CN106822167A - A kind of new application of compound and preparation method thereof - Google Patents
A kind of new application of compound and preparation method thereof Download PDFInfo
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Abstract
The invention belongs to medicinal chemistry art, specifically related to a kind of new application of compound and preparation method thereof, that is (3 β, 5 α, 22 α, 25R) new application of spirosolane 3 O β D glucopyranosyl (1 → 2) O [β D xylopyranoses base (1 → 3)] O β D glucopyranosyl (1 → 4) β D galactopyranosides and preparation method thereof, described compound can suppress lung carcinoma cell, by lung cancer cell growth cycle arrest in early stage, promote Increase Apoptosis of Lung Cancer Cells, therefore, the compound can be used for preparing cancer therapy drug, especially prepare anti-lung-cancer medicament.
Description
Technical field
The invention belongs to medicinal chemistry art, and in particular to a kind of new application of compound and preparation method thereof, i.e. (3 β, 5
α, 22 α, 25R)-spirosolane -3-O- β-D- glucopyranosyls-(1 → 2)-O- [β-D- xylopyranoses base-(1 → 3)] -
New application of O- β-D- glucopyranosyls-(1 → 4)-β-D- galactopyranosides and preparation method thereof.
Background technology
(3 β, 5 α, 22 α, 25R)-spirosolane -3-O- β-D- glucopyranosyls-(1 → 2)-O- [β-D- wood pyrans
Glycosyl-(1 → 3)]-O- β-D- glucopyranosyls-(1 → 4)-β-D- galactopyranosides, CAS:90366-11-3, white powder
End, is dissolved in methyl alcohol, is insoluble in ethanol, water, and structural formula is as follows:
(3 β, 5 α, 22 α, 25R)-spirosolane -3-O- β-D- glucopyranosyls-(1 → 2)-O- [β-D- wood pyrans
Glycosyl-(1 → 3)]-O- β-D- glucopyranosyls-(1 → 4)-β-D- galactopyranosides can be from bittersweet (for plant of Solanaceae
Bittersweet Solanum lyratum Thunb's dries herb) extract in medicinal material and obtain.But yet there are no and exist on above-claimed cpd
The report of the application aspect for the treatment of cancer disease areas, is not yet developed and is applied to clinical anticancer, is especially controlled
Lung cancer disease aspect is treated, the application is it is found by the applicant that above-claimed cpd effect is significant in terms for the treatment of cancer especially lung cancer, is
This, the present invention proposes a kind of above-claimed cpd and is preparing new application of cancer therapy drug and preparation method thereof.
The content of the invention
The technical problem to be solved in the present invention is new application and its system of a kind of compound in cancer therapy drug is prepared
Preparation Method, specially (3 β, 5 α, 22 α, 25R)-spirosolane -3-O- β-D- glucopyranosyls-(1 → 2)-O- [β-D- wood
Pyranose-(1 → 3)]-O- β-D- glucopyranosyls-(1 → 4)-β-D- galactopyranosides new application and its preparation side
Method.
Therefore, the purposes the invention provides compound of the one kind as shown in formula (I) in cancer therapy drug is prepared;
Described purposes, the cancer therapy drug includes anti-lung-cancer medicament.
Described purposes, the cancer therapy drug is anti-non-small cell lung cancer drug.
Present invention also offers a kind of cancer therapy drug, with the described compound shown in formula (I) as active ingredient.
Described cancer therapy drug, the compound shown in described formula (I), according to common process, is selected as active ingredient
Property add the customary adjuvant clinically-acceptable tablet, capsule, granule, syrup, powder, pill, tincture, the wine that are made
Agent, soft extract, lozenge or mixture.
Present invention also offers the method that one kind prepares the compound as shown in formula (I), comprise the following steps:
(1) taking during bittersweet medicinal material adds alcoholic solution carries out alcohol extracting, and the alcohol extract of acquisition is refined, and obtains final product bittersweet total
Alkali sample, it is standby;
(2) by heating for dissolving in the bittersweet total alkali sample addition alcoholic solution of the preparation in step (1), with tlc silica gel
Sample is mixed, being then splined in silica gel column chromatography carries out chromatography, with volume ratio as ethyl acetate-volumetric concentration is 93%~
97% ethanol=(2~3): 1 eluant, eluent is eluted, are detected with thin-layer chromatography, are improved bismuth potassium iodide by developer and are developed the color,
Merge identical stream part, obtain two kinds of different components of polarity, it is standby;
(3) the big component of polarity is dissolved in polar aprotic solvent in two components that will be obtained in step (2), then
Isolated and purified using high performance liquid chromatography, chromatographic condition is as follows:C18 liquid-phase chromatographic columns;With acetonitrile as mobile phase A, with 1%
The TFA aqueous solution is Mobile phase B, and gradient elution is carried out according to following procedure:0~10min, mobile phase A:The volume ratio of Mobile phase B
It is 24%~26%:76%~74% → 29%~31%:71%~69%;It is 25~35mL/min to control flow rate of mobile phase;
It is 20-30 DEG C to control column temperature;It is 0.5-1.5mL to control sampling volume;The efflux for collecting the compound carries out Mass Spectrometer Method,
The composition that relative molecular mass is 1033 is collected, the compound shown in formula (I) is obtained.
Described preparation method, in the step (1), the specific method for preparing bittersweet total alkali sample is:Bittersweet is done
Dry herb volumetric concentration is the ethanol water refluxing extraction of 65-75%, filtering, and concentrating the filtrate to 50 DEG C of relative densities is
1.05 medicinal extract, adds distilled water dispersion, filtering to take filtrate and add to D151 large pore resin absorption columns, successively with 2~4 times of cylinders
Long-pending distilled water and 2~4 times of volumetric concentrations of column volume are 93-97% ethanol waters wash-out, eluent are discarded, then with 3
~5 times of volumetric concentrations of column volume are eluted for the acidic alcohol aqueous solution of 5-7 ‰, the body of ethanol in the acidic alcohol aqueous solution
Product concentration is 93-97%, collects acidic alcohol eluent, and neutrality is neutralized to ammoniacal liquor, is filtered, and concentrates the filtrate to do, with steaming
Distilled water is disperseed, and the liquid after dispersion is added into AB-8 macroporous absorbent resins, with 6~10 times of distillation water elutions of column volume, is discarded
Eluent, is then that 93-97% ethanol waters are eluted with the volumetric concentration of 3~5 times of column volumes, collects ethanol eluate, dense
Contracting drying, obtains final product bittersweet total alkali.
Described method, in the step (2), weighs bittersweet total alkali sample 30-40 weight portions, adds 300-1000 bodies
The volumetric concentration of product part is subsequently adding 10-20 weight portions to be dissolved in the case where temperature is for 80-100 DEG C in the ethanol solution of 93-97%
Tlc silica gel be well mixed, be then splined on add 780-820 weight portions tlc silica gel silica gel column chromatography
In carry out chromatography;The weight portion is g/mL with the relation of parts by volume.
Described preparation method, in the step (2), with volume ratio as ethyl acetate-volumetric concentration be 95% ethanol
=2.5: 1 eluant, eluent is eluted.
Described preparation method, in the step (2), the aqueous hydrochloric acid solution improves the specific side of bismuth potassium iodide test solution
Method is:
The weight portion of basic bismuth nitrate 0.8~0.9 is weighed, glacial acetic acid, 39~41 parts by volume of 9~11 parts by volume are sequentially added
Water and 19~21 parts by volume liquor kalii iodide, it is well mixed to obtain final product bismuth potassium iodide test solution;
To adding mass concentration to be the aqueous hydrochloric acid solution of 0.6mol/L in the bismuth potassium iodide test solution, the bismuth potassium iodide is tried
Liquid is 1 with the volume ratio of the aqueous hydrochloric acid solution:2, obtain final product the bismuth potassium iodide test solution of the aqueous hydrochloric acid solution improvement.
The weight portion is g/mL with the relation of parts by volume.
Preferably, in the step (3), the mobile phase A:The volume ratio of Mobile phase B is 25%:75% → 30%:
70%.
Technical solution of the present invention, has the following advantages that:
1. the compound as shown in formula (I) of the present invention has the obvious effect for suppressing cancer cell, especially presses down
The effect of lung carcinoma cell processed, with the increase of the compound concentration, the inhibitory action to non-small cell lung cancer cell gradually increases
By force, by lung cancer cell growth cycle arrest in early stage such as G0/G1Phase, and with the increase of the compound concentration, delay effect
Gradually strengthen, while the effect of the compound makes the apoptosis rate of A549 cells and concentration be proportionate gradually increase, therefore, formula
(I) compound shown in can be used for preparing cancer therapy drug, especially prepare anti-lung-cancer medicament.
2. the method for compound of the preparation of the present invention as shown in formula (I), comprises the following steps:(1) bittersweet medicine is taken
Material carries out alcohol extracting in adding alcoholic solution, and the alcohol extract of acquisition is refined, and obtains final product bittersweet total alkali sample, standby;(2) will step
Suddenly the bittersweet total alkali sample of the preparation in (1) adds heating for dissolving in alcoholic solution, and sample is mixed with tlc silica gel, is then splined on
Chromatography is carried out in silica gel column chromatography, with volume ratio as ethyl acetate-volumetric concentration be 93%~97% ethanol=(2~3):
1 eluant, eluent is eluted, and is detected with thin-layer chromatography, collects the stream part containing target compound, standby;(3) by step (2)
The stream part of acquisition is dissolved in polar aprotic solvent, is then isolated and purified using high performance liquid chromatography, and chromatographic condition is such as
Under:C18 liquid-phase chromatographic columns;With acetonitrile as mobile phase A, with the 1%TFA aqueous solution as Mobile phase B, gradient is carried out according to following procedure
Wash-out:0~10min, mobile phase A:The volume ratio of Mobile phase B is 24%~26%:76%~74% → 29%~31%:71%
~69%;It is 25~35mL/min to control flow rate of mobile phase;It is 20-30 DEG C to control column temperature;It is 0.5- to control sampling volume
1.5mL;Composition is collected using mass detector, the compound shown in formula (I) is obtained;Can be from bittersweet medicinal material by the above method
In isolate and purify out compound shown in formula (I).
3. the method for compound of the preparation as shown in formula (I) of the present invention, in the step (3), the flowing
Phase A:The volume ratio of Mobile phase B is 25%:75% → 30%:70%;By selecting suitable mobile phase, formula is substantially increased
(I) compound shown in isolates and purifies purity, reduces impurity content, facilitates it to be applied to clinic.
4. the method for compound of the preparation as shown in formula (I) of the present invention, with hydrochloric acid during column chromatography for separation
The bismuth potassium iodide test solution of aqueous solution improvement is developer, and the preparation method of the developer is:To being sequentially added in basic bismuth nitrate
Glacial acetic acid, water and liquor kalii iodide, then it is water-soluble to the hydrochloric acid for adding mass concentration to be 0.6mol/L in the test solution after being well mixed
Liquid, the bismuth potassium iodide test solution is 1 with the volume ratio of the aqueous hydrochloric acid solution:2, that is, the iodine of the aqueous hydrochloric acid solution improvement is obtained
Change bismuth potassium test solution, when the developer that the preparation method is prepared causes to be detected with thin-layer chromatography, product component is relative to be carried on the back
Developed the color for scenery clear, easily determined.
Brief description of the drawings
In order to illustrate more clearly of the specific embodiment of the invention or technical scheme of the prior art, below will be to specific
The accompanying drawing to be used needed for implementation method or description of the prior art is briefly described, it should be apparent that, in describing below
Accompanying drawing is some embodiments of the present invention, for those of ordinary skill in the art, before creative work is not paid
Put, other accompanying drawings can also be obtained according to these accompanying drawings.
Fig. 1 is that the embodiment of the present invention 1 prepares (3 β, 5 α, 22 α, 25R)-spirosolane -3-O- β-D- pyrans Portugal
Grape glycosyl-(1 → 2)-O- [β-D- xylopyranoses base-(1 → 3)]-O- β-D- glucopyranosyls-(1 → 4)-β-D- pyrans half
Lactoside structural formula;
Fig. 2 is that the embodiment of the present invention 1 prepares (3 β, 5 α, 22 α, 25R)-spirosolane -3-O- β-D- glucopyras
Glycosyl-(1 → 2)-O- [β-D- xylopyranoses base-(1 → 3)]-O- β-D- glucopyranosyls-(1 → 4)-β-D- galactopyranosyls
Glucosides1HNMR spectrograms;
Fig. 3 is that the embodiment of the present invention 1 prepares (3 β, 5 α, 22 α, 25R)-spirosolane -3-O- β-D- pyrans Portugal
Grape glycosyl-(1 → 2)-O- [β-D- xylopyranoses base-(1 → 3)]-O- β-D- glucopyranosyls-(1 → 4)-β-D- pyrans half
Lactoside13C NMR spectras;
Fig. 4 is that the embodiment of the present invention 1 prepares (3 β, 5 α, 22 α, 25R)-spirosolane -3-O- β-D- pyrans Portugal
Grape glycosyl-(1 → 2)-O- [β-D- xylopyranoses base-(1 → 3)]-O- β-D- glucopyranosyls-(1 → 4)-β-D- pyrans half
Lactoside H-H cosy spectrograms;
Fig. 5 is that the embodiment of the present invention 1 prepares (3 β, 5 α, 22 α, 25R)-spirosolane -3-O- β-D- pyrans Portugal
Grape glycosyl-(1 → 2)-O- [β-D- xylopyranoses base-(1 → 3)]-O- β-D- glucopyranosyls-(1 → 4)-β-D- pyrans half
Lactoside HMQC spectrograms;
Fig. 6 is that the embodiment of the present invention 1 prepares (3 β, 5 α, 22 α, 25R)-spirosolane -3-O- β-D- pyrans Portugal
Grape glycosyl-(1 → 2)-O- [β-D- xylopyranoses base-(1 → 3)]-O- β-D- glucopyranosyls-(1 → 4)-β-D- pyrans half
Lactoside HMBC spectrograms;
Fig. 7 is that the embodiment of the present invention 1 prepares (3 β, 5 α, 22 α, 25R)-spirosolane -3-O- β-D- pyrans Portugal
Grape glycosyl-(1 → 2)-O- [β-D- xylopyranoses base-(1 → 3)]-O- β-D- glucopyranosyls-(1 → 4)-β-D- pyrans half
Lactoside Tocsy spectrograms;
Fig. 8 is that the embodiment of the present invention 1 prepares (3 β, 5 α, 22 α, 25R)-spirosolane -3-O- β-D- pyrans Portugal
Grape glycosyl-(1 → 2)-O- [β-D- xylopyranoses base-(1 → 3)]-O- β-D- glucopyranosyls-(1 → 4)-β-D- pyrans half
Lactoside Noesy spectrograms;
Fig. 9 is (3 β, 5 α, 22 α, 25R)-spirosolane -3-O- β-D- pyrans Portugal that the embodiment of the present invention 1 is prepared
Grape glycosyl-(1 → 2)-O- [β-D- xylopyranoses base-(1 → 3)]-O- β-D- glucopyranosyls-(1 → 4)-β-D- pyrans half
Impact effect figure of the lactoside to A549, H460, SK-MES-1 cell inhibitory rate;
Figure 10 a are the streaming result figures that blank control group influences on A549 Apoptosis in Apoptosis influence;
It is (3 β, 5 α, 22 α, the 25R)-spiral shell prepared containing the embodiment of the present invention 1 in Apoptosis influence that Figure 10 b are
Rotation steroid alkali alkane -3-O- β-D- glucopyranosyls-(1 → 2)-O- [β-D- xylopyranoses base-(1 → 3)]-O- β-D- glucopyras
The streaming result figure that the low concentration group of the medicine of glycosyl-(1 → 4)-β-D- galactopyranosides influences on A549 Apoptosis;
It is (3 β, 5 α, 22 α, the 25R)-spiral shell prepared containing the embodiment of the present invention 1 in Apoptosis influence that Figure 10 c are
Rotation steroid alkali alkane -3-O- β-D- glucopyranosyls-(1 → 2)-O- [β-D- xylopyranoses base-(1 → 3)]-O- β-D- glucopyras
The streaming result figure that the middle concentration group of the medicine of glycosyl-(1 → 4)-β-D- galactopyranosides influences on A549 Apoptosis;
It is (3 β, 5 α, 22 α, the 25R)-spiral shell prepared containing the embodiment of the present invention 1 in Apoptosis influence that Figure 10 d are
Rotation steroid alkali alkane -3-O- β-D- glucopyranosyls-(1 → 2)-O- [β-D- xylopyranoses base-(1 → 3)]-O- β-D- glucopyras
The streaming result figure that the high concentration group of the medicine of glycosyl-(1 → 4)-β-D- galactopyranosides influences on A549 Apoptosis.
Specific embodiment
Technical scheme is clearly and completely described below in conjunction with accompanying drawing, it is clear that described implementation
Example is a part of embodiment of the invention, rather than whole embodiments.Based on the embodiment in the present invention, ordinary skill
The every other embodiment that personnel are obtained under the premise of creative work is not made, belongs to the scope of protection of the invention.
As long as additionally, technical characteristic involved in invention described below different embodiments does not constitute conflict just each other
Can be combined with each other.
The instrument being related in following embodiments is as follows:Waters 2767/QDa prepare liquid phase mass spectrometry chromatograph:
Masslynx4.1 chromatographic work stations, Waters2767 sample managers, Waters2489 ultraviolet-visible detectors,
Waters2545 binary high pressure chromatogram pumps, QDa mass detectors, chromatographic column is Waters SunFire Prep C18OBD (10 μ
M, 19 × 250mm) Column.
Embodiment 1
The preparation method of the compound as shown in formula (I) described in the present embodiment, comprises the following steps:
(1) 10kg bittersweets are taken and dry herb, be placed in Backflow bottle, add the ethanol water that 80L volumetric concentrations are 70%,
In refluxing extraction 3 times at 100 DEG C, extract 2 hours every time, the alcohol extract of acquisition is filtered, the filtrate obtained by merging is simultaneously
It is the medicinal extract of 1.05 (50 DEG C of surveys) to carry out being concentrated under reduced pressure into relative density, to 10 times of distillations of medicinal extract quality of addition in the medicinal extract
Water is disperseed, filtering, and gained filtrate is added to D151 ion exchange large pore resin absorption columns, then successively through 3 times of cylinders
Long-pending distilled water, the ethanol water wash-out that the volumetric concentration of 3 times of column volumes is 95%, discards the eluent of outflow, then through 4
Times column volume is 95% ethanol solution wash-out containing the volumetric concentration that volumetric concentration is 6 ‰ hydrochloric acid, the eluent of outflow is collected, to institute
State and add in eluent ammoniacal liquor and be neutralized to neutrality, filter, gained filtrate decompression is concentrated to dryness, and with 1 times of steaming of crude drug amount
Distilled water is disperseed, and scattered liquid is added into AB-8 large pore resin absorption columns, then with 8 times of distillation water elutions of column volume, is discarded
The eluent of outflow, then elutes through the ethanol solution that the volumetric concentration of 4 times of column volumes is 95%, collects the eluent of outflow,
Then concentration is spin-dried for, and is spray-dried, and obtains final product 35g bittersweet total alkali samples, and yield is about 3.5 ‰, standby;
(2) the bittersweet total alkali sample 35g of the preparation in step (1) is weighed, the volumetric concentration for adding 500ml is 95% second
90 DEG C of heating water bath dissolvings, sample is mixed with tlc silica gel in alcoholic solution, is then splined on the tlc silica gel equipped with 800g
Silica gel column chromatography in carry out chromatography, with volume ratio as ethyl acetate-volumetric concentration be 95% ethanol=2.5: 1 wash-out
Agent is eluted, and is detected with thin-layer chromatography, is led to, and the bismuth potassium iodide test solution with aqueous hydrochloric acid solution improvement merges as chromogenic reagent
Identical stream part, obtains two kinds of different components of polarity, standby;
The specific method of aqueous hydrochloric acid solution improvement bismuth potassium iodide test solution is:Basic bismuth nitrate 0.85g is weighed, is added successively
Enter 10mL glacial acetic acid, 40mL water and 20mL liquor kalii iodides, it is well mixed to obtain final product bismuth potassium iodide test solution;Take the iodine that 1mL is configured
Change bismuth potassium test solution, it is the aqueous hydrochloric acid solution of 0.6mol/L to add 2mL mass concentrations, obtain final product the iodate of the aqueous hydrochloric acid solution improvement
Bismuth potassium test solution;
(3) the big component of polarity is dissolved in dimethylformamide (DMF) in the two kinds of components that will be obtained in step (2), molten
Sample solution concentration after solution is 100mg/ml, using 0.45um membrane filtrations, is then separated using high performance liquid chromatography pure
Change, chromatographic condition is as follows:C18 liquid-phase chromatographic columns are Waters SunFire Prep C18OBD (10 μm, 19 × 250mm)
Column;With acetonitrile (preparation scale) as mobile phase A, with the 1%TFA aqueous solution as Mobile phase B, carry out gradient according to following procedure and wash
It is de-:0~10min, mobile phase A:The volume ratio of Mobile phase B is 25%:75% → 30%:70%;The flow rate of mobile phase is controlled to be
30mL/min;It is 25 DEG C to control column temperature;It is 1mL to control sampling volume;Composition is collected using QDa mass detectors, collects relative
Molecular mass is 1033 composition, prepares the compound 20mg shown in formula (I).
(3 β, 5 α, 22 α, 25R)-spirosolane -3-O- β-D- glucopyranosyls-(1 that the present embodiment is prepared
→ 2)-O- [β-D- xylopyranoses base-(1 → 3)]-O- β-D- glucopyranosyls-(1 → 4)-β-D- galactopyranosides1HNMR spectrograms,13C NMR spectras, H-H cosy spectrograms, HMQC spectrograms, HMBC spectrograms, Tocsy spectrograms and Noesy spectrograms difference
See Fig. 2-8.
(3 β, 5 α, 22 α, 25R)-spirosolane -3-O- β-D- glucopyranosyls-(1 that the present embodiment is prepared
→ 2)-O- [β-D- xylopyranoses base-(1 → 3)]-O- β-D- glucopyranosyls-(1 → 4)-β-D- galactopyranosides, its
Structural formula is as shown in figure 1, wherein carbon atoms numbered 1-27 is marked in the structural formula shown in Fig. 1.Table 1 is the solution of Fig. 2-8
Analysis result.
The analysis result of the Fig. 2-8 of table 1
Wherein, in table 1 hydrogen spectrum and carbon modal data be1H-NMR (600MHz) and13C-NMR (150MHz), solvent is deuterium
For being obtained under conditions of methyl alcohol
Embodiment 2
The preparation method of the compound as shown in formula (I) described in the present embodiment, comprises the following steps:
(1) 10kg bittersweets are taken and dry herb, be placed in Backflow bottle, plus 60L volumetric concentration be 65% ethanol water,
In refluxing extraction 2 times at 90 DEG C, extract 4 hours every time, the alcohol extract of acquisition is filtered, the filtrate obtained by merging is gone forward side by side
It is the medicinal extract of 1.05 (50 DEG C of surveys) that row is concentrated under reduced pressure into relative density, to 8 times of distilled water of medicinal extract quality of addition in the medicinal extract
Disperseed, filtered, gained filtrate is added to D151 ion exchange large pore resin absorption columns, then successively through 2 times of column volumes
Distilled water, ethanol solution that the volumetric concentration of 4 times of column volumes is 93% wash-out, the eluent of outflow is discarded, then through 3 times of posts
Volume, for 93% ethanol solution is eluted, collects the eluent of outflow containing the volumetric concentration that volumetric concentration is 7 ‰ hydrochloric acid, is washed to described
Add ammoniacal liquor to be neutralized to neutrality in de- liquid, filter, gained filtrate decompression is concentrated to dryness, and with 1.5 times of distillations of crude drug amount
Moisture is dissipated, and scattered liquid is added into AB-8 large pore resin absorption columns top, then with 6 times of distillation water elutions of column volume, is discarded
The eluent of outflow, then elutes through the ethanol solution that the volumetric concentration of 5 times of column volumes is 97%, collects the eluent of outflow,
Then concentration is spin-dried for, and is spray-dried, and obtains final product 33g bittersweet total alkali samples, standby;
(2) the bittersweet total alkali sample 30g of the preparation in step (1) is weighed, the volumetric concentration for adding 300ml is 97% second
Water-bath dissolving at being 80 DEG C in temperature in alcoholic solution, adds 20g tlc silica gels to mix sample, is then splined on the thin of addition 820g
Layer chromatographic silica gel silica gel column chromatography in carry out chromatography, with volume ratio as ethyl acetate-volumetric concentration be 97% ethanol=2
: 1 eluant, eluent is eluted, with thin-layer chromatography detect, with aqueous hydrochloric acid solution improvement bismuth potassium iodide test solution as developer, with thin
Layer chromatography is detected that the identical stream part of merging obtains two kinds of different components of polarity;
The specific method of aqueous hydrochloric acid solution improvement bismuth potassium iodide test solution is:Basic bismuth nitrate 0.8g is weighed, is added successively
Enter 11mL glacial acetic acid, 39mL water and 21mL liquor kalii iodides, it is well mixed to obtain final product bismuth potassium iodide test solution;Take the iodine that 1mL is configured
Change bismuth potassium test solution, it is the aqueous hydrochloric acid solution of 0.6mol/L to add 2mL mass concentrations, obtain final product the iodate of the aqueous hydrochloric acid solution improvement
Bismuth potassium test solution;
(3) the big component of polarity is dissolved in dimethylformamide (DMF) in the two kinds of components that will be obtained in step (2), molten
Sample solution concentration after solution is 90mg/ml, using 0.45um membrane filtrations, then using high performance liquid chromatography-mass spectrum connection
With isolating and purifying, chromatographic condition is as follows:C18 liquid-phase chromatographic columns be Waters SunFire Prep C18OBD (10 μm, 19 ×
250mm)Column;With acetonitrile (preparation scale) as mobile phase A, with the 1%TFA aqueous solution as Mobile phase B, carried out according to following procedure
Gradient elution:0~10min, mobile phase A:The volume ratio of Mobile phase B is 24%:76% → 31%:69%;Control mobile phase stream
Speed is 25mL/min;It is 20 DEG C to control column temperature;It is 0.5mL to control sampling volume;Composition is collected using QDa mass detectors, is received
Collection relative molecular mass is 1033 composition, prepares the compound 20mg shown in formula (I).
The present embodiment additionally provides a kind of cancer therapy drug, the cancer therapy drug with (3 β, 5 α, 22 α, 25R) of above-mentioned preparation-
Spirosolane -3-O- β-D- glucopyranosyls-(1 → 2)-O- [β-D- xylopyranoses base-(1 → 3)]-O- β-D- pyrans Portugal
Grape glycosyl-(1 → 4)-β-D- galactopyranosides are that active ingredient adds customary adjuvant, are made according to common process and clinically may be used
The tablet of receiving.
Embodiment 3
The preparation method of the compound as shown in formula (I) described in the present embodiment, comprises the following steps:
(1) 10kg bittersweets are taken and dry herb, be placed in Backflow bottle, plus 100L volumetric concentration be 75% ethanol solution,
In refluxing extraction 3 times at 110 DEG C, extract 2 hours every time, the alcohol extract of acquisition is filtered, the filtrate obtained by merging is simultaneously
It is the medicinal extract of 1.05 (50 DEG C of surveys) to carry out being concentrated under reduced pressure into relative density, to 12 times of distillations of medicinal extract quality of addition in the medicinal extract
Water is disperseed, filtering, and gained filtrate is added to D151 ion exchange large pore resin absorption columns, then successively through 4 times of cylinders
Long-pending distilled water, the ethanol solution wash-out that the volumetric concentration of 2 times of column volumes is 97%, discards the eluent of outflow, then through 5 times
Column volume, for 97% ethanol solution is eluted, collects the eluent of outflow, to described containing the volumetric concentration that volumetric concentration is 5 ‰ hydrochloric acid
Add ammoniacal liquor to be neutralized to neutrality in eluent, filter, gained filtrate decompression is concentrated to dryness, and with 0.5 times of steaming of crude drug amount
Distilled water is disperseed, and scattered liquid is added into AB-8 large pore resin absorption columns top, then with 10 times of distillation water elutions of column volume, is abandoned
The eluent of outflow is removed, is then eluted through the ethanol solution that the volumetric concentration of 3 times of column volumes is 93%, collect the wash-out of outflow
Liquid, then concentration is spin-dried for, and is spray-dried, and obtains final product 35g bittersweet total alkali samples, and yield is about 3.5 ‰, standby;
(2) the bittersweet total alkali sample 40g of the preparation in step (1) is weighed, the volumetric concentration for adding 1000ml is 93%
Water-bath dissolving at being 100 DEG C in temperature in ethanol solution, adds 10g tlc silica gels to mix sample, is then splined on addition 780g
Tlc silica gel silica gel column chromatography in carry out chromatography, with volume ratio as ethyl acetate-volumetric concentration be 93% second
Alcohol=3: 1 eluant, eluent is eluted, are detected with thin-layer chromatography, and the bismuth potassium iodide test solution with aqueous hydrochloric acid solution improvement is colour developing
Agent, is detected with thin-layer chromatography, merges identical stream part, obtains two kinds of different components of polarity;
The specific method of aqueous hydrochloric acid solution improvement bismuth potassium iodide test solution is:Basic bismuth nitrate 0.9g is weighed, is added successively
Enter 9mL glacial acetic acid, 41mL water and 19mL liquor kalii iodides, it is well mixed to obtain final product bismuth potassium iodide test solution;Take the iodate that 1mL is configured
Bismuth potassium test solution, it is the aqueous hydrochloric acid solution of 0.6mol/L to add 2mL mass concentrations, obtains final product the bismuth iodide of the aqueous hydrochloric acid solution improvement
Potassium test solution;
(3) the big component of polarity is dissolved in dimethylformamide (DMF) in the two kinds of components that will be obtained in step (2), molten
Sample solution concentration after solution is 105mg/ml, using 0.45um membrane filtrations, is then separated using high performance liquid chromatography pure
Change, chromatographic condition is as follows:C18 liquid-phase chromatographic columns are Waters SunFire Prep C18OBD (10 μm, 19 × 250mm)
Column;With acetonitrile (preparation scale) as mobile phase A, with the 1%TFA aqueous solution as Mobile phase B, carry out gradient according to following procedure and wash
It is de-:0~10min, mobile phase A:The volume ratio of Mobile phase B is 26%:74% → 29%:71%;The flow rate of mobile phase is controlled to be
35mL/min;It is 30 DEG C to control column temperature;It is 1.5mL to control sampling volume;Composition is collected using QDa mass detectors, phase is collected
It is 1033 composition to molecular mass, prepares the compound 20mg shown in formula (I).
A kind of cancer therapy drug is present embodiments provided, the cancer therapy drug is with (3 β, 5 α, 22 α, 25R)-spiral shell of above-mentioned preparation
Rotation steroid alkali alkane -3-O- β-D- glucopyranosyls-(1 → 2)-O- [β-D- xylopyranoses base-(1 → 3)]-O- β-D- glucopyras
Glycosyl-(1 → 4)-β-D- galactopyranosides are that active ingredient adds customary adjuvant, and being made according to common process can clinically connect
The capsule received.
Embodiment 4
Present embodiments provide a kind of cancer therapy drug, the cancer therapy drug with embodiment 1 prepare (3 β, 5 α, 22 α,
25R)-spirosolane -3-O- β-D- glucopyranosyls-(1 → 2)-O- [β-D- xylopyranoses base-(1 → 3)]-O- β-D-
Glucopyranosyl-(1 → 4)-β-D- galactopyranosides are active ingredient.
Embodiment 5
Present embodiments provide a kind of cancer therapy drug, the cancer therapy drug with embodiment 1 prepare (3 β, 5 α, 22 α,
25R)-spirosolane -3-O- β-D- glucopyranosyls-(1 → 2)-O- [β-D- xylopyranoses base-(1 → 3)]-O- β-D-
Glucopyranosyl-(1 → 4)-β-D- galactopyranosides are that active ingredient adds customary adjuvant, are made according to common process and faced
Acceptable granule on bed.
Embodiment 6
Present embodiments provide a kind of cancer therapy drug, the cancer therapy drug with embodiment 1 prepare (3 β, 5 α, 22 α,
25R)-spirosolane -3-O- β-D- glucopyranosyls-(1 → 2)-O- [β-D- xylopyranoses base-(1 → 3)]-O- β-D-
Glucopyranosyl-(1 → 4)-β-D- galactopyranosides are that active ingredient adds customary adjuvant, are made according to common process and faced
Acceptable pill on bed.
Embodiment 7
Present embodiments provide a kind of cancer therapy drug, the cancer therapy drug with embodiment 1 prepare (3 β, 5 α, 22 α,
25R)-spirosolane -3-O- β-D- glucopyranosyls-(1 → 2)-O- [β-D- xylopyranoses base-(1 → 3)]-O- β-D-
Glucopyranosyl-(1 → 4)-β-D- galactopyranosides are that active ingredient adds customary adjuvant, are made according to common process and faced
Acceptable syrup on bed.
Embodiment 8
Present embodiments provide a kind of cancer therapy drug, the cancer therapy drug with embodiment 1 prepare (3 β, 5 α, 22 α,
25R)-spirosolane -3-O- β-D- glucopyranosyls-(1 → 2)-O- [β-D- xylopyranoses base-(1 → 3)]-O- β-D-
Glucopyranosyl-(1 → 4)-β-D- galactopyranosides are that active ingredient adds customary adjuvant, are made according to common process and faced
Acceptable soft extract on bed.
Experimental example
Compound prepared by the embodiment of the present invention 1 is to the inhibiting rate of non-small cell lung cancer and cell cycle, the influence of apoptosis
1 material
1.1 cellular informatics
Three kinds of non-small cell lung cancer cell strains:Human A549 cell lines, human large cell lung cancer cell H460, people's lung squamous cancer
Cell SK-MES-1, above-mentioned cell is provided by Jiangsu Kai Ji Biotechnology Ltd..A549 and H460 cells are used
Complete medium be containing volumetric concentration be that 90%RPMI640 and volumetric concentration are the mixture of 10%FBS, SK-MES-1 is thin
It containing volumetric concentration is that 90%DMEM and volumetric concentration are the mixture of 10%FBS that the complete medium that born of the same parents use is, in 37 DEG C,
Volumetric concentration 5%CO2, saturated humidity incubator in cultivate.
1.2 main agents and consumptive material
Tissue Culture Flask (U.S. FALCON 353014)
Penicillin/streptomycin solution (Jiangsu Province, China Kai Ji Biotechnology Ltd.
KGY002)
0.25%Tripsin-EDTA (Jiangsu Province, China Kai Ji Biotechnology Ltd. KGY001)
MTT (U.S. Amresco 0793)
DMSO (dissolving tested medicine) (U.S. SIGMA D2650)
PBS (Jiangsu Province, China Kai Ji Biotechnology Ltd. KGB500)
RPMI-1640 (U.S. GIBCO 31800-105)
DMEM (U.S. GIBCO 12800-082)
FBS (U.S. ExCell Biology FBS500)
96 wellcell culture plate (U.S. Corning Incorporated 3599)
6 wellcell culture plate(Corning Incorporated 3516)
Annexin V-APC/7-AAD cell apoptosis detection kits (the triumphant limited public affairs of base biotechnology share of Jiangsu Province, China
Department KGA1024)
Cell cycle detection kit (Jiangsu Province, China Kai Ji Biotechnology Ltd. KGA511)
1.3 key instruments and equipment
Superclean bench (Chinese Suzhou purifies SW-CJ-1FD)
CO2 incubators (Japanese SANYO XD-101)
Biological inverted microscope (Japanese OLYMPUS IX51)
Table-type low-speed centrifuge (Chinese Shanghai Medical Treatment Equipment Co., Ltd Medical Equipment Plant 80-2)
2.5ul, 10ul, 200ul, 1000ul pipettor (German eppendorf)
Vertical pressure pot (the rich news of Chinese Shanghai, YXQ-LS-50)
Electric drying oven with forced convection (Chinese Shanghai sage is glad, 101AS-3)
ELIASA (U.S. BioTek ELx800)
Oscillator (Chinese Shanghai Hu Xi analytical instrument factory WH-2)
Flow cytometer (U.S. Becton-Dickinson FACS Calibur)
2 methods
2.1 cell culture
(1) when the cell coverage rate in blake bottle reaches 80%~90%, original culture medium is sopped up;
(2) add appropriate trypsase (0.25%), digest 1~2min;
(3) isometric culture medium containing serum is added to terminate digestion after cell is all rounded;
(4) cell is blown and beaten with liquid-transfering gun, cell is all suspended, then cell is drawn onto in the centrifuge tube of 15ml,
1000rpm is centrifuged 5min;
(5) supernatant is outwelled, plus 1~2ml culture mediums, continuation in being transferred to blake bottle that cell is suspended again is cultivated.
2.2MTT methods detection cell propagation
(1) take the logarithm respectively growth period A549, H460, SK-MES-1 cell dissociation, count, be configured to concentration for 5 × 104
The cell suspension of individual/ml, adds 100 μ l cell suspensions per hole in 96 porocyte culture plates;
(2) 96 porocyte culture plates are placed in 37 DEG C, volumetric concentration is 5%CO224h is cultivated in incubator;
(3) diluted needed for the compound to table 2 that medicine is prepared to the dilution embodiment of the present invention 1 with complete medium
Drug concentration, the pastille culture medium of 100 μ l respective concentrations is added per hole, while setting up blank control group (adds same volume blank
Nutrient solution);
(4) 96 porocyte culture plates are placed in 37 DEG C, volumetric concentration is 5%CO272h is cultivated in incubator;
(5) 96 orifice plates are carried out into MTT dyeing, λ=490nm determines OD values;
A adds 20 μ l MTT (5mg/ml) per hole, continues to cultivate 4h in incubator;
B discards culture medium, and 150 μ l DMSO dissolvings are added per hole, and shaking table 10min is gently mixed;
C λ=490nm, ELIASA reads the absorbance OD values per hole, calculates inhibiting rate;
(6) each group inhibiting rate and medicine 503nhibiting concentration (IC50) are counted
In above formula, A is absorbance.
Pass through probit using SPSS (Staffstical Package for the Social Science) 17.0
Weighted regression method (Bliss methods) calculates IC50.
2.3PI mono- dye method detection cell cycles
Selection A549 cells carry out the detection of cell cycle and apoptosis, and are carried out by 2.2 results selection, three concentration
Follow-up test, cell is divided into four groups:Blank control group, low concentration group, middle concentration group, high concentration group, above-mentioned each group cell is dense
Spend is 5 × 106The cell of individual/ml, step is as follows
(1) the A549 cell dissociations of exponential phase are inoculated into six orifice plates, next day, after after cell attachment, according to group
Corresponding pastille culture medium concentration She Zhi not be added to be followed successively by (blank control group is mended with same volume blank nutrient solution);The pastille
Culture medium is the compound prepared using complete medium dilution embodiment 1, the compound in the pastille culture medium of low concentration group
Concentration is 0.75 μ g/ml, and middle concentration group is 1.50 μ g/ml, and high concentration group is 3.00 μ g/ml;
(2) after medicine effect 72h, each group cell is collected with 0.25% pancreatin (being free of EDTA) digestion respectively;
(3) various cells once (centrifugation 2000rpm, 5min), every group of collection 5 × 10 are washed with PBS5Individual cell, respectively
Cell concentration is configured to for single cell suspension 1ml;
(4) each group single cell suspension 1ml volumetric concentrations for preparing step (3) fix 2h (or mistakes for 70% ethanol
Night), 4 DEG C of preservations wash away fixer (if desired, 200 mesh sieve net filtrations of cell suspension are once) before dyeing with PBS;
(5) 37 DEG C of water-bath 30min of RNase A of 100 μ l are added to each group cell in step (4) respectively;
(6) add the PI of 400 μ l to dye to the cell in step (5) respectively again to mix, 4 DEG C of lucifuge 30min;
(7) machine testing on, red fluorescence at record excitation wavelength 488nm.
The double dye method detection Apoptosis of 2.4 Annexin-V APC/7-AAD
(1) the A549 cell dissociations of exponential phase are inoculated into six orifice plates, next day, after after cell attachment, according to group
The pastille culture medium of respective concentration She Zhi not be added, cell is divided into four groups:Blank control group, low concentration group, middle concentration group is high
Concentration group;(blank control group is mended with same volume blank nutrient solution);The pastille culture medium is to dilute real using complete medium
The compound of the preparation of example 1 is applied, the compound concentration in the pastille culture medium of low concentration group is 0.75 μ g/ml, and middle concentration group is
1.50 μ g/ml, high concentration group is 3.00 μ g/ml;Every group of cell concentration is 5 × 106The cell of individual/ml;
(2) after medicine effect 72h, each group cell is collected with 0.25% pancreatin (being free of EDTA) digestion;
(3) 5 × 10 are collected with PBS washed cells 2 times (centrifugation 2000rpm, 5min)5Individual cell;
(4) the Binding Buffer suspension cells of 500 μ l are added;
(5) after adding 5 μ l Annexin V-APC to mix, the 7-AAD of 5 μ l is added, is mixed;
(6) room temperature, lucifuge, 5~15min of reaction;With the situation of flow cytomery Apoptosis.
3 results
3.1 pairs of three kinds of inhibiting rates of non-small cell lung cancer cell
As the increase of the compound concentration, the inhibitory action to cell gradually strengthen, the compound is thin to A549
The 503nhibiting concentration of born of the same parents is 1.90 μ g/ml, and the 503nhibiting concentration to H460 cells is 6.38 μ g/ml, to SK-MES-1 cells
503nhibiting concentration is 4.66 μ g/ml.Concrete outcome is shown in Table 2, Fig. 9.
The medicine of table 2 to A549, H460, SK-MES-1 cell inhibitory rate influence (N=6)
3.2 pairs of influences of A549 cell cycles
The low concentration group for testing selection compound by MTT is 0.75 μ g/ml, and middle concentration group is 1.50 μ g/ml, high concentration
Group is 3.00 μ g/ml.The effect of the compound makes A549 cells arrests in G0/G1Phase, and with the compound concentration
Increase, this delay effect gradually strengthens;S phases and G2Phase cell is accordingly reduced.Concrete outcome is shown in Table 3.
The medicine of table 3 to the A549 cell cycles influence (N=3)
3.3 pairs of influences of A549 Apoptosis
Figure 10 is fluidic cell result figure, and UL represents upper left side region (being fragment and damage non-viable non-apoptotic cell) in figure, and UR is
Upper right side region (being non-viable apoptotic cell), LL is bottom-left quadrant (normal survivaling cell), and LR is that lower right region (is early stage
Apoptotic cell), apoptosis rate=cell proportion+UR areas of LR areas cell proportion.Be can be seen that by Flow cytometry result, institute
Stating the effect of compound and the apoptosis rate of A549 cells and concentration is proportionate gradually increases.Particularly, viable apoptotic cell is made
(LR areas cell proportion), non-viable apoptotic cell (UR areas cell proportion) showed increased, especially the early apoptosis of high concentration group are thin
Born of the same parents' (viable apoptotic cell of blank control group is 3.08%) have reached 35.72%.Concrete outcome is shown in Table 4.
Influence of the medicine of table 4 to A549 Apoptosis
4. experiment conclusion
(3 β, 5 α, 22 α, 25R)-spirosolane -3-O- β-D- glucopyranosyls that the present invention is prepared-(1 →
2)-O- [β-D- xylopyranoses base-(1 → 3)]-O- β-D- glucopyranosyls-(1 → 4)-β-D- galactopyranosides are to people's lung
Adenocarcinoma cell A549, human large cell lung cancer cell H460 and people's Lung Squamous Carcinoma Cells SK-MES-1 have preferable inhibition, and
Inhibition to human A549 cell lines is better than other two kinds of cancer cell effects, and the compound can influence A549 cells
Cycle, its growth cycle is stuck in early stage such as G0/G1Phase, and with the increase of the compound concentration, delay effect by
Cumulative strong, the compound can promote A549 Apoptosis, with the increase of the compound concentration, A549 apoptosis rates
It is higher.It is indicated above that (3 β, 5 α, 22 α, 25R)-spirosolane -3-O- β-D- glucopyranoses that the present invention is prepared
Base-(1 → 2)-O- [β-D- xylopyranoses base-(1 → 3)]-O- β-D- glucopyranosyls-(1 → 4)-β-D- galactopyranoses
Glycosides has significant antitumaous effect, in particular for treatment non-small cell lung cancer.
Obviously, above-described embodiment is only intended to clearly illustrate example, and not to the restriction of implementation method.It is right
For those of ordinary skill in the art, can also make on the basis of the above description other multi-forms change or
Change.There is no need and unable to be exhaustive to all of implementation method.And the obvious change thus extended out or
Among changing still in the protection domain of the invention.
Claims (10)
1. purposes of compound of the one kind as shown in formula (I) in cancer therapy drug is prepared;
2. purposes according to claim 1, it is characterised in that the cancer therapy drug includes anti-lung-cancer medicament.
3. purposes according to claim 1 and 2, it is characterised in that the cancer therapy drug is anti-non-small cell lung cancer drug.
4. a kind of cancer therapy drug, it is characterised in that with the described compound shown in formula (I) as active ingredient.
5. cancer therapy drug according to claim 4, it is characterised in that the compound shown in described formula (I) is effective
Composition, according to common process, selectively adds the clinically-acceptable preparation that customary adjuvant is made.
6. the method that one kind prepares the compound as shown in formula (I), it is characterised in that comprise the following steps:
(1) taking during bittersweet medicinal material adds alcoholic solution carries out alcohol extracting, and the alcohol extract of acquisition is refined, and obtains final product bittersweet total alkali sample
Product, it is standby;
(2) by heating for dissolving in the bittersweet total alkali sample addition alcoholic solution of the preparation in step (1), mixed with tlc silica gel
Sample, being then splined in silica gel column chromatography carries out chromatography, with volume ratio as ethyl acetate-volumetric concentration is 93%~97%
Ethanol=(2~3): 1 eluant, eluent is eluted, are detected with thin-layer chromatography, are improved bismuth potassium iodide by developer and are developed the color, and are merged
Identical stream part, obtains two kinds of different components of polarity, standby;
(3) the big component of polarity is dissolved in polar aprotic solvent in two components that will be obtained in step (2), is then used
High performance liquid chromatography is isolated and purified, and chromatographic condition is as follows:C18 liquid-phase chromatographic columns;With acetonitrile as mobile phase A, with 1%TFA water
Solution is Mobile phase B, and gradient elution is carried out according to following procedure:0~10min, mobile phase A:The volume ratio of Mobile phase B is 24%
~26%:76%~74% → 29%~31%:71%~69%;It is 25~35mL/min to control flow rate of mobile phase;Control column
Temperature is 20-30 DEG C;It is 0.5-1.5mL to control sampling volume;Collecting the efflux of the compound carries out Mass Spectrometer Method, collects phase
It is 1033 composition to molecular mass, obtains the compound shown in formula (I).
7. preparation method according to claim 6, it is characterised in that in the step (1), prepare bittersweet total alkali sample
Specific method be:Bittersweet is dried into the ethanol water refluxing extraction that herb volumetric concentration is 65-75%, is filtered, will filtered
Liquid is concentrated into the medicinal extract that 50 DEG C of relative densities are 1.05, adds distilled water dispersion, filtering to take filtrate and add to D151 macroporous absorption trees
Fat post, is successively 93-97% ethanol aqueous wash with the distilled water of 2~4 times of column volumes and the volumetric concentration of 2~4 times of column volumes
It is de-, eluent is discarded, then eluted with the acidic alcohol aqueous solution that the volumetric concentration of 3~5 times of column volumes is 5-7 ‰, the salt
The volumetric concentration of ethanol is 93-97% in sour ethanol water, collects acidic alcohol eluent, and neutrality, mistake are neutralized to ammoniacal liquor
Filter, concentrates the filtrate to do, and is disperseed with distilled water, the liquid after dispersion is added into AB-8 macroporous absorbent resins, with 6~10 times of posts
The distillation water elution of volume, discards eluent, is then 93-97% ethanol aqueous wash with the volumetric concentration of 3~5 times of column volumes
It is de-, ethanol eluate is collected, concentrate drying obtains final product bittersweet total alkali.
8. the method according to claim 6 or 7, it is characterised in that in the step (2), weigh bittersweet total alkali sample
30-40 weight portions, add 300-1000 parts by volume volumetric concentration for 93-97% ethanol solution in temperature be 80-100 DEG C
Lower dissolving, the tlc silica gel for being subsequently adding 10-20 weight portions is well mixed, and is then splined on addition 780-820 weight portions
Tlc silica gel silica gel column chromatography in carry out chromatography;The weight portion is g/mL with the relation of parts by volume.
9. preparation method according to claim 6, it is characterised in that be acetic acid second with volume ratio in the step (2)
Ester-volumetric concentration is 95% ethanol=2.5: 1 eluant, eluent is eluted.
10. preparation method according to claim 6, it is characterised in that in the step (2), the aqueous hydrochloric acid solution
Improve bismuth potassium iodide test solution specific method be:
The weight portion of basic bismuth nitrate 0.8~0.9 is weighed, glacial acetic acid, the water of 39~41 parts by volume of 9~11 parts by volume is sequentially added
With the liquor kalii iodide of 19~21 parts by volume, it is well mixed and obtains final product bismuth potassium iodide test solution;
To in the bismuth potassium iodide test solution add mass concentration for 0.6mol/L aqueous hydrochloric acid solution, the bismuth potassium iodide test solution with
The volume ratio of the aqueous hydrochloric acid solution is 1:2, obtain final product the bismuth potassium iodide test solution of the aqueous hydrochloric acid solution improvement.
The weight portion is g/mL with the relation of parts by volume.
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CN103193858A (en) * | 2013-04-12 | 2013-07-10 | 中国海洋大学 | Synthetic method of spirosolane glycoalkaloids |
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CN103193858A (en) * | 2013-04-12 | 2013-07-10 | 中国海洋大学 | Synthetic method of spirosolane glycoalkaloids |
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