectin and xylo-oligosaccharides.

[0017] Preferably, the mass ratio of the mixed probiotics to the functional additives is 3:1-1:10.

[0018] More preferably, the mass ratio of the mixed probiotics to the functional additives is 1:5.

[0019] More preferably, the functional additive is a combination of fructooligosaccharide and galacto-oligosaccharide; the mass ratio of the two is 1:1.

[0020] Still more preferably, the functional additive is a combination of fructooligosaccharide, galacto-oligosaccharide and inulin; the mass ratio of the three is 1:1:1-1:5:5; preferably 1:2:2.

[0021] Still more preferably, the functional additive is a combination of fructooligosaccharides, galacto-oligosaccharides, inulin and pectin; the mass ratio of the four is 1:1:1:1-1:5:5:5; preferably 1:2:2:2.

[0022] Still more preferably, the functional additive is a combination of fructooligosaccharides, galacto-oligosaccharides, inulin and xylo-oligosaccharides; the mass ratio of the four is 1:1:1:1-1:5:5:5; preferably 1:2:2:2.

[0023] Still more preferably, the functional additive is a combination of fructooligosaccharides, galacto-oligosaccharides, inulin, pectin and xylo-oligosaccharides; the mass ratio of the five is 1:1:1:1:1-1:5:5:5:5, preferably 1:2:2:2:2.

[0024] In a second aspect, the present invention further provides a method for preparing the above-mentioned probiotic supplement, which is specifically: mixing Lactobacillus plantarum, Bifidobacterium breve, Bifidobacterium longum subspecies longum, Weizmannella coagulans, Lactobacillus rhamnosus and a functional additive to obtain the probiotic supplement.

[0025] In a third aspect, the present invention also provides the use of the above-mentioned probiotic supplement in the preparation of a product for improving anxiety.

[0026] The product dosage form is a solid dosage form or a liquid dosage form; the solid dosage form is a powder, tablet, capsule or granule; the liquid dosage form is a decoction or a solution.

[0027] In a fourth aspect, the present invention also provides the use of the above-mentioned probiotic supplement in the preparation of a product for improving depression.

[0028] The product dosage form is a solid dosage form or a liquid dosage form; the solid dosage form is a powder, tablet, capsule or granule; the liquid dosage form is a decoction or a solution.

[0029] In a fifth aspect, the present invention also provides the use of the above-mentioned probiotic supplement in the preparation of a product for improving sleep disorders.

[0030] The product dosage form is a solid dosage form or a liquid dosage form; the solid dosage form is a powder, tablet, capsule or granule; the liquid dosage form is a decoction or a solution.

[0031] Compared with the prior art, the present invention has the following beneficial effects:

[0032] (1) The probiotic supplements involved in the present invention are composed of a variety of probiotics in different proportions and added with corresponding functional additives. The effects of the probiotic preparations in improving depression, anxiety and sleep are systematically evaluated through animal and in vitro evaluation models, and their mechanisms are explained from the perspectives of metabolites (short-chain fatty acids) and neurotransmitter precursors (tryptophan and glutamate).

[0033] (2) This probiotic supplement combines with neurotransmitters to achieve more precise effects.

[0034] (3) The multi-pathway synergistic effects of this probiotic supplement can provide more significant and lasting effects, which is more advantageous than preparations with a single mechanism of action.

[0035] (4) The bacteria used in this probiotic supplement are all probiotics, which are highly safe and not prone to drug resistance. BRIEF DESCRIPTION OF THE DRAWINGS

[0036] Figure 1 The effect of different probiotic combinations on the production of lactic acid in the supernatant of intestinal flora culture (the control is the culture medium);

[0037] Figure 2 The effect of different probiotic combinations on the production of acetic acid in the supernatant of intestinal flora culture (the control is the culture medium);

[0038] Figure 3The effect of different probiotic combinations on the amount of glutamate produced in the supernatant of intestinal flora culture (the control was the culture medium);

[0039] Figure 4 The effect of different probiotic combinations on the amount of tryptophan produced in the supernatant of intestinal flora culture (the control was the culture medium);

[0040] Figure 5 The effect of adding different proportions of functional additives to probiotics on the amount of lactic acid produced;

[0041] Figure 6 The effect of adding different proportions of functional additives to probiotics on the amount of acetic acid produced;

[0042] Figure 7 The effect of adding different proportions of functional additives to probiotics on the amount of glutamate produced;

[0043] Figure 8 The effect of adding different proportions of functional additives to probiotics on the amount of tryptophan produced;

[0044] Fig. 9 The probiotic supplementation improves the symptoms of depression model mice;

[0045] Fig.10 Probiotic supplementation promotes sleep in mice. DETAILED DESCRIPTION

[0046] The present invention is further described in detail below with reference to specific embodiments.

[0047] The purchase information of the raw materials used in the following examples is as follows:

[0048] raw material Buy from manufacturers Accession number / CAS number Lactobacillus plantarum Zhilizhongte (Wuhan) Biotechnology Co., Ltd. CGMCC 1.16089 Bifidobacterium breve Zhilizhongte (Wuhan) Biotechnology Co., Ltd. CGMCC 1.3001 Bifidobacterium longum subsp. longum Zhilizhongte (Wuhan) Biotechnology Co., Ltd. CGMCC 1.5082 Weizmannella coagulans Zhilizhongte (Wuhan) Biotechnology Co., Ltd. CGMCC 1.10823 Lactobacillus rhamnosus Zhilizhongte (Wuhan) Biotechnology Co., Ltd. CGMCC 1.12734 Fructo-oligosaccharides Shanghai Aladdin Biochemical Technology Co., Ltd. 308066-66-2 Galacto-oligosaccharides Shanghai Aladdin Biochemical Technology Co., Ltd. 6587-31-1 Inulin Shanghai Aladdin Biochemical Technology Co., Ltd. 9005-80-5 Pectin Inner Mongolia Conster Biotechnology Co., Ltd. 9000-69-5 Xylo-oligosaccharide Shanghai Aladdin Biochemical Technology Co., Ltd. 87-99-0

[0049] Example 1 A probiotic supplement and its preparation method

[0050] Lactobacillus plantarum, Bifidobacterium breve, Bifidobacterium longum subsp. longum, Weizmannella coagulans and Lactobacillus rhamnosus were prepared in a mass ratio of 20:5:10:10:1 to prepare a 100 g product.

[0051] The preparation method comprises the following steps: uniformly mixing Lactobacillus plantarum, Bifidobacterium breve, Bifidobacterium longum subspecies longum, Weizmannella coagulans and Lactobacillus rhamnosus.

[0052] Example 2 A probiotic supplement and its preparation method

[0053] Lactobacillus plantarum, Bifidobacterium breve, Bifidobacterium longum subspecies longum, Weizmannella coagulans and Lactobacillus rhamnosus were prepared into a 100 g product in a mass ratio of 12:10:5:15:4.

[0054] Preparation method: Evenly mix Lactobacillus plantarum, Bifidobacterium breve, Bifidobacterium longum subspecies longum, Weizmannella coagulans and Lactobacillus rhamnosus to obtain the product.

[0055] Example 3 A probiotic supplement and its preparation method

[0056] Lactobacillus plantarum, Bifidobacterium breve, Bifidobacterium longum subspecies longum, Weizmannella coagulans and Lactobacillus rhamnosus were prepared into a 100 g product in a mass ratio of 5:5:5:5:1.

[0057] Preparation method: Evenly mix Lactobacillus plantarum, Bifidobacterium breve, Bifidobacterium longum subspecies longum, Weizmannella coagulans and Lactobacillus rhamnosus to obtain the product.

[0058] Example 4 A probiotic supplement and its preparation method

[0059] Lactobacillus plantarum, Bifidobacterium breve, Bifidobacterium longum subspecies longum, Weizmannella coagulans and Lactobacillus rhamnosus were prepared into a 100 g product in a mass ratio of 5:40:15:15:2.

[0060] Preparation method: Evenly mix Lactobacillus plantarum, Bifidobacterium breve, Bifidobacterium longum subspecies longum, Weizmannella coagulans and Lactobacillus rhamnosus to obtain the product.

[0061] Example 5 A probiotic supplement and its preparation method

[0062] Lactobacillus plantarum, Bifidobacterium breve, Bifidobacterium longum subspecies longum, Weizmannella coagulans and Lactobacillus rhamnosus were prepared into a 100 g product in a mass ratio of 20:40:30:40:10.

[0063] Preparation method: Evenly mix Lactobacillus plantarum, Bifidobacterium breve, Bifidobacterium longum subspecies longum, Weizmannella coagulans and Lactobacillus rhamnosus to obtain the product.

[0064] Comparative Example 1

[0065] Lactobacillus plantarum, Bifidobacterium breve, Bifidobacterium longum subspecies longum, Weizmannella coagulans and Lactobacillus rhamnosus were prepared into a 100 g product in a mass ratio of 4:4:4:4:30.

[0066] Preparation method: Evenly mix Lactobacillus plantarum, Bifidobacterium breve, Bifidobacterium longum subspecies longum, Weizmannella coagulans and Lactobacillus rhamnosus to obtain the product.

[0067] Comparative Example 2

[0068] Lactobacillus plantarum, Bifidobacterium breve, Bifidobacterium longum subspecies longum, Weizmannella coagulans and Lactobacillus rhamnosus were prepared into a 100 g product in a mass ratio of 4:5:4:50:12.

[0069] Preparation method: Evenly mix Lactobacillus plantarum, Bifidobacterium breve, Bifidobacterium longum subspecies longum, Weizmannella coagulans and Lactobacillus rhamnosus to obtain the product.

[0070] Effect verification:

[0071] 1. Effects of probiotic supplementation on lactic acid and acetic acid production by intestinal flora

[0072] The activities of the probiotic supplements prepared in Examples 1-5 and Comparative Examples 1-2 were detected by in vitro bacterial culture and mass spectrometry, with the production of lactic acid and acetic acid as evaluation indicators.

[0073] Experimental methods:

[0074] The intestinal flora of the elderly population was filtered in an anaerobic workstation, and the collected filtered suspension was diluted with a flora culture medium and inoculated into a 96-well deep-well plate. According to the total bacterial count, the probiotic supplements prepared in Examples 1-5 and Comparative Examples 1-2 were added, and a control group without probiotic supplements was set up, for a total of 8 groups. The deep-well plate was placed on a shaker, rotated at a low speed, and cultured for 24 hours. After the culture was completed, the culture supernatant was obtained after centrifugation at 14000g and 4°C for 25 minutes, and the supernatant was detected for lactic acid and acetic acid using high-resolution mass spectrometry. The test results are shown in Table 1 below.

[0075] Table 1 Effects of different probiotic combinations on lactic acid and acetic acid production

[0076] Group Lactic acid (area ratio,×10000) Acetic acid (area ratio) Control group 2448±249 0.862±0.060 Example 1 7290±92*** 1.168±0.079** Example 2 7186±106*** 1.116±0.076* Example 3 6414±264*** 1.117±0.062* Example 4 6756±106*** 1.053±0.059 Example 5 4918±275*** 1.342±0.056*** Comparative Example 1 2652±416 0.864±0.060 Comparative Example 2 1874±11 0.886±0.390

[0077] Note: Compared with the control group, *p<0.05, **p<0.01, ***p<0.001

[0078] According to the results in Table 1 above, Examples 1, 2, 3, 4 and 5 can significantly increase the lactic acid and acetic acid contents in the culture supernatant of the model group, while Comparative Examples 1 and 2 do not have the effect of significantly increasing the lactic acid and acetic acid contents in the culture supernatant of the model group ( Figure 1 , Figure 2 ).

[0079] 2. Effects of probiotic supplementation on tryptophan and glutamate production by intestinal flora

[0080] The intestinal flora of the elderly population was filtered in an anaerobic workstation, and the collected filtered suspension was diluted with a flora culture medium and inoculated into a 96-well deep-well plate. According to the total bacterial count, the probiotic supplements prepared in Examples 1-5 and Comparative Examples 1-2 were added, and a control group without probiotic supplements was set up, for a total of 8 groups. The deep-well plate was placed on a shaker, rotated at a low speed, and cultured for 24 hours. After the culture was completed, the culture supernatant was obtained after centrifugation at 14000g and 4°C for 25 minutes, and the relevant neurotransmitter precursors (glutamate and tryptophan) were detected by liquid chromatography tandem mass spectrometry (LC-MS / MS). The test results are shown in Table 2 below.

[0081] Table 2 Effects of different probiotic combinations on the production of tryptophan and glutamate

[0082]

[0083]

[0084] Note: Compared with the control group, **p<0.01, ***p<0.001

[0085] According to the results in Table 2 above, Examples 1, 2, 3, 4 and 5 can significantly increase the tryptophan and glutamic acid contents in the culture supernatant of the model group, while Comparative Examples 1 and 2 have no significant effect on increasing the above amino acid contents ( Figure 3 , Figure 4 ).

[0086] Example 6 A probiotic supplement and its preparation method

[0087] The probiotic supplement comprises mixed probiotics and functional additives.

[0088] The mixed probiotics are a mixture of Lactobacillus plantarum, Bifidobacterium breve, Bifidobacterium longum subspecies longum, Weizmannella coagulans and Lactobacillus rhamnosus in a mass ratio of 20:5:10:10:1.

[0089] The functional additives are oligofructose and oligogalactose in a mass ratio of 1:2.

[0090] The mixed probiotics and functional additives were prepared into a 100 g product in a mass ratio of 1:5.

[0091] Preparation method:

[0092] The mixed probiotics and the functional additives are mixed evenly to obtain the product.

[0093] Example 7 A probiotic supplement and its preparation method

[0094] The probiotic supplement comprises mixed probiotics and functional additives.

[0095] The mixed probiotics are a mixture of Lactobacillus plantarum, Bifidobacterium breve, Bifidobacterium longum subspecies longum, Weizmannella coagulans and Lactobacillus rhamnosus in a mass ratio of 20:5:10:10:1.

[0096] The functional additives are oligofructose, oligogalactose and inulin in a mass ratio of 1:2:2.

[0097] The mixed probiotics and functional additives were prepared into a 100 g product in a mass ratio of 1:10.

[0098] Preparation method:

[0099] The mixed probiotics and the functional additives are mixed evenly to obtain the product.

[0100] Example 8 A probiotic supplement and its preparation method

[0101] The probiotic supplement comprises mixed probiotics and functional additives.

[0102] The mixed probiotics are a mixture of Lactobacillus plantarum, Bifidobacterium breve, Bifidobacterium longum subspecies longum, Weizmannella coagulans and Lactobacillus rhamnosus in a mass ratio of 20:5:10:10:1.

[0103] The functional additives are fructooligosaccharide, galacto-oligosaccharide, inulin and xylo-oligosaccharide in a mass ratio of 1:2:2:2.

[0104] The mixed probiotics and functional additives were prepared into a 100 g product in a mass ratio of 3:1.

[0105] Preparation method:

[0106] The mixed probiotics and the functional additives are mixed evenly to obtain the product.

[0107] Example 9 A probiotic supplement and its preparation method

[0108] The probiotic supplement comprises mixed probiotics and functional additives.

[0109] The mixed probiotics are a mixture of Lactobacillus plantarum, Bifidobacterium breve, Bifidobacterium longum subspecies longum, Weizmannella coagulans and Lactobacillus rhamnosus in a mass ratio of 20:5:10:10:1.

[0110] The functional additives are fructooligosaccharide, galacto-oligosaccharide, inulin and pectin in a mass ratio of 1:2:2:2.

[0111] The mixed probiotics and functional additives were prepared into a 100 g product in a mass ratio of 1:1.

[0112] Preparation method:

[0113] The mixed probiotics and the functional additives are mixed evenly to obtain the product.

[0114] Example 10 A probiotic supplement and its preparation method

[0115] The probiotic supplement comprises mixed probiotics and functional additives.

[0116] The mixed probiotics are a mixture of Lactobacillus plantarum, Bifidobacterium breve, Bifidobacterium longum subspecies longum, Weizmannella coagulans and Lactobacillus rhamnosus in a mass ratio of 20:5:10:10:1.

[0117] The functional additives are fructooligosaccharide, galacto-oligosaccharide, inulin, pectin and xylo-oligosaccharide in a mass ratio of 1:2:2:2:2.

[0118] The mixed probiotics and functional additives were prepared into a 100 g product in a mass ratio of 1:5.

[0119] Preparation method:

[0120] The mixed probiotics and the functional additives are mixed evenly to obtain the product.

[0121] Effect detection:

[0122] 3. Effects of probiotic supplementation on lactic acid and acetic acid production by intestinal flora

[0123] The activities of the probiotic supplements prepared in Examples 6-10 were detected by in vitro bacterial culture and mass spectrometry, with the production of lactic acid and acetic acid as evaluation indicators.

[0124] Experimental methods:

[0125] The intestinal flora of the elderly population was filtered in an anaerobic workstation, and the collected filtered suspension was diluted with a flora culture medium and inoculated into a 96-well deep-well plate. According to the total bacterial count, the probiotic supplements prepared in Examples 6-10 were added, and a control group without probiotic supplements was set up. The deep-well plate was placed on a shaker, rotated at a low speed, and cultured for 24 hours. After the culture was completed, the culture supernatant was obtained after centrifugation at 14000g and 4°C for 25 minutes, and the supernatant was detected for lactic acid and acetic acid using high-resolution mass spectrometry. The test results are shown in Table 3 below.

[0126] Table 3 Effect of adding different proportions of functional additives to probiotics on the production of lactic acid and acetic acid

[0127]

[0128]

[0129] Note: Compared with the control group, ***p<0.001

[0130] According to the results in Table 3 above, in Examples 6 to 10, the addition of different combinations of functional additives to probiotics can significantly increase the content of lactic acid and acetic acid in the supernatant of intestinal flora culture ( Figure 5 , Figure 6 )

[0131] 4. Effects of probiotic supplementation on tryptophan and glutamate production by intestinal flora

[0132] The intestinal flora of the elderly population was filtered in an anaerobic workstation, and the collected filtered suspension was diluted with a flora culture medium and inoculated into a 96-well deep-well plate. According to the total bacterial count, the probiotic supplement prepared in Examples 6-10 was added, and a control group without probiotic supplement was set up. The deep-well plate was placed on a shaker, rotated at a low speed, and cultured for 24 hours. After the culture was completed, the culture supernatant was obtained after centrifugation at 14000g and 4°C for 25 minutes, and the relevant neurotransmitter precursors (glutamate and tryptophan) were detected by liquid chromatography tandem mass spectrometry (LC-MS / MS). The test results are shown in Table 4 below.

[0133] Table 4 Effect of adding different proportions of functional additives to probiotics on the production of tryptophan and glutamate

[0134] Group Glutamate (area ratio, ×10000) Tryptophan (area ratio, ×10000) Control group 447±126 776±119 Example 6 2616±37*** 1628±107*** Example 7 2306±162*** 3396±37*** Example 8 1274±51*** 3238±16*** Example 9 2634±34*** 4292±241*** Example 10 2318±26*** 4850±100***

[0135] Note: Compared with the control group, ***p<0.001

[0136] According to the results in Table 4 above, in Examples 6 to 10, the addition of different combinations of functional additives to probiotics can significantly increase the content of tryptophan and glutamic acid in the intestinal flora culture supernatant ( Figure 7 , Figure 8 ).

[0137] 5. Effect of probiotic supplementation on improving symptoms of depression and anxiety model mice

[0138] Experimental methods:

[0139] Male C57BL / 6 mice aged 6-8 weeks were used and randomly divided into a model group and a probiotic supplement group according to their body weight. The probiotic supplement prepared in Example 6-8 was administered to each mouse, with a total bacterial count of 10 9 After oral administration of CFU of probiotic supplements for 10 consecutive days, the anxiety and depression model was established by forced swimming to investigate the improvement effect of probiotic supplements. The test results are shown in Table 5 below.

[0140] Table 5 Effect of probiotic supplementation on improving symptoms of depression and anxiety model mice

[0141] Group Immobility time(s) Model Group 116±13 Example 6 40±9*** Example 7 31±6*** Example 8 29±7***

[0142] Note: Compared with the model group, ***p<0.001

[0143] According to the results in Table 5 above, the probiotic supplements of Examples 6-8 can significantly reduce the immobility time of anxiety and depression model mice and improve the symptoms of anxiety and depression ( Fig. 9 ).

[0144] 6. Probiotic supplementation promotes sleep in mice

[0145] Experimental methods:

[0146] C57BL / 6 mice, 6-8 weeks old, female. The mice were kept in an environment with a temperature of 19-26°C, 12 hours (06:00-18:00) of fluorescent light and 12 hours of no light per day. The mice were randomly divided into two groups, a model group and a probiotic group. The probiotic supplement prepared in Example 6-7 was administered to each mouse, with a total bacterial count of 10 9 CFU of probiotic supplements. Probiotics were given continuously for 15 days. Three days before the test, each mouse was intraperitoneally injected with 300 mg / kg of PCPA, a 5-hydroxytryptamine (5-HT) synthesis inhibitor, once a day. The mice were transferred to the test environment and adapted for 30 minutes. The PiezoSleep non-invasive sleep detection system was used to test the sleep duration of mice within 24 hours. During the test, the mice were free to eat and drink. The percentage of total sleep time in 24 hours was used as the evaluation parameter, and the test results are shown in Table 6 below.

[0147] Table 6 Effects of probiotic supplementation on sleep in mice

[0148] Group Sleep time percentage (24 hours) Control group 63.25±2.93 Model Group 60.75±2.75 Example 6 66.60±1.63 Example 7 66.00±1.22

[0149] According to the results in Table 6 above, the probiotic supplements in Example 6 and Example 7 can both prolong the sleep time of the model mice within 24 hours, indicating that the probiotic supplement has a significant improvement effect on sleep ( Fig.10 ).

[0150] The above embodiments are only used to illustrate the technical solutions of the present invention, rather than to limit the same. Although the present invention has been described in detail with reference to the aforementioned embodiments, those skilled in the art should understand that the technical solutions described in the aforementioned embodiments may still be modified, or some of the technical features may be replaced by equivalents. However, these modifications or replacements do not deviate the essence of the corresponding technical solutions from the spirit and scope of the technical solutions of the embodiments of the present invention.

Claims

1. A probiotic supplement for improving depression, anxiety, and sleep disorders, characterized in that: The probiotic supplement comprises mixed probiotics, which are composed of lactobacillus plantarum, bifidobacterium breve, bifidobacterium longum subspecies longum, Weizmannella coagulans and lactobacillus rhamnosus, and the mass ratio thereof is 5-20:5-40:5-30:5-40:1-10.

2. The probiotic supplement according to claim 1, characterized in that The plant lactobacillus accounts for 20-70% of the total number of live bacteria; the breve bifidobacterium accounts for 5-40% of the total number of live bacteria; the long bifidobacterium subspecies long accounts for 5-30% of the total number of live bacteria; the coagulant Weizmannella accounts for 5-40% of the total number of live bacteria; the number of rhamnosus lactobacillus accounts for 5-40% of the total number of live bacteria; the total number of live bacteria in the probiotic supplement is not less than 0.5×10 10 CFU / g.

3. The probiotic supplement according to claim 1, characterized in that The probiotic supplement also includes a functional additive; the functional additive is selected from any one or more combinations of oligofructose, oligogalactose, inulin, pectin and oligoxylose; the mass ratio of the mixed probiotics to the functional additive is 3:1-1:

10.

4. The method for preparing the probiotic supplement according to any one of claims 1 to 3, characterized in that: The preparation method is obtained by mixing Lactobacillus plantarum, Bifidobacterium breve, Bifidobacterium longum subspecies longum, Weizmannella coagulans, Lactobacillus rhamnosus and functional additives.

5. Use of the probiotic supplement according to any one of claims 1 to 3 in the preparation of a product for improving sleep disorders.

6. Use of the probiotic supplement according to any one of claims 1 to 3 in the preparation of a product for improving anxiety.

7. Use of the probiotic supplement according to any one of claims 1 to 3 in the preparation of a product for improving depression.

8. A product for improving sleep disorders, characterized in that: The probiotic supplement comprises any one of claims 1 to 3, wherein the product dosage form is a solid dosage form or a liquid dosage form; the solid dosage form is a powder, tablet, capsule or granule; and the liquid dosage form is a decoction or a solution.

9. A product for improving anxiety, characterized in that: The probiotic supplement comprises any one of claims 1 to 3, wherein the product dosage form is a solid dosage form or a liquid dosage form; the solid dosage form is a powder, tablet, capsule or granule; and the liquid dosage form is a decoction or a solution.

10. A product for improving depression, characterized in that: The probiotic supplement comprises any one of claims 1 to 3, wherein the product dosage form is a solid dosage form or a liquid dosage form; the solid dosage form is a powder, tablet, capsule or granule; and the liquid dosage form is a decoction or a solution.

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