CN109528843A - A kind of extracting method of flavone of hawthorn fruit - Google Patents
A kind of extracting method of flavone of hawthorn fruit Download PDFInfo
- Publication number
- CN109528843A CN109528843A CN201811430230.6A CN201811430230A CN109528843A CN 109528843 A CN109528843 A CN 109528843A CN 201811430230 A CN201811430230 A CN 201811430230A CN 109528843 A CN109528843 A CN 109528843A
- Authority
- CN
- China
- Prior art keywords
- hawthorn
- flavone
- fruit
- liquid nitrogen
- extraction
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
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- CIWBSHSKHKDKBQ-JLAZNSOCSA-N Ascorbic acid Chemical compound OC[C@H](O)[C@H]1OC(=O)C(O)=C1O CIWBSHSKHKDKBQ-JLAZNSOCSA-N 0.000 description 2
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- REFJWTPEDVJJIY-UHFFFAOYSA-N Quercetin Chemical compound C=1C(O)=CC(O)=C(C(C=2O)=O)C=1OC=2C1=CC=C(O)C(O)=C1 REFJWTPEDVJJIY-UHFFFAOYSA-N 0.000 description 2
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- ZPWVASYFFYYZEW-UHFFFAOYSA-L dipotassium hydrogen phosphate Chemical compound [K+].[K+].OP([O-])([O-])=O ZPWVASYFFYYZEW-UHFFFAOYSA-L 0.000 description 1
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- MWDZOUNAPSSOEL-UHFFFAOYSA-N kaempferol Natural products OC1=C(C(=O)c2cc(O)cc(O)c2O1)c3ccc(O)cc3 MWDZOUNAPSSOEL-UHFFFAOYSA-N 0.000 description 1
- CXORMDKZEUMQHX-UHFFFAOYSA-N kermesic acid Chemical compound O=C1C2=C(O)C(O)=CC(O)=C2C(=O)C2=C1C=C(O)C(C(O)=O)=C2C CXORMDKZEUMQHX-UHFFFAOYSA-N 0.000 description 1
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- USHAGKDGDHPEEY-UHFFFAOYSA-L potassium persulfate Chemical compound [K+].[K+].[O-]S(=O)(=O)OOS([O-])(=O)=O USHAGKDGDHPEEY-UHFFFAOYSA-L 0.000 description 1
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Classifications
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K36/00—Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
- A61K36/18—Magnoliophyta (angiosperms)
- A61K36/185—Magnoliopsida (dicotyledons)
- A61K36/73—Rosaceae (Rose family), e.g. strawberry, chokeberry, blackberry, pear or firethorn
- A61K36/734—Crataegus (hawthorn)
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
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Abstract
The invention discloses a kind of extracting methods of flavone of hawthorn fruit, belong to hawthorn deep process technology field.It is 2-5% that this method, which is by hawthorn drying to water content, and hawthorn is put into pulverizer and carries out Freezing smashing, and Freezing smashing is that liquid nitrogen is first filled with into pulverizer, is then crushed again, obtains hawthorn powder；The ethanol solution that volume fraction 70% is added to hawthorn powder carries out ultrasonic extraction, and after extraction, centrifuging and taking supernatant repeats the above steps to the precipitating after centrifugation, supernatant will mix twice up to flavone of hawthorn fruit extracting solution.The present invention carries out broken wall treatment to hawthorn by the way of Freezing smashing, it is aided with ultrasonic technique simultaneously and carries out flavone of hawthorn fruit extraction, the implementation of this method, hawthorn grinding particle size can be improved, and degradation rate is up to 75%, and the recovery rate of flavones is improved to 9.26%, liquid nitrogen pulverization process promotes the dissolution of flavonoid component and has preferable antioxidant effect, and this method is suitable for extracting the flavones hawthorn.
Description
Technical field
The present invention relates to a kind of extracting methods of flavone of hawthorn fruit, belong to hawthorn deep process technology field.
Background technique
Hawthorn (Hawthorn) also known as red fruit, mouse are looked into, category rosaceae, Maloideae, hawthorn, and different regions are different
For example red Chinese date of call, kermes fruit, hawthorn, large-fruited Chinese hawthorn etc., reach as high as 6 meters.Contain a variety of active ingredients in hawthorn, 1950
It is relatively more to the analysis and research of the ingredients such as tannin, cellulose, triterpenes in hawthorn before year, due to plant flavone after the fifties
The fast development of substance, hawthorn are found containing more flavones ingredient, it is to cardio-cerebrovascular and some other
The therapeutic effect that disease has had.So far, people have found and are extracted 150 many kinds of substance from hawthorn, including organic
Acid, flavonoids, flavane substance and its polymer etc..
The products such as preserved fruit fruitcake can be eaten it raw or be made to Fructus Crataegi, and medical value is also very high, be the distinctive medicinal food in China
With all available plant.Hawthorn prevent and treat cardiovascular and cerebrovascular disease, improve heart vigor, reduce blood pressure with blood lipid, in terms of
With very high therapeutic efficiency；The substances such as flavonoids contained by hawthorn and vitamin C can strengthen body immune system, have and prolong
Slow aging, antitumor effect；Hawthorn can accelerate the mobility of blood circulation of human body system, activate blood circulation and disperse blood clots, and hawthorn is to treatment
Asthma, coughing with a lot of sputum, abdominal pain diarrhea have good curative effect.As functional component, Flavonoid substances, be in hawthorn content compared with
High active constituent, in the form of flavonoid glycoside substance exist, lowering blood pressure and blood fat, anticancer, in terms of have it is more
Kind effect all has significance to anti-aging, treatment cardiovascular disease etc..
But industrialization extracts flavone of hawthorn fruit and mostly uses solvent extraction method now, and time-consuming, low output, at high cost, Wu Faman
The sufficient market demand.Further, since the particularity of hawthorn cell wall, causes intracellular nutrition release difficult.And different points
Existing from, extractive technique on flavone of hawthorn fruit bioactivity influences.Therefore, during flavone of hawthorn fruit extracts, good hawthorn
Cell wall breaking technology will be the critical issue for solving flavone of hawthorn fruit extraction effect.Dried hawthorn pre-treatment at present crushes wall breaking technology
The method generallyd use includes: ambient ground, ultramicro grinding and thermophilic digestion in conjunction with alkaline process combined processing, coarse crushing and born of the same parents
Wall dissolves the technologies such as enzyme process combination, in which: ultramicro grinding generally refers to the material particles of 3mm or more being crushed to 10~25 μm
Process；The hawthorn powder that Liu Xiaoguang et al. will crushed extracts flavone of hawthorn fruit with cellulase, and recovery rate is about 5%；It is above-mentioned
All there is extraction process equipment requirement height in method, the expensive of enzyme, reaction condition are just carved, are easy inactivation, and flavones content extracts
The problems such as rate is low.
Summary of the invention
Recovery rate to solve existing flavone of hawthorn fruit extracting method is low, time-consuming, and the expensive of enzyme, reaction condition are careless
It carves, be easy inactivation, the problems such as equipment requirement is high, the present invention provides a kind of extracting method of flavone of hawthorn fruit, the technical sides of use
Case is as follows:
The purpose of the present invention is to provide a kind of extracting method of flavone of hawthorn fruit, the extracting method in accordance with the following steps into
Row:
1) it is 2-5% by dried hawthorn drying to water content, obtains pretreatment hawthorn；
2) the pretreatment hawthorn of step 1) is put into pulverizer and carries out Freezing smashing, Freezing smashing is first into pulverizer
It is filled with liquid nitrogen, is then crushed again, hawthorn powder is obtained；Wherein: liquid nitrogen charge is 5%-20% (v/m), the item of crushing
Part is 20000rpm/min-30000rpm/min, 1min-4min；
3) according to solid-liquid ratio be 1:(10-50 by the hawthorn powder that step 2) obtains) (volume fraction is addition ethanol solution
70%) ultrasonic extraction, is carried out at 40 DEG C -80 DEG C, 100W-300W, extraction time 5-15min after extraction, is centrifuged
Supernatant is taken, is repeated the above steps to the precipitating after centrifugation, supernatant will be mixed twice up to flavone of hawthorn fruit extracting solution.
It is highly preferred that drying hawthorn to water content in step 1) is 2%-5%.
It most preferably, is 2% by hawthorn drying to water content in step 1).
Preferably, step 2) the liquid nitrogen charge is 15% (v/m).
Preferably, the condition of the step 2) crushing is 25000rpm/min, 2min.
Preferably, the step 3) solid-liquid ratio is 1:30.
Preferably, the step 3) ultrasonic extraction is the ultrasonic extraction 9min at 60 DEG C, 180W.
Preferably, the step 3) centrifugation is to be centrifuged 6min at 4500r/min.
The invention has the advantages that:
Extracting method of the present invention is made by the way of liquid nitrogen frozen crushing and in conjunction with subsequent flavone of hawthorn fruit extracting method
For the hawthorn powder of acquisition while reaching low granularity level, crush efficiency is up to 75%, the recovery rate of flavones improve to
9.26%, additionally it is possible to so that extracting obtained flavone of hawthorn fruit has stronger scavenging effect to ABTS+, hydroxyl radical free radical, it is right
ABTS+ clearance rate can reach 87%, the flavone of hawthorn fruit that is reached by 83%, while being obtained for hydroxyl radical free radical clearance rate have compared with
Strong reducing power, therefore present invention energy by way of crushing liquid nitrogen frozen in conjunction with subsequent flavone of hawthorn fruit extracting method
It is enough to solve the problems, such as to improve hawthorn crushing effect (granularity, uniformity etc.) simultaneously, further solve the antioxygen for improving flavone of hawthorn fruit
Change the problem of activity, i.e., the present invention provides one kind to have the active flavone of hawthorn fruit extracting method of high anti-oxidation.
The present invention using Freezing smashing by the way of to handle hawthorn be a kind of physical method, this method is without additionally adding it
His biochemical substances, the addition for avoiding other substances introduce harmful substance；Can to avoid equipment requirement height, enzyme it is expensive,
Reaction condition is just carved, is easy the problems such as inactivation.
The method of the present invention is extracted in the flavone of hawthorn fruit obtained, and rutin content is more in the flavone of hawthorn fruit extracted, anti-oxidant
Effect is better than the hawthorn powder antioxidant effect that other processing modes obtain.
The present invention is wider using the market prospects of flavone of hawthorn fruit development functionality food additives, health care product or drug
It is wealthy, and the health that is beneficial to man, the flavone of hawthorn fruit deep processed product of high financial profit are developed, to promotion flavone of hawthorn fruit
Industry development is of great significance.
Detailed description of the invention
Fig. 1 is different grinding mode hawthorn crushing effects.
The rutin standard curve figure that Fig. 2 is measured by Sodium nitrite-aluminium nitrite method.
Fig. 3 is grinding mode flavone of hawthorn fruit yield.
Fig. 4 is embodiment 1-4 flavone of hawthorn fruit yield.
Fig. 5 is the flavone of hawthorn fruit powder particle diameter distribution map that liquid nitrogen crushes.
Fig. 6 is the flavone of hawthorn fruit powder particle diameter distribution map of ambient ground.
Fig. 7 is the flavone of hawthorn fruit powder particle diameter distribution map pulverized.
Fig. 8 is the flavone of hawthorn fruit powder particle diameter distribution map of liquid nitrogen grinding processing.
Fig. 9 is influence of the Different treatments to the Scavenging activity of ABTS.
Figure 10 is influence of the flavone of hawthorn fruit to the Scavenging activity of ABTS of different component.
Figure 11 is influence of the Different treatments to the Scavenging activity of hydroxy radical.
Figure 12 is influence of the flavone of hawthorn fruit to the Scavenging activity of hydroxy radical of different component.
Figure 13 is influence of the Different treatments to reducing power.
Figure 14 is influence of the flavone of hawthorn fruit of different component to reducing power.
Specific embodiment
The present invention will be further described combined with specific embodiments below, but the present invention should not be limited by the examples.
Experimental material used in following embodiment is as follows:
1, raw material: commercially available net hawthorn, source area Shandong, water content 7%
2, chemical reagent: rutin mark product；Dehydrated alcohol；Sodium nitrite；Aluminum nitrate；Sodium hydroxide；Liquid nitrogen；ABTS；Over cure
Change potassium；Dipotassium hydrogen phosphate；Potassium dihydrogen phosphate；The potassium ferricyanide；Trichloroacetic acid；Ferric trichloride；Ferrous sulfate；30% hydrogen peroxide；
Sodium salicylate.
3, test apparatus and equipment are shown in Table 1:
1 test apparatus of table and equipment
Embodiment 1:
A kind of extracting method of flavone of hawthorn fruit is present embodiments provided, this method carries out in accordance with the following steps:
1) it is 3% by dried hawthorn drying to water content, obtains pretreatment hawthorn；
2) the pretreatment hawthorn of step 1) is put into pulverizer and carries out Freezing smashing, Freezing smashing is first into pulverizer
It is filled with liquid nitrogen, is then crushed again, hawthorn powder is obtained；Wherein: liquid nitrogen charge is 20% (v/m), and the condition of crushing is
30000rpm/min,4min；
3) by step 2) obtain hawthorn powder according to solid-liquid ratio be 1:50 be added volume fraction be 70% ethanol solution,
80 DEG C, ultrasonic extraction is carried out under 300W, extraction time 15min after extraction, takes supernatant in 4500r/min centrifugation 6min
Liquid moves in 100mL volumetric flask, repeats the above steps to the precipitating after centrifugation, supernatant will mix and mention up to flavone of hawthorn fruit twice
Take liquid.
Embodiment 2:
A kind of extracting method of flavone of hawthorn fruit is present embodiments provided, this method carries out in accordance with the following steps:
1) it is 4% by hawthorn drying to water content, obtains pretreatment hawthorn；
2) the pretreatment hawthorn of step 1) is put into pulverizer and carries out Freezing smashing, Freezing smashing is first into pulverizer
It is filled with liquid nitrogen, is then crushed again, hawthorn powder is obtained；Wherein: liquid nitrogen charge is 5% (v/m), and the condition of crushing is
20000rpm/min,1min；
3) by step 2) obtain hawthorn powder according to solid-liquid ratio be 1:10 be added volume fraction be 70% ethanol solution,
40 DEG C, ultrasonic extraction is carried out under 100W, extraction time 5min after extraction, takes supernatant in 4500r/min centrifugation 6min
Liquid moves in 100mL volumetric flask, repeats the above steps to the precipitating after centrifugation, supernatant will mix and mention up to flavone of hawthorn fruit twice
Take liquid.
Embodiment 3:
A kind of extracting method of flavone of hawthorn fruit is present embodiments provided, this method carries out in accordance with the following steps:
1) it is 2% by hawthorn drying to water content, obtains pretreatment hawthorn；
2) the pretreatment hawthorn of step 1) is put into pulverizer and carries out Freezing smashing, Freezing smashing is first into pulverizer
It is filled with liquid nitrogen, is then crushed again, hawthorn powder is obtained；Wherein: liquid nitrogen charge is 15% (v/m), and the condition of crushing is
25000rpm/min, 2min；
3) by step 2) obtain hawthorn powder according to solid-liquid ratio be 1:30 be added volume fraction be 70% ethanol solution,
60 DEG C, ultrasonic extraction is carried out under 180W, extraction time 9min after extraction, takes supernatant in 4500r/min centrifugation 6min
Liquid moves in 100mL volumetric flask, repeats the above steps to the precipitating after centrifugation, supernatant will mix and mention up to flavone of hawthorn fruit twice
Take liquid.
Embodiment 4:
The present embodiment and the difference of embodiment 3 are: drying hawthorn to water content in step 1) is 5%, other steps
It is all the same with embodiment 3.The effect that method can obtain to illustrate the invention has carried out following experiment:
One, experimental group is arranged
Example 1 group: it is carried out according to 1 step of embodiment；
2 groups of embodiment: being carried out according to 2 step of embodiment；
3 groups of embodiment: being carried out according to 3 step of embodiment；
4 groups of embodiment: being carried out according to 4 step of embodiment；
Control group 1: using pulverizing, (Retsch ZM200, condition are to carry out commercially available net hawthorn with Universalpulverizer
Coarse crushing, sieving obtain coarse powder and Freezing smashing and handle the identical time obtaining Ultramicro-powder) replace Freezing smashing, other steps with
Embodiment 3 is identical；
Control group 2: use ambient ground (condition 25000rpm/min, 2min) replace Freezing smashing, other steps with
Embodiment 3 is identical,；
Control group 3: using liquid nitrogen grinding, (condition is sieved for commercially available net hawthorn is carried out coarse crushing with Universalpulverizer
To coarse powder, identical amount of liquid nitrogen, hand-ground when crushing with liquid nitrogen is added and handles to obtain liquid nitrogen grinding powder) Freezing smashing is replaced,
Other steps are same as Example 3.
Two, experimental method
1, crush efficiency measures:
The hawthorn powder obtained to embodiment 1-4 group and control group 1-3 carries out crush efficiency measurement, crosses classifying screen (40 mesh, 60
Mesh, 80 mesh, 100 mesh, 120 mesh), crush efficiency=screenings (g)/initial weight (g) * 100%
2, flavone of hawthorn fruit extraction process process:
Flavone of hawthorn fruit, extraction process process are extracted using ultrasonic wave added alcohol extracting method are as follows: pretreatment of raw material → alcohol steep →
Ultrasound assisted extraction → centrifugation, filtering → flavones content measurement → supernatant concentration → freeze-drying → flavones crude product.
Embodiment 1-4 group and control group 1-3 supernatant are moved in 100mL volumetric flask respectively, it is molten as sample after constant volume
Liquid carries out the measurement of flavonoid concentration, and the measuring method of flavonoid concentration is as follows:
2.1, the extraction of flavone of hawthorn fruit
2.1.1, pretreatment of raw material: dry commercially available net hawthorn is enucleated, crushes in different ways, crosses 120 meshes.
2.1.2, alcohol steep: will smashed hawthorn powder by solid-liquid ratio 1:30 and volume fraction be differently 70%
Ethanol solution mix, stand 30min after, 60 DEG C of heating water bath 20min,
2.1.3, ultrasound assisted extraction: 60 DEG C of Extracting temperature, extraction time 9min, ultrasonic power 180W is extracted.
2.1.4, the centrifugation of extract, concentration: 4500r/min is centrifuged 6min, and supernatant is taken to move in 100mL volumetric flask,
Repeat ultrasonic extraction, supernatant moves in 100mL volumetric flask, constant volume.This is sample solution.
2.2, the measuring method of flavone of hawthorn fruit content
Using sodium nitrite-aluminum nitrate Fructus Crataegi by Colorimetry flavones content.
2.2.1, the preparation of 70% volume fraction ethanol solution: the accurate 350mL dehydrated alcohol that measures pours into 500mL reagent bottle
In, then measure 150mL deionized water and continue to pour into reagent bottle, it shakes, is allowed to mix well to get the solution.
2.2.2, the preparation of standard solution: precision weighs 20mg rutin standard items, is placed in a beaker, and adds a small amount of dehydrated alcohol
It dissolves and is transferred in brown reagent bottle, then pour into remaining dehydrated alcohol, shake, be allowed to mix well to get standard solution.
2.2.3, the preparation of 5% sodium nitrite solution: precision weighs 5g sodium nitrite drug, is placed in a beaker, adds part
Deionized water dissolving, and be transferred in reagent bottle, then pour into remaining deionized water, it shakes, is allowed to mix well molten to get this
Liquid.
2.2.4, the preparation of 10% aluminum nitrate solution: precision weighs 10g aluminum nitrate drug, is placed in a beaker, and part is added to go
Ionized water is dissolved and is transferred in reagent bottle, then pours into remaining deionized water, and concussion is allowed to mix well to get the solution.
2.2.5, the preparation of 4% sodium hydroxide solution: precision weighs 4g sodium hydroxide drug, is placed in a beaker, adds part
Deionized water dissolving is simultaneously transferred in reagent bottle, then pours into remaining deionized water, and concussion is allowed to mix well molten to get this
Liquid.
2.3, the preparation of standard curve
Precision is drawn 0,1,2,3,4,5 and 6ml of standard solution and is respectively placed in 7 25ml colorimetric cylinders, adds dehydrated alcohol extremely
6ml shakes up after adding 5%NaNO2 solution 1ml, stands 6min；Add 3 solution 1ml of 10%Al (NO3), shake up, stands 6min；Add
Enter 4%NaOH solution 10ml, after shaking up plus dehydrated alcohol is to graduation mark, shakes up, stands 15min, is immediately 510nm in OD value
Absorbance value is measured at wavelength, using absorbance as abscissa, standard concentration is that ordinate draws curve.
2.4, sample measures
Flavone extractive 1ml is drawn, by above-mentioned steps time-and-motion study absorbance, with standard curve regression equation calculation flavones
Content.
Constant volume after extracted liquor capacity * is extracted when constant volume/detection when flavones yield (%)=measurement concentration * measurement
Volume/raw material dry weight
3, size distribution
Laser particle analyzes multi-purpose wet process at present, is to test sample dispersion in a solvent.This experiment will not exist together
The hawthorn powder that reason mode obtains is analyzed.
4, in hawthorn flavonoid substance component measurement
The measurement of flavonoid component: it uses high performance liquid chromatography (HPLC), chromatographic condition are as follows: Agilent HPLC-UV high
Effect liquid phase chromatogram instrument, chromatographic column Cosmosil C1 (8 4.6mm × 250mm, 5 μm)；100% methanol of mobile phase A, mobile phase
1% acetic acid of B；Flow velocity 0.6mL/min；Detection wavelength 360nm, 20 μ L of sample volume.Respectively under 30 DEG C, 40 DEG C, 50 DEG C of column temperatures,
To flavone of hawthorn fruit extracting solution, sample introduction is analyzed.
5, antioxidant activity is tested
5.1, the scavenging effect that the flavone of hawthorn fruit of Different treatments and different component tests ABTS free radical
ABTS free radical is a kind of method for detecting total free radical, while being suitable for hydrophily and lipophilic substance.
Stable blue-green radical cation is generated after the oxidation of ABTS free radical phosphate solution, determinand is added to ABTS certainly
By if there is antioxidant content, then being reacted with ABTS free radical phosphate solution, and solution takes off in based phosphates solution
Color.
7.4mmol/L ABTS and 2.6mmol/L potassium peroxydisulfate are mixed in equal volume, are configured to ABTS radical cation
Solution stands 12~16h under room temperature, dark condition, the PBS (phosphate-buffered for being then 7.4 with the pH value of 50mmol/L
Liquid) absorbance of the ABTS radical cation solution at 630nm is adjusted between 0.45~0.55.Take 1.9mL should therewith
The analyte sample fluid of solution and 0.1mL different disposal after 3min, measures its absorbance at 630nm.
Experiment sets three group sample sets, control group, background group.Flavones is replaced with distilled water in control group；In background group with
Phosphate buffer replaces ABTS working solution, separately VC is taken to do positive controls.ABTS free radical scavenging activity is calculated as follows:
E=[A0-(A-Aj)]/A0 × 100%,
It is clearance rate in E formula.A0For control group OD value, A is the OD value of sample sets, AjFor the OD value of background group.
5.2, the scavenging effect that the flavone of hawthorn fruit of Different treatments and different component tests hydroxyl radical free radical
Principle is according to FeSO4With H2O2Reaction generates OH, with the light absorption value table of OH oxidation salicylic acid products therefrom
The number of OH shown, light absorption value is bigger, and OH is more.In the presence of having scavenger, OH can be removed, to prevent centre
The accumulation of product makes the reduction of bigcatkin willow acid oxidation rates, and the absorbance for showing as system reduces, and finally measures polysaccharide to hydroxyl
The clearance rate of free radical.
By the analyte sample fluid of 1mL different disposal, the FeSO of 1mL 9mmol/L4, 1mL 9mmol/L salicylic acid and
The H of 1mL 8.8mmol/L2O2Solution is uniformly mixed, and reacts 30min at 37 DEG C, and subsequent 4000rpm is centrifuged 10min, measure its
Absorbance at 510nm.
Experiment sets three group sample sets, control group, background group.Flavones is replaced with distilled water in control group；In background group with
Distilled water replaces H2O2, separately VC is taken to do positive controls.It is returned to zero with distilled water, measured value, light absorption value is smaller, and elimination effect is better.
OH clearance rate is calculated as follows:
E=[A0-(A-Aj)]/A0 × 100%,
It is clearance rate in E formula.A0For control group OD value, A is the OD value of sample sets, AjFor the OD value of background group.5.3, no
With the flavone of hawthorn fruit reducing power measuring method of processing mode
Principle is sample by the potassium ferricyanide (K3Fe(CN)6) it is reduced into potassium ferrocyanide (K4Fe(CN)6), potassium ferrocyanide
Again with Fe3+Effect is generated ferric ferrocyanide (Prussian blue), is indicated with detecting Prussian blue absorbance at 700nm wavelength
The size of reducing power, absorbance is higher, and the reducing power of sample is stronger.
The analyte sample fluid for taking 1.0mL different disposal, with 2.5mL50mmol/L, pH value be 7.0 phosphate buffer with
And the potassium ferricyanide solution mixing of 2.5mL1%, so that it is stood 20min at 50 DEG C.Tri- chloroethene of 2.5mL10% is added later
Acid, by mixture under conditions of 4000rpm/min centrifugal treating 10min.1mL supernatant is taken, the FeCl with 3mL 0.1%3
Solution mixing.Its absorbance is measured at 700nm.
Experiment sets three group sample sets, control group, background group.Flavones is replaced with distilled water in control group；Separately take VCDo sun
Property control group.Reducing power is calculated as follows:
E=[A-A0]/A × 100%,
It is clearance rate in E formula.A0For control group OD value, A is the OD value of sample sets.
Three, experimental result
1, crushing effect:
The total hawthorn of quality Zhan of the hawthorn powder of granularity < 125 μm in the hawthorn powder that 3 liquid nitrogen frozen grinding mode of embodiment obtains
The 75% of silty amount,
The quality of the hawthorn powder of granularity < 125 μm accounts in the hawthorn powder that embodiment 1,2 and 4 liquid nitrogen frozen grinding modes obtain
48% or so of total hawthorn powder quality,
And the quality of the hawthorn powder of granularity < 125 μm accounts in the hawthorn powder obtained under similarity condition using ambient ground method
The 16% of total hawthorn powder quality,
The 35% of the total hawthorn quality of hawthorn powder quality Zhan of granularity < 125 μm in the hawthorn powder that ultramicro grinding mode obtains,
The 28% of the total hawthorn quality of granularity < 125 μm black fungus powder quality Zhan in the hawthorn powder that liquid nitrogen grinding mode obtains.No
Crush efficiency with grinding mode is as shown in Figure 1.
2, extracting flavonoids result
The drafting of 2.1 rutin standard curves
The rutin standard curve figure that Fig. 2 is measured by sodium nitrite-aluminum nitrate colorimetric method, linear fit equation are y=
0.1834x-0.0148, R2=0.9996, wherein y represents the light absorption value measured under the conditions of wavelength 510nm, and it is dense that x represents rutin
It spends (mg/ml).
Influence of the 2.2 different grinding modes to flavone of hawthorn fruit recovery rate
Hawthorn extracting flavonoids rate is up to 8.31% under 3 liquid nitrogen frozen pulverization conditions of embodiment, embodiment 1,2 and 4 liquid nitrogen frozens
Hawthorn extracting flavonoids rate is up to 7.85% or so under pulverization conditions, and is mentioned under similarity condition using ambient ground method flavone of hawthorn fruit
Taking rate is 7.54%, and pulverizing flavone of hawthorn fruit recovery rate is 6.75%, and liquid nitrogen grinding flavone of hawthorn fruit recovery rate is 7.22%.
Fig. 3 is different grinding mode flavone of hawthorn fruit yield, and compared with Freezing smashing, there are poles for Different treatments flavone of hawthorn fruit yield
Significant difference.It is as shown in Figure 4 using flavone of hawthorn fruit recovery rate obtained by embodiment 1-4.
3, the hawthorn powder particle diameter distribution that different modes are handled
By Fig. 5-8 and table 1, table 2 it is found that when being added to population of samples product 90%, particle diameter distribution that liquid nitrogen crushes
Be 191.3 μm, ambient ground takes second place, be 246.7 μm, liquid nitrogen grinding and pulverize it is then poor, respectively 254.4 μm with
273.9μm.When particle diameter distribution is up to 100 μm, liquid nitrogen pulverization process, when being added to the 67.84% of population of samples product, liquid nitrogen is ground
Mill takes second place, and is the 58.95% of total volume, ambient ground and ultramicro grinding are then poor, respectively 50.29% He of the total volume
53.52%.
1 Different treatments granularmetric analysis result of table
Note: d (0.1) is indicated adding up since minimum grain size, until critical grain when being added to population of samples product 10%
Diameter value, d (0.5), d (0.9) respectively indicate critical grain size value when being added to population of samples product 50% and 90%.
2 Different treatments particle diameter distribution table of table
4, influence of the different disposal method to flavone of hawthorn fruit component
The flavone of hawthorn fruit obtained through four kinds of different disposal methods by efficient liquid phase chromatographic analysis, in flavone of hawthorn fruit mainly at
Dividing has rutin, Vitexin and Quercetin etc..Different treatments have obtained each component proportion of flavone of hawthorn fruit not
Together, the rutin ingredient after liquid nitrogen pulverization process accounts for whole 90% or so, and ambient ground only accounts for whole with liquid nitrogen grinding rutin content
60% or so of body, rutin component content accounts for whole 70% or so after ultramicro crushing treatment, this may be since liquid nitrogen crushes
Processing promotes the dissolution of rutin component, and the result of extraction of remaining processing mode rutin component is bad.
5, antioxidant activity test result
5.1, the scavenging effect that Different treatments flavone of hawthorn fruit tests ABTS free radical
In order to indicate oxidation resistant ability effect, using the identical flavonoid concentration of Different treatments as abscissa, certainly to ABTS
It is that ordinate is mapped by the clearance rate of base.As shown in figure 9, the flavones that liquid nitrogen pulverization process obtains has very ABTS+
Significant scavenging effect, clearance rate reach 87%, and compared with positive control VC group, there was no significant difference.Ultramicro crushing treatment obtains
The flavones arrived is most weak, is 55%.Ambient ground and the Scavenging activity of liquid nitrogen grinding are lower than liquid nitrogen pulverization process to some extent
Obtained flavones, clearance rate reach 62%-66%.The flavone A BTS free radical scavenging ability that ultramicro crushing treatment obtains is weak can
It can be that high-temperature process causes the quality decline of flavones during pulverizing.Liquid nitrogen pulverization process not only ensure that low temperature, but also
Low granularity level can be reached, so flavones quality is good, ABTS free radical scavenging ability is better than other processing group.
Using different component same concentrations as abscissa, the clearance rate to ABTS free radical is that ordinate carries out making Figure 10.Reed
Flavones more than fourth content has significant scavenging effect to ABTS+, reaches 83%；The clearance rate of positive control VC is
85%, for the two compared to there was no significant difference, the low flavones Scavenging activity of rutin content is weaker, is 57%.
5.2, the scavenging effect that Different treatments flavone of hawthorn fruit tests hydroxyl radical free radical
In order to indicate oxidation resistant ability effect, using the identical flavonoid concentration of Different treatments as abscissa, certainly to hydroxyl
It is that ordinate is mapped by the clearance rate of base.As shown in figure 11, the flavones that liquid nitrogen pulverization process obtains tries hydroxyl radical free radical
The scavenging effect tested is most strong, reaches 83%, and the flavones that ultramicro crushing treatment obtains is most weak, ambient ground and liquid nitrogen grinding it is clear
Removing solid capacity takes second place, between 70%-76%.The flavones hydroxyl radical free radical Scavenging activity that ultramicro crushing treatment obtains is weak to may be
High-temperature process causes the quality decline of flavones during ultramicro grinding.Liquid nitrogen pulverization process not only ensure that low temperature, but also can reach
To low granularity level, so flavones quality is good, the scavenging effect of hydroxyl radical free radical test is better than other processing group.
Using different component same concentrations as abscissa, the clearance rate to hydroxy radical free radical is that ordinate is mapped
12.Flavones more than rutin content has significant scavenging effect to hydroxy radical, reaches 72%, positive control VC group it is clear
Except rate reaches 90%, and rutin content it is low flavones Scavenging activity it is weaker, only 56%.
The flavone of hawthorn fruit reducing power measuring method of 5.3 Different treatments
In order to indicate oxidation resistant ability effect, using the identical flavonoid concentration of Different treatments as abscissa, with also proper energy
Power is that ordinate is mapped.As shown in figure 13, the reducing power for the flavones that liquid nitrogen pulverization process obtains is most strong, reaches 93%,
Compared with positive control VC group, there was no significant difference.The flavones that ultramicro crushing treatment obtains is most weak, and only 49%, and room temperature powder
The flavones that broken and liquid nitrogen grinding Scavenging activity is obtained lower than liquid nitrogen pulverization process to some extent.Ultramicro crushing treatment obtains
To flavones reducing power it is weak may be pulverize during high-temperature process cause the quality decline of flavones.Liquid nitrogen powder
Broken processing not only ensure that low temperature, but also can reach low granularity level, so flavones quality is good, reducing power is better than other processing group.
Using different component same concentrations as abscissa, carry out making Figure 14 as ordinate using reducing power.Rutin content is high
Flavones has significant scavenging effect to reducing power, reaches 88%；Compared with positive control VC group, there was no significant difference,
The low flavones Scavenging activity of rutin content is weaker, and only 60%.
Although the present invention has been disclosed in the preferred embodiment as above, it is not intended to limit the invention, any to be familiar with this
The people of technology can do kind of change and modification, therefore protection of the invention without departing from the spirit and scope of the present invention
Range should subject to the definition of the claims.
Claims (7)
1. a kind of extracting method of flavone of hawthorn fruit, which is characterized in that the extracting method carries out in accordance with the following steps:
It is 2-5% by dried hawthorn drying to water content, obtains pretreatment hawthorn；
The pretreatment hawthorn of step 1) is put into pulverizer and carries out Freezing smashing, Freezing smashing is first to be filled with liquid into pulverizer
Then nitrogen is crushed again, obtain hawthorn powder；Wherein: liquid nitrogen charge is 5%-20%(v/m), the condition of crushing is
20000rpm/min-30000rpm/min,1min-4min；
The hawthorn powder that step 2 is obtained is 1:(10-50 according to solid-liquid ratio) ethanol solution (volume fraction 70%) is added,
40 DEG C -80 DEG C, ultrasonic extraction is carried out under 100 W-300 W, extraction time is 5-15 min, after extraction, centrifuging and taking supernatant
Liquid repeats the above steps to the precipitating after centrifugation, supernatant will mix twice up to flavone of hawthorn fruit extracting solution.
2. the method according to claim 1, wherein drying hawthorn to water content in step 1) is 2%.
3. the method according to claim 1, wherein liquid nitrogen charge described in step 2 is 15 %(v/m).
4. the method according to claim 1, wherein the condition of crushing described in step 2 be 25000rpm/min,
2min。
5. the method according to claim 1, wherein solid-liquid ratio described in step 3) is 1:30.
6. the method according to claim 1, wherein ultrasonic extraction described in step 3) is surpassed at 60 DEG C, 180W
Sound extracts 9min.
7. the method according to claim 1, wherein centrifugation described in step 3) is centrifuged at 4500r/min
6min。
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