KR20200058849A - Composition for reduction or treatment of atopic symptom comprising high immune active Lactobacillus sakei probio-65 ghost cell or Lactobacillus sakei probio-65 extract - Google Patents

Abstract

The present invention provides a composition for reduction or treatment of atopic dermatitis symptoms, which contains Lactobacillus sakei probio-65 dead cells or a Lactobacillus sakei probio-65 extract having high immune activity. The present invention has an effect of providing the composition for reduction or treatment of atopic dermatitis symptoms, which contains Lactobacillus sakei probio-65 dead cells or the Lactobacillus sakei probio-65 extract, which can be used to suppress and treat atopic dermatitis symptoms in children and adolescents suffering from atopic dermatitis symptoms.

Description

Composition for reduction or treatment of atopic dermatitis comprising high immune active Lactobacillus sakei probio- 65 ghost cell or Lactobacillus sakei probio-65 extract}

The present invention relates to a composition for reducing or treating symptoms of atopic dermatitis, which includes the extract of Lactobacillus sakei probio-65 having high immunological activity or Lactobacillus sakei probio-65, and more particularly, having atopic dermatitis. Statistical scores of SCORAD total score, SCORAD objective score, score of SCORAD objective, and dry score of SCORAD objective, which are indicators of atopic dermatitis in infants and adolescents Lactobacillus sake probio-65 or Lactobacillus sake, which has a high immunological activity that can be used for reducing and treating atopic dermatitis symptoms by enabling a significant decrease in science and a statistically significant increase in skin oil. It relates to a composition for reducing or treating atopic dermatitis symptoms comprising a probio-65 extract.

Probiotics lactic acid bacteria refers to lactic acid bacteria having health promotion, disease or disease prevention and treatment effect. Probiotic lactic acid bacteria have been widely used in Europe and other countries for a long time as an adjuvant for health promotion, and clinical trials in animals and humans have shown that they have excellent effects such as anti-cancer effect, immune-enhancing effect, and anti-allergy. Is being revealed through.

Atopic dermatitis (Atopic symptom) is a disease that occurs with a prevalence of 1 to 3% in adults, and a prevalence of around 10% in children. It is a chronic inflammatory skin disease that occurs mainly in infancy and childhood and is accompanied by severe itching. . In some patients with atopic dermatitis, asthma and rhinitis have enlarged pathology, and physical and mental pain is caused not only in young patients who develop due to persistent itching caused by dermatitis, but also in families including carers. Lowers. There is currently no adequate blood test to assess the severity of atopic dermatitis, and the most widely used to assess the severity of atopic dermatitis is the SCORing Atopic Dermatitis (SCORAD) index. The SCORAD index was created by the European Task Force on AD, which consists of more than 20 dermatologists, and its feasibility, reliability, and sensitivity were addressed and confirmed in several studies.

Itching, the most prominent symptom in patients with atopic dermatitis, is said to start from an allergic inflammatory reaction. Irritants, such as allergens outside the skin, are the source of itching. When this irritant penetrates the damaged skin barrier of atopic dermatitis patients and reaches the skin, an allergic inflammatory reaction begins in the skin. When the inflammatory response begins on the skin, several mediators, including histamine, cause itching of the skin in the inflamed area. Patients who have started itching will scratch the inflamed skin area, causing irritated skin tissue damage due to skin irritation caused by scratching and causing more severe skin irritation compared to scratching.

In terms of the mechanism of action of itching on the skin, atopic dermatitis has a different mechanism of action from itching caused by hives. In the case of itching caused by urticaria, it is mainly a result of histamine secretion, but in atopic dermatitis, various mediators are involved, and the clinical therapeutic effect of antihistamines is limited. Itching of atopic dermatitis is not a symptom as a result of the disease, but an important factor that plays an important role in pathophysiology. Once it occurs, itching and scratching accompanying it are constantly amplified.

Currently, there are no adequate treatment methods to reliably treat atopic dermatitis, and people with atopic dermatitis symptoms use self-care measures as moisturizers, avoid allergen and exacerbation factors, use of medications for external use, bath therapy, dietary care Taking care of it. On the other hand, the main drugs used as a drug treatment method for patients with severe and mild atopic dermatitis include antihistamines, topical steroid external preparations, topical immunomodulatory external preparations, and oral steroids. In severe patients, steroid injections are also used.

As a way to cope with atopic dermatitis, the effect of probiotic lactic acid bacteria has been recently reported. Some lactic acid bacteria have a significant therapeutic effect on atopic dermatitis, have a preventive effect on the development of atopic dermatitis, and have an effect of fundamentally blocking the occurrence of food allergies. Probiotics, which have been shown to be effective against atopic dermatitis, include Lactobacillus rhamnosus, Bifidobacterium lactis, Lactobacillus lutheri, and others. Probiotic lactic acid bacteria, which have been used for over 100 years, have been proven to have excellent health promoting effects such as immune-enhancing and anti-cancer effects through steady research and clinical trials, and have no harm to pregnant women or infants. The difference between the probiotic lactic acid bacteria and the general lactic acid bacteria drink is a material created by selecting only certain types of bacteria that are effective in treating and preventing atopic and allergic and food allergies. In addition, it is made in a quantity large enough to be incomparable to the number of bacteria contained in the general lactic acid bacteria drink. In order to see the true effect, a sufficient number of bacteria should be used by selecting a species that has proven to be effective in atopy.

Lactobacillus sakei subsp. Sakei is a lactic acid bacterium commonly used in fermentation of sausages and meats in livestock countries in Europe, and kimchi, a fermented food traditionally secured by Koreans for hundreds of years It plays an important role in the fermentation process, and a large amount is distributed in the kimchi being aged. Lactobacillus sakei probio-65 (Lactobacillus sakei probio-65) is a strain isolated from kimchi, exhibits acid resistance, bile resistance, and antibiotic resistance, inhibits the growth of harmful pathogenic microorganisms in the body and intestines of animals, and enhances immunity Staphylococcus aureus, which has activity and is particularly known as a disease exacerbation factor for atopic dermatitis lesions, has a strong growth inhibitory effect, and is effective in the treatment or prevention of atopic dermatitis. It is also effective, and has the effect of stabilizing the microbial flora in the intestine by suppressing abnormal growth of harmful microorganisms in the intestine. The present inventor proposed a new acid-resistant Lactobacillus sakei probio-65 having the ability to inhibit the growth of harmful pathogenic microorganisms and anti-allergic in Korean Patent Registration No. 10-0479719.

According to a study investigating the immunomodulatory properties of the Lactobacillus sakei probio-65 strain, the expression of the Foxp3 + transcription regulator as a control in the Foxp3 + transcription regulator expression increase ability evaluation experiment was not high, but the Lactobacillus sakei probio-65 The results showed that Foxp3 + transcription regulator expression was higher in the live and dead cells, and showed higher values than the live cells in the dead cells. In addition, in the evaluation of the expression of anti-inflammatory cytokine Interleukin 10 (IL-10) by Lactobacillus sakei probio-65, the expression of IL-10 as a supernatant obtained after co-culture of Lactobacillus sakei probio-65 and mesenteric lymphocytes As a result of measuring the amount, the expression level of IL-10 increased when co-cultured with Lactobacillus S. probio-65 live cells or dead cells compared to PBS, and showed higher expression levels in the case of dead cells than the live cells. In the evaluation of the expression ability of Th1, Th17-related inflammatory cytokines by Lactobacillus S. probio-65, the expression of IFN-γ and IL-17 was higher when co-cultured with Lactobacillus S. probio-65. It was confirmed to appear. Therefore, through analysis of immunomodulatory substances, Lactobacillus sakei probio-65 is considered to be more likely to be used for immunomodulation or immunosuppression (hyperimmunosuppression) than for immunity enhancement. (Lim Jung-hee et al. 2011, Korean J. Microbiol. Biotechnol. 39: 313-316).

In in vivo animal experiments using NC / Nga rats, when the rats with skin disease were fed with live or dead bacteria of Lactobacillus sakei probio-65 cells, the results showed that the skin condition of the experimental animals improved, and where the mice were itchy. The number of scratches was reduced. In addition, serum analysis showed that IgE and cutaneous T-cell-attracting chemokine (CTACK) were significantly decreased, and in the case of mycelium, IL-4 and IL-6 cytokines were reduced in serum. These animal test results indicate that when the live or dead bacteria of Lactobacillus sakei probio-65 cells are fed to the experimental animals, it suppresses the development of skin diseases such as skin inflammation and atopic dermatitis (Kim et al. 2013. J. Appl.Microbiol. 115: 517-526).

The present inventors discovered that Lactobacillus sakei probio-65 bacteria can be used as a composition for alleviating or treating symptoms of atopic dermatitis while conducting research in relation to other efficacy of Lactobacillus sakei probio-65.

The intake of functional lactic acid bacteria is known in the past to be able to exert its functionality only when the lactic acid bacteria are used in a living state, i.e., as a probiotic that maintains its viability, but recently, each of them is a dead cell or ghost probiotics, in which the viability of lactic acid bacteria has disappeared. It has been reported that the unique functionality of Lactobacillus is expressed.

In the case of Lactobacillus plantarum L-137 strain, which is a lactic acid bacterium with immunomodulatory ability, the structural stability of the cell wall increases when lactic acid bacteria are replaced with dead cells, which induces the production of interleukin-2 when compared with live cells. Even in the heat treatment, the activity of the killed lactic acid bacteria is maintained. Even when stored at low temperatures, the cell wall of Lactobacillus plantarum L-137 becomes unstable or the cell wall weakens due to contact with the digestive fluid of the human body, making it more useful to kill bacteria (Fujiki et al. 2012, Biosci.Biotechnol.Biochem.76: 918 ~ 922).

In the study using Lactobacillus paracasei NCC 2461, which has the function of promoting the production of the immunomodulatory factor IL-10, it has been reported that the fungus is effective. In studies of changes in immunomodulatory substances in Human Peripheral Blood Mononuclear Cells (PBMCs), NCC 2461 cells produced higher IL-10 when treated with dead cells than with live cells. The heat-treated NCC 2461 differentiates microRNA expression and affects the stability of IL10 mRNA, and this probiotic has an RNA fragment, and the RNA fragment released through the crushed membrane by heat treatment affects the gene expression of the host. (Demont et al. 2015, J Allergy Clin Immunol. 137: 1264-1267).

The results of the two studies mentioned above on some bioactive functions of lactic acid bacterium show that lactic acid bacterium has a better effect than live bacteria. In addition to the functional improvement, in the case of live bacteria, decomposition of cell components occurs even at low temperature storage, but heat-treated bacteria have increased stability, and the use of the bacteria is used in the distribution process such as low-temperature storage necessary to maintain the number of bacteria in the production and sale of probiotics. The number of steps can be reduced, and live bacteria are difficult to apply to the production of various types of processed products that require a heat treatment process, whereas dead bacteria can be used to produce various products.

Inhibition of atopic dermatitis using lactic acid bacteria as live bacteria has no side effects and is in vivo safe, so various studies are underway. In particular, the ability to inhibit atopic activity of lactic acid bacteria isolated from kimchi has been reported previously, but it is necessary to prove the effectiveness of atopic inhibition by dead bacteria. In addition, commercially available techniques related to atopic inhibition of lactic acid bacteria killing have not been reported so far.

The technical problem to be solved by the present invention is atopic dermatitis, which has no side effects and has high immunological activity with atopic dermatitis inhibition and treatment efficacy, including Lactobacillus sakei probio-65 or Lactobacillus sakei probio-65 extract It is intended to provide a composition for reducing or treating symptoms.

The above and other objects of the present invention can be achieved by the present invention described below.

The composition for reducing or treating atopic dermatitis including the Lactobacillus sakei probio-65 bacterium having high immunity activity of the present invention is characterized by including the Lactobacillus sakei probio-65 bacterium as an active ingredient.

Here, the dead cell is characterized in that it is a lyophilisate of the culture medium containing the Lactobacillus strain S. probio-65 dead cell.

The bacteria is characterized in that it is a Lactobacillus sakay probio-65 dead powder prepared after crushing a lyophilisate of a culture medium containing the Lactobacillus sakay probio-65 dead bacteria.

In addition, the composition for reducing or treating atopic dermatitis comprising the Lactobacillus sakei probio-65 extract of the present invention is characterized by including the Lactobacillus sakei probio-65 extract as an active ingredient.

Here, the extract is an extract of a culture medium containing Lactobacillus sakei probio-65, a liquid used after a concentration process, or an extract of a culture medium containing Lactobacillus sakei probio-65 as a dry matter of a concentrate produced in the concentration process It is characterized by being a powder prepared after crushing.

The present invention reduces or treats atopic dermatitis symptoms, including Lactobacillus sakei probio-65 bacteria or extracts thereof having high immune activity that can be used for suppression and treatment of atopic dermatitis symptoms in children and adolescents with atopic dermatitis symptoms. It has the effect of the invention to provide a composition. In addition, the composition of the present invention can be provided as a composition that is easy to store and distribute and reprocess as it is provided by heat-treated bacteria.

1 is a view showing the clinical trial period and the time of visit to the hospital for the clinical subjects ingested the test group (dead bacteria) and the control group (Placebo).
Figure 2 is a summary of the study subject's clinical trial participation status and analysis group.
3 is a graph showing a 30% or 50% improvement in atopic dermatitis symptoms by intake of Lactobacillus sakei probio-65 bacteria.
4 is a graph showing the degree of improvement in symptoms of atopic dermatitis over time by ingestion of Lactobacillus sakei probio-65 bacteria.
Figure 5 is a graph showing the degree of dermatitis in experimental animals induced atopic dermatitis.
6 is a graph showing the frequency of scratching of the experimental animal induced atopic dermatitis.
7 is a photograph showing the skin of an experimental animal induced atopic dermatitis.
8 is a graph showing the change in IgE of the experimental animal induced atopic dermatitis.

The above-described objects, features and advantages of the present invention will become more apparent by describing in detail preferred embodiments of the present invention. The following detailed description is not to be described in a limiting sense, and the scope of the present invention is limited only by the appended claims, along with all ranges equivalent to those claimed.

Hereinafter, the present invention will be described in more detail with reference to the accompanying drawings and examples.

Example

Example  One Lactobacillus Sakay probio -65 culture solution

The Lactobacillus sakei probio-65 strain was stored in a freezer at -80 ° C, and the physiological activity of the stored strain was increased by passage, and then used as a seed for preparing Lactobacillus sakei probio-65 bacteria powder. Lactobacillus sakei probio-65 taken out of the freezer was inoculated in MRS broth (Difco laboratories, USA) medium, subcultured 2-3 times for 15 hours at a temperature of 37 ° C., and the seed culture medium in which growth was vigorous was selected. . After inoculating the selected seed culture medium with MRS broth medium, the cells were cultured at 37 ° C. for 15 hours, and the cultured cells were used as a main culture starter. The main medium for culturing Lactobacillus sakei probio-65 is 1% by weight of glucose, 1% by weight of yeast extract, 0.25% by weight of soypeptone, 0.1% by weight of polyoxyethelene sorbitan mono-oleate, Liquid containing 0.1% by weight of ammonium sulfate ((NH ₄ ) ₂ SO ₄ ), 0.01% by weight of magnesium sulfate (MgSO ₄ ), 0.1% by weight of potassium dihydrogen phosphate (KH ₂ PO ₄ ) and 90 to 99% by weight of water After the culture medium was sterilized at 121 ° C for 15 minutes, the Lactobacillus sakei Probio65 culture was prepared by liquid culture of the Lactobacillus sakei probio-65 at 37 ° C for 24 hours using the medium.

Example  2 Lactobacillus Sakay probio -65 Preparation of dead powder

200 L of Lactobacillus S. probio-65 culture medium was centrifuged using a continuous centrifuge to recover the cells, and the recovered L. Bacillus S. probio-65 cells were killed by heat treatment. The water content of the killed Lactobacillus sakei probio-65 cells was lyophilized in a freeze dryer (Ilshin Lab, Korea) for 64 hours to be removed. The lyophilized sample of Lactobacillus sakei probio-65 was pulverized and used as a powder of Lactobacillus sakei probio-65.

Example  3 In children and adolescents with atopic dermatitis symptoms Lactobacillus Sakay probio -65 Deadly  Effectiveness and stability evaluation (human application test)

Example 3-1 Criteria for selection and exclusion of clinical subjects

Children and adolescents with mild to moderate atopic dermatitis symptoms aged 3 to 18 years of age who have agreed to participate in the test and signed the written consent of the test subject and the parent (or legal representative) voluntarily. For the discrimination, those who had intermittent or persistent atopic dermatitis symptoms for more than 6 months and whose SCORAD score was 20 to 50 (mild-moderate) were selected.

In addition to atopic dermatitis, people with severe skin disease, systemic disease, those who are administering antibiotics, corticosteroids, antihistamines, immunosuppressants within 4 weeks of screening criteria, medicines, herbal medicines for improving atopic dermatitis within 4 weeks of screening criteria, People who are on dietary supplements, who have received phototherapy within 4 weeks of screening criteria, who have taken probiotics within 2 weeks of screening criteria, who are planning to participate in other trials during this trial, begin this trial Persons who participated in other clinical trials within 4 weeks and those deemed inappropriate by the investigators were excluded from the selection.

Example 3-2 Observational Test and Analysis of Results by Visit of Clinical Subjects

Example 3-2-1 Clinical subjects

It was judged that recruiting 25 individuals from each of the test groups consuming Lactobacillus sakei probio-65 bacteria or the control group would have a significant result. In the clinical trial, 30 patients were recruited each considering 15% of the dropouts.

1 is a view showing the clinical trial period and the time of visit to the hospital for the clinical subjects ingested the test group (dead bacteria) and the control group (Placebo). As can be seen in Figure 1, the clinical subjects were screened at the first visit, randomized (randomization) at the second visit, and divided into a test group (bacteria) and a control group (Placebo). For the examination of the course, after the start of the trial, we visited the clinical trial hospital at Visit 3 (week 6) and Visit 4 (week 12).

Example  3-2-2 Subjects By visit  Observation test method

Example  3-2-2-1 Clinical trial subject's first visit

Subjects who wished to participate in this clinical trial were evaluated in the following order after hearing the description of the clinical trial.

① The test process was explained before participation in the test, and written consent was given to the test subject and the parent (or legal representative) of the test subject.

② Subjects were assigned research screening numbers in order.

③ The demographic information, medical history (past and current medical history including surgical history) and drug administration history of the subjects were investigated.

④ Physical examination was conducted.

⑤ Vital signs (pulse, body temperature) and body measurements (height, weight) were measured.

⑥SCORAD index was evaluated.

⑦ Clinical pathology was performed.

⑧ After the suitability evaluation of the primary test subject, the next visit date of the test subject was designated.

Example 3-2-2-2 Clinical trial subject's second visit

The visit was made within 14 days of the initial visit, and the evaluations that should be made at this visit were as follows.

① Changes in medical history and drug administration were investigated and recorded.

② Physical examination was conducted.

③ Vital signs (pulse, body temperature) were measured.

④The suitability of the selection criteria / exclusion criteria was finally evaluated by synthesizing the evaluation results of the 1st and 2nd visits.

⑤A random number was assigned and assigned.

⑥ IGA evaluation was conducted.

⑦ Skin condition was measured (moisture, oil, pH).

⑧Eosinophil, IgE, ECP, CCL17, CCL27 test was performed.

⑨ Meal instruction and dietary research were conducted.

⑩ Test food / control food was prescribed, and intake method was trained.

⑪ The primary standard moisturizer was distributed.

⑫ The next visit was designated.

Example 3-2-2-3 Subject 3 visit

This visit was made 42 days (± 7 days) after the second visit, and the evaluations to be made at this visit were as follows.

① It was confirmed whether there was an adverse reaction.

② It was confirmed whether the combination drug and the combination therapy changed.

③ Physical examination was conducted.

④ Vital signs (pulse, body temperature) were measured and body measurements (weight) were performed.

⑤SCORAD index was evaluated.

⑥ IGA evaluation was conducted.

⑦ Skin condition was measured (moisture, oil, pH).

⑧ Meal instruction and dietary research were conducted.

⑨ The intake compliance was checked.

⑩ Test food / control food was prescribed, and the intake method was explained once again.

⑪ First standard moisturizer was confirmed to be returned, and a second standard moisturizer was distributed.

⑫ The next visit was designated.

Example 3-2-2-4 4th visit of clinical trial subjects

The 4th visit was completed on the 84th day (± 5 days) after the 2nd visit as the test end date, and the evaluations to be performed at this visit were as follows.

① It was confirmed whether there was an adverse reaction.

② Confirmation of the combination drug and the combination therapy.

③ Physical examination was conducted.

④ Vital signs (pulse, body temperature) were measured and body measurements (weight) were performed.

⑤Clinical pathology examination was conducted.

⑥SCORAD index was evaluated.

⑦ IGA evaluation was conducted.

⑧ Skin condition was measured (moisture, oil, pH).

⑨Eosinophil, IgE, ECP, CCL17 and CCL27 tests were performed.

⑩A dietary investigation was conducted.

⑪ Second standard moisturizer was confirmed to be returned.

취 The intake compliance was confirmed.

Example 3-2-3 Clinical trial result analysis target number

Example 3-2-3-1 Clinical trial stability result analysis number of subjects

The safety result analysis, Safety Set, is an evaluation index derived from a total of 60 subjects, 30 test groups and 30 control groups in which safety-related information was collected among subjects who ingested food for clinical trials more than once after being randomly assigned to clinical trials. Stability results were included in the analysis.

Example 3-2-3-2 Clinical trial effectiveness evaluation analysis number of subjects

The evaluation of the effectiveness of this clinical trial was based on the PP Set as the main analysis, and the FA Set was additionally analyzed.

The FA Set was conducted for groups that did not fall under the main selection criteria violation after conducting the food for clinical trial more than once and then conducting the efficacy evaluation more than once. Therefore, among 60 randomly assigned test subjects (30 test groups, 30 control groups), 9 patients (4 test groups, 5 control groups) who did not perform the evaluation of efficacy due to withdrawal of consent or follow-up failure were excluded. Fifty-one (26 test groups and 25 control groups) were included in the FA Set.

Effectiveness evaluation The main analysis target, PP Set, is one of the subjects included in the FA Set, who has ended the clinical trial and has no significant violations affecting the results of the clinical trial, with one dropout in the test group and less than 70% compliance 1 Person, visit 3 1 without SCORAD test, 4 persons with 1 visit violation, 1 dropout from control group, 2 with less than 70% compliance, 1 visit 3 without SCORAD test, 1 person taking anti-combination drug A total of 42 patients (22 test groups and 20 control groups) were included in the PP Set.

Example  3-3 Clinical Trial Method

The clinical trial for improving atopic dermatitis by ingestion of Lactobacillus sakei probio-65 bacteria according to the present invention was designed as a randomized, double-blind, placebo-controlled clinical trial of ingested bacteria samples. When a subject who has signed a clinical trial consent form by a voluntary or legal representative of a subject with atopic dermatitis is registered in this clinical trial, it is judged whether or not the selection criteria / exclusion criteria are met through a visit evaluation, and then the test is conducted only for suitable subjects The group was randomly assigned to one of the control and control groups. The test subjects assigned were ingested test or control food for 12 weeks.

Example 3-3-1 Clinical Trial Test Food and Placebo Controlled Trials

The test group ingested a sample containing Lactobacillus sakei probio-65 lyophilized powder as a test food. The number of bacteria in the intake was 1x10 ¹⁰ CFU / day. The ingested test food was prepared after mixing a cellulose as an excipient with a 2 g / pack product. The test subjects took the test foods one packet per day 30 minutes after eating directly or in water. Cellulose was used as a control food (Placebo), and the weight and intake method of the product was the same as the test food.

Example 3-3-2 Randomization of test food

Clinical trials were randomly assigned as a test group or a control group, and conducted as a parallel test. The required number of subjects was 60 in total, 30 in each group considering the dropout rate (15%).

At Visit 2, all subjects who met the selection and exclusion criteria were assigned to each group according to the allocation code of the block randomization method according to the institution. For a balanced randomization between intake groups, a certain block size was applied, and the ratio of the number of subjects in each group was equal to 1: 1. The randomization table was applied by sequentially applying the permutation of random numbers (A and B random numbers) generated by the randomization program of the SAS® system, starting with the subject number 1, and was created by designing in advance before the clinical trial through SAS®. When the sponsors packaged the food for clinical trials, they attached the food label for the clinical trial according to the randomization table and supplied it to the testing institution before the start of the clinical trial. Subsequently, in the case of the appropriate subject as the subject of the clinical trial at the visit 2, the following subject number is assigned in order, and the test / control food having the same number as the serial number is ingested.

Example 3-3-3 Clinical Trial Method Double-blind

In order to maintain double-blindness, in addition to the contents mentioned in the production / packaging and labeling of products used in this clinical trial, the allocation of unique codes for each group was managed in a sealed state by the clinical trial investigator, and serious adverse reactions occurred. It was not disclosed until the end of the clinical trial, except in cases where it was necessary to access the code. In this clinical trial, no blinding occurred during the trial period. The clinical trial investigator supplied the food with a unique code that matches the intake allocation number of the selected test subject, and maintained the blinding by using an extra (specific code) when the food for the clinical trial was defective or damaged.

Example 3-4 Safety evaluation

The safety evaluation and analysis of ingested foods for the clinical trial subjects who consumed the test group and the control group was conducted for all 60 test subjects in the safety set.

Example  3-4-1 Adverse event evaluation

For the evaluation of adverse events, evaluation of all adverse reactions reported during the test period was calculated by calculating the incidence rate after tabulating all the adverse reactions of the subjects. The proportion of test subjects with adverse reactions between groups was calculated and compared using the Chi-square test or Fisher's exact test.

Example  3-4-2 Clinical pathology examination

Paired t-test was used for intragroup comparison of continuous type data such as hematologic and hemochemical test values, and one-way ANOVA test was performed for comparison between groups. When there was a significant result in the comparison between groups, it was analyzed using the multiple comparison method. In the case of some measurement variables of the urine test, the McNemar test was performed by dividing it into normal and abnormal groups to compare the differences within the group.

Clinical pathology was performed at visits 1 and 4 (or termination visit upon dropout) to evaluate the general health of the subjects. If the results of the clinical pathology examination can be confirmed on the same day, Visit 1 and Visit 2 could be conducted at the same time. In this case, the selection criteria and the exclusion criteria including the results of clinical pathology tests were checked before the intake of food for clinical trials was made.

The clinical pathology test of Visit 1 was applicable if there were test results within 7 days, and reexamination of abnormal results was performed at the judgment of the investigator. The inspection items included:

Example 3-4-2-1 Hematologic clinical pathology test

Hematological examination was performed on 14 WBC, RBC, Hb, Hct, MCV, MCH, MCHC, Seg.Neutrophils, Platelet, Lymphocytes, Monocytes, Eosinophils, Basophils, ESR. WBC (white blood cells), Seg.Neutrophils, Lymphocytes, Monocytes, Eosinophils, and Basophils (white blood cell percentage) tests test for the cytological and physical properties and condition of blood, suggesting bacterial infections, identifying allergic diseases, and detecting RBC, Hb, Hct and platelet tests can diagnose polycythemia, anemia, and bleeding. In addition, MCV, MCH, MCHC, and ESR are used for diagnosis of vitamin B12 deficiency anemia, iron deficiency anemia, non-deficient anemia, and the progress and prognosis of various diseases.

Example 3-4-2-2 Hemochemical clinical pathology test

Hemochemical tests include 16 types of Calcium, Phosphorus, Glucose, BUN, Uric acid, Cholesterol, Total protein, Albumin, Total Bilirubin, Alk.phos, AST (GOT), ALT (GPT), Creatinine, Na, K, Cl, etc. It was performed as a subject. Various components in the blood are measured by biochemical methods.The serum protein (Albumin), lipid (Cholesterol), serum enzyme activity (Alk.phos), Phosphorus, AST (GOT), ALT (GPT), and Total Bilirubin test are liver diseases. It plays an important role in the diagnosis of the presence and absence of the disease, its degree and prognosis, and the treatment policy. In particular, it is useful for diagnosing disorders of the liver biliary system, such as parenchymal disorders of the liver and cholestasis (gallbladder juice). Electrolytes (NA, K, CL, Calcium) can diagnose diarrhea, vomiting, dehydration, and febrile conditions, and glucose tests diagnose diabetes, infection, and hypertrophy. BUN, Uric acid, and Creatinine are indicators that reflect immature renal function in childhood, and can diagnose pyelonephritis, urinary tract disease and nephrotic syndrome.

Example 3-4-2-3 Urine test

Urine test was performed to test the components of urine, including SG, pH, Protein, Glucose, Ketone, Bilirubin, Blood, Urobilinogen, Nitrite, RBC, and WBC. As a diagnostic method to detect the condition of the whole body and the lesion by examining the components of urine, abnormalities such as turbid urination or hematuria appear when there are lesions in the urinary tract, bladder, and urethra, which are pathways that produce urine, and metabolites of the whole body in urine Because it is excreted, it is also a general test for the diagnosis of systemic diseases other than the urinary system. The purpose of this study is to identify abnormal signs through urine tests and to determine the onset through precise tests to treat early detection.

Example  3-4-3 Vital signs (Pulse, body temperature) and weight

Vital signs (pulse, body temperature) and body weight were examined at the corresponding visit, and a paired t-test was used for intragroup comparison of the difference between before and after intake, and a one-way ANOVA test was performed for comparison between groups. When there was a significant result in the comparison between groups, it was analyzed using the multiple comparison method.

Example 3-5 Efficacy evaluation method

The effectiveness of Lactobacillus sakei probio-65 for improving atopic dermatitis was evaluated by dividing it into a primary efficacy evaluation variable and a secondary efficacy evaluation variable. The primary efficacy evaluation variables were evaluated by SCORAD (SCORing of Atopic Dermatitis) total scores, and the secondary efficacy evaluation variables were SCORAD objective scores, SCORAD subjective scores, IGA (Investigator's Global) scores. Assessment), skin condition (moisture, oil, pH) change, blood Eosinophil (count), IgE, ECP, CCL17 (TARC), CCL27 (CTACK) was determined.

Example  3-5-1 SCORAD ( SCORing  of Atopic  Dermatitis) index evaluation method

SCORAD (SCORing of Atopic Dermatitis) index evaluations were conducted on the 1st, 3rd and 4th visits of the test subjects. SCORAD index was evaluated by dividing into objective and subjective SCORAD.

Table 1 is a table showing the evaluation items and score range of the SCORAD index. As can be seen in Table 1, objective SCORAD (objective SCORAD) objectively assessed the extent (intensity,%) and intensity of the subject's body lesions. The extent of lesions (extent,%) was assessed by the percentage of lesions per body part.

The severity of lesions is erythema / darkening, edema / papulation, oozing / crusting, excoriation, lichenification, and xerosis 6 Divided into three categories, the severity was evaluated by 0 to 3 points (0; none, 1; mild, 2; medium, 3; severe). Subjective SCORAD (subjective SCORAD) is a method in which the test subject himself divides and displays the degree of pruritus and sleep loss symptoms by 0 to 10 (visual analog scale). However, when it was difficult for the subject to evaluate the severity of subjective SCORAD symptoms, the subject's parents (or legal representatives) evaluated instead, and were evaluated by the same evaluator at Weeks 0, 6, and 12.

SCORAD total scores are A (range of lesions), B (degree of lesions), and C (awareness symptoms)

After calculating the score for each item, it was calculated using the formula A / 5 + 7B / 2 + C. The score range is 0-103 points, and the higher the score, the more severe the symptoms.

SCORAD total score = A / 5 + 7B / 2 + C

Example  3-5-2 Overall symptom improvement ( IGA ; Investigator's  Global Assessment)

Assessment Methods

At Visits 2-4, the investigator evaluated the subjects' overall improvement in atopic dermatitis symptoms by dividing them by 0-5. Table 2 shows the IGA criteria and score ranges. Table 2 is a table showing the overall condition and criteria for improving symptoms of atopic dermatitis.

Example  3-5-3 skin condition measurement (moisture, Oil , pH) method

Moisture (Tewameter® TM 300 and Triple TM 330T), oil (Sebumeter® SM815), pH (with Courage + Khazaka electronic's Multiprobe Adapter 9 and Probes on the fore arm and lower leg skin areas at visits 2-4 Skin-pH-meter® PH905) was measured.

Example  3-5-4 Eosinophil , IgE , ECP , CCL17 ( TARC ), CCL27 ( CTACK ) method of inspection

Eosinophil (count), IgE, and ECP tests were performed by collecting blood at visits 2 and 4 as immune-related blood indicators. After centrifugation, IgE and ECP were stored refrigerated at 4 ℃, and the blood samples were analyzed and discarded.

Eosinophil (count) is one of the most important inflammatory cells in allergic and Th2 immune responses.It is known to induce inflammatory reactions by activating by Eotaxin to the site of inflammation and secreting various cytotoxic proteins and cytokines including ECP. . IgE serves as the most important immune indicator for mediating allergic diseases, and ECP is known to act as an important inflammatory mediator in allergic and Th2 specific immune responses. Chemokines are known as low-molecular proteins, represent chemoattractants, and can be divided into four subfamily types, C, CC, CXC, and CX3C, depending on the structure and function of the proteins. Infiltrate to further induce an inflammatory reaction. Typical Th2 chemokines of the CC family include thymus & activation-regulated chemokine (TARC / CCL17), macrophagederived chemokine (MDC / CCL22), and cutaneous T cell-attracting chemokine (CTACK / CCL27). It is known to induce infiltration. It is known that chemokines such as TARC / CCL17, MDC / CCL22 and CTACK / CCL27 are detected in serum of AD patients or AD-like animal models.

Example  3-6 Effectiveness Statistical Analysis Method

Statistical analysis was performed using SAS® (Version 94, SAS Institute, Cary, North Carolina, USA). The main analysis target of the efficacy evaluation analysis was the PP set, and the FA set was additionally performed.

Example  3-6- 1 primary  Analysis of efficacy evaluation variables

The degree of change before and after intake of the primary efficacy evaluation variable, SCORAD total score, was analyzed using paired t-test, and the difference between intake groups for the degree of change was statistically significant using two sample t-test. Was evaluated.

Example  3-6-2 Analysis of secondary efficacy evaluation variables

Secondary efficacy endpoints, SCORAD score (objective, subjective), IGA, skin condition (moisture, oil, pH), blood Eosinophil (count), IgE, ECP, CCL17 (TARC), CCL27 (CTACK) The degree was analyzed using the paired t-test, and the difference between the intake groups for the degree of change was evaluated to see if there was a statistically significant difference using the two sample t-test.

Additional analysis presented the percentage of subjects using topical steroids and moisturizers during the test period, and ingested using the Chi-square test or Fisher's * s exact test. The difference in proportions between groups was compared. In addition, a one-way ANOVA test was conducted to compare the moisturizer usage between groups. Among the basis characteristics of the test subjects, items with differences between groups were corrected to perform covariance analysis (ANCOVA).

Example  3-7 Test food Safety evaluation  result

The safety assessment was randomly assigned, and among the subjects who consumed food for clinical trials at least once, the subjects who collected safety-related information were considered as safety sets.

There were 4 adverse reactions in 4 subjects in the test group administration group, and 6 in 3 subjects in the control group. The main adverse reaction was infectious disease in both groups. A total of one serious adverse reaction occurred in one of the control groups, and it was determined by the investigator that it was not related to the food for clinical trials and was confirmed to be completely cured. No test subjects were dropped out due to adverse reactions.

Example  3-7-1 Test food Safety result  Adverse reactions

In the investigation of the severity of adverse reactions, there were 4 cases of mild in the test group, 5 cases of mild in the control group, and 1 case in the moderate, and there was no statistically significant difference between the intake groups.

In the relationship with food for clinical trials, 4 cases were clearly 'not related' in the test group administration group, 2 cases were 'possibly related' in the control group, 1 case was considered 'not related', and 'not clearly related' 'The three cases were judged by the tester, and there was no statistically significant difference between the intake groups.

There were 2 adverse events that could not be excluded from the relationship with food for clinical trials, and it was confirmed that the adverse reactions of the digestive system were completely cured.

Example  3-7-2 Hematology and Hemochemical test  Analysis

As a result of hematological and hemochemical analysis, after 12 weeks of intake, Platelet increased by 29.83 ± 58.89 10 ³ / μl in the test group administration group (p = 00109), and decreased by 10.47 ± 36.07 10 ³ / μl in the control group. One difference appeared (p = 00009), but all were within the normal range. As a result of further Dunnett analysis, it was confirmed that there was a significant difference between the test group (dead cells) and the control group. There was no statistically significant difference between intake groups in all other test items.

As a result of urine analysis, there was no statistically significant difference before or after ingestion in all test items.

In the vital signs (pulse, body temperature) and weight analysis results, there were no statistically significant differences between the intake groups 6 and 12 weeks after ingestion.

Example  3-8 Atopic dermatitis improvement results

SCORAD (SCORAD of total score), SCORAD objective score, SCORAD subjective score, IGA (Investigator '* s Global Assessment), skin condition (Moisture, oil, pH) changes, changes in blood Eosinophil (count), IgE, ECP, CCL17 (TARC), CCL27 (CTACK) analyzed and compared the improvement of the test group and the control group to see if there is a statistically significant difference. Was evaluated.

Example  3-8-1 atopic dermatitis SCORAD  total score result

As a result of analyzing the changes in the SCORAD total score of the test group (dead bacteria) and the control group with the PP set, the test group (dead bacteria) administration group decreased 10.51 ± 13.69 after 12 weeks of intake (p = 00017), and the control group decreased by 3.39 ± 8.50. There was a statistically significant difference between groups (Two sample t-test; p = 0.0486, Wilcoxon rank sum test; p = 0.0242) In the FA Set analysis, the test group decreased by 9.56 ± 13.30 after 12 weeks of intake (p = 0.0012), the control group decreased by 6.15 ± 10.28 (p = 0.0064), but there was no statistically significant difference between the intake groups.

Tables 3 and 4 are tables showing the results of analysis of changes in the SCORAD total score of the test group and the control group measured at 0 weeks, 6 weeks, and 12 weeks with PP Set and FA Set.

Example  3-8-2 atopic dermatitis SCORAD  objective (range of lesions) score result

As a result of analyzing the changes in SCORAD objective (range of lesions) scores of the test group (death) and the control group with the PP set, the test group (death) dose decreased 4.34 ± 9.30 after 12 weeks of intake (p = 0.0401), and the control group was 0.16. Although it increased by ± 11.89, there was no statistically significant difference between intake groups. In the results analyzed by FA Set, after 12 weeks of ingestion, the test group (dead bacteria) administration group decreased by 3.47 ± 9.27, and the control group decreased by 0.85 ± 10.91, but there was no statistically significant difference between the intake groups.

Example  3-8-3 atopic dermatitis SCORAD  objective score

The degree of lesion (intensity) is the erythema / darkening, edema / papulation, oozing / crusting, scouring, excoriation of the subject's body lesions. Divided into 6 items (lichenification) and dryness (xerosis), the severity was evaluated by 0 to 3 points (0; none, 1; mild, 2; medium, 3; severe). As a result of analyzing the total change in SCORAD objective scores of the test group (bacterium) and the control group with the PP set, the test group decreased 2.05 ± 2.90 after 12 weeks of intake (p = 0.0034), and the control group decreased by 0.70 ± 1.78 Therefore, there was a statistically significant difference between the intake groups (p = 0.0186).

Example  3-8-3-1 atopic dermatitis SCORAD  objective (degree of lesion) abrasion  score result

As a result of analyzing the change of the scratch score of the SCORAD objective (degree of lesion) with a PP set, the test group (bacterium) decreased 0.32 ± 0.84 after 12 weeks of ingestion, and the control group increased by 0.20 ± 0.70, showing a statistically significant difference between the intake groups. Appeared (p = 0.0363).

Example  3-8-3-2 atopic dermatitis SCORAD  Dry score result of objective (degree of lesion)

As a result of analyzing the change in the dry score of the SCORAD objective (degree of lesion) with a PP set, the test group (diploid) decreased 0.55 ± 0.80 after 12 weeks of intake (p = 0.0043), and the control group increased by 0.05 ± 0.76 to statistically increase between groups. Showed a significant difference (p = 0.0180 *, p = 0.0132)

In addition, there were no statistically significant differences between the ingestion groups in the score changes of erythema / cancer, edema / pachycardia, gin / skin, and thyroidus. There was no statistically significant difference between intake groups in the results analyzed with FA Set.

Example  3-8-4 atopic dermatitis SCORAD  subjective score total result

As a result of analyzing the total change in SCORAD subjective score of the test group (bacteria) and the control group with the PP set, the test group decreased by 2.48 ± 4.70 (p = 0.0220) after 12 weeks of ingestion (p = 0.0220), and the control group decreased by 0.98 ± 3.81, but statistically between the intake groups There were no significant differences.

In the analysis of changes in the feeling of pruritus and sleep disturbance score, there was no statistically significant difference between the intake groups. There was no statistically significant difference between intake groups in the results analyzed with FA Set.

Example  3-8-5 IGA  Change amount analysis result

As a result of analyzing the change in IGA of the test group (dead bacteria) and the control group with the PP Set, the test group decreased 0.59 ± 0.85 after 12 weeks of ingestion (p = 0.0039), and the control group decreased by 0.40 ± 0.60 (p = 0.0075). There were no statistically significant differences. The FA Set analysis also showed no statistically significant difference between intake groups.

Example  3-8-6 skin condition (moisture, Oil, pH ) Change amount analysis result

In the results of analyzing the changes in the skin condition (moisture, oil, pH) of the test group (dead bacteria) and the control group with the PP set, the lower leg left oil content increased 1.68 ± 2.94 after 12 weeks of ingestion (p = 0.0226), and the control group This 0.32 ± 1.70 decrease showed a statistically significant difference between intake groups (Two sample t-test; p = 0.0159, Wilcoxon rank sum test; p = 0.0132).

As a result of analyzing the change in the lower leg left oil fraction with FA Set, the test group increased by 1.52 ± 2.76 after 12 weeks of intake (p = 0.0149), and the control group increased by 0.13 ± 2.47, showing a statistically significant difference between intake groups (Wilcoxon rank sum test; p = 0.0351).

Example  3-8-7 Eosinophil  count, IgE , ECP , CCL17 ( TARC ), CCL27 ( CTACK ) Change amount analysis result

There was no statistically significant difference between the intake groups in the results of analyzing the changes in Eosinophil count, IgE, ECP, CCL17 (TARC), and CCL27 (CTACK) of the test group (dead bacteria) and the control group with the PP set and FA set.

Example  3-8-8 analysis results of steroid use

As a result of analyzing the presence or absence of the use of steroids in the test group (dead bacteria) and the control group during the test period with the PP set and FA set, no statistically significant difference was found between the intake groups.

Example  3-8-9 Moisturizer use and usage analysis result

During the test period, the presence or absence and use of moisturizers in the test group (dead bacteria) and the control group are PP

As a result of analysis by Set and FA Set, there was no statistically significant difference between intake groups.

3, a graph showing a tester with 30% or 50% improvement in atopic dermatitis symptoms by ingesting Lactobacillus sakei probio-65 bacteria.

As can be seen in Figure 3, about 59% of the testers who ingested Lactobacillus S. probio-65 dead cells (Dead cells) according to the present invention had an effect of improving the SCORAD score of 30% or more, but the control group (Placebo). The ingested tester confirmed that only about 30% had an effect of improving the SCORAD score of 30% or more. In addition, about 32% of the testers who ingested Lactobacillus S. probio-65 dead cells according to the present invention had an effect of improving the SCORAD score of 50% or more, but about 5% of the testers who took the control (Placebo) It was confirmed that there was an improvement effect of SCORAD score of more than 50%.

 4 is a graph showing the degree of improvement in symptoms of atopic dermatitis over time by ingestion of Lactobacillus sakei probio-65 bacteria.

As can be seen in Figure 4, in the case of Visit 3 (week 6) after the start of the test, about 22% of the testers who ingested Lactobacillus sakei probio-65 according to the present invention (Dead cell) according to the present invention improve the SCORAD score However, only about 13% of the intake of the control group (Placebo) confirmed that the SCORAD score had an improvement effect. In addition, in the case of Visit 4 (Week 12) after the start of the test, about 32% of the testers who consumed Lactobacillus S. probio-65 dead cells according to the present invention had an effect of improving the SCORAD score, but the control group (Placebo) It was confirmed that only about 11% of the intake subjects had an effect of improving the SCORAD score.

Therefore, it was confirmed that the Lactobacillus sakei probio-65 bacterium according to the present invention has excellent effect of reducing or treating atopic dermatitis symptoms.

Example 3-9 Preparation of Lactobacillus sakei Probio-65 culture extract

An extractant was added to the Lactobacillus sakei Probio-65 culture medium and stirred at room temperature to extract the internal material of the strain and the active material of the culture medium. The extract is filtered and concentrated under reduced pressure to separate the extractant and the extract, and then the concentrated extract is heated or concentrated or freeze-dried with a freeze dryer (Ilshin Lab, Korea) to extract Lactobacillus sakei Probio-65 culture liquid or powder. (Hereinafter 'SEL001') was prepared.

Example 3-10 Improvement of Atopic Symptoms of Lactobacillus Sakei Probio-65 Extract Using Animal Model Animal Experiment

Animal experiments were conducted on 6-week-old ICR male mice. A total of 18 animals were purchased for each experimental group and purchased from a central laboratory animal (Adapted from Central Lab. Animal Inc.) to adapt to environmental conditions of room temperature 23 ± 2 ℃, humidity 60 ± 5%, and contrast cycle 12 hours. , Korea) for 1 week after preliminary breeding, and conducted experiments to induce atopy, drug treatment, and lactic acid bacteria extract treatment.

Example 3-10-1 Classification of experimental groups

The experimental group was treated with atopic control group (DNCB) that induced atopy with DNCB, drug treatment group (DNCB + drug) treated with clobetasol, a therapeutic drug after atopic induction, and treatment with SEL001, a lactobacillus sakei Probio-65 extract after atopic induction. Group (DNCB + SEL001).

Each test sphere consisted of 6 experimental animals, and 2 were bred in one cage.

Example 3-10-2 Atopy induction in normal mice

Atopic induction by DNCB was performed for 6 days by partially modifying the method used by Renstrom et al. (1995). The epidermal hair of the mouse's back skin was removed, and the skin was treated with 0.1 ml of a 1% DNCB solution. After 48 hours, additional dermatitis-causing drugs were treated. Further treatment was performed once daily for 3 days in 0.12 ml of DNCB solution on the skin of the experimental animal. After inducing skin sensitization, the experimental group was treated with SEL001 (40mg / cm ² ), and the drug control test group was treated with clobetasol (40mg / cm ² ), a therapeutic drug. Drug treatment was continued for 3 weeks.

Example 3-10-3 Visual evaluation of pathology in atopic dermatitis-induced animals

The degree of atopic dermatitis in all experimental animals belonging to the animal experimental group was measured by the clinical evaluation visual evaluation method used in clinical trials by Kim et al. (2008). Evaluation of clinical trial animals included erythema / bleeding; edema; Cuticle peeling / smoothing; Four items of drying were evaluated. Each item was classified into 4 grades of 0, 1, 2, and 3 according to the severity of symptoms. The total dermatitis score was expressed as the sum of four evaluation items with the highest value of 12.

After the mice were treated with 0.1 ml and 0.12 ml of 1% DNCB solution, clinical symptoms similar to atopy were observed. The frequency of scratching of the experimental animals was measured after each animal was separated and the number of scratches during the unit observation time.

It was confirmed that the topical application of extract SEL001 in atopy-induced experimental animals with DNCB showed improvement in atopy symptoms.

Figure 5 is a graph showing the degree of dermatitis in experimental animals induced atopic dermatitis.

Figure 6 is a graph showing the frequency of scratching of the experimental animal induced atopic dermatitis

7 is a view showing the skin of an experimental animal induced atopic dermatitis.

Example 3-10-4 IgE changes in plasma in atopic dermatitis-induced animals

After the clinical sample was finished, blood was collected from the inferior vena cava of each test group animal. Blood samples were left standing at room temperature for 30 minutes, and then serum was separated by centrifugation. The concentration of total IgE contained in the serum sample was confirmed with an IgE ELISA kit for mice (Shibayagi, Shibukawa, Japan). The ELISA reaction used the standard method of the reagent supplier.

8 is a graph showing the change in IgE of experimental animals induced by atopic dermatitis.

As can be seen in Figure 8, mice with atopic dermatitis induced by DNCB show a significantly higher amount of IgE than the drug or SEL001 treated test group. In the serum of mice coated with SEL001 on the skin, after atopic induction, a lower amount of IgE was found compared to that of mice treated with nothing and those treated with drugs. These results indicate that SEL001 inhibits skin inflammation caused by DNCB.

Therefore, it was confirmed that the Lactobacillus sakei Probio-65 bacterium and Lactobacillus sakei Probio-65 extract according to the present invention are excellent in reducing or treating atopic dermatitis symptoms.

Although preferred embodiments of the present invention have been described above, the present invention is not limited to the specific embodiments described above. That is, a person having ordinary knowledge in the technical field to which the present invention pertains can make a number of changes and modifications to the present invention without departing from the spirit and scope of the appended claims, and all such appropriate changes and modifications Equivalents should also be considered within the scope of the present invention.

Claims (5)

A composition for reducing or treating symptoms of atopic dermatitis comprising Lactobacillus sakei probio-65 as an active ingredient. [6] The composition for reducing or treating symptoms of atopic dermatitis according to claim 1, wherein the dead cell is a lyophilisate of a culture medium containing Lactobacillus S. probio-65 dead cell. The method of claim 1, wherein the bacteria is a Lactobacillus S. probio-65 bacterial powder produced after crushing a lyophilisate of a culture medium containing Lactobacillus S. probio-65 bacteria, reducing the symptoms of atopic dermatitis. Or therapeutic compositions. A composition for reducing or treating symptoms of atopic dermatitis comprising Lactobacillus sakei probio-65 extract as an active ingredient. The method of claim 4, wherein the extract is an extract of a culture medium containing Lactobacillus sakei probio-65, a liquid used after a concentration process, or an extract of a culture medium containing Lactobacillus sakei probio-65 produced in a concentration process. A composition for reducing or treating symptoms of atopic dermatitis, which is a powder prepared after grinding the dried product of the concentrated concentrate.

Images