CN114702467A - Aromatic casane diterpenoid compounds of flos Tiliae ananatis, extraction method and application - Google Patents

Aromatic casane diterpenoid compounds of flos Tiliae ananatis, extraction method and application Download PDF

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CN114702467A
CN114702467A CN202210514845.7A CN202210514845A CN114702467A CN 114702467 A CN114702467 A CN 114702467A CN 202210514845 A CN202210514845 A CN 202210514845A CN 114702467 A CN114702467 A CN 114702467A
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ethyl acetate
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methanol
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李春环
陈秀梅
张梓涵
卢旺
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Northwest A&F University
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Abstract

The invention discloses a jatropha aromatized casane diterpenoid compound, an extraction method and application thereof, comprising the steps of crushing stems of jatropha aromatized casane diterpenoid compounds, and extracting the crushed stems in alcohol by hot reflux to obtain a total extract; extracting the total extract with ethyl acetate, and repeatedly performing column chromatography to obtain ethyl acetate extract phase. The compound of the invention has simple preparation method, and the prepared compound or the pharmaceutically acceptable salt thereof can be used for preparing antibacterial agent preparations.

Description

Aromatic casane diterpenoid compounds of flos Tiliae ananatis, extraction method and application
Technical Field
The invention belongs to the field of natural medicinal chemistry, and particularly relates to a cymbidium aromatized cassan diterpenoid compound, an extraction method and application.
Background
The plants (Caesalpinia) belonging to the genus Caesalpinia of the family Leguminosae belong to about 100 species, are widely distributed in tropical and subtropical regions, and 17 species are distributed in China mainly from southwest to southern regions. According to literature reports, the plant of the genus is rich in chemical components, and mainly comprises diterpene, triterpene, sesquiterpene, alkaloid, saponin, glucoside and other compounds, wherein casane diterpene is a characteristic component of the genus and has wide pharmacological activity, and plays an important role in multiple aspects of antibiosis, malaria resistance, inflammation resistance, tumor resistance and the like, but few reports are provided for aromatic casane diterpene at present, and reports related to the antibacterial activity of the diterpene are not seen, so that the functional active ingredients of the diterpene still need to be further researched so as to better develop the medicinal value of the diterpene.
Disclosure of Invention
In view of the above disadvantages and drawbacks of the prior art, the present invention provides an aromatized casane diterpene compound of Tinospora japonica, an extraction method and applications thereof.
In order to achieve the purpose, the technical scheme is as follows: the invention provides a cymbidium aromatized casane diterpenoid compound, which has any one or more of the structures shown in formulas 1-12:
Figure BDA0003639102360000011
Figure BDA0003639102360000021
a method for extracting aromatic casane diterpene compounds from herba Saussureae Involueratae comprises pulverizing stem of herba Saussureae Involueratae, and extracting in alcohol under hot reflux to obtain total extract; extracting the total extract with ethyl acetate, and repeatedly performing column chromatography on the obtained ethyl acetate extract phase to obtain the final product.
Further, the alcohols include methanol and ethanol.
Furthermore, the extraction method specifically comprises the following steps:
drying stem of herba Pteridis Multifidae in the shade, pulverizing to 30 mesh, extracting with methanol under reflux to obtain extractive solution, concentrating the extractive solution under reduced pressure, recovering methanol to obtain crude extract, and extracting with ethyl acetate to obtain ethyl acetate extract phase;
dissolving an ethyl acetate extraction phase by using chloroform or acetone, carrying out column chromatography on silica gel for coarse separation, and carrying out gradient elution by using a chloroform/acetone system, wherein the volume ratio of the gradient elution of the chloroform/acetone system is 1:0, 9:1, 8:2, 7:3, 1:1 and 0:1 in sequence; taking sulfuric acid ethanol solution as a color developing agent, and carrying out color development according to TLC (thin layer chromatography) and combining similar fractions to obtain 8 components EA 1-EA 8;
after the EA2 component is subjected to silica gel column chromatography, gradient elution is carried out by using a petroleum ether/ethyl acetate system, the elution concentrations of the petroleum ether/ethyl acetate system are 30:1, 25:1, 20:1 and 10:1 in sequence, and 8 components 2A-2H are obtained according to the polarity;
component 2C sequentially undergoes forward silica gel chromatographic column and RP-C18Separating with reverse chromatographic column and Sephadx-LH20 gel chromatographic column to obtain compounds 4 and 5;
separating out crystals from the component 2D, and repeatedly recrystallizing and purifying the crystals by methanol to obtain a compound 2;
wash solution of component 2D in sequencePassing through forward chromatographic column, Sephadx-LH20 gel chromatographic column and RP-C18Separating with reverse chromatographic column to obtain compounds 3 and 9;
component 2E is separately RP-C18Separating with reverse chromatographic column, forward silica gel chromatographic column and Sephadx-LH20 gel chromatographic column to obtain compounds 1 and 10;
performing forward silica gel column chromatography on the component EA3, performing crude separation by using a petroleum ether-ethyl acetate system to obtain 9 components 69A-69G according to polarity, wherein the volume ratios of the petroleum ether-ethyl acetate system are 7:1, 5:1, 3:1 and 1:1 in sequence;
fraction 69D via RP-C18After semi-preparation of the liquid phase, the liquid phase is then subjected to RP-C18Subjecting to Sephadx-LH20 column chromatography to obtain compound 8, and subjecting to RP-C chromatography18The semi-preparative liquid phase uses methanol with volume concentration of 20%, 40%, 60%, 80% and 100% in sequence;
the component 69F sequentially passes through a chloroform-methanol gel column and an RP-C with the volume ratio of 1:118Subjecting the column to high performance liquid chromatography to obtain compounds 11 and 12;
the component 69G passes through RP-C in sequence18Performing chromatography on the chromatographic column and a Sephadx-LH20 gel column to obtain a compound 6;
carrying out forward silica gel column chromatography on the component EA-4 for crude separation, and obtaining a component 1a-1g according to the polarity; the component 1a is subjected to gradient elution by an MCI column, and the volume concentration of methanol subjected to gradient elution by the MCI column is 20%, 40%, 60%, 80% and 100% in sequence; and purifying the 60% methanol section by Sephadx-LH20 column chromatography to obtain a compound 7.
The invention also discloses application of the flos lonicerae aromatic casane diterpenoid compound or the pharmaceutically acceptable salt thereof in preparing an antibacterial agent preparation.
Furthermore, the antibacterial agent preparation is used for preventing and treating kiwifruit canker pathogen, bacillus cereus and staphylococcus aureus.
Furthermore, the antibacterial agent preparation is prepared into a pharmaceutically acceptable preparation by adding pharmaceutically acceptable auxiliary materials according to a conventional process, and the pharmaceutically acceptable preparation is a solid preparation or a liquid preparation.
Further, the solid preparation comprises granules, capsules, tablets and pills; the liquid formulation includes an injectable formulation.
Furthermore, in the antibacterial agent preparation, the content of the aromatic casane diterpenoid compound or the pharmaceutically acceptable salt thereof in the Tinospora bungeana is 2-8% by mass percentage.
Compared with the prior art, the invention has the beneficial technical effects that:
the compound of the invention has simple preparation method, and the prepared compound or the pharmaceutically acceptable salt thereof can be used for preparing antibacterial agent preparations. The antibacterial agent preparation prepared by using the compound of the present invention can be administered by oral, nasal inhalation, rectal or parenteral administration, and can be prepared into conventional solid preparations, liquid preparations or solutions for injection, etc., preferably tablets, capsules and injections.
Drawings
The accompanying drawings, which are included to provide a further understanding of the disclosure and are incorporated in and constitute a part of this specification, illustrate embodiments of the disclosure and together with the description serve to explain the disclosure without limiting the disclosure. In the drawings:
FIG. 1 is a diagram of a particular isolation scheme for compounds of the present invention.
Detailed Description
The technical solutions in the embodiments of the present invention will be clearly and completely described below with reference to the drawings in the embodiments of the present invention, and it is obvious that the described embodiments are only a part of the embodiments of the present invention, and not all of the embodiments. All other embodiments, which can be derived by a person skilled in the art from the embodiments given herein without making any creative effort, shall fall within the protection scope of the present invention.
The compounds of the present disclosure can be obtained by a variety of techniques, such as chemical synthesis methods; as another example, the extract may be obtained from other plants of the genus caesalpinia by any of a number of known techniques, such as: supercritical fluid extraction, solvent (ethanol, methanol, acetone, etc.) thermal reflux extraction, solvent (ethanol, methanol, acetone, etc.) percolation, etc.
The cichorium intybus used in the present disclosure was collected in west shuangbanna, Yunnan province in 2019 and 5 months, and was identified by the doctor of the kunming plant institute, gambir, china academy of sciences.
In the invention, 8 components are obtained by combining similar fractions according to TLC (thin layer chromatography) color development, the components 2A-2H are obtained by gradient elution through a petroleum ether/ethyl acetate system, the components 69A-69G are obtained by crude separation through the petroleum ether-ethyl acetate system, and all the components are named according to the polarity outflow sequence, namely, the components flow out firstly when the polarity is small, and then flow out when the polarity is large, and the similar components are combined according to the TLC color development condition to obtain the liquid.
The biological activity determination result and the physiological and biochemical experiment result in the embodiment of the invention prove that the aromatic casane diterpenoid compounds of the Tinospora bungeana have antibacterial activity. It will be appreciated by those skilled in the art that the results of the biological activity assay and the results of physiological and biochemical experiments establish the general utility of the aromatic casane diterpenes of Tinospora chrysantha as antibacterial agents.
For better understanding of the essence of the invention, the technical contents of the invention will be described in detail with examples, but the invention is not limited to these examples.
Example 1:
the embodiment discloses a cymbidium aromatized casane diterpenoid compound, which has any one or combination of structures shown in formulas 1 to 12:
Figure BDA0003639102360000051
physical and spectral data for compounds 1-12 are as follows:
compound 1: colorless needle crystals;
Figure BDA0003639102360000052
CD(MeOH)λmax(Δε):191(+44.8),190(-36.6)；UV(MeOH)λmax(logε)207(0.78)nm；IR(KBr)νmax 3494,2931,2384,2311,1724cm-1；HR-ESI-MS m/z 557.21478[M+Na]+[calcd for C31H34O8Na,557.21479]；1H and 13the C NMR data are shown in Table-1.
Compound 2: colorless needle crystals;
Figure BDA0003639102360000053
CD(MeOH)λmax(Δε):228(+57.9),251(-36.2)；UV(MeOH)λmax(logε)215(3.87)nm；IR(KBr)νmax 3495,2929,2384,2311,1706cm-1；HR-ESI-MS m/z471.17795[M+Na]+(calcd for C27H28O6Na,471.17781)；1H and 13the C NMR data are shown in Table-1.
Compound 3: a white amorphous powder;
Figure BDA0003639102360000054
CD(MeOH)λmax(Δε):191(+13.9),193(-7.9)；UV(MeOH)λmax(logε)207(0.65)nm；IR(KBr)νmax 3735,3554,3535,3515,2931,2385,2312,1719cm-1；HR-ESI-MS m/z 499.20929[M+Na]+(calcd for C29H32O6Na,499.20911)；1H and 13the C NMR data are shown in Table 2.
Compound 4: a white amorphous powder;
Figure BDA0003639102360000061
CD(MeOH)λmax(Δε):191(+12.5),221(-6.7)；UV(MeOH)λmax(logε)218(4.39)nm；IR(KBr)νmax 3554,2927,2862,2386,2312,1709cm-1；HR-ESI-MS m/z 467.21942[M+Na]+(calcd for C29H32O4Na,467.21928)；1H and 13the C NMR data are shown in Table 2.
Compound 5: a white amorphous powder;
Figure BDA0003639102360000062
CD(MeOH)λmax(Δε):278(+4.5),191(-19.2)；UV(MeOH)λmax(logε)214(1.23)nm；IR(KBr)νmax3737,3598,3556,3514,3338,2927,2859,2383,2311,1705cm-1.HR-ESI-MS m/z441.20383[M+Na]+(calcd for C27H30O4Na,441.20363)；1H and 13the C NMR data are shown in Table-3.
Compound 6: a brown amorphous powder;
Figure BDA0003639102360000063
CD(MeOH)λmax(Δε):190(+13.9),196(-3.6)；UV(MeOH)λmax(logε)190(4.27)nm；IR(KBr)νmax3435,2928,2362,1708cm-1；HR-ESI-MS m/z 531.19801[M+Na]+(calcd for C29H32O8Na,531.19828)；1H and 13the C NMR data are shown in Table-3.
Compound 7: a white amorphous powder;
Figure BDA0003639102360000064
CD(MeOH)λmax(Δε):191(+14.5),221(-36.7)；UV(MeOH)λmax(logε)219(2.10)nm；IR(KBr)νmax3392,2940,2877,2326,1731cm-1；HR-ESI-MS m/z 565.20380[M+Na]+(calcd for C29H30O10Na,565.20497)1H and 13the C NMR data are shown in Table 4.
Compound 8: a white amorphous powder;
Figure BDA0003639102360000065
CD(MeOH)λmax(Δε):238(+42.3),205(-18.5)；UV(MeOH)λmax(logε)202(2.53)nm；IR(KBr)νmax 3365,2928,2384,2311,1716cm-1；HR-ESI-MS m/z 657.26697[M+Na]+(calcd for C36H42O10Na,657.26702)；1H and 13c NMR data are shown inTABLE-4.
Compound 9: a white amorphous powder;
Figure BDA0003639102360000066
CD(MeOH)λmax(Δε):191(+2.11),236(-40.1)；UV(MeOH)λmax(logε)201(1.77)nm；IR(KBr)νmax 3397,2929,2868,2600,2547,2388,2309,1713cm-1；HR-ESI-MS m/z 431.25583[M+Na]+(calcd for C27H36O3Na,431.25567)；1H and 13the C NMR data are shown in Table 5.
Compound 10: a white amorphous powder;
Figure BDA0003639102360000071
CD(MeOH)λmax(Δε):190(+7.8),234(-19.5)；UV(MeOH)λmax(logε)227(1.71)nm；IR(KBr)νmax3514,2922,2856,2571,2384,2312,1742cm-1；HR-ESI-MS m/z 463.24551[M+Na]+(calcd for C27H36O5Na,463.24550)；1H and 13the C NMR data are shown in Table 5.
Compound 11: a white amorphous powder;
Figure BDA0003639102360000072
CD(MeOH)λmax(Δε):190(+20.0),193(-8.8)；UV(MeOH)λmax(logε)201(1.63)nm；IR(KBr)νmax 3454,2928,2860,2383,2311,1715cm-1；HR-ESI-MS m/z 509.25119[M+Na]+(calcd for C28H38O7Na,509.25097)；1H and 13the C NMR data are shown in Table-6.
Compound 12: colorless oil;
Figure BDA0003639102360000073
UV(MeOH)λmax(logε)230(0.71)nm；HR-ESI-MS m/z 429.22223[M+H]+(calcd for C25H33O6 429.22299)；1H and 13the C NMR data are shown in Table 6.
Figure BDA0003639102360000074
Figure BDA0003639102360000081
TABLE-1 preparation of Compounds 1 and 21H and13C-NMR data
Figure BDA0003639102360000082
Figure BDA0003639102360000091
TABLE-2 preparation of compounds 3 and 41H and13C-NMR data
Figure BDA0003639102360000092
TABLE-3 preparation of compounds 5 and 61H,13C-NMR data
Figure BDA0003639102360000101
TABLE-4 of Compounds 7 and 81H,13C-NMR data
Figure BDA0003639102360000111
TABLE-5 preparation of compounds 9 and 101H,13C-NMR data
Figure BDA0003639102360000112
Figure BDA0003639102360000121
TABLE-6 preparation of compounds 11 and 121H,13C-NMR data
Note: the nuclear magnetic data are measured by a Bruker DRX-500MHz nuclear magnetic resonance instrument.
Example 2
As shown in fig. 1, this embodiment discloses a method for extracting a spilanthol aromatized casane diterpene compound, which specifically includes:
drying stem of herba Pteridis Multifidae in the shade, pulverizing to 30 mesh, extracting with methanol under reflux for three times to obtain extractive solution, concentrating the extractive solution under reduced pressure, recovering methanol to obtain extract, concentrating under reduced pressure for five times, and mixing the extracts to obtain total extract.
Dispersing the total extract with appropriate amount of water, extracting with ethyl acetate and n-butanol for five times respectively (v/v 1:1), mixing the extractive solutions, and concentrating under reduced pressure to obtain ethyl acetate phase (400 g).
Dissolving an ethyl acetate extraction phase by using chloroform or acetone, carrying out column chromatography on silica gel for coarse separation, and carrying out gradient elution by using a chloroform/acetone system, wherein the volume ratio of solvents for gradient elution of the chloroform/acetone system is 1:0, 9:1, 8:2, 7:3, 1:1 and 0:1 in sequence; taking sulfuric acid ethanol solution as a color developing agent, carrying out color development according to TLC (thin layer chromatography) and combining similar fractions to obtain 8 components EA 1-EA 8, wherein the polarity sequence of EA 1-EA 8 is from small to large;
after the EA2 component is subjected to silica gel column chromatography, gradient elution is carried out by using a petroleum ether/ethyl acetate system, the volume ratio of elution of the petroleum ether/ethyl acetate system is 30:1, 25:1, 20:1 and 10:1 in sequence, 8 components 2A-2H are obtained according to the polarity, and the polarity sequence of the components 2A-2H is changed from small to large;
component 2C sequentially undergoes forward silica gel chromatographic column and RP-C18Separating with reverse chromatographic column and Sephadx-LH20 gel chromatographic column (methanol) to obtain compounds 4 and 5; wherein the forward silica gel chromatographic column is eluted by adopting a dichloromethane-ethyl acetate system, and the volume ratio of solvents is 50:1, 40:1, 30:1, 10:1 and 0: 1; RP-C18A reverse phase chromatographic column toEluting with methanol-water system at volume ratio of 3:2, 2:1, 3:1 and 1: 0;
separating out crystals from the component 2D, and repeatedly recrystallizing and purifying by methanol to obtain a compound 2;
the washing liquid of the component 2D sequentially passes through a forward chromatographic column, Sephadx-LH20 (acetone) and RP-C18Performing column chromatography separation to obtain compounds 3 and 9, wherein the volume ratio of n-hexane-ethyl acetate system solvent adopted by the forward chromatographic column is 20:1, 15:1, 10:1 and 5:1, and RP-C18The volume ratio of the acetone-water system solvent adopted by the column is 1:1, 3:2 and 2: 1;
component 2E is separately RP-C18Separating with column, forward silica gel chromatographic column and Sephadx-LH20 gel column (acetone) to obtain compounds 1 and 10, wherein RP-C18The column uses methanol-water as an elution solvent, the volume ratio of the solvent is 2:1, 3:1 and 1:0, and a forward silica gel chromatographic column adopts a petroleum ether-ethyl acetate system for elution, and the volume ratio is 15:1, 12:1, 8:1 and 5:1 in sequence;
performing forward silica gel column chromatography on the component EA3, performing crude separation by using a petroleum ether-ethyl acetate system to obtain 7 components 69A-69G according to the polarity, wherein the polarity sequence of the components 69A-69G is from small to large, and the solvent volume ratios of the petroleum ether-ethyl acetate system are 7:1, 5:1, 3:1 and 1:1 in sequence;
fraction 69D via RP-C18Semi-preparing liquid phase, and passing through RP-C18Performing column chromatography with Sephadx-LH20 (acetone) to obtain compound 8, wherein gradient methanol is used as solvent in the semi-preparation, the volume concentrations are 20%, 40%, 60%, 80% and 100%, and RP-C is prepared18The column takes methanol-water as an elution solvent, and the volume ratio is 2:1, 3:1, 4:1 and 1: 0;
the component 69F passes through a gel chromatographic column and RP-C in sequence18Preparing compounds 11 and 12 by column and high performance liquid chromatography, wherein the gel chromatography adopts chloroform-methanol elution system with volume ratio of 1:1, RP-C18The column uses methanol-water as an elution solvent, the volume ratio of the solvent is 3:1, 4:1, 5:1 and 1:0, and the high performance liquid chromatography preparation uses 98 percent methanol as a solvent and the flow rate is 2 ml/min;
component 69G first passes through RP-C18Column, eluting solvent is acetone-water system, volume ratioIs 4:1, 5:1, 6:1 and 1:0, and then is subjected to Sephadx-LH20 gel column (acetone) column chromatography to obtain a compound 6;
carrying out forward silica gel column chromatography on the component EA-4 for crude separation to obtain a component 1a-1 g; the component 1a is subjected to gradient elution by an MCI column, and the volume concentration of methanol subjected to gradient elution by the MCI column is 20%, 40%, 60%, 80% and 100% in sequence; and purifying the 60% methanol section by Sephadx-LH20 (acetone) column chromatography to obtain a compound 7.
Example 3
The antibacterial activity of the compound obtained by the preparation method disclosed in example 2 was tested:
1. test strains: kiwifruit canker (Pseudomonas syringae pv. Actinidae, Psa), Bacillus cereus (Bacillus cereus) and Staphylococcus aureus (Staphylococcus aureus). The strains are stored in a glycerol tube, taken out from a refrigerator at the temperature of-80 ℃, and are continuously activated twice in a fresh sterile LB culture medium in advance for use.
2. Activity test method: the bacteria were inoculated on LB medium for activation and cultured with shaking at 37 ℃ on a shaker at 170rpm/min to logarithmic phase. Diluting the bacterial liquid with LB culture medium to 2 x 10^ s6CFU/mL, add cultured bacterial liquid to sterile 96-well plate, 100. mu.l per well. The samples and controls were diluted to 200 μ M with LB medium. Test compounds are respectively dissolved in DMSO to prepare 200mM mother liquor, the mother liquor is diluted to 200 mu M by an LB culture medium, 100 mu L of diluted sample solution is added into each hole of a 96-hole plate, and after standing culture is carried out for 12-14 h in an incubator at 37 ℃, the growth condition of thalli is observed. Three groups of samples are set in parallel, an LB culture medium containing 0.5% DMSO is used as a negative control, common antibacterial drugs gentamicin and ciprofloxacin are used as positive controls, and a blank control group is set at the same time. And combining the primary screening result, further carrying out secondary screening on a series of concentration gradients (1.56-100 mu M) which are continuously diluted twice and are used for the test compound with the bacteriostatic activity. The bacteria grow in a 96-well plate LB culture medium, white precipitates can be generated due to large amount of the bacteria after a period of time, the wells with bacteria grow are turbid, the precipitates appear at the bottom, and the concentration of the compound corresponding to the wells without precipitates is the minimum inhibitory concentration. Finally, the wavelength is measured by a microplate reader under 600nmCorresponding absorbance value to verify the accuracy of MIC value (minimum inhibitory concentration). The activity data are shown in Table-7.
Figure BDA0003639102360000141
TABLE-7 bacteriostatic Activity data for Compounds 1-12
The data in Table 7 show that compounds 1-7 exhibit varying degrees of bacteriostatic action against both Actinidia canker, Bacillus cereus and Staphylococcus aureus. The compounds 2 and 3 have the most obvious bacteriostatic effect, and especially have the MIC value of 6.25 mu M for bacillus cereus and staphylococcus aureus, the inhibitory activity for kiwifruit canker germ is moderate, and the MIC values are respectively 25 and 12.5 mu M.
Meanwhile, compound 6 also showed significant antibacterial activity against bacillus cereus, with an MIC value of 6.25 μ M. According to the results, the compounds 2, 3 and 6 have obvious bacteriostatic effects and can be used as antibacterial agents for preventing and treating the bacterial infection.
Example 4:
this example discloses a tablet comprising any of the compounds disclosed in example 1, and further comprising lactose, starch, and magnesium stearate.
The preparation method comprises the following steps: the compound, lactose and starch were mixed, uniformly moistened with propylene glycol, the moistened mixture was sieved and dried, and sieved again, magnesium stearate was added, and the mixture was then tabletted to a weight of 250mg per tablet, with a compound content of 10 mg.
The obtained tablet can be used as antibacterial agent for treating bacterial infection. The daily dose may be 0.01-10 mg/kg body weight, preferably 0.1-5 mg/kg body weight, and may be administered once or more.
Example 5:
this example discloses an ampoule, which is obtained by mixing one or more of the compounds disclosed in example 1 with propylene glycol, the content of aromatic casane diterpenes being 2% to 8%;
the preparation method comprises the following steps: any of the compounds obtained in examples 1-2 was dissolved in 3 ml of propylene glycol, filtered, and the resulting solution was aseptically filled in an ampoule.
The obtained ampoule agent can be used as antibacterial agent for treating bacterial infection. The daily dose may be 0.01-10 mg/kg body weight, preferably 0.1-5 mg/kg body weight, and may be administered once or more.
Example 6:
this example discloses a capsule formulation comprising 10mg of the compound disclosed in example 1, 187mg of lactose, 3mg of magnesium stearate;
the preparation method comprises the following steps: mixing the compound with adjuvants, sieving, mixing, and encapsulating the obtained mixture into hard gelatin capsules with each capsule weighing 200mg and active ingredient content of 10 mg. The daily dosage may be 0.01-10 mg/kg body weight, preferably 0.1-5 mg/kg body weight, and may be administered once or more.
The capsule can be used as antibacterial agent for treating bacterial infection.
The preferred embodiments of the present disclosure are described in detail with reference to the accompanying drawings, however, the present disclosure is not limited to the specific details of the above embodiments, and various simple modifications may be made to the technical solution of the present disclosure within the technical idea of the present disclosure, and these simple modifications all belong to the protection scope of the present disclosure.
It should be noted that, in the foregoing embodiments, various features described in the above embodiments may be combined in any suitable manner, and in order to avoid unnecessary repetition, various combinations that are possible in the present disclosure are not described again.
In addition, any combination of various embodiments of the present disclosure may be made, and the same should be considered as the disclosure of the present disclosure, as long as it does not depart from the spirit of the present disclosure.

Claims (9)

1. The jatropha aromatic casane diterpenoid compound is characterized by having any one or more of structures shown in formulas 1-12:
Figure FDA0003639102350000011
2. a method for extracting aromatic casane diterpene compounds from herba Saussureae Involueratae comprises pulverizing stem of herba Saussureae Involueratae, and extracting in alcohol under hot reflux to obtain total extract; extracting the total extract with ethyl acetate, and repeatedly performing column chromatography on the obtained ethyl acetate extract phase to obtain the final product.
3. The method of claim 2, wherein said alcohols comprise methanol and ethanol.
4. The method for extracting the acerola aromatic casane diterpenoid compounds according to claim 2, which specifically comprises the following steps:
drying stem of herba Pteridis Multifidae in the shade, pulverizing to 30 mesh, extracting with methanol under reflux to obtain extractive solution, concentrating the extractive solution under reduced pressure, recovering methanol to obtain crude extract, and extracting with ethyl acetate to obtain ethyl acetate extract phase;
dissolving an ethyl acetate extraction phase by using chloroform or acetone, carrying out column chromatography on silica gel for coarse separation, and carrying out gradient elution by using a chloroform/acetone system, wherein the volume ratio of the gradient elution of the chloroform/acetone system is 1:0, 9:1, 8:2, 7:3, 1:1 and 0:1 in sequence; taking sulfuric acid ethanol solution as a color developing agent, and carrying out color development according to TLC (thin layer chromatography) and combining similar fractions to obtain 8 components EA 1-EA 8;
after the EA2 component is subjected to silica gel column chromatography, gradient elution is carried out by using a petroleum ether/ethyl acetate system, the elution concentrations of the petroleum ether/ethyl acetate system are 30:1, 25:1, 20:1 and 10:1 in sequence, and 8 components 2A-2H are obtained according to the polarity;
component 2C sequentially undergoes forward silica gel chromatographic column and RP-C18Separating with reverse chromatographic column and Sephadx-LH20 gel chromatographic column to obtain compounds 4 and 5;
separating out crystals from the component 2D, and repeatedly recrystallizing and purifying the crystals by methanol to obtain a compound 2;
the washing solution of the component 2D sequentially passes through a forward chromatographic column, a Sephadx-LH20 gel chromatographic column and an RP-C18Separating with reverse chromatographic column to obtain compounds 3 and 9;
component 2E is separately RP-C18Separating with reverse chromatographic column, forward silica gel chromatographic column and Sephadx-LH20 gel chromatographic column to obtain compounds 1 and 10;
performing forward silica gel column chromatography on the component EA3, performing crude separation by using a petroleum ether-ethyl acetate system to obtain 9 components 69A-69G according to polarity, wherein the volume ratios of the petroleum ether-ethyl acetate system are 7:1, 5:1, 3:1 and 1:1 in sequence;
fraction 69D via RP-C18After semi-preparation of the liquid phase, the liquid phase is then subjected to RP-C18Subjecting to Sephadx-LH20 column chromatography to obtain compound 8, and subjecting to RP-C chromatography18The semi-preparative liquid phase uses methanol with volume concentration of 20%, 40%, 60%, 80% and 100% in sequence;
the component 69F sequentially passes through a chloroform-methanol gel column and an RP-C with the volume ratio of 1:118Subjecting the column to high performance liquid chromatography to obtain compounds 11 and 12;
the component 69G passes through RP-C in sequence18Performing chromatography on the chromatographic column and a Sephadx-LH20 gel column to obtain a compound 6;
carrying out forward silica gel column chromatography on the component EA-4 for crude separation, and obtaining a component 1a-1g according to the polarity; the component 1a is subjected to gradient elution by adopting an MCI column, and the volume concentration of methanol subjected to gradient elution by the MCI column is 20%, 40%, 60%, 80% and 100% in sequence; and purifying the 60% methanol section by Sephadx-LH20 column chromatography to obtain a compound 7.
5. Use of the aromatic casane diterpene compound of claim 1 or a pharmaceutically acceptable salt thereof for the preparation of an antibacterial agent.
6. The use as claimed in claim 5, wherein the antimicrobial preparation is used for the control of Actinidia canker, Bacillus cereus and Staphylococcus aureus.
7. The use of claim 5, wherein the antibacterial agent preparation is prepared into a pharmaceutically acceptable preparation by adding pharmaceutically acceptable auxiliary materials according to a conventional process, and the pharmaceutically acceptable preparation is a solid preparation or a liquid preparation.
8. The use according to claim 7, wherein the solid formulation comprises granules, capsules, tablets, pills; the liquid formulation includes an injection formulation.
9. The use according to claim 7, wherein the antibacterial agent preparation contains 2 to 8% by mass of the aromatic casane diterpene compound or the pharmaceutically acceptable salt thereof.
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Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN114456190A (en) * 2020-11-07 2022-05-10 中国科学院昆明植物研究所 Kasan alkane diterpene phanginin A derivative, pharmaceutical composition and application thereof

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* Cited by examiner, † Cited by third party
Title
张佳瑶: "金凤花和头花杜鹃中次生代谢产物及生物活性研究", 中国优秀硕士学位论文农业科技辑西北农林科技大学硕士学位论文, pages 048 - 352 *
陈秀梅: "金凤花茎中的二萜化合物及其生物活性研究", 中国优秀硕士学位论文医药卫生科技辑西北农林科技大学硕士学位论文, pages 057 - 123 *

Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN114456190A (en) * 2020-11-07 2022-05-10 中国科学院昆明植物研究所 Kasan alkane diterpene phanginin A derivative, pharmaceutical composition and application thereof

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