CN101209254B - New use of polyhydroxy galloyl-beta-D-glucose derivatives - Google Patents
New use of polyhydroxy galloyl-beta-D-glucose derivatives Download PDFInfo
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Abstract
The invention relates to a usage of polyhydroxy galloyl-Beta-D-glucose derivatives or the medicinal salts in anti-helicobacter pylori, and the invention also relates to a usage of drugs which contain at least one extract of polyhydroxy galloyl-Beta-D-glucose derivative used for the prevention and/or treatment of diseases which are caused by helicobacter pylori.
Description
Technical field
The invention belongs to medical technical field; the new purposes that relates to the polyhydroxy galloyl-beta-D-glucose derivant is specifically related to the polyhydroxy galloyl-beta-D-glucose derivant or contains a kind of purposes of extract in anti-helicobacter pylori of this derivant at least.
Background technology
Nineteen eighty-three Australia scholar's Warren (Waren) and Marshall (Marshall) are isolated helicobacter pylori (Helicobacter pylori, abbreviation Hp) first from gastric mucosa.After studies show that in a large number: Helicobacter pylori infection is relevant with the disease of many serious harm human healths, as chronic active gastritis, gastric ulcer, duodenal ulcer, gastric mucosa dependency lymphoid tissue (MALT) malignant lymphoma and gastric cancer etc.Data shows, the whole world has the population more than 50% to infect helicobacter pylori approximately, the infection of developing country is particularly serious, and wherein adult's infection rate reaches 60～80%, 1994 international cancer research institution (IARC) classify helicobacter pylori as human I class carcinogen.At present, a line scheme of the treatment Helicobacter pylori infection of Tui Jianing mainly is based on proton pump inhibitor (PPI) or bismuth in the world, in conjunction with the triple therapy of two kinds of antibiotic compositions.Wait to solve but still exist a lot of problems in these western medicine schemes, mainly show: the expense height of western medicine, drug side effect height, eradication rate instability, easily drug resistance etc. takes place, influenced the curative effect and the application of these schemes to a great extent.Therefore, seek anti-Hp and infect the research topic that has become people's extensive concerns with treatment with the new way of its relevant disease.
The polyhydroxy galloyl-beta-D-glucose derivant is the known compound of class wide material sources; its multiple biological activity is reported in succession; as remove the activity that free radical, antioxidant activity, anti-SARS virus infect, anti-HCV NS3 protease activities, apoptotic activity, angiotensin transferase inhibitor activity, cell cycle inhibitor activity, as the tyrosine kinase inhibitor isoreactivity; but up to now; in the world wide, there is no its active any report in anti-helicobacter pylori aspect.
Summary of the invention
The object of the present invention is to provide structure suc as formula the polyhydroxy galloyl-beta-D-glucose derivant shown in the I or its pharmaceutical salts in the purposes aspect the anti-helicobacter pylori.
Wherein, R
1, R
2, R
3, R
4, R
5Be H or G or GG or GGG independently of one another, and R
1, R
2, R
3, R
4, R
5Be not H simultaneously.G, GG and GGG are defined as follows:
The preferred following chemical compound 1～13 of described polyhydroxy galloyl-beta-D-glucose derivant:
Chemical compound 1:
Chemical compound 2:
Chemical compound 3:
Chemical compound 4:
Chemical compound 5:
Chemical compound 6:
Chemical compound 7:
Chemical compound 8:
Chemical compound 9:
Chemical compound 10:
Chemical compound 11:
Chemical compound 12:
Chemical compound 13:
Described polyhydroxy galloyl-beta-D-glucose derivant most preferred compound 2:
Another object of the present invention is to provide the method for the disease that a kind of prevention or treatment helicobacter pylori cause; this method comprises polyhydroxy galloyl-beta-D-glucose derivant or its pharmaceutical salts that gives the patient treatment required dosage; the disease that described helicobacter pylori causes includes but not limited to gastritis, gastric ulcer, gastric erosion, duodenal ulcer and gastric cancer; described polyhydroxy galloyl-beta-D-glucose derivant is the chemical compound shown in the formula I; preferred compound 1～13, most preferred compound 2.
" pharmaceutical salts " of the present invention do not limit especially, can represent any medicinal salt that can be used in, for example inorganic salts such as hydrochlorate, sulfate, carbonate, bicarbonate, hydrobromate, hydriodate; And organic carboxylate such as acetate, maleate, lactate, tartrate, three fluoro acetate; Organic sulfonates such as mesylate, hydroxyl mesylate, isethionate, benzene sulfonate, toluene fulfonate, taurine; Front three amine salt, triethylamine salt, pyridiniujm, picolyl salt, hexanamine salt, N, N ,-dibenzyl ethylenediamine salt, N-methylglucosamine salt, diethanolamine salt, triethanolamine salt, three (hydroxyl methylamino) methane salt, phenethyl benzylamine salt etc.; Amino acid salts such as arginine salt, lysinate, serine salt, glycinate, aspartate, glutamate, Glu.
Polyhydroxy galloyl-beta-D-glucose derivant provided by the invention or its salt have all shown good anti-helicobacter pylori effect in external, in vivo test.External anti-Hp test shows part bacterial strain to The compounds of this invention more responsive than the ampicillin.The in vivo test result shows, and is suitable with clinical " triple therapy " commonly used effect at present to the scavenging action of helicobacter pylori.The gastritis, the gastric ulcer result of experiment that cause at anti-Hp show that the gastric mucosa injury that chemical compound of the present invention causes helicobacter pylori has obvious repair.
The invention solves current weak point for the treatment of the disease that helicobacter pylori causes clinically, as the medical expense height, poisonous side effect of medicine height, eradication rate instability, defectives such as drug resistance easily take place, thereby a kind of wide material sources, with low cost, therapeutic activity composition that toxic and side effects is low are provided.
Polyhydroxy galloyl-beta-D-glucose derivant provided by the invention or its pharmaceutical salts have good anti-helicobacter pylori activity, and the gastric mucosa injury that helicobacter pylori causes is had repair.For diseases such as treatment gastritis, gastric ulcer, gastric erosion, duodenal ulcer and gastric cancer provide wide material sources, with low cost, active ingredient that toxic and side effects is low,, provide good idea for solving the difficult problem in this type of disease of current treatment.
A further object of the present invention is to provide a kind of plant extract of anti-Helicobacter pylori infection, and described extract comprises at least a formula I compound or pharmaceutically acceptable salt thereof, preferred compound 1～13 or its pharmaceutical salts, most preferred compound 2 or its pharmaceutical salts.
Following table is the source about 13 structural formulas polyhydroxy galloyl-beta-D-glucose derivant as shown in Equation 1:
Formula I;
Wherein, R
1, R
2, R
3, R
4, R
5Be H or G or GG or GGG independently of one another, and R
1, R
2, R
3, R
4, R
5Be not H simultaneously.G, GG and GGG are defined as follows:
| Numbering | R 1 | R 2 | R 3 | R 4 | R 5 | The source |
| Chemical compound 1 | G? | G? | G? | G? | G? | Galla Chinensis |
| Chemical compound 2 | G? | G? | G? | H? | G? | Herba Erodii |
| Chemical compound 3 | G? | G? | G? | G? | H? | Rhodiola kirilowii (Regel) Maxim. |
| Chemical compound 4 | G? | H? | G? | H? | G? | The Fructus Chebulae |
| Chemical compound 5 | G? | G? | H? | H? | G? | Galla Chinensis |
| Chemical compound 6 | G? | H? | G? | G? | H? | Herba Saxifragae melanocentrae |
| Chemical compound 7 | GG? | G? | G? | G? | H? | Herba Saxifragae melanocentrae |
| Chemical compound 8 | H? | H? | G? | G? | G? | Chemosynthesis |
| Chemical compound 9 | H? | H? | G? | H? | G? | Chemosynthesis |
| Chemical compound 10 | H? | H? | H? | H? | G? | Chemosynthesis |
| Chemical compound 11 | H? | H? | G? | H? | H? | Chemosynthesis |
| Chemical compound 12 | G? | G? | H? | G? | G? | Radix Ampelopsis |
| Chemical compound 13 | G? | H? | H? | H? | G? | The angle bean |
Polyhydroxy galloyl-beta-D-glucose derivant wide material sources; it is present in the various plants, for example Herba Erodii (Geranium dahuricum) Galla Chinensis (Galla chinese), Fructus Chebulae (Terminaliachebula), Herba Saxifragae melanocentrae (Saxifraga melanocentra) and Rhodiola kirilowii (Regel) Maxim. (Rhodiolakirilowii), Herba Erodii (Geranium wilfordii) etc.It can extract by methods known in the art; say for example; the general preparation method of polyhydroxy galloyl-beta-D-glucose derivant is: make water; the lower aliphatic alcohols; aqueous lower aliphatic alcohols; the aromatic series alcohols; rudimentary ketone (acetone etc.); halogen-containing solvent and their mixed solvent about 0 ℃ to boiling spread; in decompression; normal pressure or add herb or its position RUGEN of depressing the plant that contains the polyhydroxy galloyl-beta-D-glucose derivant; stem; leaf; flower etc. extracts; can obtain containing the extract of at least a polyhydroxy galloyl-beta-D-glucose derivant, extract is carried out the polyhydroxy galloyl-beta-D-glucose derivant that various separation and purifications just can obtain relating among the present invention.The polyhydroxy galloyl-beta-D-glucose derivant can also be carried out chemical reaction by glucose and gallic acid and be obtained.
The polyhydroxy galloyl-beta-D-glucose derivant also can obtain by synthetic.
The extraction separation of polyhydroxy galloyl-beta-D-glucose derivant and synthetic method also can be with reference to following documents:
1. what is strong; Stone is green
*Yao opens; Luo Yi; Lv Yuanping.The esterification research of gallic acid and glucose, chemical research and application, 2001,13 (5): 550-553.
2. what is strong; Stone is green
*Yao opens; Sieve is proper; Lv Yuanping.6-O-3,6-two-O-3,4, the study on the synthesis of 6-three-O-Galla Turcica (Galla Helepensis) acyl-D-glucose.Sichuan University's journal (engineering science version), 2002,34 (3): 110-113.
3. what is strong; Stone is green
*Yao opens; Lv Yuanping.The esterification research (II) of gallic acid and glucose, chemical research and application, 2002,14 (6): 665-667.
4. Yu Wen wins; Chen new people; Yang Lei; Lee's word flies.Radix Ampelopsis tannin The Chemical Constituents, research and development of natural products, 1995,7 (1): 15-18.
5. fourth hilllock; Liu Yanze; Song Maoping; Zou Dapeng; Sheng Longsheng.Polyatomic phenol composition among the Fructus Chebulae, China Medicine University's journal, 2001,32 (3): 193-196.
6. Xu Xiao outstanding person; Duan Deliang; A left side is state-run; Li Zhengquan; Chen Lirong.The new purposes of polyhydroxy galloyl-β-D-glucosan derivative, Chinese patent application CN200410042882.4 (publication number: CN1704064A).
7. Xu Xiao outstanding person; Deng Hongkui; Chen Lirong; Ding Mingxiao, etc.Suppress effective components of Chinese medicinal and biological activity determination method thereof that sars coronavirus infects, Chinese patent application CN200310101980.6 (publication number: CN1602853A).
8.Deliang?Duan；Zhengquan?Li；Hongpeng?Luo；Wei?Zhang；Lirong?Chen?andXiaojie?Xu.Antiviral?compounds?from?traditional?Chinese?medicines?GallaChinese?as?inhibitors?of?HCV?NS3?protease，Bioorganic?&?Medicinal?ChemistryLetters，2004(14)：6041-6044。
9.R.W.Owen；R.Haubner；W.E.Hull；G.Erben；B.Spiegelhalder；H.Bartsch；B.Haber.Isolation?and?structure?elucidation?of?the?major?individual?polyphenols?incarob?fibre，Food?and?Chemical?Toxicology，2003(41)：1727-1738。
9.Gen-ichiro?Nonaka；Itsuo?Nishioka.Tannins?and?Related?Compounds.X.Rhubarb(2)：Isolation?and?Structures?of?a?Glycerol?Gallate，Gallic?AcidGlucoside?Gallates，Galloylglucoses?and?Isolindleyin，Chem.Pharm.Bull.，1983，31(5)：1652-1658。
10.Makoto?Nishizawa；Takashi?Yamagishi；Gen-ichiro?Nonaka；Itsuo?Nishioka；Tetsuro?Nagasawa；Hikokichi?Oura.Tannins?and?Related?Compounds.VII.Isolation?and?Characterization?of?Galloylglucoses?from?Paeoniae?Radix?andTheir?Effect?on?Urea-Nitrogen?Concentration?in?Rat?Serum，Chem.Pharm.Bull.，1983，31(8)：2593-2600。
The specific embodiment:
The following example is used for the content of the present invention of further explaining, but does not mean that the present invention is done any restriction.
Embodiment 1 prepares chemical compound 2 by Herba Erodii
Get thick root Herba Erodii (Geranium dahuricum) 2000g, pulverize, add the ethanol of 8 times of amounts 40%, reflux, extract, three times, each 2 hours.The elimination medicinal residues merge 3 extracting solution, and concentrating under reduced pressure gets extractum 280g, add the suitable quantity of water dissolving after, use petroleum ether (500ml * 4), ethyl acetate (500ml * 4), n-butyl alcohol (500ml * 4) to extract successively.Acetic acid ethyl acetate extract is merged, be evaporated to dried, divide and to pull on silicagel column (silica gel with 100～200 purpose silica gel G), use chloroform successively, chloroform: methanol=10: 1, chloroform: methanol=8: 1, chloroform: methanol=5: 1, chloroform: methanol=2: 1, methanol carries out eluting, detects with thin layer chromatography.With chloroform: the eluent of methanol=5: 1 merges, and is concentrated into dried.Gains are used gel column (Sephadex LH-20) chromatography repeatedly, and methanol is done eluent, and thin layer chromatography follow the tracks of to detect, and eluent merges, and concentrate, obtain purified chemical compound 2 after the drying.
Embodiment 2 prepares chemical compound 1 and 5 by Galla Chinensis
After Galla Chinensis 500g pulverizes, extract three times with 50% ethanol-water solution 2000ml room temperature merceration, soaked 24 hours at every turn, the filtrate decompression of extraction concentrates and boils off ethanol, obtains crude extract 260g.Use petroleum ether (500ml * 4) successively, chloroform (500ml * 4), ethyl acetate (500ml * 4), n-butyl alcohol (500ml * 4) extracts crude extract, is divided into a plurality of parts.Ethyl acetate is partly merged, concentrate evaporate to dryness, divide and pull on gel column (Sephadex LH-20), water successively, methanol: water=2: 8, methanol: water=4: 6, methanol: water=6: 4, methanol: water=8: 2, methanol, acetone carries out gradient elution, detects with thin layer chromatography, obtains 6 components.
6 components respectively with the polyamide column column chromatography that reduces pressure, are used ethyl acetate, ethyl acetate: methanol=10: 1 successively, ethyl acetate: methanol: water=10: 1: 0.5, ethanol: water=6: 4, ethanol: water=8: 2, ethanol, acetone: water=6: 4, acetone: water 8: 2, acetone carries out gradient elution, and follows the tracks of eluate with thin layer chromatography, attached gel post (Sephadex LH-20) chromatography, water successively, methanol: water 2: 8, methanol: water 4: 6, methanol: water 6: 4, methanol: water 8: 2, methanol, acetone carries out gradient elution, eluate of the same race merges concentrated, after the drying, obtain purified chemical compound 1 and 5.
Embodiment 3 prepares chemical compound 3 by Rhodiola kirilowii (Regel) Maxim.
Rhodiola kirilowii (Regel) Maxim. 3kg pulverizes, extract three times with 10000ml 80% soak with ethanol, the each immersion week, the crude extract concentrating under reduced pressure, boil off ethanol after, get extractum 300g, add the suitable quantity of water dissolving, use petroleum ether (600ml * 5) successively, chloroform (600ml * 5), ethyl acetate (600ml * 5), n-butyl alcohol (600ml * 5) extraction.Organic layer evaporated under reduced pressure respectively gets petroleum ether part 23g, chloroform part 17g, ethyl acetate part 48g, n-butyl alcohol part 100g.Ethyl acetate and n-butyl alcohol are partly gone up polyamide column, and the gained fraction is continued to obtain purified chemical compound 3 by means purification such as gel filtration chromatographies.
Embodiment 4 prepares chemical compound 4 by the Fructus Chebulae
After Fructus Chebulae's fruit 2kg pulverized, with 80% ethanol-water solution 5000mL, the room temperature merceration extracted three times, soaks a week at every turn, and the filtrate decompression of extraction concentrates and boils off ethanol, obtains crude extract 552g.With petroleum ether (600ml * 4), chloroform (600ml * 4), ethyl acetate (600ml * 4), n-butyl alcohol (600ml * 4) extracts crude extract and is divided into a plurality of parts, the evaporate to dryness organic solvent obtains petroleum ether part 25g, chloroform part 30g, ethyl acetate part 53g, n-butyl alcohol part 234g.N-butyl alcohol part (every crowd of 20g) in batches goes up polyamide column.Use chloroform successively: methanol: formic acid 100: 10: 1, chloroform: methanol: formic acid 100: 15: 1, chloroform: methanol: formic acid 60: 40: 1, methanol: formic acid carries out gradient elution at 50: 50: 1, and follow the tracks of eluate with thin layer chromatography, eluate of the same race merges and concentrates, after the drying, obtains purified chemical compound 4.
Embodiment 5 prepares chemical compound 6 and 7 by Herba Saxifragae melanocentrae
Herba Saxifragae melanocentrae 1kg pulverizes, and extracts three times with 4000ml 80% soak with ethanol, soaks a week at every turn, and the crude extract concentrating under reduced pressure boils off ethanol, gets extractum 205g, adds the suitable quantity of water dissolving.Use petroleum ether (200ml * 5) successively, chloroform (200ml * 6), ethyl acetate (300ml * 6), n-butyl alcohol (250ml * 6) extraction.Organic layer evaporated under reduced pressure respectively gets petroleum ether part 16g, chloroform part 3g, ethyl acetate part 24g, n-butyl alcohol part 45g.N-butyl alcohol partly divides pulls on polyamide column, and the gained fraction is continued to obtain purified chemical compound 6 and 7 by means purification such as gel filtration chromatographies.
The physical property and the data of the particular compound that makes by embodiment 1～5:
Chemical compound 1 is white amorphous powder, molecular ion peak m/z:939[(C among the ESI-TOF-MS (electron spray ionisation-flight time mass spectrum)
41H
32O
26)-H]
-,
Chemical compound 2 white amorphous powders, molecular ion peak m/z:787[(C among the ESI-TOF-MS
34H
28O
22)-H]
-,
Chemical compound 3 light yellow amorphous powders, molecular ion peak m/z:787[(C among the ESI-TOF-MS
34H
28O
22)-H]
-,
Chemical compound 4 light yellow amorphous powders, molecular ion peak m/z:635[(C among the ESI-TOF-MS
27H
24O
18)-H]
-,
Chemical compound 5 white amorphous powders, molecular ion peak m/z:635[(C among the ESI-TOF-MS
27H
24O
18)-H]
-
Chemical compound 6 light yellow amorphous powders, molecular ion peak m/z:635[(C among the ESI-TOF-MS
27H
24O
18)-H]
-
Chemical compound 7 light yellow amorphous powders, molecular ion peak m/z:939[(C among the ESI-TOF-MS
41H
32O
26)-H]
-
Table 1. chemical compound
1The HNMR data (δ c, ppm)
| Chemical compound | 1? | 2? | 3? | 4(CD 3OD) | 5? | 6(CD 3OD) | 7? |
| Galloyl | 6.99，s 7.01，s 7.06，s 7.11，s 7.19，s? | 7.02，s 7.09，s 7.11，s 7.19，s? | 6.90，s 6.95，s 6.97，s 7.05，s 7.11，s? | 7.09，s 7.12，s 7.15，s? | 7.06，s 7.09，s 7.16，s? | 7.00，s 7.04，s 7.10，s? | 6.99，s 7.02，s 7.08，s 7.12，s 7.19，s? |
| Glucosyl group | 6.20，d 5.49，t 5.70，t 4.07，dd 4.18，m 4.65，m 4.58，m? | 6.19，d 5.49，dd 5.70，t 4.12，t 4.16，m 4.62，dd 4.58dd? | 6.24，d 5.49，m 5.90，t 3.34，d 3.59，s 4.51，d 4.41，m? | 6.98，s 5.42，t 5.82，d 5.28，t 4.45，dd 4.95，br? | 6.00，d 4.99，s 5.27，t 3.75，m 3.79，m 4.50，m 4.48，m? | 5.92，s 4.86，s 5.21，t 3.66，t 3.83，t 4.55，d 4.47，m? | 6.35，d 5.65，m 6.03，t 3.60，dd 3.60，dd 4.58，d 4.43，m? |
Annotate: solvent indicates especially is (CD
3)
2CO
Embodiment 6:1,2,3, the anti-helicobacter pylori of 6-four-O-Galla Turcica (Galla Helepensis) acyl-β-D-glucose (chemical compound 2) is external
Experiment
1. experiment material:
Test-compound 2: by the method preparation of embodiment 1.
Experimental strain: helicobacter pylori (Helicobacter pylori, Hp) reference culture ATCC 43504, ATCC 49503 preserve center (American Type Culture Collection from U.S.'s strain, ATCC), Hp SS1 bacterial strain is from microorganism institute of University of New South Wales, all the other helicobacter pylori clinical strains digest the disease institute from Shanghai, after RUT, smear Gram, oxidase and catalase are accredited as the positive, the pure culture of going down to posterity, obtained strains is as experimental strain.
2. experimental technique
1) strain culturing method
Adopt little aerobic bag (available from Fudan University) to carry out the strain culturing of Hp.
2) biological activity determination method
Adopt agar dilution measure minimum inhibitory concentration for test agent (minimal inhibitoryconcentration, MIC).
Agar dilution is measured MIC:
(A) preparation of medicine flat board: at first with the chemical compound and the medicinal dimethyl sulfoxide of positive control (DMSO) the solution mother solution that is mixed with 3.2mg/ml of test, after the filtration sterilization, reuse sterilized water doubling dilution to 320,160,80,40,20,10,5,2.5,1.25,0.6 with the concentration series of 0.3 μ g/ml, the concentration of DMSO in medium is less than 1%.The testing drug solution that 1ml is prepared and the abundant mixing of 9ml Colombia culture medium that is incubated in 50 ℃ are cast cooling, and culture medium is Mueller-Hinton agar (Difco Co.).
(B) switching experimental bacteria (being coated with bacterium) with microscale sampler draw dilution good 1 * 10
8The bacteria suspension 0.1ml of CFU/ml Hp spreads upon the culture dish surface equably, is inverted in 37 ℃ of drying bakers to take out behind the 15min, and purpose makes the agar surface drying, and is standby.
(C) determine that MIC (contains culture dish to be measured: 85% N at little aerobic bag
2, 10% CO
2With 5% O
2) in, be incubated 37 ℃ and cultivated 72 hours, observe the Hp growing state, be minimum inhibitory concentration (minimal inhibitory concentration, MIC) value with the sample least concentration that does not have bacteria growing fully.The positive control medicine is ampicillin (Ampicillin).All test triplicate.
3. experimental result
Table 2. variable concentrations 1,2,3,6-four-O-Galla Turcica (Galla Helepensis) acyl-β-D-glucose (μ g/ml) suppresses the activity influence of helicobacter pylori
The ampicillin of variable concentrations (μ g/ml) suppresses the active influence of helicobacter pylori
+: can suppress the growth of bacterium fully;-: can not suppress the growth of bacterium fully.
4. discussion of results
Experiment in vitro shows: 1,2,3,6-four-O-Galla Turcica (Galla Helepensis) acyl-β-D-glucose has very strong anti-helicobacter pylori activity, action intensity and positive drug ampicillin are suitable substantially, the part bacterial strain to The compounds of this invention than more responsive.The compounds of this invention concentration can suppress the growth of all Hp when being 6 μ g/ml, especially the growth inhibited to Sydney strain HpSS1, H.pylori 008 is the most obvious, and minimal inhibitory concentration (MIC) is 4.0 μ g/ml.
Embodiment 7:1,2,3, anti-helicobacter pylori experiment in the body of 6-four-O-Galla Turcica (Galla Helepensis) acyl-β-D-glucose
This experiment divides two stages to carry out, and the phase I is set up Hp mice infected model, and second stage is carried out pharmacodynamics test.
1, the foundation of helicobacter pylori (Hp) mice infected model
1) cultivation of Hp bacterial strain
Hp SS1 bacterial strain is from microorganism institute of University of New South Wales.-70 ℃ of frozen strains are recovered containing on the Columbia agar plate of 5% horse blood, and transferred species is in having added 15% horse serum and antibiotic brain-heart-infusion.Cultivate amplification under little aerobic condition ,-80 ℃ of preservations are put in packing after urease, catalase, oxidase test and smear Gram's staining are identified.Before the inoculation antibacterial was cultivated 24-48 hour on the Skirrow selective medium, after evaluation and observing the antibacterial vigor, antibacterial is centrifugal in PBS liquid, washing is mixed with 1x10 with improved broth medium at last
9The bacterial suspension of CFU/ml, inoculation experiments animal immediately.
2) the Hp bacterial strain is in the intravital inoculation of mice
Laboratory animal is no special pathogen level (SPF) C57BL/6 mice, Mus 6-8 in age week, body weight 18-20g.Entrust Chinese Academy of Sciences's Shanghai Experimental Animal Center to raise.The filling stomach gives every mice 0.5ml and (contains 5 * 10
8CFU) HpSS1 bacterium liquid, the next day 1 time, totally 3 times.2 all stochastic sampling modeling animals after the modeling behind the execution mice, are got stomach and do rapid urease test, smear Gram, pathological section and antibacterial culturing detection respectively, to guarantee the infection rate of mice.
2,1,2,3, the pharmacodynamic experiment of 6-four-O-Galla Turcica (Galla Helepensis) acyl-β-D-glucose
1) experiment grouping
The male mice of the Helicobacter pylori infection for preparing is divided into 5 groups, 10 every group at random.
2) administration
Positive group: CBS 6.15mg/kg, tetracycline 50mg/kg, metronidazole 22.5mg/kg; (CBS: colloidal bismuth subcitrate)
Small dose group: 1,2,3,6-four-O-Galla Turcica (Galla Helepensis) acyl-β-D-glucose 75mg/kg;
Middle dosage group: 1,2,3,6-four-O-Galla Turcica (Galla Helepensis) acyl-β-D-Fructus Vitis viniferae saccharic 150mg/kg;
Heavy dose of group: 1,2,3,6-four-O-Galla Turcica (Galla Helepensis) acyl-β-D-glucose 450mg/kg;
Model group: irritate stomach and only give normal saline (NS) 0.4ml/.
Once a day, totally 14 days.
3) curative effect detects
Last all mices of 4 week of administration back execution.Stomach is got in dissection, vertically is cut into 4 parts along greater gastric curvature, makes rapid urease test, smear Gram's staining, pathological section and antibacterial culturing respectively and detects Hp.4 kinds of equal negative patients of detection method result are decided to be the Hp feminine gender, and detection method result positive person is decided to be the Hp infection more than a kind or a kind.
3. experimental result
Table 3.1,2,3, anti-helicobacter pylori activity in the body of 6-four-O-Galla Turcica (Galla Helepensis) acyl-β-D-glucose
*: compare with model group; #: with the positive controls ratio.
*p＜0.05；
**p＜0.01。
4. discussion of results
We use the standard triple therapy the most commonly used clinically at present positive control as drug effect in the body, and the result shows: the clearance rate of accepting the positive controls Helicobacter pylori infection of triple therapy is 70%, and there were significant differences with the model group ratio; 1,2,3,6-four-O-Galla Turcica (Galla Helepensis) acyl-β-D-glucose clearance rate when big or middle dosage is respectively 60% and 50%, and there were significant differences with the model group ratio, do not have significant difference with the positive controls ratio simultaneously.Show that chemical compound of the present invention can effectively remove the infection of mice helicobacter pylori when dosage is 450mg/Kg, 150mg/Kg, action intensity is not weaker than triple therapy commonly used at present.
Embodiment 81, and 2,3, the research of 6-four-O-Galla Turcica (Galla Helepensis) acyl-β-laboratory animal gastric ulcer that D-glucose anti-helicobacter pylori causes
1. material and method
1.1 experiment material pallasiomy (Mongolian gerbil) MGS/Sea (SPF) 6 ages in week, ♂, body weight 50～55g.Hp SS1 bacterial strain is from microorganism institute of University of New South Wales.-70 ℃ of frozen strains are recovered containing on the Columbia agar plate of 5% horse blood, and transferred species is in having added 15% horse serum and antibiotic brain-heart-infusion.Cultivate amplification under little aerobic condition ,-80 ℃ of preservations are put in packing after urease, catalase, oxidase test and smear Gram's staining are identified.Before the inoculation antibacterial was cultivated 24-48 hour on the Skirrow selective medium, after evaluation and observing the antibacterial vigor, antibacterial is centrifugal in PBS liquid, washing is mixed with 3x10 with improved broth medium at last
11The bacterial suspension of CFU/ml, standby.
1.2 method
1.2.1 animal model pallasiomy fasting 24h, 30min irritates stomach and gives 400mL/L ethanol 0.5mL before the inoculation, above-mentioned bacterial suspension 0.5mL is irritated stomach give laboratory animal in 30min, and then fasting 4 hours, inoculate for three days on end.Get the part animal at random in 3 months after the modeling, put to death, get stomach and do rapid urease test, smear Gram, pathological section and antibacterial culturing detection, guarantee the modeling success.
1.2.2 the Hp inoculation was organized as test with the large, medium and small dosage of chemical compound of the present invention after 3 months, triple therapy (CBS 6.15mg/kg, tetracycline 50mg/kg, metronidazole 22.5mg/kg) is as positive controls; Every day gastric infusion, continuous 2 weeks, stop administration after 2 weeks, raise after 1 month and to put to death all animals, get stomach, magnifier is measured gastric wall ulcer surface length and width down, with its product (mm
2) as ulcer index.
Be calculated as follows ulcer inhibition rate:
Ulcer inhibition rate (%)={ 1-(experimental group ulcer index average/matched group ulcer index mean) } * 100% data represent that with X ± S statistical procedures is carried out in the relatively employing t check of group difference.
The influence of the gastric ulcer that table 4. 1,2,3,6-four-O-Galla Turcica (Galla Helepensis) acyl-β-D-glucose infect Hp (x ± S)
Annotate:
*P＜0.05 (with comparing);
*P＜0.01 (with comparing)
Result of the test shows that the big-and-middle dosage group of The compounds of this invention all can effectively reduce modeling animal gastric ulcer area.The action intensity of heavy dose of group is organized quite with positive.Show 1,2,3,6-four-O-Galla Turcica (Galla Helepensis) acyl-β-D-glucose can effectively be treated the digestive tract ulcer that causes because of helicobacter pylori.
Claims (2)
1. 2,3 of following formula, 6-four-O-Galla Turcica (Galla Helepensis) acyl-β-D-glucose or its pharmaceutical salts purposes in the medicine of the disease that preparation treatment helicobacter pylori causes:
2. the described purposes of claim 1, the disease that wherein said helicobacter pylori causes is gastritis, gastric ulcer, gastric erosion, duodenal ulcer and gastric cancer.
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| CN101618043B (en) * | 2009-06-10 | 2011-05-25 | 暨南大学 | Use of 1,2,3,4,6-penta-O-galloyl-b-D-glucose in preparation of anti-flu drugs |
| JP5902174B2 (en) * | 2010-09-30 | 2016-04-13 | ナショナル ヤン−ミン ユニバーシティ | Anticancer extracts and compounds |
| CN105079289B (en) * | 2015-09-07 | 2019-06-04 | 北京大学第一医院 | Fermented Chinese gall herb and tea leaves are preparing the application in Anti-helicobacter pylori drugs |
| CN112159443B (en) * | 2020-09-03 | 2023-08-25 | 青海师范大学 | Alpha-glucosidase inhibitor extracted from bougainvillea spectabilis and preparation method thereof |
| CN113087608B (en) * | 2021-03-08 | 2022-11-11 | 中国科学院西北高原生物研究所 | New diaryl nonane V, VI and VII free radical inhibitor in saxifraga tangutica and separation preparation process and application thereof |
| CN113072602B (en) * | 2021-03-08 | 2023-08-01 | 广东省科学院化工研究所 | Phenolic acid glycosyl ester and preparation method thereof |
| CN113105514B (en) * | 2021-03-08 | 2023-04-07 | 中国科学院西北高原生物研究所 | Novel galloyl radical inhibitor in saxifraga tangutica and separation preparation process and application thereof |
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| JP2004352644A (en) * | 2003-05-29 | 2004-12-16 | Oriza Yuka Kk | Anti-helicobacter pylori composition |
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| CN1704064A (en) * | 2004-05-28 | 2005-12-07 | 养生堂有限公司 | Novel uses of polyhydroxy gallic acyl-beta-D-glucose derivatives |
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| JP2004315386A (en) * | 2003-04-14 | 2004-11-11 | Hayashibara Takeshi | Anti-microbial agent |
| JP2004352644A (en) * | 2003-05-29 | 2004-12-16 | Oriza Yuka Kk | Anti-helicobacter pylori composition |
| CN1602853A (en) * | 2003-10-20 | 2005-04-06 | 北京大学 | Active constituent of traditional Chinese medicine capable of inhibiting SARS coronavirus infection and its bioactivity measuring method |
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| CN103446576B (en) * | 2013-09-12 | 2014-10-08 | 光明乳业股份有限公司 | Modeling method of helicobacter pylori infection animal model |
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