CN110118736A - A method of flavones content in optimization -4 soup of sugermuller - Google Patents
A method of flavones content in optimization -4 soup of sugermuller Download PDFInfo
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Abstract
The invention belongs to anaesthetic technical fields, and this method comprises the following steps: the preparation method of -4 soup of sugermuller；Prepare control substance of Rutin solution；Choose maximum wavelength；Draw standard curve；Experiment of single factor takes single_factor method to investigate liquid-to-solid ratio, extraction time, influence of the Extracting temperature to flavones content using flavones content as index；Response phase method optimization technique；Optimized by Orthogonal Test process certification carries out orthonormal design of experiments using time A, temperature B, solid-liquid ratio C as empirical factor；Using response surface and orthonormal design of experiments, optimize extraction conditions, investigate the influence of extraction time, Extracting temperature and liquid-to-solid ratio to flavones content in -4 soup of sugermuller, using flavones yield in -4 soup of sugermuller as index, optimum process condition is that solid-liquid ratio is 1:40, and the time is 120min, 100 DEG C when temperature, it is simple and easy to do using the flavones content in heating water bath circumfluence method increase -4 soup of sugermuller, it is reliable and stable, it is suitable for industrialized production.
Description
Technical field
The invention belongs to anaesthetic technical field, the side of flavones content in specifically a kind of optimization -4 soup of sugermuller
Method.
Background technique
- 4 soup of sugermuller is the formula after being improved on the basis of conventional formulation sugermuller -3.Conventional formulation Soviet Union lattice
It is that Mongolian medicine's clinic commonly uses prescription that wood, which strangles -3, is dialled tri- taste Chinese herbal medicine of 5g by Amomum cardamomum 15g, white lettuce victory 10g and Bi and is formed, has suppression conspicuous
According to the effect of, clinic cures mainly insomnia.Existing document report: volatile oil is won with the white lettuce of Amomum cardamomum-in conventional formulation sugermuller -3
Win Extraction Process of Volatile Oil for the white lettuce of Amomum cardamomum-in -3 side of index optimization sugermuller.Optimal extraction technology can be to greatest extent
Wherein effective ingredient is extracted, so that the inherent quality and clinical therapeutic efficacy in preparation improve, enables the effect maximum of anaesthetic
The performance of limit.Although the material base of anaesthetic compound effect first is that volatile oil, traditional decoction technique boils anaesthetic process
In, it does not extract volatile oil individually and is taken, so being inaccurate by index optimization extraction process of volatile oil.According to document
Report, flavonoids is the effective component in -4 soup of anaesthetic sugermuller, so using flavonoids as index optimization extraction process be compared with
It is reasonable.
Summary of the invention
It is an object of the invention to overcome shortcoming in the prior art, provide a kind of using flavonoids as index optimization Soviet Union
The method that erythrophloeum ferdii strangles flavones content in -4 soup.
In order to achieve the object of the present invention, we will adopt the following technical scheme that and be implemented.
A method of flavones content in optimization -4 soup of sugermuller, it is characterised in that: this method comprises the following steps:
(1), first by semen ziziphi spinosae, Amomum cardamomum, white lettuce victory and Bi dial medicinal materials be respectively crushed into powder, weigh respectively semen ziziphi spinosae,
Amomum cardamomum, white lettuce victory and Bi dial medicinal powder 1.5g, 1.5g, 1.0g, 0.5g, are placed in same round-bottomed flask, are added corresponding times
Several water shakings are mixed, are impregnated, and round-bottomed flask are put into the water-bath of heating water bath reflux unit and are heated to predetermined temperature, and
Predetermined temperature is kept in the stipulated time, receives sample solution: -4 soup of sugermuller, measures absorbance, calculates flavones content.
(2), control substance of Rutin solution is prepared
Accurately weigh rutin standard items 10mg with assay balance, be placed in 100ml volumetric flask, with dehydrated alcohol dissolution and
Constant volume, it is spare.
(3), maximum wavelength is chosen
It pipettes control substance of Rutin solution and each 4ml of sample solution is put into same 25ml volumetric flask, it is 5% that mass fraction, which is added,
NaNO2Solution 1ml is placed 6 minutes after mixing, and the Al (NO that mass fraction is 10% is added3)3Solution 1ml places 6 after mixing
Minute, the NaOH solution 10ml that mass fraction is 4% is added, ultrapure water is then added and is diluted to scale, 15 points are placed after mixing
Clock is placed in visible spectrophotometer and carries out 200~800 nanometers of full wavelength scanners；In addition the chromophoric solution for taking respective volume, adds
Enter ultrapure water and be settled to scale, as blank solution；Record measurement map, to determine maximum absorption wavelength；
(4), standard curve is drawn
Precision pipettes control substance of Rutin solution 0.0,2.0,4.0,6.0,8.0,12.0ml, is respectively placed in 25ml capacity
In bottle, according to NaNO2-Al (NO3)3The colour developing of-NaOH coloration method, it is spare；With visible spectrophotometer in maximum absorption wavelength
Place carries out absorbance measurement, and each concentration of reference substance solution records five groups of absorption values respectively, takes its average value, is missed with reducing
Difference；Using absorbance A as ordinate, rutin concentration C (mg/ml) is abscissa, calibration curve equation are as follows: y=13.187x+
0.0117(R2=0.9981, R=0.9990)；
(5), experiment of single factor
Using flavones content as index, single_factor method is taken to investigate liquid-to-solid ratio, extraction time, Extracting temperature is to flavones content
It influences.
1., influence of the liquid-to-solid ratio to flavones content
According to step, (1) the method heating extraction, fixed extraction time, temperature investigate different liquid-to-solid ratios, contain to flavones
The influence of amount, each experiment are repeated 3 times, and measure absorbance, calculate flavones content；
2., influence of the Extracting temperature to flavones content
According to step, (1) the method heating extraction, fixed liquid-to-solid ratio, time are investigated different Extracting temperatures, are contained to flavones
The influence of amount, each experiment are repeated 3 times, and measure absorbance, calculate flavones content；
3., influence of the extraction time to flavones content
According to step, (1) the method heating extraction, fixed liquid-to-solid ratio, temperature are investigated different extraction times, are contained to flavones
The influence of amount, each experiment are repeated 3 times, and measure absorbance, calculate flavones content；
(6), response phase method optimization technique
According to the central combination design principle of Box-Benhnken, comprehensive experiment of single factor influences as a result, in Dan Yin
Three factors-levels Response surface methodology, analytical factor and level design and response surface analysis side are used on the basis of element experiment
Case and experimental result；
(7), Optimized by Orthogonal Test process certification
Using time A, temperature B, solid-liquid ratio C as empirical factor, L is carried out9(34) orthonormal design of experiments.
Further, the corresponding multiple includes 10 times, 20 times, 30 times, 40 times or 50 times.
Further, the predetermined temperature includes 80 DEG C, 85 DEG C, 90 DEG C, 95 DEG C or 100 DEG C.
Further, the stipulated time includes 30min, 60min, 90min, 120min or 150min.
Further, a length of 510 nanometers of the maximum absorption wave.
Further, influence of the liquid-to-solid ratio to flavones content, according to step (1) the method heating extraction, fixed extraction
Time is 60min, temperature is 100 DEG C, different liquid-to-solid ratios: 10:1,20:1,30:1,40:1 and 50:1 is investigated, to flavones content
Influence, each experiment is repeated 3 times, and measures absorbance, calculates flavones content.
Further, influence of the Extracting temperature to flavones content, according to step (1) the method heating extraction, fixer
Material is investigated different Extracting temperatures: 80 DEG C, 85 DEG C, 90 DEG C, 95 DEG C and 100 DEG C, being contained to flavones than being 40:1, time 60min
The influence of amount, each experiment are repeated 3 times, and measure absorbance, calculate flavones content.
Further, influence of the extraction time to flavones content, according to step (1) the method heating extraction, fixer
Material ratio is 40:1, temperature is 100 DEG C, investigates different extraction times: 30min, 60min, 90min, 120min and 150min, right
The influence of flavones content, each experiment are repeated 3 times, and measure absorbance, calculate flavones content.
Further, the optimal preparation method of -4 soup of sugermuller includes the following steps:
(1) semen ziziphi spinosae, Amomum cardamomum, white lettuce victory and Bi, are dialled into medicinal material and are respectively crushed into powder；
(2) semen ziziphi spinosae, Amomum cardamomum, white lettuce victory and Bi, are weighed respectively according to weight ratio 3:3:2:1 and dials medicinal powder, are placed in same
In one container；
(3) the water shaking that liquid-to-solid ratio is 40:1, is added in the above-described container to mix, impregnate；
(4), container is put into the water-bath of heating water bath reflux unit and is heated to 100 DEG C, and kept in 120min
100 DEG C, receive -4 soup of sugermuller.
Beneficial effect
The present invention uses response surface and L9(34) orthonormal design of experiments, optimize extraction conditions, investigates extraction time, extracts temperature
Degree and influence of the liquid-to-solid ratio to flavones content in -4 soup of sugermuller, according to response face and L9(34) it is orthogonal it is mutual confirmation it is found that liquid
Three kinds of material ratio, temperature and time factors are to when the influence size of flavones content in -4 soup of sugermuller is temperature > C solid-liquid ratio >
Between, using flavones yield in -4 soup of sugermuller as index, optimum process condition is that solid-liquid ratio is 1:40, and the time is 120min, temperature
It is simple and easy to do using the flavones content in heating water bath circumfluence method increase -4 soup of sugermuller 100 DEG C when spending, it is reliable and stable, it fits
In industrialized production.
Detailed description of the invention
Fig. 1 is the contour of time (B) and temperature (C) to extracting flavonoids rate；
Fig. 2 is the three-dimensional response surface design of time (B) and temperature (C) to extracting flavonoids rate；
Fig. 3 is the contour of liquid-to-solid ratio (A) and time (B) to extracting flavonoids rate；
Fig. 4 is the three-dimensional response surface design of solid-liquid ratio (A) and time (B) to extracting flavonoids rate；
Fig. 5 is the contour of solid-liquid ratio (A) and temperature (C) to extracting flavonoids rate；
Fig. 6 is the three-dimensional response surface design of solid-liquid ratio (A) and temperature (C) to extracting flavonoids rate.
Specific embodiment
In conjunction with the accompanying drawings and embodiments, the present invention is described further.
In one embodiment, as shown in figures 1 to 6, the method for flavones content includes such as in a kind of optimization -4 soup of sugermuller
Lower step:
(1), the preparation of -4 soup of sugermuller
Semen ziziphi spinosae, Amomum cardamomum, white lettuce victory and Bi are dialled into medicinal materials first and are respectively crushed into powder, weighs semen ziziphi spinosae, white respectively
Cardamom, white lettuce victory and Bi dial medicinal powder 1.5g, 1.5g, 1.0g, 0.5g be placed in same round-bottomed flask, be added 10,20,30,
40 or 50 times of water shaking is mixed, is impregnated, and round-bottomed flask is put into the water-bath of heating water bath reflux unit and is heated to 100
DEG C, and 100 DEG C of temperature is kept in 120min, and sample solution: -4 soup of sugermuller is received, it is spare；
(2), the preparation of reference substance solution
10 milligrams of rutin standard items are accurately weighed with a ten thousandth assay balance, is placed in 100 milliliters of volumetric flasks, use is anhydrous
Ethyl alcohol dissolution and constant volume, it is spare；
(3), the selection of maximum wavelength
It pipettes control substance of Rutin solution and each 4 milliliters of sample solution is placed in 25 milliliters of volumetric flasks, mass fraction, which is added, is
5% NaNO2It 1 milliliter of solution, is placed 6 minutes after mixing, the Al (NO that mass fraction is 10% is added3)3It 1 milliliter of solution, mixes
It places 6 minutes afterwards, 10 milliliters of NaOH solution that mass fraction is 4% is added, ultrapure water is then added and is diluted to scale, after mixing
It places 15 minutes, is placed in visible spectrophotometer and carries out 200~800 nanometers of full wavelength scanners；In addition the aobvious of respective volume is taken
Color solution is added ultrapure water and is settled to scale, as blank solution.Record measurement map, to determine maximum absorption wavelength；Scanning
The result shows that a length of 510 nanometers of maximum absorption wave；By the maximum absorption band for comparing control substance of Rutin solution and sample solution
Position, both discoveries absorption maximum position is in 510 nanometers, and by 510 nanometers as measurement wavelength is determined, carry out with
Lower experiment；
(4), the drafting of standard curve
Precision pipettes 0.0,2.0,4.0,6.0,8.0,12.0 milliliter of control substance of Rutin solution, is respectively placed in 25 milliliters
In volumetric flask, develop the color according to NaNO2-Al (NO3) 3-NaOH coloration method, it is spare；With visible spectrophotometer at 510 nanometers
Place carries out absorbance measurement, and each concentration of reference substance solution records five groups of absorption values respectively, takes its average value, is missed with reducing
Difference.Using absorbance A as ordinate, rutin concentration C (mg/mL) is abscissa, calibration curve equation are as follows: y=13.187x+
0.0117(R2=0.9981, R=0.9990) draw standard curve；
(5), experiment of single factor
Using flavones content as index, single_factor method is taken to investigate solid-liquid ratio, extraction time, Extracting temperature is to flavones content
It influences.
1., influence of the liquid-to-solid ratio to flavones content, according to method heating extraction under (1) item, the fixed extraction time (60min),
Temperature (100 DEG C), investigates influence of the different liquid-to-solid ratios (10,20,30,40,50 times) to flavones content, and each experiment repeats 3
It is secondary, absorbance is measured, recovery rate is calculated；According to result it is found that solid-liquid ratio be lower than 40 times when, recovery rate with the increase of solid-liquid ratio and
It increases, this is because solvent increases, the flavones extracted is more soluble, loses fewer, but recovery rate when more than 50 times
It is not further added by, has a declining tendency instead；Therefore when Response to selection face method is investigated within selection liquid-to-solid ratio 40:1；
2., influence of the Extracting temperature to flavones content according to method heating extraction under (1) item, fixed liquid-to-solid ratio (40 times), when
Between (60min), investigate influence of the different Extracting temperatures (80 DEG C, 85 DEG C, 90 DEG C, 95 DEG C, 100 DEG C) to flavones content, Mei Geshi
It tests and is repeated 3 times, measure absorbance, calculate recovery rate；It can be obtained according to result, as temperature increases, flavones content continues to increase, root
Heating water bath is used according to practical solution, so selecting 90~100 DEG C of range to be investigated when response phase method is investigated；
3., influence of the extraction time to flavones content according to method heating extraction under (1) item, fixed liquid-to-solid ratio (40 times), temperature
It spends (100 DEG C), investigates influence of the different extraction times (30min, 60min, 90min, 120min, 150min) to flavones content,
Each experiment is repeated 3 times, and measures absorbance, calculates recovery rate；Within the scope of 30min to 150min extraction time, flavones content
It is improved rapidly with the increase of extraction time, but slightly on a declining curve after 120min；To save the time, response phase method is examined
When examining, the range of 60~120min is selected to be investigated；
(6), response phase method optimization technique
According to the central combination design principle of Box-Benhnken, comprehensive experiment of single factor influences as a result, in Dan Yin
Three factors-levels Response surface methodology, analytical factor and level design and response surface analysis side are used on the basis of element experiment
Case and experimental result is shown in results under (8) item.
(7), Optimized by Orthogonal Test process certification
With time (A), temperature (B), solid-liquid ratio (C) for empirical factor, L is carried out9(3 4) orthonormal design of experiments, factor level
And experimental result is shown in results under (8) item.
(8), technical effect
1., response surface result:
1. response surface analysis factor of table and level
2. response surface analysis scheme of table and result
3. general flavone recovery rate regression equation coefficient of table and its significance test
4. general flavone recovery rate regression equation coefficient of table and its significance test
General flavone recovery rate (Y) solid-liquid ratio in -4 soup of 5. sugermuller of table, the time, temperature secondary multinomial regressive model side
Journey
The F value of quadratic regression model is 10.38, P < 0.01, shows that model has reached extremely significant level；Losing quasi- item is mould
Quasi- item P=0.9899 > 0.05 is lost in the variation of data in type, illustrates to lose quasi- item difference not significant, and test exists without pseudo-factor is lost,
It can sufficiently reflect actual conditions, regression model is suitable；The coefficient of determination R of test model2=0.9303, illustrate flavones yield
Result and model prediction result have good consistency, the correction coefficient R of test model2 Adj=0.8406, test result
There is 84.06% influence for being put to the test factor.Therefore, as a result reliably, this model can carry out analysis and pre- to flavones yield result
It surveys.F inspection may determine that influence of the independent variable to dependent variable in regression equation items variance analysis, thus obtain each factor to Huang
The primary and secondary sequence that ketone yield influences is C > B > A, is also known by regression equation and variance analysis, first order B, C are to Huang in model
The influence of ketone yield reaches extremely significant horizontal (P < 0.01)；Influence of the first order D to flavones yield reaches the level of signifiance (P <
0.05)；Quadratic term B in model2、C2, the influence to flavones yield reach extremely significant horizontal (P < 0.01).
The best factor of table 6. and peak response face amount
Using response surface contrived experiment, with the center combination experimental design principle according to Box-Benhnken, final
It is solid-liquid ratio to optimised process is 1:43.9, the time is 120min, 100 DEG C when temperature.
2., orthogonal experiment verification result
7. quadrature analysis factor of table and level
8. Orthogonal experiment results of table
It can be seen that the primary and secondary sequence of factor influenced on flavones content according to the size of R are as follows: B (temperature) > C > (feed liquid
Than) > A (time), optimum combination A3B3C2, i.e. 120min, 100 DEG C, 1:40.
Response phase method is mutually confirmed with orthogonality analysis method, and using flavones yield as index, optimised process is that solid-liquid ratio is 1:40,
Time is 120min, 100 DEG C when temperature.
Claims (9)
1. a kind of method of flavones content in optimization -4 soup of sugermuller, it is characterised in that: this method comprises the following steps:
(1), semen ziziphi spinosae, Amomum cardamomum, white lettuce victory and Bi are dialled into medicinal materials first and is respectively crushed into powder, weigh semen ziziphi spinosae, white respectively
Cardamom, white lettuce victory and Bi dial medicinal powder 1.5g, 1.5g, 1.0g, 0.5g and are placed in same round-bottomed flask, and corresponding multiple is added
Water shaking is mixed, is impregnated, and round-bottomed flask is put into the water-bath of heating water bath reflux unit and is heated to predetermined temperature, and advised
It fixes time interior holding predetermined temperature, receives sample solution: -4 soup of sugermuller, it is spare.
(2), control substance of Rutin solution is prepared
Rutin standard items 10mg is accurately weighed with assay balance, is placed in 100ml volumetric flask, is dissolved with dehydrated alcohol and fixed
Hold, it is spare.
(3), maximum wavelength is chosen
It pipettes control substance of Rutin solution and each 4ml of sample solution is put into same 25ml volumetric flask, it is 5% that mass fraction, which is added,
NaNO2Solution 1ml is placed 6 minutes after mixing, and the Al (NO that mass fraction is 10% is added3)3Solution 1ml places 6 points after mixing
Clock, is added the NaOH solution 10ml that mass fraction is 4%, and ultrapure water is then added and is diluted to scale, places 15 minutes after mixing,
It is placed in visible spectrophotometer and carries out 200~800 nanometers of full wavelength scanners；In addition the chromophoric solution of respective volume is taken, is added
Ultrapure water is settled to scale, as blank solution；Record measurement map, to determine maximum absorption wavelength；
(4), standard curve is drawn
Precision pipettes control substance of Rutin solution 0.0,2.0,4.0,6.0,8.0,12.0ml, is respectively placed in 25ml volumetric flask
In, according to NaNO2-Al (NO3)3The colour developing of-NaOH coloration method, it is spare；With visible spectrophotometer in maximum absorption wave strong point
Absorbance measurement is carried out, each concentration of reference substance solution records five groups of absorption values respectively, takes its average value, misses to reduce
Difference；Using absorbance A as ordinate, rutin concentration C (mg/ml) is abscissa, calibration curve equation are as follows: y=13.187x+
0.0117(R2=0.9981, R=0.9990)；
(5), experiment of single factor
Using flavones content as index, single_factor method is taken to investigate liquid-to-solid ratio, extraction time, shadow of the Extracting temperature to flavones content
It rings.
1., influence of the liquid-to-solid ratio to flavones content
According to step (1) the method heating extraction, fixed extraction time, temperature investigate different liquid-to-solid ratios, to flavones content
It influences, each experiment is repeated 3 times, and measures absorbance, calculates flavones content；
2., influence of the Extracting temperature to flavones content
According to step (1) the method heating extraction, fixed liquid-to-solid ratio, time investigate different Extracting temperatures, to flavones content
It influences, each experiment is repeated 3 times, and measures absorbance, calculates flavones content；
3., influence of the extraction time to flavones content
According to step (1) the method heating extraction, fixed liquid-to-solid ratio, temperature investigate different extraction times, to flavones content
It influences, each experiment is repeated 3 times, and measures absorbance, calculates flavones content；
(6), response phase method optimization technique
According to the central combination design principle of Box-Benhnken, comprehensive experiment of single factor influences as a result, in single factor test reality
Used on the basis of testing Three factors-levels Response surface methodology, analytical factor and level design and response surface analysis scheme and
Experimental result；
(7), Optimized by Orthogonal Test process certification
Using time A, temperature B, solid-liquid ratio C as empirical factor, using flavones content as inspection target, L is carried out9(34) orthogonal design reality
It tests.
2. the method for flavones content in a kind of optimization -4 soup of sugermuller according to claim 1, it is characterised in that: described
Corresponding multiple includes 10 times, 20 times, 30 times, 40 times or 50 times.
3. the method for flavones content in a kind of optimization -4 soup of sugermuller according to claim 1, it is characterised in that: described
Predetermined temperature includes 80 DEG C, 85 DEG C, 90 DEG C, 95 DEG C or 100 DEG C.
4. the method for flavones content in a kind of optimization -4 soup of sugermuller according to claim 1, it is characterised in that: described
Stipulated time includes 30min, 60min, 90min, 120min or 150min.
5. the method for flavones content in a kind of optimization -4 soup of sugermuller according to claim 1, it is characterised in that: described
A length of 510 nanometers of maximum absorption wave.
6. the method for flavones content in a kind of optimization -4 soup of sugermuller according to claim 1, it is characterised in that: described
Influence of the liquid-to-solid ratio to flavones content, according to step (1) the method heating extraction, the fixed extraction time is 60min, temperature is
100 DEG C, different liquid-to-solid ratios: 10:1,20:1,30:1,40:1 and 50:1 are investigated, the influence to flavones content, each experiment repeats 3
It is secondary, absorbance is measured, flavones content is calculated.
7. the method for flavones content in a kind of optimization -4 soup of sugermuller according to claim 1, it is characterised in that: described
Influence of the Extracting temperature to flavones content, according to step (1) the method heating extraction, fixed liquid-to-solid ratio is 40:1, the time is
60min, investigate different Extracting temperatures: 80 DEG C, 85 DEG C, 90 DEG C, 95 DEG C and 100 DEG C, the influence to flavones content, each experiment is again
It is 3 times multiple, absorbance is measured, flavones content is calculated.
8. the method for flavones content in a kind of optimization -4 soup of sugermuller according to claim 1, it is characterised in that: described
Influence of the extraction time to flavones content, according to step (1) the method heating extraction, fixed liquid-to-solid ratio is 40:1, temperature is
100 DEG C, different extraction times: 30min, 60min, 90min, 120min and 150min are investigated, the influence to flavones content, each
Experiment is repeated 3 times, and measures absorbance, calculates flavones content.
9. according to claim 1 in a kind of -8 any -4 soup of optimization sugermuller flavones content method, feature exists
In: the optimal preparation method of -4 soup of sugermuller includes the following steps:
(1), semen ziziphi spinosae, Amomum cardamomum, white lettuce victory and Bi are dialled into medicinal material and is respectively crushed into powder；
(2), semen ziziphi spinosae, Amomum cardamomum, white lettuce victory and Bi are weighed respectively and dials medicinal powder 1.5g, 1.5g, 1.0g, 0.5g, are placed in same
In one round-bottomed flask；
(3), the water shaking that liquid-to-solid ratio is 40:1 is added in the round-bottomed flask to mix, impregnate；
(4), round-bottomed flask is put into the water-bath of heating water bath reflux unit and is heated to 100 DEG C, and kept in 120min
100 DEG C, receive -4 soup of sugermuller.
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