CN113698992A - Preparation method of Eurya emarginata essential oil - Google Patents
Preparation method of Eurya emarginata essential oil Download PDFInfo
- Publication number
- CN113698992A CN113698992A CN202111185848.2A CN202111185848A CN113698992A CN 113698992 A CN113698992 A CN 113698992A CN 202111185848 A CN202111185848 A CN 202111185848A CN 113698992 A CN113698992 A CN 113698992A
- Authority
- CN
- China
- Prior art keywords
- eurya emarginata
- eurya
- emarginata
- essential oil
- supernatant
- 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
- 241000007108 Eurya emarginata Species 0.000 title claims abstract description 94
- 239000000341 volatile oil Substances 0.000 title claims abstract description 61
- 238000002360 preparation method Methods 0.000 title claims abstract description 14
- 239000000843 powder Substances 0.000 claims abstract description 28
- 239000006228 supernatant Substances 0.000 claims abstract description 28
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims abstract description 13
- 238000010438 heat treatment Methods 0.000 claims abstract description 9
- 238000004506 ultrasonic cleaning Methods 0.000 claims abstract description 9
- 238000004108 freeze drying Methods 0.000 claims abstract description 8
- 239000003814 drug Substances 0.000 claims abstract description 7
- 230000010355 oscillation Effects 0.000 claims abstract description 7
- 238000003756 stirring Methods 0.000 claims abstract description 7
- 238000000034 method Methods 0.000 claims description 10
- 239000002245 particle Substances 0.000 claims description 6
- 238000000926 separation method Methods 0.000 claims description 2
- 230000003385 bacteriostatic effect Effects 0.000 abstract description 8
- 238000012360 testing method Methods 0.000 description 35
- 241000196324 Embryophyta Species 0.000 description 11
- 230000001580 bacterial effect Effects 0.000 description 11
- 239000007788 liquid Substances 0.000 description 11
- 241000222122 Candida albicans Species 0.000 description 10
- 229940095731 candida albicans Drugs 0.000 description 10
- 238000005070 sampling Methods 0.000 description 9
- 239000000725 suspension Substances 0.000 description 9
- 238000004879 turbidimetry Methods 0.000 description 9
- 241000588724 Escherichia coli Species 0.000 description 7
- 241000191967 Staphylococcus aureus Species 0.000 description 7
- 239000001963 growth medium Substances 0.000 description 7
- 239000003153 chemical reaction reagent Substances 0.000 description 6
- 238000012258 culturing Methods 0.000 description 6
- 238000007598 dipping method Methods 0.000 description 6
- 239000011521 glass Substances 0.000 description 6
- 238000005259 measurement Methods 0.000 description 6
- 238000007789 sealing Methods 0.000 description 6
- 238000003892 spreading Methods 0.000 description 6
- 230000007480 spreading Effects 0.000 description 6
- 238000010998 test method Methods 0.000 description 6
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 6
- 230000005764 inhibitory process Effects 0.000 description 5
- 239000002609 medium Substances 0.000 description 5
- 238000002835 absorbance Methods 0.000 description 4
- 238000002156 mixing Methods 0.000 description 4
- 229920001817 Agar Polymers 0.000 description 3
- 241000894006 Bacteria Species 0.000 description 3
- 239000006137 Luria-Bertani broth Substances 0.000 description 3
- 239000001888 Peptone Substances 0.000 description 3
- 108010080698 Peptones Proteins 0.000 description 3
- 239000008272 agar Substances 0.000 description 3
- 235000015278 beef Nutrition 0.000 description 3
- 239000003795 chemical substances by application Substances 0.000 description 3
- 239000011248 coating agent Substances 0.000 description 3
- 238000000576 coating method Methods 0.000 description 3
- 238000001816 cooling Methods 0.000 description 3
- 238000005485 electric heating Methods 0.000 description 3
- 235000019441 ethanol Nutrition 0.000 description 3
- 238000000605 extraction Methods 0.000 description 3
- 230000006698 induction Effects 0.000 description 3
- 239000000203 mixture Substances 0.000 description 3
- 235000019319 peptone Nutrition 0.000 description 3
- 229910052573 porcelain Inorganic materials 0.000 description 3
- 239000002244 precipitate Substances 0.000 description 3
- 239000008223 sterile water Substances 0.000 description 3
- 238000003860 storage Methods 0.000 description 3
- 230000009471 action Effects 0.000 description 2
- 230000008859 change Effects 0.000 description 2
- 238000011160 research Methods 0.000 description 2
- 241000220317 Rosa Species 0.000 description 1
- 230000000844 anti-bacterial effect Effects 0.000 description 1
- 230000000845 anti-microbial effect Effects 0.000 description 1
- 239000010692 aromatic oil Substances 0.000 description 1
- 125000003118 aryl group Chemical group 0.000 description 1
- 230000001174 ascending effect Effects 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 238000005119 centrifugation Methods 0.000 description 1
- 238000011161 development Methods 0.000 description 1
- 238000001035 drying Methods 0.000 description 1
- 238000005516 engineering process Methods 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 235000015097 nutrients Nutrition 0.000 description 1
- 230000000144 pharmacologic effect Effects 0.000 description 1
- 238000002390 rotary evaporation Methods 0.000 description 1
- 229930000044 secondary metabolite Natural products 0.000 description 1
- 238000001228 spectrum Methods 0.000 description 1
- 235000013599 spices Nutrition 0.000 description 1
- 238000001256 steam distillation Methods 0.000 description 1
- 230000001954 sterilising effect Effects 0.000 description 1
- 238000004659 sterilization and disinfection Methods 0.000 description 1
- 230000002459 sustained effect Effects 0.000 description 1
- 230000009897 systematic effect Effects 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
- 230000000007 visual effect Effects 0.000 description 1
Classifications
-
- C—CHEMISTRY; METALLURGY
- C11—ANIMAL OR VEGETABLE OILS, FATS, FATTY SUBSTANCES OR WAXES; FATTY ACIDS THEREFROM; DETERGENTS; CANDLES
- C11B—PRODUCING, e.g. BY PRESSING RAW MATERIALS OR BY EXTRACTION FROM WASTE MATERIALS, REFINING OR PRESERVING FATS, FATTY SUBSTANCES, e.g. LANOLIN, FATTY OILS OR WAXES; ESSENTIAL OILS; PERFUMES
- C11B9/00—Essential oils; Perfumes
- C11B9/02—Recovery or refining of essential oils from raw materials
-
- C—CHEMISTRY; METALLURGY
- C11—ANIMAL OR VEGETABLE OILS, FATS, FATTY SUBSTANCES OR WAXES; FATTY ACIDS THEREFROM; DETERGENTS; CANDLES
- C11B—PRODUCING, e.g. BY PRESSING RAW MATERIALS OR BY EXTRACTION FROM WASTE MATERIALS, REFINING OR PRESERVING FATS, FATTY SUBSTANCES, e.g. LANOLIN, FATTY OILS OR WAXES; ESSENTIAL OILS; PERFUMES
- C11B9/00—Essential oils; Perfumes
- C11B9/02—Recovery or refining of essential oils from raw materials
- C11B9/022—Refining
-
- C—CHEMISTRY; METALLURGY
- C11—ANIMAL OR VEGETABLE OILS, FATS, FATTY SUBSTANCES OR WAXES; FATTY ACIDS THEREFROM; DETERGENTS; CANDLES
- C11B—PRODUCING, e.g. BY PRESSING RAW MATERIALS OR BY EXTRACTION FROM WASTE MATERIALS, REFINING OR PRESERVING FATS, FATTY SUBSTANCES, e.g. LANOLIN, FATTY OILS OR WAXES; ESSENTIAL OILS; PERFUMES
- C11B9/00—Essential oils; Perfumes
- C11B9/02—Recovery or refining of essential oils from raw materials
- C11B9/025—Recovery by solvent extraction
-
- C—CHEMISTRY; METALLURGY
- C11—ANIMAL OR VEGETABLE OILS, FATS, FATTY SUBSTANCES OR WAXES; FATTY ACIDS THEREFROM; DETERGENTS; CANDLES
- C11B—PRODUCING, e.g. BY PRESSING RAW MATERIALS OR BY EXTRACTION FROM WASTE MATERIALS, REFINING OR PRESERVING FATS, FATTY SUBSTANCES, e.g. LANOLIN, FATTY OILS OR WAXES; ESSENTIAL OILS; PERFUMES
- C11B9/00—Essential oils; Perfumes
- C11B9/02—Recovery or refining of essential oils from raw materials
- C11B9/027—Recovery of volatiles by distillation or stripping
Abstract
The invention provides a preparation method of Eurya emarginata essential oil, which comprises the following steps: s1, placing the branches and leaves of Eurya emarginata into an oven to be dried for 22-26 hours, taking out the Eurya emarginata, and then transferring the Eurya emarginata into a traditional Chinese medicine pulverizer to be pulverized to obtain Eurya emarginata dry powder; s2, adding the Eurya emarginata dry powder obtained in the step S1 into absolute ethyl alcohol, uniformly stirring, placing the Eurya emarginata dry powder into an ultrasonic cleaning instrument, carrying out ultrasonic oscillation for 20-40 minutes to obtain an extracting solution, and standing the extracting solution for 2-4 days for layering to obtain a supernatant I; s3, adding the supernatant I obtained in the step S2 into a round-bottom flask, placing the round-bottom flask into a rotary evaporator, heating, and concentrating for 3-5 hours to obtain a concentrated solution; and S4, centrifuging the concentrated solution obtained in the step S3 by using a centrifugal machine for 5-15 minutes to obtain a supernatant II, and freeze-drying the supernatant II to obtain the Eurya emarginata essential oil. The Eurya emarginata essential oil is stable and reliable, and the prepared Eurya emarginata essential oil has good bacteriostatic performance.
Description
Technical Field
The invention relates to a preparation method of Eurya emarginata essential oil.
Background
With the development of society, people are increasingly pursuing quality of life, plant essential oil which is small in harm and environment-friendly and has multiple practical purposes gradually enters the visual field of people, plant essential oil such as rose essential oil is favored by people, and the extraction and research of the plant essential oil are hot topics all the time. Plant essential oil is also called volatile oil, essential oil or aromatic oil, and is a secondary metabolite with volatile aromatic odor generated by plants. The plant essential oil serving as a novel cold sterilization technology has multiple advantages, such as wide antimicrobial spectrum, obvious antibacterial effect and low residual toxicity. As early as four hundred years ago, some plant essential oil extraction and application data are recorded in detail in Bencao gang mu drafted by Li Shizhen. At present, plant essential oil is applied and developed in the aspects of spice industry, medical treatment, agriculture and the like, and has systematic research on the aspects of bacteriostasis, components, physiological action, pharmacological action and the like. The existing plant essential oil has the problems that some plant essential oil extraction cannot be carried out by the most basic steam distillation method in a laboratory, and the bacteriostatic effect of some plant essential oil is not good.
Disclosure of Invention
The invention aims to provide a preparation method of Eurya emarginata essential oil, the preparation method is stable and reliable, and the prepared Eurya emarginata essential oil has good bacteriostatic performance.
In order to solve the technical problems, the technical scheme of the invention is as follows:
a preparation method of Eurya emarginata essential oil comprises the following steps:
s1, placing the branches and leaves of Eurya emarginata into an oven to be dried for 22-26 hours, taking out the Eurya emarginata, and then transferring the Eurya emarginata into a traditional Chinese medicine pulverizer to be pulverized to obtain Eurya emarginata dry powder;
s2, adding the Eurya emarginata dry powder obtained in the step S1 into absolute ethyl alcohol, uniformly stirring, placing the Eurya emarginata dry powder into an ultrasonic cleaning instrument, carrying out ultrasonic oscillation for 20-40 minutes to obtain an extracting solution, and standing the extracting solution for 2-4 days for layering to obtain a supernatant I;
s3, adding the supernatant I obtained in the step S2 into a round-bottom flask, placing the round-bottom flask into a rotary evaporator, heating, and concentrating for 3-5 hours to obtain a concentrated solution;
and S4, centrifuging the concentrated solution obtained in the step S3 by using a centrifugal machine for 5-15 minutes to obtain a supernatant II, and freeze-drying the supernatant II to obtain the Eurya emarginata essential oil.
Further, in step S1 of the present invention, the particle size of the Eurya emarginata dry powder is 150 μm.
Further, in step S2 of the present invention, the ratio of the Eurya emarginata dry powder to the absolute ethanol is (0.5-1.5) g:2 mL.
Further, in step S2 of the present invention, the power of the ultrasonic cleaning apparatus is 70%.
Further, in step S3 of the present invention, the heating temperature is 70-80 ℃.
Further, in step S4 of the present invention, the speed of centrifugation was 5000 rpm.
Compared with the prior art, the invention has the following beneficial effects:
the Eurya emarginata essential oil is prepared by drying and crushing the branches and leaves of Eurya emarginata, mixing the branches and leaves with absolute ethyl alcohol serving as an extractant, carrying out ultrasonic extraction to obtain an extracting solution, standing the extracting solution for layering, carrying out rotary evaporation and concentration, carrying out centrifugal separation, and carrying out freeze drying.
Detailed Description
The present invention will be described in detail with reference to specific embodiments, which are illustrative of the invention and are not to be construed as limiting the invention.
Example 1
The Eurya emarginata essential oil is prepared according to the following steps:
s1, placing the branches and leaves of Eurya emarginata into an oven to be dried for 24 hours, taking out the Eurya emarginata, and transferring the Eurya emarginata into a traditional Chinese medicine pulverizer to be pulverized to obtain Eurya emarginata dry powder with the particle size of 150 mu m;
s2, adding the Eurya emarginata dry powder obtained in the step S1 into absolute ethyl alcohol according to the proportion of 1g to 2mL, uniformly stirring, placing the Eurya emarginata dry powder into an ultrasonic cleaning instrument with the power of 70%, carrying out ultrasonic oscillation for 30 minutes to obtain an extracting solution, and standing the extracting solution for 3 days for layering to obtain a first supernatant;
s3, adding the supernatant I obtained in the step S2 into a round-bottom flask, placing the round-bottom flask into a rotary evaporator, heating to 78 ℃, and concentrating for 4 hours to obtain a concentrated solution;
and S4, centrifuging the concentrated solution obtained in the step S3 for 10 minutes at the speed of 5000rpm of a centrifuge to obtain a supernatant II, and freeze-drying the supernatant II to obtain the Eurya emarginata essential oil.
The first test example: test for Candida albicans inhibition zone
Test equipment: a culture dish; a conical flask; an ultra-clean bench; 1000mL porcelain cup; an induction cooker; an electric heating constant temperature incubator; a constant temperature shaking table; a one-ten-thousandth balance; inoculating a loop; a fine lamp; coating rods; a filter paper sheet; a glass rod; an autoclave; and (4) a liquid transferring gun.
Test reagents: candida albicans strain; eurya emarginata essential oil prepared in example 1; LB broth; agar powder; and (3) water.
The test method comprises the following steps:
(1) taking a sterilized sterile plate, pouring the LB culture medium which is dissolved and cooled to about 50 ℃ into the plate according to a sterile operation method, and cooling the plate to prepare a flat plate.
(2) Preparing bacterial suspension, taking a sterile test tube, inoculating 1-2 rings of Candida albicans into sterile water by using an inoculating ring, and fully and uniformly preparing the bacterial suspension.
(3) Inoculating, sucking 0.1mL of the prepared bacterial suspension by using a sterile pipette, inoculating on a flat plate, and uniformly spreading by using a sterile glass shovel.
(4) And (3) dipping, namely dipping the sterile filter paper sheet into the supply agent (the Eurya emarginata essential oil prepared in example 1).
(5) Adding the medicinal preparation, clamping the medicinal filter paper with sterile forceps (taking care to drain the medicinal liquid), spreading on the same bacteria-containing plate, respectively, taking care that the medicinal preparation is not contaminated, performing mark culture on the back of the plate, culturing the plate at 37 deg.C for 48-72 hr, and observing the result.
The test results are shown in table 1:
TABLE 1
Test example two: test for inhibition zone of Escherichia coli
Test equipment: a culture dish; a conical flask; an ultra-clean bench; 1000mL porcelain cup; an induction cooker; an electric heating constant temperature incubator; a constant temperature shaking table; a one-ten-thousandth balance; inoculating a loop; a fine lamp; coating rods; a filter paper sheet; a glass rod; an autoclave; and (4) a liquid transferring gun.
Test reagents: e, Escherichia coli strains; eurya emarginata essential oil prepared in example 1; LB broth; agar powder; and (3) water.
The test method comprises the following steps:
(1) taking a sterilized sterile plate, pouring the LB culture medium which is dissolved and cooled to about 50 ℃ into the plate according to a sterile operation method, and cooling the plate to prepare a flat plate.
(2) Preparing bacterial suspension, taking a sterile test tube, inoculating 1-2 rings of Escherichia coli into sterile water by using an inoculating ring, and fully and uniformly preparing the bacterial suspension.
(3) Inoculating, sucking 0.1mL of the prepared bacterial suspension by using a sterile pipette, inoculating on a flat plate, and uniformly spreading by using a sterile glass shovel.
(4) And (3) dipping, namely dipping the sterile filter paper sheet into the supply agent (the Eurya emarginata essential oil prepared in example 1).
(5) Adding the medicinal preparation, clamping the medicinal filter paper with sterile forceps (taking care to drain the medicinal liquid), spreading on the same bacteria-containing plate, respectively, taking care that the medicinal preparation is not contaminated, performing mark culture on the back of the plate, culturing the plate at 37 deg.C for 48-72 hr, and observing the result.
The test results are shown in table 2:
| blank group | 1 | 2 | 3 | 4 | 5 | 6 | Mean value | |
| Diameter of bacteriostatic circle (mm) | 0 | 1.8 | 2.2 | 2.0 | 1.8 | 2.6 | 2.2 | 2.1 |
TABLE 2
Test example three: test for inhibition zone of Staphylococcus aureus
Test equipment: a culture dish; a conical flask; an ultra-clean bench; 1000mL porcelain cup; an induction cooker; an electric heating constant temperature incubator; a constant temperature shaking table; a one-ten-thousandth balance; inoculating a loop; a fine lamp; coating rods; a filter paper sheet; a glass rod; an autoclave; and (4) a liquid transferring gun.
Test reagents: a staphylococcus aureus species; eurya emarginata essential oil prepared in example 1; LB broth; agar powder; and (3) water.
The test method comprises the following steps:
(1) taking a sterilized sterile plate, pouring the LB culture medium which is dissolved and cooled to about 50 ℃ into the plate according to a sterile operation method, and cooling the plate to prepare a flat plate.
(2) Preparing bacterial suspension, taking a sterile test tube, inoculating 1-2 rings of staphylococcus aureus to sterile water by using an inoculating ring, and fully and uniformly preparing the bacterial suspension.
(3) Inoculating, sucking 0.1mL of the prepared bacterial suspension by using a sterile pipette, inoculating on a flat plate, and uniformly spreading by using a sterile glass shovel.
(4) And (3) dipping, namely dipping the sterile filter paper sheet into the supply agent (the Eurya emarginata essential oil prepared in example 1).
(5) Adding the medicinal preparation, clamping the medicinal filter paper with sterile forceps (taking care to drain the medicinal liquid), spreading on the same bacteria-containing plate, respectively, taking care that the medicinal preparation is not contaminated, performing mark culture on the back of the plate, culturing the plate at 37 deg.C for 48-72 hr, and observing the result.
The test results are shown in table 3:
TABLE 3
As can be seen from tables 1 to 3, the Eurya emarginata essential oil prepared in the example 1 of the invention has good bacteriostatic effect on Candida albicans, Escherichia coli and Staphylococcus aureus.
Test example four: turbidimetry for testing bacteriostatic effect of Eurya emarginata essential oil on Candida albicans
Test equipment: superclean bench, alcohol burner, spectrophotometer, cell, constant temperature shaking table, pipette, erlenmeyer flask, aseptic PC pipe.
Test reagents: beef extract peptone medium, candida albicans strain; the Eurya emarginata essential oil prepared in example 1.
The test method comprises the following steps:
the Eurya emarginata essential oil, 0.5mL and 1mL, was removed with a pipette and transferred to a 250mL Erlenmeyer flask containing 94mL and 95mL of LB liquid, and labeled 1 and 2, respectively. Transferring 5mL of candida albicans overnight culture solution (cultured for 10-12h) into a triangular flask containing 95mL of LB solution, uniformly mixing the culture solution and the triangular flasks with a blank label and labels 1 and 2, placing the mixture into a shaking table for shaking culture at 37 ℃ (shaking frequency is 250r/min), respectively culturing 0, 1.5, 3, 4, 6, 8, 10, 12, 14, 16, 18 and 20h, strictly sampling 3mL according to aseptic operation, placing the sample into a sterile PC tube, sealing the tube by using a sealing film, immediately placing the sample into a refrigerator for storage, and finally determining the absorbance at the same time. And (4) immediately putting the triangular flask back to the shaking table after sampling, and continuing shaking culture for subsequent sampling.
And (3) turbidimetry determination: the measurement was carried out by photoelectric turbidimetry using an uninoculated LB medium as a control at a wavelength of 600nm, and the measurement was carried out sequentially from the earliest initial culture broth.
The test results are shown in table 4:
| Time | 0 | 1.5 | 3 | 4 | 6 | 8 | 10 | 12 | 14 | 16 | 18 | 20 |
| blank group OD | 0.087 | 0.109 | 0.382 | 0.432 | 0.731 | 1.109 | 0.898 | 1.392 | 1.581 | 1.645 | 1.622 | 1.903 |
| No. 1 bottle OD | 0.539 | 0.604 | 0.739 | 0.87 | 0.98 | 1.105 | 1.029 | 1.339 | 1.453 | 1.674 | 1.675 | 0.821 |
| No. 2 bottle OD | 0.985 | 1.049 | 2.12 | 1.242 | 1.586 | 1.632 | 1.457 | 1.727 | 1.628 | 1.807 | 1.799 | 1.782 |
TABLE 4
As can be seen from table 4:
1. under normal culture conditions, the culture medium becomes more turbid with time, the concentration of the candida albicans is increased continuously, and the concentration of the candida albicans tends to be stable after about 12 hours.
2. In the culture medium containing 0.5mL of Eurya emarginata essential oil (bottle No. 1), the concentration of Candida albicans tended to increase gradually, but at 18h the bacterial liquid concentration dropped abruptly due to the consumption of nutrients and the inhibition of Eurya emarginata essential oil.
3. In the culture medium containing 1mL of Eurya emarginata essential oil (bottle No. 2), the bacterial liquid showed a large increase at 3 hours and then tended to a gradual increase at 4 hours, probably due to the sustained inhibition of Eurya emarginata essential oil.
Test example five: turbidimetry for testing bacteriostatic effect of Eurya emarginata essential oil on escherichia coli
Test equipment: superclean bench, alcohol burner, spectrophotometer, cell, constant temperature shaking table, pipette, erlenmeyer flask, aseptic PC pipe.
Test reagents: beef extract peptone culture medium, Escherichia coli strain; the Eurya emarginata essential oil prepared in example 1.
The test method comprises the following steps:
the Eurya emarginata essential oil, 0.5mL and 1mL, was removed with a pipette and transferred to a 250mL Erlenmeyer flask containing 94mL and 95mL of LB liquid, and labeled 1 and 2, respectively. Transferring 5mL of Escherichia coli overnight culture solution (cultured for 10-12h) into a triangular flask containing 95mL of LB solution, uniformly mixing the culture solution and the triangular flasks with a blank label and labels 1 and 2, placing the mixture in a shaking table for shaking culture at 37 ℃ (shaking frequency is 250r/min), respectively culturing 0, 1.5, 3, 4, 6, 8, 10, 12, 14, 16, 18 and 20h, strictly sampling 3mL of the culture solution according to aseptic operation, placing the sample into a sterile PC tube, sealing the tube by using a sealing film, immediately placing the sample into a refrigerator for storage, and finally determining the absorbance at the same time. And (4) immediately putting the triangular flask back to the shaking table after sampling, and continuing shaking culture for subsequent sampling.
And (3) turbidimetry determination: the measurement was carried out by photoelectric turbidimetry using an uninoculated LB medium as a control at a wavelength of 600nm, and the measurement was carried out sequentially from the earliest initial culture broth.
The test results are shown in table 5:
| Time | 0 | 1.5 | 3 | 4 | 6 | 8 | 10 | 12 | 14 | 16 | 18 | 20 |
| blank group OD | 0.173 | 0.232 | 0.411 | 0.409 | 0.297 | 0.494 | 0.598 | 0.619 | 0.789 | 0.893 | 0.946 | 0.795 |
| No. 1 bottle OD | 0.639 | 0.938 | 0.931 | 0.569 | 0.968 | 0.666 | 1.141 | 1.236 | 1.322 | 1.495 | 1.08 | 0.453 |
| No. 2 bottle OD | 0.469 | 0.597 | 0.531 | 0.737 | 0.658 | 0.664 | 0.744 | 0.747 | 0.847 | 0.857 | 1.074 | 0.944 |
TABLE 5
As can be seen from table 5:
bottle No. 2 is more stable than bottle No. 1 and bottle No. 2.
The overall trend of vial No. 2 was similar to that of the blank group, probably due to the lack of significant bacteriostasis in vial No. 2.
3. The occurrence of a precipitate in vial No. 1 was found during the test, and it was presumed that the sudden rise followed by the cliff-like drop in vial No. 1 was due to the precipitate, resulting in a large change in the concentration of the absorbance test sample taken.
Test example six: turbidimetry for testing bacteriostatic effect of Eurya emarginata essential oil on staphylococcus aureus
Test equipment: superclean bench, alcohol burner, spectrophotometer, cell, constant temperature shaking table, pipette, erlenmeyer flask, aseptic PC pipe.
Test reagents: beef extract peptone medium, staphylococcus aureus strain; the Eurya emarginata essential oil prepared in example 1.
The test method comprises the following steps:
the Eurya emarginata essential oil, 0.5mL and 1mL, was removed with a pipette and transferred to a 250mL Erlenmeyer flask containing 94mL and 95mL of LB liquid, and labeled 1 and 2, respectively. Transferring 5mL of staphylococcus aureus overnight culture solution (cultured for 10-12h) into a triangular flask containing 95mL of LB solution, uniformly mixing the culture solution and the triangular flask with a blank label and labels 1 and 2, placing the mixture into a shaking table for shaking culture at 37 ℃ (shaking frequency is 250r/min), respectively culturing 0, 1.5, 3, 4, 6, 8, 10, 12, 14, 16, 18 and 20h, strictly sampling 3mL according to aseptic operation, placing the sample into a sterile PC tube, sealing the tube by using a sealing film, immediately placing the sample into a refrigerator for storage, and finally determining the absorbance at the same time. And (4) immediately putting the triangular flask back to the shaking table after sampling, and continuing shaking culture for subsequent sampling.
And (3) turbidimetry determination: the measurement was carried out by photoelectric turbidimetry using an uninoculated LB medium as a control at a wavelength of 600nm, and the measurement was carried out sequentially from the earliest initial culture broth.
The test results are shown in table 6:
TABLE 6
As can be seen from table 6:
the data fluctuation is large, especially the trough value of the bottle No. 2 is even negative, while the bottle No. 1 with 0.5mL difference of the concentration of the essential oil from the bottle No. 2 is in an ascending trend and has a large number of wave peaks, and the occurrence of the situation can be caused by the generation of precipitates.
Example 2
The Eurya emarginata essential oil is prepared according to the following steps:
s1, placing the branches and leaves of Eurya emarginata into an oven to be dried for 22 hours, taking out the Eurya emarginata, and transferring the Eurya emarginata into a traditional Chinese medicine pulverizer to be pulverized to obtain Eurya emarginata dry powder with the particle size of 150 mu m;
s2, adding the Eurya emarginata dry powder obtained in the step S1 into absolute ethyl alcohol according to the proportion of 1.5g to 2mL, uniformly stirring, placing the Eurya emarginata dry powder into an ultrasonic cleaning instrument with the power of 70 percent, carrying out ultrasonic oscillation for 40 minutes to obtain an extracting solution, and standing the extracting solution for 2 days for layering to obtain a first supernatant;
s3, adding the supernatant I obtained in the step S2 into a round-bottom flask, placing the round-bottom flask into a rotary evaporator, heating to 80 ℃, and concentrating for 3 hours to obtain a concentrated solution;
and S4, centrifuging the concentrated solution obtained in the step S3 for 15 minutes at the speed of 5000rpm of a centrifuge to obtain a supernatant II, and freeze-drying the supernatant II to obtain the Eurya emarginata essential oil.
Example 3
The Eurya emarginata essential oil is prepared according to the following steps:
s1, placing the branches and leaves of Eurya emarginata into an oven to be dried for 26 hours, taking out the Eurya emarginata, and transferring the Eurya emarginata into a traditional Chinese medicine pulverizer to be pulverized to obtain Eurya emarginata dry powder with the particle size of 150 mu m;
s2, adding the Eurya emarginata dry powder obtained in the step S1 into absolute ethyl alcohol according to the proportion of 0.5g to 2mL, uniformly stirring, placing the Eurya emarginata dry powder into an ultrasonic cleaning instrument with the power of 70 percent, carrying out ultrasonic oscillation for 20 minutes to obtain an extracting solution, and standing the extracting solution for 4 days for layering to obtain a first supernatant;
s3, adding the supernatant I obtained in the step S2 into a round-bottom flask, placing the round-bottom flask into a rotary evaporator, heating to 70 ℃, and concentrating for 5 hours to obtain a concentrated solution;
and S4, centrifuging the concentrated solution obtained in the step S3 for 5 minutes at the speed of 5000rpm of a centrifuge to obtain a supernatant II, and freeze-drying the supernatant II to obtain the Eurya emarginata essential oil.
Example 4
The Eurya emarginata essential oil is prepared according to the following steps:
s1, placing the branches and leaves of Eurya emarginata into an oven to be dried for 25 hours, taking out the Eurya emarginata, and transferring the Eurya emarginata into a traditional Chinese medicine pulverizer to be pulverized to obtain Eurya emarginata dry powder with the particle size of 150 mu m;
s2, adding the Eurya emarginata dry powder obtained in the step S1 into absolute ethyl alcohol according to the proportion of 1.2g to 2mL, uniformly stirring, placing the Eurya emarginata dry powder into an ultrasonic cleaning instrument with the power of 70 percent, carrying out ultrasonic oscillation for 35 minutes to obtain an extracting solution, and standing the extracting solution for 3 days for layering to obtain a first supernatant;
s3, adding the supernatant I obtained in the step S2 into a round-bottom flask, placing the round-bottom flask into a rotary evaporator, heating to 75 ℃, and concentrating for 3.5 hours to obtain a concentrated solution;
and S4, centrifuging the concentrated solution obtained in the step S3 for 8 minutes at the speed of 5000rpm of a centrifuge to obtain a supernatant II, and freeze-drying the supernatant II to obtain the Eurya emarginata essential oil.
The foregoing embodiments are merely illustrative of the principles and utilities of the present invention and are not intended to limit the invention. Any person skilled in the art can modify or change the above-mentioned embodiments without departing from the spirit and scope of the present invention. Accordingly, it is intended that all equivalent modifications or changes which can be made by those skilled in the art without departing from the spirit and technical spirit of the present invention be covered by the claims of the present invention.
Claims (6)
1. A preparation method of Eurya emarginata essential oil is characterized by comprising the following steps: the method comprises the following steps:
s1, placing the branches and leaves of Eurya emarginata into an oven to be dried for 22-26 hours, taking out the Eurya emarginata, and then transferring the Eurya emarginata into a traditional Chinese medicine pulverizer to be pulverized to obtain Eurya emarginata dry powder;
s2, adding the Eurya emarginata dry powder obtained in the step S1 into absolute ethyl alcohol, uniformly stirring, placing the Eurya emarginata dry powder into an ultrasonic cleaning instrument, carrying out ultrasonic oscillation for 20-40 minutes to obtain an extracting solution, and standing the extracting solution for 2-4 days for layering to obtain a supernatant I;
s3, adding the supernatant I obtained in the step S2 into a round-bottom flask, placing the round-bottom flask into a rotary evaporator, heating, and concentrating for 3-5 hours to obtain a concentrated solution;
and S4, centrifuging the concentrated solution obtained in the step S3 by using a centrifugal machine for 5-15 minutes to obtain a supernatant II, and freeze-drying the supernatant II to obtain the Eurya emarginata essential oil.
2. The method of claim 1, wherein the essential oil of Eurya emarginata comprises: in step S1, the particle size of the Eurya emarginata dry powder is 150 μm.
3. The method of claim 1, wherein the essential oil of Eurya emarginata comprises: in step S2, the ratio of the Eurya emarginata dry powder to the absolute ethyl alcohol is (0.5-1.5) g:2 mL.
4. The method of claim 1, wherein the essential oil of Eurya emarginata comprises: in step S2, the power of the ultrasonic cleaning apparatus is 70%.
5. The method of claim 1, wherein the essential oil of Eurya emarginata comprises: in the step S3, the heating temperature is 70-80 ℃.
6. The method of claim 1, wherein the essential oil of Eurya emarginata comprises: in step S4, the speed of centrifugal separation was 5000 rpm.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202111185848.2A CN113698992A (en) | 2021-10-12 | 2021-10-12 | Preparation method of Eurya emarginata essential oil |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202111185848.2A CN113698992A (en) | 2021-10-12 | 2021-10-12 | Preparation method of Eurya emarginata essential oil |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN113698992A true CN113698992A (en) | 2021-11-26 |
Family
ID=78662771
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202111185848.2A Pending CN113698992A (en) | 2021-10-12 | 2021-10-12 | Preparation method of Eurya emarginata essential oil |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN113698992A (en) |
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106038655A (en) * | 2016-05-11 | 2016-10-26 | 南京泽朗医药科技有限公司 | A method of preparing turnip volatile oil and an antibacterial application of the volatile oil |
| CN111588741A (en) * | 2020-05-13 | 2020-08-28 | 扬州大学 | Preparation method and application of folium artemisiae argyi extract |
| CN112500930A (en) * | 2020-11-26 | 2021-03-16 | 浙江海洋大学 | Preparation method and application of compound essential oil of vitex rotundifolia |
-
2021
- 2021-10-12 CN CN202111185848.2A patent/CN113698992A/en active Pending
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106038655A (en) * | 2016-05-11 | 2016-10-26 | 南京泽朗医药科技有限公司 | A method of preparing turnip volatile oil and an antibacterial application of the volatile oil |
| CN111588741A (en) * | 2020-05-13 | 2020-08-28 | 扬州大学 | Preparation method and application of folium artemisiae argyi extract |
| CN112500930A (en) * | 2020-11-26 | 2021-03-16 | 浙江海洋大学 | Preparation method and application of compound essential oil of vitex rotundifolia |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| Ma et al. | Effects of light intensity and quality on phycobiliprotein accumulation in the cyanobacterium Nostoc sphaeroides Kützing | |
| CN102965306B (en) | Bacillus subtilis and application of same in resisting aspergillus | |
| Sizonenko et al. | The New Efficiency of the «Srmp»–Listerias Growth-Promoting Factor during Factory Cultivation” | |
| CN113549582A (en) | Licorice fermentation liquor with effects of resisting oxidation, relieving acute alcoholic liver injury and regulating intestinal flora and application thereof | |
| WO2017130106A1 (en) | Process for producing starch from microalgae | |
| CN112410261B (en) | Bacillus siamensis MC2-1 and application thereof | |
| CN113755341A (en) | Liquid fermentation culture method for morchella | |
| CN108531415B (en) | Bacillus air pallidus Jh-7 for antagonizing rice bacterial blight | |
| CN113698992A (en) | Preparation method of Eurya emarginata essential oil | |
| CN113005054A (en) | Bacillus amyloliquefaciens SS-ZC-26 and preparation method and application thereof | |
| CN104293678A (en) | Forsythia endophytic fungi capable of producing forsythoside A and B and forsythin, and application thereof | |
| CN111808787A (en) | Method for improving germination rate of spores of clostridium sporogenes | |
| Naidu et al. | Antimicrobial activity of Achyranthes aspera | |
| CN104962491A (en) | Degradation strain of herbicide 2, 4-D, produced inoculum and application thereof | |
| CN113637710B (en) | Preparation method and application of chlorella extract | |
| CN113755368B (en) | Fujian chicken mycoplasma synoviae and culture medium thereof | |
| CN104830716B (en) | Microbacterium and application thereof | |
| Alkhfaji | Bee collected pollen load (BCPL) as alternative culture media for bacterial and yeast growth | |
| RU2396335C1 (en) | Method of preparing solid culture medium | |
| CN105754908B (en) | A kind of pseudomonas aeruginosa strains and its application | |
| CN1132931C (en) | Ocean bacteria capable of producing haematochrome and method for producing haematochrome using said bacteria | |
| CN109463386A (en) | Application of the mangostin in prevention and treatment bacterial wilt | |
| CN105018359B (en) | The production method of the purposes of one Yeasts and natural p-Coumaric Acid | |
| Dega et al. | Growth of Salmonella typhimurium in skim milk concentrates | |
| CN108210531A (en) | Purposes of the Essential Oil of Radix Angelicae Dahuricae in antibacterials are prepared |
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: 20211126 |