CN111494455A - Method for extracting flavonoid compounds in lamiophlomis rotata by response surface method - Google Patents
Method for extracting flavonoid compounds in lamiophlomis rotata by response surface method Download PDFInfo
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- A61K36/18—Magnoliophyta (angiosperms)
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- A—HUMAN NECESSITIES
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- A61K2236/00—Isolation or extraction methods of medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicine
- A61K2236/30—Extraction of the material
- A61K2236/33—Extraction of the material involving extraction with hydrophilic solvents, e.g. lower alcohols, esters or ketones
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- A—HUMAN NECESSITIES
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- A61K2236/30—Extraction of the material
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Abstract
The invention provides a method for extracting flavonoids from lamiophlomis rotata by a response surface method, which comprises the steps of A) crushing lamiophlomis rotata and sieving to obtain medicinal material powder, B) adding 10-90% ethanol solution into the medicinal material powder according to the solid-to-liquid ratio of 1: 10-1: 50g/m L to soak for 2-8 h, carrying out ultrasonic extraction for 10-30 min, and filtering to obtain a flavonoid extract, wherein the soaking time, the ethanol volume fraction and the ultrasonic extraction conditions of the solid-to-liquid ratio are optimized by the response surface method.
Description
Technical Field
The invention relates to the technical field of extraction of natural products, in particular to a method for extracting flavonoid compounds in lamiophlomis rotata by a response surface method.
Background
The lamiophlomis rotate (Benth.) Kudo (L amiohlomis rotate) is a specific plant from Tibet plateau, which is widely distributed in Tibet, Qinghai, Gansu and Sichuan, etc. the overground part of the lamiophlomis rotate shows wide pharmacological activity, such as treating traumatic injury, traumatic hemorrhage, yellow water disease, rheumatic arthralgia, promoting blood circulation to remove blood stasis, dispelling wind and relieving pain, etc.
The conventional extraction method of the flavonoid compound comprises an immersion method, a reflux extraction method, a Soxhlet extraction method and the like, and mainly has the following problems: consumes a large amount of organic solvent, is time-consuming and has low extraction rate. In contrast, ultrasonic extraction has been developed in recent years, not only to preserve the structure of the components and their molecular properties, but also to reduce solvent and energy consumption, save time and simplify operations.
The traditional optimization method for extracting the total flavonoids in the lamiophlomis rotata through an orthogonal experiment has low extraction rate, and a function expression between a response value and an independent variable cannot be clearly found. The development of a method is necessary to solve the problem of low extraction rate of the total flavone in the lamiophlomis rotata extracted by the traditional method.
Disclosure of Invention
In view of the above, the technical problem to be solved by the present invention is to provide a method for extracting flavonoids from lamiophlomis rotata by using a response surface method, and the method provided by the present invention has high extraction rate and high purity.
The invention provides a method for extracting total flavonoids from lamiophlomis rotata by a response surface method, which comprises the following steps:
A) pulverizing radix Lamiophlomidis Rotatae, and sieving to obtain medicinal powder;
B) adding the medicinal material powder into 10-90% ethanol solution according to the solid-to-liquid ratio of 1: 10-1: 50g/m L, soaking for 2-8 h, performing ultrasonic extraction for 10-30 min, and filtering to obtain a flavonoid compound extracting solution;
the ultrasonic extraction conditions of the soaking time, the ethanol volume fraction and the solid-liquid ratio are optimized by a response surface method, and specifically the method comprises the following steps:
i) single factor investigation: extracting total flavonoids from Lamiophlomis rotata kudo by ultrasonic extraction technology, and respectively designing single factor experiments according to soaking time, ethanol volume fraction, solid-liquid ratio, extraction time and ultrasonic power;
ii) on the basis of a single-factor experiment, adopting statistical software Design Expert11, taking the extraction rate of the flavonoid compounds in the lamiophlomis rotata as a response value, selecting soaking time, ethanol volume fraction and solid-liquid ratio which have great influence on the extraction rate of the flavonoid as response factors, and applying a central composite Design experiment Design principle to Design a response surface experiment with a five-factor level;
iii) carrying out ultrasonic extraction on the medicinal material powder to obtain the extraction rate of total flavonoids in the medicinal material under the conditions of soaking time, ethanol volume fraction and solid-to-liquid ratio; and (3) performing multiple regression fitting analysis on the data by applying Design Expert11 to obtain the optimal ultrasonic extraction conditions of the total flavonoids in the lamiophlomis rotata.
Preferably, the sieving in the step A) is specifically 40-mesh sieving.
Preferably, the extraction times in the step B) are 1-3 times.
Preferably, said step iii) comprises: and measuring the content of the flavonoid compounds in the sample solution to be measured and the standard sample solution by using an ultraviolet-visible spectrometer.
Preferably, said step iii) comprises: and (3) applying Design Expert11, establishing a quadratic polynomial equation model of the influence of experimental factors on the response value, and performing variance analysis and significance test on the response surface multiple regression model.
Preferably, the optimal ultrasonic extraction conditions in the step iii) are soaking time of 5 hours, volume fraction of ethanol in the extraction solvent of 57 percent and solid-to-liquid ratio of 1:57g/m L.
Preferably, the optimal ultrasonic extraction conditions in the step iii) are that the soaking time is 5.04h, the volume fraction of ethanol in the extraction solvent is 56.94%, and the solid-liquid ratio is 1:57.44g/m L.
Preferably, the filtration in the step B) further comprises rotary evaporation to obtain an extract.
Preferably, the quadratic polynomial equation for obtaining the extraction rate of the lamiophlomis rotata by the multivariate regression analysis is as follows:
Y＝6.59+0.19A-0.30B+0.68C+0.07AB+0.09AC+0.06BC-0.16A2-0.64B2-0.31C2。
compared with the prior art, the invention provides a method for extracting total flavonoids from lamiophlomis rotata by using a response surface method, which comprises the steps of A) crushing lamiophlomis rotata and sieving to obtain medicinal material powder, B) adding 10-90% ethanol solution into the medicinal material powder according to the solid-to-liquid ratio of 1: 10-1: 50g/m L for soaking for 2-8 h, carrying out ultrasonic extraction for 10-30 min, and filtering to obtain flavonoid extract, wherein the ultrasonic extraction conditions of the soaking time, the ethanol volume fraction and the solid-to-liquid ratio are optimized by using the response surface method, i) performing single factor investigation by using an ultrasonic extraction technology to extract total flavonoids from lamiophlomis rotata, respectively designing single factor experiments according to the soaking time, the ethanol volume fraction, the solid-to-liquid ratio, the extraction time and the ultrasonic power, ii) on the basis of the single factor experiments, using statistical software Design Expert11 to Design the extraction rate of flavonoids from lamiophlomis provided by using the statistical software, designing the extraction center on the extraction rate, and using the optimal extraction ratio of the ultrasonic extraction technology to extract total flavonoids from lamiophlomis developed according to the traditional extraction process, the response surface extraction principle of the traditional extraction technology, the extraction of the traditional Chinese medicinal materials, the extraction technology, the extraction of the traditional extraction of the general flavonoids, and the extraction of the general flavonoids in the traditional extraction technology, and the research of extracting the general flavonoids, and the general flavonoids in the research of the general flavonoids, thereby obtaining the general flavonoids by using the research of the general flavonoids in the research of the general flavonoids.
Drawings
FIG. 1 is a schematic diagram of a standard curve for rutin;
FIG. 2 is a schematic diagram showing the effect of soaking time on the extraction rate of total flavonoids;
FIG. 3 is a graph showing the effect of ethanol volume fraction on the extraction rate of total flavonoids;
FIG. 4 is a schematic diagram showing the effect of solid-liquid ratio on the extraction rate of total flavonoids;
FIG. 5 is a schematic diagram showing the effect of ultrasonic power on the extraction rate of total flavonoids;
FIG. 6 is a schematic diagram showing the effect of extraction time on the extraction rate of total flavonoids;
FIG. 7 is a graph of response plots of the interaction of soaking time and ethanol volume fraction on the extraction rate of lamiophlomis rotata;
FIG. 8 is a response surface graph of the interaction of soaking time and solid-to-liquid ratio for influencing the extraction rate of lamiophlomis rotata;
FIG. 9 is a response surface graph of ethanol volume fraction and solid-to-liquid ratio interactively influencing the extraction rate of lamiophlomis rotata.
Detailed Description
The invention provides a method for extracting total flavonoids from lamiophlomis rotata by a response surface method, and a person skilled in the art can appropriately improve process parameters by referring to the content in the text for realization. It is expressly intended that all such similar substitutes and modifications apparent to those skilled in the art are deemed to be within the scope of the invention. While the methods and applications of this invention have been described in terms of preferred embodiments, it will be apparent to those of ordinary skill in the art that variations and modifications in the methods and applications described herein, as well as other suitable variations and combinations, may be made to implement and use the techniques of this invention without departing from the spirit and scope of the invention.
The invention provides a method for extracting total flavonoids from lamiophlomis rotata by a response surface method, which comprises the following steps:
A) pulverizing radix Lamiophlomidis Rotatae, and sieving to obtain medicinal powder;
B) adding the medicinal material powder into 10-90% ethanol solution according to the solid-to-liquid ratio of 1: 10-1: 50g/m L, soaking for 2-8 h, performing ultrasonic extraction for 10-30 min, and filtering to obtain a total flavone extracting solution;
the ultrasonic extraction conditions of the soaking time, the ethanol volume fraction and the solid-liquid ratio are optimized by a response surface method, and specifically the method comprises the following steps:
i) single factor investigation: extracting total flavone compounds in lamiophlomis rotata by adopting an ultrasonic extraction technology, and respectively designing a single-factor experiment according to soaking time, ethanol volume fraction, solid-liquid ratio, extraction time and ultrasonic power;
ii) on the basis of a single-factor experiment, adopting statistical software Design Expert11, taking the extraction rate of the flavonoid compounds in the lamiophlomis rotata as a response value, selecting soaking time, ethanol volume fraction and solid-liquid ratio which have great influence on the extraction rate of the flavonoid as response factors, and applying a central composite Design experiment Design principle to Design a response surface experiment with a five-factor level;
iii) carrying out ultrasonic extraction on the medicinal material powder to obtain the extraction rate of total flavonoids in the medicinal material under the conditions of soaking time, ethanol volume fraction and solid-to-liquid ratio; and (3) performing multiple regression fitting analysis on the data by applying Design Expert11 to obtain the optimal ultrasonic extraction conditions of the total flavonoids in the lamiophlomis rotata.
The invention provides a method for extracting total flavonoids from lamiophlomis rotata by a response surface method. Firstly, the lamiophlomis rotata is crushed and sieved to obtain medicinal powder.
The preferable concrete is as follows: pulverizing radix Lamiophlomidis Rotatae, and sieving with 40 mesh sieve to obtain medicinal powder.
The source of the unique flavor is not limited in the present invention and may be commercially available as is well known to those skilled in the art.
Adding the medicinal powder into 10-90% ethanol solution according to the solid-to-liquid ratio of 1: 10-1: 50g/m L, and soaking for 2-8 h.
Preferably, the medicinal material powder is soaked for 2-8 hours in a solid-to-liquid ratio of 1: 10-1: 50g/m L, and then an ethanol solution with a volume fraction of 10-90% is added.
In a single factor experiment, the above point values may be selected, as may other values within the above point values.
Ultrasonic extracting for 10-30 min, and filtering to obtain flavonoid compound extracting solution; the extraction times are 1-3 times; more preferably 2 to 3 times.
Mixing the obtained filtrates, and rotary-steaming to obtain extract. The present invention is not limited to the specific parameters of the rotary evaporation, and the specific parameters are well known to those skilled in the art.
The ultrasonic extraction conditions of the soaking time, the ethanol volume fraction and the solid-liquid ratio are optimized by a response surface method, and the ultrasonic extraction conditions are as follows: an ultrasonic extraction process for extracting total flavonoids from lamiophlomis rotata is optimized by adopting a response surface method, and a three-factor five-level experiment is designed by applying a central composite design experiment design principle on the basis of a single-factor experiment, and specifically comprises the following steps:
i) single factor investigation: extracting total flavone from radix Lamiophlomidis Rotatae by ultrasonic extraction, and respectively designing single factor experiment according to soaking time, ethanol volume fraction, solid-liquid ratio, extraction time and ultrasonic power.
Specifically, the solid-liquid ratio is 1: 10-1: 50g/m L, the solid-liquid ratios are 1:10, 1:20, 1:30, 1:40 and 1:50g/m L are taken as experimental points respectively, and the influence of different solid-liquid ratios on the extraction rate of the total flavonoids is examined;
10-90% ethanol: respectively taking ethanol volume fractions of 10%, 30%, 50%, 70% and 90% as experimental points, and investigating the influence of different ethanol volume fractions on the extraction rate of the lamiophlomis rotata total flavonoids;
soaking for 2-8 h: taking soaking times of 0, 2, 4, 6 and 8h as experimental points respectively, and investigating the influence of different soaking times on the extraction rate of the lamiophlomis rotata total flavonoids;
respectively taking ultrasonic powers of 240W, 300W, 360W, 420W and 480W as experimental points, and investigating the influence of different ultrasonic powers on the extraction rate of the lamiophlomis rotata total flavonoids;
taking extraction time of 10min, 15min, 20min, 25 min and 30min as experimental points, and investigating the influence of different extraction time on the extraction rate of the lamiophlomis rotata total flavonoids.
ii) on the basis of a single-factor experiment, adopting statistical software Design Expert11, taking the extraction rate of the flavonoid compounds in the lamiophlomis rotata as a response value, selecting soaking time, ethanol volume fraction and solid-liquid ratio which have great influence on the extraction rate of the flavonoid as response factors, and applying a central composite Design experiment Design principle to Design a response surface experiment with a five-factor level.
iii) carrying out ultrasonic extraction on the medicinal material powder to obtain the extraction rate of the flavonoid compounds in the medicinal material under the conditions of soaking time, ethanol volume fraction and solid-to-liquid ratio; and (3) performing multiple regression fitting analysis on the data by using Design Expert11 to obtain the optimal ultrasonic extraction conditions of the flavonoid compounds in the lamiophlomis rotata.
Wherein step iii) comprises: and measuring the content of the flavonoid compounds in the sample solution to be measured and the standard sample solution by using an ultraviolet-visible spectrometer.
In the embodiment of the invention, an ultraviolet spectrophotometry method is adopted to measure flavonoid compounds in lamiophlomis rotata, wherein the extract extracted from lamiophlomis rotata is dissolved by deionized water to a constant volume, 200 mu L of extract is taken, and 0.3m L5% of NaNO is respectively added into the extract2Shaking, standing for 10min, adding 0.3m L10% Al (NO)3)3Shaking, standing for 10min, adding 4m L4% NaOH, shaking, diluting with 70% ethanol to 25m L, mixing, standing for 15min, and measuring absorbance at 505nm with 70% ethanol as blank control, and measuring total flavone extraction rate by using rutin absorbance standard curve as control.
The formula for calculating the total flavone extraction rate is as follows, wherein Y is C ×α× V/W × 100%, Y is the total flavone extraction rate in lamiophlomis rotata, C is the mass concentration (mg/m L) of the flavonoid substances in the extracting solution, α is the dilution factor, V is the volume (m L) of the extracting solution, and W is the mass (g) of the lamiophlomis rotata powder.
The rutin absorbance standard curve is prepared by weighing 50.0mg rutin standard, diluting with 70% ethanol to a constant volume of 250m L volumetric flask, preparing rutin standard mother liquor with concentration of 0.2mg/m L, accurately weighing 2.5, 5.0, 7.5, 10.0 and 12.5m L rutin standard mother liquor respectively, placing in 25m L volumetric flask with each gradient having 3 parallel concentrations, and adopting NaNO2-Al(NO3)3Preparing standard curve by color development method, adding 0.3m L5% NaNO into 25m L volume of mother liquor containing different amount of rutin standard substance2Shaking, standing for 10min, adding 0.3m L10% Al (NO)3)3Shaking, standing for 10min, adding 4m L4% NaOH, shaking, diluting with 70% ethanol to 25m L, mixing, standing for 15min, taking 70% ethanol as blank control group, measuring absorbance at 505nm, and taking the measured absorbance A (Y) as ordinate, and rutin as standardThe concentration (mg/m L) was plotted as a standard curve on the abscissa, as shown in FIG. 1.
The invention preferably applies Design Expert11, establishes a quadratic polynomial equation model of the influence of experimental factors on response values, and performs variance analysis and significance test on the response surface multiple regression model.
According to the invention, the optimal ultrasonic extraction conditions comprise soaking time of 5h, volume fraction of ethanol in the extraction solvent of 57 percent and solid-liquid ratio of 1:57g/m L.
According to the invention, the optimal ultrasonic extraction conditions in the step iii) are that the soaking time is 5.04h, the volume fraction of ethanol in the extraction solvent is 56.94%, and the solid-liquid ratio is 1:57.44g/m L.
In the invention, the quadratic polynomial equation for obtaining the extraction rate of the lamiophlomis rotata flavone by the multivariate regression analysis specifically comprises the following steps:
Y＝6.59+0.19A-0.30B+0.68C+0.07AB+0.09AC+0.06BC-0.16A2-0.64B2-0.31C2。
the method for optimizing and extracting the total flavonoids in the lamiophlomis rotata by utilizing the central combined experimental design and the response surface method obtains the optimal process condition for ultrasonically extracting the total flavonoids in the lamiophlomis rotata, has simple operation, stability and reliability, high purification efficiency and low energy consumption, and reduces the extraction cost of the flavonoids in the lamiophlomis rotata. The method provides theoretical basis for large-scale production of flavone in lamiophlomis rotata and further development and utilization of flavone compounds in plants.
The invention provides a method for extracting total flavonoids from lamiophlomis rotata by a response surface method, which comprises the steps of A) crushing lamiophlomis rotata and sieving to obtain medicinal material powder, B) adding 10-90% ethanol solution into the medicinal material powder according to a solid-to-liquid ratio of 1: 10-1: 50g/m L to soak for 2-8 h, carrying out ultrasonic extraction for 10-30 min, and filtering to obtain flavonoid extract, wherein the soaking time, the ethanol volume fraction and the solid-to-liquid ratio are optimized by the response surface method, i) single factor investigation comprises the steps of extracting the total flavonoids from lamiophlomis rotata by an ultrasonic extraction technology, respectively designing single factor experiments according to the soaking time, the ethanol volume fraction, the solid-to-liquid ratio, the extraction time and the ultrasonic power, ii) on the basis of the single factor experiments, adopting a statistical software DesignExt 11, designing an experimental principle by using the extraction rate of the flavonoid compounds in the lamiophlomis a response surface method, designing an optimal response Design principle, designing an optimal response theory of the flavonoid extraction principle, and optimizing the response surface extraction principle of the extraction of the flavonoid compounds from the lamiophlomis carried out by the ultrasonic extraction technology under the traditional ultrasonic extraction technology, so as to obtain the total flavonoids from the traditional Chinese medicinal material powder, and the traditional extraction method of the extraction of the traditional Chinese medicinal materials, thereby obtaining the total flavonoids by the extraction technology of the traditional Chinese lamiophlomis the extraction of the traditional Chinese medicinal materials.
In order to further illustrate the present invention, the method for extracting total flavonoids from lamiophlomis rotata provided by the present invention by the response surface method is described in detail below with reference to examples.
Example 1
The method comprises the steps of crushing a lamiophlomis rotata medicinal material, sieving the lamiophlomis rotata medicinal material by a 40-mesh sieve to obtain lamiophlomis rotata powder, weighing 2.0g of the preprocessed lamiophlomis rotata powder, fixing the volume fraction of ethanol to be 50%, setting the ultrasonic power to be 240W, setting the extraction time to be 20min, setting the solid-to-liquid ratio to be 1:20g/m L, and taking the soaking time of 0h, 2 h, 4h, 6 h and 8h as experimental points respectively to examine the influence of different soaking times on the extraction rate of lamiophlomis rotata total flavonoids.
Example 2
Crushing the lamiophlomis rotata medicinal material, sieving the lamiophlomis rotata medicinal material by a 40-mesh sieve to obtain lamiophlomis rotata powder, weighing 2.0g of the preprocessed lamiophlomis rotata powder, fixing the soaking time for 4h, the ultrasonic power is 240W, the extraction time is 20min, the solid-to-liquid ratio is 1:20g/m L, respectively taking the volume fractions of ethanol as 10%, 30%, 50%, 70% and 90% as experimental points, and investigating the influence of different ethanol volume fractions on the extraction rate of lamiophlomis rotata total flavone, wherein as shown in figure 3, the extraction rate of the lamiophlomis reduced after the ethanol volume fraction is increased, the extraction rate of the total flavone is the highest when the ethanol volume fraction is 50%, and then the ethanol volume fraction is further increased to 70%, and the extraction rate is slightly reduced.
Example 3
Crushing the lamiophlomis rotata medicinal material, sieving the lamiophlomis rotata medicinal material by a 40-mesh sieve to obtain lamiophlomis rotata powder, weighing 2.0g of the preprocessed lamiophlomis rotata powder, fixing the soaking time for 4h, ensuring that the volume fraction of ethanol is 50%, the ultrasonic power is 240W, the extraction time is 20min, and respectively taking solid-liquid ratios of 1:10, 1:20, 1:30, 1:40 and 1:50g/m L as experimental points to examine the influence of different solid-liquid ratios on the extraction rate of the lamiophlomis rotata total flavone, wherein the extraction rate is increased along with the increase of the solid-liquid ratio from 1:10 to 1:30g/m L from 1:30 to 1:50g/m L, however, the extraction rate is slightly reduced because the increase of the solid-liquid ratio can increase the contact area of the plant powder and the solvent, so that the flavonoid substances are more fully dissolved, but the extraction rate is not increased any more, so that the optimal solid-liquid ratio is determined to be 1:30g/m L.
Example 4
Crushing a lamiophlomis rotata medicinal material, sieving the lamiophlomis rotata medicinal material by a 40-mesh sieve to obtain lamiophlomis rotata powder, weighing 2.0g of the preprocessed lamiophlomis rotata powder, fixing the soaking time for 4h, ensuring that the volume fraction of ethanol is 50%, the solid-to-liquid ratio is 1:30g/m L, the extraction time is 20min, and respectively taking the ultrasonic power of 240 (40%), 300 (50%), 360 (60%), 420 (70%) and 480W (80%) as experimental points to examine the influence of different ultrasonic powers on the extraction rate of the lamiophlomis rotata total flavone, wherein as shown in figure 5, the extraction rate of the lamiophlomis rotata flavone is increased along with the increase of the ultrasonic power, and reaches the maximum value at 360W.
Example 5
Crushing the lamiophlomis rotata medicinal materials, sieving the lamiophlomis rotata medicinal materials by a 40-mesh sieve to obtain lamiophlomis rotata powder, weighing 2.0g of the preprocessed lamiophlomis rotata powder, fixing the soaking time for 4h, ensuring that the volume fraction of ethanol is 50%, the ultrasonic power is 360W, and the solid-to-liquid ratio is 1:30g/m L, taking extraction time of 10min, 15min, 20min, 25 min and 30min as experimental points respectively, and investigating the influence of different extraction time on the extraction rate of lamiophlomis rotata total flavone, wherein the extraction time is in positive correlation with the extraction yield of the lamiophlomis rotata as shown in figure 6, however, the extraction rate is slightly reduced from 20min to 30 min.
On the basis of the single-factor embodiment, soaking time (A), ethanol volume fraction (B) and solid-to-liquid ratio (C) are selected as experimental factors, the extraction rate of the lamiophlomis rotata total flavone is taken as a response value, and a response surface experiment of a five-factor level is designed by using design expert11 software and applying a central composite design experiment design principle.
Example 6
Three factors including soaking time (A), ethanol volume fraction (B) and solid-liquid ratio (C) are screened out through single factors to carry out a response surface method experiment, and a three-factor five-level experiment (shown in table 1) is designed by using statistical software Design Expert 11. Crushing the lamiophlomis rotata, sieving the lamiophlomis rotata with a 40-mesh sieve, weighing 2.0g of powder, respectively setting extraction variable parameters in the table 1 to perform ultrasonic extraction, filtering ultrasonic liquid, and performing rotary evaporation to obtain an extract.
TABLE 1 response surface method experimental factors and levels
The results of the experiment are shown in Table 2.
TABLE 2 Experimental design and results of response surface method
Response surface analysis and optimization: performing multiple regression analysis on the data in the table 2 by using Design Expert11 software to obtain a quadratic polynomial equation of the extraction rate of the lamiophlomis rotata flavone: lamiophlomis rotata (Benth.) kudo flavone extraction rate (%)
Y＝6.59+0.19A-0.30B+0.68C+0.07AB+0.09AC+0.06BC-0.16A2-0.64B2-0.31C2。
Table 3 shows the results of the regression analysis of variance, and the significance of the model (p) is very high (as shown by the significance analysis of variance of the model)<0.0001), the mismatching term is not significant, which shows that the interference of unknown factors on the experimental result is small, and the correlation coefficient (R) is small2) 0.9920, showing that the experimentally measured value of the extraction rate of the extracted lamiophlomis rotata has better fitting degree with the predicted value. Correction decision coefficient (R)adj 2) 0.9847, indicating that there was a change in the extraction rate of lamiophlomis rotata of about 98.47% that could be explained by this model.
TABLE 3 analysis of variance of regression models
aThe significance is as follows: indicates significance (P)<0.05); indicates very significant (P)<0.0001)
As can be seen from Table 3, A, B, C, B2、C2Has very obvious effect on the extraction rate of total flavonoids, AC and A2The influence on the extraction rate of the total flavone is obvious, and according to the F value in the regression equation, the primary and secondary sequence of the factors influencing the extraction rate of the lamiophlomis rotata flavone is as follows: solid-to-liquid ratio>Volume fraction of ethanol>And (4) soaking time.
3D response surface analysis graphs (figures 7-9) are made according to the regression equation, and the influence of the soaking time, the ethanol volume fraction and the solid-liquid ratio on the extraction rate of the lamiophlomis rotata total flavonoids can be visually seen. FIG. 7 is a three-dimensional response surface diagram of the interaction between soaking time and ethanol volume fraction to influence the extraction rate of total flavonoids. FIG. 8 shows the effect of soaking time and solid-liquid ratio on the extraction rate of total flavonoids. FIG. 9 is a 3D response surface graph of the effect of interaction of ethanol concentration and solid-to-liquid ratio on the extraction rate of flavone in Lamiophlomis rotata, showing the maximum point at the edge of the experimental region. From the above results, the interaction between the soaking time and the solid-to-liquid ratio has an important influence on the extraction rate of total flavonoids.
According to the obtained regression model, obtaining the optimal conditions for extracting the total flavonoids from the lamiophlomis rotata, namely soaking time is 5.04h, the volume fraction of ethanol is 56.94%, the solid-liquid ratio is 1:57.44g/m L, the extraction rate of the total flavonoids in the lamiophlomis rotata under the optimal extraction conditions is 7.056%, considering the feasibility of actual operation, modifying the extraction conditions into soaking time of 5h, the volume fraction of ethanol is 57%, and the solid-liquid ratio is 1:57g/m L, carrying out 3 verification experiments under the conditions, measuring the average value of the extraction rate of the total flavonoids to be 7.099%, comparing with a theoretical value, the relative error is 0.609%, and showing that the response surface method is reliable for the model established by optimizing the extraction conditions of the lamiophlomis rotata total flavonoids.
The foregoing is only a preferred embodiment of the present invention, and it should be noted that, for those skilled in the art, various modifications and decorations can be made without departing from the principle of the present invention, and these modifications and decorations should also be regarded as the protection scope of the present invention.
Claims (9)
1. A method for extracting total flavonoids from lamiophlomis rotata by a response surface method is characterized by comprising the following steps:
A) pulverizing radix Lamiophlomidis Rotatae, and sieving to obtain medicinal powder;
B) adding the medicinal material powder into 10-90% ethanol solution according to the solid-to-liquid ratio of 1: 10-1: 50g/m L, soaking for 2-8 h, performing ultrasonic extraction for 10-30 min, and filtering to obtain a total flavone extracting solution;
the ultrasonic extraction conditions of the soaking time, the ethanol volume fraction and the solid-liquid ratio are optimized by a response surface method, and specifically the method comprises the following steps:
i) single factor investigation: extracting total flavone compounds in lamiophlomis rotata by adopting an ultrasonic extraction technology, and respectively designing a single-factor experiment according to soaking time, ethanol volume fraction, solid-liquid ratio, extraction time and ultrasonic power;
ii) on the basis of a single-factor experiment, adopting statistical software Design Expert11, taking the extraction rate of the flavonoid compounds in the lamiophlomis rotata as a response value, selecting soaking time, ethanol volume fraction and solid-liquid ratio which have great influence on the extraction rate of the flavonoid as response factors, and applying a central composite Design experiment Design principle to Design a response surface experiment with a five-factor level;
iii) carrying out ultrasonic extraction on the medicinal material powder to obtain the extraction rate of the flavonoid compounds in the medicinal material under the conditions of soaking time, ethanol volume fraction and solid-to-liquid ratio; and (3) performing multiple regression fitting analysis on the data by using Design Expert11 to obtain the optimal ultrasonic extraction conditions of the flavonoid compounds in the lamiophlomis rotata.
2. The method according to claim 1, wherein the sieving in step a) is specifically a 40 mesh sieving.
3. The method according to claim 1, wherein the number of times of the extraction in the step B) is 1-3.
4. The method according to claim 1, wherein the step iii) comprises: and measuring the content of the flavonoid compounds in the sample solution to be measured and the standard sample solution by using an ultraviolet-visible spectrometer.
5. The method according to claim 1, wherein the step iii) comprises: and (3) applying Design Expert11 to establish a quadratic polynomial equation model of the influence of experimental factors on the response value, and performing variance analysis and significance test on the response surface multiple regression model.
6. The method as claimed in claim 1, wherein the optimal ultrasonic extraction conditions in step iii) are soaking time of 5h, volume fraction of ethanol in the extraction solvent of 57%, and solid-to-liquid ratio of 1:57g/m L.
7. The method as claimed in claim 1, wherein the optimal ultrasonic extraction conditions in step iii) are soaking time of 5.04h, volume fraction of ethanol in the extraction solvent of 56.94%, and solid-to-liquid ratio of 1:57.44g/m L.
8. The method of claim 1, wherein step B) further comprises rotary evaporation to an extract after said filtering.
9. The method of claim 1, wherein the second order polynomial equation for the multiple regression analysis to obtain the extraction rate of lamiophlomone is:
Y＝6.59+0.19A-0.30B+0.68C+0.07AB+0.09AC+0.06BC-0.16A2-0.64B2-0.31C2。
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