CN110835348B - Extraction, separation and purification method of strychnine - Google Patents
Extraction, separation and purification method of strychnine Download PDFInfo
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Abstract
The invention relates to a method for extracting effective components of a traditional Chinese medicine, in particular to a method for extracting brucine from traditional Chinese medicine nux vomica, belonging to the technical field of medicines. The extraction rate of the strychnine is improved by increasing the infiltration of the alkali solution and adding the alkali into the silica gel chromatographic column eluent; the extraction process is eliminated, the use of an extracting agent is reduced, an organic solvent with high toxicity and pollution to the environment is not used in the process, and the safety is improved. The technical scheme of the invention can solve the defects in the prior art, obviously improve the quality (extraction rate and purity) of the product, reduce the cost, improve the safety and production feasibility and achieve the aim of the invention.
Description
Technical Field
The invention relates to a method for extracting effective components of a traditional Chinese medicine, in particular to a method for extracting brucine from traditional Chinese medicine nux vomica, belonging to the technical field of medicines.
Background
Strychnos nux-vomica, also called Strychnos nux-vomica, was recorded in Bencao gang mu, and it is bitter in taste, cold in nature, with strong toxicity, enters liver and spleen meridians, and has the actions of activating meridians to stop pain, dissipating nodulation and relieving swelling. Is clinically used for treating diseases such as rheumatism obstinate arthralgia, numbness and paralysis, traumatic injury, carbuncle, cellulitis, gall, sequela of poliomyelitis, rheumatoid arthritis and the like; however, the toxic herbs-pungent herbs have strong toxic properties, which seriously affect the clinical application of the herbs. The treatment amount of the nux vomica is 0.3-0.6g, excessive dose is easy to cause poisoning, and how to reasonably and safely use the nux vomica becomes one of the research hotspots in recent years.
Researches show that the total alkaloids are the most main effective parts in the nux vomica, and comprise strychnine, brucine, strychnine, loganin, etc., wherein the strychnine in the main alkaloid components has the functions of anti-inflammation and analgesia, the strychnine is a central nerve stimulant and is mainly used for clinically treating facial paralysis, nerve palsy, etc., and is also a main toxic component, and the anti-inflammation and analgesia effects are basically avoided; the LD50 of the strychnine, the strychnine and the nux vomica kernel which are infused into the mouse are respectively 3.27mg/kg, 233mg/kg and 234.5mg/kg in the literature; LD50 of the medicine injected into the abdominal cavity of a mouse is 1.53mg/kg, 69.77mg/kg and 76mg/kg respectively, the oral poisoning dose is 5-10 mg, and the oral lethal dose is 30 mg. Strychnine is the main toxic component in nux vomica, has low safety, and if strychnine in the nux vomica total alkaloids can be removed, the toxicity can be reduced, the original pharmacological activity of anti-inflammation and analgesia can be kept, and the safety of clinical medication is improved. Therefore, a preparation method for improving the purity of strychnine is needed in modern research.
Chinese patent publication No. CN102432618B discloses a process for separating and purifying strychnine from total alkaloids of semen Strychni, the extracted strychnine has a purity of more than 94% and a recovery rate of more than 43%; the Chinese patent application with publication number CN106632367A discloses an extraction method for extracting strychnine from Fengshi Maqian tablets, the purity of the strychnine after extraction is 75% at most, and the recovery rate of the strychnine is 94%; the Chinese patent application with the publication number of CN107281267A discloses a method for extracting and separating nux vomica total alkaloid from nux vomica, and the maximum content of strychnine in the extracted nux vomica total alkaloid is 75 percent; the Chinese patent application with the publication number of CN108117557A discloses a method for extracting, separating and purifying strychnine, and the purity of the strychnine after extraction is more than 95%.
However, the above extraction methods have many problems, such as complex process, low yield, low purity of nux vomica, high energy consumption, use of organic solvents with high toxicity and high volatility, environmental pollution, residual toxic organic solvents, and unsuitability for industrial production.
The existing technology reports the preparation process of the strychnine with high purity and high extraction rate, and particularly, the preparation method of the strychnine with high purity and high extraction rate, which is suitable for industrial production scale, is not common.
Disclosure of Invention
The invention aims to provide a method for extracting strychnine from nux vomica with low toxicity, low cost and high efficiency.
The invention is realized by the following technical scheme:
a method for extracting, separating and purifying strychnine comprises the following steps:
step 1: soaking nux vomica serving as a raw material in an alkali solution, and extracting with ethyl acetate serving as a solvent to obtain an extracting solution A;
and 2, step: taking ethanol added with alkaline water as eluent, and separating the extract A by a silica gel chromatographic column to obtain a dry isolate B;
and step 3: dissolving the dried isolate B in acid to obtain filtrate C, and performing alkali precipitation to obtain dried powder D;
and 4, step 4: recrystallizing the dried powder D with ethanol to obtain strychnine.
Further, as a preferred embodiment, in the step 1, the base is an inorganic base, and more preferably, the base is one or a combination of two or more of sodium carbonate, potassium carbonate, sodium bicarbonate, potassium bicarbonate, sodium hydroxide, potassium hydroxide and ammonia water.
Further, as a preferred embodiment, in the step 1, the base is an organic base, more preferably, the base is one, a combination of two or more of amines, alkaloids, potassium tert-butoxide, sodium tert-butoxide and n-butyllithium, and more preferably, the amines are methylamine, ethylamine, urea, propylamine and butylamine. Further, as a preferred embodiment, in the step 1, the volume dosage of the alkali solution is 0.5-3 times of the weight of the medicinal materials, and the volume/weight ratio is: ml/g, the infiltration time is 2-4h, more preferably, the volume dosage of the alkali solution is 0.5-1.5 times of the weight of the medicinal materials, and the volume/weight is as follows: ml/g, the infiltration time is 3 h.
Further, as a preferred embodiment, the volume dosage of the ethyl acetate in the step 1 is 4-6 times of the weight of the medicinal materials, and the volume/weight ratio is: ml/g.
Further, as a preferred embodiment, the ethyl acetate is extracted under reflux in the step 1 for 1 to 4 times each for 6 to 12 hours, and more preferably, the ethyl acetate is extracted under reflux for 2 times each for 8 to 10 hours.
Further, as a preferred embodiment, the extract of step 1 is concentrated under reduced pressure to 1/3-1 of the volume of the original extract;
further, as a preferred embodiment, the alkaline water in step 2 is one or a combination of two or more of sodium carbonate solution, sodium bicarbonate solution, potassium carbonate solution, potassium bicarbonate and ammonia water.
Further, as a preferred embodiment, the concentration of the alkaline water in the step 2 is 0.1% to 2%, and more preferably, the concentration of the alkaline water is 0.5% to 1%.
Further, as a preferred embodiment, after the extract a in step 2 is separated by a silica gel chromatography column, the elution solution is collected after 5 times of the column volume.
Further, as a preferred embodiment, in the step 3, the acid is an inorganic acid, and more preferably one, two or more of hydrochloric acid, sulfuric acid and phosphoric acid.
Further, as a preferred embodiment, in the step 3, the concentration of the acid is 1% to 3%, and the volume of the acid solution is used in an amount of 20 to 50 times the weight of the dried isolate B, volume/weight: ml/g, more preferably, the acid concentration is 1% to 1.5%, the acid solution is used in an amount of 20 to 35 times by volume the weight of dry isolate B, v/w: ml/g.
Further, as a preferable embodiment, in the step 3, the base is an inorganic base, and more preferably, the base is one or a combination of two or more of sodium hydroxide solution, potassium hydroxide solution, calcium hydroxide solution, sodium carbonate solution, potassium carbonate solution, sodium bicarbonate solution and potassium bicarbonate.
Further, as a preferred embodiment, in the step 3, the pH of the filtrate C is adjusted to 8 to 13, more preferably, to 10 to 12, with the base.
Further, as a preferred embodiment, in the step 4, the dried powder D is recrystallized with a 25 to 95% ethanol solution, and more preferably, with a 25% to 35% ethanol solution.
Further, as a preferred embodiment, in the step 4, the volume usage amount of ethanol is 10 to 30 times the weight of the dry powder D, and the volume/weight ratio is: ml/g, more preferably, ethanol is used in a volume of 15 to 25 times the weight of the dry powder D, volume/weight: ml/g.
In order to comprehensively investigate the influence factors, the main factors influencing the extraction effect in the purification method are investigated, and the optimal conditions of the extraction process are determined.
The measuring method comprises the following steps: the content of strychnine obtained was measured by the following method.
The final products obtained in the embodiments of the invention are all used for measuring the content of strychnine by high performance liquid chromatography, and the chromatographic conditions are as follows: column C18(4.6 mm. times.250 mm, 5 μm) as mobile phase acetonitrile-water phase (0.01mol/L sodium heptanesulfonate mixed with 0.02mol/L potassium dihydrogen phosphate in equal amounts, pH adjusted to 2.8 with 10% phosphoric acid) (21:79), wavelength 260nm, column temperature 40 deg.C, flow rate 1.0 ml/min.
Second, Experimental example
Experimental example 1: investigation of the alkaline solution in step 1
In the test, the nux vomica medicinal materials are respectively soaked by a sodium carbonate solution, a potassium carbonate solution, a sodium bicarbonate solution, a potassium bicarbonate solution, a sodium hydroxide solution, a potassium hydroxide solution, ammonia water, a methylamine solution, an ethylamine solution and a potassium tert-butoxide solution, and different alkali solutions are investigated.
The specific experimental method is as follows:
taking 13 parts of 50g of nux vomica medicinal material, crushing, adding one or two or more of sodium carbonate solution, potassium carbonate solution, sodium bicarbonate solution, potassium bicarbonate solution, sodium hydroxide solution, potassium hydroxide solution, ammonia water, methylamine solution, ethylamine solution and potassium tert-butoxide solution for infiltration, adding ethyl acetate for reflux extraction, filtering, combining filtrates, concentrating and drying the extracting solution A to obtain a dried substance E, weighing, measuring the content of strychnine in the dried substance E, and calculating the purity and the extraction rate of strychnine (see table 1).
The extraction rate of strychnine (the content of strychnine in the dry matter E is multiplied by the weight of the concentrated dry matter E)/(the content of strychnine in the medicinal materials is multiplied by the weight of the strychnos nux-vomica medicinal materials) multiplied by 100 percent
TABLE 1 Experimental results of brucine extraction rate by alkali solution
Table 1 shows that the extraction rate of the strychnine can be more than 90% by one, two or more than two of sodium carbonate solution, potassium carbonate solution, sodium bicarbonate solution, potassium bicarbonate solution, sodium hydroxide solution, potassium hydroxide solution, ammonia water, methylamine solution, ethylamine solution and potassium tert-butoxide solution, and the purity of the strychnine is more than 20%.
Experimental example 2: step 2 Process study
The addition of alkaline water to the eluent from step 2 is preferred.
The specific experimental method is as follows:
taking 50g of the extracting solution A obtained after the strychnos nux-vomica medicinal material is subjected to the step 1, evenly dividing the extracting solution A into 14 parts, respectively passing through silica gel chromatographic columns, eluting by adopting different eluents, collecting an eluting solution with the volume being 5 times of the column volume, concentrating, drying and crushing to obtain a dry isolate B, weighing, measuring the strychnos nux-vomica alkali content in the dry isolate B, and calculating the purity and the extraction rate of the strychnos nux-vomica alkali.
1.1 examination of the alkaline Water in the eluate
In the experiment, 6 parts of 14 parts of the extracting solution of 50g of nux vomica medicinal material after the step 1 are respectively taken, ethanol added with 0.75% sodium carbonate solution, ethanol added with 0.75% sodium bicarbonate solution, ethanol added with 0.75% potassium carbonate solution, ethanol added with 0.75% potassium bicarbonate solution and ethanol added with 0.75% ammonia water are respectively used as eluent, the eluent is concentrated, dried and crushed to obtain a dried isolate B, and different eluents are examined by taking the purity and the extraction rate of the strychnine in the dried isolate B as examination indexes (see table 2).
The extraction rate of strychnine (the content of strychnine in the dry isolate B is multiplied by the weight of the dry isolate B)/(the content of strychnine in the medicinal materials is multiplied by the weight of the strychnos nux-vomica medicinal materials) is multiplied by 100 percent
TABLE 2 Experimental results of the elution solution on the extraction rate of strychnine
The experimental result shows that the extraction rate of the strychnine can be improved by adding 0.75% of sodium carbonate solution, sodium bicarbonate solution, potassium carbonate solution, potassium bicarbonate and ethanol eluent of ammonia water, the purity of the strychnine can be enabled to be more than 43%, and the extraction rate of the strychnine is more than 80%.
1.2 examination of the concentration of alkali in the eluate
In the experiment, 9 parts of 14 parts of the extracting solution of 50g of the nux vomica medicinal material after the step 1 are respectively taken as eluent, ethanol added with 0.1% of ammonia water, ethanol added with 0.25% of ammonia water, ethanol added with 0.5% of ammonia water, ethanol added with 0.75% of ammonia water, ethanol added with 1% of ammonia water, ethanol added with 1.5% of ammonia water, ethanol added with 1.75% of ammonia water and ethanol added with 2% of ammonia water, concentrated, dried and crushed to obtain a dried isolate B, and the eluent added with different concentrations of alkaline water is considered by taking the purity and the extraction rate of the nux vomica alkali in the dried isolate B as the indexes (see table 3).
The extraction rate of strychnine (the content of strychnine in the dry isolate B is multiplied by the weight of the dry isolate B)/(the content of strychnine in the medicinal materials is multiplied by the weight of the strychnos nux-vomica medicinal materials) is multiplied by 100 percent
TABLE 3 concentration of elution solution versus strychnine extraction yield
| Eluent | Brucine purity (%) | Brucine extraction rate (%) |
| Ethanol | 21.14 | 42.47 |
| Ethanol + 0.1% ammonia water | 43.22 | 70.84 |
| Ethanol + 0.25% ammonia | 43.47 | 79.65 |
| Ethanol + 0.5% ammonia water | 44.67 | 82.64 |
| Ethanol + 0.75% ammonia | 43.99 | 82.87 |
| Ethanol + 1% ammonia water | 44.67 | 81.97 |
| Ethanol + 1.5% ammonia | 44.24 | 79.41 |
| Ethanol + 1.75% ammonia | 44.96 | 78.25 |
| Ethanol + 2% ammonia water | 44.55 | 77.47 |
The experimental result shows that the extraction rate of the strychnine can be improved by adding the ethanol eluent with 0.1-2% of ammonia water, the purity of the strychnine can be more than 43%, and further, the extraction rate of the strychnine can be more than 80% by adding the ethanol eluent with 0.5-1% of ammonia water.
Experimental example 3: pH value study in step 3
In the test, sodium hydroxide solution is added to adjust the pH of the filtrate to 8, 9, 10, 11, 12 and 13, and different pH values are examined by taking the strychnine content and the extraction rate in the dry powder D as examination indexes.
The brucine extraction rate (brucine content in dry powder D multiplied by dry powder D weight)/(brucine content in medicinal material multiplied by nux vomica medicinal material weight) multiplied by 100%
The specific experimental method is as follows:
taking 50g of nux vomica, carrying out step 1, loading an extracting solution A into a silica gel chromatographic column, adopting 6 parts of an eluent eluted by ethanol added with 0.75 ammonia water, concentrating and drying to obtain 6 parts of a dried isolate B, respectively dissolving the dried isolate B by using 1% hydrochloric acid, filtering to obtain a filtrate C, respectively adjusting the pH to 8, 9, 10, 11, 12 and 13 by using a sodium hydroxide solution, standing, filtering, drying a filter cake, crushing to obtain a dried powder D, weighing, measuring the content of strychnine in the dried powder D, and calculating the purity and the extraction rate of the strychnine. The results are shown in Table 4:
TABLE 4 experimental results of pH values for strychnine
| pH value | Brucine purity (%) | Brucine extraction ratio (%) |
| 8 | 83.22 | 63.67 |
| 9 | 83.27 | 65.99 |
| 10 | 88.68 | 70.43 |
| 11 | 88.70 | 70.47 |
| 12 | 88.47 | 71.51 |
| 13 | 85.11 | 68.19 |
Table 4 shows that the extraction rate of brucine can reach more than 60% when the pH value is 8-13, the purity of brucine can reach more than 80%, furthermore, the extraction rate of brucine can reach more than 70% when the pH value is 10-12, the purity of brucine is more than 88%, and the preferred pH value is 10-12.
Experimental example 4: inspection of total extraction rate of strychnine extraction, separation and purification process
The total extraction rate of the strychnine is investigated according to a method for extracting, separating and purifying strychnine determined on the basis of the research of a series of process conditions.
Taking 50g of strychnos nux-vomica medicinal material, carrying out step 1, loading an extracting solution A into a silica gel chromatographic column, eluting 8 parts of an eluent by adopting ethanol added with 0.1% -2% ammonia water, concentrating, drying and crushing to obtain a dried isolate B, dissolving 8 parts of the dried isolate B respectively by using 1% hydrochloric acid, filtering to obtain a filtrate C, adjusting the pH of the filtrate C to 12 by using a sodium hydroxide solution, standing, filtering, drying a filter cake, crushing to obtain a dried powder D, adding 30 times of 30% ethanol into the dried powder D, heating and dissolving, filtering, crystallizing the filtrate, filtering, drying the filter cake, crushing to obtain strychnos nux-vomica alkali, and calculating the purity and the extraction rate of the strychnos nux-vomica alkali (see table 5).
The total extraction rate of strychnine (the content of strychnine in the final product is multiplied by the weight of the final product)/(the content of strychnine in the medicinal materials is multiplied by the weight of the strychnos nux-vomica medicinal materials) multiplied by 100 percent
Table 5 Total extraction Rate investigation
| Ammonia concentration (g) | Brucine purity (%) | Total extraction ratio (%) |
| 0.1％ | 95.6 | 50.1 |
| 0.25％ | 96.8 | 51.3 |
| 0.5％ | 97.3 | 52.7 |
| 0.75％ | 97.5 | 52.8 |
| 1％ | 97.0 | 51.9 |
| 1.5％ | 96.5 | 50.8 |
| 1.75％ | 95.2 | 50.7 |
| 2％ | 95.2 | 50.5 |
And (4) conclusion: through comprehensive verification of the nux vomica extraction process, the purity of the prepared nux vomica alkaloid is over 95 percent, and the total extraction rate is over 50 percent.
The applicant of the patent application with the patent publication number of CN108117557A repeatedly tests the strychnine with the purity of more than 95 percent and the extraction rate of about 23 percent. Compared with the invention, the invention provides the method for extracting and purifying the strychnine with high extraction rate and high purity.
Compared with the prior art, the extraction rate of the strychnine is improved by increasing the infiltration of the alkali solution and adding the alkali into the silica gel chromatographic column eluent; the extraction process is eliminated, the use of an extracting agent is reduced, an organic solvent with high toxicity and pollution to the environment is not used in the process, and the safety is improved.
Therefore, the technical scheme of the invention can solve the defects in the prior art, remarkably improve the quality (extraction rate and purity) of the product, reduce the cost, improve the safety and the production feasibility and achieve the aim of the invention.
Detailed Description
The present invention will be described in further detail with reference to the following examples. It should not be understood that the scope of the above-described subject matter of the present invention is limited to the following examples. All the technologies realized based on the above contents of the present invention belong to the scope of the present invention.
Example 1
Step 1: taking 50g of nux vomica medicinal material, crushing, adding 3 times of potassium carbonate solution for soaking for 4h, adding 5 times of ethyl acetate for reflux extraction for 3 times, 10h each time, filtering, combining filtrates, concentrating under reduced pressure to 1/3 of the volume of the original extract, and filtering to obtain an extract A;
step 2: loading the extract A into a silica gel chromatographic column, eluting with ethanol (added with 0.1% ammonia water solution), collecting the eluate with 5 times of column volume, concentrating and drying the eluate, and pulverizing to obtain dry isolate B;
and 3, step 3: adding 40 times of 2% sulfuric acid into the dried isolate B, heating for dissolving, filtering to obtain filtrate C, adding sodium hydroxide solution into the filtrate C to adjust the pH value of the filtrate to 11, standing at room temperature, filtering, drying the filter cake, and pulverizing to obtain dry powder D;
and 4, step 4: and adding 20 times of 35% ethanol into the dried powder D, heating for dissolving, filtering, crystallizing the filtrate, filtering, drying the filter cake, and crushing to obtain 0.2096g of strychnine with the purity of about 95.8% and the total extraction rate of about 50.2%.
Example 2
Step 1: crushing 50g of nux vomica medicinal material, adding 1.5 times of sodium bicarbonate solution for soaking for 3h, adding 6 times of ethyl acetate for reflux extraction for 2 times, each time for 9h, filtering, combining filtrates, concentrating under reduced pressure to 1/3 of the volume of the original extract, and filtering to obtain extract A;
step 2: loading the extract A into a silica gel chromatographic column, eluting with ethanol (added with 0.25% potassium carbonate solution), collecting the eluate with 5 times of column volume, concentrating and drying the eluate, and pulverizing to obtain dry isolate B;
and step 3: adding 1% phosphoric acid in an amount which is 30 times that of the dried isolate B, heating and dissolving, filtering to obtain a filtrate C, adding a potassium hydroxide solution into the filtrate C to adjust the pH value of the filtrate to 9, standing at room temperature, filtering, drying a filter cake, and crushing to obtain a dry powder D;
and 4, step 4: adding 20 times of 30% ethanol into the dry powder D, heating for dissolving, filtering, crystallizing the filtrate, filtering, drying the filter cake, and pulverizing to obtain 0.2119g of strychnine with purity of about 96.8% and total extraction rate of about 51.3%.
Example 3
Step 1: taking 50g of nux vomica medicinal material, crushing, adding 1 time of sodium carbonate solution for soaking for 2 hours, adding 5 times of ethyl acetate for reflux extraction for 2 times, 10 hours each time, filtering, combining filtrates, concentrating under reduced pressure to 1/3 of the volume of the original extract, filtering to obtain an extract A;
step 2: loading the obtained extract A into a silica gel chromatographic column, eluting with ethanol (added with 0.5% sodium carbonate solution), collecting the eluate with 5 times of column volume, concentrating and drying the eluate, and pulverizing to obtain dry isolate B;
and step 3: adding 30 times of 1.5% sulfuric acid into the dried isolate B, heating for dissolving, filtering to obtain filtrate C, adding sodium hydroxide solution into the filtrate C to adjust the pH value of the filtrate to 10, standing at room temperature, filtering, drying the filter cake, and pulverizing to obtain dry powder D;
and 4, step 4: and adding 20 times of 25% ethanol into the dried powder D, heating for dissolving, filtering, crystallizing the filtrate, filtering, drying the filter cake, and crushing to obtain 0.2177g of strychnine with the purity of about 97.2% and the total extraction rate of about 52.9%.
Example 4
Step 1: taking 50g of nux vomica medicinal material, crushing, adding 1.5 times of sodium carbonate solution for soaking for 3h, adding 5 times of ethyl acetate for reflux extraction for 2 times, each time for 8h, filtering, combining filtrates, concentrating under reduced pressure to 1/3 of the volume of the original extract, filtering to obtain extract A;
step 2: loading the extract A into a silica gel chromatographic column, eluting with ethanol (added with 0.5% ammonia water solution), collecting eluate with 5 times of column volume, concentrating and drying the eluate, and pulverizing to obtain dry isolate B;
and step 3: adding 1% hydrochloric acid in an amount which is 30 times that of the dried isolate B, heating and dissolving, filtering to obtain a filtrate C, adding a sodium hydroxide solution into the filtrate C to adjust the pH value of the filtrate to 12, standing at room temperature, filtering, drying a filter cake, and crushing to obtain a dry powder D;
and 4, step 4: adding 30 times of 30% ethanol into the dry powder D, heating for dissolving, filtering, crystallizing the filtrate, filtering, drying the filter cake, and pulverizing to obtain 0.2173g of strychnine with purity of about 97.2% and total extraction rate of about 52.8%.
Example 5
Step 1: taking 50g of nux vomica medicinal material, crushing, adding 1 time of methylamine solution for soaking for 3h, adding 5 times of ethyl acetate for reflux extraction for 2 times, each time for 9h, filtering, combining filtrates, concentrating under reduced pressure to 1/3 of the volume of the original extract, filtering to obtain extract A;
step 2: loading the extractive solution A into silica gel chromatographic column, eluting with ethanol (added with 0.75% sodium bicarbonate solution), collecting eluate 5 times of the column volume, concentrating and drying the eluate, and pulverizing to obtain dry isolate B;
and step 3: adding 35 times of 1% sulfuric acid into the dried isolate B, heating for dissolving, filtering to obtain filtrate C, adding potassium hydroxide solution into the filtrate C to adjust the pH value of the filtrate to 10, standing at room temperature, filtering, drying the filter cake, and pulverizing to obtain dry powder D;
and 4, step 4: adding 20 times of 30% ethanol into the dried powder D, heating for dissolving, filtering, crystallizing the filtrate, filtering, drying the filter cake, and pulverizing to obtain 0.2164g of strychnine with purity of about 97.4% and total extraction rate of about 52.7%.
Example 6
Step 1: crushing 50g of nux vomica medicinal material, adding 1 time of sodium bicarbonate solution for soaking for 4h, adding 5 times of ethyl acetate for reflux extraction for 2 times, each time for 9h, filtering, combining filtrates, concentrating under reduced pressure to 1/3 of the volume of the original extract, and filtering to obtain extract A;
step 2: loading the extract A into a silica gel chromatographic column, eluting with ethanol (added with 0.75% ammonia water solution), collecting the eluate with 5 times of column volume, concentrating and drying the eluate, and pulverizing to obtain dry isolate B;
and step 3: adding 35 times of 1% hydrochloric acid into the dried isolate B, heating for dissolving, filtering to obtain filtrate C, adding sodium hydroxide solution into the filtrate C to adjust the pH value of the filtrate to 10, standing at room temperature, filtering, drying the filter cake, and pulverizing to obtain dry powder D;
and 4, step 4: adding 20 times of 30% ethanol into the dried powder D, heating for dissolving, filtering, crystallizing the filtrate, filtering, drying the filter cake, and pulverizing to obtain 0.2166g of strychnine with purity of about 97.5% and total extraction rate of about 52.8%.
Example 7
Step 1: taking 50g of nux vomica medicinal material, crushing, adding 2 times of potassium bicarbonate solution for soaking for 4h, adding 4 times of ethyl acetate for reflux extraction for 3 times, each time for 9h, filtering, combining filtrates, concentrating under reduced pressure to 1/3 of the volume of the original extract, filtering to obtain extract A;
step 2: loading the extract A into a silica gel chromatographic column, eluting with ethanol (added with 1% ammonia water solution), collecting the eluate with 5 times of column volume, concentrating and drying the eluate, and pulverizing to obtain dry isolate B;
and 3, step 3: adding 20 times of 2% sulfuric acid into the dried isolate B, heating for dissolving, filtering to obtain filtrate C, adding sodium hydroxide solution into the filtrate C to adjust the pH value of the filtrate to 12, standing at room temperature, filtering, drying the filter cake, and pulverizing to obtain dry powder D;
and 4, step 4: adding 30 times of 20% ethanol into the dried powder D, heating for dissolving, filtering, crystallizing the filtrate, filtering, drying the filter cake, and pulverizing to obtain 0.2132g of strychnine with purity of about 97.0% and total extraction rate of about 51.7%.
Example 8
Step 1: taking 50g of nux vomica medicinal material, crushing, adding 3 times of ethylamine solution for soaking for 3h, adding 5 times of ethyl acetate for reflux extraction for 2 times, 10h each time, filtering, combining filtrates, concentrating under reduced pressure to 1/3 of the volume of the original extract, filtering to obtain extract A;
step 2: loading the extractive solution A into silica gel chromatographic column, eluting with ethanol (added with 1.5% sodium carbonate solution), collecting eluate 5 times of the column volume, concentrating and drying the eluate, and pulverizing to obtain dry isolate B;
and 3, step 3: adding 1.5% hydrochloric acid in an amount which is 30 times that of the dried isolate B, heating and dissolving, filtering to obtain a filtrate C, adding a potassium hydroxide solution into the filtrate C to adjust the pH value of the filtrate to 11, standing at room temperature, filtering, drying a filter cake, and crushing to obtain a dry powder D;
and 4, step 4: adding 30 times of 30% ethanol into the dry powder D, heating for dissolving, filtering, crystallizing the filtrate, filtering, drying the filter cake, and pulverizing to obtain 0.2103g of strychnine with purity of about 96.4% and total extraction rate of about 50.7%.
Example 9
Step 1: taking 50g of nux vomica medicinal material, crushing, adding 2 times of potassium carbonate solution for soaking for 2h, adding 4 times of ethyl acetate for reflux extraction for 2 times, each time for 9h, filtering, combining filtrates, concentrating under reduced pressure to 1/3 of the volume of the original extract, filtering to obtain extract A;
step 2: loading the extract A into a silica gel chromatographic column, eluting with ethanol (added with 1.75% ammonia water), collecting the eluate with 5 times of column volume, concentrating and drying the eluate, and pulverizing to obtain dry isolate B;
and 3, step 3: adding 1% hydrochloric acid 20 times the amount of the dried isolate B, heating to dissolve, filtering to obtain filtrate C, adding sodium hydroxide solution into the filtrate C to adjust the pH value of the filtrate to 11, standing at room temperature, filtering, drying the filter cake, and pulverizing to obtain dry powder D;
and 4, step 4: adding 25 times of 25% ethanol into the dried powder D, heating for dissolving, filtering, crystallizing the filtrate, filtering, drying the filter cake, and pulverizing to obtain 0.2128g of strychnine with purity of 95.3% and total extraction rate of 50.7%.
Example 10
Step 1: taking 50g of nux vomica medicinal material, crushing, adding 1 time of sodium carbonate solution for soaking for 3 hours, adding 5 times of ethyl acetate for reflux extraction for 2 times, each time for 9 hours, filtering, combining filtrates, concentrating under reduced pressure to 1/3 of the volume of the original extract, filtering to obtain an extract A;
step 2: loading the extract A into a silica gel chromatographic column, eluting with ethanol (added with 2% potassium carbonate), collecting an elution solution with the volume 5 times that of the column, concentrating and drying the eluent, and crushing to obtain a dry isolate B;
and step 3: adding 1% hydrochloric acid in an amount which is 30 times that of the dried isolate B, heating and dissolving, filtering to obtain a filtrate C, adding a sodium hydroxide solution into the filtrate C to adjust the pH value of the filtrate to 13, standing at room temperature, filtering, drying a filter cake, and crushing to obtain a dry powder D;
and 4, step 4: adding 30 times of 30% ethanol into the dry powder D, heating for dissolving, filtering, crystallizing the filtrate, filtering, drying the filter cake, and pulverizing to obtain 0.2115g of strychnine with purity of about 95.3% and total extraction rate of about 50.4%.
Claims (13)
1. A method for extracting, separating and purifying strychnine comprises the following steps:
step 1: soaking nux vomica serving as a raw material in an alkali solution, and extracting by using ethyl acetate as a solvent to obtain an extracting solution A;
step 2: taking ethanol added with alkaline water as eluent, and separating the extract A by a silica gel chromatographic column to obtain a dry isolate B;
and step 3: dissolving the dried isolate B in acid to obtain filtrate C, and performing alkali precipitation to obtain dried powder D;
and 4, step 4: recrystallizing the dry powder D with ethanol to obtain strychnine;
the alkaline water in the step 2 is one or a combination of two or more of sodium carbonate solution, sodium bicarbonate solution, potassium carbonate solution, potassium bicarbonate solution and ammonia water;
the concentration of the alkaline water in the step 2 is 0.1-2%.
2. The extraction, separation and purification method according to claim 1, wherein in the step 1, the base is an inorganic base.
3. The extraction separation method according to claim 2, wherein in step 1, the alkali is one or a combination of two or more of sodium carbonate, potassium carbonate, sodium bicarbonate, potassium bicarbonate, sodium hydroxide, potassium hydroxide, and ammonia water.
4. The extraction, separation and purification method according to claim 1, wherein the base is an organic base in the step 1.
5. The extraction, separation and purification method according to claim 4, wherein in the step 1, the base is one or a combination of two or more of amines, alkaloids, potassium tert-butoxide, sodium tert-butoxide and n-butyllithium.
6. The extraction, separation and purification method according to any one of claims 1 to 5, wherein in the step 1, the volume consumption of the alkali solution is 0.5 to 3 times of the weight of the medicinal materials, and the volume/weight ratio is as follows: ml/g, the infiltration time is 2-4 h.
7. The extraction, separation and purification method according to claim 6, wherein in the step 1, the volume consumption of the alkali solution is 0.5-1.5 times of the weight of the medicinal materials, and the volume/weight ratio is as follows: ml/g, the infiltration time is 3 h.
8. The extraction, separation and purification method according to claim 1, wherein the volume dosage of the ethyl acetate in the step 1 is 4-6 times of the weight of the medicinal materials, and the volume/weight ratio is as follows: ml/g.
9. The extraction, separation and purification method according to claim 1, wherein the ethyl acetate is extracted under reflux in step 1 for 6-12h for 1-4 times.
10. The extraction, separation and purification method according to claim 9, wherein the ethyl acetate is extracted under reflux for 8-10h for 2 times in step 1.
11. The extraction, separation and purification method according to claim 1, wherein the extract of step 1 is concentrated under reduced pressure to 1/3-1 of the volume of the original extract.
12. The extraction, separation and purification method according to claim 1, wherein the concentration of the alkaline water in the step 2 is 0.5-1%.
13. The extraction, separation and purification method according to claim 1, wherein the extraction solution A in the step 2 is separated by a silica gel chromatography column, and then an elution solution is collected after 5 times of the column volume.
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