CN102173987B - New manchurian walnut bark acid and derivatives thereof, and preparation methods and medical application thereof - Google Patents

New manchurian walnut bark acid and derivatives thereof, and preparation methods and medical application thereof Download PDF

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CN102173987B
CN102173987B CN 201110053351 CN201110053351A CN102173987B CN 102173987 B CN102173987 B CN 102173987B CN 201110053351 CN201110053351 CN 201110053351 CN 201110053351 A CN201110053351 A CN 201110053351A CN 102173987 B CN102173987 B CN 102173987B
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acid
chinese catalpa
cell
phenyl
catalpa skin
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CN102173987A (en
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李玉新
张玉伟
鲍永利
林华
郑丽华
于春雷
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Northeast Normal University
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Abstract

The invention relates to a new polyunsaturated fatty acid compound--new manchurian walnut bark acid and structurally modified products thereof, and provides the preparation methods of the new manchurian walnut bark acid and the structurally modified products thereof. The new polyunsaturated fatty acid compound--new manchurian walnut bark acid is extracted from manchurian walnut bark, and the structurally modified products of the new manchurian walnut bark acid is prepared by structural modification and reconstruction of the manchurian walnut bark acid. Simultaneously, the invention also relates to the application of the compounds as drugs, especially as anticancer drugs.

Description

Chinese catalpa skin eo-acid and derivative thereof and their preparation method and medical usage
Technical field
The present invention relates to a kind of fatty acid new compound and be the derivatives quasi-compound shown in general formula that parent was synthesized with it, also disclose their preparation method and their medical usage simultaneously.Pharmaceutically active ingredient separation and purification and structural modification field in the invention belongs to.
Background technology
Malignant tumour is common disease, the frequently-occurring disease of serious threat human health.Pharmacological agent at present remains one of main methods for the treatment of of cancer, but existing chemotherapeutics is owing to exist problems such as resistance and toxic side effect, cause the effect of cancer drug treatment unsatisfactory, therefore seeking cancer therapy drug efficient, low toxicity becomes the focus that people pay close attention to.In recent decades, from natural product, seek the focus that desirable antitumor drug or prodrug have become research, and the anti-tumor medicine that much derives from natural product goes on the market.
Juglans mandshurica ( Juglans mandshuricaMaxim), having another name called juglans mandshurica, Chinese catalpa subtree, Semen Caryae Cathayensis (northeast), be the Juglandaceae walnut, is one of important medicine source plant resource of China, the existing history for many years of cancer that is used for the treatment of among the people in China and Korea S.We adopt the active method of separating of following the trail of, from the stem skin of Juglans mandshurica, separate obtain a kind of new ωThe polyunsaturated fatty acid compounds of-9 series, namely (7 E, 9 E)-6,11-dioxononadeca-7,9-dienoic acid(is called for short: Chinese catalpa skin eo-acid).Our result of study shows that this compound has better antitumor activity.
Unique texture and better antitumor activity thereof in view of Chinese catalpa skin eo-acid, we have carried out structural modification to Chinese catalpa skin eo-acid monomeric compound, be the antitumor lead compound of parent with the natural product for seeking highly active, synthesized the new acid derivative A of Chinese catalpa skin, B, C and D compounds shown in general formula (2).We further result of study show that the new acid derivative of Chinese catalpa skin also has better antitumor activity.
Summary of the invention
One of purpose of the present invention provides the compound Chinese catalpa skin eo-acid that a kind of structural formula is formula (1).
Figure 2011100533515100002DEST_PATH_IMAGE001
Formula (1)
Described compound is white crystalline powder (petroleum ether-ethyl acetate), and mp 120-121 ℃, molecular formula is C 18H2 8O 4, molecular weight is 308, is one not see the new compound of bibliographical information.
Two of purpose of the present invention provide a kind of from Juglandaceae walnut Juglans mandshurica the method for separation and purification Chinese catalpa skin eo-acid.Its major technique feature is as follows:
(a) with Juglans mandshurica ( Juglans MandshuricaMaxim.) the dry meal of stem skin or branches and leaves or flower or fruit food or root or root skin is raw material;
(b) the 8-10 volume fraction doubly with getting raw materials quality is that 10 ~ 95% ethanolic soln continuous backflow are extracted each 2 hours 2 ~ 4 times;
(c) united extraction liquid, concentrating under reduced pressure gets black medicinal extract, it is suspended in gets suspension in the deionized water again;
(d) with this suspension with isopyknic petroleum ether extraction 3～4 times, merge each time extraction liquid, concentrating under reduced pressure;
(e) the gained petroleum ether extract is again through silica gel decompression column chromatography, obtains 6 streams of I-VI part with the petroleum ether-ethyl acetate gradient elution, and a stream part V (is separated out white crystalline powder among the Petroleum Ether:EtOAc/65:35);
(f) with white powder again with petroleum ether-ethyl acetate repeatedly recrystallization obtain Chinese catalpa skin eo-acid, products obtained therefrom carries out structure by methods such as nucleus magnetic resonance to be identified.
Three of purpose of the present invention is A, B, C and the D analog derivatives that disclose Chinese catalpa skin eo-acid, and its chemical general formula is:
Figure 2011100533515100002DEST_PATH_IMAGE002
Wherein, R in the category-A 1For phenyl or to methoxyl group-phenyl or to methyl-phenyl or to amino-phenyl or 2-pyridyl or 4-(4 '-amino-phenoxy group)-phenyl or 4-(4 '-amino-benzenesulfonyl)-phenyl or to ethanoyl-phenyl;
R in the category-B 2Be hydrogen, R 3Be ethyl or R 2Be methyl, R 3Be methyl or R 2Be methylol, R 3Be methyl or R 2Be benzyl, R 3Be methyl or R 2Be (2-methyl)-propyl group, R 3Be methyl or R 2Be sec.-propyl, R 3Be methyl
Or R 2Be 4-hydroxyl-benzyl, R 3Be methyl;
R in the C class 4For methyl or ethyl or propyl group or normal-butyl or n-pentyl or n-octyl or dodecyl or n-tetradecane base or n-hexadecyl or Octadecane base or (5-methyl-2-sec.-propyl)-cyclohexyl or phenyl or the tertiary butyl-phenyl or to methoxycarbonyl base-phenyl or acetparaminosalol-phenyl;
R in the D class 5Be hydrogen or phenyl or sec.-propyl or benzyl or methyl;
Preferred following any one compound of category-A in the new acid derivative of described Chinese catalpa skin:
Numbering Title
A-1 N (phenyl)-Chinese catalpa skin eo-acid acid amides
A-2 N (4-methoxyl group-phenyl)-Chinese catalpa skin eo-acid acid amides
A-3 N (4-methyl-phenyl)-Chinese catalpa skin eo-acid acid amides
A-4 N (4-amino-phenyl)-Chinese catalpa skin eo-acid acid amides
A-5 N (2-pyridyl)-Chinese catalpa skin eo-acid acid amides
A-6 N [4-(4'-amino-phenoxy group) phenyl]-Chinese catalpa skin eo-acid acid amides
A-7 N [4-(4'-amino-benzenesulfonyl) phenyl]-Chinese catalpa skin eo-acid acid amides
A-8 N (4-ethanoyl-phenyl)-Chinese catalpa skin eo-acid acid amides
Preferred following any one compound of category-B in the new acid derivative of described Chinese catalpa skin:
Numbering Title
B-1 N (glycine ethyl ester)-Chinese catalpa skin eo-acid acid amides
B-2 N (L-alanine methyl ester)-Chinese catalpa skin eo-acid acid amides
B-3 N (L-serine methylester)-Chinese catalpa skin eo-acid acid amides
B-4 N (L-phenylalanine methyl ester)-Chinese catalpa skin eo-acid acid amides
B-5 N (L-leucine methyl esters)-Chinese catalpa skin eo-acid acid amides
B-6 N (L-valine methyl ester)-Chinese catalpa skin eo-acid acid amides
B-7 N (L-L-Tyrosine methyl ester)-Chinese catalpa skin eo-acid acid amides
Preferred following any one compound of C class in the new acid derivative of described Chinese catalpa skin:
Numbering Title
C-1 Chinese catalpa skin eo-acid methyl esters
C-2 The new acetoacetic ester of Chinese catalpa skin
C-3 The new propyl propionate of Chinese catalpa skin
C-4 The positive butyl ester of Chinese catalpa skin eo-acid
C-5 Chinese catalpa skin eo-acid n-pentyl ester
C-6 Chinese catalpa skin eo-acid n-octyl
C-7 Chinese catalpa skin eo-acid n-dodecane ester
C-8 Chinese catalpa skin eo-acid n-tetradecane ester
C-9 Chinese catalpa skin eo-acid n-hexadecane ester
C-10 Chinese catalpa skin eo-acid Octadecane ester
C-11 Chinese catalpa skin eo-acid menthol ester
C-12 The new acid phenenyl ester of Chinese catalpa skin
C-13 Chinese catalpa skin eo-acid (to the tertiary butyl-benzene)-ester
C-14 Chinese catalpa skin eo-acid (to methoxycarbonyl base-benzene)-ester
C-15 Chinese catalpa skin eo-acid (acetparaminosalol-benzene)-ester
Preferred following any one compound of D class in the new acid derivative of described Chinese catalpa skin:
Numbering Title
D-1 [(methylol) methyl]-Chinese catalpa skin eo-acid acid amides
D-2 [(S-methylol, phenyl) methyl]-Chinese catalpa skin eo-acid acid amides
D-3 [(S-methylol, sec.-propyl) methyl]-Chinese catalpa skin eo-acid acid amides
D-4 [(S-methylol, benzyl) methyl]-Chinese catalpa skin eo-acid acid amides
D-5 [(S-methylol, methyl) methyl]-Chinese catalpa skin eo-acid acid amides
Four of purpose of the present invention provides the preparation method of Chinese catalpa skin eo-acid A, B, C and D analog derivative.
Synthesizing of category-A derivative
In the THF system, be raw material with Chinese catalpa skin eo-acid and various aromatic amine, under condensing agent DCC/HOBt effect, synthesized the category-A derivative:
Figure 2011100533515100002DEST_PATH_IMAGE003
Synthesizing of category-B derivative
In the THF system, be raw material with Chinese catalpa skin eo-acid and various L-amino acid ester, under condensing agent DCC/HOBt/NMM effect, synthesized the category-B derivative:
Figure 2011100533515100002DEST_PATH_IMAGE004
Synthesizing of C analog derivative
In the THF/ acetonitrile system, be raw material with Chinese catalpa skin eo-acid and various Fatty Alcohol(C12-C14 and C12-C18), aromatic alcohol, under condensing agent DCC/DMAP effect, synthesized the C analog derivative:
Figure 2011100533515100002DEST_PATH_IMAGE005
Synthesizing of D analog derivative
In the THF/ acetonitrile system, be raw material with Chinese catalpa skin eo-acid and various S-type chiral amino alcohol, under condensing agent DCC/DMAP effect, synthesized the D analog derivative:
Figure 2011100533515100002DEST_PATH_IMAGE006
Five of purpose of the present invention provides Chinese catalpa skin eo-acid and the purposes of derivative aspect the preparation medicine, the especially purposes aspect the preparation antitumor drug.
Chinese catalpa skin eo-acid of the present invention and the purposes of derivative in oncotherapy thereof prove in the following manner :
1. the anti tumor activity in vitro of Chinese catalpa skin eo-acid research
Adopt the MTT method to carry out the anti tumor activity in vitro research of Chinese catalpa skin eo-acid.Each tumour cell is in 96 orifice plates respectively, and cell density is 5 * 10 4Individual/ml; Add Chinese catalpa skin eo-acid next day, concentration is respectively 100 μ g/ml, 10 μ g/ml, 1 μ g/ml medicine dilutes with DMSO, establishes the DMSO control group simultaneously, establishes 3 multiple holes for every group, and the administration volume is 100 μ l/ holes; Cell is containing 5% CO 237 ℃ of incubators continue to cultivate 44h after, every hole adds 20 μ l MTT(5mg/ml), continue to cultivate 4h; Supernatant is abandoned in suction, and every hole adds 100 μ l DMSO, on microplate reader, and vibration 600s, the OD value at detection 570nm place; More than the independent experiment triplicate.The result shows, the Chinese catalpa skin eo-acid that the present invention mentions is to various tumor cell strains, comprise human leukemia cell K562, human liver cancer cell HepG2, human liver cancer cell HepG2/ADM, human colon cancer cell HCT-8 cell, human breast cancer cell MCF-7, the human lung cancer cell A549, gastric carcinoma cells SGC7901, cervical cancer Hela and prostate cancer PC-3 all have inhibited proliferation in various degree, and it is less to the influence of normal cell L02 under respective concentration, show that Chinese catalpa skin eo-acid is in certain safety range, tumour cell to vitro culture has inhibited proliferation, the results are shown in Table 1.
Table 1 Chinese catalpa skin eo-acid is to the inhibiting rate of experiment with tumour cell
Figure DEST_PATH_IMAGE007
2. the anti tumor activity in vitro of N (phenyl)-Chinese catalpa skin eo-acid acid amides research
Adopt the MTT method to carry out anti tumor activity in vitro research.Each tumour cell is in 96 orifice plates respectively, and cell density is 5 * 10 4Individual/ml; Add N (phenyl)-Chinese catalpa skin eo-acid acid amides next day, concentration is respectively 100 μ g/ml, 10 μ g/ml, 1 μ g/ml medicine dilutes with DMSO, establishes the DMSO control group simultaneously, establishes 3 multiple holes for every group, and the administration volume is 100 μ l/ holes; Cell is containing 5% CO 237 ℃ of incubators continue to cultivate 44h after, every hole adds 20 μ l MTT(5mg/ml), continue to cultivate 4h; Supernatant is abandoned in suction, and every hole adds 100 μ l DMSO, on microplate reader, and vibration 600s, the OD value at detection 570nm place; More than the independent experiment triplicate.The result shows, the N that the present invention mentions (phenyl)-Chinese catalpa skin eo-acid acid amides comprises that to various tumor cell strains human leukemia cell K562, human liver cancer cell HepG2, human colon cancer cell HCT-8 cell, human breast cancer cell MCF-7, human lung cancer cell A549, gastric carcinoma cells SGC7901, cervical cancer Hela and prostate cancer PC-3 all have inhibited proliferation in various degree.And with less to the influence of normal cell L02 under the isoconcentration, show N (phenyl)-Chinese catalpa skin eo-acid acid amides in certain safety range, the tumour cell of vitro culture there is inhibited proliferation.N (phenyl)-Chinese catalpa skin eo-acid acid amides inhibiting rate to HepG2, HepG2/ADM, HCT-8, A549, K562 and L02 etc. under high, medium and low three concentration sees Table 2.
Table 2 N (phenyl)-Chinese catalpa skin eo-acid acid amides is to the inhibiting rate of experiment with tumour cell
Figure DEST_PATH_IMAGE008
3. the anti tumor activity in vitro of N (4-methoxyl group-phenyl)-Chinese catalpa skin eo-acid acid amides research
Adopt the MTT method to carry out anti tumor activity in vitro research.Each tumour cell is in 96 orifice plates respectively, and cell density is 5 * 10 4Individual/ml; Add N (4-methoxyl group-phenyl)-Chinese catalpa skin eo-acid acid amides next day, concentration is respectively 100 μ g/ml, 10 μ g/ml, 1 μ g/ml medicine dilutes with DMSO, establishes the DMSO control group simultaneously, establishes 3 multiple holes for every group, and the administration volume is 100 μ l/ holes; Cell is containing 5% CO 237 ℃ of incubators continue to cultivate 44h after, every hole adds 20 μ l MTT(5mg/ml), continue to cultivate 4h; Supernatant is abandoned in suction, and every hole adds 100 μ l DMSO, on microplate reader, and vibration 600s, the OD value at detection 570nm place; More than the independent experiment triplicate.The result shows, the N that the present invention mentions (4-methoxyl group-phenyl)-Chinese catalpa skin eo-acid acid amides is to various tumor cell strains, comprise human liver cancer cell HepG2, people's liver cancer mdr cell (HepG2/ADM), human leukemia cell K562, human colon cancer cell HCT-8 cell, human breast cancer cell MCF-7, the human lung cancer cell A549, gastric carcinoma cells SGC7901, cervical cancer Hela and prostate cancer PC-3 all have inhibited proliferation in various degree, and relative medicine concentration is less to the influence of normal cell L02, show N (4-methoxyl group-phenyl)-Chinese catalpa skin eo-acid acid amides in certain safety range, the tumour cell of vitro culture is had inhibited proliferation.N (4-methoxyl group-phenyl)-Chinese catalpa skin eo-acid acid amides inhibiting rate to HepG2, HepG2/ADM, HCT-8, A549, K562 and L02 etc. under high, medium and low three concentration sees Table 3.
Table 3N (4-methoxyl group-phenyl)-Chinese catalpa skin eo-acid acid amides is to the inhibiting rate of experiment with tumour cell
Figure DEST_PATH_IMAGE009
4. the anti tumor activity in vitro of N (glycine ethyl ester)-Chinese catalpa skin eo-acid acid amides research
Adopt the MTT method to carry out anti tumor activity in vitro research.Each tumour cell is in 96 orifice plates respectively, and cell density is 5 * 10 4Individual/ml; Add N (glycine ethyl ester)-Chinese catalpa skin eo-acid acid amides next day, concentration is respectively 100 μ g/ml, 10 μ g/ml, 1 μ g/ml medicine dilutes with DMSO, establishes the DMSO control group simultaneously, establishes 3 multiple holes for every group, and the administration volume is 100 μ l/ holes; Cell is containing 5% CO 237 ℃ of incubators continue to cultivate 44h after, every hole adds 20 μ l MTT(5mg/ml), continue to cultivate 4h; Supernatant is abandoned in suction, and every hole adds 100 μ l DMSO, on microplate reader, and vibration 600s, the OD value at detection 570nm place; More than the independent experiment triplicate.The result shows, the N that the present invention mentions (glycine ethyl ester)-Chinese catalpa skin eo-acid acid amides is to various tumor cell strains, comprise human liver cancer cell HepG2, people's liver cancer mdr cell (HepG2/ADM), human leukemia cell K562, human colon cancer cell HCT-8 cell, human breast cancer cell MCF-7, the human lung cancer cell A549, gastric carcinoma cells SGC7901, cervical cancer Hela and prostate cancer PC-3 all have inhibited proliferation in various degree, and respective concentration is less to the influence of normal cell L02, show N (glycine ethyl ester)-Chinese catalpa skin eo-acid acid amides in certain safety range, the tumour cell of vitro culture is had inhibited proliferation.N (glycine ethyl ester)-Chinese catalpa skin eo-acid acid amides inhibiting rate to HepG2, HepG2/ADM, HCT-8, A549, K562 and L02 etc. under high, medium and low three concentration sees Table 4.
Table 4N (glycine ethyl ester)-Chinese catalpa skin eo-acid acid amides is to the inhibiting rate of experiment with tumour cell
Figure DEST_PATH_IMAGE010
5. the anti tumor activity in vitro of N (L-serine methylester)-Chinese catalpa skin eo-acid acid amides research
Adopt the MTT method to carry out anti tumor activity in vitro research.Each tumour cell is in 96 orifice plates respectively, and cell density is 5 * 10 4Individual/ml; Add N (L-serine methylester)-Chinese catalpa skin eo-acid acid amides next day, concentration is respectively 100 μ g/ml, 10 μ g/ml, 1 μ g/ml medicine dilutes with DMSO, establishes the DMSO control group simultaneously, establishes 3 multiple holes for every group, and the administration volume is 100 μ l/ holes; Cell is containing 5% CO 237 ℃ of incubators continue to cultivate 44h after, every hole adds 20 μ l MTT(5mg/ml), continue to cultivate 4h; Supernatant is abandoned in suction, and every hole adds 100 μ l DMSO, on microplate reader, and vibration 600s, the OD value at detection 570nm place; More than the independent experiment triplicate.The result shows, the N that the present invention mentions (L-serine methylester)-Chinese catalpa skin eo-acid acid amides is to various tumor cell strains, comprise human liver cancer cell HepG2, people's liver cancer mdr cell (HepG2/ADM), human leukemia cell K562, human colon cancer cell HCT-8 cell, human breast cancer cell MCF-7, the human lung cancer cell A549, gastric carcinoma cells SGC7901, cervical cancer Hela and prostate cancer PC-3 all have inhibited proliferation in various degree, and it is less to align the influence of normal cell L02 with isoconcentration, show N (L-serine methylester)-Chinese catalpa skin eo-acid acid amides in certain safety range, the tumour cell of vitro culture is had inhibited proliferation.N (L-serine methylester)-Chinese catalpa skin eo-acid acid amides inhibiting rate to HepG2, HepG2/ADM, HCT-8, A549, K562 and L02 etc. under high, medium and low three concentration sees Table 5.
Table 5N (L-serine methylester)-Chinese catalpa skin eo-acid acid amides is to the inhibiting rate of experiment with tumour cell
Figure DEST_PATH_IMAGE011
6. the anti tumor activity in vitro of Chinese catalpa skin eo-acid methyl esters research
Adopt the MTT method to carry out anti tumor activity in vitro research.Each tumour cell is in 96 orifice plates respectively, and cell density is 5 * 10 4Individual/ml; Add Chinese catalpa skin eo-acid methyl esters next day, concentration is respectively 100 μ g/ml, 10 μ g/ml, 1 μ g/ml medicine dilutes with DMSO, establishes the DMSO control group simultaneously, establishes 3 multiple holes for every group, and the administration volume is 100 μ l/ holes; Cell is containing 5% CO 237 ℃ of incubators continue to cultivate 44h after, every hole adds 20 μ l MTT(5mg/ml), continue to cultivate 4h; Supernatant is abandoned in suction, and every hole adds 100 μ l DMSO, on microplate reader, and vibration 600s, the OD value at detection 570nm place; More than the independent experiment triplicate.The result shows, the Chinese catalpa skin eo-acid methyl esters that the present invention mentions is to various tumor cell strains, comprise human liver cancer cell HepG2, people's liver cancer mdr cell (HepG2/ADM), human leukemia cell K562, human colon cancer cell HCT-8 cell, human breast cancer cell MCF-7, the human lung cancer cell A549, gastric carcinoma cells SGC7901, cervical cancer Hela and prostate cancer PC-3 all have inhibited proliferation in various degree, and same concentration is less to the influence of normal cell L02, show Chinese catalpa skin eo-acid methyl esters in certain safety range, the tumour cell of vitro culture is had inhibited proliferation.Chinese catalpa skin eo-acid methyl esters inhibiting rate to HepG2, HepG2/ADM, HCT-8, A549, K562 and L02 etc. under high, medium and low three concentration sees Table 6.
Table 6Chinese catalpa skin eo-acid methyl esters is to the inhibiting rate of experiment with tumour cell
Figure DEST_PATH_IMAGE012
7. the anti tumor activity in vitro of the new acetoacetic ester of Chinese catalpa skin research
Adopt the MTT method to carry out anti tumor activity in vitro research.Each tumour cell is in 96 orifice plates respectively, and cell density is 5 * 10 4Individual/ml; Add the new acetoacetic ester of Chinese catalpa skin next day, concentration is respectively 100 μ g/ml, 10 μ g/ml, 1 μ g/ml medicine dilutes with DMSO, establishes the DMSO control group simultaneously, establishes 3 multiple holes for every group, and the administration volume is 100 μ l/ holes; Cell is containing 5% CO 237 ℃ of incubators continue to cultivate 44h after, every hole adds 20 μ l MTT(5mg/ml), continue to cultivate 4h; Supernatant is abandoned in suction, and every hole adds 100 μ l DMSO, on microplate reader, and vibration 600s, the OD value at detection 570nm place; More than the independent experiment triplicate.The result shows, the new acetoacetic ester of Chinese catalpa skin that the present invention mentions is to various tumor cell strains, comprise human liver cancer cell HepG2, people's liver cancer mdr cell (HepG2/ADM), human leukemia cell K562, human colon cancer cell HCT-8 cell, human breast cancer cell MCF-7, the human lung cancer cell A549, gastric carcinoma cells SGC7901, cervical cancer Hela and prostate cancer PC-3 all have inhibited proliferation in various degree, and it is less to align the influence of normal cell L02 with isoconcentration, show the new acetoacetic ester of Chinese catalpa skin in certain safety range, the tumour cell of vitro culture is had inhibited proliferation.The new acetoacetic ester of Chinese catalpa skin inhibiting rate to HepG2, HepG2/ADM, HCT-8, A549, K562 and L02 etc. under high, medium and low three concentration sees Table 7.
Table 7The new acetoacetic ester of Chinese catalpa skin is to the inhibiting rate of experiment with tumour cell
Figure DEST_PATH_IMAGE013
8. the anti tumor activity in vitro of [(methylol) methyl]-Chinese catalpa skin eo-acid acid amides research
Adopt the MTT method to carry out anti tumor activity in vitro research.Each tumour cell is in 96 orifice plates respectively, and cell density is 5 * 10 4Individual/ml; Add [(methylol) methyl]-Chinese catalpa skin eo-acid acid amides next day, and concentration is respectively 100 μ g/ml, 10 μ g/ml, 1 μ g/ml medicine dilutes with DMSO, establishes the DMSO control group simultaneously, establishes 3 multiple holes for every group, and the administration volume is 100 μ l/ holes; Cell is containing 5% CO 237 ℃ of incubators continue to cultivate 44h after, every hole adds 20 μ l MTT(5mg/ml), continue to cultivate 4h; Supernatant is abandoned in suction, and every hole adds 100 μ l DMSO, on microplate reader, and vibration 600s, the OD value at detection 570nm place; More than the independent experiment triplicate.The result shows, [(methylol) methyl]-Chinese catalpa skin eo-acid acid amides that the present invention mentions is to various tumor cell strains, comprise human liver cancer cell HepG2, people's liver cancer mdr cell (HepG2/ADM), human leukemia cell K562, human colon cancer cell HCT-8 cell, human breast cancer cell MCF-7, the human lung cancer cell A549, gastric carcinoma cells SGC7901, cervical cancer Hela and prostate cancer PC-3 all have inhibited proliferation in various degree, and it is less to align the influence of normal cell L02 with isoconcentration, show [(methylol) methyl]-Chinese catalpa skin eo-acid acid amides in certain safety range, the tumour cell of vitro culture is had inhibited proliferation.[(methylol) methyl]-Chinese catalpa skin eo-acid acid amides inhibiting rate to HepG2, HepG2/ADM, HCT-8, A549, K562 and L02 etc. under high, medium and low three concentration sees Table 8.
[0019] table 8[(methylol) methyl]-Chinese catalpa skin eo-acid acid amides is to the inhibiting rate of experiment with tumour cell
9. the anti tumor activity in vitro of [(S-methylol, phenyl) methyl]-Chinese catalpa skin eo-acid acid amides research
Adopt the MTT method to carry out anti tumor activity in vitro research.Each tumour cell is in 96 orifice plates respectively, and cell density is 5 * 10 4Individual/ml; Add [(S-methylol, phenyl) methyl]-Chinese catalpa skin eo-acid acid amides next day, and concentration is respectively 100 μ g/ml, 10 μ g/ml, 1 μ g/ml medicine dilutes with DMSO, establishes the DMSO control group simultaneously, establishes 3 multiple holes for every group, and the administration volume is 100 μ l/ holes; Cell is containing 5% CO 237 ℃ of incubators continue to cultivate 44h after, every hole adds 20 μ l MTT(5mg/ml), continue to cultivate 4h; Supernatant is abandoned in suction, and every hole adds 100 μ l DMSO, on microplate reader, and vibration 600s, the OD value at detection 570nm place; More than the independent experiment triplicate.The result shows, [(the S-methylol that the present invention mentions, phenyl) methyl]-Chinese catalpa skin eo-acid acid amides is to various tumor cell strains, comprise human liver cancer cell HepG2, people's liver cancer mdr cell (HepG2/ADM), human leukemia cell K562, human colon cancer cell HCT-8 cell, human breast cancer cell MCF-7, the human lung cancer cell A549, gastric carcinoma cells SGC7901, cervical cancer Hela and prostate cancer PC-3 all have inhibited proliferation in various degree, and it is less to align the influence of normal cell L02 with isoconcentration, show [(S-methylol, phenyl) methyl]-Chinese catalpa skin eo-acid acid amides in certain safety range, the tumour cell of vitro culture is had inhibited proliferation.[(S-methylol, phenyl) methyl]-Chinese catalpa skin eo-acid acid amides inhibiting rate to HepG2, HepG2/ADM, HCT-8, A549, K562 and L02 under high, medium and low three concentration sees Table 9.
Table 9[(S-methylol, phenyl) methyl]-Chinese catalpa skin eo-acid acid amides is to the inhibiting rate of experiment with tumour cell
Figure DEST_PATH_IMAGE015
10. the anti-tumor in vivo activity research of Chinese catalpa skin eo-acid and derivative thereof
With 5 * 10 6Individual tumor cell inoculation is subcutaneous in BALB/c (nu/nu) nude mice back.10 days began intravenous injection Chinese catalpa skin eo-acid or derivatives thereof 0.4mg/kg/ days behind tumor inoculation, injected altogether 15 days, and control group is then injected isopyknic physiological saline.Put to death animal after 30 days, strip tumour and weigh, calculate tumour inhibiting rate, tumour inhibiting rate (the %)=average knurl of [it is heavy that average knurl is organized in the average knurl weight-treatment of tumour control group]/tumour control group heavy * 100%.The result shows that injection Chinese catalpa skin eo-acid and derivative thereof all have tumor-inhibiting action in the tangible body.
Table 10. Chinese catalpa skin eo-acid and derivative thereof are to the influence of HepG2 growth of xenografted
Figure DEST_PATH_IMAGE016
Figure DEST_PATH_IMAGE017
Figure DEST_PATH_IMAGE018
Figure DEST_PATH_IMAGE019
Figure DEST_PATH_IMAGE022
Figure DEST_PATH_IMAGE023
Figure DEST_PATH_IMAGE024
Advantage of the present invention and positively effect are:Separate obtaining a kind of polyunsaturated fatty acid class new compound (hereinafter to be referred as Chinese catalpa skin eo-acid) first, and this compound is carried out structural modification and transformation, synthesized derivative of fatty acid A, B, C and D class series new compound shown in general formula.The pharmacologically active experimental result shows: above-claimed cpd has remarkable antitumor effect.
Pharmaceutical composition and methods for the treatment of
The method of pharmaceutical composition and treatment tumour also within the scope of the present invention.Described pharmaceutical composition comprises Chinese catalpa skin eo-acid of the present invention and derivative and the pharmaceutically acceptable carrier for the treatment of significant quantity." pharmaceutically acceptable carrier " comprises solvent, dispersion agent (dispersion medium), dressing (a coating), antibacterium and anti-mycotic agent and isotonic agent (isotonic agent), absorption delay agent (absorption delaying agent) and absorption enhancer etc.
Pharmaceutical composition of the present invention can be made the various pharmaceutical dosage forms that are adapted to different way of administration by traditional method.For example, it can be made into oral capsule, gel seal or tablet.Capsule can comprise pharmaceutically acceptable material such as gelatin, the Mierocrystalline cellulose etc. of any standard.Tablet can be about to pharmaceutical composition and solid phase carrier and lubricant compression by traditional method and make.Described solid phase carrier comprises starch and sugared bentonite (sugar bentonite).Pharmaceutical composition of the present invention also can be made into duricrust tablet (hard shell tablet) or comprises binding agent (binder) as the capsule of lactose or N.F,USP MANNITOL, conventional weighting agent and tableting agent.Pharmaceutical composition of the present invention also can pass through the parenteral route administration.The parenteral route form of administration comprise pharmaceutical composition of the present invention aqua, etc. open the sugar soln of salts solution or 5% and the preparation that forms with other pharmaceutically acceptable vehicle well known in the art.Cyclodextrin or other chaotropic agents known in those skilled in the art all can be used as pharmaceutical excipient and come submission pharmaceutical composition of the present invention.
Briefly say, Chinese catalpa skin eo-acid of the present invention and derivative thereof can hang and be dissolved in pharmaceutically acceptable carrier (as physiological solution), by oral or intravenous infusion, or by in subcutaneous, intramuscular, the thoracic cavity, in the intraperitoneal, internal rectum, intravaginal, nose, in the stomach, in the air flue, pulmonary injection or transfusion and aerosol sucks, spraying sucks, splash into and administration such as perfusion.
All multifactor influences such as diagnosis that the selection of formulation is subjected to route of administration, preparation type, patient's (sick kind, the state of an illness, the bodily form, body weight, body surface area, age, sex), medicine influences each other and accept the doctor for medical treatment.The preparation amount ranges that is suitable for is 0.01～100.00mg/kg.Amount ranges can be done corresponding adjustment with patient and route of administration different.It will depend primarily on the diagnosis of accepting the doctor for medical treatment.For example, oral dosage generally will be higher than intravenous injection dosage.Described dosage can be adjusted by experience optimization method well known in the art.Pharmaceutical composition of the present invention is wrapped in appropriate drug submission carrier (as polymer particle body or input unit) can improves the efficient of administration, particularly oral administration.
The activity of pharmaceutical composition of the present invention can by external ( In vitro) and body in ( In vivo) test and estimate.In brief, the pharmacologically active of pharmaceutical composition of the present invention is reflected on the anti-tumor activity.In the experiment, described pharmaceutical composition is injected in animal (as the mouse model) body to estimate its pharmacologically active in vivo.On this basis, suitable dosage ranges and route of administration are determined then.
For the ease of understanding the present invention, the spy enumerates following examples.Its effect should be understood to be to explaination of the present invention but not to any type of restriction of the present invention.
Embodiment
Following examples are only better understood the present invention for help those skilled in the art, but do not limit the present invention by any way.
Embodiment 1The preparation of Chinese catalpa skin eo-acid
Get the dry meal 4kg of Juglans mandshurica stem skin, be that 70% ethanolic soln continuous backflow is extracted 3 times with 50 liters of volume fractions, each 2 hours, united extraction liquid, concentrating under reduced pressure gets black medicinal extract, it is suspended in to get the 2L suspension in the deionized water again, and this suspension is used isopyknic petroleum ether extraction 3 times successively, the difference extract layer, concentrating under reduced pressure.Get petroleum ether extract (70g) silica gel column chromatography that reduces pressure, obtain 6 stream parts of I-VI with the petroleum ether-ethyl acetate gradient elution, wherein a stream part V (is separated out white crystalline powder among the Petroleum Ether:EtOAc/65:35), with petroleum ether-ethyl acetate repeatedly recrystallization obtain Chinese catalpa skin eo-acid, its structure appraising datum sees the following form.
Figure DEST_PATH_IMAGE025
Embodiment 2Synthetic [with the example that synthesizes of A-1] of the new acid derivative of category-A Chinese catalpa skin
Chinese catalpa skin eo-acid 0.155 g (0.5mmol) and DCC 0.112g (0.55mmol) are added in the reaction vessel, the THF that adds the 8ml drying, under the ice bath behind the stirring reaction 10min, 0.082 g (0.6mmol) HOBt is dissolved among the THF of 2ml drying, adds under the whipped state in the reaction vessel, under the ice bath behind the reaction 30min, 0.55 mmol aniline is dissolved among the THF of 3ml drying, be added dropwise to reaction vessel, behind the reaction 30min, be warming up to room temperature reaction naturally under the ice bath.After the thin layer detection reaction was finished, with the reaction solution concentrating under reduced pressure, residue separated [V (sherwood oil): V (ethyl acetate)=5:1～2:1] through silica gel column chromatography, namely gets target compound N (phenyl)-Chinese catalpa skin eo-acid, all the other category-A derivatives synthetic same as above.The structure of final category-A derivative all by ESI MS with 1Analytical procedures such as H NMR identify, with the formal specification of table:
Figure DEST_PATH_IMAGE026
Embodiment 3Synthetic [with the example that is collectively referred to as of B-5] of the new acid derivative of category-B Chinese catalpa skin
Chinese catalpa skin eo-acid 0.154g (0.5mmol) and DCC 0.112g (0.55mmol) are dropped in the reaction vessel, the THF that adds the 8ml drying, under the ice bath behind the stirring reaction 10min, 0.081g (0.6mmol) HOBt is dissolved among the THF of 2ml drying, add in the reaction vessel under the whipped state, after reacting 30min under the ice bath, 0.55 mmol L-leucine methyl ester hydrochloride and 0.1mlNMM are dissolved among the THF of 3ml drying, be added dropwise to reaction vessel, behind the reaction 30min, be warming up to room temperature reaction naturally under the ice bath.After the thin layer detection reaction was finished, with the reaction solution concentrating under reduced pressure, residue separated [V (sherwood oil): V (ethyl acetate)=5:1～2:1] through silica gel column chromatography, namely gets target compound N (phenyl)-Chinese catalpa skin eo-acid, all the other category-B derivatives synthetic same as above.The structure of final category-B derivative all by ESI MS with 1Analytical procedures such as H NMR identify, with the formal specification of table:
Embodiment 4Synthetic [with the example that synthesizes of C-8] of the new acid derivative of C class Chinese catalpa skin
Chinese catalpa skin eo-acid 0.156g (0.5mmol) and DCC0.113g (0.55mmol) are added in the reaction vessel, add the THF of 8ml drying, ice bath stirs down 0.032 g (0.25mmol) DMAP is added reaction system.After reacting 30min under the ice bath, the positive tetradecyl alcohol of 0.129g (0.6mmol) is added reaction system, naturally after being warming up to room temperature reaction 8～10h behind the stirring reaction 30min under the ice bath, filter out deposit D CU, filtrate decompression concentrates, residue separates [V (sherwood oil): V (ethyl acetate)=5:1～2:1] through silica gel column chromatography, namely gets target compound C-8, all the other C analog derivatives synthetic same as above.The structure of final C analog derivative all by ESI MS with 1Analytical procedures such as H NMR identify, with the formal specification of table:
Figure DEST_PATH_IMAGE028
Embodiment 5Synthetic [with the example that synthesizes of D-2] of the new acid derivative of D class Chinese catalpa skin
Chinese catalpa skin eo-acid 0.154 g (0.5mmol) and DCC0.112 g (0.55mmol) are added in the reaction vessel, the THF that adds the 8ml drying, under the ice bath behind the stirring reaction 10min, 0.081g (0.6mmol) HOBt is dissolved among the THF of 2ml drying, adds under the whipped state in the reaction vessel, under the ice bath behind the reaction 30min, 0.6 mmol benzene glycinol is dissolved among the THF of 3ml drying, be added dropwise to reaction vessel, behind the reaction 30min, be warming up to room temperature reaction naturally under the ice bath.After the thin layer detection reaction was finished, with the reaction solution concentrating under reduced pressure, residue separated [V (sherwood oil): V (ethyl acetate)=5:1～2:1] through silica gel column chromatography, namely gets target compound D-2, the synthetic experimentation with D-2 of all the other D analog derivatives.The structure of final D analog derivative all by ESI MS with 1Analytical procedures such as H NMR identify, with the formal specification of table:
Figure DEST_PATH_IMAGE029
Embodiment 6The inorganic acid salt of the new acid amide derivatives of Chinese catalpa skin or the preparation of organic acid salt
(1) preparation of inorganic acid salt, with the example that is prepared as of N [4-(4'-amino-phenoxy group) phenyl]-Chinese catalpa skin eo-acid acid amides (A-6) hydrochloride:
0.5 mmol N [4-(4'-amino-phenoxy group) phenyl]-Chinese catalpa skin eo-acid acid amides is joined in the aqueous hydrochloric acid of 20ml5%, after low-grade fever stirs and makes its dissolving, add adequate amount of ethanol or acetone, the refrigeration crystallization, filter, vacuum-drying namely gets N [4-(4'-amino-phenoxy group) phenyl]-Chinese catalpa skin eo-acid amide hydrochloride, yield 58%.
(2) preparation of organic acid salt, with the example that is prepared as of [(S-methylol, sec.-propyl) methyl]-Chinese catalpa skin eo-acid acid amides (D-3) acetate:
With 0.5 mmol [(S-methylol, sec.-propyl) methyl]-Chinese catalpa skin eo-acid acid amides joins in the reaction vessel that fills the 10ml methylene dichloride, add the 2ml Glacial acetic acid again, 30-40 ℃ of following stirring reaction 1-2h, the refrigeration crystallization of cooling back filters, and vacuum-drying namely gets [(S-methylol, sec.-propyl) methyl]-Chinese catalpa skin eo-acid acid amides acetate, yield 60%.
Figure DEST_PATH_IMAGE030
Figure DEST_PATH_IMAGE031
Figure DEST_PATH_IMAGE032
The anti tumor activity in vitro research of experimental example 1 Chinese catalpa skin eo-acid
Adopt the MTT method to carry out the anti tumor activity in vitro research of Chinese catalpa skin eo-acid.Each tumour cell is in 96 orifice plates respectively, and cell density is 5 * 10 4Individual/ml; Add Chinese catalpa skin eo-acid next day, concentration is respectively 100 μ g/ml, 10 μ g/ml, 1 μ g/ml medicine dilutes with DMSO, establishes the DMSO control group simultaneously, establishes 3 multiple holes for every group, and the administration volume is 100 μ l/ holes; Cell is containing 5% CO 237 ℃ of incubators continue to cultivate 44h after, every hole adds 20 μ l MTT(5mg/ml), continue to cultivate 4h; Supernatant is abandoned in suction, and every hole adds 100 μ l DMSO, on microplate reader, and vibration 600s, the OD value at detection 570nm place; More than the independent experiment triplicate.The result shows, the Chinese catalpa skin eo-acid that the present invention mentions is to various tumor cell strains, comprise human leukemia cell K562, human liver cancer cell HepG2, human liver cancer cell HepG2/ADM, human colon cancer cell HCT-8 cell, human breast cancer cell MCF-7, the human lung cancer cell A549, gastric carcinoma cells SGC7901, cervical cancer Hela and prostate cancer PC-3 all have inhibited proliferation in various degree, and it is less to the influence of normal cell L02 under respective concentration, show that Chinese catalpa skin eo-acid is in certain safety range, tumour cell to vitro culture has inhibited proliferation, the results are shown in Table 1.
The anti tumor activity in vitro research of experimental example 2 N (phenyl)-Chinese catalpa skin eo-acid acid amides
Adopt the MTT method to carry out anti tumor activity in vitro research.Each tumour cell is in 96 orifice plates respectively, and cell density is 5 * 10 4Individual/ml; Add N (phenyl)-Chinese catalpa skin eo-acid acid amides next day, concentration is respectively 100 μ g/ml, 10 μ g/ml, 1 μ g/ml medicine dilutes with DMSO, establishes the DMSO control group simultaneously, establishes 3 multiple holes for every group, and the administration volume is 100 μ l/ holes; Cell is containing 5% CO 237 ℃ of incubators continue to cultivate 44h after, every hole adds 20 μ l MTT(5mg/ml), continue to cultivate 4h; Supernatant is abandoned in suction, and every hole adds 100 μ l DMSO, on microplate reader, and vibration 600s, the OD value at detection 570nm place; More than the independent experiment triplicate.The result shows, the N that the present invention mentions (phenyl)-Chinese catalpa skin eo-acid acid amides comprises that to various tumor cell strains human leukemia cell K562, human liver cancer cell HepG2, human colon cancer cell HCT-8 cell, human breast cancer cell MCF-7, human lung cancer cell A549, gastric carcinoma cells SGC7901, cervical cancer Hela and prostate cancer PC-3 all have inhibited proliferation in various degree.And with less to the influence of normal cell L02 under the isoconcentration, show N (phenyl)-Chinese catalpa skin eo-acid acid amides in certain safety range, the tumour cell of vitro culture there is inhibited proliferation.N (phenyl)-Chinese catalpa skin eo-acid acid amides inhibiting rate to HepG2, HepG2/ADM, HCT-8, A549, K562 and L02 etc. under high, medium and low three concentration sees Table 2.
The anti tumor activity in vitro research of experimental example 3 N (4-methoxyl group-phenyl)-Chinese catalpa skin eo-acid acid amides
Adopt the MTT method to carry out anti tumor activity in vitro research.Each tumour cell is in 96 orifice plates respectively, and cell density is 5 * 10 4Individual/ml; Add N (4-methoxyl group-phenyl)-Chinese catalpa skin eo-acid acid amides next day, concentration is respectively 100 μ g/ml, 10 μ g/ml, 1 μ g/ml medicine dilutes with DMSO, establishes the DMSO control group simultaneously, establishes 3 multiple holes for every group, and the administration volume is 100 μ l/ holes; Cell is containing 5% CO 237 ℃ of incubators continue to cultivate 44h after, every hole adds 20 μ l MTT(5mg/ml), continue to cultivate 4h; Supernatant is abandoned in suction, and every hole adds 100 μ l DMSO, on microplate reader, and vibration 600s, the OD value at detection 570nm place; More than the independent experiment triplicate.The result shows, the N that the present invention mentions (4-methoxyl group-phenyl)-Chinese catalpa skin eo-acid acid amides is to various tumor cell strains, comprise human liver cancer cell HepG2, people's liver cancer mdr cell (HepG2/ADM), human leukemia cell K562, human colon cancer cell HCT-8 cell, human breast cancer cell MCF-7, the human lung cancer cell A549, gastric carcinoma cells SGC7901, cervical cancer Hela and prostate cancer PC-3 all have inhibited proliferation in various degree, and relative medicine concentration is less to the influence of normal cell L02, show N (4-methoxyl group-phenyl)-Chinese catalpa skin eo-acid acid amides in certain safety range, the tumour cell of vitro culture is had inhibited proliferation.N (4-methoxyl group-phenyl)-Chinese catalpa skin eo-acid acid amides inhibiting rate to HepG2, HepG2/ADM, HCT-8, A549, K562 and L02 etc. under high, medium and low three concentration sees Table 3.
The anti tumor activity in vitro research of experimental example 4 N (glycine ethyl ester)-Chinese catalpa skin eo-acid acid amides
Adopt the MTT method to carry out anti tumor activity in vitro research.Each tumour cell is in 96 orifice plates respectively, and cell density is 5 * 10 4Individual/ml; Add N (glycine ethyl ester)-Chinese catalpa skin eo-acid acid amides next day, concentration is respectively 100 μ g/ml, 10 μ g/ml, 1 μ g/ml medicine dilutes with DMSO, establishes the DMSO control group simultaneously, establishes 3 multiple holes for every group, and the administration volume is 100 μ l/ holes; Cell is containing 5% CO 237 ℃ of incubators continue to cultivate 44h after, every hole adds 20 μ l MTT(5mg/ml), continue to cultivate 4h; Supernatant is abandoned in suction, and every hole adds 100 μ l DMSO, on microplate reader, and vibration 600s, the OD value at detection 570nm place; More than the independent experiment triplicate.The result shows, the N that the present invention mentions (glycine ethyl ester)-Chinese catalpa skin eo-acid acid amides is to various tumor cell strains, comprise human liver cancer cell HepG2, people's liver cancer mdr cell (HepG2/ADM), human leukemia cell K562, human colon cancer cell HCT-8 cell, human breast cancer cell MCF-7, the human lung cancer cell A549, gastric carcinoma cells SGC7901, cervical cancer Hela and prostate cancer PC-3 all have inhibited proliferation in various degree, and respective concentration is less to the influence of normal cell L02, show N (glycine ethyl ester)-Chinese catalpa skin eo-acid acid amides in certain safety range, the tumour cell of vitro culture is had inhibited proliferation.N (glycine ethyl ester)-Chinese catalpa skin eo-acid acid amides inhibiting rate to HepG2, HepG2/ADM, HCT-8, A549, K562 and L02 etc. under high, medium and low three concentration sees Table 4.
The anti tumor activity in vitro research of experimental example 5 N (L-serine methylester)-Chinese catalpa skin eo-acid acid amides
Adopt the MTT method to carry out anti tumor activity in vitro research.Each tumour cell is in 96 orifice plates respectively, and cell density is 5 * 10 4Individual/ml; Add N (L-serine methylester)-Chinese catalpa skin eo-acid acid amides next day, concentration is respectively 100 μ g/ml, 10 μ g/ml, 1 μ g/ml medicine dilutes with DMSO, establishes the DMSO control group simultaneously, establishes 3 multiple holes for every group, and the administration volume is 100 μ l/ holes; Cell is containing 5% CO 237 ℃ of incubators continue to cultivate 44h after, every hole adds 20 μ l MTT(5mg/ml), continue to cultivate 4h; Supernatant is abandoned in suction, and every hole adds 100 μ l DMSO, on microplate reader, and vibration 600s, the OD value at detection 570nm place; More than the independent experiment triplicate.The result shows, the N that the present invention mentions (L-serine methylester)-Chinese catalpa skin eo-acid acid amides is to various tumor cell strains, comprise human liver cancer cell HepG2, people's liver cancer mdr cell (HepG2/ADM), human leukemia cell K562, human colon cancer cell HCT-8 cell, human breast cancer cell MCF-7, the human lung cancer cell A549, gastric carcinoma cells SGC7901, cervical cancer Hela and prostate cancer PC-3 all have inhibited proliferation in various degree, and it is less to align the influence of normal cell L02 with isoconcentration, show N (L-serine methylester)-Chinese catalpa skin eo-acid acid amides in certain safety range, the tumour cell of vitro culture is had inhibited proliferation.N (L-serine methylester)-Chinese catalpa skin eo-acid acid amides inhibiting rate to HepG2, HepG2/ADM, HCT-8, A549, K562 and L02 etc. under high, medium and low three concentration sees Table 5.
The anti tumor activity in vitro research of experimental example 6 Chinese catalpa skin eo-acid methyl esters
Adopt the MTT method to carry out anti tumor activity in vitro research.Each tumour cell is in 96 orifice plates respectively, and cell density is 5 * 10 4Individual/ml; Add Chinese catalpa skin eo-acid methyl esters next day, concentration is respectively 100 μ g/ml, 10 μ g/ml, 1 μ g/ml medicine dilutes with DMSO, establishes the DMSO control group simultaneously, establishes 3 multiple holes for every group, and the administration volume is 100 μ l/ holes; Cell is containing 5% CO 237 ℃ of incubators continue to cultivate 44h after, every hole adds 20 μ l MTT(5mg/ml), continue to cultivate 4h; Supernatant is abandoned in suction, and every hole adds 100 μ l DMSO, on microplate reader, and vibration 600s, the OD value at detection 570nm place; More than the independent experiment triplicate.The result shows, the Chinese catalpa skin eo-acid methyl esters that the present invention mentions is to various tumor cell strains, comprise human liver cancer cell HepG2, people's liver cancer mdr cell (HepG2/ADM), human leukemia cell K562, human colon cancer cell HCT-8 cell, human breast cancer cell MCF-7, the human lung cancer cell A549, gastric carcinoma cells SGC7901, cervical cancer Hela and prostate cancer PC-3 all have inhibited proliferation in various degree, and same concentration is less to the influence of normal cell L02, show Chinese catalpa skin eo-acid methyl esters in certain safety range, the tumour cell of vitro culture is had inhibited proliferation.Chinese catalpa skin eo-acid methyl esters inhibiting rate to HepG2, HepG2/ADM, HCT-8, A549, K562 and L02 etc. under high, medium and low three concentration sees Table 6.
The anti tumor activity in vitro research of the new acetoacetic ester of experimental example 7 Chinese catalpa skins
Adopt the MTT method to carry out anti tumor activity in vitro research.Each tumour cell is in 96 orifice plates respectively, and cell density is 5 * 10 4Individual/ml; Add the new acetoacetic ester of Chinese catalpa skin next day, concentration is respectively 100 μ g/ml, 10 μ g/ml, 1 μ g/ml medicine dilutes with DMSO, establishes the DMSO control group simultaneously, establishes 3 multiple holes for every group, and the administration volume is 100 μ l/ holes; Cell is containing 5% CO 237 ℃ of incubators continue to cultivate 44h after, every hole adds 20 μ l MTT(5mg/ml), continue to cultivate 4h; Supernatant is abandoned in suction, and every hole adds 100 μ l DMSO, on microplate reader, and vibration 600s, the OD value at detection 570nm place; More than the independent experiment triplicate.The result shows, the new acetoacetic ester of Chinese catalpa skin that the present invention mentions is to various tumor cell strains, comprise human liver cancer cell HepG2, people's liver cancer mdr cell (HepG2/ADM), human leukemia cell K562, human colon cancer cell HCT-8 cell, human breast cancer cell MCF-7, the human lung cancer cell A549, gastric carcinoma cells SGC7901, cervical cancer Hela and prostate cancer PC-3 all have inhibited proliferation in various degree, and it is less to align the influence of normal cell L02 with isoconcentration, show the new acetoacetic ester of Chinese catalpa skin in certain safety range, the tumour cell of vitro culture is had inhibited proliferation.The new acetoacetic ester of Chinese catalpa skin inhibiting rate to HepG2, HepG2/ADM, HCT-8, A549, K562 and L02 etc. under high, medium and low three concentration sees Table 7.
The anti tumor activity in vitro research of experimental example 8 [(methylol) methyl]-Chinese catalpa skin eo-acid acid amides
Adopt the MTT method to carry out anti tumor activity in vitro research.Each tumour cell is in 96 orifice plates respectively, and cell density is 5 * 10 4Individual/ml; Add [(methylol) methyl]-Chinese catalpa skin eo-acid acid amides next day, and concentration is respectively 100 μ g/ml, 10 μ g/ml, 1 μ g/ml medicine dilutes with DMSO, establishes the DMSO control group simultaneously, establishes 3 multiple holes for every group, and the administration volume is 100 μ l/ holes; Cell is containing 5% CO 237 ℃ of incubators continue to cultivate 44h after, every hole adds 20 μ l MTT(5mg/ml), continue to cultivate 4h; Supernatant is abandoned in suction, and every hole adds 100 μ l DMSO, on microplate reader, and vibration 600s, the OD value at detection 570nm place; More than the independent experiment triplicate.The result shows, [(methylol) methyl]-Chinese catalpa skin eo-acid acid amides that the present invention mentions is to various tumor cell strains, comprise human liver cancer cell HepG2, people's liver cancer mdr cell (HepG2/ADM), human leukemia cell K562, human colon cancer cell HCT-8 cell, human breast cancer cell MCF-7, the human lung cancer cell A549, gastric carcinoma cells SGC7901, cervical cancer Hela and prostate cancer PC-3 all have inhibited proliferation in various degree, and it is less to align the influence of normal cell L02 with isoconcentration, show [(methylol) methyl]-Chinese catalpa skin eo-acid acid amides in certain safety range, the tumour cell of vitro culture is had inhibited proliferation.[(methylol) methyl]-Chinese catalpa skin eo-acid acid amides inhibiting rate to HepG2, HepG2/ADM, HCT-8, A549, K562 and L02 etc. under high, medium and low three concentration sees Table 8.
The anti tumor activity in vitro research of experimental example 9 [(S-methylol, phenyl) methyl]-Chinese catalpa skin eo-acid acid amides
Adopt the MTT method to carry out anti tumor activity in vitro research.Each tumour cell is in 96 orifice plates respectively, and cell density is 5 * 10 4Individual/ml; Add [(S-methylol, phenyl) methyl]-Chinese catalpa skin eo-acid acid amides next day, and concentration is respectively 100 μ g/ml, 10 μ g/ml, 1 μ g/ml medicine dilutes with DMSO, establishes the DMSO control group simultaneously, establishes 3 multiple holes for every group, and the administration volume is 100 μ l/ holes; Cell is containing 5% CO 237 ℃ of incubators continue to cultivate 44h after, every hole adds 20 μ l MTT(5mg/ml), continue to cultivate 4h; Supernatant is abandoned in suction, and every hole adds 100 μ l DMSO, on microplate reader, and vibration 600s, the OD value at detection 570nm place; More than the independent experiment triplicate.The result shows, [(the S-methylol that the present invention mentions, phenyl) methyl]-Chinese catalpa skin eo-acid acid amides is to various tumor cell strains, comprise human liver cancer cell HepG2, people's liver cancer mdr cell (HepG2/ADM), human leukemia cell K562, human colon cancer cell HCT-8 cell, human breast cancer cell MCF-7, the human lung cancer cell A549, gastric carcinoma cells SGC7901, cervical cancer Hela and prostate cancer PC-3 all have inhibited proliferation in various degree, and it is less to align the influence of normal cell L02 with isoconcentration, show [(S-methylol, phenyl) methyl]-Chinese catalpa skin eo-acid acid amides in certain safety range, the tumour cell of vitro culture is had inhibited proliferation.[(S-methylol, phenyl) methyl]-Chinese catalpa skin eo-acid acid amides inhibiting rate to HepG2, HepG2/ADM, HCT-8, A549, K562 and L02 under high, medium and low three concentration sees Table 9.
The anti-tumor in vivo activity research of experimental example 10 Chinese catalpa skin eo-acids and derivative thereof
With 5 * 10 6Individual tumor cell inoculation is subcutaneous in BALB/c (nu/nu) nude mice back.10 days began intravenous injection Chinese catalpa skin eo-acid or derivatives thereof 0.4mg/kg/ days behind tumor inoculation, injected altogether 15 days, and control group is then injected isopyknic physiological saline.Put to death animal after 30 days, strip tumour and weigh, calculate tumour inhibiting rate, tumour inhibiting rate (the %)=average knurl of [it is heavy that average knurl is organized in the average knurl weight-treatment of tumour control group]/tumour control group heavy * 100%.The result shows that injection Chinese catalpa skin eo-acid and derivative thereof all have tumor-inhibiting action (seeing Table 10-18) in the tangible body.

Claims (1)

1. polyunsaturated fatty acid compounds Chinese catalpa skin eo-acid, its structural formula is as the formula (1);
Figure 2011100533515100001DEST_PATH_IMAGE001
。
Formula (1)
2. the preparation method of compound according to claim 1 is characterized in that:
(a) with Juglans mandshurica ( Juglans MandshuricaMaxim.) the dry meal of stem skin or branches and leaves or flower or fruit food or root or root skin is raw material;
(b) the 8-10 volume fraction doubly with getting raw materials quality is that 10 ~ 95% ethanolic soln continuous backflow are extracted each 2 hours 2 ~ 4 times;
(c) united extraction liquid, concentrating under reduced pressure gets black medicinal extract, it is suspended in gets suspension in the deionized water again;
(d) with this suspension with isopyknic petroleum ether extraction 3～4 times, merge each time extraction liquid, concentrating under reduced pressure;
(e) the gained petroleum ether extract through silica gel decompression column chromatography, obtains 6 cuts of I-VI with the petroleum ether-ethyl acetate gradient elution again, and the cut V is to separate out white crystalline powder among petroleum ether-ethyl acetate/65:35;
(f) with white powder again with petroleum ether-ethyl acetate repeatedly recrystallization obtain Chinese catalpa skin eo-acid, products obtained therefrom carries out structure by magnetic nuclear resonance method to be identified.
3. new acid derivative of Chinese catalpa skin, it is characterized in that with Chinese catalpa skin eo-acid monomeric compound and aromatic amine or amino acid ester or alkyl alcohol or hand-type amino alcohol be raw material, adopt conventional condensation reaction to prepare this new fats acid derivative category-A, category-B, C class and D compounds, its general formula is:
Figure 599178DEST_PATH_IMAGE002
Wherein, R in the category-A 1For phenyl or to methoxyl group-phenyl or to methyl-phenyl or to amino-phenyl or 2-pyridyl-phenyl or 4-(4 '-amino-phenoxy group)-phenyl or 4-(4 '-amino-benzenesulfonyl)-phenyl or to ethanoyl-phenyl;
R in the category-B 2Be hydrogen, R 3Be ethyl or R 2Be methyl, R 3Be methyl or R 2Be methylol, R 3Be methyl or R 2Be benzyl, R 3Be methyl or R 2Be (2-methyl)-propyl group, R 3Be methyl or R 2Be sec.-propyl, R 3Be methyl or R 2Be 4-hydroxyl-benzyl, R 3Be methyl;
R in the C class 4For methyl or ethyl or propyl group or normal-butyl or n-pentyl or n-octyl or dodecyl or n-tetradecane base or n-hexadecyl or Octadecane base or (5-methyl-2-sec.-propyl)-cyclohexyl or phenyl or the tertiary butyl-phenyl or to methoxycarbonyl base-phenyl or acetparaminosalol-phenyl;
R in the D class 5Be hydrogen or phenyl or sec.-propyl or benzyl or methyl.
4. the new acid derivative of Chinese catalpa skin according to claim 3 is characterized in that A, B and D compounds and medically acceptable acid form salt, and wherein inorganic acid salt is hydrochloride, vitriol, nitrate or phosphoric acid salt; Organic acid salt is formate, acetate, oxalate, Citrate trianion, maleate, fumarate or amino acid salts.
5. the application of the new acid derivative of the described Chinese catalpa skin of the Chinese catalpa skin eo-acid described in the claim 1 and claim 3 in the anti-liver cancer of preparation, lung cancer, colorectal carcinoma, leukemia, cancer of the stomach, mammary cancer, cervical cancer and prostate cancer medicine.
6. be activeconstituents with any one compound in claim 1 described Chinese catalpa skin eo-acid and the new acid derivative of the described Chinese catalpa skin of claim 3, become pharmaceutical composition in conjunction with pharmaceutically acceptable vehicle or vehicle group.
7. the described pharmaceutical composition of claim 6 is characterized in that, described pharmaceutical composition can be injection, oral preparation, creme, spray.
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* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN1931816A (en) * 2006-10-11 2007-03-21 东北林业大学 Juglone extracting process
CN101456797A (en) * 2007-12-13 2009-06-17 中国科学院兰州化学物理研究所 Walnut powder with anticancer activity as well as preparation method and use thereof

Patent Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN1931816A (en) * 2006-10-11 2007-03-21 东北林业大学 Juglone extracting process
CN101456797A (en) * 2007-12-13 2009-06-17 中国科学院兰州化学物理研究所 Walnut powder with anticancer activity as well as preparation method and use thereof

Non-Patent Citations (3)

* Cited by examiner, † Cited by third party
Title
周靖等.山核桃青果皮中脂肪酸成分的色谱－质谱分析.《中国林副特产》.2002,(第3期), *
曲中原等.青龙衣不同萃取部位抗肿瘤活性研究.《上海中医药杂志》.2009,第43卷(第1期),
青龙衣不同萃取部位抗肿瘤活性研究;曲中原等;《上海中医药杂志》;2009;第43卷(第1期);87-90 *

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