CN1760202A - Anticancer new compound of Xiacaogan I and preparation method and the purposes in pharmacy thereof - Google Patents
Anticancer new compound of Xiacaogan I and preparation method and the purposes in pharmacy thereof Download PDFInfo
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Abstract
A kind of new compound of Xiacaogan I with anticancer activity, with the Gypsophila elegans is raw material, by pulverizing, solvent extraction, refining, silica gel column chromatography separate, and are that eluent gradient elution and C-18 reversed-phase silica gel column chromatography and dextrane gel column chromatography obtain the pure product of Xiacaogan I with chloroform-methanol-water mixed solvent.Experimental results show that this compound has significant anti-cancer activity, toxic side effect is little, thereby confirms that Xiacaogan I can be used for preparing the pharmaceutical composition for the treatment of cancer.
Description
Technical field:
The present invention relates to field of medicaments, is about a kind of compound with anticancer activity.
Background technology:
Following definition is applicable to whole specification sheets and claims:
Xiacaogan I=Chinese: 3-O-[β-D-galactopyranose-(1 → 2)]-[β-D-xylopyranose-(1 → 3)]-beta d glucopyranosiduronic acid base-23-aldehyde-Oleanolic Acid.
=English name: Gypsogenin 3-O-[β-D-galactopyranosyl-(1 → 2)]-[β-D-xylopyranosyl-(1 → 3)]-β-D-glucuronopyranosde.
Human life in the cancer serious threat.According to World Health Organization statistics, in 5,800,000,000 populations of the whole world, there are nearly 4,000 ten thousand people to suffer from cancer, and along with the increasing the weight of of industrialization, quickening of urbanization process and environmental pollution, cancer morbidity also rises continuing, and annual growth is up to 2-3%.Present global annual New Development cancer patient 8,700,000, therefore Si Wang number reaches 600 to 7,000,000, accounts for 1/10th of the total death toll of population.The tumor incidence of China is equally very surprising, and according to Ministry of Health's statistics, China's tumor incidence totally was the gesture (the more stable or decline of sickness rate of minority cancer is also arranged) of rising in past 20 years; The nineties, China's tumor incidence rose to 0.127%.Increase tumour patient 2,000,000 people newly domestic every year in recent years, about dead more than 130 ten thousand people, present national tumour patient Estimate of Total Number is about 4,500,000 people.In recent years, China's cancer morbidity continues to rise, and most common solid tumors such as lung cancer, liver cancer, colorectal carcinoma and carcinoma of the pancreas etc. also lack active drug, and many antitumour drugs produce resistance in process of clinical application.Therefore, antitumor drug research remains the very important problem of pendulum in face of researcher.
From natural product, seek new activeconstituents, and and then to develop new medicine be the important channel of capturing clinical persistent ailment now; The present invention uses up-to-date technological means, and in conjunction with modern isolation and identification method, chemical constitution study by the rosy clouds grass being carried out system and anti-tumor activity research are in the hope of the foundation that provides science for the medicine of seeking new treatment tumour or lead compound.
The rosy clouds grass is the root of Caryophyllaceae Gypsophila plant rosy clouds grass Gypsophila oldhamiana Miq..This product whole nation is distributed more widely, main product in Shandong, ground such as Shanxi, Shaanxi, Ningxia, the Inner Mongol, Gansu, Xinjiang, Hebei, Heilungkiang.This plant is China's herbal medicine commonly used among the people, and " Chinese medicine voluminous dictionary " records, and is used as medicine as the surrogate of Starwort Root in some area.This property of medicine is sweet, be slightly cold, and has effects such as clearing heat and cooling blood, promoting blood circulation to remove blood stasis, swelling and pain relieving, removing necrosis, promoting granulation.Illness such as main treatment consumptive disease flesh heat, hectic fever due to yin, deficiency of Yin chronic malaria, children's's infantile malnutrition due to digestive disturbances or intestinalparasites heat can be used for also simultaneously that wound, bone are torn open, wound etc.The vegetalization scholar in Europe carried out some researchs to this plant, got compounds such as triterpene, triterpenoid saponin, flavones, sterol, polysaccharide, organic acid.Pharmacological experiment study shows, the triterpenoid saponin that extracts in the rosy clouds grass gives rabbit forming the atherosclerotic while or taking orally every day later on, can reduce serum ornitrol concentration, and cholesterol/kephalin coefficient is reduced, and aorta lipoids content is reduced.Someone thinks that saponin can act on plasma lipoprotein, stops cholesteric esterification and in the deposition of vessel wall, and also the someone thinks and can stop cholesterol from intestinal absorption.There are bibliographical information gypsoside, nine glucosides not to have antibacterial effect, remove the glucoside key on the decarboxylate behind the enzymolysis, remove 5 sugar, generate time saponin and have significant anti-microbial effect.Domestic less to this plant research, the research of chemical ingredients and pharmacological action aspect there is no report.
In Chinese patent application 200510044301.5, the applicant discloses the rosy clouds grass extract that extracts from Gypsophila elegans, has good anticancer, antivirus action can be used for preparing cancer therapy drug and anti-viral pharmaceutical compositions.We have therefrom extracted a kind of new compound with antitumour activity again on the basis of above-mentioned research.
Summary of the invention:
The purpose of this invention is to provide a kind of new compound---Xiacaogan I with antitumour activity.
Another object of the present invention provides a kind of have the Xiacaogan I new compound of antitumour activity and the preparation method of physiologically acceptable salt thereof.
A further object of the invention provides has antitumour activity Xiacaogan I new compound and the new purposes of physiologically acceptable salt in preparation treatment cancer drug thereof.
The present invention extracts in Gypsophila elegans, obtain this that separate, purify has an anti-cancer active compound, our called after Xiacaogan I (Oldhamianoside I), and its structural formula is:
Chemistry is by name: Chinese: 3-O-[β-D-galactopyranose-(1 → 2)]-[β-D-xylopyranose-(1 → 3)]-beta d glucopyranosiduronic acid base-23-aldehyde-Oleanolic Acid.
=English name: Gypsogenin 3-O-[β-D-galactopyranosyl-(1 → 2)]-[β-D-xylopyranosyl-(1 → 3)]-β-D-glucuronopyranosde.
Xiacaogan I of the present invention is the new compound that extracts from the rosy clouds grass, separates, purifies and obtain.The present invention extracts preparation Xiacaogan I compound and comprises following key step from Gypsophila elegans:
1, producing of rosy clouds grass crude extract:
(1), get dry Gypsophila elegans and pulverize after, use solvent extraction, said solvent is selected from alcohol, water or water and the pure mixture or the mixture of water and acetonitrile of acetonitrile, a 1-4 carbon;
(2), the extracting solution concentrating under reduced pressure gets medicinal extract, is rosy clouds grass crude extract;
2, rosy clouds grass crude extract is refining:
A), above-mentioned medicinal extract water makes suspension, uses n-butanol extraction again, isolates propyl carbinol and distributes phase, obtains refining rosy clouds grass extract;
Perhaps
B), medicinal extract macroporous adsorbent resin excessively, this moment, the relative density of medicinal extract was controlled at 1.04-1.06, with polar solvent flush away impurity, used the alcoholic solution wash-out more earlier, collected elutriant respectively, obtained purified rosy clouds grass extract; Here said polar solvent is generally water or 1%-10% alcohol solution, and said alcoholic solution eluent then is the 20-60% alcohol solution;
3, the Separation ﹠ Purification of Xiacaogan I:
(C), separate with silica gel column chromatography:
I), above-mentioned refining extract with silica gel mixed sample after, separate with silica gel column chromatography; And be eluent with chloroform-methanol-water mixed solvent, carry out gradient elution, collect elutriant respectively, obtain the Xiacaogan I crude product;
Ii), obtain pure Xiacaogan I with C-18 reversed-phase silica gel column chromatography and dextrane gel column chromatography purification again; Perhaps
(D) separate with the solvent segregation process:
A), above-mentioned purified rosy clouds grass extract being condensed into relative density is 1.18 thick medicinal extract;
B), thick medicinal extract lyophilize, add 2-5 times of water, heating for dissolving;
C), add 10-30 times of organic solvent in the solution it separated out, organic solvent can be selected acetone or alcohol, collects the precipitate airing and obtains Xiacaogan I.
According to the Xiacaogan I compound step that the present invention proposes, can design the preparation technology of multiple Xiacaogan I.First kind of preferred Xiacaogan I compound technology of the present invention may further comprise the steps:
1, get dry Gypsophila elegans and pulverize after, with the alcoholic solvent extraction, alcohol extract concentrate pure medicinal extract;
2, pure medicinal extract adds water and makes suspension, uses n-butanol extraction again;
3, n-butanol extracting liquid silica gel mixed sample separates with silica gel column chromatography, and is eluent with chloroform-methanol-water mixed solution, carries out gradient elution, is earlier 9: 1: 0.1 mixed solvent wash-out with chloroform-methanol-water ratio; Follow mixed solvent wash-out with 8: 2: 0.2; Use 65: 35: 10 mixed solvent wash-out again; Collect 65: 35: 10 elutriant, obtain Xiacaogan I;
4, obtain Xiacaogan I with C-18 reversed-phase silica gel column chromatography and dextrane gel column chromatography purification.
Second kind of Xiacaogan I preferred preparation of the present invention technology is:
(1), get the Gypsophila elegans of pulverizing, extract with alcoholic solvent, it is rare medicinal extract of 1.06 that extracting solution is condensed into relative density;
(2), rare medicinal extract crosses macroporous adsorbent resin, said polymeric adsorbent be in low polar macroporous resin, its specific surface area is 50～600m
2/ g, to colourless, first decon discards washings with the polar solvent washing;
(3), use alcoholic solution wash-out, collection elutriant, methyl alcohol or ethanolic soln that said alcoholic solution is 20-60% here;
(4), with elutriant with silica gel mixed sample after, separate with silica gel column chromatography, and be eluent with chloroform-methanol-water mixed solution, carry out gradient elution, collect elutriant respectively, concentrate eluant obtains Xiacaogan I;
(5), further use solvent recrystallization, obtain the Xiacaogan I elaboration.
Another Xiacaogan I preferred preparation technology of the present invention is:
1, get the Gypsophila elegans of pulverizing, with alcoholic solvent extraction, it is rare medicinal extract of 1.06 that extracting solution is condensed into relative density;
2, cross macroporous adsorbent resin with the rare medicinal extract of above-mentioned method, with polar solvent flush away impurity, use the alcoholic solution wash-out more earlier, collect elutriant respectively;
3, to be condensed into relative density be 1.18 thick medicinal extract to elutriant;
4, thick medicinal extract lyophilize adds 2-5 times of water dissolution, filters;
5, filtrate adds 10-30 times of acetone or alcohol, leaves standstill, and collects precipitate, and airing obtains Xiacaogan I.
With modern spectral methods such as chemical reaction, infrared, nucleus magnetic resonance and mass spectrums the new bonded structure of extracting is identified that qualification result is as follows:
Xiacaogan I pale yellow powder (aqueous ethanol).Infrared spectra is shown with hydroxyl (3381cm
-1), carbonyl (1711,1702,1694cm
-1) and carbon-carbon double bond (1604cm
-1).It is m/z979.8[M+k that FABMS provides quasi-molecular ion peak]
+And 963.8[M+Na]
+, determine that in conjunction with hydrogen spectrum and carbon spectrum its molecular composition is C
4H
72O
19, also can see the ion fragment peak m/z847.7[M+K-132 that other is important]
+, 831.7[M+Na-132]
+, 669.5[M+Na-132-162)]
+, 493.4[M+Na-132-162-176]
+, 471.6[M+H-132-162-176]
+And 453.6[M+H-132-162-176-H
2O], Liebermann-Burchard and Molish reaction are all positive.
After this idic acid hydrolysis, water liquid detects D-wood sugar, D-semi-lactosi through TLC.Tentatively be inferred as a triterpene saponin componds that has three sugar.
1Occur six angular methyl(group) signals (δ 0.68,0.81,0.88,0.94,1.19,1.36) in the HNMR spectrum, be unimodal, δ 4.66 is an a pair of key hydrogen, and δ 9.78 is an aldehyde radical hydrogen;
13An aldehyde radical carbon signal (δ 210.1) and a carboxyl carbon signal (δ 181.2) and two double key carbon signals (δ 144.7,121.9) appear in the CNMR spectrum.According to
1HNMR,
13CNMR, DEPT belong to all the carbon spectrum data on this compound aglycon, on the aglycon
13CNMR data and Gypsogenin basically identical, the aglycon of determining compound is Gypsogenin.
13In the CNMR spectrum, the end group carbon signal (δ 103.2,103.7,104.4) of three sugar is arranged.There are 3 carbon of the carbon signal hint aglycon of δ 84.5 to be connected to sugar chain.According to DEPT spectrum and FABMS and relevant document, the structure of inferring this compound is: 3-O-β-D-xylopyranosyl-(1 → 3)-[β-D-galactopyr-anosyl-(1 → 2)]-β-D-glucuronopyranosylGypsogenin.According to
1HNMR,
13CNMR, DEPT, FABMS also belong in conjunction with relevant document all carbon spectrum data to this compound.The results are shown in Table 2.
The carbon spectrum of table 2 Xiacaogan I
| Numbering | A | B | C | Numbering | A | B |
| 1 2 3 4 5 6 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 COO CH 3 | 37.7 24.8 84.5 54.9 48.1 19.0 32.2 39.6 47.5 35.9 23.4 121.9 144.7 41.9 28.0 20.1 46.3 41.8 46.5 30.7 34.0 32.9 210.1 10.8 15.3 17.1 25.9 181.2 33.1 23.6 | 38.1 23.7 84.6 55.1 47.8 20.5 32.7 40.1 48.7 36.5 23.9 122.3 144.9 42.3 28.4 23.8 46.6 42.1 46.7 31.0 34.3 33.3 210.1 11.1 15.7 17.6 26.2 180.2 33.3 23.9 52.1 | 38.4 27.0 71.6 56.2 47.9 21.0 33.1 40.0 47.6 36.1 23.8 122.1 144.8 42.2 28.2 23.8 46.5 41.9 46.4 30.9 34.2 32.5 207.0 9.6 15.7 17.3 26.1 180.0 33.3 23.8 | Glur-1 Glur-2 Glur-3 Glur-4 Glur-5 Glur-6 Gal-1 Gal-2 Gal-3 Gal-4 Gal-5 Gal-6 Xyl-1 Xyl-2 Xyl-3 Xyl-4 Xyl-5 | 103.2 77.9 85.9 69.9 76.4 175.4 103.7 73.1 74.7 70.4 75.5 61.5 104.4 74.9 77.8 71.5 66.8 | 104.1 78.7 86.1 70.3 76.9 170.0 104.4 73.8 75.4 71.1 75.7 61.9 105.1 75.4 77.2 71.4 67.4 |
A：Gypsogenin 3-O-[β-D-galactopyranosyl-(1→2)][β-D-xylopyranosyl-(1→3)-β-D-glucuronopyranosde
B：Gypsogenin methyl ester 3-O-[β-D-galactopyranosyl-(1→2)][β-D-xylopyranosyl-(1→3)-β-D-glucuronopyranosde
C：Gypsogenin
Structure is as follows:
Compd B=compd A CH
3
Compound C H H
Identify that through above structure definite this chemical combination is a new compound.Called after Xiacaogan I (Oldhamianoside I).
Chinese chemical name: 3-O-[β-D-galactopyranose-(1 → 2)]-[β-D-xylopyranose-(1 → 3)]-beta d glucopyranosiduronic acid base-23-aldehyde-Oleanolic Acid.
=English name: Gypsogenin3-O-[β-D-galactopyranosyl-(1 → 2)]-[β-D-xylopyranosyl-(1 → 3)]-β-D-glucuronopyranosde.
The following physical chemical characteristics of this compound exhibits:
The relevant data of Xiacaogan I is as follows: pale yellow powder (aqueous ethanol), C
47H
72O
19IRυ
maxcm
-1：3381，2944，2721，1711，1702，1694，1604，1392，1075，1040，841。
1HNMR(C
5D
5N，500MHz，δ，ppm)δ：9.78(1H，s，H-23)，5.16(1H，d，J＝7.0Hz，H-1of xylose)，4.66(1H，br s，H-12)，1.36(3H，s，H-24)，1.19(3H，s，H-27)，0.94(3H，s，H-29)，0.88(3H，s，H-30)，，0.81(3H，s，H-26)，0.68(3H，s，H-25)。Table 2.
13C Chemical shifts(±0.1)of compounds A.B and Cin pyridine-d
5。
Further measured the anti-tumor activity of new compound of Xiacaogan I.
Testing 1. Xiacaogan Is observes the cell in vitro toxic action of human cancer cell
7 kinds of human cancer cell strains are adopted in experiment, comprising: Bel-7402 (liver cancer), A2780 (ovarian cancer), KB (mouth epithelial cells cancer), BGC-823 (cancer of the stomach), MCF-7 (mammary cancer), A549 (adenocarcinoma of lung), Hela (cervical cancer).Experimentize with mtt assay, the results are shown in Table 1.
Table 1. Xiacaogan I is in external influence to growth of human tumor cells
| Sample | IC50(μg/ml) | ||||||
| A2780 | KB | A549 | Bel-7402 | BGC-823 | MCF-7 | Hela | |
| Xiacaogan I | 15.25 | 22.13 | 28.46 | ＞50 | ＞50 | ＞50 | ＞50 |
Xiacaogan I is to A2780, KB, and A549, three-type-person's tumor cell line has certain cytotoxicity; To Bel-7402, BGC-823, MCF-7, four kinds of human tumor cell lines of Hela are not seen obvious cytotoxicity.
Test the restraining effect of 2. Xiacaogan Is to the mice transplanted tumor growth
Observed the restraining effect of Xiacaogan I with the experimental technique of routine, the results are shown in Table 2, table 3 mouse S180 sarcoma, rat liver cancer H22.
Table 2. Xiacaogan I is to the influence of mouse S180 sarcoma growth
| Group | Dosage (mg/kg) | Number of animals (beginning/end) | Knurl heavy (g) | Inhibiting rate (%) | The P value |
| Control group FT207 Xiacaogan I Xiacaogan I Xiacaogan I | - 130 50 100 200 | 10/10 10/10 10/10 10/10 10/10 | 2.32±0.34 0.95±0.46 ** 2.11±0.66 1.56±0.32 * 1.32±0.37 * | - 59.1 13.4 32.8 43.1 | - ＜0.01 ＞0.05 ＜0.05 ＜0.05 |
Annotate: " * " compares p＜0.05 with control group; " * * " compares p＜0.01 with control group.
Table 3. Xiacaogan I is to the influence of rat liver cancer H22 growth
| Group | Dosage (mg/kg) | Number of animals (beginning/end) | Knurl heavy (g) | Inhibiting rate (%) | The P value |
| Contrast FT207 Xiacaogan I Xiacaogan I Xiacaogan I | - 130 50 100 200 | 10/10 10/10 10/10 10/10 10/10 | 2.13±0.82 0.98±0.38 ** 1.87±0.97 1.21±1.02 * 1.12±0.50 * | - 54.0 12.2 43.2 47.4 | - ＜0.01 ＞0.05 ＜0.05 ＜0.05 |
Annotate: " * " compares p＜0.05 with control group; " * * " compares p＜0.01 with control group.
Experimental result shows that under this experiment condition, Xiacaogan I all has certain growth-inhibiting effect to mouse S180 sarcoma, rat liver cancer H22, and is certain dose-effect relationship, illustrates that this sample has certain anti-tumor activity.
Test the influence that 3. Xiacaogan Is transform mouse lymphocyte.The results are shown in Table 4.
The influence that table 4. Xiacaogan I transforms mouse spleen lymphocyte
| Group | Dosage (mg/kg) | Number of animals (only) | The OD value (X ± SD) | Increment rate (%) |
| Negative control group Xiacaogan I Xiacaogan I Xiacaogan I | -- 50 100 200 | 10 10 10 10 | 1.44±0.59 1.46±0.34 1.51±0.85 1.50±0.74 | 1.4 4.9 4.2 |
Experimental result: each organizes lymphopoiesis does not have significant difference.The result shows that Xiacaogan I do not see the effect that strengthens the lymphocyte transformation ability at tumor-bearing mice.
Test 4. Xiacaogan Is and tumor-bearing mice is cleaned up the influence of ability.The results are shown in Table 5.
Table 5. Xiacaogan I influences normal mouse carbon clearance exponential
| Group | Dosage (mg/kg) | The K value | The α value |
| Blank Xiacaogan I Xiacaogan I Xiacaogan I | -- 50 100 200 | 0.058±0.040 0.063±0.042 0.074±0.046 0.076±0.025 | 4.920±2.303 5.746±1.352 5.885±1.659 5.799±1.061 |
Experimental result: three dosage groups clean up the index difference of comparing with the blank group that there are no significant.Xiacaogan I does not have obvious influence to the clearance in mice ability.
Test of the synergism of 5. Xiacaogan Is to the endoxan tumor-inhibiting action.The results are shown in Table 6.
Table 6 Xiacaogan I is to the synergism of endoxan tumor-inhibiting action
| Group | Dosage mg/kg | Number of animals | The knurl weight (g, X ± SD) | Tumour inhibiting rate (%) | The P value |
| Control group endoxan endoxan 10mg/kg endoxan+Xiacaogan I | - 20 10 50 100 200 | 18 10 10 10 10 10 | 2.53±0.84 1.16±0.56 ** 2.22±0.78 1.57±0.49 1.37±0.52 1.04±0.55 | 54.2 12.3 37.9 45.8 58.9 | ＜0.01 ＞0.05 ＜0.05 ＜0.05 ＞0.05 |
Annotate: compare " * " P＜0.05, " * * " P＜0.01 with control group.
Experimental result shows that this product and chemotherapeutics reduce the consumption of chemotherapeutics, thereby reduce its toxic side effect with the tumor-inhibiting action that can increase chemotherapeutics.
In sum:
1. Xiacaogan I is to A2780, KB, and A549, three-type-person's tumor cell line has certain cytotoxicity.
2. the tumour inhibiting rate of Xiacaogan I 50mg/kg group is 13.4%, and the tumour inhibiting rate of 100mg/kg group is 32.8%, and the tumour inhibiting rate of 200mg/kg group is 43.1%.Show that growth has certain restraining effect to Xiacaogan I to mouse 180 sarcomas, and be certain dose-effect relationship.。
3. Xiacaogan I 50mg/kg group is 12.2% to the tumour inhibiting rate of mouse H22, and the tumour inhibiting rate of 100mg/kg group is 43.2%, and the tumour inhibiting rate of 200mg/kg group is 47.4%.Show that growth has certain restraining effect to Xiacaogan I to rat liver cancer H22, and be certain dose-effect relationship.
4. synergy test result: give tumor-bearing mice Xiacaogan I 50-200mg/kg, its tumour inhibiting rate is respectively 37.9%, 45.8%, 58.9%, can obviously improve the tumor-inhibiting action of 10mg/kg endoxan, illustrate that Xiacaogan I and endoxan share and have the obvious synergistic effect mouse S180 sarcoma.At present, the antitumor action research of rosy clouds grass saponin(e there is no report both at home and abroad, and has better antitumor activity, and this has the prospect of researching and developing preferably in antitumor research.
Experimental results show that by above, Xiacaogan I has the good anticancer activity, toxic side effect is little, thereby confirm Xiacaogan I and at physiologically acceptable salt, can be used to prepare the pharmaceutical composition for the treatment of cancer, in described pharmaceutical composition, contain Xiacaogan I with anticancer activity, and acceptable carrier pharmaceutically.
Embodiment
Below in conjunction with embodiment the present invention is described in further detail.
Embodiment one,
Pulverize after getting the Gypsophila elegans seasoning, get the 5kg alcohol reflux, the ethanol extract concentrating under reduced pressure gets ethanol extract 500g, with the ethanol extract water dissolution, and n-butanol extraction, propyl carbinol part 150g; N-butanol portion with silica gel mixed sample after, silica gel column chromatography separates, with the different gradient mixed solvent of chloroform-methanol-water wash-out, the thin-layer chromatography monitoring, same composition merges, and in conjunction with C-18 reversed-phase silica gel column chromatography and Portugal's coagel column chromatography, obtains Xiacaogan I.
Embodiment two,
Pulverize after getting the Gypsophila elegans seasoning, get the 5kg methanol extraction, the methanol extract liquid concentrating under reduced pressure gets methyl alcohol medicinal extract 580g, with methyl alcohol medicinal extract water dissolution, and n-butanol extraction, propyl carbinol part 170g; N-butanol portion with silica gel mixed sample after, silica gel column chromatography separates, with the different gradient mixed solvent of chloroform-methanol-water wash-out, the thin-layer chromatography monitoring, same composition merges, and in conjunction with C-18 reversed-phase silica gel column chromatography and Portugal's coagel column chromatography, obtains Xiacaogan I.
Embodiment three,
Pulverize after getting the Gypsophila elegans seasoning, add 50% methyl alcohol 7kg, boil and extracted 2 hours, filter, merge filtrate twice, the filtrate after merging is condensed into rare medicinal extract of relative density 1.06, this rare medicinal extract is crossed macroporous adsorbent resin, earlier be washed till colourless with 1% methyl alcohol, this meoh eluate is discarded, use 40% methanol-eluted fractions again, collect this 40% meoh eluate to colourless, after will reclaiming the methyl alcohol silica gel mixed sample, silica gel column chromatography separates, with the different gradient mixed solvent of chloroform-methanol-water wash-out, thin-layer chromatography monitoring, same composition merges, and obtains Xiacaogan I.
Embodiment four:
Pulverize after getting 1 kilogram of seasoning of Gypsophila elegans, add 1% ethanol 10kg, boil and extracted 2 hours, filter filtrate for later use, the dregs of a decoction add 1% ethanol 8kg again and boil and extracted 2 hours, filter, merge filtrate twice, the filtrate after merging is condensed into rare medicinal extract of relative density 1.06, this rare medicinal extract is crossed macroporous adsorbent resin, elder generation is washed till colourless with 10% ethanol, this ethanol eluate is discarded, and uses 60% ethanol elution to colourless again, collect this 60% ethanol eluate, with reclaiming the thick medicinal extract that elutriant behind the ethanol is condensed into relative density 1.18,, add 2 times water in the Chinese medical extract with this thick medicinal extract lyophilize, heating makes dissolving, the acetone that adds 24 times again left standstill 24 hours, collected precipitate, dry, obtain containing the elaboration of Xiacaogan I 90%.
Embodiment five:
Get Xiacaogan I 2 grams and put adding ethanol 50ml in the 250ml flask, stir the ethanolic soln that adds sodium hydroxide down, transfer pH value to be about 8,0 ℃ of stirrings 30 minutes, separate out crystallization, filter, the ice washing with alcohol, the ether washing, drying obtains the Xiacaogan I sodium salt.
Claims (9)
1, a kind of tool anti-cancer active compound is characterized in that said anti-cancer active compound is an Xiacaogan I, and its structural formula is:
2,, it is characterized in that comprising following key step according to the preparation method of the described tool anti-cancer active compound of claim 1:
(1), producing of rosy clouds grass crude extract:
I), get dry Gypsophila elegans and pulverize after, use solvent extraction;
Ii), the extracting solution concentrating under reduced pressure gets medicinal extract;
(2), rosy clouds grass crude extract is refining:
(A), medicinal extract with water dissolution after, use solvent extraction again, obtain refining rosy clouds grass extract; Perhaps
(B), medicinal extract crosses macroporous adsorbent resin, with 1%-10% solvent flush away impurity, uses 20-60% alcoholic solution wash-out again, collects elutriant, obtains purified rosy clouds grass extract;
(3), the Separation ﹠ Purification of Xiacaogan I:
(C), separate with silica gel column chromatography:
I), refining extract with silica gel mixed sample after, silica gel column chromatography separates, and is eluent with chloroform-methanol-water mixed solvent, carries out gradient elution, collects elutriant respectively, obtains Xiacaogan I;
Ii), further obtain the Xiacaogan I elaboration with solvent recrystallization.Perhaps
(D) separate with the solvent segregation process:
I), will make with extra care extract and be condensed into the thick medicinal extract that relative density is 1.1-1.26;
Ii), with thick medicinal extract lyophilize, add 2-5 times of water, dissolving, filter;
Iii), add 10-30 times of acetone or alcohol in the solution, leave standstill, collect precipitate, airing obtains Xiacaogan I.
3,, it is characterized in that said solvent in the step (1) is selected from alcohol, water or water and the mixture of alcohol or the mixture of water and acetonitrile of acetonitrile, a 1-4 carbon according to the described preparation method of claim 2.
4,, it is characterized in that said solvent is selected from Virahol, propyl carbinol in the step (2-A) according to the described preparation method of claim 2.
5, according to the preparation method of the described tool anti-cancer active compound of claim 2, it is characterized in that the preparation technology of said Xiacaogan I compound, comprise following key step:
(1), get dry Gypsophila elegans and pulverize after, extract with alcoholic solvent, alcohol extract concentrate pure medicinal extract;
(2), after the pure medicinal extract water dissolving, again with propyl carbinol or Virahol extraction;
(3), extracting solution is with silica gel mixed sample, silica gel column chromatography separates, and is eluent with chloroform-methanol-water mixed solution, carries out gradient elution and collects elutriant respectively, obtains the Xiacaogan I crude product;
(4), obtain the Xiacaogan I elaboration with C-18 reversed-phase silica gel column chromatography and dextrane gel column chromatography purification.
6, according to the preparation method of the biochemical compound of the anticancer work of the described tool of claim 2, it is characterized in that the preparation technology of said Xiacaogan I compound, comprise following key step:
(1), get the Gypsophila elegans of pulverizing, with the alcoholic solvent extraction, alcohol extract is condensed into rare medicinal extract that relative density is 1.04-1.06;
(2), rare medicinal extract crosses macroporous adsorbent resin, water or the washing of 1% methanol aqueous solution discard washings to colourless;
(3), with 20-40% methanol aqueous solution wash-out, collect elutriant respectively;
(4), elutriant with silica gel mixed sample after, silica gel column chromatography separates, and is eluent with chloroform-methanol-water mixed solution, carries out gradient elution, collects elutriant respectively, obtains Xiacaogan I;
(5), further use solvent recrystallization, obtain the Xiacaogan I elaboration.
7, according to the preparation method of the described tool anti-cancer active compound of claim 2, it is characterized in that the preparation technology of said Xiacaogan I compound, comprise following key step:
(1), get the Gypsophila elegans of pulverizing, with the alcoholic solvent extraction, alcohol extract is condensed into rare medicinal extract that relative density is 1.04-1.06;
(2), rare medicinal extract crosses macroporous adsorbent resin, water or less than the washing of 10% aqueous ethanolic solution to colourless, discard washings;
(3), be that eluent carries out wash-out with the 40-60% aqueous ethanolic solution, collect elutriant respectively, being condensed into relative density is the thick medicinal extract of 1.1-1.26;
(4), with behind the thick extract dry, add 2-5 times of water dissolution, filter;
(5), filtrate adds 10-30 times of acetone or alcohol, leaves standstill, and collects precipitate, airing obtains Xiacaogan I.
8,, it is characterized in that said macroporous resin hangs down polar macroporous resin in being according to the preparation method of the described tool anti-cancer active compound of claim 2.
9, according to the purposes of the described tool anti-cancer active compound of claim 1 in preparation treatment cancer drug composition, contain the anticancer Xiacaogan I of living and giving birth to of tool in the wherein said pharmaceutical composition, and acceptable carrier pharmaceutically.
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1944454B (en) * | 2006-09-29 | 2012-05-02 | 山东省医学科学院药物研究所 | Anti-cancer compound, method for extracting said compound, composition containing said compound, and its use in medicine production |
| CN103908483A (en) * | 2014-03-27 | 2014-07-09 | 青岛大学 | Extracting method of radix gypsophila saponin |
| CN104688798A (en) * | 2015-03-24 | 2015-06-10 | 山东省医学科学院药物研究所 | Gypsophila oldhamiana Miq extractive preparation method and application |
-
2005
- 2005-11-11 CN CNB2005100450767A patent/CN100341888C/en not_active Expired - Fee Related
Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1944454B (en) * | 2006-09-29 | 2012-05-02 | 山东省医学科学院药物研究所 | Anti-cancer compound, method for extracting said compound, composition containing said compound, and its use in medicine production |
| CN103908483A (en) * | 2014-03-27 | 2014-07-09 | 青岛大学 | Extracting method of radix gypsophila saponin |
| CN104688798A (en) * | 2015-03-24 | 2015-06-10 | 山东省医学科学院药物研究所 | Gypsophila oldhamiana Miq extractive preparation method and application |
| CN104688798B (en) * | 2015-03-24 | 2018-09-21 | 山东省医学科学院药物研究所 | A kind of preparation method and application of rosy clouds grass extract |
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| CN100341888C (en) | 2007-10-10 |
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