Pharmaceutical compositions comprising osmanthus glycoside h and osmanthin a and their use for the treatment or prevention of inflammatory bowel disease

Abstract

This invention relates to a pharmaceutical composition comprising osmanthin H and scutellarin, and its use in treating or preventing inflammatory bowel disease, belonging to the pharmaceutical field. This invention establishes a rat model of inflammatory bowel disease to evaluate the therapeutic effects of osmanthin H and scutellarin. The results show that osmanthin H and scutellarin can inhibit pro-inflammatory factors (TNF-α, IL-1β, IL-6), upregulate anti-inflammatory factors (IL-10), and regulate LPS and Lcn2 levels in vivo, restoring intestinal immune balance and achieving an effective therapeutic effect on inflammatory bowel disease. In vivo pharmacodynamic tests fully confirm the efficacy of the composition of this invention in treating inflammatory bowel disease.

Description

Technical Field

[0001] This invention relates to pharmaceutical compositions comprising cinnamyl glycoside H and scutellarin, and their use in treating or preventing inflammatory bowel disease, belonging to the pharmaceutical field. Background Technology

[0002] Inflammatory bowel disease (IBD) is a group of chronic inflammatory bowel diseases with unknown etiologies, which are related to genetic susceptibility, intestinal immune disorders, dysbiosis, and environmental factors. It mainly includes ulcerative colitis (UC) and Crohn's disease (CD). Clinically, it is necessary to rule out other factors that cause intestinal inflammation, such as infectious enteritis and intestinal malignancies, and make a diagnosis based on characteristic lesions such as chronic inflammation and ulceration of the intestinal mucosa. UC mainly affects the colorectal region, with continuous superficial ulcers and mucus-containing bloody stools as typical manifestations. CD can affect the entire gastrointestinal tract, but is more common in the terminal small intestine and colon, with discontinuous deep mucosal ulcers and cobblestone-like changes as typical manifestations. Long-term recurrent attacks can easily progress to intestinal stricture, perforation, and even colorectal cancer. It is worth noting that IBD is not a disease limited to the intestines, but a multisystem disease. Approximately 25% to 40% of patients will experience extraintestinal manifestations, which can affect multiple extraintestinal organs and cause chronic complications. It not only significantly increases the risk of autoimmune diseases and thromboembolic diseases, but is also associated with various diseases such as chronic kidney disease, fatty liver, anemia, arthritis, and psoriasis. At the same time, it can increase the risk of colorectal cancer. Its mechanism may be related to chronic intestinal inflammation, mucosal barrier damage, and excessive release of pro-inflammatory cytokines, which seriously affect the patient's quality of life and prognosis.

[0003] Given the increasing prevalence of IBD in recent decades, and considering that, as mentioned above, there is currently no cure for IBD besides lifestyle modifications and supportive care, existing drug treatments suffer from significant side effects, large individual differences in efficacy, and ineffectiveness in some patients, while surgical treatment has a high recurrence rate and numerous complications, the rapid increase in IBD incidence and its severe impact on patients' quality of life and the social healthcare burden necessitates new and effective prevention and treatment strategies. This includes in-depth research into the pathogenesis of IBD, identifying new therapeutic targets, developing novel therapeutic drugs with proven efficacy and fewer side effects, and standardizing treatment protocols to improve treatment outcomes for IBD patients, reduce disease recurrence and complications, improve patients' quality of life, and alleviate the social healthcare burden. Summary of the Invention

[0004] The purpose of this invention is to provide a pharmaceutical composition containing osmanthin H and scutellarin, and its use in treating or preventing inflammatory bowel disease (IBD), addressing the problems of limited efficacy and significant side effects of existing therapeutic drugs. This invention selects two natural active ingredients, osmanthin H and scutellarin, as research subjects, systematically investigating the intervention effects of monomers and compound compositions on IBD, aiming to achieve simultaneous improvement of intestinal inflammation and barrier function through synergistic effects, providing a safe, efficient, and multi-targeted new natural drug solution for IBD treatment.

[0005] To achieve the above objectives, the first aspect of the present invention adopts the following technical solution: the use of osmanthus glycoside H in the preparation of a medicament for treating or preventing inflammatory bowel disease.

[0006] Among them, Osmanthuside H belongs to the class of phenylethanol glycosides and has the molecular formula C. 19 H 28 O 11 CAS No.: 149155-70-4, the structural formula of cinnamyl glycoside H is:

[0007] .

[0008] A second aspect of the invention discloses the use of scutellarin in the preparation of medicaments for the treatment or prevention of inflammatory bowel disease.

[0009] Bracteatin, a flavonoid natural compound, has the molecular formula C2. 15 H 10 O7, CAS No.: 3260-50-2; the structural formula of bractin is:

[0010] .

[0011] A third aspect of the invention provides a pharmaceutical composition comprising osmanthin H and scutellarin (as an active ingredient). Preferably, the active ingredient is a combination of osmanthin H and scutellarin.

[0012] In a preferred embodiment, the pharmaceutical composition further includes a pharmaceutically acceptable excipient.

[0013] In the pharmaceutical composition, the ratio of the dose of cinnamyl glycoside H to the dose of scutellarin is (1-10):1, preferably a mass ratio of 4:1.

[0014] This invention has discovered that when osmanthus glycoside H and stigmacin are used together, they can exert a synergistic effect. The two work at different levels, such as inhibiting inflammation, regulating intestinal flora, and protecting the intestinal mucosal barrier. They improve the pathological damage of inflammatory bowel disease through multiple targets and pathways, and their effects are significantly better than those of using a single component alone.

[0015] The dosage of the drug is as follows: the dosage of cinnamyl glycoside H is 1-40 mg / kg / day, preferably 10-30 mg / kg / day, more preferably 20 mg / kg / day; the dosage of stigmacin is 1-40 mg / kg / day, preferably 10-30 mg / kg / day, more preferably 20 mg / kg / day; the total dosage of the two combined is 5-50 mg / kg / day, preferably 10-30 mg / kg / day, more preferably 20 mg / kg / day.

[0016] In some feasible implementations, the pharmaceutical composition can be formulated into conventional dosage forms such as injections, capsules, granules, and tablets, or into sustained-release formulations or enteric formulations (such as enemas and rectal suppositories); it can be prepared using methods known in the art, combining the active ingredient with excipients.

[0017] In some feasible implementations, the pharmaceutical composition is administered via one or more of the following methods: oral, injection, implantation, spray, and / or enteric administration.

[0018] In a fourth aspect, the use of the aforementioned pharmaceutical composition in the preparation of a medicament for treating or preventing inflammatory bowel disease is disclosed.

[0019] A fifth aspect of the invention provides a method for treating or preventing inflammatory bowel disease, the method comprising administering, via enema, a therapeutically effective amount of cinnamyl glycoside H, scutellarin, a combination thereof, or a pharmaceutical composition comprising the composition to a subject.

[0020] The advantages of this invention are as follows: This invention establishes a rat model of inflammatory bowel disease to evaluate the therapeutic effects of osmanthin H and scutellarin, showing that osmanthin H and scutellarin can inhibit pro-inflammatory factors (TNF-α, IL-1β, IL-6), upregulate anti-inflammatory factors (IL-10), and regulate LPS and Lcn2 levels in vivo, thereby restoring intestinal immune balance and achieving an effective therapeutic effect on inflammatory bowel disease. In vivo pharmacodynamic tests fully confirm the efficacy of the composition of this invention in treating inflammatory bowel disease.

[0021] Furthermore, this invention provides a monomeric compound composition composed of osmanthin H and scutellarin, and investigates the therapeutic effect of the monomeric compound composition on inflammatory bowel disease. It proves that the overall therapeutic effect of the monomeric compound composition of this invention is better than that of osmanthin H and scutellarin administered alone. That is, osmanthin H, scutellarin and their monomeric compound composition all have good pharmacological effects in the treatment of inflammatory bowel disease, and the monomeric compound composition has a better improvement effect. It is expected that the above-mentioned active ingredients can be developed into new drugs for improving and treating inflammatory bowel disease. Attached Figure Description

[0022] Figure 1The effect of drug intervention on body weight in mice with inflammatory bowel disease;

[0023] Figure 2 The effect of drug intervention on the DAI score of mice with inflammatory bowel disease;

[0024] Figure 3 The effect of drug intervention on colon length in mice with inflammatory bowel disease;

[0025] Figure 4 Effects of drug intervention on serum levels of the pro-inflammatory factor TNF-α in mice with inflammatory bowel disease;

[0026] Figure 5 Effects of drug intervention on serum levels of the pro-inflammatory factor IL-1β in mice with inflammatory bowel disease;

[0027] Figure 6 Effects of drug intervention on serum levels of the pro-inflammatory factor IL-6 in mice with inflammatory bowel disease;

[0028] Figure 7 Effects of drug intervention on serum IL-10 levels in mice with inflammatory bowel disease;

[0029] Figure 8 Effects of drug intervention on serum LPS levels in mice with inflammatory bowel disease;

[0030] Figure 9 Effects of drug intervention on serum Lcn2 levels in mice with inflammatory bowel disease. Detailed Implementation

[0031] To make the objectives, technical solutions, and advantages of this invention clearer, the technical solutions of this invention will be clearly and completely described below in conjunction with embodiments. Obviously, the described embodiments are only some, not all, of the embodiments of this invention. All other embodiments obtained by those skilled in the art based on the embodiments of this invention without creative effort are within the scope of protection of this invention.

[0032] Furthermore, to better illustrate the present invention, numerous specific details are provided in the following detailed embodiments. Those skilled in the art should understand that the present invention can be practiced without certain specific details. In some embodiments, materials, elements, methods, and means well known to those skilled in the art are not described in detail in order to highlight the spirit of the invention.

[0033] The present invention will be described below through exemplary embodiments.

[0034] Example 1

[0035] This embodiment is a composition for treating inflammatory bowel disease, which comprises the following components in parts by weight:

[0036] 12 parts of osmanthus glycoside H and 6 parts of stigmacin.

[0037] Example 2

[0038] This embodiment is a composition for treating inflammatory bowel disease, which comprises the following components in parts by weight:

[0039] 12 parts of osmanthus glycoside H1 and 2 parts of stigmacin.

[0040] Example 3

[0041] This embodiment is a composition for treating inflammatory bowel disease, which comprises the following components in parts by weight:

[0042] Six parts of osmanthus glycoside H1 and four parts of stigmacin.

[0043] Experimental Example 1

[0044] In this experiment, SPF-grade C57BL / 6J male mice were purchased from Beijing Vital River Laboratory Animal Technology Co., Ltd. Mesalazine is a first-line drug for the clinical treatment of mild to moderate ulcerative colitis. It can reduce the release of pro-inflammatory factors and protect the intestinal mucosal barrier, and is currently the mainstream positive control drug for the treatment of inflammatory bowel disease.

[0045] 1. Grouping and administration of experimental animals

[0046] In this experiment, 7-8 week old male C57BL / 6J mice were randomly divided into a blank control group and a model group. After 5 days of acclimatization, the model group was administered a combination of antibiotics (vancomycin: 50 mg / kg, neomycin sulfate: 100 mg / kg, metronidazole: 100 mg / kg, ampicillin: 100 mg / kg) by gavage for 5 days. Following this, they were administered a humanized bacterial suspension from patients with inflammatory bowel disease by gavage for 14 days (5 × 10⁻⁶ mcg). 8 A mouse model of inflammatory bowel disease (IBD) was established by administering 2.5% DSS to the mice freely for 5 days. On day 5 after modeling, three mice were randomly selected from both the normal group and the model group and sacrificed. The colonic tissue structure, inflammation, and ulceration were observed under a microscope. The success of the model was determined based on the macroscopic signs and the pathological changes in the colonic mucosa observed under a microscope.

[0047] In this experiment, mice that successfully developed the model were randomly divided into the following groups: model control group, mesalazine positive drug control group (mesalazine enema solution, specification: 60g:4g), osmanthus glycoside H group, scutellarin group, and osmanthus glycoside H and scutellarin combined group, with 15 mice in each group.

[0048] After successful model replication, the treatment groups were administered the corresponding doses via whole colonic enema. The control group and the model group were simultaneously given an equal volume of sterile saline via enema, a single intervention, for 14 consecutive days, with normal diet maintained during the treatment period. The dosages are shown in Table 1.

[0049] Table 1. Grouping and Dosage of Experimental Animals

[0050]

[0051] 2. Phenotypic evaluation

[0052] Mice were monitored daily for body weight, fecal condition, and fecal blood loss, and the Disease Activity Index (DAI) score was calculated to preliminarily assess the therapeutic effect of the drug on inflammatory bowel disease. One hour after the last administration, mice in each group were weighed, blood was collected by enucleation, colon tissue was dissected, and the colon length of each group was measured.

[0053] After drug administration, blood was collected from the eyeballs of mice, and whole blood was collected from each group of mice. The blood was centrifuged at 3000 rpm for 15 min at 4 ℃, and the supernatant was transferred to a new centrifuge tube and stored at -80 ℃. The serum levels of TNF-α, IL-1β, IL-6, IL-10, LPS, and Lcn2 were detected using an enzyme-linked immunosorbent assay (ELISA) kit, strictly following the kit instructions.

[0054] 3. Comparative Example

[0055] Table 2 Comparison of experimental animal groups and drug dosages

[0056]

[0057] In Table 2, Group 1 is Comparative Example 1, Group 2 is Comparative Example 2, and Group 3 is Comparative Example 3.

[0058] The results of the comparative experiments were determined according to 1. Animal grouping and administration, and 2. Phenotypic evaluation.

[0059] 4. Experimental Results

[0060] Table 1 shows the various indicators of each group of mice as follows: Figure 1-9 And as described in 4.1-4.6 below.

[0061] 4.1 Effects of drug intervention on body weight in mice with inflammatory bowel disease

[0062] Figure 1Table 1 shows the changes in body weight of mice in each group at the end of the experiment. Compared with the blank control group, the body weight of mice in the model group decreased significantly, indicating that intestinal inflammation severely affected the digestive and absorptive functions of mice, leading to a significant decrease in body weight. Compared with the model group, the body weight of mice in each treatment group was increased to varying degrees (p<0.01), with the osmanthin H group, scutellarin group, and monomeric compound group showing increases of 10.09%, 8.50%, and 15.21%, respectively, demonstrating significant body weight improvement. Comparing the two monomeric treatment groups, the monomeric compound group showed a more prominent effect at the same dosage, with its body weight recovering to near the level of the blank control group. This indicates that osmanthin H and scutellarin can effectively improve intestinal dysfunction and promote body weight recovery in mice, with the monomeric compound group showing better results.

[0063] 4.2 Effects of drug intervention on DAI score in mice with inflammatory bowel disease

[0064] The control group mice had formed, uniform stools with clean, uncontaminated perianal area. The model group mice exhibited significantly loose, bloody stools, perianal soiling, and stools adhering to hair, indicating a poor overall condition. All treatment groups showed a significant improvement trend compared to the model group. The DAI (Disseminated Intake Index) was calculated by comprehensively evaluating three factors: weight change rate, stool characteristics, and bloody stools, providing an objective assessment of the severity of colitis. The DAI score of the model group mice was significantly higher than that of the control group (p<0.01). Compared to the model group, the DAI scores of all treatment groups were significantly lower (p<0.01), with the osmanthus glycoside H group, scutellarin group, and single-component combination group showing reductions of 59.68%, 56.45%, and 76.61%, respectively. Therefore, the single-component combination group showed better improvement than the osmanthus glycoside H or scutellarin treatment groups alone, indicating a more ideal comprehensive intervention effect. Specific results are as follows: Figure 2 As shown.

[0065] 4.3 Effects of drug intervention on colon length in mice with inflammatory bowel disease

[0066] Chronic intestinal inflammation can lead to intestinal mucosal damage, tissue fibrosis, and abnormal intestinal wall structure, ultimately resulting in colon shortening. Therefore, colon length can directly reflect the degree of intestinal inflammation and damage. Compared with the blank control group, the colon length in the model group was significantly shortened (p<0.01); compared with the model group, the colon length in each treatment group was significantly improved (p<0.01), with improvements of 34.83%, 33.87%, and 42.58% in the osmanthus glycoside H group, the scutellarin group, and the monomer-compound group, respectively. Comparing the two monomer-administered groups, the monomer-compound group showed the most significant effect on colon length recovery, suggesting that osmanthus glycoside H and scutellarin have good effects in inhibiting intestinal inflammation and protecting intestinal tissue, with the monomer-compound group showing a more prominent effect. (Specific details are as follows...) Figure 3 As shown.

[0067] 4.4 Effects of drug intervention on serum pro-inflammatory factor levels in mice with inflammatory bowel disease

[0068] The anti-inflammatory effects of the drugs were evaluated by detecting serum levels of TNF-α, IL-1β, and IL-6. Compared with the blank control group, the levels of the above-mentioned pro-inflammatory factors in the model group mice were significantly increased (p<0.01); compared with the model group, the levels of pro-inflammatory factors in each treatment group were significantly decreased (p<0.01). For TNF-α, the levels in the osmanthus glycoside H group, scutellarin group, and monomeric compound group decreased by 44.51%, 30.58%, and 55.21%, respectively; for IL-1β, the levels in the osmanthus glycoside H group, scutellarin group, and monomeric compound group decreased by 59.78%, 52.96%, and 67.78%, respectively; and for IL-6, the levels in the osmanthus glycoside H group, scutellarin group, and monomeric compound group decreased by 65.75%, 61.93%, and 74.68%, respectively. Therefore, the monomeric compound group showed a better downregulation effect on pro-inflammatory factors than the osmanthus glycoside H and scutellarin monomeric treatment groups. This suggests that osmanthus glycoside H, scutellarin, and monomeric compound groups can all exert strong anti-inflammatory activity. Specific results are as follows Figure 4 , Figure 5 , Figure 6 As shown.

[0069] 4.5 Effects of drug intervention on serum IL-10 levels in mice with inflammatory bowel disease

[0070] The regulatory effect of drugs on immune balance was assessed by detecting serum anti-inflammatory factor IL-10 levels. Compared with the blank control group, the IL-10 level in the model group was significantly reduced (p<0.01). Compared with the model group, the IL-10 levels in each treatment group were upregulated to varying degrees, with the osmanthus glycoside H group, scutellarin group, and monomeric compound group showing upregulations of 34.86%, 4.50%, and 97.60%, respectively. The monomeric compound group showed a more significant enhancement effect than the osmanthus glycoside H and scutellarin monomeric treatment groups. This indicates that osmanthus glycoside H, scutellarin, and the monomeric compound group can all effectively regulate immune imbalance and enhance the body's anti-inflammatory capacity. Specifically... Figure 7 As shown.

[0071] 4.6 Effects of drug intervention on serum LPS and Lcn2 levels in mice with inflammatory bowel disease

[0072] Serum LPS and Lcn2 levels reflect intestinal barrier integrity, bacterial endotoxin translocation, and intestinal inflammatory activity. Compared with the blank control group, the LPS and Lcn2 levels in the model group were significantly increased (P<0.01). Compared with the model group, all treatment groups reduced LPS and Lcn2 levels to varying degrees. For LPS, the reductions in the osmanthus glycoside H group, scutellarin group, and monomeric compound group were 41.09%, 37.42%, and 56.89%, respectively; for Lcn2, the reductions in the osmanthus glycoside H group, scutellarin group, and monomeric compound group were 40.68%, 40.40%, and 53.31%, respectively. This indicates that the model mice have significant intestinal barrier damage and persistent inflammatory response. The monomeric compound group showed the best effect, with index levels approaching those of the blank control group. This indicates that osmanthus glycoside H, scutellarin, and monomeric compound groups can effectively repair the intestinal barrier, reduce bacterial translocation, and alleviate intestinal inflammation. (Details follow...) Figure 8 , Figure 9 As shown.

[0073] Figure 1-9 In comparison with the blank control group, * p<0.05, ** p<0.01, *** p < 0.001; compared with the model group, # p<0.05, ## p<0.01, ### p<0.001.

[0074] The results of the comparative experiment are shown below.

[0075] Compared with the model group, the body weight of inflammatory bowel disease mice in Comparative Examples 1, 2, and 3 was increased by 12.64%, 11.58%, and 11.38%, respectively; the DAI scores of Comparative Examples 1, 2, and 3 were decreased by 68.02%, 62.73%, and 60.05%, respectively; the colon length of Comparative Examples 1, 2, and 3 was increased by 38.95%, 35.32%, and 34.73%, respectively; the TNF-α content of inflammatory bowel disease mice in Comparative Examples 1, 2, and 3 was decreased by 49.12%, 44.65%, and 40.12%, respectively; and the IL-1β content of inflammatory bowel disease mice in Comparative Examples 1, 2, and 3 was decreased by 12.64%, 11.58%, and 11.38%, respectively. The levels of IL-6 in mice with inflammatory bowel disease (IBD) decreased by 62.09%, 60.56%, and 57.15%, respectively. The levels of IL-6 in mice with IBD in Comparative Examples 1, 2, and 3 decreased by 68.16%, 65.80%, and 63.44%, respectively. The levels of IL-10 in mice with IBD in Comparative Examples 1, 2, and 3 increased by 80.40%, 51.79%, and 40.52%, respectively. The levels of LPS in mice with IBD in Comparative Examples 1, 2, and 3 decreased by 50.39%, 43.74%, and 40.12%, respectively. The levels of Lcn2 in mice with IBD in Comparative Examples 1, 2, and 3 decreased by 47.10%, 42.24%, and 42.08%, respectively.

[0076] Comparative Example 1 shows that replacing osmanthin H with a component similar to the parent nucleus and possessing anti-inflammatory activity affects its therapeutic effect in synergistically with scutellarin in inflammatory bowel disease. Comparative Examples 2 and 3 demonstrate that the mass ratio of osmanthin H to scutellarin is more important for promoting the recovery of inflammatory bowel disease indicators in mice.

[0077] 5. Overall Conclusion

[0078] This invention uses a DSS-induced inflammatory bowel disease model in C57BL / 6J mice to evaluate the intervention effects of osmanthus glycoside H, scutellarin, and their individual and combined administrations at multiple levels, including overall phenotype, immune inflammation level, and intestinal barrier function. Results showed that osmanthus glycoside H, scutellarin, and the single-drug combination group all improved the phenotypes of decreased body weight, increased DAI score, and shortened colon in the model mice to varying degrees; they significantly reduced serum levels of pro-inflammatory factors TNF-α, IL-1β, and IL-6, and increased the content of anti-inflammatory factor IL-10; simultaneously, they reduced LPS and Lcn2 levels, improving intestinal barrier function impairment. The single-drug combination group showed the best effect.

[0079] Overall, the combined use of cinnamyl glycoside H and stigmacin has a synergistic effect, and its improvement effect on inflammatory bowel disease is significantly better than that of a single monomer, showing good therapeutic potential and application prospects.

[0080] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, and not to limit them; although the present invention has been described in detail with reference to the foregoing embodiments, those skilled in the art should understand that modifications can still be made to the technical solutions described in the foregoing embodiments, or equivalent substitutions can be made to some of the technical features; and these modifications or substitutions do not cause the essence of the corresponding technical solutions to deviate from the spirit and scope of the technical solutions of the embodiments of the present invention.

Claims

1. Use of cinnamyl glycoside H in the preparation of drugs for the treatment or prevention of inflammatory bowel disease.

2. The use according to claim 1, characterized in that, The structural formula of cinnamyl glycoside H is: 。 3. Use of scutellarin in the preparation of drugs for the treatment or prevention of inflammatory bowel disease.

4. The use according to claim 3, characterized in that, The structural formula of sphagnum motif is: 。 5. A pharmaceutical composition, characterized in that, Including cinnamyl glycoside H and stigmacin.

6. The pharmaceutical composition according to claim 5, characterized in that, The pharmaceutical composition also includes pharmaceutically acceptable excipients.

7. The pharmaceutical composition according to claim 5, characterized in that, In the pharmaceutical composition, the ratio of the dose calculated as osmanthin H to the dose calculated as stigmacin is (1-10):

1.

8. The pharmaceutical composition according to claim 5, characterized in that, The pharmaceutical composition can be formulated as an injection, capsule, granule, tablet, sustained-release formulation, or enteric formulation.

9. A pharmaceutical composition according to claim 5, characterized in that, The pharmaceutical composition may be administered via one or more of the following methods: oral, injection, implantation, spray, and / or enteric administration.

10. Use of the pharmaceutical composition according to any one of claims 5-9 in the preparation of a medicament for treating or preventing inflammatory bowel disease.

Images