A lactic acid bacterium, a fermentation broth and their application in the preparation of a drug for contact dermatitis

Abstract

The present invention belongs to the field of biological medicine technology, and relates to a lactic acid bacterium, a fermentation broth and their application in the preparation of drugs for contact dermatitis. The preservation number of the strain described in the present invention is: CCTCC M 2019547, and the taxonomic name is Lactobacillus NT_Lac_YF1. This lactic acid bacterium has a high conversion ability for traditional Chinese medicine raw material liquid. The present invention also provides a method for preparing a fermentation broth using this Lactobacillus NT_Lac_YF1. The prepared fermentation broth has a significant effect on improving the symptoms of contact dermatitis mouse models.

Description

Technical Field

[0001] The invention belongs to the technical field of biomedicine and relates to lactic acid bacteria, fermentation liquid and application of the lactic acid bacteria in preparing a contact dermatitis medicine. Background Art

[0002] Contact dermatitis is an allergic disease caused by internal and external factors. It is characterized by polymorphic rash and exudative tendency, is prone to recurrence, and is accompanied by skin inflammation such as severe itching, which affects the patient's quality of life and may even cause mental illnesses such as depression. The disease occurs in about 15-20% of the population, and the incidence rate in children is as high as over 25%.

[0003] In clinical medicine, the treatment of contact dermatitis is mainly symptomatic treatment. There are some problems in its use: (1) Antihistamines have significant effects in the acute phase, but their effects gradually decrease in the chronic phase. Long-term use will cause drug dependence and mental effects, and have significant central nervous system inhibitory effects and large side effects. (2) Although hormone drugs can quickly control symptoms, they are prone to relapse or even aggravation after discontinuation of the drug, so oral administration is not recommended. (3) Some drugs contain immunomodulators. Long-term use or excessive dosage can cause kidney poisoning or leukopenia. In severe cases, toxic side effects such as aplastic anemia and toxic hepatitis may occur.

[0004] Contact dermatitis is well known in the field of traditional Chinese medicine. This disease belongs to the category of "invasion sore", "wet sore", "hydrangea wind", "rotating ear sore" in traditional Chinese medicine. Traditional Chinese medicine believes that contact dermatitis is caused by wind, dampness and heat blocking the skin. In treatment, both internal and external treatments are often used, among which external treatments are mainly used to eliminate and dredge, and also to control symptoms such as itching and skin damage.

[0005] How to give full play to the advantages of traditional Chinese medicine, screen out Chinese herbal preparations that are non-toxic, easy to take, have clear mechanisms of action, and can effectively improve the symptoms of contact dermatitis, replace or synergize with the current clinical treatment of the disease, and avoid the side effects caused by the large-scale use of antihistamines, glucocorticoids, and immunomodulators, which plays a vital role in improving patients' symptoms and improving their quality of life.

[0006] The combination of Chinese and Western medicine in the treatment of contact dermatitis not only takes advantage of the characteristics of Western medicine, which is quick and simple, has a rapid effect, focuses on the pathological mechanism of the disease, has strong targeting and good compliance, but also highlights the fact that Chinese medicine takes into account the external manifestations of the disease and improves the symptoms of the disease to achieve an organic combination of syndrome differentiation and disease differentiation. On the basis of syndrome differentiation and treatment, Chinese and Western medicine are combined to reduce the dosage of Western medicine and reduce the toxic side effects and adverse reactions of Western medicine.

[0007] In recent years, for the treatment of contact dermatitis, especially Western medicine treatment, there is still a lack of effective therapies. Clinically, topical glucocorticoids and oral antihistamines are mostly used. Although the symptoms can be well controlled in the short term, long-term use of topical hormonal drugs is likely to cause a series of adverse reactions such as skin atrophy, keratinization thickening, and telangiectasia. A large number of clinical studies have shown that traditional Chinese medicine compound treatment of contact dermatitis has good effects in relieving the condition, maintaining the stability of the condition, reducing disease recurrence, and improving the quality of life, with fewer adverse reactions and being easily accepted by patients, which is worthy of further clinical promotion. How to clarify the exact efficacy of traditional Chinese medicine and how to improve the utilization of the active ingredients of traditional Chinese medicine are current hotspots. At present, the main drugs for treating contact dermatitis are Phellodendron amurense, Cnidium monnieri, Atractylodes lancea, Galla chinensis, Prunella vulgaris, Chrysanthemum morifolium, Lonicera japonica, Sophora flavescens, Borneol, etc. Fermented traditional Chinese medicine, as a modern microbial transformation technology, has good effects on increasing the content and absorption of the active ingredients of traditional Chinese medicine and degrading the toxic and side effect components in traditional Chinese medicine.

[0008] In addition, from the perspective of the existing technology, CN 116270924 A discloses a fermentation extraction process for a drug composition for eczema, but the classification status and source of the fermentation strain are not clear. Similarly, CN 116585397 A discloses a drug composition for treating eczema-like skin diseases, its preparation method and application. The prepared drug has the effects of antibacterial, anti-inflammatory, clearing heat and detoxifying, and has significant therapeutic effects on acute, chronic and pediatric eczema-like skin diseases caused by the environment, pathogenic bacteria and immune disorders. The fermentation strain described is high-efficiency koji. As is well known, koji is formed by natural fermentation of grains, and there are many types of microorganisms in the microbial flora and its exact classification status cannot be determined. Whether it can be used in the pharmaceutical industry remains to be discussed. Earlier, CN 114774315 A discloses a strain of Lactobacillus rhamnosus, which has strong antibacterial ability, good anti-inflammatory and immunomodulatory functions, and can regulate the transformation value of mouse spleen lymphocytes, cellular inflammatory factors, and intestinal short-chain fatty acid levels; it can promote the activity of immune cells, reduce the inflammatory response, and significantly improve the symptoms of eczema patients. The lactic acid bacteria preparation described in this invention is a simple bacterial agent, which can be used as a drug or a health food, but whether it can be used for traditional Chinese medicine fermentation is not specifically described. To sum up, there is currently no report on using lactic acid bacteria to ferment traditional Chinese medicine to prepare a fermented traditional Chinese medicine preparation to improve the symptoms of contact dermatitis. Summary of the Invention

[0009] In view of this, the purpose of the present invention is to provide a lactic acid bacterium, which can effectively transform the traditional Chinese medicine raw material liquid to obtain a fermentation broth, and the obtained fermentation broth can effectively improve the symptoms of contact dermatitis.

[0010] To achieve the purpose of the present invention, the technical scheme adopted by the present invention is as follows:

[0011] A lactic acid bacterium NT_Lac_YF1, the Latin name of the lactic acid bacterium is Lactobacillus sp., which was deposited at the China Center for Type Culture Collection on July 12, 2019. The address is: Wuhan University, Wuhan, China, and the deposit number is CCTCC M2019547.

[0012] A fermentation broth is obtained by fermenting a traditional Chinese medicine raw material broth with the above-mentioned lactic acid bacterium NT_Lac_YF1.

[0013] Preferably, the traditional Chinese medicine raw materials contained in the above-mentioned traditional Chinese medicine raw material broth are: 1-2 parts by weight of Phellodendron amurense, 3 parts by weight of Cnidium monnieri, 2-5 parts by weight of Atractylodes lancea, 2 parts by weight of Chinese gall, 4 parts by weight of Prunella vulgaris, 1 part by weight of Chrysanthemum morifolium, 1-3 parts by weight of Lonicera japonica, 1-3 parts by weight of Sophora flavescens, and 0.5 part by weight of Borneol.

[0014] A preparation method of the above-mentioned fermentation broth is to amplify and culture the lactic acid bacterium NT_Lac_YF1, centrifuge it to prepare a bacterial sludge, inoculate it into the traditional Chinese medicine raw material broth, carry out microbial transformation for 3-5 days, and obtain a traditional Chinese medicine fermentation broth after filtering and sterilizing.

[0015] Preferably, in the present invention, the centrifugation condition is 8000 r / min.

[0016] Preferably, in the above-mentioned preparation method, the preparation method of the traditional Chinese medicine raw material broth is: mix the above-mentioned traditional Chinese medicine raw materials, add 8-10 times the mass of distilled water and soak for 1-2 hours, decoct with slow fire for 50-70 minutes to obtain the first decoction, filter with 6-8 layers of gauze, decoct the remaining solid phase with 5-6 times the mass of distilled water of the traditional Chinese medicine raw materials for another 30-40 minutes, and combine the two decoctions to form the traditional Chinese medicine raw material broth.

[0017] Preferably, in the above-mentioned preparation method, the amplification and culture of the lactic acid bacterium NT_Lac_YF1 is specifically: after the lactic acid bacterium NT_Lac_YF1 is resuscitated on a plate, inoculate it into a strain culture medium and carry out gradient amplification culture.

[0018] In the present invention, the strain culture medium is: 10.0 g of peptone, 10.0 g of beef extract, 5.0 g of yeast extract, 2.0 g of diammonium hydrogen citrate, 20.0 g of glucose, 1.0 mL of Tween 80, 5.0 g of sodium acetate, 2.0 g of dipotassium hydrogen phosphate, 0.58 g of magnesium sulfate, 0.25 g of manganese sulfate, 18.0 g of agar, 1000 mL of distilled water, and pH 6.2-6.6.

[0019] In the present invention, 10-fold gradient amplification culture is adopted.

[0020] Preferably, the culture conditions for the gradient amplification culture are: the inoculation amount is 3%-5%, static culture at 37°C for 2-4 days, and the cell content is 1.0×10 8 -1.0×10 9CFU / mL.

[0021] Preferably, the inoculation amount of the inoculation is 1.11×10 4 -1.11×10 5 CFU / L of the traditional Chinese medicine raw material liquid; the specific conditions for the microbial transformation are as follows: the initial pH value of the fermentation is 5.0 - 5.4, at 37°C, anaerobic fermentation is carried out for 3 - 5 days in a closed state, and the end point of the fermentation is based on the pH value of 3 - 3.5.

[0022] Application of the above fermentation broth in the preparation of drugs for contact dermatitis.

[0023] Compared with the prior art, the present invention has the following beneficial effects:

[0024] (1) Most microorganisms cannot grow and reproduce in the traditional Chinese medicine liquid. The lactic acid bacteria NT_Lac_YF1 described in the present invention also has good tolerance to the traditional Chinese medicine raw material liquid used in the present invention (can grow and reproduce in the traditional Chinese medicine raw material liquid), and has high conversion ability to the traditional Chinese medicine raw material liquid.

[0025] (2) The traditional Chinese medicine fermentation preparation prepared by fermenting with lactic acid bacteria NT_Lac_YF1 has a good improvement effect on the symptoms of the contact dermatitis mouse disease model. Description of the Drawings

[0026] Figure 1 For the staining characteristics of NT_Lac_YF1;

[0027] Figure 2 For the culture characteristics of NT_Lac_YF1. The left figure is the culture of lactic acid bacteria NT_Lac_YF1 in an anaerobic culture tank, and the right figure is the colony of lactic acid bacteria NT_Lac_YF1 on the MRS culture medium plate;

[0028] Figure 3 For the auricle symptoms after sensitization and excitation of different treatment groups, where: 1. Ear thickness difference; 2. Ear weight difference;

[0029] Figure 4 For the comparison of ear thickness difference and ear weight difference of mice in different treatment groups, where: 1. Ear thickness difference, 2. Ear weight difference;

[0030] Figure 5 For the comparison of serum inflammatory factors of different treatment groups, where: 1. IL-4, 2. LTB4; 3. LTC4. Detailed Embodiments

[0031] The present invention will be further described below in conjunction with embodiments. It should be understood that the specific embodiments described herein are only used to explain the present invention and are not used to limit the present invention. Any simple improvement to the preparation method of the present invention under the premise of the concept of the present invention belongs to the protection scope of the present invention. For the experimental methods without specific conditions in the following examples, they are usually carried out according to the well-known means in the art.

[0032] The test materials used in the following examples, unless otherwise specified, were all obtained by purchasing from conventional biochemical reagent stores and pharmaceutical stores.

[0033] The chemicals used in this example meet the chemical pure standard.

[0034] Unless otherwise specified in the following examples, the strain medium (MRS medium) used for culturing Lactobacillus NT_Lac_YF1 is as follows: peptone 10.0 g, beef extract 10.0 g, yeast extract 5.0 g, diammonium hydrogen citrate 2.0 g, glucose 20.0 g, Tween 80 1.0 mL, sodium acetate 5.0 g, dipotassium hydrogen phosphate 2.0 g, magnesium sulfate 0.58 g, manganese sulfate 0.25 g, agar 18.0 g, distilled water 1000 mL, pH 6.2 - 6.6.

[0035] Example 1: Obtaining of Lactobacillus NT_Lac_YF1 of the present invention

[0036] Sampling: Lactobacillus NT_Lac_YF1 of the present invention is from fermented pickled Chinese cabbage purchased from the vegetable market. Take 100 g, cut it into pieces with a sterilized scissors, grind it in a sterilized mortar, transfer it to a 50 mL sterile centrifuge tube, shake and mix well, centrifuge at 800 rpm for 5 minutes, and take the supernatant for standby. Isolation and purification: Take 1 mL of the above supernatant and inoculate it into 9 mL of MRS medium, anaerobically culture at 35 °C for about 3 days, perform gradient dilution, spread it on the MRS solid medium plate, anaerobically culture at 35 °C for 2 days, streak the suspected colonies on the MRS plate to obtain pure colonies, and perform staining and microscopic examination (such as Figure 1 ).

[0037] Identification: Culture the strain in MRS liquid medium, centrifuge, wash, obtain the bacterial cells to extract DNA, amplify the specific band with 16S rDNA universal primers, sequence, compare, and identify it as a new strain of Lactobacillus, named: NT_Lac_YF1. It was deposited at the China Center for Type Culture Collection on July 12, 2019. The address is: Wuhan University, Wuhan, China. Its deposit number is: CCTCC M 2019547, and the taxonomic name is Lactobacillus NT_Lac_YF1 (Lactobacillus sp.). This bacterium has good acid resistance and reproductive ability in traditional Chinese medicine. The culture temperature is 32 - 35 °C, and it is Gram-positive (such as Figure 1), cultured anaerobically in MRS medium, with good growth (such as Figure 2 ).

[0038] Example 2: Preparation of fermentation broth

[0039] 1) Mix 40 g of Phellodendron amurense, 60 g of Cnidium monnieri, 60 g of Atractylodes lancea, 40 g of Galla chinensis, 80 g of Prunella vulgaris, 20 g of Chrysanthemum morifolium, 60 g of Lonicera japonica, and 40 g of Sophora flavescens, add 3200 g of distilled water, soak for 2 hours, and decoct over low heat for 60 minutes to obtain the first decoction. Filter with 8-layer gauze, decoct the remaining solid phase with 2400 g of distilled water for another 30 minutes, and combine the two decoctions to form the traditional Chinese medicine raw material liquid;

[0040] 2) Take 10.0 g of peptone, 10.0 g of beef extract, 5.0 g of yeast extract, 2.0 g of diammonium hydrogen citrate, 20.0 g of glucose, 1.0 mL of Tween-80, 5.0 g of sodium acetate, 2.0 g of dipotassium hydrogen phosphate, 0.58 g of magnesium sulfate, 0.25 g of manganese sulfate, 18.0 g of agar, 1000 mL of distilled water, and pH 6.2 - 6.6 to prepare MRS medium as the strain medium;

[0041] 3) Inoculate the lactic acid bacterium NT_Lac_YF1 obtained in step 2) onto the plate of the strain medium and culture it by streaking. Pick a single colony and inoculate it into 10 mL of MRS medium, and culture it statically anaerobically at 35 °C for 3 days; after obtaining the liquid strain, perform 10-fold gradient expansion culture. The culture conditions are: inoculation amount 3% - 5%, static culture at 35 °C for 3 days, and the cell content is 1.0×10 8 -1.0×10 9 CFU / mL; obtain the bacterial liquid;

[0042] 4) Centrifuge the cultured bacterial liquid at 8000 r / min to prepare bacterial sludge; inoculate the traditional Chinese medicine raw material liquid with the bacterial sludge, and inoculate it into the traditional Chinese medicine raw material liquid at an inoculation amount of 1.0×10 4 -1.0×10 5 CFU / L, with an initial pH value of 5.0 - 5.4. After inoculation, select a fermentation container according to the prepared amount, ferment anaerobically at 35 °C for 3 - 5 days, and take the end point of fermentation as the standard when the pH value is 3 - 3.5 to obtain the traditional Chinese medicine fermentation broth;

[0043] 5) Centrifuge the fermentation broth at 8000 r / min for 10 min, and then filter it with a 0.45 nm filter membrane to obtain the traditional Chinese medicine fermentation broth;

[0044] 6) Divide 2000 mL of the prepared traditional Chinese medicine fermentation broth into two equal parts, seal one part and store it refrigerated at 4 °C, and add 5 g of borneol to the other part, mix well, and store it refrigerated at 4 °C.

[0045] Test Example 1 Experiment on the effect of fermentation broth on improving the symptoms of contact dermatitis mouse model

[0046] 1) Preparation of DNFB solution: 2,4-dinitrofluorobenzene (DNFB) was dissolved in a mixed solution of acetone: olive oil (4:1) to prepare a 0.5% DNFB solution.

[0047] 2) Establishment of contact dermatitis model: This method was carried out with reference to the "Specification for the Preparation of Eczema Animal Models (Draft)" of the Professional Committee of Traditional Chinese Medicine Experimental Pharmacology of the China Association of Chinese Medicine and in combination with the literature "Comparative Study on the Construction of Allergic Contact Dermatitis in Mice by Two Methods" published by Zhu Xiaoxiao et al. The specific method is as follows: Thirty-six C57BL / 6 male mice at 6-8 weeks of age and weighing 18-20 g were taken. After 1 week of adaptive feeding, the abdominal hair of the mice was shaved, with an area of 3 cm × 3 cm. They were randomly divided into 6 groups, with 6 mice in each group, and were housed separately in cages. One group was set as the blank group without any treatment. After 2 days of adaptation for the remaining 5 groups of mice after shaving, the construction of the allergic contact dermatitis model was started. The method is as follows: 25 μL of 0.5% DNFB was dropped on the shaved abdominal area of each mouse and evenly smeared for sensitization for 2 consecutive days.

[0048] 3) Drug treatment: The 5 groups of sensitized mice were respectively set as the model group, compound flumetasone ointment group, traditional Chinese medicine raw material liquid group, traditional Chinese medicine fermented liquid group, and traditional Chinese medicine fermentation preparation group. Different treatments were carried out on all groups of mice. Among them, in the blank group, 25 μL of distilled water was smeared on the abdominal depilated area of each mouse twice a day, with an interval of 12 hours each time, for 5 consecutive days; in the model blank group, 25 μL of distilled water was smeared on the abdominal depilated area for 5 consecutive days; in the compound flumetasone ointment group, compound flumetasone ointment was smeared on the abdominal depilated area for 5 consecutive days; in the traditional Chinese medicine raw material liquid group, 25 μL of traditional Chinese medicine raw material liquid was smeared on the abdominal depilated area for 5 consecutive days; in the traditional Chinese medicine fermented liquid group, 25 μL of traditional Chinese medicine fermented liquid was smeared on the abdominal depilated area for 5 consecutive days; in the traditional Chinese medicine fermentation preparation group, 25 μL of traditional Chinese medicine fermentation preparation was smeared on the abdominal depilated area for 5 consecutive days. The above treatments for each group were carried out twice a day, with an interval of 12 hours each time, for 5 consecutive days. After 5 days of treatment, the right ears of the 6 groups of mice were challenged with 20 μL of 0.1% DNFB on both sides, and the left ears were used as a control with an acetone: olive oil (4:1) matrix solution.

[0049] 4) Detection indicators

[0050] Corresponding detections were carried out according to the following indicators

[0051] Scoring of the skin lesion degree of the animal model: The degree of auricle skin damage was observed and scored 4 days after challenge in the mice of each experimental group. The scoring criteria are as follows: Obvious erythema, edema and exudation, or obvious keratosis and crusting are scored 3 points; Mild erythema, edema, with no obvious exudation, or moderate keratosis are scored 2 points; Erythema, edema and exudation are not obvious, or mild keratosis is scored 1 point; Normal skin is scored 0 points (such as Figure 3 )

[0052] Detection of cytokine content in mouse serum: 4 days after stimulation of mice, canthal vein serum was collected, and ELISA kits were used to determine the levels of interleukin 4 (IL-4), leukotriene B4 (LTB4), and leukotriene C4 (LTC4) in serum, and the differences in related inflammatory factors among the groups were compared.

[0053] Measurement of mouse auricle swelling: 4 days after the mice were stimulated, the skin damage of the mouse auricle was observed and recorded, and then the mice were killed, and the thickness of the same part of the left and right ears of each mouse was measured. The thickness of the middle part of the left and right ears of the mouse was measured with a micro-caliper, and the difference in thickness between the left and right ears was calculated, ear thickness difference = right ear thickness average (mm) - left ear thickness average (mm), and the ear thickness difference between the experimental groups was compared; the ear pieces on both sides of the mouse were cut off, and the same part of the left and right ear pieces was taken with a 6mm punch, and they were immediately weighed with a 1 / 10,000 balance, and the weight difference between the left and right ear pieces was calculated, ear weight difference = right ear thickness average (mm) - left ear thickness average (mm), and the ear weight difference between the experimental groups was compared.

[0054] The model group, compound flumethasone ointment group, Chinese medicine raw material liquid group, Chinese medicine fermentation liquid group and Chinese medicine fermentation preparation group all had different degrees of ear swelling, and the ear thickness difference increased. The results are shown in Table 1. Figure 4 As shown:

[0055] 5) Experimental results and analysis

[0056] The ear thickness and swelling of mice in each group are shown in Table 1:

[0057] Table 1 Comparison of ear thickness and swelling of mice in each group (n=6)

[0058] Group n Degree of skin lesion Difference in ear thickness (mm) Difference in ear weight (mg) Blank 6 0(6 / 6) 0.008±0.004 0.133±0.137 Model 6 3(5 / 6)2(1 / 6) 0.175±0.012* 5.017±0.796* Compound flumetasone ointment 6 2(3 / 6)1(3 / 6) 0.055±0.021*▲ 1.067±0.299*▲ Original Chinese medicine solution 6 2(4 / 6)1(2 / 6) 0.080±0.021*▲ 2.383±0.578*▲ Fermented Chinese medicine solution 6 2(3 / 6)1(2 / 6) 0.060±0.013*▲★☆ 1.833±0.539*▲★☆ Fermented Chinese medicine preparation 6 2(3 / 6)1(2 / 6) 0.058±0.021*▲★■☆ 1.6±0.438*▲★■☆

[0059] Notes: *Compared with the blank group, P < 0.01; ▲Compared with the model group, P < 0.01; ★Compared with the Chinese medicine raw material group, P < 0.01; ■Compared with the Chinese medicine fermentation liquid group, P > 0.05; ☆Compared with the compound flumethasone ointment group, P > 0.05

[0060] The results of serum-related inflammatory factors in mice in each experimental group are shown in Table 2. Figure 5 As shown:

[0061] Table 2 Blood IL-4, LTB4, LTC4 levels in each group (x±s, pg / mL, n=6)

[0062] Group IL-4 LTB4 LTC4 Blank 14.87±1.66 67.35±7.26 229.37±27.54 Model 28.17±3.94* 135.97±14.03* 471.26±43.54* Compound flumetasone ointment 18.74±2.82*▲ 84.79±8.24*▲ 279.04±33.49*▲ Original Chinese medicine solution 20.11±2.10*▲ 96.20±9.33*▲ 314±31.13*▲ Fermented Chinese medicine solution 18.64±2.11*▲★☆ 86.91±7.90*▲★☆ 276.04±23.53*▲★☆ Fermented Chinese medicine preparation 17.45±1.65*▲★■☆ 80.44±5.25*▲★■☆ 279.69±22.72*▲★■☆

[0063] Note: *P < 0.01 compared with the blank group; ▲P < 0.01 compared with the model group; ★P < 0.01 compared with the traditional Chinese medicine raw material group; ■P > 0.05 compared with the traditional Chinese medicine fermentation broth group; ☆P > 0.05 compared with the compound flumetasone ointment group

[0064] Test conclusion: In the above test, when the fermentation broth was topically applied to contact dermatitis (on a mouse model), it could effectively reduce the incidence of induced dermatitis in the ears of mice, reduce the degree of skin damage on the epidermis of the ears of the mouse model; reduce the ear thickness difference and ear weight difference; effectively inhibit the increase of interleukin-4 (IL-4), leukotriene B4 (LTB4), and leukotriene C4 (LTC4) in the blood. This preparation has a good prevention and treatment effect on both clinical symptoms and inflammatory factors.

[0065] The above are only the preferred embodiments of the present application and are not used to limit the present application. For those skilled in the art, various changes and modifications can be made to the present application. Any modification, equivalent replacement, improvement, etc. made within the spirit and principle of the present application shall be included within the protection scope of the present application.

Claims

1. A lactic acid bacterium ( Lactobacillus sp. ), NT_Lac_YF1, characterized in that It is preserved in the China Center for Type Culture Collection with the preservation number of CCTCC M 2019547.

2. A fermentation broth, characterized in that, It is obtained by fermenting the traditional Chinese medicine raw material liquid with the lactic acid bacterium NT_Lac_YF1 described in claim 1. The traditional Chinese medicine raw materials contained in the traditional Chinese medicine raw material liquid are: 1-2 parts by weight of Phellodendri Chinensis Cortex, 3 parts by weight of Fructus Cnidii, 2-5 parts by weight of Atractylodis Rhizoma, 2 parts by weight of Galla Chinensis, 4 parts by weight of Prunellae Spica, 1 part by weight of Chrysanthemi Flos, 1-3 parts by weight of Lonicerae Japonicae Flos, 1-3 parts by weight of Sophorae Flavescentis Radix, and 0.5 part by weight of Borneolum Syntheticum.

3. The preparation method of the fermentation broth according to claim 2, characterized in that, The lactic acid bacterium NT_Lac_YF1 is amplified and cultured and then centrifuged to prepare a bacterial sludge, which is inoculated into the traditional Chinese medicine raw material liquid, and undergoes microbial transformation for 3-5 days. After filtration and sterilization, the traditional Chinese medicine fermentation liquid is obtained.

4. The preparation method according to claim 3, wherein, The preparation method of the traditional Chinese medicine raw material liquid is: mixing the traditional Chinese medicine raw materials, adding 8-10 times the mass of distilled water and soaking for 1-2 hours, decocting with a gentle fire for 50-70 minutes to obtain the first decoction, filtering with 6-8 layers of gauze, decocting the remaining solid phase with 5-6 times the mass of the traditional Chinese medicine raw materials of distilled water for another 30-40 minutes, and combining the two decoctions to form the traditional Chinese medicine raw material liquid.

5. The preparation method according to claim 3, characterized in that, The amplification and culture of the lactic acid bacterium NT_Lac_YF1 specifically is: after the lactic acid bacterium NT_Lac_YF1 is resuscitated on a plate, it is inoculated into a strain culture medium for gradient amplification culture.

6. The preparation method according to claim 5, characterized in that, The culture conditions for the gradient enrichment culture are as follows: inoculum size 3%-5%, static culture at 37°C for 2-4 days, and the cell content is 1.0×10 8 -1.0×10 9 CFU / mL.

7. The preparation method according to claim 3, characterized in that, The inoculation amount of the inoculation is 1.11×10 4 -1.11×10 5 CFU / L of the traditional Chinese medicine raw material liquid; the specific conditions for the microbial transformation are as follows: the initial pH value of the fermentation is 5.0 - 5.4, the temperature is 32°C - 37°C, and anaerobic fermentation is carried out in a closed state for 3 - 5 days. The end point of the fermentation is based on the pH value of 3 - 3.

5.

8. Use of the fermentation liquid described in claim 2 in the preparation of a drug for treating contact dermatitis.

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