CN112107586A - Application of pulsatilla saponin B4 in preparation of medicine for treating ulcerative colitis - Google Patents

Application of pulsatilla saponin B4 in preparation of medicine for treating ulcerative colitis Download PDF

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CN112107586A
CN112107586A CN202010030434.1A CN202010030434A CN112107586A CN 112107586 A CN112107586 A CN 112107586A CN 202010030434 A CN202010030434 A CN 202010030434A CN 112107586 A CN112107586 A CN 112107586A
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ulcerative colitis
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medicine
saponin
pulsatilla
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李艳红
韩倩
唐华争
邹敏
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South China University of Technology SCUT
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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/70—Carbohydrates; Sugars; Derivatives thereof
    • A61K31/7028—Compounds having saccharide radicals attached to non-saccharide compounds by glycosidic linkages
    • A61K31/7034—Compounds having saccharide radicals attached to non-saccharide compounds by glycosidic linkages attached to a carbocyclic compound, e.g. phloridzin
    • A61K31/704—Compounds having saccharide radicals attached to non-saccharide compounds by glycosidic linkages attached to a carbocyclic compound, e.g. phloridzin attached to a condensed carbocyclic ring system, e.g. sennosides, thiocolchicosides, escin, daunorubicin
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P1/00—Drugs for disorders of the alimentary tract or the digestive system
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P1/00—Drugs for disorders of the alimentary tract or the digestive system
    • A61P1/04—Drugs for disorders of the alimentary tract or the digestive system for ulcers, gastritis or reflux esophagitis, e.g. antacids, inhibitors of acid secretion, mucosal protectants

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Abstract

The invention discloses application of pulsatilla chinensis saponin B4 in preparation of a medicine for treating ulcerative colitis. The pulsatilla chinensis saponin B4 can remarkably relieve the symptoms of ulcerative colitis model mice, has better effect than the latest UC western medicine tofacitinib and the traditional Chinese medicine monomer berberine with larger treatment prospect on UC, does not cause spleen swelling accompanied by the two medicines in the process of treating the ulcerative colitis, has certain relieving effect, and has no structural damage to other visceral organs. The pulsatilla chinensis saponin B4 has definite curative effect on UC, low toxic and side effects and high safety, is expected to be used for clinical treatment of UC, and solves the problems of insufficient curative effect, more side effects and high toxicity of the existing medicines.

Description

Application of pulsatilla saponin B4 in preparation of medicine for treating ulcerative colitis
Technical Field
The invention relates to application of pulsatilla chinensis saponin B4, in particular to application of pulsatilla chinensis saponin B4 in preparation of a medicine for treating ulcerative colitis.
Background
Ulcerative Colitis (UC) is a chronic inflammatory bowel disease occurring in the colon, and is an autoimmune disease with the clinical manifestations of subacute or chronic abdominal pain, diarrhea with mucous bloody stool. The pathogenesis of ulcerative colitis is unknown, and genetic susceptibility, epithelial barrier defect, immune response disorder, environmental factors and the like are involved. Clinical treatment drugs mainly comprise anti-inflammatory drugs, corticosteroids, biological agents and immunosuppressants, however, most of the drugs have serious toxic and side effects, and the cure rate of patients is low and the drugs are easy to repeat. The tumor necrosis factor TNF-alpha antibody is widely used in nearly two decades, but has the problems of high toxic and side effects, high price, intramuscular or subcutaneous injection and the like.
The JAK (JAK kinase)/STAT (Signal transducer and activator of transcription) Signal channel inhibitor Tofacitinib (Tofacitinib) is a non-selective orally-taken JAK inhibitor and can inhibit JAK-STAT Signal channel cascade reaction, thereby obviously inhibiting inflammatory reaction mediated by various inflammatory factors. Tofacitinib was approved by the U.S. drug and food administration (FDA) for the treatment of the autoimmune disease rheumatoid arthritis in 11 months 2012 and for the treatment of moderate to severe UC in 5 months 2018 by the U.S. FDA with a high dose (10mg/kg) clinical remission rate of about 40%. But because the compound acts on the upstream target of the signal, a plurality of non-specific reactions can be caused, including toxic and side effects such as hyperlipemia, serious infection, malignant tumor, pulmonary embolism and the like, wherein part of the toxic side effects can endanger life, and the toxicity is in direct proportion to the dosage. In view of this, the european commission on drug administration safety began a comprehensive evaluation of tofacitinib at 2019, while the U.S. FDA also issued a black box warning (a rare safety warning) of the risk of tofacitinib causing blood clotting and death, while cautiously taking or discontinuing the clinical dose of 10mg/kg and compromising the clinical dose of 5 mg/kg. In view of the low clinical remission rate and serious adverse reactions of the latest medicament tofacitinib, the clinical treatment dilemma of UC is not well solved at present, and a new alternative medicament still needs to be researched urgently.
In recent years, the pharmacological activity of a traditional Chinese medicine monomer taking artemisinin as an example is continuously discovered, and the traditional Chinese medicine provides a huge treasury for the discovery of new medicines. Pulsatillae saponin B4(Anemoside B4, AB4) is the main component of Pulsatilla chinensis (Pulsatilla) of Pulsatilla of Ranunculaceae, and has biological activities of anti-inflammatory, anti-cytotoxicity, anti-tumor, etc., but its therapeutic effect on UC has not been reported.
Disclosure of Invention
The invention aims to overcome the defects in the prior art and provides the application of the pulsatilla saponin B4 in preparing the medicine for treating the ulcerative colitis.
The purpose of the invention is realized by the following technical scheme: application of pulsatilla saponin B4 in preparing medicine for treating ulcerative colitis is provided.
The medicine contains anemonin B4 and one of its physiologically acceptable salts.
The medicine contains pharmaceutically acceptable carriers or auxiliary materials.
The carrier is preferably at least one of a microcapsule, a microsphere, a nanoparticle and a liposome.
The auxiliary materials are preferably at least one of cosolvent, solubilizer, preservative, wetting agent, emulsifier, surfactant, sustained-release agent, excipient, disintegrant and lubricant.
The medicine for treating ulcerative colitis can be prepared into various dosage forms by adopting a conventional method, such as tablets, granules, capsules, pills and other oral dosage forms.
Compared with the prior art, the invention has the following advantages and effects:
1. the pulsatilla chinensis saponin B4 can significantly relieve the symptoms of mice in an ulcerative colitis model, and has better effect than the latest UC western medicine tofacitinib and traditional Chinese medicine monomer berberine with larger treatment prospect on UC.
2. The pulsatilla chinensis saponin B4 can not cause spleen swelling accompanying the treatment of ulcerative colitis by the tofacitinib and berberine, has a certain relieving effect, and has no structural damage to other organs. Therefore, the pulsatilla chinensis saponin B4 has definite curative effect on UC, low toxic and side effects and high safety, is expected to be used for clinical treatment of UC, and solves the problems of insufficient curative effect, more side effects and high toxicity of the existing medicines.
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FIG. 1 is a diagram of a mouse molding method.
FIG. 2 is a statistical graph of mouse body weights; wherein, in comparison with the control group,#P<0.05，##P<0.01，###P＜0.001，####P<0.0001；n＝4。
FIG. 3 is a statistical plot of the consistency scores of the mouse feces; wherein, in comparison with the control group,#P＜0.05，###P＜0.001，####P<0.0001; comparison with model group<0.05，**P<0.01；n＝4。
FIG. 4 is a statistical chart of mouse fecal occult blood scores; wherein, in comparison with the control group,####P<0.0001; comparison with model group<0.05，**P<0.01；n＝4。
FIG. 5 is a statistical plot of disease activity index scores for mice; wherein, in comparison with the control group,##P＜0.01，###P＜0.001，####P<0.0001; comparison with model group<0.05，**P<0.01；n＝4。
FIG. 6 is a measurement and statistical plot of colon length in mice; wherein A is a colon measurement photograph of a mouse; b is the colon length statistical result; compared with the control group, the compound of the formula,##p is less than 0.01; comparison with model group<0.01，***P＜0.001；n＝4。
FIG. 7 is a statistical plot of colon and spleen weights per unit length for mice; wherein A is a statistical chart of colon weight per unit length of a mouse; b is a weight statistical chart of the spleen of the mouse; wherein, in comparison with the control group,##P<0.01，###P<0.001; comparison with model group<0.01；n＝4。
FIG. 8 is a photograph of HE staining of mouse colon and spleen; wherein A is colon; b is spleen.
FIG. 9 is a photograph (20X) of HE staining of heart, liver, lung, kidney and brain of a mouse; wherein A is heart; b is liver; c is lung; d is kidney; e is brain.
Detailed Description
Embodiments of the present invention will be described in detail below with reference to embodiments, but it will be understood by those skilled in the art that the following embodiments and examples are only illustrative of the present invention and should not be construed as limiting the scope of the present invention. Those who do not specify the conditions are performed according to the conventional conditions or the conditions recommended by the manufacturer. The reagents or instruments used are not indicated by the manufacturer, and are all conventional products available commercially.
Example 1
First, mice (purchased from slykh scenda laboratory animals limited, china chansha, hannan) were randomly divided into 7 groups of 4 mice each. The mice are induced to generate ulcerative colitis symptoms by adopting dextran sulfate (DSS), then the mice are administered in groups, and the alleviation and treatment effects of the drugs in each group are examined by taking the alleviation condition of the inflammation of the mice after administration as an index. The experimental sets were as follows:
(1) normal group: during the experiment (18 days), free drinking water was given, and no drug was added to the diet.
(2) Model group (DSS group): adopting 2% (w/v) DSS circulation to induce mice to generate ulcerative colitis, and a molding method comprises the following steps: dissolving DSS in drinking water at a concentration of 2% (w/v), wherein the 2% (w/v) DSS solution is administered on days 1-4, 8-11 and 15-18, respectively; water was given on days 5-7 and 12-14 and the experiment was ended on day 19 (figure 1).
(3) Tofacitinib group (DSS + tofacitinib): the molding method and period were the same as those of the model group, and administration of 30mg/kg tofacitinib (purchased from MCE, USA) by gavage was started on the fifth day of molding.
(4) Berberine group (DSS + berberine): the molding method and period are the same as those of the model group, and 40mg/kg berberine (purchased from Yuanye, Shanghai, China, and not less than 95%) is used for intragastric administration on the fifth day of molding.
(5) Pulsatillae saponin B4 group (DSS + AB 4): the molding method and the period are the same as the model group, and 100mg/kg of pulsatilla saponin B4 (purchased from Rifangsi, Chinese Chengdu, and is not less than 98%) is used for intragastric administration at the fifth day of molding.
And secondly, recording the body weight, diarrhea and hematochezia conditions of all mice in the experimental process, and calculating a disease activity index score (DAI). DAI (8 points) — weight loss score (2 points) + fecal consistency score (3 points) + fecal occult blood score (3 points). The total DAI score ranged from 0 (unaffected) to 8 (severe colitis). The specific calculation method is as follows:
A) body weight
The body weight of the mice was monitored daily. Weight loss was calculated as the percentage difference between the original weight (day 0) and the daily weight. The scores were as follows: no change is 0; 0-1% ═ 0.1; 1-2% ═ 0.2; 2-3% ═ 0.3; 3-4% ═ 0.3 … … and so on, 19-20% ═ 0.19; 2 is larger than or equal to 20 percent.
B) Excrement and urine
The consistency of the mouse feces was measured on the 5 th, 7 th, 12 th, 15 th and 19 th days (first day after completion of molding). The scores were as follows: normal is 0; soft but still forming ═ 1; very soft-2; diarrhea was 3.
C) Occult blood in stool
The occult blood condition of the mouse feces is synchronously detected, and the score is as follows: negative blood is 0; positive blood hidden-2 min is 1, 1min is 1.25, and 10s internal color development is 1.5; 1.75 parts of instant color development; feces bloodstain can be seen by naked eyes 2; rectal bleeding is 3.
According to the scoring standard, all scores of the normal group are zero, which indicates that the mice in the normal group have no related abnormal reaction. Based on the disease activity indexes obtained by further comparing the weight, the loose stool and the blood of each group of mice and the integral accumulation of the indexes: compared with the normal group, the weight of the model group is obviously reduced from day 2, the weight of each drug treatment group is relieved to different degrees, and particularly, the weight of the pulsatilla saponin B4 treatment group is obviously increased from day 12 compared with the model group (figure 2); the model group significantly increased the stool consistency score at five test times (days 5, 7, 12, 15, 19) compared to the normal group; compared to the model group, only pasqueflower saponin B4 significantly reduced the stool consistency score at day 12, and berberine and anemonin B4 significantly reduced the score at day 19 (fig. 3); the model group significantly increased the hematochezia score at five test times (days 5, 7, 12, 15, 19) compared to the normal group; compared with the model group, the tofacitinib group significantly reduced the hematochezia score on days 12, 15 and 19, and the pulsatilla saponin B4 group reduced the hematochezia score on days 12 and 19 (fig. 4); the DAI scores of the model groups at each test time point were significantly increased compared to the normal group; compared to the model group, 3 administration groups all significantly reduced the DAI score on the first day (day 19) after the end of administration, while the pasqueflower saponin B4 group also significantly reduced the DAI score on day 12 and was lower than the other groups (fig. 5). The results show that the Chinese pulsatilla root saponin B4 has obvious effects of reducing the body weight, and improving the degree of loose stool and hematochezia, and the effect is slightly better than that of two positive drug groups.
Three, each group of mice was sacrificed on day 19, the colons were rapidly taken out and measured for length, weighed after washing with physiological saline, and the spleens were separated and weighed.
The results show that compared with the normal group, the colon of the mouse in the model group is obviously shortened and swollen; compared with the model group, the three administration groups significantly improve the colon shortening condition, the pulsatilla saponin B4 group has the best effect of improving colon shortening (figure 6), and the pulsatilla saponin B4 improves the colon swelling condition, and the effect is better than that of two positive control drugs, namely tofacitinib and berberine (figure 7A). The spleen of the model mouse is obviously swelled compared with the normal group; compared to the model group, spleen swelling was somewhat relieved in the anemonin B4 group, and no further spleen swelling was observed in the tofacitinib and berberine groups (fig. 7B).
And fourthly, performing hematoxylin-eosin (HE) staining on the colon, the spleen, the liver, the heart, the kidney, the lung and the brain of each group of mice, detecting pathological changes of the colon and the spleen, and simultaneously detecting whether the organs have structural damage or not so as to determine the potential toxicity of the medicines used in each administration group.
The results show that: the colon of the model group mice has severe structural damage relative to the normal group; compared with the model group, the three administration groups obviously improved the colon structure destruction, and the pulsatilla saponin B4 group had the best improvement effect (figure 8A). Splenocyte proliferation appears in the model group and the three positive medicine groups relative to the normal group; pulsatillae saponin B4 group was milder (FIG. 8B). Structural tissues of liver, heart, kidney, lung, brain and other organs of the pulsatilla saponin B4 group have no obvious injury compared with the model group (fig. 9A-E), which indicates that the pulsatilla saponin B4 group has no obvious toxicity.
The results of the second to fourth steps show that compared with the normal group, the scores and indexes of all indexes of the clinical symptoms of the mice in the model group are obviously improved, the colon is obviously shortened and swollen, and the spleen is obviously swollen, so that the DSS can effectively induce the mice to generate the symptoms of ulcerative colitis, and all indexes of the DSS accord with the symptoms and development rules of the colitis. Compared with the model group, the three administration groups can improve the symptoms of the ulcerative colitis, and the shortening degree of the colon of the mouse is reduced, wherein the score of each index of the pulsatilla saponin B4 group is slightly better than that of the other two positive control groups. Therefore, the drug AB4 provided by the invention has obvious curative effect on UC, can relieve spleen swelling, has no structural damage to organs, has curative effect on relieving and treating UC, and has better safety.
The above embodiments are preferred embodiments of the present invention, but the present invention is not limited to the above embodiments, and any other changes, modifications, substitutions, combinations, and simplifications which do not depart from the spirit and principle of the present invention should be construed as equivalents thereof, and all such changes, modifications, substitutions, combinations, and simplifications are intended to be included in the scope of the present invention.

Claims (6)

1. Application of pulsatilla saponin B4 in preparing medicine for treating ulcerative colitis is provided.
2. The use according to claim 1, wherein the medicament comprises one of anemonin B4 and a physiologically acceptable salt thereof.
3. The use according to claim 1 or 2, wherein the medicament comprises a pharmaceutically acceptable carrier or excipient.
4. The use of claim 3, wherein the carrier is at least one of a microcapsule, a microsphere, a nanoparticle, and a liposome.
5. The use of claim 3, wherein the adjuvant is at least one of a cosolvent, a solubilizer, a preservative, a wetting agent, an emulsifier, a surfactant, a sustained release agent, an excipient, a disintegrant, and a lubricant.
6. The use of claim 1, wherein the medicament is formulated as a tablet, granule, capsule, pill.
CN202010030434.1A 2020-01-13 2020-01-13 Application of pulsatilla saponin B4 in preparation of medicine for treating ulcerative colitis Pending CN112107586A (en)

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Cited By (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN112656802A (en) * 2021-01-14 2021-04-16 华南理工大学 Application of pulsatilla saponin B4 in preparation of medicine for treating multiple sclerosis
CN114209739A (en) * 2021-12-16 2022-03-22 江西中医药大学 Application of pulsatilla chinensis extract in preparation of antidepressant drug
CN115844911A (en) * 2022-12-02 2023-03-28 华南理工大学 Combined pharmaceutical composition for treating ulcerative colitis
CN116196270A (en) * 2023-05-05 2023-06-02 江西中医药大学 In-situ gel preparation for treating ulcerative colitis and preparation method thereof

Citations (1)

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Publication number Priority date Publication date Assignee Title
CN109549944A (en) * 2019-01-30 2019-04-02 苏州大学 Application of the anemoside B4 in preparation inflammatory enteropathy drug

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CN109549944A (en) * 2019-01-30 2019-04-02 苏州大学 Application of the anemoside B4 in preparation inflammatory enteropathy drug

Non-Patent Citations (1)

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Title
杨连荣 等: "白头翁皂苷 B4 在正常大鼠和溃疡性结肠炎大鼠体内组织分布情况的比较", 《中国实验方剂学杂志》 *

Cited By (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN112656802A (en) * 2021-01-14 2021-04-16 华南理工大学 Application of pulsatilla saponin B4 in preparation of medicine for treating multiple sclerosis
CN114209739A (en) * 2021-12-16 2022-03-22 江西中医药大学 Application of pulsatilla chinensis extract in preparation of antidepressant drug
CN114209739B (en) * 2021-12-16 2023-05-26 江西中医药大学 Application of pulsatilla chinensis bunge extract in preparation of antidepressant treatment drugs
CN115844911A (en) * 2022-12-02 2023-03-28 华南理工大学 Combined pharmaceutical composition for treating ulcerative colitis
CN116196270A (en) * 2023-05-05 2023-06-02 江西中医药大学 In-situ gel preparation for treating ulcerative colitis and preparation method thereof

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