阅读说明：本技术 一种高抑菌活性平卧菊三七生物碱的提取方法 (Method for extracting gynura procumbens alkaloid with high antibacterial activity ) 是由 李安平 余江帆 唐玉妹 王乐 李刚 于 2020-04-13 设计创作，主要内容包括：一种高抑菌活性平卧菊三七生物碱的提取方法,包括以下步骤：(1)粉碎过筛；(2)超临界CO-2萃取；(3)分离；(4)真空干燥；(5)高压结晶；(6)洗涤干燥；(7)电磁场结晶；(8)过滤；(9)洗涤干燥。本发明先用超临界CO-2萃取和大孔吸附树脂分离,然后通过高压结晶和电磁场结晶获得纯度≥95%的平卧菊三七生物碱晶体。本发明所提取到的平卧菊三七生物碱抑菌活性较强,对病毒RSV和HSV-1均有较好的抑制作用,其效果均接近黄连碱晶体,能广泛应用于食品和药品领域,市场前景广阔。(A method for extracting Gynura procumbens alkaloid with high antibacterial activity comprises the following steps: (1) pulverizing and sieving; (2) supercritical CO 2 Extracting; (3) separating; (4) vacuum drying; (5) high-pressure crystallization; (6) washing and drying; (7) crystallizing by an electromagnetic field; (8) filtering; (9) and (5) washing and drying. The invention uses supercritical CO firstly 2 Extracting, separating with macroporous adsorbent resin, and crystallizing under high pressure and electromagnetic field to obtain Gynura procumbens alkaloid crystal with purity of 95% or more. The gynura procumbens alkaloid extracted by the invention has stronger antibacterial activity, has better inhibiting effect on viruses RSV and HSV-1, has the effect similar to that of coptisine crystals, can be widely applied to the fields of food and medicine, and has wide market prospect.)

1. A method for extracting gynura procumbens alkaloid with high bacteriostatic activity is characterized by comprising the following steps:

(1) crushing and sieving: cleaning whole plant of Gynura procumbens, removing impurities, draining, drying, and pulverizing to obtain Gynura procumbens whole plant powder;

(2) supercritical CO₂And (3) extraction: placing the gynura procumbens powder obtained in the step (1) into an extraction kettle, then respectively preheating the extraction kettle, a separation kettle and a storage tank, pressurizing the system when the set temperature is reached, adding absolute ethyl alcohol as an entrainer, performing circular extraction, and obtaining an extracting solution through the separation kettle;

(3) separation: adsorbing the extracting solution obtained in the step (2) by using macroporous adsorption resin, eluting and collecting to obtain eluent;

(4) and (3) vacuum drying: vacuum drying the eluent obtained in the step (3) to obtain total alkaloid dry matter;

(5) high-pressure crystallization: adding absolute ethyl alcohol into the total alkaloid dry matter obtained in the step (4), dissolving to form a total alkaloid saturated solution, pouring the total alkaloid saturated solution into a pressure container, pressurizing, fully cooling and crystallizing, and then relieving pressure to obtain a high-pressure crystallization mixed solution;

(6) washing and drying: filtering the high-pressure crystallization mixed solution obtained in the step (5) by using a filter screen, washing and filtering the obtained crystal by using anhydrous glacial ethanol, and then drying the crystal in vacuum to constant weight to obtain a crude crystal;

(7) electromagnetic field crystallization: putting the coarse crystal obtained in the step (6) into a container, dissolving the coarse crystal with a solvent, then putting the container into an electromagnetic field for vacuum concentration and crystallization, cooling to normal temperature after crystallization, and destroying the vacuum degree to obtain a crystal mixed solution;

(8) and (3) filtering: filtering the crystal mixed solution obtained in the step (7) by using filter cloth to obtain crystals;

(9) washing and drying: and (4) washing the crystal obtained in the step (8) with anhydrous glacial ethanol, and then carrying out vacuum drying to obtain the gynura procumbens alkaloid crystal with high purity.

2. The method for extracting gynura procumbens alkaloids with high antibacterial activity according to claim 1, wherein in the step (1), the drying temperature is 65-75 ℃, the drying is carried out until the moisture content is 8-14%, and the mesh number of the sieving mesh is preferably 60 meshes.

3. The method for extracting gynura procumbens alkaloids with high antibacterial activity as claimed in claim 1 or 2, wherein in step (2), supercritical CO in the extraction kettle₂The extraction conditions were: the extraction temperature is 45-55 ℃, the extraction pressure is 22-25 MPa, and the extraction time is 6-10 min; the addition mode of the absolute ethyl alcohol is preferably as follows: adding gynura procumbens powder at an interval of 50-60 min according to a mass ratio of 1 g: 1 ml-1 g: 3 ml; the conditions in the separation vessel were: the temperature is 35-40 ℃, the pressure is 13-16 MPa, and the time is 4-6 min.

4. The method for extracting gynura procumbens alkaloids with high antibacterial activity according to any one of claims 1 to 3, wherein in step (3), the macroporous adsorbent resin is D101 type macroporous adsorbent resin; the adsorption flow rate of the macroporous adsorption resin is 2-6 BV/h, and the temperature is 30-60 ℃; the elution is performed by taking ethanol with the volume concentration of 10-40% as an eluent, the flow rate is 1-4 BV/h, and the temperature is 10-40 ℃.

5. The method for extracting gynura procumbens alkaloids with high antibacterial activity according to any one of claims 1 to 4, wherein in the step (4), the temperature of vacuum drying is 55-60 ℃, the vacuum degree is 50-90 KPa, and the drying time is 4-6 h.

6. The method for extracting gynura procumbens alkaloid with high bacteriostatic activity according to one of claims 1 to 5, wherein in the step (5), the preparation method of the total alkaloid saturated solution comprises the following steps: adding absolute ethyl alcohol according to the proportion of 1 g: 2 ml-1 g: 5ml of total alkaloid dry matter, and then dissolving at the temperature of 50-60 ℃; the pressure for pressurizing is preferably 40-65 Mpa, and the time for maintaining the pressure is preferably 30-100 min.

7. The method for extracting gynura procumbens alkaloids with high antibacterial activity according to any one of claims 1 to 6, wherein in the step (6), the mesh number of the filtering screen is 200-250 meshes; the temperature of the anhydrous glacial ethanol is preferably-25 to-5 ℃.

8. The method for extracting gynura procumbens alkaloid with high bacteriostatic activity according to one of claims 1 to 7, wherein in the step (7), the solvent is dissolved by adding absolute ethyl alcohol according to the mass ratio of 1 g: 2ml to 1 g: 5ml of coarse crystals; the magnetic flux density of the electromagnetic field is preferably 0.2-0.7T; the vacuum degree during vacuum concentration and crystallization is preferably 80-100 Kpa.

9. The method for extracting gynura procumbens alkaloids with high antibacterial activity according to any one of claims 1 to 8, wherein in the step (8), the mesh number of the filter cloth is 200-250 meshes.

10. The method for extracting gynura procumbens alkaloids with high antibacterial activity according to any one of claims 1 to 9, wherein in the step (9), the vacuum drying is performed at a temperature of 40 to 45 ℃, a vacuum degree of 50 to 90KPa and a time of 20 to 24 hours.

Technical Field

The invention relates to a method for extracting and separating plant alkaloid, in particular to a method for extracting and separating alkaloid with high bacteriostatic activity from gynura procumbens.

Background

Gynura procumbens (Lour.) Merr is Compositae, and Gynura procumbens is climbing herbaceous plant, and is approved as new resource food by the national ministry of health in 2012. A large number of researches show that gynura procumbens contains a large number of chlorogenic acid, flavonoid, alkaloid and other active ingredients, and has the effects of clearing away heat and toxic materials, promoting blood circulation to remove meridian obstruction, improving human immunity, resisting viruses and the like.

At present, the work of extracting and separating the functional active ingredients of gynura procumbens mainly focuses on the aspects of chlorogenic acid, flavonoid compounds, polysaccharide and the like, and the research on the extraction method of alkaloid is less. Some plant alkaloids have antibacterial, antitumor and blood sugar lowering effects, and are low in cost, so that they are hot spots for research. In particular, in the food industry, food preservatives are often added in order to prolong the shelf life of products, and excessive consumption of artificially synthesized preservatives is not beneficial to human health. Therefore, the efficient and safe bacteriostatic preservative extracted from the gynura procumbens is urgently expected.

The alkaloid is a nitrogen-containing basic organic compound in plant secondary metabolites, most of the alkaloid has complex nitrogen-containing heterocycles, and the alkaloid is one of important effective components in Chinese herbal medicines. At present, 5000 to 7000 kinds of alkaloids separated from plants can be classified into organic amines, indoles, isoquinolines, pyridines, and tropanes based on their structures. Most alkaloids are insoluble in water or water, and are easily soluble in organic solvents such as chloroform, diethyl ether, ethanol, acetone, etc. The use of organic solvents, although helpful for the extraction and separation of alkaloids, also causes environmental pollution and poor safety of alkaloids.

Therefore, it is desirable to have a method for extracting and separating gynura procumbens alkaloids with safety and high efficiency, the extracted and separated alkaloids have strong antibacterial activity, and the extracted and separated alkaloids can be added into food to inhibit the growth of various harmful microorganisms, and simultaneously can help consumers to resist viruses and keep the health of the bodies.

Disclosure of Invention

The technical problem to be solved by the invention is to overcome the defects in the prior art and provide the method for extracting and separating the gynura procumbens alkaloid, which has the advantages of less environmental pollution in the extraction process, high extraction, separation and purification efficiency, good product safety and strong bacteriostatic activity.

The molecular formula of the gynura procumbens alkaloid compound extracted by the invention is C₇H₁₁NO₂Named as Arecaidine, has a chemical structural formula as follows:

the technical scheme adopted by the invention for solving the technical problems is as follows:

a method for extracting Gynura procumbens alkaloid with high antibacterial activity comprises the following steps:

(1) crushing and sieving: cleaning whole plant of Gynura procumbens, removing impurities, draining, drying, and pulverizing to obtain Gynura procumbens whole plant powder;

(2) supercritical CO₂And (3) extraction: placing the gynura procumbens powder obtained in the step (1) into an extraction kettle, then respectively preheating the extraction kettle, a separation kettle and a storage tank, pressurizing the system when the set temperature is reached, adding absolute ethyl alcohol as an entrainer, performing circular extraction, and separating through the separation kettle to obtain an extracting solution;

(3) separation: adsorbing the extracting solution obtained in the step (2) by using macroporous adsorption resin, eluting and collecting to obtain eluent;

(4) and (3) vacuum drying: vacuum drying the eluent obtained in the step (3) to obtain total alkaloid dry matter;

(5) high-pressure crystallization: adding absolute ethyl alcohol into the total alkaloid dry matter obtained in the step (4) to form a total alkaloid absolute ethyl alcohol saturated solution, pouring the total alkaloid absolute ethyl alcohol saturated solution into a pressure container, pressurizing, fully cooling and crystallizing the solution at the temperature, and then releasing pressure to obtain a high-pressure crystallization mixed solution;

(6) washing and drying: filtering the high-pressure crystallization mixed solution obtained in the step (5), washing wet crystals obtained by filtering for 1-2 times by using anhydrous glacial ethanol, and drying in vacuum to constant weight to obtain coarse crystals;

(7) electromagnetic field crystallization: putting the coarse crystal obtained in the step (6) into a container, dissolving the coarse crystal by using a solvent, then putting the container into an electromagnetic field for vacuum concentration and crystallization, cooling to normal temperature after crystallization is finished, and destroying the vacuum degree to obtain a crystal mixed solution;

(8) and (3) filtering: filtering the crystal mixed solution obtained in the step (7) by using filter cloth to obtain crystals;

(9) washing and drying: and (4) washing the crystal obtained in the step (8) with anhydrous glacial ethanol, and then carrying out vacuum drying to obtain the gynura procumbens alkaloid crystal with high purity.

Further, in the step (1), the drying temperature is 65-75 ℃, the drying is carried out until the moisture content is 8-14% by mass, and the mesh number of the sieving screen is preferably 60 meshes.

Further, in the step (2), supercritical CO in the extraction kettle₂The extraction conditions were: the extraction temperature is 45-55 ℃, the extraction pressure is 22-25 MPa, and the extraction time is 6-10 min.

Further, in the step (2), the anhydrous ethanol is preferably added in a manner of: adding the gynura procumbens powder at an interval of 50-60 min according to the mass ratio of 1 g: 1 ml-1 g: 3 ml.

Further, in the step (2), the conditions in the separation kettle are as follows: the temperature is 35-40 ℃, the pressure is 13-16 MPa, and the time is 4-6 min.

Further, in the step (3), the macroporous adsorption resin is D101 type macroporous adsorption resin; the adsorption flow rate of the macroporous adsorption resin is 2-6 BV/h, and the temperature is 30-60 ℃; the elution is performed by taking ethanol with the volume concentration of 10-40% as an eluent, the flow rate is 1-4 BV/h, and the temperature is 10-40 ℃.

Further, in the step (4), the temperature of vacuum drying is 55-60 ℃, the vacuum degree is 50-90 KPa, and the drying time is 4-6 h.

Further, in the step (5), the preparation method of the total alkaloid saturated solution comprises the following steps: adding absolute ethyl alcohol according to the proportion of 1 g: 2 ml-1 g: 5ml of total alkaloid dry matter, and then dissolving at the temperature of 50-60 ℃; the pressure for pressurizing is preferably 40-65 Mpa, and the time for maintaining the pressure is preferably 30-100 min.

Further, in the step (6), the mesh number of the filter screen for filtering is 200-250 meshes. The temperature of the anhydrous glacial ethanol is-25 to-5 ℃.

Further, in the step (7), the solvent is dissolved by adding absolute ethyl alcohol according to the proportion that the mass of the crude crystals is 1 g: 2 ml-1 g: 5 ml. The cooling to the normal temperature means that the temperature is slowly reduced from 40-50 ℃ to the normal temperature at the speed of 1-2 ℃/min.

Further, in the step (7), the magnetic flux density of the electromagnetic field is 0.2-0.7T.

Further, in the step (7), the vacuum degree during vacuum concentration and crystallization is 80-100 KPa.

Further, in the step (8), the mesh number of the filter cloth is 200-250 meshes.

Further, in the step (9), the temperature of the vacuum drying is 40-45 ℃, the vacuum degree is 50-90 KPa, and the time is 20-24 h.

Compared with the prior art, the invention has the beneficial effects that:

1) the invention passes supercritical CO₂The extraction technology obtains high-purity alkaloid extract, and then the extract is put under high pressure to accelerate the formation of crystal nuclei and the growth of crystals in the cooling process, so that alkaloid crystals are quickly obtained;

2) the method comprises the steps of placing a crude crystallization solution in an electromagnetic field for reduced pressure concentration crystallization separation, influencing the aggregation state, migration speed and direction of molecules and atoms through the action of the magnetic field, increasing the collision chance among alkaloids, promoting the formation of crystal nuclei and the growth of crystals, and simultaneously increasing the separation degree among substances by means of the influence on the various substances caused by the difference of polarity, charge, magnetic sensitivity and the like among the various substances in the crude crystallization solution, so as to achieve the purposes of separation and purification;

3) the invention does not use any toxic and harmful chemical reagent, and has no pollution to the environment;

in a word, the invention overcomes the defects of the prior art, provides the extraction method of the high-purity gynura procumbens alkaloid, and the prepared alkaloid has stronger bacteriostatic activity, has better inhibiting effect on viruses RSV and HSV-1, and has the effect similar to that of coptisine crystals.

Detailed Description

The present invention will be further described with reference to the following examples.

The chemical reagents used in the examples of the present invention, unless otherwise specified, are commercially available in a conventional manner.

Example 1

The embodiment comprises the following steps:

(1) crushing and sieving: cleaning whole plant of Gynura procumbens, removing impurities, draining, drying at 70 deg.C until water content is 10%, pulverizing with pulverizer, and sieving with 60 mesh sieve to obtain Gynura procumbens whole plant powder;

(2) supercritical CO₂And (3) extraction: placing the gynura procumbens powder obtained in the step (1) into an extraction kettle, then respectively preheating the extraction kettle, a separation kettle and a storage tank, pressurizing the system when the set temperature is reached, adding absolute ethyl alcohol as an entrainer according to the mass ratio of 1g to 1ml of gynura procumbens powder every 50min, circularly extracting, and obtaining an extracting solution through the separation kettle; supercritical CO in extraction kettle₂The extraction conditions were: the temperature is 50 ℃, the pressure is 23MPa, and the time is 8 min; the working conditions of the separation kettle are as follows: the temperature is 35 ℃, the pressure is 15MPa, and the time is 5 min;

(3) separation: adsorbing the extracting solution obtained in the step (2) by using D101 type macroporous adsorption resin, eluting and collecting to obtain eluent; the adsorption flow rate of the macroporous adsorption resin is 3BV/h, and the temperature is 40 ℃; during elution, ethanol with the volume concentration of 25% is adopted as an eluent, the flow rate is 2BV/h, and the temperature is 35 ℃;

(4) and (3) vacuum drying: vacuum drying the eluent obtained in the step (3) at 55 ℃ and 70KPa for 5h to obtain total alkaloid dry matter;

(5) high-pressure crystallization: adding absolute ethanol into the total alkaloid dry matter obtained in the step (4) according to the proportion of 1 g: 2ml, dissolving at 50 ℃ to form a saturated solution, pouring the saturated solution into a pressure container, maintaining the pressure condition for 50min at 45MPa, and releasing pressure after the solution is sufficiently cooled and crystallized to obtain a high-pressure crystallization solution;

(6) washing and drying: filtering the high-pressure crystallization solution obtained in the step (5) by using a 200-mesh screen, washing the crystals for 1 time by using anhydrous glacial ethanol at the temperature of minus 10 ℃, and then drying in vacuum to constant weight to obtain coarse crystals; the vacuum drying conditions were: the temperature is 55 ℃, the vacuum degree is 60KPa, and the time is 5 h;

(7) electromagnetic field crystallization: dissolving the coarse crystal obtained in the step (6) in absolute ethyl alcohol according to the mass ratio of 1 g: 3ml, then placing the dissolved coarse crystal in an electromagnetic field with the magnetic flux density of 0.3T for vacuum concentration crystallization, wherein the vacuum degree is controlled to be 85KPa, slowly cooling the solution from 45 ℃ to normal temperature at the speed of 1 ℃/min, and destroying the vacuum degree to obtain a crystal solution;

(8) and (3) filtering: filtering the crystal solution obtained in the step (7) by using 200-mesh filter cloth to obtain crystals;

(9) washing and drying: washing the crystal obtained in the step (8) with anhydrous ice ethanol at-15 ℃ for 2 times, and then drying for 22 hours at 45 ℃ and under the vacuum degree of 70KPa to obtain Gynura procumbens alkaloid crystals with the purity of 95%, wherein the yield is 73%.

Comparative example 1

Comparative example 1 differs from example 1 only in that the crystallization pressure in step (5) was adjusted from 45MPa to atmospheric pressure, and crude crystals were obtained after completion of the experimental step (6) of washing and drying as described in example 1 above, and then the time for starting the occurrence of crystals in step (5), the purity of the crude crystals and the recovery rate were measured respectively, and the results are shown in Table 1.

Example 2

Example 2 differs from example 1 only in that the crystallization pressure in step (5) was adjusted from 45MPa to 40MPa, and crude crystals were obtained after completion of the experimental step (6) of washing and drying as described in the above examples, and then the time for starting the appearance of crystals in step (5), the purity of the crude crystals and the recovery rate were measured, respectively, and the results are shown in Table 1.

Example 3

Example 3 differs from example 1 only in that the crystallization pressure in step (5) was adjusted from 45MPa to 60MPa, and crude crystals were obtained after completion of the experimental step (6) of washing and drying as described in the above examples, and then the time for starting the appearance of crystals in step (5), the purity of the crude crystals and the product recovery were measured, respectively, and the results are shown in Table 1.

TABLE 1 influence of the pressure of the high-pressure crystallization on the crystallization of crude alkaloid crystals

Note: the upper right-hand corner of the data in the table indicates that there is a very significant difference between examples 2 and 3 compared to comparative example 1, P < 0.01.

As can be seen from Table 1, there were very significant differences in the onset of crystallization time, crude purity and recovery rate of examples 2 and 3 as compared to comparative example 1, P < 0.01. The crystallization of alkaloid is assisted by applying certain pressure during the crystallization process, wherein crystallization is started at 41min and 38min after 40MPa and 60MPa are applied, and crystallization is started after 63min at normal temperature. The purity of the obtained alkaloid crystals is improved from 46.52% of comparative example 1 to 78.14% of example 2 and 79.77% of example 3, and the product recovery rate is improved from 56.08% of comparative example 1 to 68.81% of example 2 and 70.33% of example 3.

Comparative example 2

Comparative example 2 differs from example 1 only in that the electromagnetic field intensity in step (7) was adjusted from a magnetic flux density of 0.3T to no additional magnetic field intensity, gynura procumbens alkaloid crystals were obtained as described in example 1 above, and then the time to start appearance of crystals, the purity of final crystals and the recovery rate were measured in step (7), and the results are shown in table 2.

Example 4

Inventive example 4 differs from example 1 only in that the electromagnetic field intensity in step (7) was adjusted from a magnetic flux density of 0.3T to 0.6T, gynura procumbens alkaloid crystals were obtained as described in example 1 above, and then the time to start appearance of crystals, the purity of final crystals and the recovery rate were measured in step (7), and the results are shown in table 2.

TABLE 2 influence of electromagnetic field intensity on alkaloid crystallization

Note: the upper right-hand corner of the data in the table indicates that there is a very significant difference between examples 1 and 4 compared to comparative example 1, P < 0.01.

As can be seen from Table 2, the crystal onset time, product purity and product recovery of inventive examples 1 and 4 were all very significantly different from those of comparative example 2, P < 0.01. Compared with comparative example 2 without electromagnetic field, the alkaloid crystals obtained under the conditions of example 1 with the magnetic flux density of 0.3T and example 4 with the magnetic flux density of 0.5T respectively begin to crystallize at a time earlier from 51min, which is the time taken by comparative example 2, to 27min, which is example 1, and 26min, which is example 4, the alkaloid crystals obtained have a purity increased from 81.52% of comparative example 2 to 95.14% of example 1 and 97.01% of example 4, and a product recovery increased from 61.94% of comparative example 2 to 72.40% of example 1 and 78.26% of example 4.

TABLE 3 influence of Gynura procumbens alkaloid crystals on zone diameter/mm

Test sample	Escherichia coli	Staphylococcus aureus	Bacillus subtilis	Salmonella	Aspergillus niger
						EXAMPLE 1 Crystal	12.54±0.25b	13.29±0.17c	10.86±0.25b	13.47±0.49a	12.86±0.25a
EXAMPLE 4 crystals	15.59±0.12a	14.02±0.42b	11.95±0.34a	13.65±0.16a	12.82±0.28a
						Sterile water (blank control)	7.00±0.01c	7.00±0.01d	7.00±0.01c	7.00±0.01b	7.00±0.01b
Coptisine (positive control)	16.34±0.63a	15.11±0.12a	12.12±0.15a	13.10±0.12a	12.10±0.12a

Note: the numerical values of different letters marked on the same line of data have significant difference, and P is less than 0.05;

2 Gynura procumbens alkaloid crystals prepared by the methods of the embodiment 1 and the embodiment 4 and coptisine (alkaloid standard sample) are respectively prepared into a sample solution of 2mg/mL by using sterile water, the sterile water is used as blank control, and the bacteriostatic effect of the sample solution is tested by adopting a flat plate perforation method. Putting 100 μ L of the prepared sample solution into a round hole with a plate aperture of 7mm, putting a bacteria culture medium into a constant temperature incubator at 37 ℃ for culturing for 24h, and putting a mould culture medium into a constant temperature incubator at 28 ℃ for culturing for 48 h. The diameter of the zone of inhibition of the dish was observed and measured with a vernier caliper, the results are shown in Table 3.

Referring to table 3, the 2 kinds of alkaloid crystals with different purities extracted in example 1 and example 4 of the present invention have significantly higher bacteriostatic effect on 5 kinds of common bacteria than the sterile water control (P < 0.05), which indicates that they have broad-spectrum bacteriostatic effect. The alkaloid crystals extracted by the method in the example 4 and coptisine have the strongest bacteriostatic effect, but no significant difference (P is more than 0.05) exists between the alkaloid crystals and coptisine, which shows that the gynura procumbens alkaloid crystals extracted by the method in the example 4 have stronger bacteriostatic activity.

TABLE 4 inhibitory Effect of Gynura procumbens alkaloid crystals on RSV and HSV-1 viruses

The crystal of example 1, the crystal of example 4 and the crystal of coptisine (alkaloid standard) are respectively prepared into different concentrations by sterile water, and the sterile water is used as a blank control. Experimental viruses Respiratory Syncytial Virus (RSV) and herpes simplex virus type I (HSV-1) are adopted, and the virus inhibition effect is evaluated to select the half Toxic Concentration (TC) of the drug₅₀) Half Effective Concentration (EC)₅₀) And therapeutic index [ TI ═ half Toxic Concentration (TC)₅₀) Effective concentration at half maximum (EC)₅₀)]And 3 indexes are evaluated. The larger the therapeutic index TI, the better the virus inhibition effect of this sample.

As can be seen from Table 4, the crystals of example 1 and the crystals of example 4 have a large therapeutic index TI (therapeutic index of viral RSV and HSV-1), a strong inhibitory effect and an effect close to that of coptisine crystals. In the effective concentration range of the gynura procumbens alkaloid crystals, the alkaloid concentration is reduced along with the increase of the dilution concentration, the inhibition rate is reduced, and a relatively obvious dose-effect relationship is shown.