CN102772639A - Preparation method of thunberg fritillary bulb total alkaloid extract - Google Patents
Preparation method of thunberg fritillary bulb total alkaloid extract Download PDFInfo
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- CN102772639A CN102772639A CN2012102834441A CN201210283444A CN102772639A CN 102772639 A CN102772639 A CN 102772639A CN 2012102834441 A CN2012102834441 A CN 2012102834441A CN 201210283444 A CN201210283444 A CN 201210283444A CN 102772639 A CN102772639 A CN 102772639A
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Abstract
The invention discloses a preparation method of a thunberg fritillary bulb total alkaloid extract. The preparation method includes the following steps of step one, taking and crushing thunberg fritillary bulbs, adding 65%-85% of an ethanol aqueous solution to the thunberg fritillary bulbs, subjecting the mixture to countercurrent extraction for 4-6 hours at the temperature of 30-60 DEG C, filtering the mixture, combining and collecting filter liquor and concentrating the filter liquor to obtain a concentrated solution; step two, adding muriatic acid with a mass fraction of 2% to the concentrated solution, performing full stirring and filtering, collecting filter liquor, using a muriatic acid solution with a mass fraction of 2% to wash residues for two times and combining the filter liquor; and step three, regulating the potential of hydrogen (pH) value of the filter liquor to be 7.0-9.0, using ethyl acetate to extract the obtained solution and subjecting the extracted solution to decompression concentration to obtain the thunberg fritillary bulb total alkaloid extract. By means of a countercurrent extraction method, alkaloid ingredients can be fully extracted, and compared with existing technical methods, the countercurrent extraction method has the advantages of being high in extraction efficiency, low in cost, capable of saving solvents, applicable to industrial production and the like.
Description
Technical field
The present invention relates to the technical field that effective ingredients in plant extracts, the method for preparing of total alkaloids extract in particularly a kind of Bulbus Fritillariae Thunbergii.
Background technology
Bulbus Fritillariae Thunbergii is the Liliaceae Fritillaria, and the dry bulb of Bulbus Fritillariae Thunbergii is one of conventional Chinese medicine, is classified as one of famous " Zhejiang eight flavors ", and its bitter in the mouth, cold in nature has effects such as clearing heat and moistening lung, preventing phlegm from forming and stopping coughing, mass dissipating and swelling eliminating.The main active of Bulbus Fritillariae Thunbergii is an alkaloid, contains peimine, verticinone, former peimine, Bulbus Fritillariae Thunbergii glycosides, different peimine etc.
Total alkaloid content in the Bulbus Fritillariae Thunbergii is very low, only contains 0.12%-0.25% (mass fraction), and this extracts for the total alkaloids of Bulbus Fritillariae Thunbergii and has brought special difficulty.At present, few to the literature research of Bulbus Fritillariae Thunbergii total alkaloids extract method for preparing, and extraction process consumption quantity of solvent is excessive, extraction efficiency is low, cost is too high, be not suitable for industrialized great production in the prior art.
Summary of the invention
Technical problem to be solved of the present invention provides the method for preparing of total alkaloids extract in a kind of Bulbus Fritillariae Thunbergii; The present invention uses counter-current to extract the total alkaloids in the Bulbus Fritillariae Thunbergii; It can extract the total alkaloids composition more fully; Compare with art methods and to have characteristics such as the solvent of saving, extraction efficiency is high, cost is low, be suitable for suitability for industrialized production.
The present invention solves the problems of the technologies described above the technical scheme that is adopted: the method for preparing of total alkaloids extract in a kind of Bulbus Fritillariae Thunbergii may further comprise the steps successively:
Step (1): get Bulbus Fritillariae Thunbergii, pulverizing back adding mass fraction is the ethanol water of 65-85%, and 30-60 ℃ of following countercurrent extraction 4-6h merges to collect after filtering and filtrate, and concentrates, and gets concentrated solution;
Step (2): the adding mass fraction is 2% hydrochloric acid in the said concentrated solution, and abundant the stirring filtered, and collects and filtrates, and residue use mass fraction is 2% hydrochloric acid solution washed twice, merges above-mentioned filtrating;
Step (3): adopt potassium hydroxide alkalization step (2) the gained filtrating of mass fraction 5%, transfer pH value to 7.0-9.0, gained solution is with ethyl acetate extraction, and the extract concentrating under reduced pressure gets the Bulbus Fritillariae Thunbergii total alkaloids extract.
In said step (1), adding mass fraction was 75% ethanol water, 40 ℃ of following countercurrent extraction 6 hours.
In the said step (2), the mass fraction that adds 1/4 times of raw medicinal herbs amount in the said concentrated solution is 2% hydrochloric acid.Raw medicinal herbs amount described herein is meant by Mass Calculation.Said " 1/4 times of raw medicinal herbs amount " promptly refers to add 1/4 times the hydrochloric acid solution that quality is the raw medicinal herbs amount.
In the said step (3), transfer pH value to 8.0.
The present invention compared with prior art has following advantage:
(1) the present invention adopts the countercurrent extraction method, can extract the alkaloids composition more fully, compares with art methods to have characteristics such as the solvent of saving, extraction efficiency is high, cost is low, is suitable for suitability for industrialized production.
Description of drawings
Fig. 1 is the graph of a relation of total alkaloids quality (mg) in alcoholic acid mass percent (%) and the leachate.
Fig. 2 is the graph of a relation of total alkaloids quality (mg) in time of contact (h) and the leachate.
Fig. 3 be temperature (℃) with leachate in the graph of a relation of total alkaloids quality (mg).
Fig. 4 is the schematic representation of apparatus that the method for preparing of total alkaloids extract in a kind of Bulbus Fritillariae Thunbergii of the present invention is used.
The specific embodiment
One, the extraction process condition is investigated
1, leaching condition experiment
(1) concentration of alcohol is to the influence of alkaloid compound leaching effect
In the commercial production; Owing to often contain certain moisture in the raw material; Ethanol also contains certain moisture after recycling; There is very big influence in concentration of alcohol to the leaching of alkaloid compound, so with different concentration ethanol liquid alkaloids substance is carried out leaching experiment, analyzes with the total alkaloids follow-up of quality.In concentration of alcohol is tested the influence of from raw material, leaching alkaloids, get 5 parts of raw materials respectively, every part of 40.00g experimentizes, with each 180.00ml of Different concentrations of alcohol aqueous solution, lixiviate 6h, relatively gained total alkaloids quality; Relatively concentration of alcohol is to the solid-liquid separation after leaching and the influence of slag content simultaneously, and its result is as shown in Figure 1.
Can know that by Fig. 1 under the same terms, leaching ability power is followed successively by 75%>80%>70%>85%>65%, therefore, select 75% ethanol water as leaching agent.
(2) time of contact is to the influence of alkaloid compound leaching effect
Get 6 parts of raw materials respectively, every part of 40.00g, each adds concentration is that 75% ethanol water 180.00ml leaches, and is respectively 1h time of contact, 2h, 3h, 4h, 5h, 6h, it is as shown in Figure 2 that it leaches the result.
Can know that by Fig. 2 under the room temperature (26 ℃), extraction time is 6h, leach reaction and reach balance basically; Prolong extraction time, leaching rate is constant basically.
(3) temperature is to the influence of alkaloid compound leaching effect
Get 6 parts of raw material powders respectively, every part of 40.00g, each adds 180.00ml concentration is 75% ethanol water, under different temperature, leaches, and extraction time is 2h, and sampling detects total alkaloids quality in the leachate, and the result is as shown in Figure 3.
Can know that by Fig. 3 along with the increase of extraction temperature, the leaching rate of alkaloids also increases accordingly; When temperature reaches 40 ℃, increase temperature, the leaching rate increase of alkaloids is tending towards slow.
2, four sections adverse current percolations (process diagram is seen Fig. 4)
6 ф 25.4mm * 1000mm glass have been installed have altogether been drawn together a mouthful pipe, as the infuser (leaching post) of four sections adverse current percolation leaching experiments, wherein four operations are done week for two and are migrated.The 150.00g Bulbus Fritillariae Thunbergii raw material powder particle (be called for short raw material, as follows) of packing in the every post, with dosing pump with leaching agent (65-85% ethanol water), with about 5mlmin
-1Flow squeeze in first leaching post, ethanol water is with about 1cmmin
-1The speed percolation after first post flows out the 450ml leachate, is connected first post and second post.The new leaching agent that dosing pump is squeezed into flows out entering second post through first post; With last identical, after second post flows out 450ml black leachate, second post and the 3rd post are connected.The rest may be inferred, after the 4th post flows out the 450ml effluent by the time, the 4th post and the 5th post connected.Simultaneously, the dosing pump outlet is moved to second post upper end from first post upper end and connect (first post is withdrawn from) from system with second post.At this moment, new leaching agent advances second post through dosing pump, behind three, four, five posts, flows out from the 5th post, gets the 450ml leachate equally; After this, new leaching agent advances the 3rd post, goes out leachate from the 6th post.The rest may be inferred, and constantly circulation is carried out four sections adverse current percolation leaching experiments to reaching the leaching balance.The 9th post begins to end to the 12 post in this test; These four effusive leachates of post are the leachate during approximate equilibrium, thus when new leaching agent from advancing, through ten, 11,12 posts from the 9th post; And after the following outflow of the 12 post 450ml black leachate, stop leaching test.With the effusive leachate sampling of nine, ten, 11,12 posts, the total alkaloids quality is measured in censorship.(annotate: cylinder is and recycles, and nine, ten, 11,12 only represent label, do not have practical significance)
3, four sections adverse current percolation leaching test results
Counter-current as above-mentioned makes an experiment.Every duplicate samples is 150.0g, is leachate with 75% ethanol, 40 ℃ of following lixiviate 6h, and liquid-solid ratio is 3.Get the effusive leachate of nine, ten, 11,12 posts and measure, its result is as shown in table 1.
Four sections adverse current percolation of table 1 leaching test result
Can know that from table 1 total alkaloid content is in 0.25% in the Bulbus Fritillariae Thunbergii, then its leaching rate of four sections adverse current percolation solvent extraction method (in leachate) reaches 94.03% (in total alkaloids).
Embodiment: following through specific embodiment, the present invention is made further detailed description.
Embodiment 1:
Get Bulbus Fritillariae Thunbergii medical material 150g, adding 4.5 times of amount mass fractions is 70% alcoholic solution, 30 ℃ of following countercurrent extraction 4h, filters the back and merges collection filtrating, concentrates; 1/4 times of raw medicinal herbs amount mass fraction of concentrated solution adding is 2% hydrochloric acid, fully stirs, and filters; Collect filtrating, residue use mass fraction is 2% hydrochloric acid solution washed twice, merges above-mentioned filtrating; It is 5% NaOH solution adjust pH to 7 that filtrating is used with mass fraction, and gained solution ethyl acetate extraction is collected ethyl acetate layer; The extract concentrating under reduced pressure gets the Bulbus Fritillariae Thunbergii total alkaloids extract, and drying obtains the dried cream 790.3mg of total alkaloids crude extract.
Embodiment 2:
Get Bulbus Fritillariae Thunbergii medical material 150g, adding 4.5 times of amount mass fractions is 75% alcoholic solution, 40 ℃ of following countercurrent extraction 6h, filters the back and merges collection filtrating, concentrates; 1/4 times of raw medicinal herbs amount mass fraction of concentrated solution adding is 2% hydrochloric acid, fully stirs, and filters; Collect filtrating, residue use mass fraction is 2% hydrochloric acid solution washed twice, merges above-mentioned filtrating; It is 5% NaOH solution adjust pH to 8 that filtrating is used with mass fraction, and gained solution ethyl acetate extraction is collected ethyl acetate layer; The extract concentrating under reduced pressure gets the Bulbus Fritillariae Thunbergii total alkaloids extract, and drying obtains the dried cream 784.5mg of total alkaloids crude extract.
Embodiment 3:
Get Bulbus Fritillariae Thunbergii medical material 150g, adding 4.5 times of amount mass fractions is 80% alcoholic solution, 50 ℃ of following countercurrent extraction 5h, filters the back and merges collection filtrating, concentrates; 1/4 times of raw medicinal herbs amount mass fraction of concentrated solution adding is 2% hydrochloric acid, fully stirs, and filters; Collect filtrating, residue use mass fraction is 2% hydrochloric acid solution washed twice, merges above-mentioned filtrating; It is 5% NaOH solution adjust pH to 9 that filtrating is used with mass fraction, and gained solution ethyl acetate extraction is collected ethyl acetate layer; The extract concentrating under reduced pressure gets the Bulbus Fritillariae Thunbergii total alkaloids extract, and drying obtains the dried cream 792.6mg of total alkaloids crude extract.
Assay
Get the dried cream 12mg of total alkaloids extract of the Bulbus Fritillariae Thunbergii of gained among the embodiment 1～3, be settled to 100mL with chloroform and use need testing solution as Determination of Total Alkaloid in the dried cream of total alkaloids extract.
Determination of Total Alkaloid method in the medical material: Thunberg Fritillary Bulb (crossing sieve No. 4) 2g decided in accurate title, behind ammonia 1mL moistening 15min, adds 20mL E-C-dehydrated alcohol (volume ratio is 25: 8: 2.5); Weigh, supersound process 40min weighs; Supply weight with the extraction solvent, 4000rmin
-1Centrifugal, the accurate supernatant 5mL that draws, evaporate to dryness, residue add the chloroform dissolving and are settled to 10mL, shake up, and use need testing solution as Determination of Total Alkaloid in the Bulbus Fritillariae Thunbergii medical material.
Precision takes by weighing peimine reference substance 5.52mg, adds the chloroform dissolving and is settled to 50mL, as reference substance solution.
Accurate draw in the dried cream of total alkaloids extract Determination of Total Alkaloid with Determination of Total Alkaloid in need testing solution 1.5mL, the Bulbus Fritillariae Thunbergii with need testing solution 1.5mL, reference substance solution 1mL, 2mL and blank solution chloroform; Place the 25mL volumetric flask respectively, the adding pH value is Potassium Hydrogen Phthalate-sodium hydrate buffer solution 5mL and 0.001molL of 5.0
-1Bromothymol blue test solution 2mL, chloroform is settled to scale, fully shakes up the back and moves in the separatory funnel; Leave standstill 40min; Obtain chloroform layer and move in the tool plug conical flask that is placed with 0.20g anhydrous sodium sulfate (in advance in 105 ℃ of heating 1h) in advance, airtight shaking up got supernatant and measured absorbance at the 416nm place; According to total alkaloid content in the standard curve calculating Bulbus Fritillariae Thunbergii medical material is 0.25%, calculates the total alkaloid content in the dried cream of total alkaloids extract according to following formula:
Total alkaloid content such as table 2 show in the dried cream of embodiment 1～3 gained total alkaloids extract.
Table 2 assay result
As stated, just can realize the present invention preferably.
Claims (4)
1. the method for preparing of total alkaloids extract in the Bulbus Fritillariae Thunbergii is characterized in that, may further comprise the steps successively:
Step (1): get Bulbus Fritillariae Thunbergii, pulverizing back adding mass fraction is the ethanol water of 65-85%, and 30-60 ℃ of following countercurrent extraction 4-6h merges to collect after filtering and filtrate, and concentrates, and gets concentrated solution;
Step (2): the adding mass fraction is 2% hydrochloric acid in the said concentrated solution, and abundant the stirring filtered, and collects and filtrates, and residue use mass fraction is 2% hydrochloric acid solution washed twice, merges above-mentioned filtrating;
Step (3): adopt potassium hydroxide alkalization step (2) the gained filtrating of mass fraction 5%, transfer pH value to 7.0-9.0, gained solution is with ethyl acetate extraction, and the extract concentrating under reduced pressure gets the Bulbus Fritillariae Thunbergii total alkaloids extract.
2. the method for preparing of total alkaloids extract is characterized in that in a kind of according to claim 1 Bulbus Fritillariae Thunbergii, and in said step (1), adding mass fraction is 75% ethanol water, 40 ℃ of following countercurrent extraction 6 hours.
3. the method for preparing of total alkaloids extract is characterized in that in a kind of according to claim 1 Bulbus Fritillariae Thunbergii, and in the said step (2), the mass fraction that adds 1/4 times of raw medicinal herbs amount in the said concentrated solution is 2% hydrochloric acid.
4. the method for preparing of total alkaloids extract is characterized in that in a kind of according to claim 1 Bulbus Fritillariae Thunbergii, in the said step (3), transfers pH value to 8.0.
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