CN103012559A - Peptaibol compound and application thereof - Google Patents
Peptaibol compound and application thereof Download PDFInfo
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- CN103012559A CN103012559A CN2012105607219A CN201210560721A CN103012559A CN 103012559 A CN103012559 A CN 103012559A CN 2012105607219 A CN2012105607219 A CN 2012105607219A CN 201210560721 A CN201210560721 A CN 201210560721A CN 103012559 A CN103012559 A CN 103012559A
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- peptaibol
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- antineoplastic
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Abstract
The invention relates to a peptaibol compound and an application thereof. Trichodermaasperellum is cultured by fermentation, a fermentation product is obtained, and the compound is separated and purified from the fermentation product. The peptaibol compound belongs to an active linear small peptide, and a C end and an N end are unusually acetylated simultaneously. The peptaibol compound has antineoplastic activity. The compound can be used for preparing cell proliferation inhibitors or antineoplastic agents and for antineoplastic researches.
Description
Technical field
The present invention relates to a kind of sour jujube spore peptides and application thereof.
Background technology
Sour jujube spore peptides (peptaibol) mainly is the active linear little peptide of a class that derives from Trichoderma, and often being in the news has antibiotic, antimycotic, antiviral, parasiticide isoreactivity, and the C end is very rare with N end acetylizad sour jujube spore peptide of while.Inventor's research is learnt, trichoderma asperellum (Trichoderma asperellum) (was deposited in Wuhan University Chinese Typical Representative culture collection center on October 31st, 2012, deposit number is: CCTCC M 2012438) crude extract of liquid fermentation production has good cell inhibitory effect active, then its activeconstituents is studied.Sour jujube spore peptides had anti-tumor activity shown in research was found, had not yet to see the report of chemical structure and the cell inhibitory effect activity of this compound, so also there is not yet relevant therewith medicine on the market.
Summary of the invention
The object of the present invention is to provide a kind of sour jujube spore peptides and application thereof.This compound has the inhibition tumor cell proliferation function, has anti-tumor activity.Its structural formula is:
Its constitutional features is: contain six methylalanine residues, a α-amino-isovaleric acid residue, an Isoleucine residue, an alanine residue, and C end and the simultaneously acetylize of N end.
The present invention has also protected the purposes of described compound in the preparation inhibition of cell proliferation.And the purposes of this compound in the preparation antitumor drug.
Remarkable advantage of the present invention: the spore of sour jujube shown in research peptides belongs to active linear little peptide, and is rare C end and the simultaneously acetylize of N end.Described sour jujube spore peptides has anti-tumor activity, has not yet to see the report of chemical structure and the cell inhibitory effect activity of this compound, so also there is not yet relevant therewith medicine on the market.
Embodiment
The chemical structure of the compound of indication in following embodiment:
Fermentative production and the separation and purification of embodiment 1 this compound
1 fermentative production
Produce the fermentation culture of bacterium: by the ordinary method of culturing micro-organisms, get trichoderma asperellum (
Trichoderma asperellum) IBPT-2 (has been deposited in Chinese Typical Representative culture collection center on October 31st, 2012, address: Wuhan Wuhan University, deposit number is: CCTCC M 2012438) an amount of, and be inoculated on the PDA solid slant culture base and in 28 degrees centigrade of incubators, cultivated 4 days.
Get 4 days trichoderma asperellum of slant culture (Trichoderma asperellum) an amount of, be inoculated into 400mL nutrient solution [substratum composition (grams per liter): N.F,USP MANNITOL 20.0, yeast extract paste 3.0 is housed, maltose 20.0, monosodium glutamate 10.0, glucose 10.0, KH
2PO
40.5, MgSO
40.3, seawater] the 1000mL Erlenmeyer flask in, 28 ℃ of static cultivations are after 30 days, obtain mycelium and fermented liquid.
The acquisition of 2 medicinal extract
With gauze with mycelium and separation of fermentative broth.With fermented liquid and ethyl acetate 1:2 extracting twice, the extraction liquid underpressure distillation obtains the ethyl acetate extract of fermented liquid to doing.
The separation and purification of 3 compounds
Medicinal extract with 100-200 order silica gel mixed sample after, take sherwood oil: methylene dichloride: the methyl alcohol gradient composition is as elutriant, by the 300-400 order silica gel silica gel chromatography column chromatography that reduces pressure, obtains 5 components (A-E).Collect B component (methylene chloride-methanol 20:1 eluate), take sherwood oil: methylene dichloride: the methyl alcohol gradient composition by the pressurized silica gel column chromatography, gets 4 components (B1-B4) as elutriant.Collecting B component 3(methylene chloride-methanol is the eluate of 20:1) carry out Sephadex LH-20 gel filtration chromatography by chloroform-methanol (1:2) for solvent, pass through again the RP-18 reversed-phase silica gel column chromatography take methanol-water (3:2) as solvent elution, at last by half preparative liquid chromatography (1010 type ODS-A, 10 * 250 mm, 5 μ m): separating flow velocity is 5 mL/min, moving phase is 55 % acetonitrile solutions, compound shown in obtaining.
The compound clear crystal, positive ion HR-ESI-MS
M/z: 979.6203 [M+H]
+, molecular formula C
47H
82N
10O
12;[
α]
25 D-45 (c=0.1, MeOH);
1H and
13The NMR data such as C-NMR see Table 1.Negative ion second order ms fragmention (-) ESIMS/MS
M/z978,936,851,765,666,581,468; Positive ion second order ms fragmention (+) ESIMS/MS
M/z1002 [M+Na]+, 831,745,674,589,504,391.
Table 1 compound
1H and
13The C-NMR data (500 and 125MHz, in DMSO-d
6 )
a
1) this table signal ownership is based on DEPT, HMQC and HMQC spectrum analysis result.The multiple degree of carbon signal utilizes the DEPT method to determine.
2) numeral and the code name in this hurdle represents respectively
1H-
1In the H COSY spectrum with corresponding line in
1H provides the coupling coherent signal
1H nuclear.
3) numeral in this hurdle and code name represent respectively in HMBC spectrum with corresponding line in
1H provides the coupling coherent signal
13C nuclear.
4) A=Aib in this hurdle, V=Val, I=Ile, Al=Ala.
The test of embodiment 2 anti tumor activity in vitro
1 laboratory sample and experimental technique
The preparation specimen of sample solution is the pure compounds of separation and purification in the above-mentioned enforcement 1.Precision takes by weighing an amount of sample, is mixed with the solution of desired concn with methyl alcohol, and is active for surveying.
The succeeding transfer culture of clone and cell adopts people's lung cancer A549 cell system.Cell is with the RPMI-1640 substratum that contains 10%FBS, at 37 ℃ of succeeding transfer culture in the incubator that passes into 5% carbonic acid gas.
The cell inhibitory effect activity test method
People's lung cancer A549 cell that Sulforhodamine B (SRB) method is taken the logarithm vegetative period, being mixed with density with fresh RPMI-1640 substratum is every milliliter 2 * 10
5The cell suspension of individual cell is inoculated in the 96 porocyte culture plates by every hole 200 microlitres, and every hole adds sample or the blank solution of 2 microlitre different concns, in 37 ℃ of lower cultivations 24 hours.Be taken at the cell after cultivating under the drug effect, the morphological change that at first causes in optical microphotograph Microscopic observation drug treating is judged to have or not the cell cycle to suppress the morphological feature of necrocytosis, then 4 ℃, 3000 rev/mins centrifugal 3 minutes, suck supernatant liquor.Add 20% Tricholroacetic Acid, 50 microlitres in every porocyte, place 4 ℃ to fix 1 hour, water flushing 5 times and dry air.Every hole adds acetum 50 microlitres of 0.4% SRB and left standstill 30 minutes in room temperature.Clean 4 times with 1% acetic acid water, remove unconjugated free SRB dyestuff.Every hole adds 150 microlitre Tris and changes (100mmol/L, pH 10.5) soluble protein combination dye into and utilize microplate reader to measure every hole in optical density(OD) (OD) value at 520nm place.Each concentration of sample all arranges three holes in same 96 orifice plates, and other establishes three hole blanks and acellular zeroing hole (if medicine has color will do the acellular zeroing of relative medicine concentration).Each hole OD value is done first corresponding acellular zeroing, gets the average OD value in three holes again and calculates the sample on cell proliferation inhibiting rate (IR%) under each concentration by IR (%)=(OD blank-OD sample)/OD blank * 100% formula.Inhibiting rate according to different concns calculates, and draws IC
50Value.
2. experimental result
Cell inhibitory effect active testing result
In the srb assay test, this compound suppresses result: IC to the propagation of people's lung cancer A549 cell
50=0.52mg/mL.
3. conclusion
Compound has obvious Cytostatic to tumor cell effect, can be used as preparation inhibition of cell proliferation or antineoplastic agent and is used for antineoplastic research.
Claims (4)
1. compound
2. compound claimed in claim 1 is in the purposes of preparation in the inhibition of cell proliferation.
3. the purposes of described compound according to claim 2 is characterized in that: cell behaviour lung cancer A549 cell.
4. compound claimed in claim 1 is in the purposes of preparation in the antitumor drug.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201210560721.9A CN103012559B (en) | 2012-12-21 | 2012-12-21 | A kind of Peptaibol and use thereof and its application |
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| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201210560721.9A CN103012559B (en) | 2012-12-21 | 2012-12-21 | A kind of Peptaibol and use thereof and its application |
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| Publication Number | Publication Date |
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| CN103012559A true CN103012559A (en) | 2013-04-03 |
| CN103012559B CN103012559B (en) | 2018-07-17 |
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| CN201210560721.9A Expired - Fee Related CN103012559B (en) | 2012-12-21 | 2012-12-21 | A kind of Peptaibol and use thereof and its application |
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104211779A (en) * | 2013-05-30 | 2014-12-17 | 福州大学 | Heterocycle peptaibol compound, preparation method and purpose thereof |
| CN104211780B (en) * | 2013-05-30 | 2016-09-07 | 福州大学 | A kind of cyclic depsipeptides compound and its production and use |
Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102071148A (en) * | 2010-12-01 | 2011-05-25 | 山东大学 | Trichoderma pseudokoningii (SMF2) strain and application thereof |
-
2012
- 2012-12-21 CN CN201210560721.9A patent/CN103012559B/en not_active Expired - Fee Related
Patent Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102071148A (en) * | 2010-12-01 | 2011-05-25 | 山东大学 | Trichoderma pseudokoningii (SMF2) strain and application thereof |
Non-Patent Citations (3)
| Title |
|---|
| REN J.等: "Asperelines A-F, Peptaibols from the Marine-Derived Fungus Trichoderma asperellum", 《J. NAT. PROD.》 * |
| WILHELM C.等: "New Peptaibols from Mycogone cervina", 《J. NAT. PROD.》 * |
| 宋晓妍 等: "木霉peptaibols抗菌肽的研究进展", 《微生物学报》 * |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104211779A (en) * | 2013-05-30 | 2014-12-17 | 福州大学 | Heterocycle peptaibol compound, preparation method and purpose thereof |
| CN104211780B (en) * | 2013-05-30 | 2016-09-07 | 福州大学 | A kind of cyclic depsipeptides compound and its production and use |
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| Publication number | Publication date |
|---|---|
| CN103012559B (en) | 2018-07-17 |
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Address after: 350116, No. 2 School Road, Minhou New District, Fuzhou County, Fuzhou, Fujian Applicant after: Fuzhou University Address before: 350002 Fuzhou, Gulou District, Fujian Industrial Road, No. 523 Applicant before: Fuzhou University |
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Granted publication date: 20180717 Termination date: 20191221 |