CN110105421A - A kind of oxidation uncaria indoles diterpene alkaloid and the preparation method and application thereof - Google Patents
A kind of oxidation uncaria indoles diterpene alkaloid and the preparation method and application thereof Download PDFInfo
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- CN110105421A CN110105421A CN201910500313.6A CN201910500313A CN110105421A CN 110105421 A CN110105421 A CN 110105421A CN 201910500313 A CN201910500313 A CN 201910500313A CN 110105421 A CN110105421 A CN 110105421A
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- C07J—STEROIDS
- C07J73/00—Steroids in which the cyclopenta[a]hydrophenanthrene skeleton has been modified by substitution of one or two carbon atoms by hetero atoms
- C07J73/001—Steroids in which the cyclopenta[a]hydrophenanthrene skeleton has been modified by substitution of one or two carbon atoms by hetero atoms by one hetero atom
- C07J73/003—Steroids in which the cyclopenta[a]hydrophenanthrene skeleton has been modified by substitution of one or two carbon atoms by hetero atoms by one hetero atom by oxygen as hetero atom
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Abstract
The present invention relates to a kind of oxidation uncaria indoles diterpene alkaloids and the preparation method and application thereof, after the preparation method of the oxidation uncaria indoles diterpene alkaloid includes the following steps: that (1) crushes dry uncaria buckle stem branch, 95% ethyl alcohol, hydrogen peroxide are added at room temperature and after glacial acetic acid stirring 2-3 hours, add isometric dehydrated alcohol, it is warming up to reflux temperature, after refluxing extraction 12-14 hours, filtering, filtrate are concentrated to give extract；(2) extract that step (1) obtains is through chromatographic isolation up to the oxidation uncaria indoles diterpene alkaloid.
Description
Technical field
The invention belongs to extracted form natural plant fields, and in particular to a kind of oxidation uncaria indoles diterpene alkaloid and its preparation
Method and application.
Background technique
Uncaria Uncaria rhynchophylla (Miq.) Jacks. is Rubiaceae (Rubiaceae) wild gambier
(Uncaria) the plant plant is one of main Original plant of traditional Chinese medicine uncaria, and medicinal part is buckle stem branch, has breath
Wind arresting convulsion, The flat liver of heat-clearing and other effects.A variety of indoles alkaloids is isolated from uncaria at present (based on rhynchophyllin).In the recent period
Applicant is extracted using alcohol reflux, and binding silica gel column chromatography, high performance liquid chromatography preparation obtain one from uncaria buckle stem branch
The uncaria indoles diterpene alkaloid compound of kind structure novel, and test its cytotoxic activity.It is new in order to obtain more structures
The uncaria indoles diterpene alkaloid compound of grain husk, applicant, which improves said extracted method, obtains a kind of increased activity
Aoxidize uncaria indoles diterpene alkaloid.
Summary of the invention
The present invention provides a kind of oxidation uncaria indoles diterpene alkaloid or its pharmaceutically acceptable salt, it is characterised in that institute
Stating oxidation uncaria indoles diterpene alkaloid has structure shown in Formulas I:
The present invention provides a kind of uncaria indoles diterpene alkaloid or its pharmaceutically acceptable salt, it is characterised in that the hook
Rattan indoles diterpene alkaloid has structure shown in Formula II:
Another embodiment of the present invention provides a kind of while preparing above-mentioned Formulas I, Formula II compound method, it is characterised in that
Include the following steps:
(1) after crushing dry uncaria buckle stem branch, 95% ethyl alcohol, hydrogen peroxide and glacial acetic acid stirring 2- are added at room temperature
After 3 hours, isometric dehydrated alcohol is added, is warming up to reflux temperature, after refluxing extraction 12-14 hours, filtering, filtrate are dense
Contract to obtain extract；
(2) extract that step (1) obtains is through chromatographic isolation up to Formulas I, Formula II compound.
The preferred 20-100 mesh of crushing described in step (1), it is preferable to use 95% second for every kilogram of dry uncaria buckle stem branch
Alcohol 3-4L, hydrogen peroxide 1.0-1.5L, glacial acetic acid 0.5-0.8L.
The preferred silica gel column chromatography of step (2) chromatographic isolation is combined with high performance liquid chromatography preparation；Specially extract
First through silica gel column chromatography, stationary phase is 200-300 mesh silica gel, mobile phase using petroleum ether-ethyl acetate by 100:0,90:10,
80:20,70:30,60:40,50:50,40:60,30:70,20:80,10:90,0:100 gradient elution, each gradient collect 3
Column volume is prepared into Formulas I, formula through high performance liquid chromatography after the fraction merging concentration that wherein 90:10 and 70:30 gradient elution obtains
II compound；The high performance liquid chromatography prepares the chromatographic condition used: chromatographic column for Agilent Eclipse XDB-C18,
9.4 × 250mm, 5 μm, flow velocity 1.5mL/min, mobile phase MeOH:H2O=65:35.
Another embodiment of the present invention provide uncaria buckle stem branch prepare above-mentioned Formulas I, the application in Formula II compound.
Another embodiment of the present invention provides above-mentioned Formulas I, Formula II compound or its pharmaceutically acceptable salt and is preparing
Application in anti-tumor drug.
Another embodiment of the present invention provides a kind of pharmaceutical composition, it is characterised in that described pharmaceutical composition is with above-mentioned
Formulas I, Formula II compound or its pharmaceutically acceptable salt are as effective component.It is antitumor that the pharmaceutical composition may also include other
Drug.The pharmaceutical composition may also include the auxiliary material that can pharmaceutically connect.
95% ethyl alcohol of the present invention refers to that volume fraction is 95% ethanol solution；The hydrogen peroxide preferred mass point
Number is the hydrogen peroxide of 30%-50%.
Detailed description of the invention
Fig. 1 is compound of formula I1H NMR(CDCl3, 600MHz) and figure；
Fig. 2 is compound of formula I13C NMR(CDCl3, 150MHz) and figure；
Fig. 3 is the MS figure of compound of formula I；
Fig. 4 is Formula II compound1H NMR(CDCl3, 600MHz) and figure；
Fig. 5 is Formula II compound13C NMR(CDCl3, 150MHz) and figure；
Fig. 6 is the MS figure of Formula II compound.
Specific embodiment
For the ease of a further understanding of the present invention, examples provided below has done more detailed description to it.But
It is that these embodiments are only not supposed to be a limitation to the present invention or implementation principle for better understanding invention, reality of the invention
The mode of applying is not limited to the following contents.
Embodiment 1
(1) it takes dry uncaria buckle stem branch (100g) to be crushed to 20-100 mesh, 95% ethyl alcohol is added at room temperature
After (300mL), 30% hydrogen peroxide (100mL) and glacial acetic acid (50mL) stir 2 hours, isometric dehydrated alcohol is added
(450mL) is warming up to reflux temperature, and after refluxing extraction 12 hours, filtering, filtrate are concentrated to give extract；
(2) for the extract that step (1) obtains first through silica gel column chromatography, stationary phase is 200-300 mesh silica gel, and mobile phase uses
Petroleum ether-ethyl acetate press 100:0,90:10,80:20,70:30,60:40,50:50,40:60,30:70,20:80,10:90,
0:100 gradient elution, each gradient collect 3 column volumes, and the fraction that wherein 90:10 and 70:30 gradient elution obtains merges dense
Prepare that (chromatographic condition: chromatographic column is Agilent Eclipse XDB-C18,9.4 × 250mm, 5 μ after contracting through high performance liquid chromatography
M, flow velocity 1.5mL/min, mobile phase MeOH:H2O=65:35 compound of formula I (faint yellow solid, 22mg), Formula II) are obtained
It closes object (white solid, 15mg).
Compound of formula I:1H NMR(CDCl3, 600MHz) and δ: 7.78 (1H, s, NH-1), 7.46 (1H, d, J=7.2Hz, H-
20), 7.32 (1H, d, J=7.2Hz, H-23), 7.13-7.07 (2H, m, H-21,22), 6.04 (1H, s, H-11), 5.76 (1H,
M, H-6), 4.23 (1H, s, H-9), 3.26 (1H, d, J=17.8Hz, Ha-5), 2.83-2.74 (2H, m, H-16, Ha-17),
2.47 (1H, dd, J=12.0,10.1Hz, Hb-17), 2.16-2.04 (2H, m, Hb-5, H-15a), 2.00-1.92 (2H, m, H-
14),1.86-1.80(1H,m,H-15b),1.37(3H,s,CH3-25),1.33(3H,s,CH3-28),1.28(3H,s,CH3-
29),1.06(3H,s,CH3-26)；13C NMR(CDCl3,125MHz)δ:196.8,154.1,151.3,144.8,139.9,
125.1,120.7,119.8,118.6,117.7,116.6,111.5,111.5,84.9,75.8,73.8,50.7,49.4,
43.2,32.8,32.1,27.2,26.5,25.2,21.1,21.0,16.3；ESIMS m/z[M+H]+434.2.
Formula II compound:1H NMR(CDCl3,600MHz)δ:7.78(1H,s,NH-1),7.46-7.44(1H,m,H-20),
7.33-7.30 (1H, m, H-23), 7.12-7.10 (2H, m, H-21,22), 3.26-3.22 (1H, dd, J=12.0,2.4Hz, H-
9), 3.08-3.04 (1H, dd, J=12.0,3.6Hz, H-7), 2.82-2.75 (1H, m, H-16), 2.72-2.68 (1H, dd, J
=13.3,6.6Hz, Ha-17), 2.38-2.33 (1H, dd, J=13.3,10.9Hz, Hb-17), 2.02-1.95 (1H, m, Ha-
5),1.87-1.77(2H,m,Hb-11,Ha-15),1.72-1.60(4H,m,Ha-14,Ha-10,Hb-5,Hb-15),1.52-
1.39(3H,m,H-13,Hb-14,Hb-10),1.22(3H,s,CH3-29),1.21(3H,s,CH3-28),1.16(3H,s,CH3-
26),1.15-1.14(1H,m,Ha-11),1.05(3H,s,CH3-25),0.91(3H,s,CH3-30)；13C NMR(CDCl3,
125MHz)δ:150.8,140.0,125.1,120.4,119.5,118.4,118.2,111.4,85.7,84.7,72.0,53.0,
48.8,46.4,40.0,37.7,36.6,33.9,26.1,25.3,24.6,23.7,22.0,21.9,20.0,14.6,12.7；
ESIMS m/z[M+H]+422.61,[M+H]+444.60.
Embodiment 2
The cytotoxic activity of hepatoma H22 cells is tested in this experiment according to mtt assay.It is thin with pancreatin digestion logarithmic phase first
Cell is uniformly diluted and is made 2 × 10 by born of the same parents4The single cell suspension of a/mL is inoculated in 96 orifice plates and 100 μ L is respectively added, then puts
Enter constant temperature incubation in cell incubator (37 DEG C of constant temperature, 5%CO2Under the conditions of cultivate), for 24 hours after it is adherent to cell, respectively plus
Enter Formulas I, II compound, final compound concentration is made to be followed successively by 0.625 μ g/mL, 1.25 μ g/mL, 2.5 μ g/mL, 5 μ g/mL, 25 μ g/
ML, and blank control wells and each three of Positive control wells (positive control drug cis-platinum) are set, it is used after 48h and uses automatic ELISA
Plate reader measures absorbance of each hole at 490nm.In triplicate, compared with negative control, compound pair is calculated separately
The inhibiting rate and IC of tumour cell50Value.The result shows that the IC of formula Compound I50Value is 6.3 ± 1.1 μ g/mL, Formula II
The IC of compound50Value is 14.2 ± 2.1 μ g/mL.
Claims (9)
1. a kind of oxidation uncaria indoles diterpene alkaloid or its pharmaceutically acceptable salt, it is characterised in that the oxidation uncaria Yin
Diindyl diterpene alkaloid has structure shown in Formulas I:
2. a kind of preparation formula I, Formula II compound method simultaneously, it is characterised in that include the following steps:
(1) after crushing dry uncaria buckle stem branch, it is small that 95% ethyl alcohol, hydrogen peroxide and glacial acetic acid stirring 2-3 are added at room temperature
Shi Hou adds isometric dehydrated alcohol, is warming up to reflux temperature, after refluxing extraction 12-14 hours, filtering, filtrate concentration
Obtain extract；
(2) extract that step (1) obtains is through chromatographic isolation up to Formulas I, Formula II compound；
Formulas I, Formula II compound structure are as follows:
3. method as claimed in claim 2, it is characterised in that the preferred 20-100 mesh of crushing, every kilogram of drying described in step (1)
Uncaria buckle stem branch it is preferable to use 95% ethyl alcohol 3-4L, hydrogen peroxide 1.0-1.5L, glacial acetic acid 0.5-0.8L.
4. the described in any item methods of claim 2-3, it is characterised in that the preferred silica gel column chromatography of step (2) chromatographic isolation
It is combined with high performance liquid chromatography preparation；For specially extract first through silica gel column chromatography, stationary phase is 200-300 mesh silica gel, flowing
Mutually using petroleum ether-ethyl acetate press 100:0,90:10,80:20,70:30,60:40,50:50,40:60,30:70,20:80,
10:90,0:100 gradient elution, each gradient collect 3 column volumes, the fraction that wherein 90:10 and 70:30 gradient elution obtains
Merge and is prepared into Formulas I, Formula II compound through high performance liquid chromatography after being concentrated；The chromatostrip that the high performance liquid chromatography preparation uses
Part: chromatographic column is Agilent Eclipse XDB-C18,9.4 × 250mm, 5 μm, and flow velocity 1.5mL/min, mobile phase is
MeOH:H2O=65:35.
5. application of the uncaria buckle stem branch in preparation formula I, Formula II compound.
6. compound of formula I described in claim 1 or its pharmaceutically acceptable salt application in preparation of anti-tumor drugs.
7. a kind of pharmaceutical composition, it is characterised in that described pharmaceutical composition is with compound of formula I described in claim 1 or its medicine
Acceptable salt is as effective component on.
8. pharmaceutical composition as claimed in claim 7, it is characterised in that the pharmaceutical composition may also include other anti-tumor drugs.
9. the described in any item pharmaceutical compositions of claim 7-8, it is characterised in that the pharmaceutical composition may also include pharmaceutically
The auxiliary material that can be connect.
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| CN201910500313.6A CN110105421B (en) | 2019-06-12 | 2019-06-12 | Oxidation uncaria indoxyl diterpenoid alkaloid and preparation method and application thereof |
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Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPH09291091A (en) * | 1996-04-25 | 1997-11-11 | Soyaku Gijutsu Kenkyusho:Kk | Indol alkaloid derivative, its production and antiviral agent |
| US20030191173A1 (en) * | 2000-01-18 | 2003-10-09 | Garcia Maria L. | Method for treating ocular hypertension |
| CN102311435A (en) * | 2011-07-05 | 2012-01-11 | 南京泽朗农业发展有限公司 | Preparation method for high purity rhynchophylline |
| CN107827940A (en) * | 2017-11-20 | 2018-03-23 | 中国科学院昆明植物研究所 | Yncaria stem with hooks acid amides A and its pharmaceutical composition and application |
-
2019
- 2019-06-12 CN CN201910500313.6A patent/CN110105421B/en active Active
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPH09291091A (en) * | 1996-04-25 | 1997-11-11 | Soyaku Gijutsu Kenkyusho:Kk | Indol alkaloid derivative, its production and antiviral agent |
| US20030191173A1 (en) * | 2000-01-18 | 2003-10-09 | Garcia Maria L. | Method for treating ocular hypertension |
| CN102311435A (en) * | 2011-07-05 | 2012-01-11 | 南京泽朗农业发展有限公司 | Preparation method for high purity rhynchophylline |
| CN107827940A (en) * | 2017-11-20 | 2018-03-23 | 中国科学院昆明植物研究所 | Yncaria stem with hooks acid amides A and its pharmaceutical composition and application |
Non-Patent Citations (1)
| Title |
|---|
| NA GAO ET AL.: "Alkaloids from the endophytic fungus Penicillium brefeldianum and their cytotoxic activities", 《CHINESE CHEMICAL LETTERS》 * |
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