A probiotic lysate composition and its application in preparing a product with anti-enteritis effect

Abstract

The present invention specifically discloses a probiotic lysate composition and its application in the preparation of a product with anti-enteritis effect. The probiotic lysate composition comprises a Lactobacillus rhamnosus lysate, a Bifidobacterium lysate and a Lactobacillus paracasei lysate. Studies have shown that the probiotic lysate composition of the present invention has anti-enteritis effects and also has the effect of alleviating intestinal damage caused by enteritis. Therefore, using the probiotic lysate composition of the present invention as an effective ingredient for preparing foods, functional foods or medicines with anti-enteritis effects and / or alleviating intestinal damage caused by enteritis has important application value.

Description

Technical Field

[0001] The present invention relates to the technical field of biomedicine, and in particular to a probiotic lysate composition and application thereof in preparing a product with anti-enteritis effect. Background Art

[0002] Enteritis is an inflammation of the small intestine and colitis caused by bacteria, viruses, fungi, and parasites. Abdominal pain, diarrhea, and ulcers and erosions are the primary clinical manifestations of enteritis. Treatments for enteritis primarily include Western medicine, traditional Chinese medicine, microbial preparations, and colon dialysis. However, these treatments are often unsatisfactory and prone to relapse.

[0003] Studies have shown that the balance of intestinal flora is crucial to human intestinal health, and an imbalance in intestinal flora can lead to the occurrence of intestinal inflammation and other related diseases, such as chronic proctitis. Probiotics, as common microorganisms, can colonize the human intestine, regulate the intestinal microenvironment, and are crucial to human intestinal health. Live probiotics or active substances such as short-chain fatty acids they secrete play an important role in the treatment of intestinal diseases. A large amount of research data shows that supplementing with probiotics is an effective method for preventing and treating gastrointestinal diseases. Live probiotics or active substances such as short-chain fatty acids they secrete play an important role in enhancing cellular immune responses, regulating cytokine secretion, and stabilizing the intestinal barrier. Whether dead probiotics or their lysates have biological activity, especially in terms of anti-inflammatory properties, and whether they can relieve enteritis are rarely reported, and their mechanism of action is not very clear. Summary of the Invention

[0004] In order to overcome at least one of the technical problems existing in the prior art, the present invention first provides a probiotic lysate composition.

[0005] The above technical problems to be solved by the present invention are achieved through the following technical solutions:

[0006] The present invention first provides a probiotic lysate composition, which comprises a Lactobacillus rhamnosus lysate, a Bifidobacterium lysate and a Lactobacillus paracasei lysate.

[0007] Studies have shown that a probiotic lysate composition comprising a Lactobacillus rhamnosus lysate, a Bifidobacterium lysate and a Lactobacillus paracasei lysate has anti-enteritis activity and also has the effect of alleviating intestinal damage caused by enteritis.

[0008] Preferably, the mass ratio of the Lactobacillus rhamnosus lysate, the Bifidobacterium lysate and the Lactobacillus paracasei lysate is 1:1-100:1-100.

[0009] More preferably, the mass ratio of the Lactobacillus rhamnosus lysate, the Bifidobacterium lysate and the Lactobacillus paracasei lysate is 1:1-25:1-25.

[0010] Most preferably, the mass ratio of the Lactobacillus rhamnosus lysate, the Bifidobacterium lysate and the Lactobacillus paracasei lysate is 1:12:5.

[0011] Preferably, the Lactobacillus rhamnosus lysate is prepared by the following method:

[0012] S11. Lactobacillus rhamnosus was added to PBS buffer and ultrasonically lysed to obtain a preliminary lysate of Lactobacillus rhamnosus;

[0013] S12. The preliminary Lactobacillus rhamnosus lysate is first subjected to ultrafiltration and dialysis using a 1 kDa ultrafiltration membrane; then, the retentate after ultrafiltration and dialysis using the 1 kDa ultrafiltration membrane is subjected to ultrafiltration and dialysis using a 3 kDa ultrafiltration membrane. The ultrafiltrate after ultrafiltration and dialysis using the 3 kDa ultrafiltration membrane is lyophilized to obtain the Lactobacillus rhamnosus lysate (i.e., the Lactobacillus rhamnosus lysate having a molecular weight of 1-3 kDa).

[0014] More preferably, the mass amount of PBS buffer is 4 to 6 times that of Lactobacillus rhamnosus.

[0015] Further preferably, the PBS buffer is a PBS buffer with a pH of 6.5; and the ultrasonic lysis has an ultrasonic time of 10 to 20 minutes.

[0016] Preferably, the bifidobacterium lysate is prepared by the following method:

[0017] S21. Bifidobacterium was added to PBS buffer and ultrasonically lysed to obtain a preliminary lysate of bifidobacteria;

[0018] S22. The preliminary lysate of bifidobacteria was first subjected to ultrafiltration and dialyzation using a 1 kDa ultrafiltration membrane; then the retentate after ultrafiltration and dialyzation using a 1 kDa ultrafiltration membrane was subjected to ultrafiltration and dialyzation using a 3 kDa ultrafiltration membrane, and the ultrafiltrate after ultrafiltration and dialyzation using a 3 kDa ultrafiltration membrane was lyophilized to obtain the bifidobacterium lysate (i.e., a bifidobacterium lysate having a molecular weight of 1-3 kDa).

[0019] More preferably, the mass amount of PBS buffer is 4 to 6 times that of Bifidobacterium.

[0020] Further preferably, the PBS buffer is a PBS buffer with a pH of 6.5; and the ultrasonic lysis has an ultrasonic time of 10 to 20 minutes.

[0021] Preferably, the Lactobacillus paracasei lysate is prepared by the following method:

[0022] S31. Lactobacillus paracasei was added to PBS buffer and ultrasonically lysed to obtain a preliminary lysate of Lactobacillus paracasei;

[0023] S32. the preliminary lysate of Lactobacillus paracasei is first subjected to ultrafiltration dialysis with the ultrafiltration membrane of 1kDa; then the retentate after the ultrafiltration dialysis with the ultrafiltration membrane of 1kDa is subjected to ultrafiltration dialysis with the ultrafiltration membrane of 3kDa; again the retentate after the ultrafiltration dialysis with the ultrafiltration membrane of 3kDa is subjected to ultrafiltration dialysis with the ultrafiltration membrane of 5kDa, and the ultrafiltrate after the ultrafiltration dialysis with the ultrafiltration membrane of 5kDa is freeze-dried to obtain the described Lactobacillus paracasei lysate (i.e., the Lactobacillus paracasei lysate with a molecular weight of 3-5kDa).

[0024] More preferably, the mass amount of PBS buffer is 4 to 6 times that of Lactobacillus paracasei.

[0025] Further preferably, the PBS buffer is a PBS buffer with a pH of 6.5; and the ultrasonic lysis has an ultrasonic time of 10 to 20 minutes.

[0026] The inventors surprisingly found in a large number of experiments that when the Lactobacillus rhamnosus lysate with a molecular weight of 1-3 kDa, the Bifidobacterium lysate with a molecular weight of 1-3 kDa, and the Lactobacillus paracasei lysate with a molecular weight of 3-5 kDa prepared according to the above-mentioned method of the present invention are combined, their anti-enteritis and intestinal damage relief effects caused by enteritis are significantly better than those of the Lactobacillus rhamnosus lysate with a molecular weight of 1-3 kDa, the Bifidobacterium lysate with a molecular weight of 1-3 kDa, or the Lactobacillus paracasei lysate with a molecular weight of 3-5 kDa alone; this shows that the combined use of the three has a significant synergistic effect of anti-enteritis and intestinal damage relief caused by enteritis.

[0027] In addition, the inventor has also found in research, after any two combinations of the lactobacillus rhamnosus lysate that molecular weight is 1-3kDa, the bifidobacterium lysate that molecular weight is 1-3kDa and the lactobacillus paracasei lysate that molecular weight is 3-5kDa, or after the lactobacillus rhamnosus lysate, bifidobacterium lysate and the lactobacillus paracasei lysate combination of other molecular weights that adopt alternative method to prepare, can not produce the effect of collaborative anti-enteritis and the intestinal tract injury that enteritis causes between them.The effect of their anti-enteritis and the intestinal tract injury that enteritis causes will be significantly less than the compositions obtained after the combination of the lactobacillus rhamnosus lysate that molecular weight is 1-3kDa, the bifidobacterium lysate that molecular weight is 1-3kDa and the lactobacillus paracasei lysate that molecular weight is 3-5kDa that prepare according to the above method of the present invention.

[0028] The present invention also provides a use of the probiotic lysate composition in preparing a product with anti-enteritis effect.

[0029] More preferably, the anti-enteritis agent is anti-intestinal epithelial cell inflammation.

[0030] Preferably, the probiotic lysate composition is used in the preparation of a product having an enteritis effect and simultaneously having the effect of alleviating intestinal damage caused by enteritis.

[0031] Preferably, the product is a medicine.

[0032] Beneficial effects: The present invention provides a new probiotic lysate composition; studies have shown that the probiotic lysate composition has an anti-enteritis effect and also has the effect of alleviating intestinal damage caused by enteritis. Therefore, using the probiotic lysate composition of the present invention as an effective ingredient to prepare a medicine with an anti-enteritis effect and / or an effect of alleviating intestinal damage caused by enteritis has important application value. In addition, the probiotic lysate composition of the present invention can be obtained by lysing Lactobacillus rhamnosus, Bifidobacterium and Lactobacillus paracasei, and the preparation process is simple and easy to operate, and large-scale production and application can be achieved. BRIEF DESCRIPTION OF THE DRAWINGS

[0033] In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the following briefly introduces the drawings required for use in the embodiments or the description of the prior art. Obviously, the drawings described below are only drawings of some embodiments of the present invention. For ordinary technicians in this field, other drawings can be obtained based on these drawings without paying any creative work.

[0034] Figure 1 The experimental results show that the probiotic lysate composition improves the vitality of intestinal epithelial cells, inhibits the DSS-induced decrease in SOD activity of intestinal epithelial cells, and inhibits the increase in MDA secretion.

[0035] Figure 2 Figure 3. Experimental results of probiotic lysate composition regulating DSS-induced inflammatory cytokine secretion in intestinal epithelial cells.

[0036] Figure 3 Figure 3. Experimental results of the effect of probiotic lysate composition on the expression of NF-κB signaling pathway proteins in DSS-induced intestinal epithelial cells.

[0037] Figure 4 Figure 3. Experimental results showing the effect of the probiotic lysate composition on the body weight and colon length of DSS-induced mice.

[0038] Figure 5Figure 3. Experimental results of probiotic lysate composition inhibiting DSS-induced secretion of inflammatory factors in mouse cells. DETAILED DESCRIPTION

[0039] The following is a clear and complete description of the technical solutions of the present invention in conjunction with the embodiments. Obviously, the embodiments described are only some of the embodiments of the present invention, not all of them. All other embodiments derived by persons of ordinary skill in the art based on the embodiments of the present invention without inventive effort are also within the scope of protection of the present invention.

[0040] Example 1 Preparation method of Lactobacillus rhamnosus lysate

[0041] S11. Take 20 g of Lactobacillus rhamnosus and suspend it in 100 g of PBS buffer (pH 6.5) and ultrasonically lyse it for 15 min to obtain a preliminary lysate of Lactobacillus rhamnosus.

[0042] S12. The preliminary Lactobacillus rhamnosus lysate is first subjected to ultrafiltration and dialysis using a 1 kDa ultrafiltration membrane; then, the retentate after ultrafiltration and dialysis using the 1 kDa ultrafiltration membrane is subjected to ultrafiltration and dialysis using a 3 kDa ultrafiltration membrane. The ultrafiltrate after ultrafiltration and dialysis using the 3 kDa ultrafiltration membrane is lyophilized to obtain the Lactobacillus rhamnosus lysate (i.e., the Lactobacillus rhamnosus lysate having a molecular weight of 1-3 kDa).

[0043] Example 2 Preparation method of bifidobacterium lysate

[0044] S21. Take 20 g of Bifidobacterium, suspended in 100 g of PBS buffer at pH 6.5, and ultrasonically lysed for 15 min to obtain a preliminary lysate of Bifidobacterium;

[0045] S22. The preliminary lysate of bifidobacteria was first subjected to ultrafiltration and dialyzation using a 1 kDa ultrafiltration membrane; then the retentate after ultrafiltration and dialyzation using a 1 kDa ultrafiltration membrane was subjected to ultrafiltration and dialyzation using a 3 kDa ultrafiltration membrane, and the ultrafiltrate after ultrafiltration and dialyzation using a 3 kDa ultrafiltration membrane was lyophilized to obtain the bifidobacterium lysate (i.e., a bifidobacterium lysate having a molecular weight of 1-3 kDa).

[0046] Example 3 Preparation method of Lactobacillus paracasei

[0047] S31 take Lactobacillus paracasei 20g, suspended in 100gpH 6.5 PBS buffer, ultrasonic lysis 15min, to give Lactobacillus paracasei preliminary lysate;

[0048] S32. the preliminary lysate of Lactobacillus paracasei is first subjected to ultrafiltration dialysis with the ultrafiltration membrane of 1kDa; then the retentate after the ultrafiltration dialysis with the ultrafiltration membrane of 1kDa is subjected to ultrafiltration dialysis with the ultrafiltration membrane of 3kDa; again the retentate after the ultrafiltration dialysis with the ultrafiltration membrane of 3kDa is subjected to ultrafiltration dialysis with the ultrafiltration membrane of 5kDa, and the ultrafiltrate after the ultrafiltration dialysis with the ultrafiltration membrane of 5kDa is freeze-dried to obtain the described Lactobacillus paracasei lysate (i.e., the Lactobacillus paracasei lysate with a molecular weight of 3-5kDa).

[0049] Example 4 Preparation Method of Probiotic Lysate Composition

[0050] Take the Lactobacillus rhamnosus lysate (1 g) with a molecular weight of 1-3 kDa, the Bifidobacterium lysate (12 g) with a molecular weight of 1-3 kDa, and the Lactobacillus paracasei lysate (5 g) with a molecular weight of 3-5 kDa prepared according to Example 1, Example 2, and Example 3, and mix them to obtain the probiotic lysate composition of the present invention.

[0051] Effect Example 1 Evaluation of in vitro anti-inflammatory activity of probiotic lysate composition

[0052] The experimental subjects in the following effect experimental examples are the preliminary lysate of Lactobacillus rhamnosus, the preliminary lysate of Bifidobacterium, the preliminary lysate of Lactobacillus paracasei, the lysate of Lactobacillus rhamnosus with a molecular weight of 1-3 kDa, the lysate of Bifidobacterium with a molecular weight of 1-3 kDa, the lysate of Lactobacillus paracasei with a molecular weight of 3-5 kDa, and the probiotic lysate composition prepared according to Example 4.

[0053] Hcoepic cells were collected at 5×10 3 / mL was inoculated in a Costa 96-well plate and cultured for 24 hours. Thereafter, it was incubated with a final concentration of 0.5% DSS for 12 hours. The cells were collected and washed twice with cold PBS. The Hcoepic cells were incubated with a preliminary lysate of Lactobacillus rhamnosus (1.0 mg / mL), a preliminary lysate of Bifidobacterium (1.0 mg / mL), a preliminary lysate of Lactobacillus paracasei (1.0 mg / mL), a lysate of Lactobacillus rhamnosus with a molecular weight of 1-3 kDa (1.0 mg / mL), a lysate of Bifidobacterium with a molecular weight of 1-3 kDa (1.0 mg / mL), a lysate of Lactobacillus paracasei with a molecular weight of 3-5 kDa (1.0 mg / mL) and a probiotic lysate composition (1.0 mg / mL) for 24 hours, and the cells and culture medium were collected. The CCK 8 method was used to detect cell viability, the ELISA kit was used to measure the SOD activity, MDA content and inflammatory cytokine content; the Western blotting method was used to detect changes in related protein expression.

[0054] The blank control group was marked as Control; the DSS group was marked as DSS; the preliminary lysate of Lactobacillus rhamnosus was marked as group A, the preliminary lysate of Bifidobacterium was marked as group B, the preliminary lysate of Lactobacillus paracasei was marked as group C, the lysate of Lactobacillus rhamnosus with a molecular weight of 1-3 kDa was marked as group D, the lysate of Bifidobacterium with a molecular weight of 1-3 kDa was marked as group E, the lysate of Lactobacillus paracasei with a molecular weight of 3-5 kDa was marked as group F, and the probiotic lysate composition was marked as group G.

[0055] The experimental results showed that ( Figure 1 A), the probiotic lysate composition of the present invention can effectively alleviate cell damage caused by DSS exposure and improve cell viability. Compared with the DSS group, the cell viability of the probiotic lysate composition increased by 17.4% after treatment (p<0.01); under the same concentration conditions, the probiotic lysate composition has a stronger cell protection activity against DSS-induced cells than a single probiotic preliminary lysate; under the same concentration conditions, the probiotic lysate composition has a stronger cell protection activity against DSS-induced cells than a Lactobacillus rhamnosus lysate with a molecular weight of 1-3kDa or a Bifidobacterium lysate with a molecular weight of 1-3kDa or a Lactobacillus paracasei lysate with a molecular weight of 3-5kDa when used alone. We believe that the combination of Lactobacillus rhamnosus lysate with a molecular weight of 1-3kDa, a Bifidobacterium lysate with a molecular weight of 1-3kDa and a Lactobacillus paracasei lysate with a molecular weight of 3-5kDa can improve the cell's resistance to damage through a synergistic effect.

[0056] DSS-induced oxidative damage to intestinal epithelial cells will further induce cell apoptosis and aggravate intestinal damage. Therefore, inhibiting DSS-induced oxidative damage to intestinal epithelial cells is of great significance for inhibiting intestinal inflammation and alleviating damage. The antioxidant system of cells includes enzymes and non-enzymatic antioxidants, such as SOD, CAT and GSH. In the present invention, DSS exposure significantly reduced the SOD activity of intestinal epithelial cells, while the secreted MDA was significantly increased. Compared with the blank control group, the SOD activity of the DSS-exposed group decreased by 46.8% (p<0.01), and the MDA content increased by 289.6% (p<0.01). After treatment with the probiotic lysate composition, the DSS-induced decrease in epithelial cell SOD activity and increase in MDA content were significantly inhibited. Compared with the DSS group, the SOD activity increased by 38.3% (p<0.01) and the MDA content decreased by 63.5% (p<0.01, Figure 1B, C). Analysis of the experimental results showed that under the same concentration conditions, the probiotic lysate composition had a stronger effect on the SOD activity and MDA content of DSS-induced intestinal epithelial cells than a single probiotic preliminary lysate; under the same concentration conditions, the probiotic lysate composition had a stronger effect on the SOD activity and MDA content of DSS-induced intestinal epithelial cells than the use of a 1-3kDa Lactobacillus rhamnosus lysate, a 1-3kDa Bifidobacterium lysate, or a 3-5kDa Lactobacillus paracasei lysate alone. We believe that the combination of a 1-3kDa Lactobacillus rhamnosus lysate, a 1-3kDa Bifidobacterium lysate, and a 3-5kDa Lactobacillus paracasei lysate can improve the antioxidant capacity of cells through a synergistic effect.

[0057] It is reported that the presence of cellular inflammation and oxidative damage will cause a vicious cycle and aggravate enteritis. DSS exposure induces an inflammatory response in intestinal epithelial cells and induces the release of cellular inflammatory factors. Therefore, inhibiting the inflammatory response induced by DSS can reduce inflammatory damage to cells. In this experiment, DSS exposure significantly increased the secretion of pro-inflammatory cytokines by intestinal epithelial cells. Compared with the blank control group, the contents of the DSS-exposed group (TNF-α, IL-1β, IL-6, IL-8, IL-13, IL-18) increased by 159.3% (p<0.01), 201.2% (p<0.01), 238.2% (p<0.01), 118.1% (p<0.01), 91.6% (p<0.01), 231.1% (p<0.01), respectively. Figure 2 A, B, C, D, E, F); In contrast, compared with the blank control group, DSS exposure reduced the levels of anti-inflammatory cytokines IL-4 and IL-10 by 34.6% (p < 0.01) and 29.6% (p < 0.01, Figure 2G, H). The above results show that DSS exposure significantly induced the secretion of a large number of inflammatory factors, while the secretion of anti-inflammatory factors was inhibited. After treatment with the probiotic lysate composition, the DSS-induced cellular inflammatory factor secretion disorder was effectively inhibited. Compared with the DSS group, the TNF-α, IL-1β, IL-6, IL-8, IL-13, and IL-18 levels in the probiotic lysate composition group decreased by 48.6% (p<0.01), 55.4% (p<0.01), 59.5% (p<0.01), 13.1% (p<0.05), 24.6% (p<0.01), and 53.1% (p<0.01); compared with the DSS-exposed group, the IL-4 and IL-10 levels in the probiotic lysate composition group increased by 41.6% (p<0.01) and 8.21%. Further analysis of the experimental results revealed that, under the same concentration conditions, the probiotic lysate combination had a stronger activity in regulating the secretion of inflammatory factors in intestinal epithelial cells induced by DSS than a single probiotic preliminary lysate; under the same concentration conditions, the probiotic lysate combination had a stronger activity in regulating the secretion of inflammatory factors in intestinal epithelial cells induced by DSS than the use of a 1-3kDa Lactobacillus rhamnosus lysate, a 1-3kDa Bifidobacterium lysate, or a 3-5kDa Lactobacillus paracasei lysate alone. We believe that the combination of a 1-3kDa Lactobacillus rhamnosus lysate, a 1-3kDa Bifidobacterium lysate, and a 3-5kDa Lactobacillus paracasei lysate can synergistically regulate the secretion of inflammatory factors in intestinal epithelial cells, thereby exerting a more excellent anti-inflammatory activity.

[0058] Inflammation is a defensive response of the body involving multiple cells and cytokines, but severe or long-term inflammation can cause damage to the body. Nuclear factor Kappa B (NF-κB) is closely related to the inflammatory response. In the present invention, DSS exposure significantly increased the phosphorylation of IKKα, IKKβ and IκBα proteins in intestinal epithelial cells, while the content of IKKα, IKKβ and IκBα proteins was significantly reduced, indicating that DSS exposure activated the NF-κB signaling pathway in intestinal epithelial cells. In the probiotic lysate composition group, the phosphorylation of IKKα, IKKβ and IκBα proteins in intestinal epithelial cells induced by DSS exposure was effectively inhibited ( Figure 3A). Experimental results suggest that the probiotic lysate combination can mitigate DSS-induced inflammatory damage by inhibiting DSS-induced activation of the NF-κB signaling pathway in intestinal epithelial cells. Similarly, analysis of experimental results revealed that, at the same concentration, the probiotic lysate combination exhibited greater inhibitory activity against DSS-induced activation of the NF-κB signaling pathway in intestinal epithelial cells than a single probiotic primary lysate. Furthermore, at the same concentration, the probiotic lysate combination exhibited greater inhibitory activity against DSS-induced activation of the NF-κB signaling pathway in intestinal epithelial cells than either a 1-3 kDa Lactobacillus rhamnosus lysate, a 1-3 kDa Bifidobacterium lysate, or a 3-5 kDa Lactobacillus paracasei lysate alone. We believe that the combination of a 1-3 kDa Lactobacillus rhamnosus lysate, a 1-3 kDa Bifidobacterium lysate, and a 3-5 kDa Lactobacillus paracasei lysate may synergistically inhibit NF-κB signaling pathway activation, thereby exerting enhanced anti-inflammatory activity.

[0059] Effect Example 2 Evaluation of the in vivo anti-inflammatory activity of probiotic lysate composition

[0060] The experimental subjects in the following effect experimental examples are the preliminary lysate of Lactobacillus rhamnosus, the preliminary lysate of Bifidobacterium, the preliminary lysate of Lactobacillus paracasei, the lysate of Lactobacillus rhamnosus with a molecular weight of 1-3 kDa, the lysate of Bifidobacterium with a molecular weight of 1-3 kDa, the lysate of Lactobacillus paracasei with a molecular weight of 3-5 kDa, and the probiotic lysate composition prepared according to Example 4.

[0061] C57BL / 6 mice were randomly divided into Group A, 8 mice per group, and the following groups: blank control (Control), DSS model group (DSS), Lactobacillus rhamnosus preliminary lysate group, Bifidobacterium preliminary lysate group, Lactobacillus paracasei preliminary lysate group, Lactobacillus rhamnosus lysate group with a molecular weight of 1-3 kDa, Bifidobacterium lysate group with a molecular weight of 1-3 kDa, Lactobacillus paracasei lysate group with a molecular weight of 3-5 kDa, and a probiotic lysate combination group. Except for the control group, DSS powder was added to the drinking water of the remaining groups to prepare a DSS solution with a mass concentration of 30 g / L, which was freely consumed by the mice for 7 consecutive days to establish an enteritis model (mice in the DSS group continued to drink the DSS solution until the end of the experiment). After the modeling was completed, mice were gavaged and administered the drug once a day starting on the 8th day for 14 consecutive days. The preliminary lysate of Lactobacillus rhamnosus, the preliminary lysate of Bifidobacterium, the preliminary lysate of Lactobacillus paracasei, the lysate of Lactobacillus rhamnosus with a molecular weight of 1-3 kDa, the lysate of Bifidobacterium with a molecular weight of 1-3 kDa, the lysate of Lactobacillus paracasei with a molecular weight of 3-5 kDa and the probiotic lysate composition were dissolved in water at a concentration of 50 mg / mL. 0.2 mL / 10 g was administered by gavage each time, and the dosage was calculated according to the weight of the mice. The blank control group was marked as Control; the DSS group was marked as DSS; the preliminary lysate of Lactobacillus rhamnosus was marked as group a, the preliminary lysate of Bifidobacterium was marked as group b, the preliminary lysate of Lactobacillus paracasei was marked as group c, the lysate of Lactobacillus rhamnosus with a molecular weight of 1-3 kDa was marked as group d, the lysate of Bifidobacterium with a molecular weight of 1-3 kDa was marked as group e, the lysate of Lactobacillus paracasei with a molecular weight of 3-5 kDa was marked as group f, and the probiotic lysate composition was marked as group g.

[0062] After the experiment, blood was collected from the eyeballs and centrifuged, and the inflammatory factor content was detected according to the instructions of the ELISA kit; the mice were killed by dislocation, and the colon was completely removed from the cecum to the anus and the length was measured.

[0063] During the experiment, mice in the DSS group developed diarrhea and bloody stools, while mice in the probiotic lysate composition-treated group did not experience diarrhea or bloody stools. The experimental results showed that the DSS group had reduced activity and disordered hair. At the end of the experiment, the mice had significantly decreased in weight. However, in the probiotic lysate composition-treated group, DSS-induced weight loss was effectively suppressed. At the end of the experiment, there was no significant difference in weight compared with the blank group ( Figure 4 A). The DAI score of the probiotic lysate composition treatment group was significantly lower than that of the DSS group ( Figure 4 B). Furthermore, the colon length of mice was compared. The colon of mice in the DSS group became thinner and shorter. After treatment with the probiotic lysate composition, the thinning and shortening of the colon of mice caused by DSS exposure was effectively alleviated ( Figure 4C). We further found that, under the same concentration conditions, the probiotic lysate combination had stronger therapeutic activity against DSS-induced diarrhea, hematochezia, increased DAI scores, and colon thinning and shortening in mice than a single probiotic primary lysate, and stronger than the use of a 1-3 kDa Lactobacillus rhamnosus lysate, a 1-3 kDa Bifidobacterium lysate, or a 3-5 kDa Lactobacillus paracasei lysate alone. We believe that the combination of a 1-3 kDa Lactobacillus rhamnosus lysate, a 1-3 kDa Bifidobacterium lysate, and a 3-5 kDa Lactobacillus paracasei lysate can synergistically improve the mice's resistance to damage and reduce DSS-induced intestinal damage in mice.

[0064] DSS exposure can induce an inflammatory response in the intestinal epithelial cells of mice and aggravate inflammatory damage. In this experiment, DSS stimulation significantly increased the secretion of inflammatory factors in mice. Compared with the blank control group, the levels of TNF-α, IL-1β, IL-6, IL-13, and IL-18 in the DSS exposure group increased by 176.2% (p<0.01), 89.5% (p<0.01), 154.2% (p<0.01), 116.6% (p<0.01), and 201.2% (p<0.01). Figure 5 A, B, C, D, E); In contrast, the IL-4 content in the serum of mice exposed to DSS decreased by 45.6% (p < 0.01, Figure 5 F); It shows that DSS exposure significantly induced intestinal inflammation in mice. After treatment with the probiotic lysate composition, the imbalance of inflammatory factor secretion induced by DSS was effectively alleviated. Compared with the DSS group, the TNF-α, IL-1β, IL-6, IL-13, and IL-18 levels in the probiotic lysate composition group decreased by 35.9% (p<0.01), 28.6% (p<0.01), 45.3% (p<0.01), 38.9% (p<0.01), and 54.8% (p<0.01), respectively; the IL-4 content increased by 49.1% (p<0.01). Similarly, under the same concentration conditions, the probiotic lysate composition has a stronger secretion-regulating activity on DSS-induced inflammatory factors than the single probiotic preliminary lysate used alone, and is stronger than the use of a 1-3kDa Lactobacillus rhamnosus lysate or a 1-3kDa Bifidobacterium lysate or a 3-5kDa Lactobacillus paracasei lysate alone. The animal experimental results in this part once again prove that the combination of a 1-3kDa Lactobacillus rhamnosus lysate, a 1-3kDa Bifidobacterium lysate, and a 3-5kDa Lactobacillus paracasei lysate has a synergistic effect, can effectively regulate the secretion of inflammatory factors in intestinal epithelial cells, and exert anti-enteritis activity.

[0065] The above disclosure is only a preferred embodiment of the present invention, and it is certainly not intended to limit the scope of the present invention. A person skilled in the art can understand that all or part of the processes of the above embodiment and equivalent changes made in accordance with the claims of the present invention are still within the scope of the invention.

Claims

1. A probiotic lysate composition, characterized in that: The probiotic lysate composition comprises Lactobacillus rhamnosus lysate, Bifidobacterium lysate and Lactobacillus paracasei lysate; The Lactobacillus rhamnosus lysate is prepared by the following method: S11. Lactobacillus rhamnosus was added to PBS buffer and ultrasonically lysed to obtain a preliminary lysate of Lactobacillus rhamnosus; S12. The initial lysate of Lactobacillus rhamnosus was first subjected to ultrafiltration and dialysis using a 1 kDa ultrafiltration membrane; the retentate after ultrafiltration and dialysis using a 1 kDa ultrafiltration membrane was then subjected to ultrafiltration and dialysis using a 3 kDa ultrafiltration membrane, and the ultrafiltrate after ultrafiltration and dialysis using a 3 kDa ultrafiltration membrane was lyophilized to obtain the Lactobacillus rhamnosus lysate; The bifidobacterium lysate is prepared by the following method: S21. Bifidobacterium was added to PBS buffer and ultrasonically lysed to obtain a preliminary lysate of bifidobacteria; S22. The preliminary lysate of bifidobacteria was first subjected to ultrafiltration and dialysis using a 1kDa ultrafiltration membrane; the retentate after ultrafiltration and dialysis using a 1kDa ultrafiltration membrane was then subjected to ultrafiltration and dialysis using a 3kDa ultrafiltration membrane, and the ultrafiltrate after ultrafiltration and dialysis using a 3kDa ultrafiltration membrane was lyophilized to obtain the bifidobacterium lysate; The Lactobacillus paracasei lysate is prepared by the following method: S31. Lactobacillus paracasei was added to PBS buffer and ultrasonically lysed to obtain a preliminary lysate of Lactobacillus paracasei; S32. The preliminary lysate of Lactobacillus paracasei is first subjected to ultrafiltration and dialysis using a 1 kDa ultrafiltration membrane; then the retentate after ultrafiltration and dialysis using a 1 kDa ultrafiltration membrane is subjected to ultrafiltration and dialysis using a 3 kDa ultrafiltration membrane; then the retentate after ultrafiltration and dialysis using a 3 kDa ultrafiltration membrane is subjected to ultrafiltration and dialysis using a 5 kDa ultrafiltration membrane, and the ultrafiltrate after ultrafiltration and dialysis using a 5 kDa ultrafiltration membrane is freeze-dried to obtain the Lactobacillus paracasei lysate.

2. The probiotic lysate composition according to claim 1, characterized in that The mass ratio of the Lactobacillus rhamnosus lysate, the Bifidobacterium lysate and the Lactobacillus paracasei lysate is 1:1-100:1-100.

3. The probiotic lysate composition according to claim 1, characterized in that The mass ratio of the Lactobacillus rhamnosus lysate, the Bifidobacterium lysate and the Lactobacillus paracasei lysate is 1:1-25:1-25.

4. The probiotic lysate composition according to claim 1, characterized in that The mass dosage of PBS buffer is 4 to 6 times that of Lactobacillus rhamnosus; The PBS buffer solution is a PBS buffer solution with a pH of 6.5; and the ultrasonic lysis process has an ultrasonic time of 10 to 20 minutes.

5. The probiotic lysate composition according to claim 1, characterized in that The mass dosage of PBS buffer is 4 to 6 times that of Bifidobacterium; The PBS buffer solution is a PBS buffer solution with a pH of 6.5; and the ultrasonic lysis process has an ultrasonic time of 10 to 20 minutes.

6. The probiotic lysate composition according to claim 1, characterized in that The mass dosage of PBS buffer is 4 to 6 times that of Lactobacillus paracasei; The PBS buffer solution is a PBS buffer solution with a pH of 6.5; and the ultrasonic lysis process has an ultrasonic time of 10 to 20 minutes.

7. Use of the probiotic lysate composition according to any one of claims 1 to 6 in the preparation of a product with anti-enteritis effect.

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