CN107312059A - Steroid saponin derivative and its preparation method and application - Google Patents
Steroid saponin derivative and its preparation method and application Download PDFInfo
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Abstract
The invention discloses steroid saponin derivative and its preparation method and application.The present invention is that biological transformed bacteria carries out bioconversion with C.olivaceum bacterial strain, is finally recovered the steroid saponin derivative shown in the formula V of acquisition formula I using Pseudoprodioscin as substrate.Pharmacological experiment is proved; steroid saponin derivative shown in the formula V of formula I has excellent antitumor activity and thrombus dissolving activity; there is significant protection activity to cardiac muscle cell; toxicity to normal cell is low, has important application prospect in terms of antitumor, treatment disease of cardiovascular system, raising immunity, reduction blood glucose, antifertility, antibacterial.
Description
Technical field
The present invention relates to steroid saponin derivative, more particularly to using Pseudoprodioscin as substrate, using bioconversion
Steroid saponin derivative is obtained prepared by method, the invention further relates to the steroid saponin derivative in treatment cardiovascular system
Application in terms of disease, antitumor, raising immunity or reduction blood glucose, belongs to steroid saponin derivative field.
Background technology
Steroid saponin is the important chemical composition of a class in natural products, is broadly divided into furostanol saponin and spirostanol saponin
(FENG B,KANG L-P,MA B-P,et al.The substrate specificity of a glucoamylase
with steroidal saponin-rhamnosidase activity from Curvularia lunata[J]
.Tetrahedron,2007,63(29):6796-812.), it is main to include treatment angiocarpy mostly with certain pharmacological activity
Systemic disease, antitumor, raising immunity, reduction blood glucose, antifertility, antibacterial action etc..The research of steroid saponin is naturally being produced
Occupied an important position always in thing chemistry.It is broadly divided into two major classes：Furostan type (furostanoside) and spirostan type
(spirostanoside).Structural modification is carried out to such material by bioconversion, the novel material of structure can be obtained.
Bioconversion has the advantages that a lot compared to traditional chemical method, and the condition of bioconversion is gentleer, room temperature, often
Pressure and the progress in the pH for being bordering on neutrality, course of reaction also more environmentally-friendly green, and the regioselectivity with height and bottom
Thing selectivity (HEGAZY M-E F, MOHAMED T A, ELSHAMY A I, et al.Microbial
biotransformation as a tool for drug development based on natural products
from mevalonic acid pathway:A review[J].Journal of Advanced Research,2015,6
(1):17-33).The course of reaction of steroid saponin bioconversion mainly includes desugar and reacted, sugaring reaction, plus hydroxyl, demethoxy
Base formation double bond, double bond oxidation scission are carbonyl etc..Structural modification is carried out to steroid saponin with the method for bioconversion, can be obtained
The compound for being relatively difficult to obtain to some chemical methodes, the more gentle environmental protection of reaction condition, and the regional choice with height
Property, therefore bioconversion increasingly payes attention to and obtained many development.
Pseudoprodioscin is widely present in yam, be treat angiocardiopathy Diaoxinxue Kang it is effective into
One of part, the activity with cell toxicant, antimycotic, antiviral and cardiovascular aspect.Mei Dong et al. (DONG M, FENG
X,WANG B X,et al.Microbial metabolism of pseudoprotodioscin[J].Planta Med,
2004,70(7):Bioconversion, four changes of separation identification 637-41) are carried out to Pseudoprodioscin (compound 1) with aspergillus fumigatus
Compound, wherein compound 2 and 3 are noval chemical compound (Fig. 1).
It is also furostanol saponin that protodioscin, which is, and (all-wave, Ma Baiping aspergillus oryzaes are to steroidal soap in rhizoma dioscoreae nipponicae for all-wave et al.
The bioconversion [D] of glycosides；Shenyang Pharmaceutical University, 2006.) bioconversion is carried out to it with aspergillus oryzae, and it is biological to have studied it
Path for transformation (Fig. 2).
The preparation method that bioconversion is carried out by substrate of Pseudoprodioscin has that biotransformation efficiency is low, product mostly
The low defect of yield, in addition, the different degrees of presence pharmacology of the steroid saponin derivative prepared using the bioconversion is lived
The problems such as property is relatively low, toxicity is larger, has much room for improvement.
The content of the invention
An object of the present invention is to provide a class with higher pharmacological activity and the more preferable steroid saponin derivative of security
Thing；
The second object of the present invention is to provide a kind of preparation method of the steroid saponin derivative of high transformation efficiency；
The third object of the present invention be by described steroid saponin derivative be applied to prepare treatment disease of cardiovascular system,
In terms of antitumor, raising immunity, reduction blood glucose, antifertility, antibacterial.
The above-mentioned purpose of the present invention is achieved through the following technical solutions：
Steroid saponin derivative, its structural formula is any compound shown in I-formula of following formula V：
R1=-β-D-Glu
The salt of the steroid saponin derivative is also certainly included within protection scope of the present invention.
Invention further provides a kind of Biotransfer process for preparing of described steroid saponin derivative, including it is following
Step：
(1) it is biology with C.olivaceum (Chaetomium olivaceum) bacterial strain using Pseudoprodioscin as substrate
Transformed bacteria carries out bioconversion reaction；Reaction product is extracted, extract concentration obtains crude extract；
(2) after silicagel column on crude extract, eluted with eluent；Eluent be chloroform-methanol-water, polarity by
12.5:1:0.08 to 1:1:0.1；After point plate merges, 1-12 parts of samples are finally given；
Upper silicagel column elution liquid is CHCl to sample 9 again3–MeOH–H2O, ratio is 7:1:0.1,5:1:0.1,4:1:0.1,
3:1:0.1,2:1:0.1 and 1:1:0.1, obtain 7 parts of sample A9-G9；D9And E9It is principal component point, by D9Upper preparation liquid phase, flowing
It is mutually MeOH-H2O, ratio is 71:39, obtain type I compound and the compound of formula II；By E9Upper preparation liquid phase, mobile phase is
MeOH-H2O, ratio is 67:33, obtain the compound of formula III；
Silicagel column on sample 8, eluent is CHCl3-MeOH-H2O, ratio is 10:1:0.07,8:1:0.07,6:1:0.1,
4:1:0.1,3:1:0.1；18 parts of components have been connect altogether, and the 8th and the 9th part of component is upper after merging to prepare liquid phase, and mobile phase is MeOH-
H2O(62:38,2mL/min) obtain the compound of formula IV；
By gel column on sample 7, mobile phase is MeOH, finally obtains 6 parts of sample A7-F7；By sample B7Upper preparation liquid phase,
Mobile phase is MeOH-H2O, ratio is 80:20, obtain the compound of formula V.
As preferred, the extraction described in step (1) is that the n-butanol that equivalent is added in every bottle of culture medium carries out ultrasound
Extraction；Silicagel column is the silica gel mixed sample with 100-200 mesh on crude extract in step (2), and post is filled with 200-300 mesh silica gel.
The present invention proves that the compound shown in I-formula of formula V is to HePG2 (liver cancer cells), MCF-7 by pharmacological experiment
The JEG-3 such as (human breast cancer cell), NCI-H460 (people's lung ED-SCLC), SF-268 (human glioma cell) has
Strong active anticancer, but the toxicity to normal cell is far below toxicity of the Pseudoprodioscin to normal cell.
Tumour of the present invention includes：In primary solid tumor, drug-resistant tumor, metastatic tumo(u)r, malignant tissue or cell
Any one or more；It is preferred that, the cancer includes：Nasopharyngeal carcinoma, oral cavity epidermoid carcinoma, carcinoma sarcomatodes, oophoroma, pancreas
Cancer, prostate cancer, colon cancer (including colorectal cancer), the carcinoma of the rectum, metastatic colorectal cancer, non-squamous non-small cell lung
Cancer, breast cancer, metastatic breast cancer, metastatic HER2 negative breast cancers, the cancer of the brain, be grown up cerebral glioma, children's brain colloid
Cytoma, child-resistance cerebral glioma, glioma, astrocytoma, medulloblastoma, children are thin into nerve channel
Born of the same parents' knurl, glioma, oligodendroglioma, meningioma, kidney (such as advanced renal cell carcinoma), carcinoma of urinary bladder, cervical carcinoma, oesophagus
Cancer, stomach cancer, incidence cancer, liver cancer, lung cancer (ED-SCLC and non-small cell lung cancer), squamous non-small cell lung cancer, melanoma,
Myeloma, sarcoma (including osteosarcoma), cutaneum carcinoma (including squamous cell carcinoma), stomach cancer, carcinoma of testis, thyroid cancer, uterine cancer,
In rind gall, cholangiocarcinoma, leiomyosarcoma, embryonal-cell lipoma, fibrosarcoma, neuroendocrine carcinoma, salivary-gland carcinoma any one or
It is a variety of.
Invention further provides a kind of antitumor medicine composition, the steroid saponin containing the upper effective dose for the treatment of spreads out
Biological and pharmaceutically acceptable carrier.The carrier includes：Solvent, surfactant, solubilizer, cosolvent, lubricant, fill out
Fill any one or more in agent, isotonic regulator, stabilizer or supporting agent.
The formulation of anti-tumor agent of the present invention includes：Injection, tablet, capsule, granule, supensoid agent, emulsion,
Solution, glue, freeze drying powder injection, mucilage, aerosol, micro-capsule, microballoon, nanometer formulation, sustained release preparation or controlled release preparation
In any one or more；Preferably nanometer formulation.Wherein, the nanometer formulation includes：Liposome, nanocapsule, nanosphere,
Any one or more in lipid nano particle or micella, preferably any one in liposome or micella or two kinds.This hair
The particle diameter of the bright nanometer formulation is 10 nanometers -300 nanometers.Preparing the conventional material of the nanometer formulation includes：Lipid, phosphorus
Fat, cholesterol, high molecular polymer, bi-block copolymer, triblock polymer, albumen, polypeptide, nucleic acid, chitosan, surface are lived
Property agent, solubilizer, cosolvent, filler, isotonic regulator, stabilizer, supporting agent etc..
Anti-tumor compositions or anticancer preparation of the present invention can be administered by clinically-acceptable therapeutic modality, including injection,
Orally, suck, be implanted into or irrigate etc..
The present invention is using the influence experimental evaluation the compounds of this invention to hypoxia-reoxygenation H9C2 myocyte survival rates to the heart
The protection activity of flesh.Pass through the survey of LDH measure, cTn-I protein delivery amounts in the measure (MTT) of cell survival rate, supernatant
It is fixed, determine that the compound shown in I-formula of formula V there is obvious protection to make the rat myocardial cell of anoxia/reoxygenation
With.
Zoopery is proved, compared to Pseudoprodioscin, and the compound shown in I-formula of formula V has excellent
Thrombus dissolving activity, can be applied to treatment disease of cardiovascular system.
It is upper effective containing treatment invention further provides a kind of pharmaceutical composition prevented or treat angiocardiopathy
The steroid saponin derivative and pharmaceutically acceptable carrier of amount；The carrier includes：Solvent, surfactant, solubilizer, help
Any one or more in solvent, lubricant, filler, isotonic regulator, stabilizer or supporting agent.
It is of the present invention prevention or treatment angiocardiopathy pharmaceutical composition can according to this area conventional formulation side
Method is prepared into clinically suitable preparation formulation, including：Injection, tablet, capsule, granule, supensoid agent, emulsion, solution
In agent, glue, freeze drying powder injection, mucilage, aerosol, micro-capsule, microballoon, nanometer formulation, sustained release preparation or controlled release preparation
Any one or more；Preferably nanometer formulation.Wherein, the nanometer formulation includes：Liposome, nanocapsule, nanosphere, lipid
Any one or more in nanoparticle or micella, preferably any one in liposome or micella or two kinds.Institute of the present invention
The particle diameter for stating nanometer formulation is 10 nanometers -300 nanometers.Preparing the conventional material of the nanometer formulation includes：Lipid, phosphatide, courage
Sterol, high molecular polymer, bi-block copolymer, triblock polymer, albumen, polypeptide, nucleic acid, chitosan, surfactant,
Solubilizer, cosolvent, filler, isotonic regulator, stabilizer, supporting agent etc..
The pharmaceutical preparation of present invention treatment angiocardiopathy can be administered by clinically-acceptable therapeutic modality, including note
Penetrate, orally, suction, implantation or perfusion etc..
In addition, the compound shown in I-formula of formula V also has the pharmacology work improved in terms of immunity, antibacterial, antifertility
Property.
Brief description of the drawings
Fig. 1 is the biotransformation pathway of Pseudoprodioscin in the prior art；
Fig. 2 is the biotransformation pathway of protodioscin；
Fig. 3 is the structural formula of carrier saponin derivative of the present invention；
The lamellae figure of Fig. 4 runic things, is from left to right followed successively by sample 1-12；
The lamellae figure of Fig. 5 samples 9；
The lamellae figure of Fig. 6 samples 8；
The lamellae figure of Fig. 7 samples 7；
The experimental result to scheming cytoprotection of Fig. 8 steroid saponin derivatives；
Fig. 9 H9C2 Myocytes Anoxia reoxygenation models set up result；
Figure 10 lactic dehydrogenases (LDH:Lactic Dehydrogenase) burst size measurement result；
The measurement result of Figure 11 cardiac troponins (cTnI) burst size；
Influence measurement results of the Figure 12 to H9C2 cardiac muscle cell's mitochondrial membrane potential of anoxia/reoxygenation.
Embodiment
The invention will now be further described with reference to specific embodiments, advantages of the present invention and feature will be with description and
It is apparent.It should be understood that the embodiment is only exemplary, any limitation is not constituted to the scope of the present invention.This area
Technical staff should be understood that without departing from the spirit and scope of the invention can to the details of technical solution of the present invention and
Form is modified or replaced, but these modifications or substitutions each fall within protection scope of the present invention.
The preparation and identification of the steroid saponin derivative of embodiment 1
1. the expansion and extraction of bacterial strain
The bacterium (C.olivaceum Chaetomium olivaceum CGMCC 3.3604) in test tube is first seeded in one bottle
Cultivate 2 days, the bacterium in this triangular flask has been transferred into 17 1L triangular flask with glue head dropper in 250mL triangular flask then
In.Culture medium used is PDA culture medium in experiment.Shaking table condition is 28 DEG C, 160r/min；
Continue to add substrate Pseudoprodioscin after cultivating 2 days.The amount of addition is that 2.5g (uses 2.5g Pseudoprodioscins
17mL methanol dissolving, then adding 1mL in each triangular flask).
Addition substrate terminating reaction after 4 days：The n-butanol ultrasonic extraction of equivalent (400mL) is added in every bottle of culture medium
30min, in triplicate.Extract concentrated by rotary evaporation.Crude extract 19.32g is obtained.2. the separation of crude extract
Silica gel mixed sample of the crude extract with 35g100-200 mesh, 500g200-300 mesh silica gel dress post.Eluent be chloroform-
Methanol-water, polarity is by 12.5:1:0.08 to 1:1:0.1 (table 1).Sample is connect with 100mL conical flask, the conjunction of sample spot plate is connect
And after 12 parts.Lamellae figure is shown in Fig. 4.Lamellae is from left to right followed successively by sample 1-12, and ((laboratory is existing for Pseudoprodioscin
Standard items), Gracillin (the existing standard items in laboratory), Dioscin (the existing standard items in laboratory), crude extract, blank
Control).
Table 1
Sample 9 is 1.952g, with the silica gel 5g pressings of 100-200 mesh, and the silica gel 60g of 300-400 mesh fills post, and eluent is
CHCl3–MeOH–H2O(7:1:0.1,5:1:0.1,4:1:0.1,3:1:0.1,2:1:0.1 and 1:1:0.1) (table 2), then again
To 7 parts of samples, seven parts of sample (A are obtained after merging9-G9), lamellae figure is Fig. 5；Lamellae is followed successively by A from left to right9-G9It is (pseudo- former
Dioscin, Gracillin, Dioscin).
Table 2
| Eluent | Chloroform | Methanol | Water |
| Ratio (mL) | 280 | 40 | 4 |
| 300 | 60 | 6 | |
| 240 | 60 | 6 | |
| 300 | 100 | 10 | |
| 260 | 130 | 13 | |
| 150 | 150 | 15 |
D9And E9It is principal component point, because the color is too dark, has first gone up gel depigmentation.Then D9Upper preparation liquid phase, mobile phase
For MeOH-H2O(71:39,2mL/min) obtaining sterling compound 1 (type I compound) (20.59mg) and compound 2, (formula II is changed
Compound) (4mg).E9Also with liquid phase is prepared, mobile phase is MeOH-H2O(67:33,2mL/min) obtain (chemical combination of formula III of compound 3
Thing) (15.8mg).
Sample 8 is 468mg, and sample, 300-400 mesh silica gel 20g dress posts are mixed with the silica gel 1.5g of 100-200 mesh.Upper silicagel column
(300-400 mesh, 20g) eluent is CHCl3-MeOH-H2O ratios are 10:1:0.07,8:1:0.07,6:1:0.1,4:1:0.1,
3:1:0.1 (table 3), has connect 18 parts of components altogether, and lamellae figure is Fig. 6.
Table 3
| Eluent | Chloroform | Methanol | Water |
| Ratio (mL) | 100 | 10 | 0.7 |
| 160 | 20 | 1.4 | |
| 180 | 30 | 3 | |
| 200 | 50 | 5 | |
| 180 | 60 | 6 |
Upper after 8th and the 9th part of merging to prepare liquid phase, mobile phase is MeOH-H2O(62:38,2mL/min) obtain sterling
Compound 4 (compound of formula IV) (1.59mg).
Sample 7 (496mg) first goes up gel, and mobile phase is MeOH..Finally obtain 6 parts of sample (A7-F7)；Lamellae figure is shown in figure
7 (are followed successively by A from left to right7-F7)。
Sample B7Upper preparation liquid phase, mobile phase is MeOH-H2O(80:20,2mL/min) obtain (chemical combination of formula V of compound 5
Thing) (12.09mg).
Compound 1：White powder.UV absorbs for end.[α]D 20=-20.86 (c0.110, MeOH) IR (Neat) vmax
(cm-1):3372,2935,1364,1033。HR-ESI-MS m/z:937.4758[M+Na]+(calcd for C46H74O18Na,
937.4773)。
Compound 2：White powder.UV absorbs for end.[α]D 20=-20.41 (c0.074, MeOH) IR (Neat) vmax
(cm-1):3362,2932,1448,1379,1039。HR-ESI-MS m/z:937.4782[M+Na]+(calcd for
C46H74O18Na,937.4773)。
Compound 3：White powder.UV absorbs for end.[α]D 20=-18.82 (c0.090, MeOH) IR (Neat) vmax
(cm-1):3369,2935,1363,1033。HR-ESI-MS m/z:955.4919[M+Na]+(calcd for C46H76O19Na,
955.4879)。
Compound 4：White powder.UV absorbs for end.[α]D 20=-34.71 (c 0.081, MeOH) IR (Neat) vmax
(cm-1):3384,2920,1762,1645,1033。HR-ESI-MS m/z:691.3331[M+Na]+(calcd for
C34H52O13Na,691.3306)。
Compound 5：White powder.UV absorbs for end.[α]D 20=-20.86 (c 0.110, MeOH) IR (Neat) vmax
(cm-1):3370,2935,1652,1456,1380,1033。HR-ESI-MS m/z:761.4097[M+Na]+(calcd for
C39H62O13Na,761.4088)。
The anti tumor activity in vitro experiment of the steroid saponin derivative of experimental example 1
Anti tumor activity in vitro experiment shows that compound 1-5 prepared by the embodiment of the present invention 1 antitumor activity is substantially excellent
In Pseudoprodioscin or the antitumor activity of structurally similar compounds.
The vitro cytotoxicity experiment of the steroid saponin derivative of experimental example 2
Vitro cytotoxicity experimental data proves that compound 1-5 prepared by the embodiment of the present invention 1 toxicity is significantly lower than puppet
The toxicity of protodioscin or structurally similar compounds.
The experiment acted on myocardial cell protection of the steroid saponin derivative of experimental example 3
Take the logarithm growth period cell with 1 × 104The density in individual/hole is inoculated in 96 orifice plates, in 5%CO2, 37 DEG C of conditions
Nutrient solution is removed after culture 30h, the compound 1-5 for the various concentrations (μ g/mL) being made into DMEM is added, trained under similarity condition
Support 24h.After processing terminates, nutrient solution is removed, 500 μM of H are added2O2Nutrient solution is carefully removed after placing 1h, 10 μ are added in every hole
L CCK-8, then place after 2h, survey absorbance (OD values) at 450nm in microplate ELIASA.By the OD values of each instrument connection
Background OD values (serum free medium adds CCK-8, acellular) are subtracted, average value ± SD is calculated according to each hole OD values.
Cell survival rate %=(dosing cell OD values-background OD values)/(control cell OD values-background OD values) × 100%.
Cell survivaling number is as shown in Figure 8.As seen from Figure 8, the compound 1-5 that prepared by the embodiment of the present invention 1 by hydrogen peroxide to causing damage
The cardiac muscle cell of wound, there is significant protective effect.
Protective effect of the steroid saponin derivative of experimental example 4 to H9C2 Myocytes Anoxia reoxygenations is tested
First, experiment material
1st, test compound：Compound 1-5 (S1) prepared by the embodiment of the present invention 1；
2nd, positive control sample：Diaoxinxue Kang.
2nd, experimental method and result
(1) foundation of H9C2 Myocytes Anoxias reoxygenation model
To H9c2 Myocytes Anoxia 12h, difference reoxygenation 0,8,12,16 and 24h, when detecting that reoxygenation is different using mtt assay
Between H9c2 myocyte survival rates.After reoxygenation 12h, myocyte survival rate is 51.3%.Show anoxic 8h, reoxygenation 12h damage
Hinder condition optimal.Now cell survival rate is more than 50%, and degree of injury meets the requirement (figure that cell model is set up in experiment
9)。
(2) lactic dehydrogenase (LDH:Lactic Dehydrogenase) burst size measure
H9c2 cells are divided into five groups and are administered, control group (Con), model group (H/R), S1 low dose groups (H/R+L), S1
High dose group (H/R+H), positive drug group (Diaoxinxue Kang, DA).Each group uses the DMEM high glucose mediums without serum to dilute
Certain density medicine is pre-processed (incubation) 24h to cell.Then tested according to the method for hypoxia-reoxygenation, experiment knot
Beam collects whole cell conditioned mediums, and determining the assay method in kit according to LDH is detected.
Measurement result shows that sample S1 low dosages and high dose can significantly reduce the LDH burst sizes (Figure 10) of cell conditioned medium.
(3) measure of cardiac troponin (cTnI) burst size
H9c2 cells are divided into five groups and are administered, control group (Con), model group (H/R), S1 low dose groups (H/R+L), S1
High dose group (H/R+H), positive drug group (Diaoxinxue Kang, DA).It is certain using the DMEM high glucose mediums dilution without serum
The medicine of concentration is pre-processed (incubation) 24h to cell.Then tested according to the method for hypoxia-reoxygenation, experiment terminates to receive
The whole cell conditioned mediums of collection, determine the assay method in kit according to cTnI and are detected.
Testing result shows that sample S1 low dosages and high dose can significantly reduce cell conditioned medium cTnI burst size (figure
11)。
(4) influence to H9C2 cardiac muscle cell's mitochondrial membrane potential of anoxia/reoxygenation is detected
H9c2 cells are divided into five groups and are administered, control group (Con), model group (H/R), S1 low dose groups (H/R+L), S1
High dose group (H/R+H), positive drug group (Diaoxinxue Kang, DA).Each group uses the DMEM high glucose mediums without serum to dilute
Certain density medicine is pre-processed (incubation) 24h to cell.Then tested according to the method for hypoxia-reoxygenation, experiment knot
Beam, which is removed, adds JC-1 fluorescence probes liquid processing cell after cell conditioned medium, cleaning cell, fluorescence microscopy Microscopic observation simultaneously records reality
Test.Test result indicates that sample S1 low dosages and high dose can substantially reduce the line of the H9c2 cardiac muscle cell of anoxia/reoxygenation
Mitochondrial membrane potential and protect cardiac muscle cell (Figure 12).
Claims (10)
1. steroid saponin derivative, it is characterised in that its structural formula is shown in I-formula of following formula V：
R1=-β-D-Glu
2. the salt of steroid saponin derivative described in claim 1.
3. the Biotransfer process for preparing of the steroid saponin derivative described in claim 1, it is characterised in that including following step
Suddenly：
(1) it is bioconversion with C.olivaceum (Chaetomium olivaceum) bacterial strain using Pseudoprodioscin as substrate
Bacterium carries out bioconversion reaction；Reaction product is extracted, extract concentration obtains crude extract；
(2) after silicagel column on crude extract, eluted with eluent；Eluent is chloroform-methanol-water, and polarity is by 12.5:1:
0.08 to 1:1:0.1；After point plate merges, 1-12 parts of samples are finally given；
Upper silicagel column elution liquid is CHCl to sample 9 again3–MeOH–H2O, ratio is 7:1:0.1,5:1:0.1,4:1:0.1,3:1:
0.1,2:1:0.1 and 1:1:0.1, obtain 7 parts of sample A9-G9；D9And E9It is principal component point, by D9Upper preparation liquid phase, mobile phase is
MeOH-H2O, ratio is 71:39, obtain type I compound and the compound of formula II；By E9Upper preparation liquid phase, mobile phase is MeOH-
H2O, ratio is 67:33, obtain the compound of formula III；
Silicagel column on sample 8, eluent is CHCl3-MeOH-H2O, ratio is 10:1:0.07,8:1:0.07,6:1:0.1,4:1:
0.1,3:1:0.1；18 parts of components have been connect altogether, and the 8th and the 9th part of component is upper after merging to prepare liquid phase, and mobile phase is MeOH-H2O
(62:38,2mL/min) obtain the compound of formula IV；
By gel column on sample 7, mobile phase is MeOH, finally obtains 6 parts of sample A7-F7；By sample B7Upper preparation liquid phase, flowing
It is mutually MeOH-H2O, ratio is 80:20, obtain the compound of formula V.
4. according to the Biotransfer process for preparing described in claim 3, it is characterised in that the extraction described in step (1) is every
The n-butanol that equivalent is added in bottle culture medium carries out ultrasonic extraction；Silicagel column is to use 100-200 purposes on crude extract in step (2)
Silica gel mixed sample, post is filled with 200-300 mesh silica gel.
5. purposes of the steroid saponin derivative in antineoplastic is prepared described in claim 1.
6. steroid saponin derivative described in claim 1 is in preparing treatment disease of cardiovascular system or preparing the myocardium medicine of protection
Purposes.
7. steroid saponin derivative described in claim 1 is preparing the purposes in improving immunity medicine.
8. purposes of the steroid saponin derivative described in claim 1 in antibacterials are prepared.
9. purposes of the steroid saponin derivative in antifertility drug is prepared described in claim 1.
10. a kind of antitumor medicine composition, it is characterised in that：Steroid described in claim 1 containing the upper effective dose for the treatment of
Body saponin derivative and pharmaceutically acceptable carrier.
Applications Claiming Priority (2)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN2016102700182 | 2016-04-27 | ||
| CN201610270018 | 2016-04-27 |
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| CN112194704A (en) * | 2020-10-21 | 2021-01-08 | 中国人民解放军空军军医大学 | Steroid saponin compound and preparation method and application thereof |
| CN113759036A (en) * | 2021-08-10 | 2021-12-07 | 北京康仁堂药业有限公司 | Method for measuring content of protodioscin in rhizoma dioscoreae septemlobae |
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| CN1727358A (en) * | 2004-07-29 | 2006-02-01 | 成都地奥九泓制药厂 | Steroid saponin of alpha, beta double bond structure outside furan nucleus, preparation method and application |
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Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN112194704A (en) * | 2020-10-21 | 2021-01-08 | 中国人民解放军空军军医大学 | Steroid saponin compound and preparation method and application thereof |
| CN112194704B (en) * | 2020-10-21 | 2021-06-01 | 中国人民解放军空军军医大学 | Steroid saponin compound and preparation method and application thereof |
| CN113759036A (en) * | 2021-08-10 | 2021-12-07 | 北京康仁堂药业有限公司 | Method for measuring content of protodioscin in rhizoma dioscoreae septemlobae |
| CN113759036B (en) * | 2021-08-10 | 2023-09-08 | 北京康仁堂药业有限公司 | Method for measuring content of protodioscin in rhizoma Dioscoreae Septemlobae |
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