CN104984035B - A kind of preparation method of fritillaria flower extract - Google Patents
A kind of preparation method of fritillaria flower extract Download PDFInfo
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- CN104984035B CN104984035B CN201510443153.8A CN201510443153A CN104984035B CN 104984035 B CN104984035 B CN 104984035B CN 201510443153 A CN201510443153 A CN 201510443153A CN 104984035 B CN104984035 B CN 104984035B
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Abstract
The present invention relates to a kind of preparation methods of fritillaria flower extract, it is characterised in that and include the following steps: at -65~-35 DEG C to take out fresh fritillary flower after fully charge, be placed in freeze drying equipment, the dry 12-24h at -50~-30 DEG C, 1-20Pa；Dry fritillary flower is crushed, 20-100 mesh is crossed, obtains fritillary flower powder；0.5~5.0g fritillary flower powder is taken, is placed in 50ml supercritical extraction reactor, with CO2For extractant, static extracting is carried out by entrainer of 0.05~5.00v/v% ammonium hydroxide-ethanol solution；55~65 DEG C of extraction temperature, 35~45MPa of pressure, entrainer flow velocity 0.2ml/min；After soaking time 60-120min, by the supercritical CO containing extract2Release, obtains supercritical extract；By extract at 40~50 DEG C, drying is concentrated under reduced pressure into get fritillary flower activity extract is arrived.In extract other than containing alkaloids, also containing the Flavonoid substances not having in fritillaria extract.
Description
Technical field
The present invention relates to a kind of preparation methods of fritillaria flower extract.
Background technique
Fritillaria thunbergii has the effect of clearing heat and eliminating phlegm, powder detoxifcation.It is usually used in treating cough due to wind-heat evil, lung carbuncle larynx numbness, sore swells
The diseases such as poison.Alkaloid in fritillaria thunbergii mainly includes peimine, peiminine, Zhebeinine, zhebeirine, peimisine
With the substances such as different peimine.Currently, alkaloid is mainly extracted from Bulbs of Fritillaria thunbergii Miq.
The method for extracting bioactive ingredients in the existing bulb from fritillaria mainly has: sour water-organic solvent extracts
Method, alcohols-sour water-organic solvent extraction, lipophylic organic solvents extraction method.Extraction process has decoction, dipping, diacolation,
Reflux etc..These extracting methods and technique have all largely used toxic, inflammable and explosive organic solvent, do not only result in production cost
It is excessively high, and have biggish harm to environment and human body；And extraction process is complicated, and impurity content is high in extract.
The source of fritillaria thunbergii is mainly artificial growth.Between the annual 3-4 month, fritillaria thunbergii, which blooms, often plucks fritillary flower or petal
It removes, so that underground bulb obtains more nutrients.That extracts spends general all arbitrarily discarding or burials, causes to waste.
Summary of the invention
The technical problem to be solved by the present invention is to for the prior art status provide it is a kind of from discarded fritillary flower
Extract the preparation method of the fritillaria flower extract of active material.
The technical scheme of the invention to solve the technical problem is: the preparation method of the fritillaria flower extract,
It is characterized in that including the following steps:
Fresh fritillary flower is taken out after fully charge at -65~-35 DEG C, is placed in freeze drying equipment, -50~-
30 DEG C, 1-20Pa 12-24h of lower drying time；
Dry fritillary flower is crushed, 20-100 mesh is crossed, obtains fritillary flower powder；
0.5~5.0g fritillary flower powder is taken, is placed in 50ml supercritical extraction reactor, with CO2For extractant, with 0.05~
5.00v/v% ammonium hydroxide-ethanol solution is that entrainer carries out static extracting；55~65 DEG C of extraction temperature, 35~45MPa of pressure, folder
Band agent flow velocity 0.2ml/min；After soaking time 60-120min, by the supercritical CO containing extract2Release, obtains overcritical
Extract；
By extract at 40~50 DEG C, drying is concentrated under reduced pressure into get fritillary flower activity extract is arrived.
Alternatively, the preparation method of the fritillaria flower extract, it is characterised in that include the following steps:
Fresh fritillary flower is taken out after fully charge at -65~-35 DEG C, is placed in freeze drying equipment, -50~-
30 DEG C, 1-20Pa 12-24h of lower drying time；
Dry fritillary flower is crushed, 20-100 mesh is crossed, obtains fritillary flower powder；
Fritillary flower powder described in 10~100g is taken, 0.05~5.00v/v% of 1-10ml ammonium hydroxide-ethanol solution, stirring is added
Uniformly, it is placed in 500ml supercritical extraction reactor, with CO2It is folder with 0.05~5.00v/v% ammonium hydroxide-ethanol solution for extractant
Band agent progress dynamic extraction, 60~70 DEG C of extraction temperature, 40~50MPa of pressure, entrainer flow velocity 0.5ml/min, CO2Flow velocity
5L/min, extraction time 60-150min；
By supercritical extract at 40~50 DEG C, drying is concentrated under reduced pressure into get fritillary flower activity extract is arrived.
Compared with prior art, preparation method provided by the present invention makes full use of discarded fritillary flower, mentions from fritillary flower
Effective alkaloid component has been taken out, and has also extracted the flavones of high-content, extract is in addition to having effects that alkaloid
Outside, also there is excellent inoxidizability；Both the high production cost in the presence of traditional fritillaria extraction process had been solved, solvent is waved
Hair pollutes the problem of ambient enviroment, and organic solvent usage amount is few in this method extraction process, has largely used nontoxic titanium dioxide
Carbon, it is environmental-friendly；And discarded fritillary flower is made to be fully utilized, the new way using fritillary flower is opened, to filling
Divide and has great importance using living resources.
Detailed description of the invention
Fig. 1 is the LC-MS total ion current figure of extract in the embodiment of the present invention 1；
Fig. 2 is extraction figure of the extract at m/z 432.35 in the embodiment of the present invention 1, and wherein peak 1 is Zhebeinine
Zhebeinine, peak 2 are peimine peimine, and peak 3 is different peimine isoverticine；
Fig. 3 is extraction figure of the extract at m/z 430.33 in the embodiment of the present invention 1, and wherein peak 1 is Peininine
Peiminine, peak 2 are bulb of thunberg fritillary ketone zhebeinone；
The mass spectrogram of extract peimine peimine in Fig. 4 embodiment of the present invention 1；
The mass spectrogram of extract Peininine peiminine in Fig. 5 embodiment of the present invention 1；
The ordinate of Fig. 1 to Fig. 5 is abundance %；
In Fig. 6 embodiment of the present invention 1 in the anti-oxidant test of extract in lard POV value and time relational graph.
Specific embodiment
The present invention will be described in further detail below with reference to the embodiments of the drawings.
Embodiment 1
The preparation method of the fritillaria flower extract includes the following steps:
Fresh fritillary flower is taken out after fully charge at -50 DEG C, is placed in freeze drying equipment, at -35 DEG C, 1Pa
Drying time 20h；
Dry fritillary flower is crushed, 60 meshes is crossed, obtains fritillary flower powder；
5g fritillary flower powder is taken, is placed in 50ml supercritical extraction reactor, with CO2For extractant, with 0.1v/v% ammonium hydroxide-second
Alcoholic solution is that entrainer carries out static extracting；60 DEG C of extraction temperature, pressure 40MPa, entrainer flow velocity 0.2ml/min；When immersion
Between after 120min, by the supercritical CO containing extract2Release, obtains supercritical extract；
By extract at 45 DEG C, drying is concentrated under reduced pressure into get alkaloid is arrived.
Obtained alkaloid is detected.
1, effective component in Detection and Extraction object:
Use Acquity UPLC synapt G2 chromatograph-mass spectrometer；
Chromatographic column: BEH C18 (2.1 × 50mm, 1.7 μm)；
Mobile phase: methanol -10mmol/L ammonium acetate (35: 65), flow velocity: 0.2ml/min, column temperature: 30 DEG C, 2 μ L of sample volume
Using electric spray ion source (ESI), cation acquisition mode, capillary voltage 2.8kV, orifice potential 40V, ion
100 DEG C of source temperature.
Testing result is as shown in Figures 1 to 5.
Detect each activity substance content:
Using quantified by external standard method method, standard curve is done with the concentration and peak area of standard items peimine and Peininine,
The peak area of the respective substance measured in sample is substituted into standard curve to calculate.
After measured, peimine and Peininine total content 20.48-35.59% in the present embodiment extract.
Flavones content uses determined by ultraviolet spectrophotometry, is conventional method.It is summarized as follows:
Rutin standard items 20mg is accurately weighed, 20% ethyl alcohol dissolution is added, is placed in 100mL volumetric flask, constant volume is spare.
It is accurate draw rutin standard solution 0,1.0,2.0,3.0,4.0,5.0mL be respectively placed in 10mL volumetric flask, it is each to be added 30%
Ethyl alcohol makes its volume 5mL；Then 5% NaNO is added2Solution 0.3mL, shakes up, and places 6min；Add 10% Al
(NO3)3Solution 0.3mL, shakes up, and places 6min；It again plus the NaOH solution 4mL of 1mol/L, 30% ethyl alcohol constant volume and shakes up, places
15min。
It is control with practical blank, absorbance is measured at 510nm wavelength.Using absorbance as ordinate, with rutin concentration
For abscissa, calibration curve is drawn, obtains regression equation are as follows: A=0.0379C+0.0003, R2=0.9994.
General flavone content 15.98-29.50% in supercritical extract after measured.Rutin is as determination of total flavonoids
In standard items, rutin with six concentration of concentration (include 0 concentration) then plus color developing agent (NaNO2、Al(NO3)3The colour developing such as NaOH,
With spectrophotometric determination light absorption value, the size of light absorption value with concentration be it is corresponding, straight line equation can be made, then sample
Product add identical chromogenic reagent measurement light absorption value, and general flavone content in sample can be calculated by substituting into equation.
To sum up, the present invention prepared by alkaloid in containing Zhebeinine (zhebeinine), peimine (peimine),
Different peimine (isoverticine) m/z 432.35；Peininine (peiminine), bulb of thunberg fritillary ketone (zhebeinone) m/z
430.33 and Flavonoid substances.
2, extract is tested for the property
The anti-oxidant experiment peroxide value POV of lard is measured according to GB/T 5009.37-2003 method.
Sample 1: 0.10wt%SFE extract is added into 20g pure lard
Sample 2: 0.20wt%SFE extract is added into 20g pure lard
Sample 3: 0.50wt%SFE extract is added into 20g pure lard
Sample 4: 0.05wt% vitamin E is added into 20g pure lard
Sample 5: 0.05wt%BHT (2,6 di tert butyl 4 methyl phenol) is added into 20g pure lard
Each sample is stirred evenly respectively, is placed in accelerated oxidation in 65 DEG C of constant temperature oven, every measuring its POV for 24 hours.Pig
POV value and the relationship of time are as shown in Figure 6 in oil.
As seen from Figure 6, the active material that the present invention is extracted from fritillary flower has significant inoxidizability, Neng Gouxian
Writing reduces lard peroxide value, and can learn that the additional amount of various concentration can inhibit pig according to the curve of sample 1 to sample 3
The generation of peroxide in oil, and with the increase that concentration is added, antioxidant activity enhancing.
Embodiment 2
The preparation method of the fritillaria flower extract includes the following steps:
Fresh fritillary flower is taken out after fully charge at -60 DEG C, is placed in freeze drying equipment, at -40 DEG C, 20Pa
Drying time 15h；
Dry fritillary flower is crushed, 80 meshes is crossed, obtains fritillary flower powder；
Fritillary flower powder described in 100g is taken, 10ml 0.1v/v% ammonium hydroxide-ethanol solution is added, stirs evenly, is placed in
In 500ml supercritical extraction reactor, with CO2For extractant, dynamic extraction is carried out by entrainer of 0.1v/v% ethanol containing ammonia,
65 DEG C of extraction temperature, pressure 45MPa, entrainer flow velocity 0.5ml/min, CO2Flow velocity 5L/min, extraction time 150min；
By supercritical extract at 45 DEG C, drying is concentrated under reduced pressure into get alkaloid is arrived.
Composition detection is carried out to obtained alkaloid and performance test, test method and test result are same as Example 1.
Claims (2)
1. a kind of preparation method of fritillaria flower extract, it is characterised in that include the following steps:
Fresh fritillary flower is taken out after fully charge at -65~-35 DEG C, is placed in freeze drying equipment, -50~-30 DEG C,
Dry 12-24h under 1-20Pa；
Dry fritillary flower is crushed, 20-100 mesh is crossed, obtains fritillary flower powder；
0.5~5.0g fritillary flower powder is taken, is placed in 50ml supercritical extraction reactor, with CO2For extractant, with 0.05~5.00v/
V% ammonium hydroxide-ethanol solution is that entrainer carries out static extracting；55~65 DEG C of extraction temperature, 35~45MPa of pressure, entrainer stream
Fast 0.2ml/min；After soaking time 60-120min, by the supercritical CO containing extract2Release, obtains supercritical extract；
By extract at 40~50 DEG C, drying is concentrated under reduced pressure into get fritillary flower activity extract is arrived.
2. a kind of preparation method of fritillaria flower extract, it is characterised in that include the following steps:
Fresh fritillary flower is taken out after fully charge at -65~-35 DEG C, is placed in freeze drying equipment, -50~-30 DEG C,
1-20Pa 12-24h of lower drying time；
Dry fritillary flower is crushed, 20-100 mesh is crossed, obtains fritillary flower powder；
Fritillary flower powder described in 10~100g is taken, 0.05~5.00v/v% of 1-10ml ammonium hydroxide-ethanol solution is added, stirring is equal
It is even, it is placed in 500ml supercritical extraction reactor, with CO2It is entrainment with 0.05~5.00v/v% ammonium hydroxide-ethanol solution for extractant
Agent progress dynamic extraction, 60~70 DEG C of extraction temperature, 40~50MPa of pressure, entrainer flow velocity 0.5ml/min, CO2Flow velocity 5L/
Min, extraction time 60-150min；
By supercritical extract at 40~50 DEG C, drying is concentrated under reduced pressure into get fritillary flower activity extract is arrived.
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| CN101768160A (en) * | 2010-02-01 | 2010-07-07 | 南京泽朗医药科技有限公司 | Preparation method of sipeimine |
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Non-Patent Citations (2)
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| CO2超临界萃取川贝母游离生物碱工艺研究;张良等;《西华大学学报 自然科学版》;20080131;第27卷(第1期);第39-41页及第3页，尤其是第3节第3段 |
| 浙贝母花、地上茎与鳞茎总生物碱、总皂苷含量测定的比较;陈文君等;《浙江中医药大学学报》;20080731;第32卷(第4期);第530-531页，尤其是第530页左栏第1段、第2.1节，第531页表1、第2.3节、第4.1节 |
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