LU502836B1 - Linderae radix composition with antagonistic effect on inflammatory bowel diseases and application thereof - Google Patents
Linderae radix composition with antagonistic effect on inflammatory bowel diseases and application thereof Download PDFInfo
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- LU502836B1 LU502836B1 LU502836A LU502836A LU502836B1 LU 502836 B1 LU502836 B1 LU 502836B1 LU 502836 A LU502836 A LU 502836A LU 502836 A LU502836 A LU 502836A LU 502836 B1 LU502836 B1 LU 502836B1
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- linderae
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- 239000000203 mixture Substances 0.000 title claims abstract description 37
- 208000022559 Inflammatory bowel disease Diseases 0.000 title claims abstract description 11
- 230000003042 antagnostic effect Effects 0.000 title abstract description 7
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims abstract description 22
- 150000004676 glycans Chemical class 0.000 claims abstract description 22
- 239000005017 polysaccharide Substances 0.000 claims abstract description 21
- 229920001282 polysaccharide Polymers 0.000 claims abstract description 21
- 241001076416 Dendrobium tosaense Species 0.000 claims abstract description 20
- 239000000284 extract Substances 0.000 claims abstract description 17
- 238000002360 preparation method Methods 0.000 claims abstract description 5
- 238000001556 precipitation Methods 0.000 claims abstract description 3
- 238000000605 extraction Methods 0.000 claims abstract 4
- 238000010992 reflux Methods 0.000 claims abstract 4
- 238000002156 mixing Methods 0.000 claims abstract 2
- 239000003814 drug Substances 0.000 claims description 10
- 229940079593 drug Drugs 0.000 claims description 7
- 239000002244 precipitate Substances 0.000 claims description 5
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 5
- 239000006228 supernatant Substances 0.000 claims description 3
- 238000001816 cooling Methods 0.000 claims description 2
- 239000000706 filtrate Substances 0.000 claims 4
- 238000001035 drying Methods 0.000 claims 2
- 238000001914 filtration Methods 0.000 claims 2
- 238000005119 centrifugation Methods 0.000 claims 1
- 238000000034 method Methods 0.000 claims 1
- 238000005406 washing Methods 0.000 claims 1
- 210000001072 colon Anatomy 0.000 abstract description 21
- 230000000968 intestinal effect Effects 0.000 abstract description 21
- 230000037396 body weight Effects 0.000 abstract description 9
- 208000025865 Ulcer Diseases 0.000 abstract description 8
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- 230000006378 damage Effects 0.000 abstract description 5
- 208000014674 injury Diseases 0.000 abstract description 5
- 238000003809 water extraction Methods 0.000 abstract description 2
- 238000010438 heat treatment Methods 0.000 abstract 1
- 241000700159 Rattus Species 0.000 description 34
- NHJVRSWLHSJWIN-UHFFFAOYSA-N 2,4,6-trinitrobenzenesulfonic acid Chemical compound OS(=O)(=O)C1=C([N+]([O-])=O)C=C([N+]([O-])=O)C=C1[N+]([O-])=O NHJVRSWLHSJWIN-UHFFFAOYSA-N 0.000 description 15
- 210000001519 tissue Anatomy 0.000 description 14
- 208000004232 Enteritis Diseases 0.000 description 8
- 230000002550 fecal effect Effects 0.000 description 8
- 230000000694 effects Effects 0.000 description 7
- 210000001165 lymph node Anatomy 0.000 description 7
- 239000000243 solution Substances 0.000 description 7
- 230000003247 decreasing effect Effects 0.000 description 6
- 230000014509 gene expression Effects 0.000 description 6
- 238000012360 testing method Methods 0.000 description 6
- 102100027456 Cytochrome c oxidase subunit 2 Human genes 0.000 description 5
- 101000725401 Homo sapiens Cytochrome c oxidase subunit 2 Proteins 0.000 description 5
- 101000605127 Homo sapiens Prostaglandin G/H synthase 2 Proteins 0.000 description 5
- 241001465754 Metazoa Species 0.000 description 5
- 210000000265 leukocyte Anatomy 0.000 description 5
- 241001523681 Dendrobium Species 0.000 description 4
- 206010061218 Inflammation Diseases 0.000 description 4
- 210000001744 T-lymphocyte Anatomy 0.000 description 4
- 230000002489 hematologic effect Effects 0.000 description 4
- 230000004054 inflammatory process Effects 0.000 description 4
- 210000001616 monocyte Anatomy 0.000 description 4
- 210000000440 neutrophil Anatomy 0.000 description 4
- 230000001575 pathological effect Effects 0.000 description 4
- 210000000662 T-lymphocyte subset Anatomy 0.000 description 3
- 230000000112 colonic effect Effects 0.000 description 3
- 238000001514 detection method Methods 0.000 description 3
- 210000003608 fece Anatomy 0.000 description 3
- 230000002055 immunohistochemical effect Effects 0.000 description 3
- 231100000915 pathological change Toxicity 0.000 description 3
- 230000036285 pathological change Effects 0.000 description 3
- 239000000126 substance Substances 0.000 description 3
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 2
- 210000000436 anus Anatomy 0.000 description 2
- 230000036528 appetite Effects 0.000 description 2
- 235000019789 appetite Nutrition 0.000 description 2
- 238000004820 blood count Methods 0.000 description 2
- 210000003743 erythrocyte Anatomy 0.000 description 2
- 210000002443 helper t lymphocyte Anatomy 0.000 description 2
- 230000008946 inflammatory intestinal reaction Effects 0.000 description 2
- 230000003902 lesion Effects 0.000 description 2
- 210000000713 mesentery Anatomy 0.000 description 2
- 210000004877 mucosa Anatomy 0.000 description 2
- 230000002093 peripheral effect Effects 0.000 description 2
- 210000001995 reticulocyte Anatomy 0.000 description 2
- 210000002966 serum Anatomy 0.000 description 2
- 206010009900 Colitis ulcerative Diseases 0.000 description 1
- 102000001554 Hemoglobins Human genes 0.000 description 1
- 108010054147 Hemoglobins Proteins 0.000 description 1
- 206010067482 No adverse event Diseases 0.000 description 1
- QGMRQYFBGABWDR-UHFFFAOYSA-M Pentobarbital sodium Chemical compound [Na+].CCCC(C)C1(CC)C(=O)NC(=O)[N-]C1=O QGMRQYFBGABWDR-UHFFFAOYSA-M 0.000 description 1
- UIIMBOGNXHQVGW-UHFFFAOYSA-M Sodium bicarbonate Chemical compound [Na+].OC([O-])=O UIIMBOGNXHQVGW-UHFFFAOYSA-M 0.000 description 1
- 208000031737 Tissue Adhesions Diseases 0.000 description 1
- 201000006704 Ulcerative Colitis Diseases 0.000 description 1
- 210000001015 abdomen Anatomy 0.000 description 1
- 230000005856 abnormality Effects 0.000 description 1
- 230000001476 alcoholic effect Effects 0.000 description 1
- 230000003110 anti-inflammatory effect Effects 0.000 description 1
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- 210000004027 cell Anatomy 0.000 description 1
- 239000006285 cell suspension Substances 0.000 description 1
- 239000003153 chemical reaction reagent Substances 0.000 description 1
- 239000003795 chemical substances by application Substances 0.000 description 1
- 230000001684 chronic effect Effects 0.000 description 1
- 210000001151 cytotoxic T lymphocyte Anatomy 0.000 description 1
- 230000013872 defecation Effects 0.000 description 1
- 238000002224 dissection Methods 0.000 description 1
- 230000003628 erosive effect Effects 0.000 description 1
- 238000002474 experimental method Methods 0.000 description 1
- 230000037406 food intake Effects 0.000 description 1
- 235000012631 food intake Nutrition 0.000 description 1
- 210000003405 ileum Anatomy 0.000 description 1
- 230000005934 immune activation Effects 0.000 description 1
- 208000026278 immune system disease Diseases 0.000 description 1
- 239000012535 impurity Substances 0.000 description 1
- 230000008595 infiltration Effects 0.000 description 1
- 238000001764 infiltration Methods 0.000 description 1
- 208000027866 inflammatory disease Diseases 0.000 description 1
- 230000002757 inflammatory effect Effects 0.000 description 1
- 230000003871 intestinal function Effects 0.000 description 1
- 230000006742 locomotor activity Effects 0.000 description 1
- 239000000463 material Substances 0.000 description 1
- 238000005259 measurement Methods 0.000 description 1
- WSFSSNUMVMOOMR-NJFSPNSNSA-N methanone Chemical compound O=[14CH2] WSFSSNUMVMOOMR-NJFSPNSNSA-N 0.000 description 1
- 230000027939 micturition Effects 0.000 description 1
- 230000004660 morphological change Effects 0.000 description 1
- 230000017074 necrotic cell death Effects 0.000 description 1
- 238000001543 one-way ANOVA Methods 0.000 description 1
- 239000012188 paraffin wax Substances 0.000 description 1
- 230000036961 partial effect Effects 0.000 description 1
- 229960002275 pentobarbital sodium Drugs 0.000 description 1
- 230000000144 pharmacologic effect Effects 0.000 description 1
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- 150000004804 polysaccharides Polymers 0.000 description 1
- 239000000047 product Substances 0.000 description 1
- 230000035755 proliferation Effects 0.000 description 1
- 102000004169 proteins and genes Human genes 0.000 description 1
- 108090000623 proteins and genes Proteins 0.000 description 1
- 210000000664 rectum Anatomy 0.000 description 1
- 230000002829 reductive effect Effects 0.000 description 1
- 210000003289 regulatory T cell Anatomy 0.000 description 1
- 230000001105 regulatory effect Effects 0.000 description 1
- 210000000813 small intestine Anatomy 0.000 description 1
- 235000014347 soups Nutrition 0.000 description 1
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Classifications
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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
- A61K31/715—Polysaccharides, i.e. having more than five saccharide radicals attached to each other by glycosidic linkages; Derivatives thereof, e.g. ethers, esters
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K36/00—Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
- A61K36/18—Magnoliophyta (angiosperms)
- A61K36/185—Magnoliopsida (dicotyledons)
- A61K36/54—Lauraceae (Laurel family), e.g. cinnamon or sassafras
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- A—HUMAN NECESSITIES
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- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K36/00—Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
- A61K36/18—Magnoliophyta (angiosperms)
- A61K36/88—Liliopsida (monocotyledons)
- A61K36/898—Orchidaceae (Orchid family)
- A61K36/8984—Dendrobium
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K2236/00—Isolation or extraction methods of medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicine
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Abstract
Disclosed are a Linderae Radix composition with an antagonistic effect on inflammatory bowel diseases and an application thereof. The Linderae Radix composition is prepared by mixing a Linderae Radix extract and Dendrobium officinale polysaccharide, and a mass percent of the Linderae Radix extract in the Linderae Radix composition is 1-99%. A preparation method for the Linderae Radix extract includes the following steps: adding Linderae Radix decoction pieces to 50-90% (volume concentration) ethanol, heating the Linderae Radix decoction pieces, and performing reflux extraction and concentration to obtain the Linderae Radix extract; a preparation method for the Dendrobium officinale polysaccharide includes the following steps: adding Dendrobium officinale to ethanol, and performing water extraction and alcohol precipitation to obtain the Dendrobium officinale polysaccharide. The composition has the functions of improving the body weight decrease, and significantly alleviating colon hyperemia and edema, intestinal wall expansion, and mucosal ulcer and injuries.
Description
BL-5563
LINDERAE RADIX COMPOSITION WITH ANTAGONISTIC EFFECT ON LUs02836
INFLAMMATORY BOWEL DISEASES AND APPLICATION THEREOF
[01] The present invention relates to the technical field of traditional Chinese medicine products, and relates to a Linderae Radix composition with an antagonistic effect on inflammatory bowel diseases and an application thereof.
[02] The inflammatory bowel disease (IBD) is a chronic nonspecific intestinal inflammatory disease involving the ileum, rectum and colonic mucosa.
[03] However, there is few reports on the application of traditional Chinese medicine compositions in treating inflammatory bowel diseases in the market.
[04] In view of current gaps in the market and shortcomings in the prior art, the present invention is intended to provide a Linderae Radix composition with an antagonistic effect on inflammatory bowel diseases and an application thereof.
[05] In the present invention, the Linderae Radix composition includes a Linderae
Radix extract and Dendrobium officinale polysaccharide, wherein the Linderae Radix extract is extracted with an alcoholic solution, and centrifugated after cooling and standing, supernatant are collected, and precipitates and water-soluble impurities are removed; Dendrobium officinale is subjected to water extraction and alcohol precipitation, and is rich in polysaccharide components. These two substances are combined to produce a significant effect on preventing TNBS-induced ulcerative colitis, and can regulate the proportion of T-cell subsets in mesenteric lymph nodes, improve intestinal immune disorders, inhibit inflammation-associated protein expressions in colon tissue, reduce intestinal inflammatory reactions, improve pathological changes in colon tissue affected by the inflammatory bowel disease, and restore intestinal functions; while improving the roughness, largeness and blackness of traditional Chinese medicine extracts, reducing doses of drugs, and facilitating improvements in the patient’s compliance.
[06] Pharmacological (animal) studies have demonstrated that the Linderae Radix composition provided by the present invention can significantly reduce the intestinal index and intestinal weight per unit length of the enteritis model animals, and improve pathological changes in colon tissue. Administering the composition to enteritis model rats can effectively improve the body weight decrease, reduce the fecal property index, reduce numbers of white blood cells, neutrophils and monocytes in serum, significantly alleviate colon hyperemia and edema, intestinal wall expansion, and mucosal ulcer and injuries, and reduce pathological tissue scores.
[07] FIG 1 shows the morphology of colon tissue of rats in various groups: A: normal control; B: model control; C: Linderae Radix extract, D: crude Dendrobium officinale polysaccharide; E: Linderae Radix-Dendrobium officinale composition. 1
BL-5563
[08] FIG 2 shows immunohistochemical results of COX2 protein expressions in LU502836 colon tissue of rats in various groups: A: normal control; B: model control; C: Linderae
Radix extract; D: crude Dendrobium officinale polysaccharide; E: Linderae
Radix-Dendrobium officinale composition.
[09] Example 1: Study of effect on 2,4,6-trinitrobenzene sulfonic acid (TNBS)-induced enteritis
[10] 1. Experimental materials
[11] 1.1. Animals: Male SD rats, purchased from the Experimental Animal Center of
Zhejiang Academy of Medical Sciences.
[12] 1.2. Drugs and reagents: Linderae Radix was provided by Tiantaishan Wuyao
Co., Ltd; a Linderae Radix extract and Dendrobium officinale polysaccharide were prepared and mixed at a mass ratio of 1:1, and prepared with pure water into a medicinal solution at the corresponding concentration for later use.
[13] Modeling drug: 2,4,6-trinitrobenzene sulfonic acid (TNBS) (batch No.:
SLBK1620V, Sigma). Preparation of modeling solution (freshly prepared): 40 ml of a
TNBS stock solution (50 mg/ml) was mixed with an equal volume of anhydrous ethanol to obtain a TNBS 50% ethanol solution (at a TNBS concentration of 25 mg/ml).
[14] 1.3. Instruments: ITACHI 7100 biochemical analyzer; BPH-9082 microplate reader, BD Calibur flow cytometer; a complete set of pathological equipment (Leica,
Germany)
[15] 2. Experimental method
[16] 2.1. Grouping and administration: After adaptively fed for 3 days, SD rats were weighed and randomly grouped by body weight into a normal control group, a model control group, a Linderae Radix extract group, a Dendrobium officinale polysaccharide group and a Linderae Radix-Dendrobium officinale composition group. After being fasted but given water for 24 h, SD rats were anesthetized with 1% pentobarbital sodium (at a dose of 40 mg/kg). A TNBS modeling group was given a TNBS 50% ethanol solution for coloclysis. A fully lubricated rubber tube with a diameter of 2 mm was gently inserted from the anus to a depth of about 8 cm, a prepared TNBS solution (25 mg/ml, 0.36 ml/100 g) was delivered at a dose of 90 mg/kg, and then the plastic rubber tube was slowly pulled out. The rats were inverted for 1 min by holding the anus and lifting the tail by hand, so that the modeling agent fully penetrated into the intestinal lumen. The normal control group was given an equal volume of normal saline for coloclysis. The administration was started 1 day after TNBS modeling and continued for 21 days. The normal control group and model group were given the corresponding doses of pure water.
[17] 2.2. Index detection
[18] 2.2.1. General observation: For rats in various groups, the general condition, survival status, body weight, food intake and fecal shape were observed and recorded, and fecal occult blood was detected. Fecal properties were scored according to the criteria: normal (dark brown, hard and ellipsoidal) = O point; soft = 2 points; loose or bloody = 4 points. 2
BL-5563
[19] 2.22. Gross observation and pathological examination of tissue: During LU502836 dissection, the appearance of the colon was observed with naked eyes for morphological changes (such as color, edema, intestinal wall expansion and adhesion, ulcer and erosion), and scored according to the criteria: no significant abnormality (0 point), hyperemia and edema (1 point), intestinal wall expansion (2 points), and ulcer and adhesion (3 points), tissue taken from a lesion was placed in 10% formaldehyde, embedded in paraffin, sectioned, stained with hematoxylin-eosin (HE), histologically observed under a microscope, and scored according to the criteria: no obvious injury (0 point), inflammatory infiltration and vascular proliferation (1 point), mucosal cell necrosis (2 points), and ulcer (3 points), COX2 protein expressions in colon tissue were observed by an immunohistochemical DAB staining method.
[20] 2.2.3. Determination of colon length, weight and index: The rats were weighed before sacrifice, and the colon was taken after sacrifice. The intestinal lumen was cut along the mesentery, and rinsed with ice normal saline. The colon was blotted with filter paper, its total length and weight were measured, and its weight per unit length and index were calculated.
[21] 2.2.4. Detection of T-cell subsets in mesenteric lymph nodes by flow cytometer:
The rats were dissected along the center line of the abdomen, so that the small intestine was exposed. A lymph node was taken from a deep part of the mesentery, placed in a pre-cooled PBS solution, ground, filtered with a 200-mesh screen, centrifuged at 1,000 r/min, re-suspended and counted. 100 pL of cell suspension was taken, serum of rats was blocked, the corresponding antibody was added, and detection was performed by a flow cytometer.
[22] 2.3. Statistical methods: Measurement data was expressed in X, +s. The #-test was used for a comparison between two groups, and the one-way analysis of variance was used for multiple comparisons among the model group and various administration groups. P < 0.05 indicated a statistically significant difference.
[23] 3. Experimental results
[24] 3.1. General observation: The rats in the normal control group were active, with glossy hair, good appetite, normal urination and defecation, and hard, dark brown and ellipsoidal feces. No adverse reactions were observed from a normal administration group, which was similar to the normal control group throughout the test. The rats in the model control group had less locomotor activities, liked to gather at edges, looked tired and lazy, responded slowly and lost appetite, with dirty crissum, and mucosanguineous or loose feces, which was significantly different from normal rat feces, on the next day after modeling. The fecal property index significantly increased on the third day after modeling (P < 0.01), and gradually recovered after 2 weeks. Compared with the model group, all test substances can effectively reduce the fecal property index, which significantly decreased in the second week of administration in the Linderae
Radix-Dendrobium officinale composition group (P < 0.05) (see Table 1-1). 3
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[25] Table 1-1. Fecal property indexes of rats in various groups (x,~ + SD) LU502836
Fecal property index
Group _— 3d 11d 19d
Normal control group 0.00+0.00 0.00+0.00 0.00+0.00
Model control group 3.61+0.60°° 2.58+1.34%" 1.33+0.76°
Linderae Radix group 2.95+1.00 2.14+1.22 0.85+0.38
Dendroblum officinale 2.98+1.02 2.44+1.43 0.94+0.64 polysaccharide group
Composition group 2.85+1.02 1.43+1.68" 0.76+0.54
[26] Note: Compared with the normal control group, “P < 0.05, and “°P < 0.01; compared with the model control group, * P <0.05.
[27] 3.2. Effect on body weight compared with normal control group: The Linderae
Radix extract, crude Dendrobium officinale polysaccharide and their composition can effectively increase body weights of model rats, and improve body weight decrease symptoms in TNBS-induced enteritis model rats (see Table 1-2).
[28] Table 1-2. Body weights of rats in various groups at time points (x, 7 = SD)
Body weight (g)
Group -_— a ————————— 1d 3d 6d 9d 11d 14d 17d 19d
Normal control 465.4 461.4 4704+ 4758+ 496.6 5104+ 5190 5292+ group +254 £194 21.8 20.3 19.7 19.5 +249 24.2
Model control 4704 4276 423.8 49104 4480+ 465.8 470.6 481.2+ group +282 +360 36.1° 37.8" 439 384° +414 386"
Linderae Radix 4558 431.2 4305+ 4288+ 430.0 4455+ 463.8 483.7+ group +202 £255 44.2 47.4 53.9 62.7 +60.8 30.5
D . endroiun 469.4 4346 4224+ 4324+ 4378+ 4626+ 4838 498.0% polysacchaflde 4193 +207 256 253 247 175 £150 167 group
Composition erou 465.2 4384 4288+ 4270+ 42804 4410+ 471.0 4908+
P SOUP 4207 +220 284 298 380 318 +340 377
[29] Note: Compared with the normal control group, **P < 0.05 (the same below).
[30] 3.3. Effect on colon length, weight and intestinal index of model rats: Compared with the model control group, the test drugs can effectively increase the colon length, and decrease the weight per unit length and colon coefficient, which is more significant in the composition group; this suggests that the composition can effectively improve pathological changes in colon tissue of enteritis model animals (see Table 1-3).
[31] Table 1-3. Intestinal lengths, weights and indexes in various groups (x,7 + SD)
I inal . . igh
Group Dose eat Intestinal Intestinal enn (g/kg) length (cm) index (%) (2) (g/cm)
Normal control - 23402 174429 0434003 0.13+0.02 group 4
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Model control - 4.1416° 128:20° 0864037 03200185 CI group
Linderae Radix 1 3.240 6 141410 0.654013 0.22+0.06 group
Dendrobium polysaccharide 0.24 4.0+1.1 14.8+3.1 0.8+0.24 0.27+0.07 group
Composition 1 3240.7 162425 0.63+0.14 0.20+0.04 group
[32] 3.4. Gross observation and histopathological examination of intestinal tissue:
For rats in the normal control group, the colonic mucosa was generally observed to be light red, smooth and intact; for rats in the model control group, the colonic wall was thickened, and colon specimens exhibited hyperemia and edema, ulceration, colon tissue adhesion, partial intestinal wall expansion, and obvious mucosal ulcer and injuries (see FIG 1). It can be seen through the observation of HE-stained histopathological sections under a light microscope that, compared with the model control group, administering the Linderae Radix extract, Dendrobium officinale polysaccharide and their composition for interventions can effectively improve colon hyperemia and edema, intestinal wall expansion, and mucosal ulcer and injuries, which suggests that the composition has an antagonistic effect on TNBS-induced intestinal tissue lesions (see Table 1-4). The immunohistochemical results show that inflammation-associated COX2 protein expressions significantly increased in colon tissue of model rats, and the test drugs can effectively inhibit COX2 expressions in the colon and inhibit inflammatory reactions in enteritis model rats (see FIG. 2).
[33] Table 1-4. Gross observation and pathological scores of intestinal tissue of rats in various groups(x,~ + SD)
Examination indexes
Gross . .
Group observation Histopathological Overall score score score
Normal control group 0.0+0.0 0.0+0.0 0.0+0.0
Model control group 1.71+0.99°® 1.89+0.9°* 3.33+1.80°*
Linderae Radix group 1.26+0.81 1.62+0.81 3.24+1.53
Dendrobium 1.16+0.27 1.61+1.08 2.88+1.62 polysaccharide group
Composition group 0.90+0.80 1.26+0.72 2.07+1.35
[34] Note: Compared with the normal control group, ““P < 0.01 (the same below).
[35] 3.5. Effect on hematological indexes of model rats: Compared with the normal group, the white blood cell count as well as the total number and proportion of neutrophils, monocytes and reticulocytes significantly increased in model rats (P < 0.05 and P < 0.01), and the red blood cell count and hemoglobin significantly decreased (P < 0.05), which suggests that there are inflammatory reactions in model animals; compared
BL-5563 with the model group, the test substances can effectively reduce numbers of peripheral LU502836 white blood cells, neutrophils, monocytes and reticulocytes, and numbers of white blood cells, neutrophils and monocytes decreased more significantly in the composition group (Tables 1-5, 6 and 7), which suggests that the composition is anti-inflammatory to some extent.
[36] Table 1-5. Determination results of hematological indexes of rats in various groups -- 1 (x, + SD)
Dose
Group (ke) WBC RBC HGB HCT
Normal control - 8.9+2.1 8.11+0.33 138.7£7.2 44.942 3 group
Model control - 15.449,25 7.33+056% 1274114% 41.7439 group
Linderae Radix 1 123447 7.374056 1293+140 42.6+47 group
Dendrobium polysaccharide 1 11.6+£1.7 7.81+£0.73 133.3+7.4 41.1£2.4 group
Composition 1 10.741.6 7.624087 125.8415.2 42.2447 group
[37] Table 1-6. Determination results of hematological indexes of rats in various groups -- 2(x,~ + SD)
Group Dose %NEU %LYM %MON %EOS %BA %LU %RETI (g/kg) T PH O so C C
Normal 19.244. 77.04. 1.940. 1.200. 0.18 0564 2.41+0. control group | 1 9 64 40 0.07 0.21 46
Model 40.11 54.342 33641. 1.520. 020+ 048+ 5.77Æ2. control group ‘ 9.5% 1.1 31° 73 0.07 026 93°°
Linderae 1 32.341 62.4*1 3.13+1. 1.230. 0.20 0.57 2.6940.
Radix group 5.0 5.3 03 21 0.00 0.32 58 an | 29.041 66.641 336+1. 1.3340. 0.184 050+ 3.96+0. 7.3 7.7 11 91 0.08 0.23 93 e group
Composition 1 28.941 65.941 2.460. 11.240. 0.17% 0.60+ 3.88+0. group 7.0 7.6 65 58 0.06 0.26 95
[38] Table 1-7. Determination results of hematological indexes of rats in various groups -- 5(x, 7 + SD)
Group oo #NEUT je #EOS RAS #LUC Qu
Normal 1.690. 683+ 017 0.11& 0.02 005 1964+ control group | 49 1.82 0.06 0.04 0.01 0.03 42.7
Model i TAS+6. 695+ 058 0.28+ 0.03 0.07+ oo control group 82% 1.72 043° 031 0.03 0.04 X 6
BL-5563
Linderae | 3.9342. 7.004 038+ 0.174 0.024 0.074 2077+ LUS02836
Radix group 37 0.87 0.17 0.08 0.01 0.04 30.2 an | 4003. 7.74% 0.314 015: 0024 005 288.1% 40 2.42 0.17 0.10 0.01 0.04 50.0 e group
Composition 1 2.9241. 7.174 035+ 0.12+ 0.03 0.05 289.7+ group 47 2.73 0.11 0.07 0.01 0.03 42.7
[39] 3.6. Effect on T-cell subtypes in mesenteric lymph nodes of model rats:
Compared with the normal group, T-lymphocytes (CD3*) in mesenteric lymph nodes of model rats significantly decreased (P < 0.05), the proportion of helper T-cell subsets (Th,
CD4") increased, the proportion of cytotoxic T-cell subtypes (Tc, CD8 ¥) decreased, and the ratio of regulatory T-cells (FoxP3) to helper T-cell subsets decreased, which suggests that model rats were in a state of excessive immune activation. Compared with the model group, the Dendrobium officinale polysaccharide and composition can increase the number of T-lymphocytes in mesenteric lymph nodes, decrease the proportion of Th, increase the proportion of Tc, up-regulate the FoxP3/Th ratio, and improve a state of intestinal immune hyperfunction in model rats (see Table 1-8).
[40] Table 1-8. Determination results of peripheral T-cell subtypes of rats in various groups(x,- + SD)
Group oho CD3* (%) CD4*/CD3* CD8*/CD3* CD4*/CDS* foxP3/CD4*
Normal - 46.1465 644460 317465 22407 77412 control group
Model control - 40.5+2.6* 669432 286431 2404 61#11° group
Linderae . 1 37.34+10.3 67.644.1 28.6+4.0 2.4+0.5 6.2+0.8
Radix group
Dendrobium polysaccharide 1 41.244 8 62.8+6.7 30.0+4.3 2.1+0.3 6.3+1.0 group
Composition 1 52.0410.3* 65.6453 30.6452 2.240.5 6.3+1.0 group
[41] Note: Compared with the normal control group, “P < 0.05; compared with the model control group, * P <0.05.
[42] These results show that the Linderae Radix extract, Dendrobium officinale polysaccharide and their composition have a significant antagonistic effect on
TNBS-induced enteritis, which is more significant in the composition group; such an anti-enteritis effect is related to inhibited COX2 expressions, regulated T-cell subsets in mesenteric lymph nodes, improved intestinal immune hyperfunction and reduced intestinal inflammatory reactions. 7
Claims (3)
1. An application of a Linderae Radix composition in preparing drugs for treating inflammatory bowel diseases, wherein the Linderae Radix composition is prepared by mixing a Linderae Radix extract and Dendrobium officinale polysaccharide at a mass ratio of 1:1; a preparation method for the Linderae Radix extract comprises the following steps: adding Linderae Radix decoction pieces to 50-90% (volume concentration) ethanol in a volume of 8-12 times a volume of the decoction pieces, and conducting reflux extraction at 90-100°C 2-3 times, 1-1.5 h each time; collecting a filtrate through filtering, leaving the filtrate to stand for cooling, and centrifuging the filtrate at 1,500-3,500 rpm for 5-10 min, collecting supernatant, and vacuum-recovering ethanol to obtain the Linderae Radix extract through drying; a preparation method for the Dendrobium officinale polysaccharide comprises the following steps: adding Dendrobium officinale to 85-90% (volume concentration) ethanol in a volume of 8-12 times a volume of the Dendrobium officinale, and conducting reflux extraction at 85-90°C 2-3 times, 1-1.5 h each time; collecting a residue through filtering, drying the residue, adding water in a volume of 8-12 times the volume of the residue, conducting reflux extraction 2-3 times, 1-1.5 h each time, and combining filtrates; vacuum-concentrating the solution to 1/3-1/2 of the original volume, adding anhydrous ethanol until the alcohol concentration of the solution reaches 80-85%, placing the solution in a 4°C refrigerator for 24-36 h, centrifuging the solution at 1,500-3,500 rpm for 5-10 min, discarding supernatant, collecting precipitate and dissolving the precipitate in 60-80°C hot water at a ratio of 1.0-2.0 g/ml, repeating the alcohol precipitation process 1-3 times, collecting precipitate through centrifugation, and washing the precipitate with 80-90% ethanol 2-3 times to obtain crude Dendrobium officinale polysaccharide.
2. The application of claim 1, wherein a purity of the prepared Dendrobium officinale polysaccharide is 70-85%.
3. The application of claim 1, wherein a mass content of the Linderae Radix composition in prepared drugs for treating inflammatory bowel diseases is 1-100%. 8
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