CN111840156A - Extraction method of active substance of cinnamomum camphora flower and determination method of oxidation resistance - Google Patents
Extraction method of active substance of cinnamomum camphora flower and determination method of oxidation resistance Download PDFInfo
- Publication number
- CN111840156A CN111840156A CN202010679067.8A CN202010679067A CN111840156A CN 111840156 A CN111840156 A CN 111840156A CN 202010679067 A CN202010679067 A CN 202010679067A CN 111840156 A CN111840156 A CN 111840156A
- Authority
- CN
- China
- Prior art keywords
- solution
- flower
- camphor
- free radical
- measuring
- 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
- 241000723346 Cinnamomum camphora Species 0.000 title claims abstract description 67
- DSSYKIVIOFKYAU-XCBNKYQSSA-N (R)-camphor Chemical compound C1C[C@@]2(C)C(=O)C[C@@H]1C2(C)C DSSYKIVIOFKYAU-XCBNKYQSSA-N 0.000 title claims abstract description 65
- 238000000034 method Methods 0.000 title claims abstract description 26
- 239000013543 active substance Substances 0.000 title claims abstract description 20
- 238000000605 extraction Methods 0.000 title claims abstract description 18
- 230000003647 oxidation Effects 0.000 title claims abstract description 10
- 238000007254 oxidation reaction Methods 0.000 title claims abstract description 10
- CIWBSHSKHKDKBQ-JLAZNSOCSA-N Ascorbic acid Chemical compound OC[C@H](O)[C@H]1OC(=O)C(O)=C1O CIWBSHSKHKDKBQ-JLAZNSOCSA-N 0.000 claims abstract description 44
- 229960000846 camphor Drugs 0.000 claims abstract description 41
- 229930008380 camphor Natural products 0.000 claims abstract description 41
- 239000000243 solution Substances 0.000 claims abstract description 38
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims abstract description 26
- OHDRQQURAXLVGJ-HLVWOLMTSA-N azane;(2e)-3-ethyl-2-[(e)-(3-ethyl-6-sulfo-1,3-benzothiazol-2-ylidene)hydrazinylidene]-1,3-benzothiazole-6-sulfonic acid Chemical compound [NH4+].[NH4+].S/1C2=CC(S([O-])(=O)=O)=CC=C2N(CC)C\1=N/N=C1/SC2=CC(S([O-])(=O)=O)=CC=C2N1CC OHDRQQURAXLVGJ-HLVWOLMTSA-N 0.000 claims abstract description 26
- 239000011550 stock solution Substances 0.000 claims abstract description 24
- ZZZCUOFIHGPKAK-UHFFFAOYSA-N D-erythro-ascorbic acid Natural products OCC1OC(=O)C(O)=C1O ZZZCUOFIHGPKAK-UHFFFAOYSA-N 0.000 claims abstract description 22
- 229930003268 Vitamin C Natural products 0.000 claims abstract description 22
- 239000000284 extract Substances 0.000 claims abstract description 22
- 239000011718 vitamin C Substances 0.000 claims abstract description 22
- 235000019154 vitamin C Nutrition 0.000 claims abstract description 22
- 230000007760 free radical scavenging Effects 0.000 claims abstract description 17
- 238000007865 diluting Methods 0.000 claims abstract description 11
- VTLYFUHAOXGGBS-UHFFFAOYSA-N Fe3+ Chemical compound [Fe+3] VTLYFUHAOXGGBS-UHFFFAOYSA-N 0.000 claims abstract description 10
- 229910001447 ferric ion Inorganic materials 0.000 claims abstract description 10
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims abstract description 9
- 239000013641 positive control Substances 0.000 claims abstract description 5
- 238000001914 filtration Methods 0.000 claims abstract description 4
- 238000004140 cleaning Methods 0.000 claims abstract description 3
- 238000004108 freeze drying Methods 0.000 claims abstract description 3
- 238000010025 steaming Methods 0.000 claims abstract description 3
- 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 claims abstract 7
- 238000002835 absorbance Methods 0.000 claims description 25
- 239000012224 working solution Substances 0.000 claims description 13
- 239000003085 diluting agent Substances 0.000 claims description 11
- -1 ABTS free radical Chemical class 0.000 claims description 10
- 239000004480 active ingredient Substances 0.000 claims description 5
- 230000003078 antioxidant effect Effects 0.000 claims description 5
- 238000002156 mixing Methods 0.000 claims description 5
- 229910021578 Iron(III) chloride Inorganic materials 0.000 claims description 4
- RBTARNINKXHZNM-UHFFFAOYSA-K iron trichloride Chemical compound Cl[Fe](Cl)Cl RBTARNINKXHZNM-UHFFFAOYSA-K 0.000 claims description 4
- 239000002904 solvent Substances 0.000 claims description 4
- YNJBWRMUSHSURL-UHFFFAOYSA-N trichloroacetic acid Chemical compound OC(=O)C(Cl)(Cl)Cl YNJBWRMUSHSURL-UHFFFAOYSA-N 0.000 claims description 4
- 238000001816 cooling Methods 0.000 claims description 3
- MGJZITXUQXWAKY-UHFFFAOYSA-N diphenyl-(2,4,6-trinitrophenyl)iminoazanium Chemical compound [O-][N+](=O)C1=CC([N+](=O)[O-])=CC([N+]([O-])=O)=C1N=[N+](C=1C=CC=CC=1)C1=CC=CC=C1 MGJZITXUQXWAKY-UHFFFAOYSA-N 0.000 claims description 3
- 239000012153 distilled water Substances 0.000 claims description 3
- 239000012064 sodium phosphate buffer Substances 0.000 claims description 3
- 239000006228 supernatant Substances 0.000 claims description 3
- 239000011259 mixed solution Substances 0.000 claims description 2
- 238000012360 testing method Methods 0.000 claims description 2
- 239000011149 active material Substances 0.000 claims 2
- 239000003963 antioxidant agent Substances 0.000 claims 2
- 235000006708 antioxidants Nutrition 0.000 claims 2
- 241000628997 Flos Species 0.000 abstract description 2
- 239000000523 sample Substances 0.000 description 12
- 150000003254 radicals Chemical class 0.000 description 7
- 239000000463 material Substances 0.000 description 4
- 238000010790 dilution Methods 0.000 description 3
- 239000012895 dilution Substances 0.000 description 3
- 238000002474 experimental method Methods 0.000 description 3
- 238000011160 research Methods 0.000 description 3
- 241000196324 Embryophyta Species 0.000 description 2
- QKUSRAKPUWQSJS-UHFFFAOYSA-N diazanium 3-ethyl-2H-1,3-benzothiazole-6-sulfonate Chemical compound [NH4+].[NH4+].[O-]S(=O)(=O)C1=CC=C2N(CC)CSC2=C1.[O-]S(=O)(=O)C1=CC=C2N(CC)CSC2=C1 QKUSRAKPUWQSJS-UHFFFAOYSA-N 0.000 description 2
- 239000003205 fragrance Substances 0.000 description 2
- XEEYBQQBJWHFJM-UHFFFAOYSA-N iron Substances [Fe] XEEYBQQBJWHFJM-UHFFFAOYSA-N 0.000 description 2
- 229910052742 iron Inorganic materials 0.000 description 2
- 238000004519 manufacturing process Methods 0.000 description 2
- 239000000203 mixture Substances 0.000 description 2
- 230000002000 scavenging effect Effects 0.000 description 2
- 239000000126 substance Substances 0.000 description 2
- 238000005303 weighing Methods 0.000 description 2
- 230000002292 Radical scavenging effect Effects 0.000 description 1
- 244000082946 Tarchonanthus camphoratus Species 0.000 description 1
- 235000005701 Tarchonanthus camphoratus Nutrition 0.000 description 1
- 230000000844 anti-bacterial effect Effects 0.000 description 1
- 239000013043 chemical agent Substances 0.000 description 1
- 239000012141 concentrate Substances 0.000 description 1
- 238000010586 diagram Methods 0.000 description 1
- BNIILDVGGAEEIG-UHFFFAOYSA-L disodium hydrogen phosphate Chemical compound [Na+].[Na+].OP([O-])([O-])=O BNIILDVGGAEEIG-UHFFFAOYSA-L 0.000 description 1
- 230000000694 effects Effects 0.000 description 1
- 238000005265 energy consumption Methods 0.000 description 1
- 238000005516 engineering process Methods 0.000 description 1
- 235000019441 ethanol Nutrition 0.000 description 1
- 235000013305 food Nutrition 0.000 description 1
- 238000007710 freezing Methods 0.000 description 1
- 230000008014 freezing Effects 0.000 description 1
- 239000012535 impurity Substances 0.000 description 1
- 238000005259 measurement Methods 0.000 description 1
- 229910000403 monosodium phosphate Inorganic materials 0.000 description 1
- 235000019799 monosodium phosphate Nutrition 0.000 description 1
- 229930014626 natural product Natural products 0.000 description 1
- 239000002245 particle Substances 0.000 description 1
- 230000001766 physiological effect Effects 0.000 description 1
- USHAGKDGDHPEEY-UHFFFAOYSA-L potassium persulfate Chemical compound [K+].[K+].[O-]S(=O)(=O)OOS([O-])(=O)=O USHAGKDGDHPEEY-UHFFFAOYSA-L 0.000 description 1
- OTYBMLCTZGSZBG-UHFFFAOYSA-L potassium sulfate Chemical compound [K+].[K+].[O-]S([O-])(=O)=O OTYBMLCTZGSZBG-UHFFFAOYSA-L 0.000 description 1
- 229910052939 potassium sulfate Inorganic materials 0.000 description 1
- 235000011151 potassium sulphates Nutrition 0.000 description 1
- 239000000047 product Substances 0.000 description 1
- 239000012488 sample solution Substances 0.000 description 1
- 238000000926 separation method Methods 0.000 description 1
- AJPJDKMHJJGVTQ-UHFFFAOYSA-M sodium dihydrogen phosphate Chemical compound [Na+].OP(O)([O-])=O AJPJDKMHJJGVTQ-UHFFFAOYSA-M 0.000 description 1
- 238000000638 solvent extraction Methods 0.000 description 1
- 229910001220 stainless steel Inorganic materials 0.000 description 1
- 239000010935 stainless steel Substances 0.000 description 1
- 238000010998 test method Methods 0.000 description 1
- 238000003828 vacuum filtration Methods 0.000 description 1
- 238000005406 washing Methods 0.000 description 1
Images
Classifications
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K8/00—Cosmetics or similar toiletry preparations
- A61K8/18—Cosmetics or similar toiletry preparations characterised by the composition
- A61K8/96—Cosmetics or similar toiletry preparations characterised by the composition containing materials, or derivatives thereof of undetermined constitution
- A61K8/97—Cosmetics or similar toiletry preparations characterised by the composition containing materials, or derivatives thereof of undetermined constitution from algae, fungi, lichens or plants; from derivatives thereof
- A61K8/9783—Angiosperms [Magnoliophyta]
- A61K8/9789—Magnoliopsida [dicotyledons]
-
- 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
- A23L33/00—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof
- A23L33/10—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof using additives
- A23L33/105—Plant extracts, their artificial duplicates or their derivatives
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61Q—SPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
- A61Q19/00—Preparations for care of the skin
-
- C—CHEMISTRY; METALLURGY
- C09—DYES; PAINTS; POLISHES; NATURAL RESINS; ADHESIVES; COMPOSITIONS NOT OTHERWISE PROVIDED FOR; APPLICATIONS OF MATERIALS NOT OTHERWISE PROVIDED FOR
- C09K—MATERIALS FOR MISCELLANEOUS APPLICATIONS, NOT PROVIDED FOR ELSEWHERE
- C09K15/00—Anti-oxidant compositions; Compositions inhibiting chemical change
- C09K15/34—Anti-oxidant compositions; Compositions inhibiting chemical change containing plant or animal materials of unknown composition
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/17—Systems in which incident light is modified in accordance with the properties of the material investigated
- G01N21/25—Colour; Spectral properties, i.e. comparison of effect of material on the light at two or more different wavelengths or wavelength bands
- G01N21/31—Investigating relative effect of material at wavelengths characteristic of specific elements or molecules, e.g. atomic absorption spectrometry
- G01N21/33—Investigating relative effect of material at wavelengths characteristic of specific elements or molecules, e.g. atomic absorption spectrometry using ultraviolet light
-
- 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
- A61K2800/00—Properties of cosmetic compositions or active ingredients thereof or formulation aids used therein and process related aspects
- A61K2800/40—Chemical, physico-chemical or functional or structural properties of particular ingredients
- A61K2800/52—Stabilizers
- A61K2800/522—Antioxidants; Radical scavengers
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K2800/00—Properties of cosmetic compositions or active ingredients thereof or formulation aids used therein and process related aspects
- A61K2800/80—Process related aspects concerning the preparation of the cosmetic composition or the storage or application thereof
- A61K2800/805—Corresponding aspects not provided for by any of codes A61K2800/81 - A61K2800/95
Landscapes
- Life Sciences & Earth Sciences (AREA)
- Health & Medical Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Physics & Mathematics (AREA)
- Botany (AREA)
- Engineering & Computer Science (AREA)
- General Health & Medical Sciences (AREA)
- Mycology (AREA)
- Spectroscopy & Molecular Physics (AREA)
- Animal Behavior & Ethology (AREA)
- Public Health (AREA)
- Veterinary Medicine (AREA)
- Dermatology (AREA)
- Biotechnology (AREA)
- Analytical Chemistry (AREA)
- Biochemistry (AREA)
- General Physics & Mathematics (AREA)
- Immunology (AREA)
- Pathology (AREA)
- Organic Chemistry (AREA)
- Microbiology (AREA)
- Materials Engineering (AREA)
- Birds (AREA)
- Epidemiology (AREA)
- Nutrition Science (AREA)
- Food Science & Technology (AREA)
- Polymers & Plastics (AREA)
- Agricultural Chemicals And Associated Chemicals (AREA)
Abstract
The invention discloses an extraction method of camphor flower active substances and a determination method of oxidation resistance. The extraction method of the active substances of the camphor flower comprises the following steps: cleaning fresh picked camphor flower, and freeze-drying to remove water; extracting with a flash extractor, standing overnight, filtering to obtain extractive solution, and steaming to obtain flos Cinnamomi Camphorae extract; dissolving the extract of the flowers of Cinnamomum camphora in absolute ethanol to obtain stock solution. The method for measuring the oxidation resistance of the active substance of the cinnamomum camphora flower comprises the following steps: diluting stock solution prepared by the method for extracting the active substances of the camphor flowers into different concentration gradients, and measuring DPPH free radical scavenging capacity, ABTS free radical scavenging capacity and ferric ion reducing capacity by using vitamin C with the same concentration as a positive control. The invention efficiently extracts the active substances in the camphor flower, and the extract has stronger DPPH and ABTS free radical scavenging capacity and ferric ion reducing power.
Description
Technical Field
The invention relates to an extraction method of camphor flower active substances and a determination method of oxidation resistance, and belongs to the technical field of extraction of natural products.
Background
The flowers of Cinnamomum camphora (Cinnamomum camphora) are flowers of Cinnamomum camphora (Cinnamomum camphora), which are usually distributed in southern and southwest provinces of China in 4-5 months. The full camphor tree has camphor fragrance and is mainly used for extraction and production of camphor. The overall shape of the camphor flower is small, the flower length is about 5mm, the camphor flower is light yellow when in blooming, and the fragrance is fresh and elegant. At present, researches on camphor trees mostly concentrate on branches, leaves and trunks, and the researches show that the branches, leaves and the like of the camphor trees have strong physiological activities such as oxidation resistance, antibacterial property and the like, but relevant reports on extraction and activity research of camphor flowers are not provided.
Flash extraction is a plant tissue crushing extraction method, materials are crushed at a high speed to a lower particle size at room temperature, active ingredients in the materials can be efficiently extracted by being assisted with a proper solvent, the extraction efficiency is higher compared with that of a conventional solvent extraction method, the required solvent amount is less, and the flash extraction method is widely applied to primary separation and extraction of plant tissues.
Disclosure of Invention
The technical problem to be solved by the invention is as follows: how to extract the active substances in the camphor flower with high efficiency and initially explore the antioxidant activity.
In order to solve the technical problem, the invention provides a method for extracting active substances from camphor flowers, which is characterized by comprising the following steps of:
Step 1), taking freshly picked camphor flowers, cleaning, and freeze-drying to remove water;
step 2): selecting absolute ethyl alcohol as a solvent, extracting by using a flash extractor, standing overnight, filtering to obtain an extracting solution, and performing rotary steaming on the extracting solution to obtain a camphor flower extract;
step 3): dissolving the extract of the flowers of Cinnamomum camphora in absolute ethanol to obtain stock solution.
Preferably, the mass ratio of the camphor flower to the absolute ethyl alcohol in the step 2) is 1: 10; the power of the flash extractor is 150W, and the extraction time is 1 min.
Preferably, the concentration of the stock solution in the step 3) is 40 mg/mL.
The invention also provides a method for measuring the oxidation resistance of the camphor flower active substance, which is characterized in that stock solution prepared by the method for extracting the camphor flower active substance is diluted into different concentration gradients, and the DPPH free radical scavenging capacity, ABTS free radical scavenging capacity and ferric ion reducing capacity are measured by using the same concentration vitamin C as a positive control.
Preferably, when determining the DPPH free radical scavenging ability and ABTS free radical scavenging ability, the concentration of the dilution and the concentration of the vitamin C in the stock solution are not more than 320 mu g/mL.
More preferably, the DPPH radical scavenging ability is measured by: respectively diluting the stock solution and the vitamin C into different gradient diluents; putting 150 mu L of each sample with concentration and 150 mu L of absolute ethanol solution of DPPH with concentration of 1.5mM into a 96-well plate, uniformly mixing, storing at 30 ℃ in a dark place, and measuring the absorbance value by using a microplate reader after 30 min; the DPPH radical clearance formula is shown as formula 1:
in the formula 1, ABlank spaceAbsorbance of 150. mu.L of absolute ethanol and 150. mu.L of DPPH solution, ASample (I)The absorbance was 150. mu.L of sample and 150. mu.L of DPPH solution.
More preferably, the determination method of the ABTS free radical scavenging ability is as follows: preparing needed ABTS working solution in advance, diluting the camphor flower stock solution and vitamin C into different gradient diluents respectively, placing 0.15mL samples and 2.85mL ABTS working solution of each concentration into a test tube, standing for 8min at 30 ℃, taking absolute ethyl alcohol as a blank, and measuring absorbance at 734nm by using an ultraviolet spectrophotometer; the formula for ABTS free radical clearance is shown in formula 2:
in the formula 2, ABlank spaceThe absorbance of 0.15mL of absolute ethanol and 2.85mL of ABTS working solution, ASample (I)Absorbance was 0.15mL of sample and 2.85mL of ABTS working solution.
Preferably, when the reduction capability of the ferric ions is measured, the concentration of the diluent and the concentration of the vitamin C in the stock solution are not more than 1 mg/mL.
More preferably, the method for measuring the reduction capability of the ferric ions comprises the following steps: respectively diluting the stock solution and the vitamin C into different gradient diluents; mixing the samples with each concentration evenly with 2.5mL of 0.2mM sodium phosphate buffer solution with pH value of 6.6 and 2.5mL of 1 wt% potassium ferricyanide solution, and incubating the mixed solution for 20min at 50 ℃; after rapid cooling, 2.5mL of 10 wt% trichloroacetic acid solution is added, and then the solution is centrifuged at 6000r/min for 10min, and the upper solution is taken; 2.5mL of the supernatant, 2.5mL of distilled water, and 0.5mL of a 0.1 wt% ferric chloride solution were mixed well, and the absorbance value was measured at 700nm using an ultraviolet spectrophotometer.
The invention efficiently extracts the active substances in the camphor flowers and performs preliminary exploration on the antioxidant activity. The method has the advantages of simple process, low energy consumption and high extraction rate, and can quickly obtain the active substances in the camphor flowers. Experiments show that compared with positive control vitamin C, the extract has stronger DPPH and ABTS free radical scavenging capacity and ferric ion reducing power, and has potential application prospects in the fields of daily chemicals and foods.
Drawings
FIG. 1 is a schematic representation of the scavenging effect of Cinnamomum camphora flower extract and vitamin C on 1, 1-diphenyl-2-picrylhydrazino (free radical) (DPPH) free radicals;
FIG. 2 is a schematic representation of the scavenging effect of Cinnamomum camphora flower extract and vitamin C on free radicals of 2, 2-diaza-bis (3-ethyl-benzothiazole-6-sulfonic acid) diammonium salt (ABTS);
fig. 3 is a schematic diagram of the total reducing power of the cinnamomum camphora flower extract and vitamin C to ferric ions.
Detailed Description
In order to make the invention more comprehensible, preferred embodiments are described in detail below with reference to the accompanying drawings.
Example 1
Extracting active ingredients of the camphor flower:
materials and methods
1.1 materials and Main instruments
Picking the camphor flowers from Shanghai city Fengxian Bay town; anhydrous ethanol was purchased from merck chemical technology (shanghai) ltd, 1-diphenyl-2-picrylhydrazino (free radical) (DPPH), 2-diaza-bis (3-ethyl-benzothiazole-6-sulfonic acid) diammonium salt (ABTS), potassium persulfate, disodium hydrogen phosphate, sodium dihydrogen phosphate, trichloroacetic acid, potassium ferricyanide, ferric chloride, and the like, and purchased from shanghai nationality chemical agents ltd.
Flash extractor JHBE-50T type (Henan Zhi Jing Biotech, Inc.); model FD-1A-50 of the freeze dryer (shanghai belang instruments manufacturing ltd); electronic balance PL203 type (mettler-toledo instruments (shanghai) ltd); ultraviolet spectrophotometer UV-6000 type (Shanghai Meta analytical instruments Limited); a multifunctional microplate reader TECAN-M200PRO type (Austrian Dirken Co., Ltd.); micropipette guns (france GILSON), rotary evaporators, vacuum pumps, various glassware, etc.
1.2 test methods
1.2.1 flash extraction of active substances from flowers of Cinnamomum camphora
Picking fresh camphor flowers, washing impurities and rotten petals in the camphor flowers with clean water;
primarily filtering the washed camphor flower to remove water, placing the camphor flower in a lower layer of a refrigerator for freezing, and removing the water in the camphor flower by using a freeze dryer for later use;
(2) accurately weighing 50 g of freeze-dried camphor flower in a stainless steel extraction tank, adding 500mL of absolute ethyl alcohol, fully extracting by using a flash extractor under the crushing power of 150W and the crushing time of 1min, and standing the extracting solution overnight;
(3) removing flower residue by vacuum filtration with a vacuum pump to obtain extractive solution, removing ethanol with a rotary evaporator to obtain flos Cinnamomi Camphorae extract 2.08g, weighing 400mg, dissolving in 10mL anhydrous ethanol to obtain high concentration stock solution, and placing in refrigerator for use.
Example 2
Determination of DPPH free radical clearance rate of camphor flower extract:
the high-concentration stock solution and the reference vitamin C are respectively diluted into gradient dilution solutions with the concentrations of 20 mug/mL, 40 mug/mL, 80 mug/mL, 160 mug/mL and 320 mug/mL.
Samples with the concentration of 150 mu L and 150 mu L of DPPH solution (diluted by absolute ethyl alcohol) with the concentration of 1.5mM are placed in a 96-well plate, mixed evenly, stored in a dark place at 30 ℃, and the absorbance value is measured by using a microplate reader after 30 min. Vitamin C was used as a positive control and DPPH free radical clearance formula was as follows:
In the formula, ABlank spaceAbsorbance of 150. mu.L of absolute ethanol and 150. mu.L of DPPH solution, ASample (I)Absorbance for 150. mu.L of sample and 150. mu.L of DPPH solution; IC (integrated circuit)50The concentration of the sample at which DPPH free radical scavenging rate reached 50%. The IC of the experiments for eliminating DPPH free radical from the cinnamomum camphora flower extract can be calculated by the curve fitting formula in figure 150＝59.9μg/mL。
Example 3
Determination of the ABTS free radical clearance rate of the cinnamomum camphora flower extract:
(1) diluting the high-concentration camphor flower extract stock solution and a reference product vitamin C into gradient diluents with the concentrations of 20 mug/mL, 80 mug/mL, 160 mug/mL and 320 mug/mL respectively.
(2) A7 mmol/L ABTS solution and a 140mmol/L high potassium sulfate solution were mixed at a ratio of 62.5: 1, standing overnight in the dark at room temperature to form ABTS stock solution, and diluting with anhydrous ethanol to obtain ABTS working solution before use, wherein the absorbance of the working solution is required to be 0.7 +/-0.02 at 734nm for later use.
(3) Respectively diluting the camphor flower extract stock solution and vitamin C into diluents with different gradients in concentration; during measurement, 0.15mL of sample solution is mixed with 2.85mL of ABTS working solution, the mixture is kept stand at 30 ℃ for 8min, and the absorbance is measured at 734nm by taking absolute ethyl alcohol as a blank.
The formula for ABTS free radical clearance is shown in formula 2:
In the formula 2, ABlank spaceThe absorbance of 0.15mL of absolute ethanol and 2.85mL of ABTS working solution, ASample (I)Absorbance was 0.15mL of sample and 2.85mL of ABTS working solution. IC (integrated circuit)50Is the concentration of the sample at which the absorbance value measured was 0.5. The IC of the camphorwood flower extract ABTS free radical scavenging experiment can be calculated by a curve fitting formula in figure 250＝94.81μg/mL。
Example 4
And (3) measuring the total reducing power of the iron ions of the cinnamomum camphora flower extract:
(1) diluting the high-concentration stock solution and the reference vitamin C into gradient dilution solution with concentration of 0-1mg/mL respectively.
(2) Mixing the samples with each concentration with sodium phosphate buffer solution (2.5mL, 0.2mM, pH 6.6) and 2.5mL of 1 wt% potassium ferricyanide solution, and incubating the mixture at 50 deg.C for 20 min;
(3) after rapid cooling, 2.5mL of 10 wt% trichloroacetic acid solution is added, and then the solution is centrifuged for 10min at 6000r/min, and the upper solution is taken;
(4) 2.5mL of the supernatant, 2.5mL of distilled water and 0.5mL of a 0.1 wt% ferric chloride solution were mixed well, and the absorbance value was measured at 700nm using an ultraviolet spectrophotometer. A high absorbance indicates a high reducing power. EC50 is the concentration of the sample at which the absorbance value measured was 0.5. EC of the total reducing power of the iron ions of the camphor flower extract can be calculated by a curve fitting formula in figure 3 50＝46.87μg/mL。
Claims (9)
1. The method for extracting the active substances of the cinnamomum camphora flowers is characterized by comprising the following steps:
step 1), taking freshly picked camphor flowers, cleaning, and freeze-drying to remove water;
step 2): selecting absolute ethyl alcohol as a solvent, extracting by using a flash extractor, standing overnight, filtering to obtain an extracting solution, and performing rotary steaming on the extracting solution to obtain a camphor flower extract;
step 3): dissolving the extract of the flowers of Cinnamomum camphora in absolute ethanol to obtain stock solution.
2. The method for extracting active substance from cinnamomum camphora flower according to claim 1, wherein the mass ratio of cinnamomum camphora flower to absolute ethyl alcohol in step 2) is 1: 10; the power of the flash extractor is 150W, and the extraction time is 1 min.
3. The method for extracting active ingredients from cinnamomum camphora flower according to claim 1, wherein the concentration of the stock solution in step 3) is 40 mg/mL.
4. A method for measuring oxidation resistance of a camphor flower active substance, which is characterized in that stock solutions prepared by the method for extracting the camphor flower active substance according to any one of claims 1 to 3 are diluted into different concentration gradients, and the DPPH free radical scavenging capacity, ABTS free radical scavenging capacity and ferric ion reducing capacity are measured by using vitamin C with the same concentration as a positive control.
5. The method for measuring the antioxidant activity of the active ingredient of cinnamomum camphora flower according to claim 4, wherein the concentrations of the diluent and vitamin C in the stock solution are not more than 320 μ g/mL when the DPPH free radical scavenging ability and ABTS free radical scavenging ability are measured.
6. The method for measuring the antioxidant capacity of the camphor flower active material according to claim 5, wherein the DPPH free radical scavenging capacity is measured by: respectively diluting the stock solution and the vitamin C into different gradient diluents; putting 150 mu L of each sample with concentration and 150 mu L of absolute ethanol solution of DPPH with concentration of 1.5mM into a 96-well plate, uniformly mixing, storing at 30 ℃ in a dark place, and measuring the absorbance value by using a microplate reader after 30 min; the DPPH radical clearance formula is shown as formula 1:
in the formula 1, ABlank spaceAbsorbance of 150. mu.L of absolute ethanol and 150. mu.L of DPPH solution, ASample (I)The absorbance was 150. mu.L of the sample and 150. mu.L of the LDPPH solution.
7. The method for measuring the oxidation resistance of the camphor flower active substance according to claim 5, wherein the ABTS free radical scavenging capacity is measured by: preparing needed ABTS working solution in advance, diluting the camphor flower stock solution and vitamin C into different gradient diluents respectively, placing 0.15mL samples and 2.85mL ABTS working solution of each concentration into a test tube, standing for 8min at 30 ℃, taking absolute ethyl alcohol as a blank, and measuring absorbance at 734nm by using an ultraviolet spectrophotometer; the formula for ABTS free radical clearance is shown in formula 2:
In the formula 2, ABlank spaceThe absorbance of the working solution was 0.15mL of absolute ethanol and 2.85mL of ABTS, ASample (I)Absorbance was 0.15mL of sample and 2.85mL of ABTS working solution.
8. The method for measuring the antioxidant ability of the active ingredient of cinnamomum camphora flower according to claim 4, wherein the concentration of the diluent and the concentration of vitamin C in the stock solution are not more than 1mg/mL when the reducing power of ferric ions is measured.
9. The method for measuring the oxidation resistance of the camphor flower active material according to claim 8, wherein the method for measuring the reduction capacity of the ferric ions comprises the following steps: respectively diluting the stock solution and the vitamin C into different gradient diluents; uniformly mixing the samples with various concentrations with 2.5mL of 0.2mM sodium phosphate buffer solution with the pH value of 6.6 and 2.5mL of 1 wt% potassium ferricyanide solution, and incubating the mixed solution for 20min at 50 ℃; after rapid cooling, 2.5mL of 10 wt% trichloroacetic acid solution is added, and then the solution is centrifuged at 6000r/min for 10min, and the upper solution is taken; 2.5mL of the supernatant, 2.5mL of distilled water, and 0.5mL of a 0.1 wt% ferric chloride solution were mixed well, and the absorbance value was measured at 700nm using an ultraviolet spectrophotometer.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202010679067.8A CN111840156A (en) | 2020-07-15 | 2020-07-15 | Extraction method of active substance of cinnamomum camphora flower and determination method of oxidation resistance |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202010679067.8A CN111840156A (en) | 2020-07-15 | 2020-07-15 | Extraction method of active substance of cinnamomum camphora flower and determination method of oxidation resistance |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN111840156A true CN111840156A (en) | 2020-10-30 |
Family
ID=72983511
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202010679067.8A Pending CN111840156A (en) | 2020-07-15 | 2020-07-15 | Extraction method of active substance of cinnamomum camphora flower and determination method of oxidation resistance |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN111840156A (en) |
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN110954379A (en) * | 2018-09-27 | 2020-04-03 | 深圳钜运生物科技有限公司 | Method for evaluating antioxidant capacity of fruits |
| CN113075016A (en) * | 2021-04-01 | 2021-07-06 | 上海应用技术大学 | Extraction process and activity determination method for ultrasonically-assisted flash extraction of osmanthus hydrolat |
| CN113149177A (en) * | 2021-05-26 | 2021-07-23 | 华侨大学 | Method for quickly removing indole in water |
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| KR20030024284A (en) * | 2001-09-17 | 2003-03-26 | 한불화장품주식회사 | A COSMETIC COMPOSITION CONTAINING AN EXTRACT OF Cinnamomum camphora |
| CN104830537A (en) * | 2015-04-30 | 2015-08-12 | 东莞波顿香料有限公司 | Preparation method and application of aquilaria wood hydrolat |
| CN106226431A (en) * | 2016-08-08 | 2016-12-14 | 西北农林科技大学 | A kind of persons of royal lineage's Caulis et folium euphorbiae milii leaf tea antioxidant content composition and Quality Analysis Methods |
-
2020
- 2020-07-15 CN CN202010679067.8A patent/CN111840156A/en active Pending
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| KR20030024284A (en) * | 2001-09-17 | 2003-03-26 | 한불화장품주식회사 | A COSMETIC COMPOSITION CONTAINING AN EXTRACT OF Cinnamomum camphora |
| CN104830537A (en) * | 2015-04-30 | 2015-08-12 | 东莞波顿香料有限公司 | Preparation method and application of aquilaria wood hydrolat |
| CN106226431A (en) * | 2016-08-08 | 2016-12-14 | 西北农林科技大学 | A kind of persons of royal lineage's Caulis et folium euphorbiae milii leaf tea antioxidant content composition and Quality Analysis Methods |
Non-Patent Citations (5)
| Title |
|---|
| 吴学文等: "湖南产樟树不同部位精油分析", 《天然产物研究与开发》 * |
| 昝丽霞: "香樟果皮花色苷提取工艺优化及抗氧化研究", 《广西植物》 * |
| 李嘉欣等: "樟树叶精油组成分析及抗氧化活性研究", 《林产化学与工业》 * |
| 杨玉红等: "《生命科学综合实验指导》", 31 July 2016, 东北大学出版社 * |
| 林翔云: "《香樟开发利用》", 31 January 2010, 化学工业出版社 * |
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN110954379A (en) * | 2018-09-27 | 2020-04-03 | 深圳钜运生物科技有限公司 | Method for evaluating antioxidant capacity of fruits |
| CN113075016A (en) * | 2021-04-01 | 2021-07-06 | 上海应用技术大学 | Extraction process and activity determination method for ultrasonically-assisted flash extraction of osmanthus hydrolat |
| CN113149177A (en) * | 2021-05-26 | 2021-07-23 | 华侨大学 | Method for quickly removing indole in water |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN111840156A (en) | Extraction method of active substance of cinnamomum camphora flower and determination method of oxidation resistance | |
| CN101906030B (en) | Method for extracting juglone from walnut green husk residue by microwave reflux | |
| CN105560888B (en) | A kind of method of polyphenol component leaching rate in raising dendrobium candidum | |
| CN110467528A (en) | A method of extracting carnosic acid from rosemary | |
| CN102512354B (en) | Bamboo fungus alcohol extract and preparation method and application thereof | |
| CN103766913B (en) | A kind of method extracting total stilbene compound from seed of Flos Moutan | |
| Al-Muwaly et al. | Antioxidant and free radical scavenging effects of Iraqi sumac (Rhus coriaria L) | |
| CN104106627A (en) | Vegetable fresh-keeping composition containing fructus forsythiae leaf extract and preparation method thereof | |
| CN103254321B (en) | The method for extraction and purification of medicinal fungi Phellinus vaninii polysaccharide | |
| Deepak et al. | Phenolic quantification and anti-oxidant activity of Morchella esculenta | |
| CN107936265A (en) | The extracting method of plant condensed tannin high polymer | |
| CN101941998A (en) | Method for extracting and measuring anthocyanin with special biological activity | |
| CN110507677A (en) | A kind of extracting method of sargassum fusifome total polyphenols | |
| CN113367166B (en) | Eugenia jambolana extract bactericide as well as preparation method and application thereof | |
| EP3243828A1 (en) | Method for extracting stilbene compounds | |
| CN106421047B (en) | Honey pomelo leaf extract and preparation method and application thereof | |
| CN104628878A (en) | Crateva unilocalaris Buch. polysaccharide extraction and purification method and application of crateva unilocalaris Buch. polysaccharide | |
| CN107556348A (en) | Acryloyl acid esters compound and preparation method thereof | |
| CN113925081A (en) | Natural plant anticorrosion composition, preparation method and application thereof | |
| CN113440551A (en) | Salvia miltiorrhiza residue extract with antioxidant activity and application thereof | |
| CN111407699A (en) | Camellia extract, preparation method and application thereof, and cosmetic containing camellia extract | |
| CN110845637A (en) | Extraction method of dendrobium pohuanense polysaccharide | |
| Wang et al. | Microwave-ultrasonic synergistic extraction of crude se-polysaccharides from se-enriched tea | |
| CN111544494A (en) | HPEF extraction method of tea seed phenolic acid | |
| CN111096456A (en) | Preparation method of active component for inhibiting formation of late glycosylation product in lotus leaf |
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 | ||
| RJ01 | Rejection of invention patent application after publication |
Application publication date: 20201030 |
|
| RJ01 | Rejection of invention patent application after publication |