CN111481645A - Method for extracting amomum tsao-ko total flavonoids - Google Patents
Method for extracting amomum tsao-ko total flavonoids Download PDFInfo
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- 229930003935 flavonoid Natural products 0.000 title claims abstract description 33
- 235000017173 flavonoids Nutrition 0.000 title claims abstract description 33
- 238000000034 method Methods 0.000 title claims abstract description 31
- 150000002215 flavonoids Chemical class 0.000 title claims abstract description 30
- 241001346334 Amomum tsao-ko Species 0.000 title claims description 6
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims abstract description 76
- 238000001035 drying Methods 0.000 claims abstract description 38
- WEVYAHXRMPXWCK-UHFFFAOYSA-N Acetonitrile Chemical compound CC#N WEVYAHXRMPXWCK-UHFFFAOYSA-N 0.000 claims abstract description 33
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims abstract description 29
- 235000019441 ethanol Nutrition 0.000 claims abstract description 28
- 239000011347 resin Substances 0.000 claims abstract description 27
- 229920005989 resin Polymers 0.000 claims abstract description 27
- 239000003480 eluent Substances 0.000 claims abstract description 26
- GAMYVSCDDLXAQW-AOIWZFSPSA-N Thermopsosid Natural products O(C)c1c(O)ccc(C=2Oc3c(c(O)cc(O[C@H]4[C@H](O)[C@@H](O)[C@H](O)[C@H](CO)O4)c3)C(=O)C=2)c1 GAMYVSCDDLXAQW-AOIWZFSPSA-N 0.000 claims abstract description 24
- 238000000605 extraction Methods 0.000 claims abstract description 24
- 229930003944 flavone Natural products 0.000 claims abstract description 24
- 150000002212 flavone derivatives Chemical class 0.000 claims abstract description 24
- 235000011949 flavones Nutrition 0.000 claims abstract description 24
- 238000001179 sorption measurement Methods 0.000 claims abstract description 24
- VHBFFQKBGNRLFZ-UHFFFAOYSA-N vitamin p Natural products O1C2=CC=CC=C2C(=O)C=C1C1=CC=CC=C1 VHBFFQKBGNRLFZ-UHFFFAOYSA-N 0.000 claims abstract description 24
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 claims abstract description 20
- 239000000843 powder Substances 0.000 claims abstract description 19
- 239000000284 extract Substances 0.000 claims abstract description 18
- 239000000287 crude extract Substances 0.000 claims abstract description 16
- 239000002904 solvent Substances 0.000 claims abstract description 16
- 235000013399 edible fruits Nutrition 0.000 claims abstract description 14
- 239000006228 supernatant Substances 0.000 claims abstract description 13
- 238000010992 reflux Methods 0.000 claims abstract description 11
- NVNLLIYOARQCIX-MSHCCFNRSA-N Nisin Chemical compound N1C(=O)[C@@H](CC(C)C)NC(=O)C(=C)NC(=O)[C@@H]([C@H](C)CC)NC(=O)[C@@H](NC(=O)C(=C/C)/NC(=O)[C@H](N)[C@H](C)CC)CSC[C@@H]1C(=O)N[C@@H]1C(=O)N2CCC[C@@H]2C(=O)NCC(=O)N[C@@H](C(=O)N[C@H](CCCCN)C(=O)N[C@@H]2C(NCC(=O)N[C@H](C)C(=O)N[C@H](CC(C)C)C(=O)N[C@H](CCSC)C(=O)NCC(=O)N[C@H](CS[C@@H]2C)C(=O)N[C@H](CC(N)=O)C(=O)N[C@H](CCSC)C(=O)N[C@H](CCCCN)C(=O)N[C@@H]2C(N[C@H](C)C(=O)N[C@@H]3C(=O)N[C@@H](C(N[C@H](CC=4NC=NC=4)C(=O)N[C@H](CS[C@@H]3C)C(=O)N[C@H](CO)C(=O)N[C@H]([C@H](C)CC)C(=O)N[C@H](CC=3NC=NC=3)C(=O)N[C@H](C(C)C)C(=O)NC(=C)C(=O)N[C@H](CCCCN)C(O)=O)=O)CS[C@@H]2C)=O)=O)CS[C@@H]1C NVNLLIYOARQCIX-MSHCCFNRSA-N 0.000 claims abstract description 10
- 108010053775 Nisin Proteins 0.000 claims abstract description 10
- 108010059820 Polygalacturonase Proteins 0.000 claims abstract description 10
- 108010093305 exopolygalacturonase Proteins 0.000 claims abstract description 10
- 239000004309 nisin Substances 0.000 claims abstract description 10
- 235000010297 nisin Nutrition 0.000 claims abstract description 10
- 239000011780 sodium chloride Substances 0.000 claims abstract description 10
- 235000002639 sodium chloride Nutrition 0.000 claims abstract description 10
- 108010059892 Cellulase Proteins 0.000 claims abstract description 9
- 229940106157 cellulase Drugs 0.000 claims abstract description 9
- 239000012153 distilled water Substances 0.000 claims abstract description 8
- 239000000725 suspension Substances 0.000 claims abstract description 8
- 239000000047 product Substances 0.000 claims abstract description 7
- 238000002791 soaking Methods 0.000 claims abstract description 7
- 238000003756 stirring Methods 0.000 claims abstract description 7
- 241001127714 Amomum Species 0.000 claims abstract description 5
- 239000003463 adsorbent Substances 0.000 claims abstract description 5
- 239000003109 Disodium ethylene diamine tetraacetate Substances 0.000 claims abstract description 4
- 235000019301 disodium ethylene diamine tetraacetate Nutrition 0.000 claims abstract description 4
- 238000010438 heat treatment Methods 0.000 claims abstract description 4
- 238000000967 suction filtration Methods 0.000 claims abstract description 3
- 238000011068 loading method Methods 0.000 claims description 19
- 239000007788 liquid Substances 0.000 claims description 10
- KCXVZYZYPLLWCC-UHFFFAOYSA-N EDTA Chemical compound OC(=O)CN(CC(O)=O)CCN(CC(O)=O)CC(O)=O KCXVZYZYPLLWCC-UHFFFAOYSA-N 0.000 claims description 8
- 238000010828 elution Methods 0.000 claims description 7
- XEKOWRVHYACXOJ-UHFFFAOYSA-N Ethyl acetate Chemical compound CCOC(C)=O XEKOWRVHYACXOJ-UHFFFAOYSA-N 0.000 claims description 6
- 238000004108 freeze drying Methods 0.000 claims description 5
- 229960001484 edetic acid Drugs 0.000 claims description 4
- 238000010298 pulverizing process Methods 0.000 claims description 4
- 238000011049 filling Methods 0.000 claims description 2
- 239000005452 food preservative Substances 0.000 abstract description 4
- 235000019249 food preservative Nutrition 0.000 abstract description 4
- 239000003963 antioxidant agent Substances 0.000 abstract description 3
- 230000003078 antioxidant effect Effects 0.000 abstract description 3
- 235000006708 antioxidants Nutrition 0.000 abstract description 3
- 229940079593 drug Drugs 0.000 abstract description 3
- 239000003814 drug Substances 0.000 abstract description 3
- ZGTMUACCHSMWAC-UHFFFAOYSA-L EDTA disodium salt (anhydrous) Chemical compound [Na+].[Na+].OC(=O)CN(CC([O-])=O)CCN(CC(O)=O)CC([O-])=O ZGTMUACCHSMWAC-UHFFFAOYSA-L 0.000 abstract 1
- 238000005070 sampling Methods 0.000 abstract 1
- JMGZEFIQIZZSBH-UHFFFAOYSA-N Bioquercetin Natural products CC1OC(OCC(O)C2OC(OC3=C(Oc4cc(O)cc(O)c4C3=O)c5ccc(O)c(O)c5)C(O)C2O)C(O)C(O)C1O JMGZEFIQIZZSBH-UHFFFAOYSA-N 0.000 description 8
- IVTMALDHFAHOGL-UHFFFAOYSA-N eriodictyol 7-O-rutinoside Natural products OC1C(O)C(O)C(C)OC1OCC1C(O)C(O)C(O)C(OC=2C=C3C(C(C(O)=C(O3)C=3C=C(O)C(O)=CC=3)=O)=C(O)C=2)O1 IVTMALDHFAHOGL-UHFFFAOYSA-N 0.000 description 8
- FDRQPMVGJOQVTL-UHFFFAOYSA-N quercetin rutinoside Natural products OC1C(O)C(O)C(CO)OC1OCC1C(O)C(O)C(O)C(OC=2C(C3=C(O)C=C(O)C=C3OC=2C=2C=C(O)C(O)=CC=2)=O)O1 FDRQPMVGJOQVTL-UHFFFAOYSA-N 0.000 description 8
- IKGXIBQEEMLURG-BKUODXTLSA-N rutin Chemical compound O[C@H]1[C@H](O)[C@@H](O)[C@H](C)O[C@@H]1OC[C@H]1[C@H](O)[C@@H](O)[C@H](O)[C@@H](OC=2C(C3=C(O)C=C(O)C=C3OC=2C=2C=C(O)C(O)=CC=2)=O)O1 IKGXIBQEEMLURG-BKUODXTLSA-N 0.000 description 8
- ALABRVAAKCSLSC-UHFFFAOYSA-N rutin Natural products CC1OC(OCC2OC(O)C(O)C(O)C2O)C(O)C(O)C1OC3=C(Oc4cc(O)cc(O)c4C3=O)c5ccc(O)c(O)c5 ALABRVAAKCSLSC-UHFFFAOYSA-N 0.000 description 8
- 235000005493 rutin Nutrition 0.000 description 8
- 229960004555 rutoside Drugs 0.000 description 8
- 238000011160 research Methods 0.000 description 6
- 241000196324 Embryophyta Species 0.000 description 5
- 238000001914 filtration Methods 0.000 description 5
- 238000002835 absorbance Methods 0.000 description 4
- -1 flavonoid compound Chemical class 0.000 description 4
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 description 3
- 239000000126 substance Substances 0.000 description 3
- 239000001913 cellulose Substances 0.000 description 2
- 229920002678 cellulose Polymers 0.000 description 2
- 238000012986 modification Methods 0.000 description 2
- 230000004048 modification Effects 0.000 description 2
- 238000012545 processing Methods 0.000 description 2
- 239000012488 sample solution Substances 0.000 description 2
- LPXPTNMVRIOKMN-UHFFFAOYSA-M sodium nitrite Chemical compound [Na+].[O-]N=O LPXPTNMVRIOKMN-UHFFFAOYSA-M 0.000 description 2
- 238000006467 substitution reaction Methods 0.000 description 2
- BNGXYYYYKUGPPF-UHFFFAOYSA-M (3-methylphenyl)methyl-triphenylphosphanium;chloride Chemical compound [Cl-].CC1=CC=CC(C[P+](C=2C=CC=CC=2)(C=2C=CC=CC=2)C=2C=CC=CC=2)=C1 BNGXYYYYKUGPPF-UHFFFAOYSA-M 0.000 description 1
- BDOYKFSQFYNPKF-UHFFFAOYSA-N 2-[2-[bis(carboxymethyl)amino]ethyl-(carboxymethyl)amino]acetic acid;sodium Chemical compound [Na].[Na].OC(=O)CN(CC(O)=O)CCN(CC(O)=O)CC(O)=O BDOYKFSQFYNPKF-UHFFFAOYSA-N 0.000 description 1
- 241000234299 Zingiberaceae Species 0.000 description 1
- 239000004480 active ingredient Substances 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 238000004140 cleaning Methods 0.000 description 1
- 230000000052 comparative effect Effects 0.000 description 1
- 238000007865 diluting Methods 0.000 description 1
- 238000004090 dissolution Methods 0.000 description 1
- HHEAADYXPMHMCT-UHFFFAOYSA-N dpph Chemical compound [O-][N+](=O)C1=CC([N+](=O)[O-])=CC([N+]([O-])=O)=C1[N]N(C=1C=CC=CC=1)C1=CC=CC=C1 HHEAADYXPMHMCT-UHFFFAOYSA-N 0.000 description 1
- 238000002481 ethanol extraction Methods 0.000 description 1
- 235000013305 food Nutrition 0.000 description 1
- 239000000463 material Substances 0.000 description 1
- 239000001814 pectin Substances 0.000 description 1
- 229920001277 pectin Polymers 0.000 description 1
- 235000010987 pectin Nutrition 0.000 description 1
- 238000002360 preparation method Methods 0.000 description 1
- 238000004321 preservation Methods 0.000 description 1
- 235000010288 sodium nitrite Nutrition 0.000 description 1
- 235000013599 spices Nutrition 0.000 description 1
- 238000012360 testing method Methods 0.000 description 1
- 229940126680 traditional chinese medicines Drugs 0.000 description 1
- 239000000341 volatile oil Substances 0.000 description 1
- 238000005303 weighing Methods 0.000 description 1
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)
- A61K36/88—Liliopsida (monocotyledons)
- A61K36/906—Zingiberaceae (Ginger family)
- A61K36/9064—Amomum, e.g. round cardamom
-
- 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
- A23L3/00—Preservation of foods or foodstuffs, in general, e.g. pasteurising, sterilising, specially adapted for foods or foodstuffs
- A23L3/34—Preservation of foods or foodstuffs, in general, e.g. pasteurising, sterilising, specially adapted for foods or foodstuffs by treatment with chemicals
- A23L3/3454—Preservation of foods or foodstuffs, in general, e.g. pasteurising, sterilising, specially adapted for foods or foodstuffs by treatment with chemicals in the form of liquids or solids
- A23L3/3463—Organic compounds; Microorganisms; Enzymes
- A23L3/3472—Compounds of undetermined constitution obtained from animals or plants
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23V—INDEXING SCHEME RELATING TO FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES AND LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION
- A23V2002/00—Food compositions, function of food ingredients or processes for food or foodstuffs
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K2236/00—Isolation or extraction methods of medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicine
- A61K2236/10—Preparation or pretreatment of starting material
- A61K2236/17—Preparation or pretreatment of starting material involving drying, e.g. sun-drying or wilting
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K2236/00—Isolation or extraction methods of medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicine
- A61K2236/10—Preparation or pretreatment of starting material
- A61K2236/19—Preparation or pretreatment of starting material involving fermentation using yeast, bacteria or both; enzymatic treatment
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K2236/00—Isolation or extraction methods of medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicine
- A61K2236/30—Extraction of the material
- A61K2236/33—Extraction of the material involving extraction with hydrophilic solvents, e.g. lower alcohols, esters or ketones
- A61K2236/333—Extraction of the material involving extraction with hydrophilic solvents, e.g. lower alcohols, esters or ketones using mixed solvents, e.g. 70% EtOH
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K2236/00—Isolation or extraction methods of medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicine
- A61K2236/30—Extraction of the material
- A61K2236/39—Complex extraction schemes, e.g. fractionation or repeated extraction steps
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K2236/00—Isolation or extraction methods of medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicine
- A61K2236/50—Methods involving additional extraction steps
- A61K2236/51—Concentration or drying of the extract, e.g. Lyophilisation, freeze-drying or spray-drying
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K2236/00—Isolation or extraction methods of medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicine
- A61K2236/50—Methods involving additional extraction steps
- A61K2236/55—Liquid-liquid separation; Phase separation
Abstract
The invention discloses a method for extracting tsaoko total flavonoids, which comprises the steps of drying and crushing fresh tsaoko fruits to obtain tsaoko powder, adding ethanol, adding cellulase, pectinase, sodium chloride, disodium ethylenediamine tetraacetate and nisin, uniformly stirring, and soaking for 0.5-2 hours to obtain an extracting solution; heating the extracting solution for reflux extraction, performing suction filtration, and concentrating by a rotary evaporator to remove the solvent to obtain a crude extract product; dissolving the crude extract in distilled water to form suspension, centrifuging, and sampling the supernatant on a macroporous adsorbent resin column at a flow rate; after adsorption, standing, eluting with water until no turbidity exists, eluting with acetonitrile and ethyl alcohol eluent, collecting eluent, and concentrating under reduced pressure to recover the solvent; obtaining extract, and drying to obtain the tsaoko amomum fruit total flavone. The method has the advantages that the extraction amount of the total flavonoids reaches more than 28mg/g, the purity of the total flavonoids reaches more than 85%, and the extracted fructus tsaoko total flavonoids have wide application in food preservatives, antioxidant drugs and other aspects.
Description
Technical Field
The invention relates to the technical field of extraction of active ingredients of natural plants, in particular to a method for extracting amomum tsao-ko total flavonoids.
Background
The tsaoko (scientific name: Amomum tsaoko Crevost et L emarie) is a perennial herbaceous plant of Amomum in Zingiberaceae, mainly distributed in provinces such as Yunnan, Guangxi and Guizhou, etc., the tsaoko is one of bulk varieties of medicinal and edible Chinese medicinal materials, is mainly applied to the field of food processing as a spice, and is also applied to the fields of traditional Chinese medicines and other fields.
In recent years, researches on tsaoko mainly focus on extraction and identification of tsaoko essential oil, researches on other components of tsaoko are few, with the deep research on tsaoko, the medicinal value of tsaoko is increasingly emphasized by scholars at home and abroad, the extraction of other chemical components of tsaoko has become a research hotspot, a plant flavonoid compound is a natural antioxidant, as the plant flavonoid compound can clear free radicals in human bodies and inhibit the generation of free radicals, the plant flavonoid compound can be used as a natural food preservative and is widely applied to the preservation and processing of products, scholars have a certain research on the extraction of the tsaoko total flavonoids, such as Yuanyuan, Zhangian and the like, the research on the removal of the DPPH free radicals is disclosed, the extraction process is that the tsaoko total flavonoids are extracted by cleaning, drying, crushing, passing through a 40-mesh sieve, adding an ethanol extracting solution, adding an ultrasonic wave assisted extraction, centrifuging, an extracting solution, 201710656974.9 discloses the tsaoko total flavonoid extract, the preparation method comprises the steps of (1) heating, extracting, ①, drying, extracting, concentrating, extracting, a column, a BV-57, concentrating, extracting, a macroporous column, a macroporous adsorption column is used for extracting, a macroporous column for extracting, the ethanol extraction column is used for extracting, the ethanol concentration of the ethanol is used for extracting, the ethanol is used for extracting, the ethanol is used for 2-90 column, the concentration is used for extracting, the ethanol for extracting, the concentration is used for extracting, the ethanol for extracting is used for extracting, the ethanol for extracting, the purity of the concentration is used for extracting is used for 2 concentration is used for extracting, the purity of the concentration is used.
Disclosure of Invention
The invention aims at the problems and provides a method for extracting the amomum tsao-ko total flavonoids. The method has the advantages that the extraction amount of the total flavonoids reaches more than 28mg/g, the purity of the total flavonoids reaches more than 95%, and the extracted fructus tsaoko total flavonoids have wide application in food preservatives, antioxidant drugs and other aspects.
In order to achieve the above purpose, the technical scheme adopted by the invention is as follows:
a method for extracting fructus Tsaoko total flavone comprises the following steps:
(1) drying fresh fructus Tsaoko, pulverizing to obtain fructus Tsaoko powder, adding ethanol according to the dosage of 1:15-25 g/L of the material-liquid ratio, adding cellulase, pectinase, sodium chloride, disodium ethylenediamine tetraacetate and nisin, stirring uniformly, and soaking for 0.5-2h to obtain an extracting solution;
(2) heating the extracting solution for reflux extraction, performing suction filtration, and concentrating by a rotary evaporator to remove the solvent to obtain a crude extract product;
(3) dissolving the crude extract in distilled water to form suspension, centrifuging, and loading the supernatant onto macroporous adsorbent resin column at a loading flow rate of 1.0-3.0 BV/h; standing after adsorption, eluting at an elution speed of 1.0-3.0BV/h, eluting with water until no turbidity exists, eluting with an acetonitrile and ethyl alcohol eluent, collecting eluent, and concentrating under reduced pressure to recover a solvent; obtaining extract, and drying to obtain the tsaoko amomum fruit total flavone.
Preferably, the drying in step (1) and step (3) is performed by freeze drying and then microwave drying.
Preferably, the freeze drying is carried out for 2-4h at the temperature of-25 to-45 ℃, and then the product is dried under the power of 150 minus one hundred thousand hours until the product is qualified; drying the tsaoko fruit in the step (1) until the water content of the tsaoko fruit is 12-15%, and drying the extract in the step (3) until the water content of the extract is 4-6%.
Preferably, the weight ratio of the tsaoko powder to the cellulase, the pectinase, the sodium chloride, the ethylene diamine tetraacetic acid and the nisin is 100:0.5-1.0:0.3-0.5:1-3:0.8-1.5: 0.2-0.5.
Preferably, the reflux extraction in step (2) is reflux extraction at 60-70 deg.C for 3 times, each time for 1.5-2.5 h.
Preferably, the macroporous adsorption resin column is HPD-100 type, L D605 type or HP-20 type macroporous adsorption resin column.
Preferably, the mass ratio of the resin filling amount to the tsaoko powder is 1: 2-5.
Preferably, the volume ratio of the acetonitrile to the ethyl acetate in the eluent is 1: 3-5.
Preferably, the amount of the eluent is 8 to 12 times of the volume of the filled macroporous adsorption resin.
Compared with the prior art, the invention has the advantages and beneficial effects that:
1. according to the invention, the cellulose and the pectin in the tsaoko powder are dissolved by using the cellulose and the pectinase, so that the dissolution of the total flavone is facilitated, and the tsaoko powder is soaked by using the sodium chloride, the disodium ethylene diamine tetraacetate and the nisin, so that not only is the extraction rate of the total flavone effectively improved, but also the stability of the total flavone can be effectively improved.
2. The method can improve the purity of the total flavone by passing through a macroporous adsorption resin column and eluting with acetonitrile and ethyl alcohol ethyl ester eluent, and is simple and low in cost.
3. The method firstly performs freeze drying and then performs microwave drying in the whole drying process of the tsaoko amomum fruits, can prevent effective components such as total flavonoids and the like from being damaged, and has the advantages of thorough drying, simple operation and low cost.
4. The method has the advantages that the extraction amount of the total flavonoids reaches more than 28mg/g, the purity of the total flavonoids reaches more than 85%, and the extracted fructus tsaoko total flavonoids have wide application in the fields of food preservatives, medicines and the like.
Detailed Description
The present invention will be described in further detail with reference to specific embodiments. It should be emphasized that the following description is merely exemplary in nature and is not intended to limit the scope of the invention or its application.
Example 1
A method for extracting fructus Tsaoko total flavone comprises the following steps:
(1) drying fresh fructus Tsaoko (purchased from Guangxi Yulin) at-30 deg.C for 3.5h, drying at power of 200W until water content is 13.5%, pulverizing to 80 mesh, collecting fructus Tsaoko powder 100g, adding 95% ethanol according to dosage of material-liquid ratio of 1:20 g/L, adding cellulase 0.8g, pectinase 0.5g, sodium chloride 2.5g, disodium edetate 1.2g, and nisin 0.3g, stirring well, soaking for 1h to obtain extract;
(2) extracting the extractive solution at 65 deg.C under reflux for 3 times (2 hr each time), vacuum filtering, and concentrating with rotary evaporator to remove solvent to obtain crude extract;
(3) dissolving the crude extract in distilled water to form a suspension, centrifuging, taking supernatant, loading the supernatant on an HPD-100 type macroporous adsorption resin column at a loading flow rate of 2.0BV/h, wherein the mass ratio of the resin loading to the tsaoko powder is 1:3, standing after adsorption, then eluting at an elution speed of 2.0BV/h, firstly eluting with water until no turbidity exists, then eluting with acetonitrile and ethyl alcohol eluent at a volume ratio of 1:4, wherein the using amount of the eluent is 10 times of the volume of the filled macroporous adsorption resin, collecting the eluent, and concentrating under reduced pressure to recover the solvent; obtaining extract, drying at-30 deg.C for 4h, and drying at power of 200W until water content is 4.6%, to obtain 3.283mg fructus Tsaoko total flavone with total flavone content of 87.34%.
The content of the total flavone is calculated according to the following method, the specific method comprises the following steps of accurately weighing 10mg of rutin standard substance, dissolving the rutin standard substance by absolute ethyl alcohol, and then fixing the volume to 100 m L to obtain 0.10 mg/m L rutin contrast liquid, respectively and accurately sucking 0, l, 2, 3, 4 and 5m L rutin contrast liquid, placing the rutin contrast liquid in a 10m L measuring flask for later use, measuring the absorbance of the solution according to the following method operations, namely adding 5% sodium nitrite solution 0.5 m L, shaking up, standing for 6 min, adding 10% aluminum nitrate solution 0.5 m L, shaking up, standing for 6Illin, adding 4% sodium hydroxide inner line solution 4 m L, diluting the solution to scale by 60% ethanol, shaking up, standing for 15 min, measuring the absorbance value at 510nm, drawing a standard curve, namely, the solubility of the rutin solution is respectively 0, 0.01, 0.02, 0.03, 0.04, 0.05mg/m L, the linear relation of the rutin content in the range of 0-0.05 mg/m L, regression, and the equation of R + 3629 is good, and the linear relation of the rutin content in the2=0.9993。
Measuring 10mg of tsaoko total flavonoids, metering to 100 m L with absolute ethyl alcohol, obtaining a sample solution of 0.1mg/m L, measuring 1 ml of the sample solution in a 10 ml test tube, measuring absorbance according to the method, substituting the measured absorbance value into a human standard curve equation, and finding out the content of the total flavonoids.
Example 2
A method for extracting fructus Tsaoko total flavone comprises the following steps:
(1) drying fresh tsaoko fruits at the temperature of minus 35 ℃ for 3 hours, then drying at the power of 250W until the water content is 12.5%, crushing to 80 meshes, taking 100g of tsaoko fruit powder, adding 90% ethanol according to the dosage of 1:25 g/L of the feed-liquid ratio, then adding 0.6g of cellulase, 0.4g of pectinase, 2g of sodium chloride, 1.0g of ethylene diamine tetraacetic acid and 0.3g of nisin, stirring uniformly, and soaking for 1.5 hours to obtain an extract;
(2) extracting the extractive solution at 70 deg.C under reflux for 3 times (each for 1.5 hr), vacuum filtering, and concentrating with rotary evaporator to remove solvent to obtain crude extract;
(3) dissolving the crude extract in distilled water to form a suspension, centrifuging, taking supernatant, loading the supernatant on an HPD-100 type macroporous adsorption resin column at a loading flow rate of 3.0BV/h, wherein the mass ratio of the resin loading to the tsaoko powder is 1:4, standing after adsorption, then eluting at an elution speed of 3.0BV/h, firstly eluting with water until no turbidity exists, then eluting with acetonitrile and ethyl alcohol eluent at a volume ratio of 1:2, wherein the using amount of the eluent is 12 times of the volume of the filled macroporous adsorption resin, collecting the eluent, and concentrating under reduced pressure to recover the solvent; obtaining extract, drying at-35 deg.C for 3.5h, and drying at power of 250W until water content is 5.2%, to obtain 3.114g fructus Tsaoko total flavone with total flavone content of 86.07%.
Example 3
A method for extracting fructus Tsaoko total flavone comprises the following steps:
(1) drying fresh tsaoko fruits at-40 ℃ for 2.5h, then drying under the power of 200W until the water content is 15%, crushing to 80 meshes, taking 100g of tsaoko powder, adding 80% ethanol according to the dosage of 1:22 g/L of the feed-liquid ratio, then adding 1.0g of cellulase, 0.3g of pectinase, 1.5g of sodium chloride, 1.2g of ethylene diamine tetraacetic acid and 0.5g of nisin, stirring uniformly, and soaking for 0.5h to obtain an extract;
(2) extracting the extractive solution at 65 deg.C under reflux for 3 times (2 hr each time), vacuum filtering, and concentrating with rotary evaporator to remove solvent to obtain crude extract;
(3) dissolving the crude extract in distilled water to form a suspension, centrifuging, taking supernatant, loading the supernatant on an HP-20 type macroporous adsorption resin column at a loading flow rate of 1.0BV/h, wherein the mass ratio of the resin loading to the tsaoko powder is 1:3, standing after adsorption, then eluting at an elution speed of 3.0BV/h, firstly eluting with water until no turbidity exists, then eluting with an acetonitrile and ethyl alcohol eluent with a volume ratio of 1:1, wherein the dosage of the eluent is 10 times of the volume of the macroporous adsorption resin, collecting the eluent, and concentrating under reduced pressure to recover the solvent; obtaining extract, drying at-40 deg.C for 3h, and drying at power of 250W until water content is 4.8%, to obtain 2.814g fructus Tsaoko total flavone with total flavone content of 85.14%.
Example 4
A method for extracting fructus Tsaoko total flavone comprises the following steps:
(1) drying fresh tsaoko fruits at-30 ℃ for 4h, then drying under the power of 150W until the water content is 14.5%, crushing to 80 meshes, taking 100g of tsaoko fruit powder, adding 95% ethanol according to the dosage of 1:25 g/L of the feed-liquid ratio, then adding 0.6g of cellulase, 0.5g of pectinase, 2.5g of sodium chloride, 1.0g of ethylene diamine tetraacetic acid disodium and 0.4g of nisin, stirring uniformly, and soaking for 2h to obtain an extract;
(2) extracting the extractive solution at 60 deg.C under reflux for 3 times (2.5 hr each time), vacuum filtering, and concentrating with rotary evaporator to remove solvent to obtain crude extract;
(3) dissolving the crude extract in distilled water to form a suspension, centrifuging, taking supernatant, loading the supernatant on an HP-20 type macroporous adsorption resin column at a loading flow rate of 2.0BV/h, wherein the mass ratio of the resin loading to the tsaoko powder is 1:5, standing after adsorption, then eluting at an elution speed of 2.0BV/h, firstly eluting with water until no turbidity exists, then eluting with an acetonitrile and ethyl alcohol eluent with a volume ratio of 1:2, wherein the dosage of the eluent is 10 times of the volume of the macroporous adsorption resin, collecting the eluent, and concentrating under reduced pressure to recover the solvent; obtaining extract, drying at-30 deg.C for 4h, and drying at power of 150W until water content is 5.4%, to obtain 3.069g fructus Tsaoko total flavone with total flavone content of 86.78%.
Comparative example
A method for extracting fructus Tsaoko total flavone comprises the following steps:
(1) drying fresh fructus Tsaoko at-30 deg.C for 4 hr, pulverizing to 80 mesh, collecting fructus Tsaoko powder 100g, and adding 95% ethanol according to the material-liquid ratio of 1:25 g/L to obtain extract;
(2) extracting the extractive solution at 60 deg.C under reflux for 3 times (2.5 hr each time), vacuum filtering, and concentrating with rotary evaporator to remove solvent to obtain crude extract;
(3) dissolving the crude extract in distilled water to form a suspension, centrifuging, taking supernatant, loading the supernatant on an HP-20 type macroporous adsorption resin column at a loading flow rate of 2.0BV/h, wherein the mass ratio of the resin loading to the tsaoko powder is 1:5, standing after adsorption, then eluting at an elution speed of 2.0BV/h, firstly eluting with water until no turbidity exists, then eluting with an acetonitrile and ethyl alcohol eluent with a volume ratio of 1:2, wherein the dosage of the eluent is 10 times of the volume of the macroporous adsorption resin, collecting the eluent, and concentrating under reduced pressure to recover the solvent; obtaining extract, drying at-30 deg.C for 4h, and drying at power of 150W until water content is 5.4%, to obtain 2.358mg fructus Tsaoko total flavone with total flavone content of 82.19%.
The foregoing is a more detailed description of the invention in connection with specific/preferred embodiments and is not intended to limit the practice of the invention to those descriptions. It will be apparent to those skilled in the art that various substitutions and modifications can be made to the described embodiments without departing from the spirit of the invention, and such substitutions and modifications are to be considered as within the scope of the invention.
Claims (10)
1. A method for extracting amomum tsao-ko total flavonoids is characterized by comprising the following steps: the method comprises the following steps:
(1) drying fresh fructus Tsaoko, pulverizing to obtain fructus Tsaoko powder, adding ethanol according to the dosage of 1:15-25 g/L of the material-liquid ratio, adding cellulase, pectinase, sodium chloride, disodium ethylenediamine tetraacetate and nisin, stirring uniformly, and soaking for 0.5-2h to obtain an extracting solution;
(2) heating the extracting solution for reflux extraction, performing suction filtration, and concentrating by a rotary evaporator to remove the solvent to obtain a crude extract product;
(3) dissolving the crude extract in distilled water to form suspension, centrifuging, and loading the supernatant onto macroporous adsorbent resin column at a loading flow rate of 1.0-3.0 BV/h; standing after adsorption, eluting at an elution speed of 1.0-3.0BV/h, eluting with water until no turbidity exists, eluting with an acetonitrile and ethyl alcohol eluent, collecting eluent, and concentrating under reduced pressure to recover a solvent; obtaining extract, and drying to obtain the tsaoko amomum fruit total flavone.
2. The method for extracting the tsaoko total flavonoids according to claim 1, wherein the extraction process comprises the following steps: and (3) the drying in the step (1) and the step (3) is to perform microwave drying after freeze drying.
3. The method for extracting the tsaoko total flavonoids according to claim 2, wherein the extraction process comprises the following steps: the freeze drying is to dry for 2 to 4 hours at the temperature of minus 25 to minus 45 ℃, and then dry under the power of 150 minus one hundred thousand centimeters (W) and 250W until the product is qualified; drying the tsaoko fruit in the step (1) until the water content of the tsaoko fruit is 12-15%, and drying the extract in the step (3) until the water content of the extract is 4-6%.
4. The method for extracting the tsaoko total flavonoids according to claim 1, wherein the extraction process comprises the following steps: the weight ratio of the tsaoko powder to the cellulase, the pectinase, the sodium chloride, the ethylene diamine tetraacetic acid and the nisin is 100:0.5-1.0:0.3-0.5:1-3:0.8-1.5: 0.2-0.5.
5. The method for extracting the tsaoko total flavonoids according to claim 1, wherein the extraction process comprises the following steps: the reflux extraction in the step (2) is carried out for 3 times at the temperature of 60-70 ℃, and each extraction time is 1.5-2.5 h.
6. The method for extracting the tsaoko total flavonoids according to claim 1, wherein the macroporous adsorbent resin column is HPD-100 type, L D605 type or HP-20 macroporous adsorbent resin column.
7. The method for extracting the tsaoko total flavonoids according to claim 1, wherein the extraction process comprises the following steps: the mass ratio of the resin filling amount to the tsaoko powder is 1: 2-5.
8. The method for extracting the tsaoko total flavonoids according to claim 1, wherein the extraction process comprises the following steps: the volume ratio of acetonitrile to ethyl acetate in the eluent is 1: 3-5.
9. The method for extracting tsaoko total flavonoids as claimed in claim 8, wherein: the dosage of the eluent is 8 to 12 times of the volume of the filled macroporous adsorption resin.
10. The method for extracting the tsaoko total flavonoids according to claim 1, wherein the extraction process comprises the following steps: the ethanol is 70-95% ethanol.
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