CN108066349A - Applications of IV a of panax japonicus saponin in treatment ephritis or kidney fibrosis drug is prepared - Google Patents
Applications of IV a of panax japonicus saponin in treatment ephritis or kidney fibrosis drug is prepared Download PDFInfo
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- CN108066349A CN108066349A CN201711487645.2A CN201711487645A CN108066349A CN 108066349 A CN108066349 A CN 108066349A CN 201711487645 A CN201711487645 A CN 201711487645A CN 108066349 A CN108066349 A CN 108066349A
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- panax japonicus
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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
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- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07H—SUGARS; DERIVATIVES THEREOF; NUCLEOSIDES; NUCLEOTIDES; NUCLEIC ACIDS
- C07H1/00—Processes for the preparation of sugar derivatives
- C07H1/06—Separation; Purification
- C07H1/08—Separation; Purification from natural products
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- C07—ORGANIC CHEMISTRY
- C07H—SUGARS; DERIVATIVES THEREOF; NUCLEOSIDES; NUCLEOTIDES; NUCLEIC ACIDS
- C07H15/00—Compounds containing hydrocarbon or substituted hydrocarbon radicals directly attached to hetero atoms of saccharide radicals
- C07H15/20—Carbocyclic rings
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- C07J63/00—Steroids in which the cyclopenta(a)hydrophenanthrene skeleton has been modified by expansion of only one ring by one or two atoms
- C07J63/008—Expansion of ring D by one atom, e.g. D homo steroids
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Abstract
The invention discloses new opplications of IV a of panax japonicus saponin in treatment ephritis or kidney fibrosis drug is prepared.IV a of panax japonicus saponin is the extract of Tai Bai Aralia woods, and inventor, which studies, finds that IV a of panax japonicus saponin has renal protection.IV a extracting methods of panax japonicus saponin are simple, the expression of the MCP 1,6 inflammatory factor of TNF α, IL of the NRK 49F cells of the inductions of TGF β 1 are can inhibit, reduces the kidney region fibrosis of UUO mouse, ephritis and kidney fibrosis, significant effect caused by a variety of causes can be treated.
Description
Technical field
It is used to prevent and treat the new application of ephritis or kidney fibrosis the present invention relates to IV a of panax japonicus saponin.
Background technology
Chronic renal failure（CRF）One of major disease for influencing global human health is become.Kidney region fibrosis
It is one of Chinese pathology feature of CRF.How to be the key that prevention CRF using the development of pharmaceutical intervention kidney region fibrosis.Kidney
Fibrosis is the necessary approach of CRF, and various kidney troubles are developed to the co-channel of End-stage renal disease.It is it is now recognized that last eventually
The pathological change of phase kidney trouble is irreversible, and therefore, the occurrence and development for controlling kidney fibrosis are most important.The hair of kidney fibrosis
Interpretation of the cause, onset and process of an illness system is extremely complex, is too many levels, the multifactor slow process to interact, and main mechanism includes：Inflammatory reaction, oxidation
Stress, the activation of Apoptosis, myofibroblast and the synthesis of multiplication, extracellular matrix it is unbalance etc. with degrading.
Clinically mainly include for the treatment of kidney fibrosis at present：The treatment (such as obstructive nephropathy) of protopathy, phase
Close risk factor elimination and intervention (such as infect, drug) and for pathogenesis treatment (such as non-steroidal anti-inflammatory drug,
Antihypertensive drugs).Although above-mentioned therapy can play kidney fibrosis certain therapeutic effect, can not prevent completely slow
Property nephrosis was gradually in progress to whole latter stage.Since the pathogenesis of fibrotic disease is complicated, it is more to be related to factor, searching one is needed therefore
Kind can resist the drug of ephritis and kidney fibrosis.
The content of the invention
The object of the present invention is to provide the new application of IV a of panax japonicus saponin, which is that IV a of panax japonicus saponin is used to make
The drug of standby treatment ephritis or kidney fibrosis.
Realization process of the present invention is as follows：
Following structural formula（I）Applications of shown IV a of panax japonicus saponin in treatment ephritis or kidney fibrosis drug is prepared,
（I）.
Above-mentioned treatment kidney fibrosis drug can be conventional capsule, tablet or pill.
Above-mentioned IV a of panax japonicus saponin is prepared by the following method to obtain：
（1）The too white Aralia wood medicinal powder of 20 kg adds in 10 times of 80% ethyl alcohol of amount of mass volume ratio, and 60 min of refluxing extraction at 80 DEG C is returned
Stream extraction 3 times, filtering merge, and concentration and recovery ethyl alcohol obtains coarse extract；
（2）By step（1）Coarse extract disperseed with distilled water, remove oil-soluble impurities with the chloroform extraction of 3 times of volumes respectively, then
With the water saturated extracting n-butyl alcohol 3 times of 3 times of volumes, merge 3 extract liquors, be recovered under reduced pressure at 60 DEG C, steamed in 60 DEG C of water-baths
It is dry；
（3）By step（2）Obtained n-butyl alcohol extract utilizes silica gel column chromatography, Sephadex LH-20 column chromatographys, ODS reverse phases
Column chromatography, Lobar column chromatographys and reversed-phase HPLC preparation means are separated, and obtain IV a of panax japonicus saponin；
（4）Tai Bai Aralia wood saponin(e molecular weight is larger, and structure is complex.Utilize 1D NMR (1H and 13C NMR), 2D NMR
The spectroscopic techniques such as (DQCOSY, TOCSY, HMQC, HMBC, NOESY and ROESY) and MS (ESI-MS) are to isolated soap
Glycosides ingredient carries out structure elucidation, and with reference to chemical means（Sour water solution, basic hydrolysis are digested, methylated with acetylation etc.）It determines
Its plane and stereochemical structure.
Tai Bai Aralia woods (A ralia taibaiensisZ. Wang et H. C. Zheng) specialty is in China Chinese and Western
Portion's Qin_Ba mountain areas and remaining arteries and veins, main chemical compositions are triterpenoid saponin, and aglycon is oleanolic acid, is mainly deposited in the form of into glycosides
.IV a of panax japonicus saponin is the extract of Tai Bai Aralia woods, and inventor has been surprisingly found that IV a of panax japonicus saponin has kidney by research
Protective effect.IV a extracting methods of panax japonicus saponin are simple, can inhibit MCP-1, TNF- of the NRK-49F cells of the induction of TGF-β 1
The expression of α, IL-6 inflammatory factor reduces the kidney region fibrosis of UUO mouse, treats ephritis and kidney fiber caused by a variety of causes
Change, significant effect.
Description of the drawings
Fig. 1 is the block diagram of each group animal renal function index；
Fig. 2 is the block diagram of each group animal renal function index；
Fig. 3 is the microphoto of the nephridial tissue HE stained slices of each group animal, including sham-operation group, model group, and low dose group, in
Dosage group, high dose group；
Fig. 4 is the microphoto of the nephridial tissue Masson stained slices of each group animal, including sham-operation group, model group, low dosage
Group, middle dose group, high dose group；
The MCP-1 for the NRK-49F cells that Fig. 5 induces for TGF-β 1, TNF-α, the block diagram of the expression of IL-6 inflammatory factors.
Specific embodiment
Embodiment 1 (prepares IV a of panax japonicus saponin)
The too white Aralia woods of 20kg is taken to add in 10 times of amount volumetric concentrations to extract 3 times for 80% alcohol reflux, the refluxing extraction at 80 DEG C
60min, 60 DEG C are concentrated under reduced pressure into relative density as 1.01-1.05 Tai Bai Aralia wood crude extracts；Crude extract distilled water is disperseed, point
It Yong not chloroform 1:3 (v/v) extractions remove oil-soluble impurities, with water saturated extracting n-butyl alcohol 3 times (1:3, v/v) 3, are merged
Secondary extract liquor is recovered under reduced pressure at 60 DEG C with Rotary Evaporators, is evaporated in 60 DEG C of water-baths.N-butyl alcohol extract utilizes silicagel column
The means such as chromatography, Sephadex LH-20 column chromatographys, ODS reversed-phase column chromatographies, Lobar column chromatographys and reversed-phase HPLC preparation are divided
From.Utilize 1D NMR (1H and 13C NMR), 2D NMR (DQCOSY, TOCSY, HMQC, HMBC, NOESY and ROESY) and MS
(ESI-MS) etc. spectroscopic techniques carry out isolated saponin constituent structure elucidation, and with reference to chemical means（Sour water solution, alkali
Hydrolyze, digest, methylating with acetylation etc.）Its plane and stereochemical structure is determined, obtains compound panax japonicus saponin IV
a。
Embodiment 2 (capsule)
The panax japonicus saponin IV a powder 10mg for weighing prepared by embodiment 1, starch 50-100mg, microcrystalline cellulose 50-70mg are interior
Add dextrin 70mg, lactose 70mg, sodium carboxymethylcellulose 7mg, cross 60 mesh sieves, be uniformly mixed, be wetting agent system with 85% ethyl alcohol
Softwood, excessively 40 mesh sieves, 60 DEG C of 5 h of oven drying, 40 mesh sieve whole grains, 60 mesh sieves weed out fine powder to get 40-60 mesh particles.
The critical relative moisture of particle selects No. 1 capsule below 78%（0.50 ml）Be filled to get.
Embodiment 3 (tablet)
Using the technique of wet granulation, IV a powder of panax japonicus saponin prepared by embodiment 1, lactose, microcrystalline cellulose, carboxylic are weighed
The mass ratio of sodium carboxymethylcellulose pyce and magnesium stearate is：10：25:83:6:0.8.IV a powder of panax japonicus saponin is crossed into 100 mesh sieves,
Lactose, microcrystalline cellulose, sodium carboxymethylcellulose and magnesium stearate are crossed into 60 mesh sieves respectively, obtained uniform powder is put clean respectively
In net container.The soft ability of 75% ethyl alcohol system, soft ability are pelletized with 18-24 mesh sieves, and particle puts 60 DEG C of dry 2-4h, 18-24 mesh sieve whole grains,
Additional superfine silica gel powder and magnesium stearate mixing, tabletting is to get label.
Embodiment 4 (pill)
The prescription of screening is：Drug：Matrix mass ratio is 1：150, PEG4000:PEG6000 mass ratioes be 2 ︰ 1,85 DEG C of material temperature,
Drop is away from 5cm, and 10 DEG C of cooling temperature, condensate liquid is vegetable oil.The panax japonicus saponin IV of the preparation of embodiment 1 is weighed by recipe quantity precision
A is in appropriate purified water, after dissolving, adds in the mixture of PEG4000 and PEG6000, is placed in 80 DEG C or more of magnetic stirring apparatus
In water-bath, to after all melting, the two is uniformly mixed heating stirring.It is instilled with dropper in dripping pill device, controls suitable drop
Speed makes it fall in condensate liquid, treats its cooling and shaping, takes out dripping pill, with filter paper throw away the refrigerant, naturally dry selects ball.I.e.
.
Embodiment 5 (effect experiment)
Wild type, male C57BL/6,18-20g, adaptability are fed one week, are randomly divided into sham-operation group, model group, low dosage
Group, middle dose group, high dose group, every group of equal 8 rat.In addition to sham-operation group, ligation of ureter on the left of remaining four groups of row
(UUO) kidney fibrosis model is constructed.Pre-operative anxiety 12h, can't help water, with 3.5% chloraldurate（350mg/kg）Intraperitoneal injection fiber crops
After liquor-saturated, prone position is fixed on surgical plate, preserved skin under the right rib in back, conventional iodophor disinfection operating field skin, other on the right side of backbone to open
0.5cm along costal margin downlink stringer notch, successively cuts integumentary musculature, exposure simultaneously separates right side ureter, with 4-0 suture rows
It ligatures twice, observation surgical field of view is without bleeding, layer-by-layer suture subcutaneous tissue and skin.Sham-operation group only Ureter dissection,
It does not ligature, remaining surgical procedure is identical.IV a intervention groups postoperative half an hour of panax japonicus saponin starts to give panax japonicus saponin IV a 10
Mg/kg/d, 20 mg/kg/d and 40 mg/kg/d gavages, continuous 14 days.Sham-operation and model group are given isometric solvent and are filled
Stomach, each group mouse were put to death respectively in postoperative 14th day on request.The 14th day after modeling success, each group mouse is put to death, is received respectively
Collect Bilateral Renal tissue samples.With 3.5% chloraldurate（350mg/kg）It is fixed after intraperitoneal injection of anesthesia mouse.Use blood taking needle neck
Arterial blood drawing, each 1ml are spare after centrifugation.Abdomen center line position successively splits thoracic cavity, abdominal cavity, exposure heart and Bilateral Renal
It is dirty.Atrium dextrum is cut off in eye scissors side, and catheter needle penetrates left ventricle and fixation, with the physiological saline of precooling（About 200ml）It is filled
Note, after bilateral renal integral color bleaches, cuts off bilateral renal, and immerses in the physiological saline of precooling at once.Rapid removal
After adipose tissue and kidney peplos, row longitudinal cut is to hemisection respectively for bilateral renal, and then part kidney, inserts -80 DEG C of refrigerators and use
In Immunofluorescence test, after part nephridial tissue puts into liquid nitrogen immediately, -80 DEG C of refrigerators are stored in for Western Blotting
And PCR detections, part nephridial tissue, which is immersed in 10% formalin solution, fixes, for light microscope inspection.
First, IV a of panax japonicus saponin is to the renal function of UUO rat kidney fibrosis
The blood serum sample of each group rat is taken, serum creatinine content (Nanjing in rat blood serum is detected according to serum creatinine kit specification
Build up Bioengineering Research Institute).Each group mouse mouse urine is taken, (biological work is built up in Nanjing according to urea nitrogen and Urine proteins kit
Journey research institute) specification operation, measure urea nitrogen and urine protein content.As a result (table 1-3, Fig. 1) shows：Model group is compared with sham-operation
Group have significant difference (p<0 .01), basic, normal, high dosage administration group can significantly reduce UUO mouse kidney fibrosis serum creatinine,
Urea nitrogen, urine volume expression (p<0 .01), show that IV a of panax japonicus saponin has certain protective effect to damage kidney.
2nd, influences of IV a of panax japonicus saponin to UUO Rat renal fibrosing kidneys indexes
Each group mouse peritoneal injects 3.5% chloral hydrate anesthesia, takes out separated mouse kidney tissue and weighs, and it is left to calculate kidney
Kidney and weight ratio.As a result (table 4-5, Fig. 2) shows：Model group compared with blank group renal index significant difference (p<0 .01)；In,
High dose administration group can significantly reduce left kidney index, the expression of Urine proteins (p<0 .01)；Prompt panax japonicus saponin IV a
There is certain inhibitory action to kidney fibrosis caused by UUO models.
3rd, the inflammatory reaction and kidney fibrosis that panax japonicus saponin IV a generates renal tissue after unilateral ureteral obstruction
It influences.
The another left side nephridial tissue for taking each group mouse, crosscutting half are placed in 10% formalin solution and fix, and HE is respectively adopted
Decoration method and Masson decoration methods, as a result (Fig. 3-4) show：In HE stained slices, each UUO groups nephridial tissue is shown significantly
Interstitial inflammation and fibrosis, renal interstitial gap substantially expand, inflammatory cell infiltration, glomerular capillary expansion, and renal tubule is bad
Extremely, gradual atrophy.UUO renal tissues of rats degree of injury can be improved after giving the treatment of IV a various doses of panax japonicus saponin, with high agent
Amount is the most apparent.Masson dyes display sham-operation group collagenous fibres without hyperplasia, and tubular substrate film is high-visible.UUO groups, can
See a large amount of collagen stainings of nephridial tissue, collagenous fibres substantially increase in renal interstitial, and interstitial fibers tissue is in pencil and netted hyperplasia
It is formed.After giving the treatment of IV a various doses of panax japonicus saponin, UUO renal tissues of rats kidney fibrosis degree mitigates, and prompts panax japonicus
IV a of saponin(e can be effectively improved the kidney region fibrosis generated after nephridial tissue obstruction.
4th, the influence for the NRK-49F Cellular inflammatory factor expressions that IV a of panax japonicus saponin induces TGF-β 1
Cell discards cell culture medium, with cold PBS rinsings once.37 DEG C of 1-2min of pancreatin digestive juice.Complete medium is added to neutralize
Pancreatin, 1000rpm 6min, abandons supernatant.Blood cell counting plate counts, and spreads 96 orifice plates, per hole 1x104A cell.Be placed on 37
DEG C, 5%CO2It is cultivated for 24 hours in cell incubator.It is pre-processed respectively with the IV a solution of panax japonicus saponin of 20,40 and 80 μ g/ml thin
Born of the same parents 1h.Then with TGF-β 1 (10ng/ml) processing for 24 hours, cell culture supernatant is collected.Using monocyte chemotactic factor -1
(MCP-1) ELISA kit (U.S. R＆D), human tumor necrosis factor-alpha (TNF-α) ELISA kit (U.S. R＆D) and
Human interleukin-6 (IL-6) ELISA kit (U.S. R＆D) detects MCP-1 in cell liquid, TNF-α and IL-6's respectively
Concentration.As a result (table 6-8, Fig. 5) shows：MCP-1, TNF-α in model group, IL-6 significantly raise (p<0 .01), panax japonicus saponin
The basic, normal, high dosage groups of IV a can significantly inhibit TGF-β 1 induction cell MCP-1, TNF-α, IL-6 levels rise (p<0
.01), IV a of panax japonicus saponin, which can effectively inhibit the expression of inflammatory factor, to be shown without influence on normal cell.
Claims (3)
1. structural formula（I）Applications of shown IV a of panax japonicus saponin in treatment ephritis or kidney fibrosis drug is prepared,
（I）.
2. application according to claim 1, it is characterised in that：The treatment kidney fibrosis drug is conventional capsule
Agent, tablet or pill.
3. application according to claim 1 or 2, it is characterised in that：IV a of panax japonicus saponin is by following methods system
：
（1）The too white Aralia wood medicinal powder of 20 kg adds in 10 times of 80% ethyl alcohol of amount of mass volume ratio, and 60 min of refluxing extraction at 80 DEG C is returned
Stream extraction 3 times, filtering merge, and concentration and recovery ethyl alcohol obtains coarse extract；
（2）By step（1）Coarse extract disperseed with distilled water, remove oil-soluble impurities with the chloroform extraction of 3 times of volumes respectively, then
With the water saturated extracting n-butyl alcohol 3 times of 3 times of volumes, merge 3 extract liquors, be recovered under reduced pressure at 60 DEG C, steamed in 60 DEG C of water-baths
It is dry；
（3）By step（2）Obtained n-butyl alcohol extract utilizes silica gel column chromatography, Sephadex LH-20 column chromatographys, ODS reverse phases
Column chromatography, Lobar column chromatographys and reversed-phase HPLC preparation means are separated, and obtain IV a of panax japonicus saponin.
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| CN201711487645.2A CN108066349B (en) | 2017-12-30 | 2017-12-30 | Application of panax japonicus saponin IVa in preparing medicine for treating nephritis or renal fibrosis |
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109336942A (en) * | 2018-10-23 | 2019-02-15 | 吉林大学 | A method of the extraction purification panax japonicus saponin VI from ginseng |
Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102824355A (en) * | 2012-08-22 | 2012-12-19 | 中国人民解放军第四军医大学 | Use of panax japonicus saponin IV a in preparation of alpha-glycosidase inhibitor |
-
2017
- 2017-12-30 CN CN201711487645.2A patent/CN108066349B/en not_active Expired - Fee Related
Patent Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102824355A (en) * | 2012-08-22 | 2012-12-19 | 中国人民解放军第四军医大学 | Use of panax japonicus saponin IV a in preparation of alpha-glycosidase inhibitor |
Non-Patent Citations (3)
| Title |
|---|
| H WANG, ET AL.: "Inhibitory effects of Chikusetsusaponin IVa on lipopolysaccharide-induced pro-inflammatory responses in THP-1 cells", 《INTERNATIONAL JOURNAL OF IMMUNOPATHOLOGY AND PHARMACOLOGY》 * |
| 吴忻晶 等: "竹节参皂苷IVa及其衍生物的药理活性及作用机制研究进展", 《中国药房》 * |
| 盖丰丰 等: "补肾益气类方药对慢性肾脏病中IL-6干预作用的研究进展", 《中国中西医结合肾病杂志》 * |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109336942A (en) * | 2018-10-23 | 2019-02-15 | 吉林大学 | A method of the extraction purification panax japonicus saponin VI from ginseng |
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