CN106317157A - Polyhydroxy diketone cucuribitane triterpene as well as preparation method and application thereof - Google Patents

Polyhydroxy diketone cucuribitane triterpene as well as preparation method and application thereof Download PDF

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CN106317157A
CN106317157A CN201610701251.1A CN201610701251A CN106317157A CN 106317157 A CN106317157 A CN 106317157A CN 201610701251 A CN201610701251 A CN 201610701251A CN 106317157 A CN106317157 A CN 106317157A
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compound
methanol
chloroform
diketone
preparation
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CN106317157B (en
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魏华
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Zhongzhitang Pharmaceutical Shandong Co ltd
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Jishou University
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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07J—STEROIDS
    • C07J9/00—Normal steroids containing carbon, hydrogen, halogen or oxygen substituted in position 17 beta by a chain of more than two carbon atoms, e.g. cholane, cholestane, coprostane

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Abstract

The invention discloses polyhydroxy diketone cucuribitane triterpene separated and purified from rhizomes of hemsleya pengxianensis W. J. Chang var. jinfushanensis L. D. Shen & W. J. Chang as well as a preparation method and application thereof. The chemical structure and physicochemical properties of polyhydroxy diketone cucuribitane triterpene are determined by utilizing modern analytical means. Polyhydroxy diketone cucuribitane triterpene is named 2beta, 3alpha, 7beta, 16alpha, 20beta, 25-hexahydroxy cucurbitane terpene-5(E)-monoene-11,22-dione according to the naming rules of related compounds, is colorless powder, is freely soluble in chloroform and methanol and is a new compound. Functional tests prove that 2beta, 3alpha, 7beta, 16alpha, 20beta, 25-hexahydroxy cucurbitane terpene-5(E)-monoene-11,22-dione has stronger inhibiting effects on tumor cells, such as HeLa cells and KB cells, can serve as a raw material for preparing medicines for preventing and treating tumors and has stronger application value and a market prospect.

Description

A kind of polyhydroxy diketone class calabash alkane type triterpenoid and preparation method thereof and purposes
Technical field
The present invention relates to a kind of polyhydroxy diketone class cucurbitane type tetraterpene derivatives and preparation method thereof and purposes, refer specifically to 2β, 3α, 7β, 16α, 20β, 25-hexahydroxy cucurbitane terpene-5 (E)-monoene-11,22-diketone and preparation method thereof and purposes.
Background technology
Jinfo Shan Mountain Radix Hemsleyae Macrospermae (Hemsleya pengxianensis W. J. Chang var. jinfushanensis L. D. Shen & W. J. Chang) it is Cucurbitaceae (Fabaceae) Genus Hemsleya plant, originate in China region of Southeast, be born in sea Pull out in border and the mountain valley shrubbery of about 2000 meters.The fruit of Jinfo Shan Mountain Radix Hemsleyae Macrospermae is avette, long 4-5 centimetre, diameter 2.5-3.5 li Rice, peel surface has tiny verruca to distinguish with former mutation, and main active is respectively cucurbitane type Fourth Ring three Terpene and saponin thereof and Triterpenoids sapogenins and saponin thereof, have the multiple efficacies such as heat-clearing and toxic substances removing, anti-inflammation, clinically It is mainly used in treating the multiple diseases such as bacillary dysentery, various inflammation, ulcer, jaundice.
The drug effect of Jinfo Shan Mountain Radix Hemsleyae Macrospermae is mainly derived from cucurbitane type triterpenoid compound therein, therefore, development and utilization In Radix Hemsleyae Macrospermae cucurbitane type monomeric compound, excavate further its potential medical value, and the knot to monomer whose compound Structure and physicochemical property are determined and characterize, significant for developing Jinfo Shan Mountain Radix Hemsleyae Macrospermae resource.
Summary of the invention
The present invention overcomes the deficiencies in the prior art exactly, it is provided that a kind of the having of isolated from Jinfo Shan Mountain Radix Hemsleyae Macrospermae rhizome Important biomolecule activity and the polyhydroxy diketone class Cucurbitanes of industrialization value.This monomeric compound is from gold Buddhist Isolated first in the Radix Hemsleyae Macrospermae of mountain, after utilizing modern analysis means to characterize its structure and confirm its biological activity, according to relevantization The naming rule named 2 of compoundβ, 3α, 7β, 16α, 20β, 25-hexahydroxy cucurbitane terpene-5 (E)-monoene-11,22- Diketone, this compound is a noval chemical compound.
A kind of from Jinfo Shan Mountain Radix Hemsleyae Macrospermae rhizome the 2 of isolatedβ, 3α, 7β, 16α, 20β, 25-hexahydroxy cucurbitane Terpene-5 (E)-monoene-11,22-diketone, there is the structure that is shown below:
Above-mentioned from Jinfo Shan Mountain Radix Hemsleyae Macrospermae rhizome the 2 of isolatedβ, 3α, 7β, 16α, 20β, 25-hexahydroxy cucurbitane Terpene-5 (E)-monoene-11,22-diketone obtains as follows:
Jinfo Shan Mountain Radix Hemsleyae Macrospermae rhizome crushed after being dried is sieved, and adds 95% alcohol heating reflux and extracts 3 times, and each 1-3 hour, merging carried Take liquid, decompression and solvent recovery, after concentration total extractum, total extractum with after water-dispersible, use successively petroleum ether, chloroform, ethyl acetate, N-butanol extraction, extract is concentrated to dryness；Take ethyl acetate extract extractum silica gel column chromatography to separate, chloroform-methanol (1:0-0: 1) gradient elution, obtains 12 fraction Fr A-L, Fr.F part and uses chloroform-methanol as eluting after gel chromatography eluting again Liquid eluting removes pigment, and then the inverted middle pressure chromatographic column of sample is through MeOH-H2O (60:40; 70:30; 80:20; 90: 10) gradient elution, obtains four part Fr. F1-4, and wherein Fr. F3 separates through high-efficient liquid phase chromatogram purification, uses methanol-water Eluting, the eluent collected 16.7 minutes crystallizes and get final product.
In described heating and refluxing extraction, Jinfo Shan Mountain Radix Hemsleyae Macrospermae rhizome is 1:8-1:12 with the mass volume ratio of ethanol.
In described chloroform-methanol eluent, chloroform is 45:55-60:40 with the volume ratio of methanol.
In described methanol-water eluent, methanol is 80:20 with the volume ratio of water.
2β, 3α, 7β, 16α, 20β, 25-hexahydroxy cucurbitane terpene-5 (E)-monoene-11,22-diketone is without toner End, is soluble in chloroform, and methanol, by 2β, 3α, 7β, 16α, 20β, 25-hexahydroxy cucurbitane terpene-5 (E)-monoene-11, 22-diketone carries out extracorporeal anti-tumor pharmacodynamic experiment, and extracorporeal anti-tumor pharmacodynamic experiment utilizes MTT colorimetry.
With 2β, 3α, 7β, 16α, 20β, 25-hexahydroxy cucurbitane terpene-5 (E)-monoene-11,22-diketone is experiment Group, with Doxorubicin(doxorubicin, antitumor drug) be matched group, set up blank group simultaneously, experimental group, matched group and Blank group chooses HeLa(human cervical carcinoma) cell and KB(human mouth epidermoid carcinoma) it is experimental subject, after culture medium dilution, with 6 × The density of 104/ml is inoculated in 96 orifice plates, every hole 100 μ l, and after normally cultivating 24 hours in incubator, each group adds corresponding Medicine, makes the ultimate density of each group of medicine be respectively 2.5 μ g/ml (1 group), 5 μ g/ml (2 groups), 10 μ g/ml (3 groups), 20 μ g/ml (four groups), 40 μ g/ml (5 groups), set 5 concentration altogether, the multiple hole of each concentration 3；After cultivating 48 hours, in often Hole adds MTT 10 μ l dyeing；After continuing to cultivate four hours, inhaling and abandon original fluid, every hole adds DMSO 100 μ l, puts on shaking table low Speed vibration 10 min, make crystal fully dissolve, and detect optical density value, root at enzyme-linked immunosorbent assay instrument 570 nm wavelength 50% inhibition concentration (IC is calculated according to optical density value50, μ g/mL), optical density value calculates IC50Computational methods be existing known skill Art.Experimental group, matched group are to HeLa cell and the IC of KB cell50As shown in table 1.
Table 1
Group HeLa cell KB cell
Experimental group 2.9 ± 0.8 14.7 ± 1.6
Matched group 1.3 ± 0.11 0.89 ± 0.03
By the data of upper table it can be seen that of the present invention 2β, 3α, 7β, 16α, 20β, 25-hexahydroxy cucurbitane Terpene-5 (E)-monoene-11,22-diketone is respectively provided with certain inhibitory action to HeLa cell and KB cell, it is possible to anti-as preparation The raw material of curing oncoma medicine, possesses stronger commercial application and is worth.
Compared with prior art, the beneficial effects of the present invention is:
Isolated has the 2 of important anti-tumor activity from the Jinfo Shan Mountain Radix Hemsleyae Macrospermae rhizome firstβ, 3α, 7β, 16α, 20β, 25-hexahydroxy cucurbitane terpene-5 (E)-monoene-11,22-diketone, and utilize modern analysis means determine its chemical constitution and Physicochemical property.Prove through functional trial: 2β, 3α, 7β, 16α, 20β, 25-hexahydroxy cucurbitane terpene-5 (E)-monoene- 11,22-diketone have stronger inhibitory action to tumor cell, it is possible to as the raw material of preparation preventing and treating tumour medicine, have relatively Strong using value and market prospect.
Accompanying drawing explanation
Fig. 1 is 2β, 3α, 7β, 16α, 20β, 25-hexahydroxy cucurbitane terpene-5 (E)-monoene-11,22-diketone Schematic arrangement.
Fig. 2 is 2β, 3α, 7β, 16α, 20β, 25-hexahydroxy cucurbitane terpene-5 (E)-monoene-11,22-diketone Proton nmr spectra (1H-NMR).
Fig. 3 is 2β, 3α, 7β, 16α, 20β, 25-hexahydroxy cucurbitane terpene-5 (E)-monoene-11,22-diketone Carbon-13 nmr spectra (13C-APT).
Detailed description of the invention
Below in conjunction with specific embodiment, the present invention is further illustrated
The first step: Jinfo Shan Mountain Radix Hemsleyae Macrospermae rhizome (5.0 kg) crushed after being dried crosses 80 mesh sieves.Second step: medicinal powder adds 10 times amount second Alcohol heating and refluxing extraction 3 times, each 2 hours, united extraction liquid, decompression and solvent recovery, obtain total extractum 1033 g after concentration.3rd Step: the total extractum of Jinfo Shan Mountain Radix Hemsleyae Macrospermae rhizome adds after suitable quantity of water carries out dispersion process, respectively by petroleum ether, chloroform, ethyl acetate, positive fourth Alcohol extracts, and is extracted to colourless, is evaporated to do by extract, weighs to obtain petroleum ether part total extractum 56g, chloroform extract Total extractum 302g, ethyl acetate extract total extractum 151g, n-butanol portion total extractum 409 g.4th step: take ethyl acetate layer leaching Cream 151 g separates through silica gel column chromatography (100～200 mesh), and chloroform-methanol (1:0-0:1) gradient elution obtains 12 fractions Fr A-L.5th step: Fr.F part removes pigment through gel chromatography eluting, chloroform-methanol (45:55) as elution, Then the inverted middle pressure chromatographic column of sample is through MeOH-H2O (60:40; 70:30; 80:20;90:10) gradient elution, obtains Four part Fr. F1-4. the 6th steps: wherein Fr. F3 separates through high-efficient liquid phase chromatogram purification, uses methanol-water (80:20) Eluting, the eluent crystallization collected 16.7 minutes i.e. obtains colourless powder, is soluble in chloroform, methanol.
The structural characterization of above-mentioned colourless powder and confirmation are as follows:
Above-mentioned gained colourless powder is carried out proton nmr spectra (1H-NMR) and carbon-13 nmr spectra (13C-APT) analyze,1H- NMR spectra as in figure 2 it is shown,13C-APT spectrogram is as shown in Figure 3.
Fig. 2 and Fig. 3 being carried out spectrum analysis, is belonged at each for Fig. 2 and Fig. 3 peak, the peak of Fig. 2 and Fig. 3 belongs to such as table 2 institute Showing, by Fig. 2, Fig. 3 and the data of table 1, the chemical structural formula of colourless powder is as it is shown in figure 1, according to there being related compounds Naming rule named 2β, 3α, 7β, 16α, 20β, 25-hexahydroxy cucurbitane terpene-5 (E)-monoene-11,22-two Ketone.
English entitled 2β, 3α, 7β, 16α, 20β, 25-hexahydroxycucurbita-5(E)-ene-11, 22-dione。
Table 2 compound 11H-NMR and13C-NMR (150MHz, C5D5N) modal data
Above-mentioned simply presently preferred embodiments of the present invention, not makees any pro forma restriction to the present invention.Any it is familiar with this area Technical staff, in the case of without departing from technical solution of the present invention scope, all may utilize the technology contents of the disclosure above to this Inventive technique scheme makes many possible variations and modification, or is revised as the Equivalent embodiments of equivalent variations.Therefore, every not Depart from technical solution of the present invention content, according to the technology of the present invention essence to any simple modification made for any of the above embodiments, etc. With change and modification, all should fall in the range of technical solution of the present invention is protected.

Claims (8)

1. a compound, it is characterised in that: there is the structure that is shown below.
Compound the most according to claim 1, it is characterised in that: this compound is colourless powder, is soluble in chloroform, first Alcohol.
Compound the most according to claim 1, it is characterised in that: HeLa cell and KB cell are respectively provided with relatively by this compound Strong inhibitory action.
4. compound described in claim 1 or 2 or 3 in prophylaxis of tumours and prepares the application in antitumor drug.
5. the preparation method of compound described in claim 1-3, it is characterised in that: Jinfo Shan Mountain Radix Hemsleyae Macrospermae rhizome crushed after being dried mistake Sieve, add 95% alcohol heating reflux extract 3 times, each 1-3 hour, united extraction liquid, decompression and solvent recovery, after concentration always soak Cream, total extractum is with after water-dispersible, and successively with petroleum ether, chloroform, ethyl acetate, n-butanol extraction, extract is concentrated to dryness；Take second Extractum silica gel column chromatography in acetoacetic ester position separates, and chloroform-methanol (1:0-0:1) gradient elution obtains 12 fraction Fr A- L, Fr.F part removes pigment with chloroform-methanol as elution after gel chromatography eluting again, and then sample is inverted Middle pressure chromatographic column is through MeOH-H2O (60:40; 70:30; 80:20;90:10) gradient elution, obtains four part Fr. F1- 4, wherein Fr. F3 separates through high-efficient liquid phase chromatogram purification, uses methanol-water eluting, collects the eluent crystallization of 16.7 minutes i.e. ?.
The preparation method of compound the most according to claim 5: it is characterized in that: Jinfo Shan Mountain snow in described heating and refluxing extraction Gallbladder rhizome is 1:8-1:12 with the mass volume ratio of ethanol.
The preparation method of compound the most according to claim 5: it is characterized in that: chloroform in described chloroform-methanol eluent It is 45:55-60:40 with the volume ratio of methanol.
The preparation method of compound the most according to claim 5: it is characterized in that: in described methanol-water eluent methanol with The volume ratio of water is 80:20.
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Cited By (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN114133422A (en) * 2021-12-16 2022-03-04 中国医学科学院药用植物研究所 Cucurbitane triterpenoid compound and preparation method and application thereof
CN115160396A (en) * 2022-08-03 2022-10-11 河南中医药大学 Cucurbitane tetracyclic triterpenoid extracted from Hemsleya chinensis with anti-enteritis activity, and preparation method and application thereof

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CN103665082A (en) * 2013-12-10 2014-03-26 山东省医学科学院药物研究所 Hemsleya cucurbitane tetracyclic triterpenoid compound, pharmaceutical compositions containing same and application of compound and pharmaceutical composition

Patent Citations (1)

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CN103665082A (en) * 2013-12-10 2014-03-26 山东省医学科学院药物研究所 Hemsleya cucurbitane tetracyclic triterpenoid compound, pharmaceutical compositions containing same and application of compound and pharmaceutical composition

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Cited By (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN114133422A (en) * 2021-12-16 2022-03-04 中国医学科学院药用植物研究所 Cucurbitane triterpenoid compound and preparation method and application thereof
CN114133422B (en) * 2021-12-16 2022-12-20 中国医学科学院药用植物研究所 Cucurbitane triterpenoid compound and preparation method and application thereof
CN115160396A (en) * 2022-08-03 2022-10-11 河南中医药大学 Cucurbitane tetracyclic triterpenoid extracted from Hemsleya chinensis with anti-enteritis activity, and preparation method and application thereof
CN115160396B (en) * 2022-08-03 2024-04-05 河南中医药大学 Cucurbitane-type tetracyclic triterpene compound with anti-enteritis activity extracted from Chinese hemsleya root, and preparation method and application thereof

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