Bifidobacterium longum subsp. Infantis YLGB-1496 strain and application thereof in reducing cortisol content and / or improving appetite

Abstract

The invention provides a bifidobacterium longum subsp. Infantis YLGB-1496 strain and application of the bifidobacterium longum subsp. Infantis YLGB-1496 strain in reducing cortisol content and / or improving appetite.

Description

Technical Field

[0001] The invention relates to the technical field of microorganisms, and in particular to a Bifidobacterium longum subspecies infantis YLGB-1496 strain and an application thereof in reducing cortisol content and / or improving appetite. Background Art

[0002] Cortisol is a stress hormone that is secreted by the adrenal glands in response to stressful conditions. It plays a key role in regulating the body’s stress response, metabolic function, and immune system. Although cortisol helps cope with stress in the short term, chronically high cortisol levels have been linked to a variety of health problems, including anxiety, depression, obesity, high blood pressure, and decreased immune function. Additionally, changes in cortisol levels are closely linked to fluctuations in appetite, and cortisol also plays an important role in regulating appetite. Therefore, regulating cortisol levels is essential for maintaining physical and mental health.

[0003] In recent years, probiotics have received widespread attention for their positive effects on gut health. However, there are currently limited probiotic products on the market for lowering cortisol.

[0004] Therefore, developing a probiotic product that can effectively reduce cortisol levels would help meet this growing health need. Summary of the invention

[0005] The present invention provides a Bifidobacterium longum subspecies infantis YLGB-1496 strain and an application thereof in reducing cortisol content and / or improving appetite, and has a good effect in reducing cortisol content and / or improving appetite.

[0006] The present invention provides use of Bifidobacterium longum subspecies infantis YLGB-1496 strain in preparing a product for reducing cortisol content and / or improving appetite.

[0007] Optionally, the Bifidobacterium longum subsp. infantis YLGB-1496 strain is deposited in the China Type Culture Collection of Wuhan University with a deposit number of CCTCC NO: M2011122.

[0008] Optionally, the product comprises the Bifidobacterium longum subsp. infantis YLGB-1496 strain.

[0009] Optionally, in the product, the viable count of the Bifidobacterium longum subspecies infantis YLGB-1496 strain is 10 6 CFU / g~10 11 CFU / g.

[0010] Optionally, the product is administered to the subject via the gastrointestinal route.

[0011] Optionally, the subjects include infants aged 0 to 3 years old.

[0012] Optionally, the product includes one or more of ordinary food, health food, solid beverage, nutritional supplement, infant formula, special medical purpose formula food and medicine.

[0013] The present invention provides a composition for reducing the content of cortisol and / or improving appetite, wherein the composition comprises Bifidobacterium longum subspecies infantis YLGB-1496 strain.

[0014] Optionally, the composition further comprises an excipient, wherein the excipient comprises maltodextrin.

[0015] The present invention provides a method for preparing the composition as described above, comprising: mixing a raw material system including the Bifidobacterium longum subspecies infantis YLGB-1496 strain to obtain the composition.

[0016] The present invention provides a Bifidobacterium longum subspecies infantis YLGB-1496 strain and an application thereof in reducing cortisol content and / or improving appetite. The YLGB-1496 strain is deposited in the China Type Culture Collection of Wuhan University with a deposit number of CCTCC NO: M2011122. Experiments have confirmed that the YLGB-1496 strain has a good effect in reducing cortisol content and / or improving appetite, and also has a good effect in improving body weight. BRIEF DESCRIPTION OF THE DRAWINGS

[0017] Figure 1 Specific technology roadmap for clinical trials;

[0018] Figure 2 A bar graph showing the number of days in Example 1 in which the subjects reported that they had no appetite in the past month;

[0019] Figure 3 This is a statistical chart of the cumulative number of days of decreased appetite for the subjects in Example 2 over a period of 3 months (N is the number of samples, and the rank mean is the average of the cumulative number of days of decreased appetite);

[0020] Figure 4 The bar graph is a graph of the cortisol content in the saliva of the subjects in Example 1 (Probiotic is the probiotic intervention group, and Placebo is the placebo group). DETAILED DESCRIPTION

[0021] In order to enable those skilled in the art to better understand the scheme of the present invention, the present invention is further described in detail below. The specific implementation methods listed below are only for describing the principles and features of the present invention. The examples are only used to explain the present invention and are not intended to limit the scope of the present invention. Based on the embodiments of the present invention, all other implementation methods obtained by ordinary technicians in the field without creative work are within the scope of protection of the present invention.

[0022] Intestinal microecology refers to the complex ecosystem formed by the interaction between microorganisms that parasitize the human intestine and the human body.

[0023] The formation of intestinal microecology is a complex process. The initial microorganisms in the newborn's body mainly come from the mother's birth canal, feces and the surrounding environment. 0-3 years old is the key time window for the establishment and evolution of intestinal flora, and breastfeeding is the most important factor in determining the early intestinal microecology of infants. Full-term, vaginally delivered newborns who are breastfed are more likely to form healthy intestinal flora, such as intestinal flora with more appropriate content and types of bifidobacteria and lactobacilli.

[0024] Breast milk contains probiotics and prebiotics such as human milk oligosaccharides (HMOs). In 2003, the literature first reported that breast milk contains bacteria and directly affects the intestinal flora of infants; many subsequent studies have provided definitive evidence that the intestinal microorganisms of breastfed infants come from breast milk; there are now many studies on probiotics in breast milk, including the discovery, isolation and research of probiotics in breast milk, and patents have been applied for them. For example, the University of Granada in Spain has applied for global and even Chinese patents, and it discovered three probiotic strains in breast milk (two strains of Lactobacillus and one strain of Bifidobacterium Lactobacillus rhamnosus HERO 22A (CNCM I-4036), Lactobacillusparacasei HERO7 (CNCM I-4034) and Bifidobacterium breve HERO 15B (CNCM I-4035)).

[0025] The list of bacteria that can be added to food in many countries includes Bifidobacterium, Lactobacillus, Streptococcus, etc., and is specified to specific genera. The "List of Probiotics that Can Be Used in Health Foods" and the "List of Bacteria that Can Be Used in Foods" specify edible probiotics to bacteria. The "List of Bacteria that Can Be Used in Infant Foods" released in 2011 refines each probiotic that can be used in food to strains. Lactobacillus and Bifidobacterium have been recognized as probiotics.

[0026] Currently, there is only one strain derived from human breast milk in the list of bacteria that can be used in infant food, which is Lactobacillus mucosa CECT5716, which originated in Spain. The rest of the strains are not derived from breast milk. In addition, in the latest "List of Bacteria that Can Be Used in Food" in August 2022, Bifidobacterium longum infantis is included, and the European Food Safety Authority Qualification Permit (QPS) list also includes Bifidobacterium longum infantis, indicating that this strain is safe for consumption. In recent years, the importance of Bifidobacterium longum infantis (Bifidobacterium infantis) to the intestines of infants and young children has been confirmed by many studies.

[0027] Since breast milk contains microorganisms and is very important for the establishment of intestinal flora in early life, especially Bifidobacterium longum infantis subsp. infantis in breast milk is of great significance to the health of infants and young children. Therefore, Bifidobacterium longum infantis subsp. infantis in breast milk has become a research hotspot.

[0028] Based on this, an embodiment of the present invention provides a use of Bifidobacterium longum subspecies infantis YLGB-1496 strain in the preparation of a product that reduces cortisol content and / or improves appetite.

[0029] The above-mentioned Bifidobacterium longum subsp. Infantis GB-1496 strain was deposited in the China Type Culture Collection of Wuhan University on April 10, 2011, with the deposit number of CCTCC NO: M2011122.

[0030] The Bifidobacterium longum subspecies infantis (B. infantis) YLGB-1496 strain was cultured and isolated from the milk of a mother who was exclusively breastfeeding. Its gene sequencing results were compared with the published gene sequences of Bifidobacterium infantis, and it was determined to be a new strain of Bifidobacterium infantis.

[0031] The experiment proved that the long Bifidobacterium infantis subspecies YLGB-1496 strain has a good effect in reducing cortisol content and / or improving appetite, and also has a good effect on improving body weight. Therefore, the long Bifidobacterium infantis subspecies YLGB-1496 strain can be used to prepare products that reduce cortisol content and / or improve appetite.

[0032] In some embodiments, the above products include Bifidobacterium longum subsp. infantis YLGB-1496 strain.

[0033] Specifically, in the above product, the viable count of Bifidobacterium longum subspecies infantis YLGB-1496 strain may be greater than or equal to 10 6 CFU / g.

[0034] Furthermore, in the above product, the viable count of the Bifidobacterium longum subspecies infantis YLGB-1496 strain can be 10 6 CFU / g~10 11 CFU / g.

[0035] In some embodiments, the above-mentioned Bifidobacterium longum subsp. infantis YLGB-1496 strain is administered to the subject via the gastrointestinal tract.

[0036] At present, many studies have confirmed that the intestinal microecology in early life plays a vital role in the maturation of the immune system, and the diversity and maturity of the intestinal flora in infancy are closely related to health. Existing research data have confirmed that the above-mentioned Bifidobacterium longum infantis subspecies YLGB-1496 strain is resistant to the strong acidity and bile salts of the artificial gastrointestinal tract, which means that it can pass through the stomach and upper small intestine of the human digestive tract, and eventually reach the lower small intestine and colon to exert its activity. For infants and young children (such as infants and young children aged 0 to 3 years), because the acidity of their gastric acid and the concentration of bile salts are lower than those of adults, the above-mentioned Bifidobacterium longum infantis subspecies YLGB-1496 strain as a probiotic is easier to colonize in the intestines of infants and young children at an early stage.

[0037] In a specific embodiment, the above-mentioned subjects include infants aged 0 to 3 years (or 0 to 36 months). Specifically, infants aged 0 to 3 years may include infants aged 0 to 3 years and infants aged 1 to 3 years.

[0038] Clinical trial data confirms that the above-mentioned Bifidobacterium longum infantis subspecies YLGB-1496 strain can effectively reduce the cortisol content in the saliva of infants and young children aged 0 to 3 years old, improve appetite and stomach, and also has a good effect on improving body weight.

[0039] It is understandable that the embodiments of the present invention do not limit the types of the above-mentioned products. For example, the above-mentioned products may include one or more of ordinary food, health food, solid beverages, nutritional supplements, infant formula, special medical purpose formula and medicines.

[0040] The present invention also provides a composition for reducing cortisol content and / or improving appetite, the composition comprising the above-mentioned Bifidobacterium longum subspecies infantis YLGB-1496 strain. Based on the above-mentioned Bifidobacterium longum subspecies infantis YLGB-1496 strain, the composition can effectively reduce cortisol content and / or improve appetite.

[0041] Additionally, the composition may include excipients such as maltodextrin.

[0042] Furthermore, in the above composition, the viable count of the Bifidobacterium longum subspecies infantis YLGB-1496 strain may be greater than or equal to 10 6 CFU / g.

[0043] Furthermore, in the above composition, the viable count of Bifidobacterium longum subspecies infantis YLGB-1496 strain can be 10 6 CFU / g~10 11 CFU / g.

[0044] The embodiment of the present invention further provides a method for preparing the above composition, comprising: mixing a raw material system including Bifidobacterium longum subspecies infantis YLGB-1496 strain to obtain the above composition.

[0045] In a specific implementation, the bacterial powder of Bifidobacterium longum subspecies infantis YLGB-1496 strain can be prepared first, and then the raw material system including the bacterial powder is mixed evenly to obtain the above-mentioned composition.

[0046] The bacterial powder of the above-mentioned Bifidobacterium longum infantis subspecies YLGB-1496 strain must meet the requirement of viable lactic acid bacteria count ≥ 1.5×10 11 CFU / g, and a water activity of 0.05-0.1, such as 0.05, 0.07, 0.1 or a range consisting of any two thereof.

[0047] In addition, the above raw material system may further include an excipient, such as maltodextrin.

[0048] The present invention is further described below by specific examples and comparative examples. Unless otherwise specified, the reagents, materials and instruments used below are all conventional reagents, conventional materials and conventional instruments, which can be commercially available, and the reagents and materials involved can also be synthesized by conventional synthesis methods.

[0049] Through clinical trials on infants aged 0-3 years, specifically targeting the long Bifidobacterium infantis subspecies YLGB-1496 strain, the clinical manifestations of supplementation and intervention in infants aged 0-3 years were explored, and the effects of the YLGB-1496 strain on the saliva cortisol content and appetite of infants aged 0-3 years (early life population) were studied. The method used in the trial was a multi-center randomized double-blind controlled study. The specific technical route is as follows: Figure 1 shown.

[0050] Example 1

[0051] This study has been registered on ClinicalTrials.gov (identifier number NCT05794815). The inclusion and exclusion criteria of the subjects are as follows:

[0052] 1) Children aged 1 to 3 years old were recruited as subjects. Before the study began, all parents of the subjects signed written informed consent. The inclusion and exclusion criteria of the subjects are as follows:

[0053] Inclusion criteria included: current weight was P20-P80 on the children's percentile table, currently using infant formula without probiotics, not allowing the use of prebiotics or probiotics, and willing to participate in the entire experiment;

[0054] Exclusion criteria included any malformation, maternal metabolic and / or chronic disease, current or previous medical conditions that could interfere with the study conditions or interventions, oral antibiotic use within 2 weeks before the intervention, known probiotic allergy or possible food allergy, participation in other clinical studies within 4 weeks before the intervention, or unwillingness to participate for any specific reason.

[0055] 2) Preparation of probiotic products and placebo

[0056] The intervention substance used in this study was Bifidobacterium longum infantis subspecies YLGB-1496 powder, and the excipient was maltodextrin, with a specification of 2g / bag; the placebo was maltodextrin, 2g / bag, and its appearance, packaging, weight, and color were consistent with the intervention substance. The intervention substance and placebo were commissioned to SGS Standards Technical Services (Shanghai) Co., Ltd. to conduct tests on Escherichia coli, Salmonella, Staphylococcus aureus, and mold, and the results were in line with national standards. In addition, heavy metals and additives such as lead (Pb), benzoic acid, sorbic acid, sodium saccharin cyclohexylaminosulfonate (cyclamate), and tartrazine were tested, and the results were not detected.

[0057] The intervention material (bacterial powder) was stored in a -20 degrees Celsius refrigerator; after the intervention material was distributed to the subjects (research subjects), it was stored in a home refrigerator at -18 degrees Celsius.

[0058] 3) Research implementation

[0059] After enrollment, subjects were randomized according to the inclusion and exclusion criteria. Eligible subjects were randomly assigned to the two groups of the study (probiotic group or placebo group) in a 1:1 ratio according to a computer-generated list and identified by a separate code. Randomization was performed by the study statistician, who had no contact with the participants. No member of the research team had access to the allocation sequence until the study was completed.

[0060] The researchers distributed the intervention items to all included infants and young children according to the group information.

[0061] Probiotic intervention group (intervention group or control group): once a day, 1 pack (bag) each time (containing 1.5×10 YLGB-1496 live bacteria 10 CFU / pack), starting from the day of using the probiotic product and continuing for 3 months.

[0062] Placebo control group (placebo group): 1 pack (maltodextrin, does not contain Bifidobacterium infantis YLGB-1496 strain) once a day, starting from the day of using the placebo product and continuing for 3 months.

[0063] The two groups of infants and their main feeders were interviewed at three follow-up points (0w, 6w, and 12w) to collect information such as diet and health records, as well as saliva samples. The cortisol content in the saliva, appetite, and weight improvement of the subjects at the three follow-up points were recorded and tested truthfully.

[0064] 4) Data entry and statistical analysis

[0065] The research data was entered using Epidata software by two people on two computers, and then the database was compared twice. If any problems were found during the process, the monitor was notified in time and the researcher was required to provide answers. A question sheet was used to record various questions and answers during the data entry process, and the question sheet was saved for future reference.

[0066] The data were analyzed using SPSS software version 20.0. Given the skewed distribution and nonparametric nature of the data, the Mann-Whitney U test was used to compare differences between groups, while the chi-square test was used to compare routine data. Spearman rank correlation was used in nonparametric correlation analysis, and the correlation coefficient was rho (r). Negative binomial regression was also used to evaluate the effects of probiotic intervention on cortisol content, appetite, and weight. All tests were two-sided, and *P<0.05 was considered statistically significant. Unless otherwise stated, data are presented as mean ± standard error.

[0067] Example 2

[0068] 1) Healthy infants aged 0-3 years were recruited as subjects. All parents of the subjects were informed and signed informed consent. This study has been registered in the ISRCTN registry (registration number ISRCTN12824613). The inclusion and exclusion criteria of the subjects are as follows:

[0069] Inclusion criteria include:

[0070] Infants aged 0-3 years born at 37-42 weeks of gestation, with a birth weight of 2500 g or more and less than 4000 g, regardless of gender;

[0071] The mother of the infant or young child has no diagnosed metabolic diseases such as diabetes, and no infectious diseases such as hepatitis B and HIV;

[0072] The parents or primary guardians of the infants and young children agreed to collect stool samples from the infants and young children during this study;

[0073] No allergic diseases diagnosed by clinical physicians at the time of enrollment (including but not limited to eczema, asthma, allergic proctitis, allergic rhinitis, hay fever, etc.);

[0074] Family members or primary guardians promised not to add additional probiotic products (including formula milk containing probiotics) to infants and young children during the intervention period;

[0075] The guardians of the enrolled subjects agreed to participate in this intervention study and signed a written informed consent form, and were able to understand and fill in infant diary and other forms as required.

[0076] Exclusion criteria include:

[0077] Infants and young children included due to missed diagnosis;

[0078] Violation of inclusion criteria was discovered during the trial;

[0079] Infants and young children who do not have any test records available for evaluation;

[0080] During the trial, the patient took other probiotics not in the study, so the effect of the probiotics used in the study cannot be evaluated;

[0081] Subjects with poor compliance, taking less than 80% of the full dose of probiotics;

[0082] If a subject experiences an AE, laboratory abnormality, or other medical condition related to the probiotic Bifidobacterium infantis YLGB-1496, it would not be in the best interest of the subject to continue participating in the study;

[0083] Withdrawal of informed consent during the intervention period;

[0084] Subjects who have obtained informed consent and are eligible for randomized trials but fail to complete the course of treatment, observation period, and data collection specified in this protocol for some reason will be treated as dropouts;

[0085] Infants and young children who need to use antibiotics or immunosuppressive drugs due to illness during the intervention period;

[0086] Infants and young children who developed allergies to the probiotic strains and / or maltodextrin components used in this study during the intervention period.

[0087] 2) The probiotic product and placebo were prepared, the study was implemented, data were entered and statistically analyzed according to Example 1, and the number of days of decreased appetite and weight improvement of the above infants within 3 months were statistically analyzed.

[0088] Test results

[0089] Table 1 Number of days in Example 1 when the subjects reported no appetite in the past month (days / week)

[0090]

[0091] Combined with Table 1, Figure 2 , Figure 3 It can be seen that the number of days without appetite in the probiotic intervention group (intervention group) was significantly reduced, which can confirm that Bifidobacterium longum infantis subspecies YLGB-1496 can effectively improve the appetite of the subjects.

[0092] Table 2 Cortisol content in saliva of subjects in Example 1 (unit: ng / µg)

[0093]

[0094] Combined with Table 2, Figure 4 It can be seen that the salivary cortisol content in the probiotic intervention group (intervention group) was significantly reduced, which can confirm that Bifidobacterium longum infantis subspecies YLGB-1496 can effectively reduce the salivary cortisol content of the subjects.

[0095] Table 3 Body weight of subjects in Example 1 (unit: kg)

[0096]

[0097] Table 4 Body weight of subjects in Example 2 (unit: kg)

[0098]

[0099] Combining Tables 3 and 4, it can be seen that the weight gain of the probiotic intervention group (intervention group) was significant, which can confirm that Bifidobacterium longum infantis subspecies YLGB-1496 can effectively improve the weight of the subjects.

[0100] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, rather than to limit it. Although the present invention has been described in detail with reference to the aforementioned embodiments, those skilled in the art should understand that they can still modify the technical solutions described in the aforementioned embodiments, or replace some or all of the technical features therein with equivalents. However, these modifications or replacements do not cause the essence of the corresponding technical solutions to deviate from the scope of the technical solutions of the embodiments of the present invention.

Claims

1. Use of Bifidobacterium longum subspecies infantis YLGB-1496 strain in the preparation of products for reducing cortisol content and / or improving appetite.

2. The use according to claim 1, characterized in that The Bifidobacterium longum subsp. infantis YLGB-1496 strain is deposited in the China Center for Type Culture Collection of Wuhan University with a deposit number of CCTCC NO: M2011122.

3. The use according to claim 1 or 2, characterized in that The product comprises the Bifidobacterium longum subspecies infantis YLGB-1496 strain.

4. The use according to claim 3, characterized in that In the product, the viable count of the Bifidobacterium longum subspecies infantis YLGB-1496 strain is 10 6 CFU / g~10 11 CFU / g.

5. The use according to claim 1, characterized in that: The product is administered to the subject via the gastrointestinal route.

6. The use according to claim 5, characterized in that The subjects include infants and young children aged 0 to 3 years old.

7. The use according to claim 1, characterized in that The products include one or more of ordinary food, health food, solid beverages, nutritional supplements, infant formula food, special medical purpose formula food and medicines.

8. A composition for reducing cortisol content and / or improving appetite, characterized in that: The composition includes Bifidobacterium longum subsp. infantis YLGB-1496 strain.

9. The composition according to claim 8, characterized in that The composition further comprises an excipient, wherein The excipients include maltodextrin.

10. A method for preparing the composition according to claim 8 or 9, characterized in that: include: The raw material system including the Bifidobacterium longum subspecies infantis YLGB-1496 strain is mixed to obtain the composition.

Images