CN109970593B - The extracting method and its extract of a kind of oat extract and application - Google Patents
The extracting method and its extract of a kind of oat extract and application Download PDFInfo
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P1/00—Drugs for disorders of the alimentary tract or the digestive system
- A61P1/16—Drugs for disorders of the alimentary tract or the digestive system for liver or gallbladder disorders, e.g. hepatoprotective agents, cholagogues, litholytics
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P29/00—Non-central analgesic, antipyretic or antiinflammatory agents, e.g. antirheumatic agents; Non-steroidal antiinflammatory drugs [NSAID]
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P3/00—Drugs for disorders of the metabolism
- A61P3/04—Anorexiants; Antiobesity agents
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P3/00—Drugs for disorders of the metabolism
- A61P3/08—Drugs for disorders of the metabolism for glucose homeostasis
- A61P3/10—Drugs for disorders of the metabolism for glucose homeostasis for hyperglycaemia, e.g. antidiabetics
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P35/00—Antineoplastic agents
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P39/00—General protective or antinoxious agents
- A61P39/06—Free radical scavengers or antioxidants
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P9/00—Drugs for disorders of the cardiovascular system
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- B—PERFORMING OPERATIONS; TRANSPORTING
- B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
- B01D—SEPARATION
- B01D15/00—Separating processes involving the treatment of liquids with solid sorbents; Apparatus therefor
- B01D15/08—Selective adsorption, e.g. chromatography
- B01D15/10—Selective adsorption, e.g. chromatography characterised by constructional or operational features
- B01D15/22—Selective adsorption, e.g. chromatography characterised by constructional or operational features relating to the construction of the column
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- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
- C07C231/00—Preparation of carboxylic acid amides
- C07C231/22—Separation; Purification; Stabilisation; Use of additives
- C07C231/24—Separation; Purification
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- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
- C07C235/00—Carboxylic acid amides, the carbon skeleton of the acid part being further substituted by oxygen atoms
- C07C235/02—Carboxylic acid amides, the carbon skeleton of the acid part being further substituted by oxygen atoms having carbon atoms of carboxamide groups bound to acyclic carbon atoms and singly-bound oxygen atoms bound to the same carbon skeleton
- C07C235/32—Carboxylic acid amides, the carbon skeleton of the acid part being further substituted by oxygen atoms having carbon atoms of carboxamide groups bound to acyclic carbon atoms and singly-bound oxygen atoms bound to the same carbon skeleton the carbon skeleton containing six-membered aromatic rings
- C07C235/38—Carboxylic acid amides, the carbon skeleton of the acid part being further substituted by oxygen atoms having carbon atoms of carboxamide groups bound to acyclic carbon atoms and singly-bound oxygen atoms bound to the same carbon skeleton the carbon skeleton containing six-membered aromatic rings having the nitrogen atom of at least one of the carboxamide groups bound to a carbon atom of a six-membered aromatic ring
Abstract
The present invention relates to the processing of cereal functional component and applied technical fields, and in particular to the extracting method and its extract and application, the anthracene amide of a kind of oat extract are extracted using molecularly imprinted solid phase extraction column.Extracting method of the invention has the advantages that effective component extraction rate is high, impurity content is few, and oat extract anthracene amide can be used for preparing in the drug for the treatment of obesity and its associated disease.
Description
Technical field
The present invention relates to the processing of cereal functional component and applied technical fields, and in particular to a kind of extraction of oat extract
Method and its extract and application.
Background technique
In recent decades, obesity be lead to one of Major risk factors of global health problem, and the generation of the disease with
A series of chronic diseases are closely related, such as type II diabetes, cardiovascular disease, non-alcohol fatty liver (NAFLD), reproduction
With gastrointestinal cancer etc..Obesity be due to metabolic dysfunction and calorie intake exhaust between secular disequilibrium caused by,
Also systemic low grade inflammation and response to oxidative stress can be generated while showing as weight gain.
Food and drug administration's approval only has Orlistat for treating fat related drugs, Belviq and
Lorcaserin is several.However, adverse reaction and high secondary failure rates that these drugs are shown in succession also demonstrate drug
The limitation for the treatment of.Therefore, extracting in natural products, there is the functional component for inhibiting obesity action to be increasingly becoming now
The hot spot of research.To fat relevant metabolic disorder illness such as cardiovascular disease, hypertension and atherosclerosis are answered with oxidation
Swash related.In addition, can partially be implemented by the targeting intervention of oxidative stress transmitting inflammation approach to fat relevant low grade inflammation
Prevention to fat relevant metabolic dysfunction illness.Therefore, by adjusting oxidative stress and inflammation come regulating lipid metabolism
It is a kind of approach that effective treatment is fat.
Contain in oat there are many antioxidant, including a kind of unique nitrogenous phenolic acid analog derivative, by Canada
Scientist Collins (1989) first identified is simultaneously named as avenanthramide.Newest research results show avenanthramide except tool
Have very strong in vitro and other than internal anti-oxidation function, also there are anti-inflammatory, antiproliferative and the multiple biological activities such as antipruritic.
Patent CN200880115107.3 avenanthramide is applied to or prevent with the ectoparasite biological infection of animal or
In the pharmaceutical composition for infecting relevant skin, inflammation, stimulation or allergy etc..Patent CN200680005534.7 is public
The substance with avenanthramide structure has been opened in the application for treating immune allergy, hyper-proliferative and inflammation etc..The above research is said
Bright avenanthramide has application value very outstanding as natural and inflammation inhibitor.But at present for anthracene
The research of amide extracting method is less.
Patent CN201711221474.9 discloses the purposes of a kind of avenanthramide extract and its extracting method and it,
The extracting method of anthracene amide includes the following steps: oat pretreatment, oat germination, germinated oat processing, ultrasonic extraction, purifying,
Anthracene amide is also disclosed in edible oil or high-fat foods as the purposes of natural.But the extracting method is selective
Poor, impurity is more, and there is presently no be used to treat fat and its associated disease research report about avenanthramide.
Molecular imprinting technology is as an emerging technology, the molecularly imprinted polymer (molecularly of synthesis
Imprinted polymers, MIP) have the characteristics that highly selective, high stability, high applicability.Solid phase extraction techniques conduct
A kind of sorption extraction technology is extensive due to it can be well adapted to the actual sample of various composition complexity and less reagent dosage
It uses.The two is combined to the interference matrix that not only can remove in actual sample, but also various complexity can be efficiently solved
The endogenous of actual sample interferes.
Therefore, it is mentioned using a kind of avenanthramide that can solve above-mentioned technical problem of molecular engram solid phase extraction technological development
Method is taken to be very important.
Summary of the invention
There is provided that a kind of effective component extraction rate is high, impurity content is few the purpose of the present invention is overcome the deficiencies in the prior art
Oat extract anthracene amide extracting method.
The present invention is achieved by the following technical programs:
A kind of extracting method of oat extract anthracene amide, the anthracene amide are mentioned using molecularly imprinted solid phase extraction column
It takes.
The extraction process of the oat extract anthracene amide is as follows:
S1: ammonium phosphate is dissolved in the in the mixed solvent of water and ethyl alcohol, obtains mixed solution A；
S2: it stands after being adjusted with acid the pH value to acidity of solution A as extract liquor B；
S3: it is mixed after oat is ground with extract liquor B, is centrifuged after ultrasound, takes supernatant liquor；
S4: the remainder particulate after taking supernatant liquor repeats step S3, is concentrated to give substance C after merging supernatant liquor；
S5: by substance C by molecularly imprinted solid phase extraction column, up to oat extract anthracene amide after elution, elution.
Preferably, in the step S1 molal volume of ammonium phosphate and mixed solvent ratio be 1:300-600mol/L, water and
The volume ratio of ethyl alcohol is 1:3-5, and ethyl alcohol mass fraction is 60-90%.
It is highly preferred that the molal volume of ammonium phosphate and mixed solvent ratio is 1:500mol/L, water and second in the step S1
The volume ratio of alcohol is 1:4, and ethyl alcohol mass fraction is 80%.
Preferably, the acid in the step S2 is phosphoric acid, and the pH value range of solution A is 2.6-3.2.
It is highly preferred that the pH value range of solution A is 2.8 in the step S2.
Preferably, the mass volume ratio of oat and extract liquor is 1:5-20g/mL, ultrasonic time 10- in the step S3
40min, centrifugal rotational speed 2000-2400r/min, centrifugation time 10-15min.
It is highly preferred that the mass volume ratio of oat and extract liquor is 1:10g/mL, ultrasonic time in the step S3
30min, centrifugal rotational speed 2250r/min, centrifugation time 10min.
Preferably, step S3, coextraction 2-4 times are repeated in the step S4.
It is highly preferred that step S3 is repeated in the step S4, coextraction 3 times.
The molecularly imprinted solid phase extraction column preparation process is as follows:
(1) preparation of anthracene amide:
S1: p-Coumaric Acid is dissolved in pyridine, and acetic anhydride is added, and then plus water cooling is hidden, and collects precipitate A；
S2: precipitate A is washed with water, then DMF and triethylamine are added in precipitate A, obtains mixing molten for dry, placement
Liquid B；
S3: being dissolved in methylene chloride for BOP, is added in the mixed solution B under conditions of ice-water bath, obtains solution C；
S4: 2- amino -5- hydroxybenzoic acid is dissolved in DMF, is added in the solution C, stirs under conditions of ice-water bath
It mixes and is stored at room temperature, then react to obtain solution D with hydrochloric acid termination again, stand；
S5: extracting solution D with organic solvent, be concentrated to give substance E, and two containing pyrrolidines are added in substance E
Then chloromethanes solution reaction is added hydrochloric acid and terminates reaction, finally extracted with organic solvent, be concentrated and dried, obtain anthracene acyl
Amine solid powder；
(2) preparation of covalent organic frame material:
S1: under a nitrogen atmosphere, dry phloroglucin and methenamine being added in trifluoroacetic acid, and heating carries out anti-
It answers；
S2: hydrochloric acid is added in the solution after above-mentioned reaction, and heating continues to react, and is cooled to after complete reaction
Room temperature；
S3: being extracted with organic solvent, is concentrated, is dried, and a kind of substance TP is obtained；
S4: TP and p-phenylenediamine are added in acetic acid, mixed solvent is made with 1,3,5-trihydroxybenzene and dioxane and is also added to vinegar
In acid, heating reaction is dried in vacuo after complete reaction, obtains covalent organic frame material；
(3) preparation of the amine-modified Mn-ZnS quantum dot of polyethyleneimine:
S1: by ZnSO4·7H2O and Mn (CH3COO)2Mixed dissolution Yu Shuizhong, and polyethyleneimine is dissolved in the water,
Two kinds of solution are mixed to get solution A；
S2: under inert gas conditions, it is adjusted with acid the pH value of solution A, obtains solution B；
S3: under inert gas conditions by Na2S·9H2O is dissolved in the water, and is added in solution B and reacts, obtains solution C；
S4: dehydrated alcohol is added in solution C, and centrifugation obtains supernatant liquor and lower sediment, takes the lower sediment to be
The amine-modified Mn-ZnS quantum dot of polyethyleneimine；
(4) preparation of molecularly imprinted polymer:
S1: the polyethyleneimine prepared in the covalent organic frame material prepared in methanol, step (2), step (3) is repaired
The Mn-ZnS quantum dot mixing of decorations is reacted；
S2: the anthracene amide and ethylene glycol dimethacrylate prepared in step (1) is added, it is different to be eventually adding azo two
Butyronitrile, heating continue to react, and obtain solid F；
S3: carrying out soxhlet type after solid F is ground up, sieved, up to molecularly imprinted polymer after drying；
(5) molecularly imprinted polymer fills out column: the molecularly imprinted polymer being prepared in step (4) is added to solid phase extraction
It takes in void column, does not release the polymer in extraction column to get molecularly imprinted solid phase extraction column.
Preferably, in the preparation process of the amine-modified Mn-ZnS quantum dot of step (3) polyethyleneimine, used in step S2
Acid is hydrochloric acid, combines auxiliary dissolution using microwave and ultrasound.
It is highly preferred that step S2 is adopted in the preparation process of the amine-modified Mn-ZnS quantum dot of step (3) polyethyleneimine
With microwave ultrasonic wave combinatorial compound/abstraction instrument auxiliary dissolution.
Preferably, in the step (1) in the preparation process of anthracene amide:
S1: the p-Coumaric Acid of 1-10mmol is dissolved in the pyridine of 1-20mL, the acetic anhydride of 10-30mL is added, instead
3-8h between seasonable, is then added the water of 40-100mL, and 0-4 DEG C of the temperature of water refrigerates, 2-6 DEG C of refrigerated storage temperature, cold preservation time 1-
2h collects precipitate A；
S2: precipitate A is cleaned 2-4 times with 2-6 DEG C of water, then 5- is added in dry, placement 12-24h in precipitate A
The triethylamine of the DMF and 1-10mL of 40mL, obtain mixed solution B；
S3: the BOP of 1-10mmol is dissolved in the methylene chloride of 1-20mL, with 1-2 drop/s under conditions of ice-water bath
Rate is added drop-wise to dropwise in the mixed solution B, obtains solution C；
S4: the 2- amino -5- hydroxybenzoic acid of 1-10mmol is dissolved in the DMF of 10-80mL, under conditions of ice-water bath
It is added drop-wise in the solution C dropwise with 1-2 drop/s rate, stirs 20-40min, be stored at room temperature 1-3h, then use 50- again
The 0.5M hydrochloric acid of 150mL terminates reaction, obtains solution D, stands, 3-6 DEG C of dwell temperature, time of repose 12-24h；
S5: solution D is extracted with the organic solvent of 50-300mL, is concentrated to give substance E, is added and contains in substance E
The 5-30mL dichloromethane solution of volume fraction 3-6% pyrrolidines reacts, reaction time 20-40min, is then added 5-40mL's
1M hydrochloric acid terminates reaction, is finally extracted with the organic solvent of 50-200mL, is concentrated and dried, obtains anthracene amide solid powder.
Preferably, in the step (2) in the preparation process of covalent organic frame material:
S1: under a nitrogen atmosphere, dry 4-10g phloroglucin and 10-20g methenamine are added to 60-150mL's
In trifluoroacetic acid, heating is reacted, and 90-120 DEG C of heating temperature, reaction time 2-4h；
S2: the 3M hydrochloric acid of 100-300mL is added in the solution after above-mentioned reaction, and heating continues to react, and is heated
90-120 DEG C of temperature, reaction time 1-2h is cooled to room temperature after complete reaction；
S3: being extracted with the organic solvent of 200-500mL, is concentrated, is dried, and a kind of substance TP is obtained；
S4: the p-phenylenediamine of the TP of 50-100mg and 30-60mg is added in the 3M acetic acid of 60-100mL, uses 30-50mL
1,3,5-trihydroxybenzene and the dioxane of 40-80mL make mixed solvent and be also added in acetic acid, heating reaction, heating temperature 110-
130 DEG C, heating time 70-80h is dried in vacuo after complete reaction, obtains covalent organic frame material.
Preferably, in the step (3) in the preparation process of the amine-modified Mn-ZnS quantum dot of polyethyleneimine:
S1: by 250-350mg ZnSO4·7H2O and 5-25mg Mn (CH3COO)2Mixed dissolution in 5-15mL water, and
700-800mg polyethyleneimine is dissolved in 10-20mL water, two kinds of solution are mixed to get solution A；
S2: under inert gas conditions, super using microwave with the pH value of the hydrochloric acid conditioning solution A of 6mol/L to 2.5-5.5
Sound wave combinatorial compound/abstraction instrument auxiliary dissolution, is arranged 50-80 DEG C of temperature, microwave power 400-700W, ultrasonic power 800-
1200W, dissolution time 10-30min, obtains solution B；
S3: under inert gas conditions by the Na of 0.2-1mmol2S·9H2O is dissolved in 1-10mL water, is added in solution B
Reaction, reaction time 0.3-1h obtain solution C；
S4: the dehydrated alcohol of 5-15mL is added in solution C, centrifugation, centrifugation rate 5000-7000r/min, when centrifugation
Between 5-20min, obtain supernatant liquor and lower sediment, taking lower sediment is the amine-modified Mn-ZnS quantum dot of polyethyleneimine.
Preferably, in the step (4) in the preparation process of molecularly imprinted polymer:
S1: will be in the covalent organic frame material that prepared in 2-10mL methanol, 6-20mg step (2), 5-30mL step (3)
The amine-modified Mn-ZnS quantum dot mixing of the polyethyleneimine of preparation is reacted, reaction time 0.5-1h；
S2: adding the anthracene amide and 0.1-3mL ethylene glycol dimethacrylate prepared in 200-400g step (1),
It is eventually adding 30-80mg azodiisobutyronitrile, heating continues to react, and 60-80 DEG C of heating temperature, reaction time 20-30h,
Obtain solid F；
S3: soxhlet type, extraction times 10-20h, molecular engram polymerization obtained by drying are carried out after solid F is ground up, sieved
Object.
The present invention relates to the oat extract anthracene amides that the above-mentioned extracting method of any one is extracted.
The present invention relates to the applications for the oat extract anthracene amide that any one said extracted method is extracted, which is characterized in that
The oat extract anthracene amide is used to prepare in the drug for the treatment of obesity and its associated disease, and associated disease includes II type glycosuria
Disease, cardiovascular disease, non-alcohol fatty liver, reproduction and gastrointestinal cancer etc..
The beneficial effects of the present invention are:
The present invention is handled oat, first before using molecularly imprinted solid phase extraction column extraction avenanthramide
The oat after grinding is extracted with the extract liquor of preparation, and with ultrasound on extracting, centrifugation, concentration, as the pH of extract liquor
When value range is 2.6-3.2, effect of extracting is preferable, handles oat with the method, is conducive to remove most of impurity, keeps oat fast
Speed passes through molecularly imprinted solid phase extraction column and realizes molecularly imprinted solid phase extraction column to effective component anthracene amide with high selectivity
Absorption improves extraction efficiency.
The present invention is combined using molecular engram and solid phase extraction techniques prepares molecularly imprinted solid phase extraction column, for extracting swallow
Anthracene amide in wheat can not only remove the interference matrix in oat, but also can efficiently solve in oat complicated ingredient
Endogenous interference, realizes the absorption with high selectivity to effective component anthracene amide, greatly improves the recovery rate of anthracene amide
And separation rate, and the anthracene amide impurities content extracted is less.
The present invention prepares molecularly imprinted polymer in such a way that covalent organic frame material and molecular engram combine, altogether
Valence organic framework materials stable structure, thermal stability are strong, combine with molecularly imprinted polymer and are capable of providing more adsorption potentials
The stability for putting and reinforcing integral material is conducive to the enrichment of anthracene amide in the removing and oat of template molecule, further increases
The extraction efficiency of anthracene amide, reduces impurity content.
Anthracene amide has the function of stronger anti-oxidant and anti-inflammatory, can treat fat and its associated disease according to this,
Compared with existing drug, anthracene amide is natural grain extract, nontoxic, has both ensured the validity of its weight-reducing, and energy
It enough prevents from causing this side effect of insulin resistance.
Specific embodiment
Embodiment 1
The preparation of molecularly imprinted solid phase extraction column:
(1) preparation of anthracene amide:
S1: the p-Coumaric Acid of 1mmol is dissolved in the pyridine of 1mL, is added the acetic anhydride of 10mL, reaction time 3h,
Then the water of 40mL, 0 DEG C of the temperature of water is added, refrigeration, collects precipitate A by 2 DEG C of refrigerated storage temperature, cold preservation time 1h；
S2: precipitate A is cleaned 2 times with 2 DEG C of water, dry, place 12h, be then added in precipitate A 5mL DMF and
The triethylamine of 1mL obtains mixed solution B；
S3: the BOP of 1mmol is dissolved in the methylene chloride of 1mL, under conditions of ice-water bath dropwise with 1 drop/s rate
It is added drop-wise in the mixed solution B, obtains solution C；
S4: 1mmol 2- amino -5- hydroxybenzoic acid is dissolved in the DMF of 10mL, with 1 drop/s under conditions of ice-water bath
Rate be added drop-wise in the solution C dropwise, stir 20min, be stored at room temperature 1h, then terminated again with the 0.5M hydrochloric acid of 50mL anti-
It answers, obtains solution D, stand, 3 DEG C of dwell temperature, time of repose 12h；
S5: extracting solution D with the organic solvent of 50mL, be concentrated to give substance E, is added in substance E and contains volume
The 5mL dichloromethane solution of 3% pyrrolidines of score reacts, reaction time 20min, and the 1M hydrochloric acid that 5mL is then added terminates reaction,
It is finally extracted with the organic solvent of 50mL, is concentrated and dried, obtains anthracene amide solid powder.
(2) preparation of covalent organic frame material:
S1: under a nitrogen atmosphere, dry 4g phloroglucin and 10g methenamine are added to the trifluoroacetic acid of 60mL
In, heating is reacted, and 90 DEG C of heating temperature, reaction time 2h；
S2: the 3M hydrochloric acid of 100mL is added in the solution after above-mentioned reaction, and heating continues to react, heating temperature
90 DEG C, reaction time 1h is cooled to room temperature after complete reaction；
S3: being extracted with the organic solvent of 200mL, is concentrated, is dried, and a kind of substance TP is obtained；
S4: the p-phenylenediamine of the TP of 50mg and 30mg being added in the 3M acetic acid of 60mL, with the 1,3,5-trihydroxybenzene of 30mL and
The dioxane of 40mL makees mixed solvent and is also added in acetic acid, heating reaction, and 110 DEG C of heating temperature, heating time 70h, to
It is dried in vacuo after fully reacting, obtains covalent organic frame material.
(3) preparation of the amine-modified Mn-ZnS quantum dot of polyethyleneimine:
S1: by 250mg ZnSO4·7H2O and 5mg Mn (CH3COO)2Mixed dissolution is in 5mL water, and by the poly- second of 700mg
Alkene imines is dissolved in 10mL water, and two kinds of solution are mixed to get solution A；
S2: it under the conditions of argon gas, with the pH value of the hydrochloric acid conditioning solution A of 6mol/L to 2.5, is combined using microwave ultrasonic wave
Synthesis/abstraction instrument auxiliary dissolution, is arranged temperature 50 C, microwave power 400W, ultrasonic power 800W, dissolution time 10min is obtained
To solution B；
S3: by the Na of 0.2mmol under the conditions of argon gas2S·9H2O is dissolved in 1mL water, is added in solution B and is reacted, and is reacted
Time 0.3h, obtains solution C；
S4: the dehydrated alcohol of 5mL is added in solution C, centrifugation, centrifugation rate 5000r/min, centrifugation time 5min,
Supernatant liquor and lower sediment are obtained, taking lower sediment is the amine-modified Mn-ZnS quantum dot of polyethyleneimine.
(4) preparation of molecularly imprinted polymer:
S1: poly- by what is prepared in the covalent organic frame material prepared in 2mL methanol, 6mg step (2), 5mL step (3)
The Mn-ZnS quantum dot mixing of aziridine modification is reacted, reaction time 0.5h；
S2: adding the anthracene amide and 0.1mL ethylene glycol dimethacrylate prepared in 200g step (1), finally plus
Enter 30mg azodiisobutyronitrile, heating continues to react, and 60 DEG C of heating temperature, reaction time 20h obtains solid F；
S3: soxhlet type, extraction times 10h, molecularly imprinted polymer obtained by drying are carried out after solid F is ground up, sieved.
(5) molecularly imprinted polymer fills out the preparation of column: the molecularly imprinted polymer being prepared in step (4) is added to
In Solid Phase Extraction void column, do not release the polymer in extraction column to get molecularly imprinted solid phase extraction column.
Molecularly imprinted solid phase extraction column extracts oat extract anthracene amide:
S1: 1mmol ammonium phosphate is dissolved in the in the mixed solvent of 300mL water and 60% ethyl alcohol, the body of water and 60% ethyl alcohol
Product obtains mixed solution A than being 1:3；
S2: the pH value for using phosphoric acid to adjust solution A is stood to after 2.6 as extract liquor B；
S3: it will be mixed after the grinding of 1g oat with 5mL extract liquor B, be centrifuged 10min, centrifugal rotational speed again after ultrasonic 10min
2000r/min takes supernatant liquor；
S4: the remainder particulate after taking supernatant liquor is repeated 2 times step S3, is concentrated into relative density after merging supernatant liquor
1.20 obtaining substance C；
S5: by substance C by molecularly imprinted solid phase extraction column, up to oat extract anthracene amide after elution, elution.
Embodiment 2
Identical with the preparation process of molecularly imprinted solid phase extraction column in embodiment 1, molecularly imprinted solid phase extraction column extracts swallow
Wheat extract anthracene amide process is as follows:
S1: 1mmol ammonium phosphate is dissolved in the in the mixed solvent of 600mL water and 90% ethyl alcohol, the body of water and 90% ethyl alcohol
Product obtains mixed solution A than being 1:5；
S2: the pH value for using phosphoric acid to adjust solution A is stood to after 3.2 as extract liquor B；
S3: it will be mixed after the grinding of 1g oat with 20mL extract liquor B, be centrifuged 15min, centrifugal rotational speed again after ultrasonic 40min
2400r/min takes supernatant liquor；
S4: the remainder particulate after taking supernatant liquor is repeated 4 times step S3, is concentrated into relative density after merging supernatant liquor
1.40 obtaining substance C；
S5: by substance C by molecularly imprinted solid phase extraction column, up to oat extract anthracene amide after elution, elution.
Embodiment 3
Identical with the preparation process of molecularly imprinted solid phase extraction column in embodiment 1, molecularly imprinted solid phase extraction column extracts swallow
Wheat extract anthracene amide process is as follows:
S1: 1mmol ammonium phosphate is dissolved in the in the mixed solvent of 450mL water and 80% ethyl alcohol, the body of water and 80% ethyl alcohol
Product obtains mixed solution A than being 1:4；
S2: the pH value for using phosphoric acid to adjust solution A is stood to after 2.8 as extract liquor B；
S3: it will be mixed after the grinding of 1g oat with 10mL extract liquor B, be centrifuged 10min, centrifugal rotational speed again after ultrasonic 30min
2250r/min takes supernatant liquor；
S4: the remainder particulate after taking supernatant liquor is repeated 3 times step S3, is concentrated into relative density after merging supernatant liquor
1.32 obtaining substance C；
S5: by substance C by molecularly imprinted solid phase extraction column, up to oat extract anthracene amide after elution, elution.
Embodiment 4
It is identical with the molecularly imprinted solid phase extraction column extraction process of oat extract anthracene amide in embodiment 3, molecule used
The preparation process of trace solid-phase extraction column is as follows:
(1) preparation of anthracene amide:
S1: the p-Coumaric Acid of 10mmol is dissolved in the pyridine of 20mL, and the acetic anhydride of 30mL, reaction time is added
8h, is then added the water of 100mL, 4 DEG C of the temperature of water, and refrigeration, collects precipitate A by 6 DEG C of refrigerated storage temperature, cold preservation time 2h；
S2: precipitate A is cleaned 4 times with 6 DEG C of water, it is dry, place the DMF that 40mL is then added in precipitate A for 24 hours and
The triethylamine of 10mL obtains mixed solution B；
S3: the BOP of 10mmol is dissolved in the methylene chloride of 20mL, under conditions of ice-water bath with 2 drops/s rate by
It is added drop-wise in the mixed solution B, obtains solution C；
S4: 10mmol 2- amino -5- hydroxybenzoic acid is dissolved in the DMF of 80mL, under conditions of ice-water bath with 2 drop/
The rate of s is added drop-wise to dropwise in the solution C, is stirred 40min, is stored at room temperature 3h, is then terminated again with the 0.5M hydrochloric acid of 150mL
Reaction, obtains solution D, stands, 6 DEG C of dwell temperature, time of repose is for 24 hours；
S5: extracting solution D with the organic solvent of 300mL, be concentrated to give substance E, is added in substance E and contains volume
The 30mL dichloromethane solution of 6% pyrrolidines of score reacts, reaction time 40min, and the 1M hydrochloric acid that 40mL is then added terminates instead
It answers, is finally extracted with the organic solvent of 200mL, be concentrated and dried, obtain anthracene amide solid powder.
(2) preparation of covalent organic frame material:
S1: under a nitrogen atmosphere, dry 10g phloroglucin and 20g methenamine are added to the trifluoroacetic acid of 150mL
In, heating is reacted, and 120 DEG C of heating temperature, reaction time 4h；
S2: the 3M hydrochloric acid of 300mL is added in the solution after above-mentioned reaction, and heating continues to react, heating temperature
120 DEG C, reaction time 2h is cooled to room temperature after complete reaction；
S3: being extracted with the organic solvent of 500mL, is concentrated, is dried, and a kind of substance TP is obtained；
S4: the p-phenylenediamine of the TP of 100mg and 60mg being added in the 3M acetic acid of 100mL, with the 1,3,5-trihydroxybenzene of 50mL and
The dioxane of 80mL makees mixed solvent and is also added in acetic acid, heating reaction, and 130 DEG C of heating temperature, heating time 80h, to
It is dried in vacuo after fully reacting, obtains covalent organic frame material.
(3) preparation of the amine-modified Mn-ZnS quantum dot of polyethyleneimine:
S1: by 350mg ZnSO4·7H2O and 25mg Mn (CH3COO)2Mixed dissolution gathers in 15mL water, and by 800mg
Aziridine is dissolved in 20mL water, and two kinds of solution are mixed to get solution A；
S2: it under the conditions of argon gas, with the pH value of the hydrochloric acid conditioning solution A of 6mol/L to 5.5, is combined using microwave ultrasonic wave
Synthesis/abstraction instrument auxiliary dissolution, is arranged 80 DEG C of temperature, microwave power 700W, ultrasonic power 1200W, dissolution time 30min,
Obtain solution B；
S3: by the Na of 1mmol under the conditions of argon gas2S·9H2O is dissolved in 10mL water, is added in solution B and is reacted, and is reacted
Time 1h, obtains solution C；
S4: the dehydrated alcohol of 15mL is added in solution C, centrifugation, centrifugation rate 7000r/min, centrifugation time
20min obtains supernatant liquor and lower sediment, and taking lower sediment is the amine-modified Mn-ZnS quantum dot of polyethyleneimine.
(4) preparation of molecularly imprinted polymer:
S1: by what is prepared in the covalent organic frame material prepared in 10mL methanol, 20mg step (2), 30mL step (3)
The amine-modified Mn-ZnS quantum dot mixing of polyethyleneimine is reacted, reaction time 1h；
S2: the anthracene amide and 3mL ethylene glycol dimethacrylate prepared in 400g step (1) is added, is eventually adding
80mg azodiisobutyronitrile, heating continue to react, and 80 DEG C of heating temperature, reaction time 30h obtains solid F；
S3: soxhlet type, extraction times 20h, molecularly imprinted polymer obtained by drying are carried out after solid F is ground up, sieved.
(5) molecularly imprinted polymer fills out the preparation of column: the molecularly imprinted polymer being prepared in step (4) is added to
In Solid Phase Extraction void column, do not release the polymer in extraction column to get molecularly imprinted solid phase extraction column.
Embodiment 5
It is identical with the molecularly imprinted solid phase extraction column extraction process of oat extract anthracene amide in embodiment 3, molecule used
The preparation process of trace solid-phase extraction column is as follows:
(1) preparation of anthracene amide:
S1: the p-Coumaric Acid of 5mmol is dissolved in the pyridine of 10mL, is added the acetic anhydride of 20mL, reaction time 5h,
Then the water of 70mL, 2 DEG C of the temperature of water is added, refrigeration, collects precipitate A by 4 DEG C of refrigerated storage temperature, cold preservation time 1.5h；
S2: precipitate A is cleaned 3 times with 4 DEG C of water, dry, place 18h, be then added in precipitate A 30mL DMF and
The triethylamine of 5mL obtains mixed solution B；
S3: the BOP of 5mmol is dissolved in the methylene chloride of 10mL, under conditions of ice-water bath dropwise with 2 drops/s rate
It is added drop-wise in the mixed solution B, obtains solution C；
S4: 5mmol2- amino -5- hydroxybenzoic acid is dissolved in the DMF of 40mL, with 2 drops/s under conditions of ice-water bath
Rate be added drop-wise in the solution C dropwise, stir 30min, be stored at room temperature 2h, then terminated again with the 0.5M hydrochloric acid of 100mL
Reaction, obtains solution D, stands, 4 DEG C of dwell temperature, time of repose 18h；
S5: extracting solution D with the organic solvent of 200mL, be concentrated to give substance E, is added in substance E and contains volume
The 20mL dichloromethane solution of 5% pyrrolidines of score reacts, reaction time 30min, and the 1M hydrochloric acid that 25mL is then added terminates instead
It answers, is finally extracted with the organic solvent of 130mL, be concentrated and dried, obtain anthracene amide solid powder.
(2) preparation of covalent organic frame material:
S1: under a nitrogen atmosphere, dry 7g phloroglucin and 15g methenamine are added to the trifluoroacetic acid of 100mL
In, heating is reacted, and 100 DEG C of heating temperature, reaction time 3h；
S2: the 3M hydrochloric acid of 200mL is added in the solution after above-mentioned reaction, and heating continues to react, heating temperature
110 DEG C, reaction time 1.5h is cooled to room temperature after complete reaction；
S3: being extracted with the organic solvent of 300mL, is concentrated, is dried, and a kind of substance TP is obtained；
S4: the p-phenylenediamine of the TP of 70mg and 45mg being added in the 3M acetic acid of 80mL, with the 1,3,5-trihydroxybenzene of 40mL and
The dioxane of 60mL makees mixed solvent and is also added in acetic acid, heating reaction, and 120 DEG C of heating temperature, heating time 75h, to
It is dried in vacuo after fully reacting, obtains covalent organic frame material.
(3) preparation of the amine-modified Mn-ZnS quantum dot of polyethyleneimine:
S1: by 300mg ZnSO4·7H2O and 15mg Mn (CH3COO)2Mixed dissolution gathers in 10mL water, and by 750mg
Aziridine is dissolved in 15mL water, and two kinds of solution are mixed to get solution A；
S2: it under the conditions of argon gas, with the pH value of the hydrochloric acid conditioning solution A of 6mol/L to 4.0, is combined using microwave ultrasonic wave
Synthesis/abstraction instrument auxiliary dissolution, is arranged 65 DEG C of temperature, microwave power 600W, ultrasonic power 1000W, dissolution time 20min,
Obtain solution B；
S3: by the Na of 1.5mmol under the conditions of argon gas2S·9H2O is dissolved in 5mL water, is added in solution B and is reacted, and is reacted
Time 0.6h, obtains solution C；
S4: the dehydrated alcohol of 10mL is added in solution C, centrifugation, centrifugation rate 6000r/min, centrifugation time
10min obtains supernatant liquor and lower sediment, and taking lower sediment is the amine-modified Mn-ZnS quantum dot of polyethyleneimine.
(4) preparation of molecularly imprinted polymer:
S1: by what is prepared in the covalent organic frame material prepared in 6mL methanol, 12mg step (2), 15mL step (3)
The amine-modified Mn-ZnS quantum dot mixing of polyethyleneimine is reacted, reaction time 0.8h；
S2: the anthracene amide and 1mL ethylene glycol dimethacrylate prepared in 300g step (1) is added, is eventually adding
52mg azodiisobutyronitrile, heating continue to react, and 70 DEG C of heating temperature, the reaction time for 24 hours, obtains solid F；
S3: soxhlet type, extraction times 16h, molecularly imprinted polymer obtained by drying are carried out after solid F is ground up, sieved.
(5) molecularly imprinted polymer fills out the preparation of column: the molecularly imprinted polymer being prepared in step (4) is added to
In Solid Phase Extraction void column, do not release the polymer in extraction column to get molecularly imprinted solid phase extraction column.
Embodiment 6
It is identical with the molecularly imprinted solid phase extraction column extraction process of oat extract anthracene amide in embodiment 3, molecule used
The preparation process of trace solid-phase extraction column is as follows:
(1) preparation of anthracene amide:
S1: the p-Coumaric Acid of 5mmol is dissolved in the pyridine of 2mL, is added the acetic anhydride of 10mL, reaction time 5h,
Then the water of 50mL, 3 DEG C of the temperature of water is added, refrigeration, collects precipitate A by 4 DEG C of refrigerated storage temperature, cold preservation time 1h；
S2: precipitate A is cleaned 3 times with 4 DEG C of water, dry, place 12h, be then added in precipitate A 10mL DMF and
The triethylamine of 1.25mL obtains mixed solution B；
S3: the BOP of 5mmol is dissolved in the methylene chloride of 5mL, under conditions of ice-water bath dropwise with 2 drops/s rate
It is added drop-wise in the mixed solution B, obtains solution C；
S4: 5mmol 2- amino -5- hydroxybenzoic acid is dissolved in the DMF of 20mL, with 2 drops/s under conditions of ice-water bath
Rate be added drop-wise in the solution C dropwise, stir 30min, be stored at room temperature 2h, then terminated again with the 0.5M hydrochloric acid of 80mL anti-
It answers, obtains solution D, stand, 4 DEG C of dwell temperature, time of repose 14h；
S5: extracting solution D with the organic solvent of 200mL, be concentrated to give substance E, is added in substance E and contains volume
The 10mL dichloromethane solution of 5% pyrrolidines of score reacts, reaction time 20min, and the 1M hydrochloric acid that 20mL is then added terminates instead
It answers, is finally extracted with the organic solvent of 100mL, be concentrated and dried, obtain anthracene amide solid powder.
(2) preparation of covalent organic frame material:
S1: under a nitrogen atmosphere, dry 6g phloroglucin and 15g methenamine are added to the trifluoroacetic acid of 90mL
In, heating is reacted, and 100 DEG C of heating temperature, reaction time 2.5h；
S2: the 3M hydrochloric acid of 150mL is added in the solution after above-mentioned reaction, and heating continues to react, heating temperature
100 DEG C, reaction time 1h is cooled to room temperature after complete reaction；
S3: being extracted with the organic solvent of 350mL, is concentrated, is dried, and a kind of substance TP is obtained；
S4: the p-phenylenediamine of the TP of 63mg and 48mg being added in the 3M acetic acid of 75mL, with the 1,3,5-trihydroxybenzene of 40mL and
The dioxane of 40mL makees mixed solvent and is also added in acetic acid, heating reaction, and 120 DEG C of heating temperature, heating time 72h, to
It is dried in vacuo after fully reacting, obtains covalent organic frame material.
(3) preparation of the amine-modified Mn-ZnS quantum dot of polyethyleneimine:
S1: by 288mg ZnSO4·7H2O and 10mg Mn (CH3COO)2Mixed dissolution gathers in 10mL water, and by 750mg
Aziridine is dissolved in 10mL water, and two kinds of solution are mixed to get solution A；
S2: it under the conditions of argon gas, with the pH value of the hydrochloric acid conditioning solution A of 6mol/L to 4.0, is combined using microwave ultrasonic wave
Synthesis/abstraction instrument auxiliary dissolution, setting temperature 60 C, microwave power 500W, ultrasonic power 1000W, dissolution time 15min,
Obtain solution B；
S3: by the Na of 1mmol under the conditions of argon gas2S·9H2O is dissolved in 2.5mL water, is added in solution B and is reacted, and is reacted
Time 0.5h, obtains solution C；
S4: the dehydrated alcohol of 10mL is added in solution C, centrifugation, centrifugation rate 6000r/min, centrifugation time
10min obtains supernatant liquor and lower sediment, and taking lower sediment is the amine-modified Mn-ZnS quantum dot of polyethyleneimine.
(4) preparation of molecularly imprinted polymer:
S1: poly- by what is prepared in the covalent organic frame material prepared in 2mL methanol, 10mg step (2), 5mL step (3)
The Mn-ZnS quantum dot mixing of aziridine modification is reacted, reaction time 0.5h；
S2: the anthracene amide and 1mL ethylene glycol dimethacrylate prepared in 300g step (1) is added, is eventually adding
45mg azodiisobutyronitrile, heating continue to react, and 60 DEG C of heating temperature, the reaction time for 24 hours, obtains solid F；
S3: soxhlet type, extraction times 15h, molecularly imprinted polymer obtained by drying are carried out after solid F is ground up, sieved.
(5) molecularly imprinted polymer fills out the preparation of column: the molecularly imprinted polymer being prepared in step (4) is added to
In Solid Phase Extraction void column, do not release the polymer in extraction column to get molecularly imprinted solid phase extraction column.
Comparative example 1
During difference with embodiment 6 is that molecularly imprinted solid phase extraction column extracts oat extract anthracene amide, use
Phosphoric acid adjusts the pH value of solution A to 5.0, remaining condition is all the same.
Comparative example 2
During difference with embodiment 6 is that molecularly imprinted solid phase extraction column extracts oat extract anthracene amide, swallow
The mass volume ratio of wheat and extract liquor is 1:45g/mL, remaining condition is all the same.
Comparative example 3
During difference with embodiment 6 is that molecularly imprinted solid phase extraction column extracts oat extract anthracene amide, institute
It is 40% with ethanol content, remaining condition is all the same.
Comparative example 4
Difference with embodiment 6 is in the preparation process of molecularly imprinted solid phase extraction column used, step (4) molecular engram
The anthracene amide prepared in step (1) is not added in the preparation of polymer, and non-imprinted polymer and non-trace Solid Phase Extraction is prepared
Column, remaining condition is all the same, specific as follows:
It is identical with the molecularly imprinted solid phase extraction column extraction process of oat extract anthracene amide in embodiment 3, using non-print
Mark solid-phase extraction column replaces molecularly imprinted solid phase extraction column to extract oat extract anthracene amide, non-trace solid-phase extraction column used
Preparation process is as follows:
(1) preparation of covalent organic frame material:
S1: under a nitrogen atmosphere, dry 6g phloroglucin and 15g methenamine are added to the trifluoroacetic acid of 90mL
In, heating is reacted, and 100 DEG C of heating temperature, reaction time 2.5h；
S2: the 3M hydrochloric acid of 150mL is added in the solution after above-mentioned reaction, and heating continues to react, heating temperature
100 DEG C, reaction time 1h is cooled to room temperature after complete reaction；
S3: being extracted with the organic solvent of 350mL, is concentrated, is dried, and a kind of substance TP is obtained；
S4: the p-phenylenediamine of the TP of 63mg and 48mg being added in the 3M acetic acid of 75mL, with the 1,3,5-trihydroxybenzene of 40mL and
The dioxane of 40mL makees mixed solvent and is also added in acetic acid, heating reaction, and 120 DEG C of heating temperature, heating time 72h, to
It is dried in vacuo after fully reacting, obtains covalent organic frame material.
(2) preparation of the amine-modified Mn-ZnS quantum dot of polyethyleneimine:
S1: by 288mg ZnSO4·7H2O and 10mg Mn (CH3COO)2Mixed dissolution gathers in 10mL water, and by 750mg
Aziridine is dissolved in 10mL water, and two kinds of solution are mixed to get solution A；
S2: it under the conditions of argon gas, with the pH value of the hydrochloric acid conditioning solution A of 6mol/L to 4.0, is combined using microwave ultrasonic wave
Synthesis/abstraction instrument auxiliary dissolution, setting temperature 60 C, microwave power 500W, ultrasonic power 1000W, dissolution time 15min,
Obtain solution B；
S3: by the Na of 1mmol under the conditions of argon gas2S·9H2O is dissolved in 2.5mL water, is added in solution B and is reacted, and is reacted
Time 0.5h, obtains solution C；
S4: the dehydrated alcohol of 10mL is added in solution C, centrifugation, centrifugation rate 6000r/min, centrifugation time
10min obtains supernatant liquor and lower sediment, and taking lower sediment is the amine-modified Mn-ZnS quantum dot of polyethyleneimine.
(3) preparation of non-imprinted polymer:
S1: poly- by what is prepared in the covalent organic frame material prepared in 2mL methanol, 10mg step (2), 5mL step (3)
The Mn-ZnS quantum dot mixing of aziridine modification is reacted, reaction time 0.5h；
S2: adding 1mL ethylene glycol dimethacrylate, is eventually adding 45mg azodiisobutyronitrile, heating continue into
Row reaction, 60 DEG C of heating temperature, the reaction time for 24 hours, obtains solid F；
S3: soxhlet type, extraction times 15h, non-imprinted polymer obtained by drying are carried out after solid F is ground up, sieved.
(4) non-imprinted polymer fills out the preparation of column: the non-imprinted polymer being prepared in step (4) is added to solid phase
It extracts in void column, does not release the polymer in extraction column to get non-trace solid-phase extraction column.
Test case 1
Using methanol as solvent, the anthracene amide extracted in embodiment 1-6 and comparative example 1-4 is configured to 35 μ g/mL's
Anthracene amide standard solution, using 10mL methanol as eluent, 100mL n-hexane is leacheate, and nitrogen is blown under the conditions of 50 DEG C, is then adopted
Redissolve with methanol and anthracene amide content in 10 kinds of samples is detected using high performance liquid chromatograph.Liquid phase chromatogram condition is
Xtiamate C18 (4.6X 250mm, 5 μm), column temperature are 30 DEG C, flow velocity 0.6mL/min, B phase in mobile phase (methanol+
0.1% formic acid): A phase (+0.1% formic acid of water)=8:2,5 μ L of sampling volume).Gradient elution program is 0-5min B phase 80%, A
Phase 20%；6-10min B phase 70%, A phase 30%；11-15min B phase 60%, A phase 40%；16-20min B phase 50%, A phase
50%；21-25min B phase 40%, A phase 60%；26-30min B phase 30%, A phase 70%, test result is as shown in table 1.
The test of 1 anthracene amide content of table
| Sample | Anthracene amide content μ g/g |
| Embodiment 1 | 219 |
| Embodiment 2 | 224 |
| Embodiment 3 | 227 |
| Embodiment 4 | 238 |
| Embodiment 5 | 232 |
| Embodiment 6 | 247 |
| Comparative example 1 | 201 |
| Comparative example 2 | 193 |
| Comparative example 3 | 210 |
| Comparative example 4 | 195 |
Test case 2
60 male C57BL/6 mouse (3 week old) are selected, alternately 12h light/12h is dark at 23 DEG C ± 3 DEG C by mouse follows
Ring, and identical diet is provided.After 1 week laundering period, mouse is based on RANDOMIZED BLOCK DESIGN and is divided into two groups, wherein 10 small
Mouse is organized as low fat diet (LFD) provides energy (10% fat, 20% protein and the 70% carbon aquation of 3.85 kilocalories/gram
Close object), and remaining 50 mouse be then used as high fat diet (HFD) group provide 4.73 kilocalories/gram energy (35% fat,
24% protein and 41% carbohydrate), it persistently provides 8 weeks, to establish obese model.According to formula bluntness (%)=
The average weight of [average weight of the actual weight-LFD group mouse of HFD group mouse]/LFD group mouse, when HFD group mouse obesity
Degree value is greater than 20% and is considered as modeling success.
The successful 22 HFD group mouse of modeling is randomly divided into 11 groups, every group 2, wherein 10 groups of mouse fill respectively daily
The anthracene amide extracted in stomach 100mg/kg embodiment 1-6 and comparative example 1-4, remaining 1 group of mouse as a control group, fill daily
The water of stomach 100mg/kg.After experiment periods 8 weeks, all mouse fasting 12 hours weigh Mice Body then by sacrificed by decapitation
Weight, every group is averaged, and the results are shown in Table 2.Then the blood sample for collecting mouse, is stored in spare at -80 DEG C.
Influence of the 2 anthracene amide of table to HFD group mouse weight
| Sample | Mouse weight reduces percentage % |
| Embodiment 1 | 12.6 |
| Embodiment 2 | 13.7 |
| Embodiment 3 | 13.9 |
| Embodiment 4 | 14.3 |
| Embodiment 5 | 14.2 |
| Embodiment 6 | 14.8 |
| Comparative example 1 | 11.1 |
| Comparative example 2 | 10.3 |
| Comparative example 3 | 11.5 |
| Comparative example 4 | 10.5 |
| Control group | 0.2 |
As can be seen from Table 2, the weight of mouse high in fat can be effectively reduced in stomach-filling anthracene amide, improves fat situation, and
Anthracene amide content is higher, and the effect of losing weight is more obvious.
Test case 3
The mouse blood sample of anthracene amide and control group that the stomach-filling embodiment 6 collected in test case 2 is extracted with
The rate of 3000r/min is centrifuged 15 minutes, and the glucose and insulin then collected in supernatant liquor use business enzyme reagent kit
It is detected.Based on formula HOMA-IR=[serum glucose (mmol/L) × serum insulin (mU/L)]/22.4, HOMA-IR
Index represents the ability of insulin resistance, and the glucose and insulin concentration in supernatant liquor are serum glucose and serum pancreas
Island element concentration, every group of test result are averaged, and the HOMA-IR index results being calculated according to formula are as shown in table 3.
Influence of the 3 anthracene amide of table to glucose and insulin in HFD group mice serum
| Sample | Glucose | Insulin | HOMA-IR index |
| Embodiment 6 | 4.11±0.49a | 38.72±2.57a | 7.08±0.55a |
| Control group | 5.10±0.22b | 43.93±2.07b | 9.83±0.67b |
Note: there is no the subscript of common letter different in table.
As can be seen from Table 3, the concentration of glucose in blood and insulin, HOMA- can be effectively reduced in stomach-filling anthracene amide
IR index also has apparent reduction, this shows that avenanthramide has the energy for mitigating the insulin resistance as caused by high fat diet
Power.
The technical means disclosed in the embodiments of the present invention is not limited to the technical means disclosed in the above technical means, and further includes
Technical solution consisting of any combination of the above technical features.The foregoing is a specific embodiment of the present invention, should refer to
Out, for those skilled in the art, without departing from the principle of the present invention, can also make several
Improvements and modifications, these modifications and embellishments are also considered to be within the scope of the present invention.
Claims (7)
1. a kind of extracting method of avenanthramide, which is characterized in that the avenanthramide uses molecular engram solid phase extraction
Column extracts；
Wherein, the extraction process of the avenanthramide is as follows:
S1: ammonium phosphate is dissolved in the in the mixed solvent of water and ethyl alcohol, obtains mixed solution A；
S2: it stands after being adjusted with acid the pH value to acidity of solution A as extract liquor B；
S3: it is mixed after oat is ground with extract liquor B, is centrifuged after ultrasound, takes supernatant liquor；
S4: the remainder particulate after taking supernatant liquor repeats step S3, is concentrated to give substance C after merging supernatant liquor；
S5: by substance C by molecularly imprinted solid phase extraction column, up to avenanthramide after elution, elution；
The molecularly imprinted solid phase extraction column preparation process is as follows:
(1) preparation of anthracene amide:
S1: p-Coumaric Acid is dissolved in pyridine, and acetic anhydride is added, and then plus water cooling is hidden, and collects precipitate A；
S2: precipitate A is washed with water, then DMF and triethylamine are added in precipitate A, obtains mixed solution B for dry, placement；
S3: being dissolved in methylene chloride for BOP, is added in the mixed solution B under conditions of ice-water bath, obtains solution C；
S4: being dissolved in DMF for 2- amino -5- hydroxybenzoic acid, be added in the solution C under conditions of ice-water bath, and stirring is simultaneously
It is stored at room temperature, then reacts to obtain solution D with hydrochloric acid termination again, stand；
S5: extracting solution D with organic solvent, be concentrated to give substance E, and the dichloromethane containing pyrrolidines is added in substance E
Then alkane solution reaction is added hydrochloric acid and terminates reaction, finally extracted, be concentrated and dried with organic solvent, it is solid to obtain anthracene amide
Body powder；
(2) preparation of covalent organic frame material:
S1: under a nitrogen atmosphere, dry phloroglucin and methenamine being added in trifluoroacetic acid, and heating is reacted；
S2: hydrochloric acid is added in the solution after above-mentioned reaction, and heating continues to react, and is cooled to room after complete reaction
Temperature；
S3: being extracted with organic solvent, is concentrated, is dried, and a kind of substance TP is obtained；
S4: TP and p-phenylenediamine being added in acetic acid, mixed solvent is made with 1,3,5-trihydroxybenzene and dioxane and is also added in acetic acid,
Heating reaction, is dried in vacuo after complete reaction, obtains covalent organic frame material；
(3) preparation of the amine-modified Mn-ZnS quantum dot of polyethyleneimine:
S1: by ZnSO4·7H2O and Mn (CH3COO)2Mixed dissolution Yu Shuizhong, and polyethyleneimine is dissolved in the water, two kinds are molten
Liquid is mixed to get solution A；
S2: under inert gas conditions, it is adjusted with acid the pH value of solution A, obtains solution B；
S3: under inert gas conditions by Na2S·9H2O is dissolved in the water, and is added in solution B and reacts, obtains solution C；
S4: dehydrated alcohol is added in solution C, and centrifugation obtains supernatant liquor and lower sediment, and taking lower sediment is poly- second
The Mn-ZnS quantum dot of alkene imines modification；
(4) preparation of molecularly imprinted polymer:
S1: the polyethyleneimine prepared in the covalent organic frame material prepared in methanol, step (2), step (3) is amine-modified
The mixing of Mn-ZnS quantum dot is reacted；
S2: the anthracene amide and ethylene glycol dimethacrylate prepared in step (1) is added, two isobutyl of azo is eventually adding
Nitrile, heating continue to react, and obtain solid F；
S3: carrying out soxhlet type after solid F is ground up, sieved, up to molecularly imprinted polymer after drying；
(5) molecularly imprinted polymer fills out column: the molecularly imprinted polymer being prepared in step (4) is added to Solid Phase Extraction sky
In column, do not release the polymer in extraction column to get molecularly imprinted solid phase extraction column.
2. extracting method according to claim 1, which is characterized in that phosphorus in step S1 in the extraction process of the avenanthramide
The molal volume of sour ammonium and mixed solvent ratio is 1:300-600mol/L, and the volume ratio of water and ethyl alcohol is 1:3-5, ethyl alcohol quality point
Number is 60-90%；Acid in step S2 is phosphoric acid, and the pH value range of solution A is 2.6-3.2.
3. extracting method according to claim 1, which is characterized in that swallow in step S3 in the extraction process of the avenanthramide
The mass volume ratio of wheat and extract liquor be 1:5-20g/mL, ultrasonic time 10-40min, centrifugal rotational speed 2000-2400r/min, from
Heart time 10-15min；Step S3, coextraction 2-4 times are repeated in step S4.
4. extracting method according to claim 1, which is characterized in that the molecularly imprinted solid phase extraction column preparation step
(1) in the preparation process of anthracene amide:
S1: the p-Coumaric Acid of 1-10mmol is dissolved in the pyridine of 1-20mL, is added the acetic anhydride of 10-30mL, when reaction
Between 3-8h, then be added 40-100mL water, 0-4 DEG C of the temperature of water, refrigeration, 2-6 DEG C of refrigerated storage temperature, cold preservation time 1-2h, receive
Collect precipitate A；
S2: precipitate A is cleaned 2-4 times with 2-6 DEG C of water, then dry, placement 12-24h is added 5-40mL's in precipitate A
The triethylamine of DMF and 1-10mL obtains mixed solution B；
S3: the BOP of 1-10mmol is dissolved in the methylene chloride of 1-20mL, with 1-2 drop/s rate under conditions of ice-water bath
It is added drop-wise in the mixed solution B dropwise, obtains solution C；
S4: the 2- amino -5- hydroxybenzoic acid of 1-10mmol is dissolved in the DMF of 10-80mL, with 1- under conditions of ice-water bath
2 drops/s rate is added drop-wise to dropwise in the solution C, is stirred 20-40min, is stored at room temperature 1-3h, then again with 50-150mL's
0.5M hydrochloric acid terminates reaction, obtains solution D, stands, 3-6 DEG C of dwell temperature, time of repose 12-24h；
S5: extracting solution D with the organic solvent of 50-300mL, be concentrated to give substance E, is added in substance E and contains volume
The 5-30mL dichloromethane solution of score 3-6% pyrrolidines reacts, reaction time 20-40min, and the 1M salt of 5-40mL is then added
Acid terminates reaction, is finally extracted with the organic solvent of 50-200mL, is concentrated and dried, obtains anthracene amide solid powder；
In the step (2) in the preparation process of covalent organic frame material:
S1: under a nitrogen atmosphere, dry 4-10g phloroglucin and 10-20g methenamine are added to the trifluoro of 60-150mL
In acetic acid, heating is reacted, and 90-120 DEG C of heating temperature, reaction time 2-4h；
S2: the 3M hydrochloric acid of 100-300mL is added in the solution after above-mentioned reaction, and heating continues to react, heating temperature
90-120 DEG C, reaction time 1-2h is cooled to room temperature after complete reaction；
S3: being extracted with the organic solvent of 200-500mL, is concentrated, is dried, and a kind of substance TP is obtained；
S4: the p-phenylenediamine of the TP of 50-100mg and 30-60mg being added in the 3M acetic acid of 60-100mL, equal with 30-50mL
The dioxane of benzenetriol and 40-80mL make mixed solvent and are also added in acetic acid, heating reaction, and 110-130 DEG C of heating temperature,
Heating time 70-80h, is dried in vacuo after complete reaction, obtains covalent organic frame material.
5. extracting method according to claim 1, which is characterized in that the molecularly imprinted solid phase extraction column preparation process step
Suddenly in the preparation process of the amine-modified Mn-ZnS quantum dot of (3) polyethyleneimine, acid used in step S2 is hydrochloric acid, using microwave and is surpassed
Sound wave combined auxiliary dissolution.
6. extracting method according to claim 5, which is characterized in that the molecularly imprinted solid phase extraction column preparation process step
Suddenly in (3) in the preparation process of the amine-modified Mn-ZnS quantum dot of polyethyleneimine:
S1: by 250-350mg ZnSO4·7H2O and 5-25mg Mn (CH3COO)2Mixed dissolution is in 5-15mL water, and by 700-
800mg polyethyleneimine is dissolved in 10-20mL water, and two kinds of solution are mixed to get solution A；
S2: under inert gas conditions, with the pH value of the hydrochloric acid conditioning solution A of 6mol/L to 2.5-5.5, microwave and ultrasound connection
It closes auxiliary to dissolve, 50-80 DEG C of solution temperature, microwave power 400-700W, ultrasonic power 800-1200W, dissolution time 10-
30min obtains solution B；
S3: under inert gas conditions by the Na of 0.2-1mmol2S·9H2O is dissolved in 1-10mL water, is added anti-in solution B
It answers, reaction time 0.3-1h obtains solution C；
S4: the dehydrated alcohol of 5-15mL is added in solution C, centrifugation, centrifugation rate 5000-7000r/min, centrifugation time 5-
20min obtains supernatant liquor and lower sediment, and taking lower sediment is the amine-modified Mn-ZnS quantum dot of polyethyleneimine；
In the step (4) in the preparation process of molecularly imprinted polymer:
S1: by preparation in the covalent organic frame material prepared in 2-10mL methanol, 6-20mg step (2), 5-30mL step (3)
Polyethyleneimine it is amine-modified Mn-ZnS quantum dot mixing reacted, reaction time 0.5-1h；
S2: the anthracene amide and 0.1-3mL ethylene glycol dimethacrylate prepared in 200-400g step (1) is added, finally
30-80mg azodiisobutyronitrile is added, heating continues to react, and 60-80 DEG C of heating temperature, reaction time 20-30h is obtained
Solid F；
S3: soxhlet type, extraction times 10-20h, molecularly imprinted polymer obtained by drying are carried out after solid F is ground up, sieved.
7. a kind of application for the avenanthramide that extracting method as described in any one of claim 1-6 is extracted, which is characterized in that
The avenanthramide is used to prepare in the drug for the treatment of obesity and its associated disease, and associated disease is type II diabetes.
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