CN114591925A - Method for extracting fructus Rosae Normalis superoxide dismutase - Google Patents
Method for extracting fructus Rosae Normalis superoxide dismutase Download PDFInfo
- Publication number
- CN114591925A CN114591925A CN202210265569.5A CN202210265569A CN114591925A CN 114591925 A CN114591925 A CN 114591925A CN 202210265569 A CN202210265569 A CN 202210265569A CN 114591925 A CN114591925 A CN 114591925A
- Authority
- CN
- China
- Prior art keywords
- superoxide dismutase
- centrifuging
- rosa roxburghii
- extraction
- crude extract
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
- 102000019197 Superoxide Dismutase Human genes 0.000 title claims abstract description 84
- 108010012715 Superoxide dismutase Proteins 0.000 title claims abstract description 84
- 238000000034 method Methods 0.000 title claims abstract description 25
- 238000000605 extraction Methods 0.000 claims abstract description 37
- 239000007788 liquid Substances 0.000 claims abstract description 34
- 240000002547 Rosa roxburghii Species 0.000 claims abstract description 31
- 235000000640 Rosa roxburghii Nutrition 0.000 claims abstract description 31
- 239000000287 crude extract Substances 0.000 claims abstract description 29
- 238000001914 filtration Methods 0.000 claims abstract description 28
- OKTJSMMVPCPJKN-UHFFFAOYSA-N Carbon Chemical compound [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 claims abstract description 25
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims abstract description 15
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims abstract description 15
- 238000005406 washing Methods 0.000 claims abstract description 14
- 238000001816 cooling Methods 0.000 claims abstract description 13
- 238000002156 mixing Methods 0.000 claims abstract description 10
- 229910052799 carbon Inorganic materials 0.000 claims abstract description 5
- 239000003153 chemical reaction reagent Substances 0.000 claims abstract description 5
- 239000012266 salt solution Substances 0.000 claims abstract description 5
- 150000003839 salts Chemical class 0.000 claims abstract description 5
- 238000001035 drying Methods 0.000 claims abstract description 3
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 claims description 60
- 239000000243 solution Substances 0.000 claims description 33
- 239000011780 sodium chloride Substances 0.000 claims description 31
- 238000003756 stirring Methods 0.000 claims description 25
- 239000007787 solid Substances 0.000 claims description 22
- 239000000047 product Substances 0.000 claims description 13
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 claims description 12
- 239000000706 filtrate Substances 0.000 claims description 11
- 238000004108 freeze drying Methods 0.000 claims description 11
- 239000000203 mixture Substances 0.000 claims description 6
- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 claims description 4
- 239000007864 aqueous solution Substances 0.000 claims description 4
- 238000005119 centrifugation Methods 0.000 claims description 4
- 238000000227 grinding Methods 0.000 claims description 2
- 238000010979 pH adjustment Methods 0.000 claims description 2
- 241000220317 Rosa Species 0.000 abstract description 15
- 230000000694 effects Effects 0.000 abstract description 11
- 235000004789 Rosa xanthina Nutrition 0.000 abstract description 8
- 241000109329 Rosa xanthina Species 0.000 abstract description 8
- 241000196324 Embryophyta Species 0.000 abstract description 6
- 239000003960 organic solvent Substances 0.000 abstract description 5
- 102000004169 proteins and genes Human genes 0.000 abstract description 2
- 108090000623 proteins and genes Proteins 0.000 abstract description 2
- 238000003912 environmental pollution Methods 0.000 abstract 1
- 230000000052 comparative effect Effects 0.000 description 19
- 239000012535 impurity Substances 0.000 description 9
- 102000004190 Enzymes Human genes 0.000 description 6
- 108090000790 Enzymes Proteins 0.000 description 6
- 238000002474 experimental method Methods 0.000 description 5
- 238000012360 testing method Methods 0.000 description 5
- 235000013399 edible fruits Nutrition 0.000 description 4
- WQGWDDDVZFFDIG-UHFFFAOYSA-N pyrogallol Chemical compound OC1=CC=CC(O)=C1O WQGWDDDVZFFDIG-UHFFFAOYSA-N 0.000 description 4
- 241001465754 Metazoa Species 0.000 description 3
- 239000008280 blood Substances 0.000 description 3
- 210000004369 blood Anatomy 0.000 description 3
- 238000004458 analytical method Methods 0.000 description 2
- 238000006701 autoxidation reaction Methods 0.000 description 2
- 235000013339 cereals Nutrition 0.000 description 2
- 230000002708 enhancing effect Effects 0.000 description 2
- 230000002255 enzymatic effect Effects 0.000 description 2
- 230000002265 prevention Effects 0.000 description 2
- 229940079877 pyrogallol Drugs 0.000 description 2
- 240000002234 Allium sativum Species 0.000 description 1
- 241000219357 Cactaceae Species 0.000 description 1
- 208000003643 Callosities Diseases 0.000 description 1
- 208000035473 Communicable disease Diseases 0.000 description 1
- 206010011409 Cross infection Diseases 0.000 description 1
- 244000241257 Cucumis melo Species 0.000 description 1
- 235000015510 Cucumis melo subsp melo Nutrition 0.000 description 1
- 208000007212 Foot-and-Mouth Disease Diseases 0.000 description 1
- 241000710198 Foot-and-mouth disease virus Species 0.000 description 1
- 206010020649 Hyperkeratosis Diseases 0.000 description 1
- 206010028980 Neoplasm Diseases 0.000 description 1
- 206010029803 Nosocomial infection Diseases 0.000 description 1
- 240000007594 Oryza sativa Species 0.000 description 1
- 235000007164 Oryza sativa Nutrition 0.000 description 1
- 240000006079 Schisandra chinensis Species 0.000 description 1
- 235000008422 Schisandra chinensis Nutrition 0.000 description 1
- OUUQCZGPVNCOIJ-UHFFFAOYSA-M Superoxide Chemical compound [O-][O] OUUQCZGPVNCOIJ-UHFFFAOYSA-M 0.000 description 1
- 208000018756 Variant Creutzfeldt-Jakob disease Diseases 0.000 description 1
- 240000004922 Vigna radiata Species 0.000 description 1
- 235000010721 Vigna radiata var radiata Nutrition 0.000 description 1
- 235000011469 Vigna radiata var sublobata Nutrition 0.000 description 1
- 240000008042 Zea mays Species 0.000 description 1
- 235000005824 Zea mays ssp. parviglumis Nutrition 0.000 description 1
- 235000002017 Zea mays subsp mays Nutrition 0.000 description 1
- 230000032683 aging Effects 0.000 description 1
- 230000004075 alteration Effects 0.000 description 1
- QVGXLLKOCUKJST-UHFFFAOYSA-N atomic oxygen Chemical compound [O] QVGXLLKOCUKJST-UHFFFAOYSA-N 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 230000000975 bioactive effect Effects 0.000 description 1
- 230000036772 blood pressure Effects 0.000 description 1
- 208000005881 bovine spongiform encephalopathy Diseases 0.000 description 1
- 201000011510 cancer Diseases 0.000 description 1
- 238000006243 chemical reaction Methods 0.000 description 1
- 235000005822 corn Nutrition 0.000 description 1
- 239000002537 cosmetic Substances 0.000 description 1
- 230000007547 defect Effects 0.000 description 1
- 238000004925 denaturation Methods 0.000 description 1
- 230000036425 denaturation Effects 0.000 description 1
- 239000003814 drug Substances 0.000 description 1
- 230000007159 enucleation Effects 0.000 description 1
- 229930003944 flavone Natural products 0.000 description 1
- 150000002213 flavones Chemical class 0.000 description 1
- 235000011949 flavones Nutrition 0.000 description 1
- 235000012055 fruits and vegetables Nutrition 0.000 description 1
- 235000004611 garlic Nutrition 0.000 description 1
- 238000003306 harvesting Methods 0.000 description 1
- 235000013402 health food Nutrition 0.000 description 1
- 230000036039 immunity Effects 0.000 description 1
- 239000004615 ingredient Substances 0.000 description 1
- 244000005700 microbiome Species 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 230000002107 myocardial effect Effects 0.000 description 1
- 150000007524 organic acids Chemical class 0.000 description 1
- 235000005985 organic acids Nutrition 0.000 description 1
- 229910052760 oxygen Inorganic materials 0.000 description 1
- 239000001301 oxygen Substances 0.000 description 1
- 238000001556 precipitation Methods 0.000 description 1
- 238000012545 processing Methods 0.000 description 1
- 230000002035 prolonged effect Effects 0.000 description 1
- 238000010298 pulverizing process Methods 0.000 description 1
- 238000000746 purification Methods 0.000 description 1
- 239000012629 purifying agent Substances 0.000 description 1
- 230000005855 radiation Effects 0.000 description 1
- 235000009566 rice Nutrition 0.000 description 1
- 230000009758 senescence Effects 0.000 description 1
- 238000000926 separation method Methods 0.000 description 1
- 230000007480 spreading Effects 0.000 description 1
- 239000000126 substance Substances 0.000 description 1
- 238000006467 substitution reaction Methods 0.000 description 1
- 239000000758 substrate Substances 0.000 description 1
- 239000000606 toothpaste Substances 0.000 description 1
- 229940034610 toothpaste Drugs 0.000 description 1
- 230000001988 toxicity Effects 0.000 description 1
- 231100000419 toxicity Toxicity 0.000 description 1
- 238000002137 ultrasound extraction Methods 0.000 description 1
- 235000013311 vegetables Nutrition 0.000 description 1
- 235000019156 vitamin B Nutrition 0.000 description 1
- 239000011720 vitamin B Substances 0.000 description 1
Classifications
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
- C12N9/0004—Oxidoreductases (1.)
- C12N9/0089—Oxidoreductases (1.) acting on superoxide as acceptor (1.15)
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Y—ENZYMES
- C12Y115/00—Oxidoreductases acting on superoxide as acceptor (1.15)
- C12Y115/01—Oxidoreductases acting on superoxide as acceptor (1.15) with NAD or NADP as acceptor (1.15.1)
- C12Y115/01001—Superoxide dismutase (1.15.1.1)
Landscapes
- Chemical & Material Sciences (AREA)
- Organic Chemistry (AREA)
- Life Sciences & Earth Sciences (AREA)
- Health & Medical Sciences (AREA)
- Engineering & Computer Science (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Zoology (AREA)
- Wood Science & Technology (AREA)
- Genetics & Genomics (AREA)
- Biochemistry (AREA)
- General Health & Medical Sciences (AREA)
- General Engineering & Computer Science (AREA)
- Medicinal Chemistry (AREA)
- Molecular Biology (AREA)
- Biomedical Technology (AREA)
- Biotechnology (AREA)
- Microbiology (AREA)
- Medicines Containing Plant Substances (AREA)
- Enzymes And Modification Thereof (AREA)
Abstract
The invention provides a method for extracting superoxide dismutase from rosa roxburghii, and relates to the technical field of plant extraction. A method for extracting fructus Rosae Normalis superoxide dismutase comprises the following steps: removing cores of roxburgh roses, crushing, mixing with an extracted salt solution, extracting, centrifuging, filtering and decoloring with active carbon under the condition of pH 8.5-9.0 to obtain a crude extract, adjusting the pH value of the crude extract to 6.0-7.0, centrifuging, filtering, adding a small amount of absolute ethyl alcohol into the liquid, concentrating under reduced pressure and vacuum at 45-50 ℃ until the concentration of the extracted salt is 15-20%, cooling, standing, centrifuging, washing and drying to obtain the superoxide dismutase. By utilizing the principle that protein has good water solubility under alkaline conditions, the method realizes the high-efficiency extraction of the superoxide dismutase in the roxburgh rose under the alkaline conditions of pH8.5-9.0, the obtained superoxide dismutase has high activity, the environmental pollution caused by organic solvents is avoided, the used reagents are common inorganic reagents, the operation is simple, and the cost advantage is achieved.
Description
Technical Field
The invention relates to the technical field of plant extraction, in particular to a method for extracting Rosa roxburghii superoxide dismutase.
Background
The rosa roxburghii tratt is a fruit of perennial deciduous frutescens flower of rosaceous plants, grows in sunny hills, valleys, roadside and bushes with the altitude of 500-2500 m, is a natural wild fruit in places such as Guizhou, Huxi mountain, Xiangxi, Liangshan, and Ming mountain, is artificially planted in large areas in Kaifeng cities of Guizhou province and Henan province, has good edible value and medicinal value, contains rich B vitamins, VC, flavones, organic acids and other ingredients, can protect the heart, and has the effects of relieving fatigue, enhancing myocardial vitality, reducing blood pressure, enhancing immunity, delaying senescence, resisting cancer and the like. The roxburgh rose has short harvest period and is the fruit with the highest content of superoxide dismutase discovered at present, so that the method for extracting the superoxide dismutase from the roxburgh rose has wide prospect in industry.
Superoxide dismutase is widely present in animals, plants and microorganisms, is metalloenzyme taking superoxide anion free radicals as a substrate, and can remove free radicals, so the superoxide dismutase plays an important role in the aspects of oxygen toxicity prevention, radiation damage resistance, aging prevention and the like, is widely used clinically as a new biochemical medicament, is widely used as a bioactive component in daily chemical products such as health food, cosmetics, toothpaste and the like, and is extracted from human blood or animal blood, but the international spreading of infectious diseases such as bovine spongiform encephalopathy, foot-and-mouth disease and the like and the use of SOD extracted from animal blood brings about a plurality of risk factors for people. SOD is extracted from plants, especially pollution-free vegetables, melons and fruits, wild plants and grains which are eaten by people in daily life, such as cactus, garlic, schisandra chinensis, pollen, Xiaobailai rice and other fruits and vegetables, and the SOD also contains abundant SOD in grains such as mung beans, corns and the like, has rich resources and very high use safety, avoids possible cross infection and has low cost.
The extraction and separation method of the rosa roxburghii superoxide dismutase is developed, the additional value of the rosa roxburghii is improved, the deep processing of the rosa roxburghii is facilitated, the industrial chain of the rosa roxburghii is prolonged, the prior extraction method of the rosa roxburghii superoxide dismutase mainly adopts an ultrasonic-assisted extraction method, a thermal denaturation method, an isoelectric point method, an organic solvent precipitation method and the like, and the common extraction method has the problems of low extraction rate or limitation of organic solvent pollution, high cost and the like.
Disclosure of Invention
Technical problem to be solved
Aiming at the defects of the prior art, the invention provides a method for extracting Rosa roxburghii superoxide dismutase, which solves the problems of low extraction rate or limitation of organic solvent pollution, high cost and the like of the common extraction method.
(II) technical scheme
In order to realize the purpose, the invention is realized by the following technical scheme: a method for extracting fructus Rosae Normalis superoxide dismutase comprises the following steps: removing cores of roxburgh roses, crushing, mixing with an extracted salt solution, extracting, centrifuging, filtering and decoloring with active carbon under the condition of pH 8.5-9.0 to obtain a crude extract, adjusting the pH value of the crude extract to 6.0-7.0, centrifuging, filtering, adding a small amount of absolute ethyl alcohol into the liquid, concentrating under reduced pressure and vacuum at 45-50 ℃ until the concentration of the extracted salt is 15-20%, cooling, standing, centrifuging, washing and drying to obtain the superoxide dismutase.
Preferably, the mixing ratio of the roxburgh rose and the extraction saline water is 1: 3-5 in g: mL.
Preferably, the extraction salt is NaCl, and the extraction salt solution is a 3% NaCl solution.
Preferably, the extraction pH is 8.5-9.0.
Preferably, the volume of the absolute ethyl alcohol added is not more than 10% of the volume of the purification solution.
Preferably, the extraction method of the Rosa roxburghii superoxide dismutase comprises the following steps:
s1, removing cores of roxburgh roses, putting the roxburgh roses into a grinder, grinding the roxburgh roses for 3-5 min at the rotation speed of 3500 r/min-4000 r/min to obtain roxburgh rose pulp, adding the roxburgh roses into a stirring container, adjusting the pH to 8.5-9.0 by using a 3% NaCl solution and a 3.5% NaOH aqueous solution, stirring the roxburgh roses for 5h at the rotation speed of 400-500 r/min at room temperature, wherein the mixing ratio of the roxburgh roses to the 3% NaCl solution is 1: 3-5 in g: mL, filtering the mixture by using a 100-mesh screen after mixing and stirring, adding filtrate into a centrifuge for centrifugation, filtering centrifugating the centrifugate by using filter paper and decolorizing the centrifugate by using activated carbon to prepare a crude extract;
s2, adjusting the pH of the crude extract obtained in the step S1 to 6.0-7.0 by using a 3.5% HCL aqueous solution, standing, adding the crude extract into a centrifuge again for centrifugation to obtain a clear liquid, adding 10% volume of absolute ethyl alcohol into the clear liquid, and stirring the mixture for 10-15 min at a rotating speed of 300-400r/min by using a stirrer to uniformly mix the mixture to obtain a purified liquid.
S3, vacuum concentrating the purified liquid at 45-50 ℃ until the NaCl concentration is 15-20%, cooling to room temperature, standing for 12h to obtain a superoxide dismutase precipitated liquid, centrifuging by using a centrifuge to obtain a solid, washing by using pure water, centrifuging to obtain the solid to obtain a wet superoxide dismutase product, and freeze-drying the wet superoxide dismutase product to obtain the superoxide dismutase.
Preferably, the centrifugal condition is that the rotating speed is 4500 r/min-5000 r/min, and the centrifugal time is 15 min-20 min.
Preferably, the reagents used for pH adjustment are 3.5% hydrochloric acid and 3.5% sodium hydroxide.
(III) advantageous effects
The invention provides a method for extracting superoxide dismutase from rosa roxburghii tratt. The method has the following beneficial effects:
the method avoids the influence of nuclear impurities on the extraction of superoxide dismutase, realizes the high-efficiency extraction of the superoxide dismutase in the roxburgh rose under the alkaline condition of pH8.5-9.0 by utilizing the principle that protein has better water solubility under the alkaline condition, has high activity of the obtained superoxide dismutase, avoids the pollution of organic solvents to the environment, adopts common inorganic reagents, and has the advantages of simple operation and cost.
Detailed Description
The technical solutions in the embodiments of the present invention will be clearly and completely described below with reference to the embodiments of the present invention, and it is obvious that the described embodiments are only a part of the embodiments of the present invention, and not all of the embodiments. All other embodiments, which can be obtained by a person skilled in the art without making any creative effort based on the embodiments in the present invention, belong to the protection scope of the present invention.
Example 1:
this example provides a test of the extraction method of superoxide dismutase from Rosa roxburghii Tratt.
Removing cores of 50g of roxburgh rose, crushing, stirring for 5 hours at room temperature by using 150mL of 3% NaCl solution with the pH value of 8.5-9.0, filtering by using a 100-mesh screen, centrifuging filtrate, filtering centrifugate by using filter paper and decolorizing by using activated carbon to prepare a crude extract; adjusting the pH value of the crude extract to 6.0-7.0 by using 3.5% HCL solution, stirring, standing at room temperature for 15min, pouring into a centrifuge tube, centrifuging, and removing precipitated impurities; adding 10% volume of absolute ethyl alcohol into the clear liquid to obtain purified liquid; and (3) concentrating the purified liquid at 45-50 ℃ under reduced pressure and in vacuum until the NaCl concentration is 15%, cooling to room temperature, standing for 12h to obtain a superoxide dismutase eluate, centrifuging to obtain a solid, washing with pure water, centrifuging to obtain the solid to obtain a wet superoxide dismutase product, and freeze-drying to obtain the superoxide dismutase.
Example 2:
this example provides a test of the extraction method of superoxide dismutase from Rosa roxburghii Tratt.
Removing cores of 50g of roxburgh rose, crushing, stirring for 5 hours at room temperature by using 150mL of 3% NaCl solution with the pH value of 8.5-9.0, filtering by using a 100-mesh screen, centrifuging filtrate, filtering centrifugate by using filter paper and decolorizing by using activated carbon to prepare a crude extract; adjusting the pH value of the crude extract to 6.0-7.0 by using 3.5% HCL solution, stirring, standing at room temperature for 15min, pouring into a centrifuge tube, centrifuging, and removing precipitated impurities; adding 10% volume of absolute ethyl alcohol into the clear liquid to obtain purified liquid; and (3) concentrating the purified liquid at 45-50 ℃ under reduced pressure and in vacuum until the NaCl concentration is 20%, cooling to room temperature, standing for 12h to obtain a superoxide dismutase eluate, centrifuging to obtain a solid, washing with pure water, centrifuging to obtain a solid to obtain a wet superoxide dismutase product, and freeze-drying to obtain the superoxide dismutase.
Example 3:
this example provides a test of the extraction method of superoxide dismutase from Rosa roxburghii Tratt.
Removing cores of 50g of roxburgh rose, crushing, stirring for 5 hours at room temperature by using 200mL of 3% NaCl solution with the pH value of 8.5-9.0, filtering by using a 100-mesh screen, centrifuging the filtrate, filtering the centrifugate by using filter paper and decoloring by using activated carbon to prepare a crude extract; adjusting the pH value of the crude extract to 6.0-7.0 by using 3.5% HCL solution, stirring, standing at room temperature for 15min, pouring into a centrifuge tube, centrifuging, and removing precipitated impurities; adding 10% volume of absolute ethyl alcohol into the clear liquid to obtain purified liquid; and (3) concentrating the purified solution at 45-50 ℃ under reduced pressure and in vacuum until the NaCl concentration is 15-20%, cooling to room temperature, standing for 12h to obtain a superoxide dismutase eluate, centrifuging to obtain a solid, washing with pure water, centrifuging to obtain a solid to obtain a wet superoxide dismutase product, and freeze-drying to obtain the superoxide dismutase.
Example 4:
this example provides a test of the extraction method of superoxide dismutase from Rosa roxburghii Tratt.
Removing cores of 50g of roxburgh rose, crushing, stirring for 5 hours at room temperature by using 250mL of 3% NaCl solution with the pH value of 8.5-9.0, filtering by using a 100-mesh screen, centrifuging the filtrate, filtering the centrifugate by using filter paper and decoloring by using activated carbon to prepare a crude extract; adjusting the pH value of the crude extract to 6.0-7.0 by using 3.5% HCL solution, stirring, standing at room temperature for 15min, pouring into a centrifuge tube, centrifuging, and removing precipitated impurities; adding 10% volume of absolute ethyl alcohol into the clear liquid to obtain purified liquid; and (3) concentrating the purified liquid at 45-50 ℃ under reduced pressure and in vacuum until the NaCl concentration is 15% -20%, cooling to room temperature, standing for 12 hours to obtain a superoxide dismutase eluate, centrifuging to obtain a solid, washing with pure water, centrifuging to obtain a solid to obtain a wet superoxide dismutase product, and freeze-drying to obtain the superoxide dismutase.
Comparative example 1:
this example provides a comparative experiment of a method for extracting superoxide dismutase from Rosa roxburghii Tratt without adjusting pH 8.5-9.0.
Removing core of 50g fructus Rosae Normalis, pulverizing, stirring with 150mL 3% NaCl solution at room temperature for 5 hr, filtering with 100 mesh screen, centrifuging the filtrate, filtering the centrifugate with filter paper and decolorizing with active carbon to obtain crude extractive solution; adjusting the pH value of the crude extract to 6.0-7.0 by using 3.5% HCL solution, stirring, standing at room temperature for 15min, pouring into a centrifuge tube, centrifuging, and removing precipitated impurities; adding 10% volume of absolute ethyl alcohol into the clear liquid to obtain purified liquid; and (3) concentrating the purified solution at 45-50 ℃ under reduced pressure and in vacuum until the NaCl concentration is 15-20%, cooling to room temperature, standing for 12h to obtain a superoxide dismutase eluate, centrifuging to obtain a solid, washing with pure water, centrifuging to obtain a solid to obtain a wet superoxide dismutase product, and freeze-drying to obtain the superoxide dismutase.
Comparative example 2:
this example provides a comparative experiment of the extraction process of superoxide dismutase from Rosa roxburghii Tratt without activated carbon treatment.
Removing cores of 50g of roxburgh rose, crushing, stirring for 5 hours at room temperature by using 150mL of 3% NaCl solution with the pH value of 8.5-9.0, filtering by using a 100-mesh screen, centrifuging filtrate, and filtering centrifugate by using filter paper to prepare a crude extract; adjusting the pH value of the crude extract to 6.0-7.0 by using a 3.5% HCL solution, stirring, standing at room temperature for 15min, pouring into a centrifuge tube, centrifuging, and removing precipitated impurities; adding 10% volume of absolute ethyl alcohol into the clear liquid to obtain purified liquid; and (3) concentrating the purified solution at 45-50 ℃ under reduced pressure and in vacuum until the NaCl concentration is 15-20%, cooling to room temperature, standing for 12h to obtain a superoxide dismutase eluate, centrifuging to obtain a solid, washing with pure water, centrifuging to obtain a solid to obtain a wet superoxide dismutase product, and freeze-drying to obtain the superoxide dismutase.
Comparative example 3:
this example provides a comparative experiment of a method for extracting superoxide dismutase from Rosa roxburghii Tratt, in which the purified solution is vacuum concentrated at 45-50 deg.C under reduced pressure until the NaCl concentration is 30%, and an analysis of the extraction results of superoxide dismutase from examples 1-4 and comparative examples 1-3.
This example provides a test of the extraction method of superoxide dismutase from Rosa roxburghii Tratt.
Removing cores of 50g of roxburgh rose, crushing, stirring for 5 hours at room temperature by using 150mL of 3% NaCl solution with the pH value of 8.5-9.0, filtering by using a 100-mesh screen, centrifuging filtrate, filtering centrifugate by using filter paper and decolorizing by using activated carbon to prepare a crude extract; adjusting the pH value of the crude extract to 6.0-7.0 by using 3.5% HCL solution, stirring, standing at room temperature for 15min, pouring into a centrifuge tube, centrifuging, and removing precipitated impurities; adding 10% volume of absolute ethyl alcohol into the clear liquid to obtain purified liquid; and (3) concentrating the purified liquid at 45-50 ℃ under reduced pressure and in vacuum until the NaCl concentration is 30%, cooling to room temperature, standing for 12h to obtain a superoxide dismutase eluate, centrifuging to obtain a solid, washing with pure water, centrifuging to obtain the solid to obtain a wet superoxide dismutase product, and freeze-drying to obtain the superoxide dismutase.
Comparative example 4:
this example provides a comparative experiment of a method for extracting superoxide dismutase from Rosa roxburghii without enucleation treatment.
Crushing 50g of roxburgh rose, stirring for 5 hours at room temperature by using 150mL of 3% NaCl solution with pH of 8.5-9.0, filtering by using a 100-mesh screen, centrifuging the filtrate, filtering the centrifugate by using filter paper and decolorizing the centrifugate by using activated carbon to prepare a crude extract; adjusting the pH value of the crude extract to 6.0-7.0 by using a 3.5% HCL solution, stirring, standing at room temperature for 15min, pouring into a centrifuge tube, centrifuging, and removing precipitated impurities; adding 10% volume of absolute ethyl alcohol into the clear liquid to obtain purified liquid; and (3) concentrating the purified solution at 45-50 ℃ under reduced pressure and in vacuum until the NaCl concentration is 15-20%, cooling to room temperature, standing for 12h to obtain a superoxide dismutase eluate, centrifuging to obtain a solid, washing with pure water, centrifuging to obtain a solid to obtain a wet superoxide dismutase product, and freeze-drying to obtain the superoxide dismutase.
Comparative example 5:
this example provides a comparative experiment of a method for extracting superoxide dismutase from Rosa roxburghii Tratt and an analysis of the extraction results of superoxide dismutase from examples 1-4 and comparative examples 1-5.
Removing cores of 50g of roxburgh rose, crushing, stirring for 5 hours at room temperature by using 150mL of 3% NaCl solution with the pH value of 8.5-9.0, filtering by using a 100-mesh screen, centrifuging filtrate, filtering centrifugate by using filter paper and decolorizing by using activated carbon to prepare a crude extract; adding NaCl into the crude extract until the overall salinity of the solution reaches 20% to obtain superoxide dismutase eluate, centrifuging to obtain solid, washing with pure water, centrifuging to obtain solid to obtain wet superoxide dismutase, and freeze drying to obtain the superoxide dismutase.
The enzyme specific activities of the superoxide dismutase obtained in examples 1-3 and comparative examples 1-5 were determined by using pyrogallol autoxidation method (enzyme activity unit is defined as 1 enzyme activity unit, which is the amount of enzyme that inhibits pyrogallol autoxidation rate by 50% in 1mL of reaction solution at 25 ℃ for 1 min), as shown in Table 1.
TABLE 1 enzymatic specific Activity of Rosa roxburghii superoxide dismutase
| Specific activity of enzyme (U/g) | |
| Example 1 | 18997 |
| Example 2 | 18585 |
| Example 3 | 16780 |
| Example 4 | 17050 |
| Comparative example 1 | 9161 |
| Comparative example 2 | 8020 |
| Comparative example 3 | 2243 |
| Comparative example 4 | 10400 |
| Comparative example 5 | 2174 |
As shown in Table 1, the enzymatic specific activity of the superoxide dismutase obtained by the extraction method of the roxburgh rose superoxide dismutase provided by the invention can reach 18997U/g at most. Compared with the embodiments 1-4 and the comparative examples 1-5, the method for extracting the rosa roxburghii superoxide dismutase, which is provided by the invention, can effectively improve the enzyme activity of the rosa roxburghii superoxide dismutase extraction by the methods of active carbon treatment, purifying agent selection, vacuum concentration, denucleation treatment and the like.
Although embodiments of the present invention have been shown and described, it will be appreciated by those skilled in the art that changes, modifications, substitutions and alterations can be made in these embodiments without departing from the principles and spirit of the invention, the scope of which is defined in the appended claims and their equivalents.
Claims (8)
1. A method for extracting Rosa roxburghii superoxide dismutase is characterized by comprising the following steps: removing cores of rosa roxburghii tratt, crushing, mixing with an extracted salt solution, extracting, centrifuging, filtering and decoloring by active carbon under the condition of pH8.5-9.0 to obtain a crude extract, adjusting the pH value of the crude extract to 6.0-7.0, centrifuging, filtering, adding a small amount of absolute ethyl alcohol into liquid, concentrating under reduced pressure and vacuum at 45-50 ℃ until the concentration of the extracted salt is 15-20%, cooling, standing, centrifuging, washing and drying to obtain the superoxide dismutase.
2. The extraction method of fructus Rosae Normalis superoxide dismutase as claimed in claim 1, wherein: the mixing ratio of the roxburgh rose to the extraction saline water is 1: 3-5 in g: mL.
3. The extraction method of fructus Rosae Normalis superoxide dismutase as claimed in claim 1, wherein: the extraction salt is NaCl, and the extraction salt solution is 3% NaCl solution.
4. The extraction method of fructus Rosae Normalis superoxide dismutase according to claim 1, which is characterized in that: the extraction pH is 8.5-9.0.
5. The extraction method of fructus Rosae Normalis superoxide dismutase as claimed in claim 1, wherein: the addition volume of the absolute ethyl alcohol is not more than 10% of the volume of the purified liquid.
6. The extraction method of fructus Rosae Normalis superoxide dismutase as claimed in claims 1-5, wherein: the method comprises the following steps:
s1, removing kernels of rosa roxburghii tratt, putting the rosa roxburghii tratt into a grinder, grinding at a rotation speed of 3500 r/min-4000 r/min for 3 min-5 min to obtain rosa roxburghii tratt pulp, adding the rosa roxburghii tratt pulp into a stirring container, adjusting the pH to 8.5-9.0 by using a 3% NaCl solution and a 3.5% NaOH aqueous solution, stirring at a rotation speed of 400-500 r/min for 5h at room temperature, wherein the mixing ratio of the rosa roxburghii tratt to the 3% NaCl solution is 1: 3-5 in g/mL, filtering through a 100-mesh screen after mixing and stirring, adding filtrate into a centrifuge for centrifugation, filtering centrifugate through filter paper and decolorizing through activated carbon to prepare a crude extract;
s2, adjusting the pH of the crude extract obtained in the step S1 to 6.0-7.0 by using a 3.5% HCL aqueous solution, standing, adding the crude extract into a centrifuge again for centrifugation to obtain a clear liquid, adding 10% by volume of absolute ethyl alcohol into the clear liquid, stirring the mixture at a rotating speed of 300-400r/min for 10-15 min by using a stirrer, and uniformly mixing the mixture to obtain a purified liquid.
S3, vacuum concentrating the purified liquid at 45-50 ℃ until the NaCl concentration is 15-20%, cooling to room temperature, standing for 12h to obtain a superoxide dismutase precipitated liquid, centrifuging by using a centrifuge to obtain a solid, washing by using pure water, centrifuging to obtain the solid to obtain a wet superoxide dismutase product, and freeze-drying the wet superoxide dismutase product to obtain the superoxide dismutase.
7. The extraction method of fructus Rosae Normalis superoxide dismutase as claimed in claim 6, wherein: the centrifugal condition is 4500 r/min-5000 r/min, and the centrifugal time is 15 min-20 min.
8. The extraction method of fructus Rosae Normalis superoxide dismutase as claimed in claim 1, wherein: the reagents used for the pH adjustment were 3.5% hydrochloric acid and 3.5% sodium hydroxide.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202210265569.5A CN114591925A (en) | 2022-03-17 | 2022-03-17 | Method for extracting fructus Rosae Normalis superoxide dismutase |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202210265569.5A CN114591925A (en) | 2022-03-17 | 2022-03-17 | Method for extracting fructus Rosae Normalis superoxide dismutase |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN114591925A true CN114591925A (en) | 2022-06-07 |
Family
ID=81810731
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202210265569.5A Pending CN114591925A (en) | 2022-03-17 | 2022-03-17 | Method for extracting fructus Rosae Normalis superoxide dismutase |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN114591925A (en) |
Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101812432A (en) * | 2010-04-21 | 2010-08-25 | 张卫国 | Process for extracting superoxide dismutase (SOD) from houttuynia, hovenia dulcis thumb, rosa roxburghii, myrobalan, bottle gourd and chayote |
| CN102747054A (en) * | 2012-07-27 | 2012-10-24 | 田瑛 | Process for extracting superoxide dismutase (SOD) |
| CN110540973A (en) * | 2018-05-29 | 2019-12-06 | 江苏芝能生物科技有限公司 | method for extracting SOD (superoxide dismutase) in roxburgh rose |
| CN114149982A (en) * | 2021-12-03 | 2022-03-08 | 西安全奥生物科技有限公司 | Method for extracting fructus Rosae Normalis superoxide dismutase |
-
2022
- 2022-03-17 CN CN202210265569.5A patent/CN114591925A/en active Pending
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101812432A (en) * | 2010-04-21 | 2010-08-25 | 张卫国 | Process for extracting superoxide dismutase (SOD) from houttuynia, hovenia dulcis thumb, rosa roxburghii, myrobalan, bottle gourd and chayote |
| CN102747054A (en) * | 2012-07-27 | 2012-10-24 | 田瑛 | Process for extracting superoxide dismutase (SOD) |
| CN110540973A (en) * | 2018-05-29 | 2019-12-06 | 江苏芝能生物科技有限公司 | method for extracting SOD (superoxide dismutase) in roxburgh rose |
| CN114149982A (en) * | 2021-12-03 | 2022-03-08 | 西安全奥生物科技有限公司 | Method for extracting fructus Rosae Normalis superoxide dismutase |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN107488515B (en) | Extracting solution and method for extracting tea tree flower essential oil by using same | |
| AU2020101467A4 (en) | Preparation method of lycium ruthenicum murr spray powder capsule | |
| CN111334540B (en) | Method for extracting dendrobium officinale polysaccharide by utilizing biological fermentation | |
| CN106749735A (en) | A kind of process of extracting pueraria polysaccharide | |
| CN113249421A (en) | Golden-silk-royal chrysanthemum protein polypeptide and preparation and application thereof | |
| CN107573438A (en) | A kind of method that polysaccharide is extracted in the pericarp from passion fruit | |
| CN114149982B (en) | Extraction method of rosa roxburghii tratt superoxide dismutase | |
| CN113797261A (en) | Preparation method of camellia flower/leaf extract | |
| CN108300561A (en) | Method for extracting peony seed oil by aqueous enzymatic method | |
| WO2017215313A1 (en) | Method for the preparation of antioxidant peptide using gingko nut shells | |
| CN109043117B (en) | Acidic macadamia nut glycoprotein and production method thereof | |
| CN114591925A (en) | Method for extracting fructus Rosae Normalis superoxide dismutase | |
| CN112442136A (en) | Method for extracting functional components from tremella | |
| CN114249666B (en) | Extraction method of betaine in phoenix tree leaves | |
| CN111808905B (en) | Method for extracting walnut oligopeptide from low-temperature squeezed hickory cake and application thereof | |
| CN106923350B (en) | Method for preparing water-soluble dietary fiber from corn stigma | |
| CN114904294A (en) | Preparation method of high-yield tea flavone | |
| CN110484577B (en) | Method for extracting and preparing mannose from dragon fruit stems | |
| CN114939084A (en) | Russule extract and preparation method and application thereof | |
| CN108084290A (en) | A kind of method of polysaccharide in microwave radiation exaraction common vetch dish | |
| CN108125209B (en) | Black rice anthocyanin salt with antioxidant effect and preparation method thereof | |
| CN113559236A (en) | Selenium-rich walnut and dendrobium composition, tablet and application thereof | |
| CN111171112A (en) | Enzymatic extraction method of bitter melon seed protein | |
| CN112843103A (en) | Method for culturing Polyporus leucoderma, extracting mycelium flavone and flavone compound product thereof | |
| CN107183704B (en) | Preparation and application of edible water lily ethanol extract with antioxidant effect |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination |