CN100999720A - Plant medium composition containing bee glue bacteriostatic agent and preparation process thereof - Google Patents
Plant medium composition containing bee glue bacteriostatic agent and preparation process thereof Download PDFInfo
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- CN100999720A CN100999720A CN 200610172193 CN200610172193A CN100999720A CN 100999720 A CN100999720 A CN 100999720A CN 200610172193 CN200610172193 CN 200610172193 CN 200610172193 A CN200610172193 A CN 200610172193A CN 100999720 A CN100999720 A CN 100999720A
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- propolis
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Abstract
The present invention relates to one kind of plant culture medium composition containing propolis bacteriostat and its preparation process. The culture medium consists of propolis clathrate in 1-3 g in per 100 ml, and the propolis clathrate consists of propolis ethanol glue and beta-cyclodextrin in the weight ratio of 1 to 2.7. The culture medium can reduce microbial contamination for plant to grow normally and maintain the effective components in propolis, and its beta-cyclodextrin can increase the water solubility of the effective antibiotic components in propolis. The present invention is simple, feasible and significant in plant tissue culture.
Description
Technical field
The invention belongs to field of plant tissue culture technique, be specifically related to a kind of propolis-containing fungistat plant medium composition and preparation method.
Background technology
Patent about plant culture is a lot, for example: the method for Chinese patent CN1422948 plant cell culture medium and use culture medium culturing vegetable cell, in substratum, add a certain amount of superoxide, reach the promotion growth and development of plant.Chinese patent CN1193344 prevents the composition and the method for microbial contamination of plant tissue culture media, in plant tissue culture media, add methylchloroisothiazandnone, methylisothiazolinone, magnesium chloride and magnesium nitrate, these materials can effectively reduce or prevent the microbial contamination of this plant tissue culture media, guarantee plant tissue normal growth or growth.
Patent of invention about the water-soluble bee glue preparation is a lot, for example: the preparation method of Chinese patent CN94118328.9 water-soluble bee glue, adopt glycerine to soak, obtain water-soluble bee glue.Chinese patent CN03141394.3 water-soluble bee glue and preparation method thereof, it is made up of propolis, polyoxyethylene glycol and distilled water.Chinese patent CN200380104801.2 comprises the four-tuple compound of propolis as active substance, adopt the propolis active substance, hydrophilic support and a kind of be amino acid, another kind is the four-tuple compound of fine-powder form of the common grinding of glycyrrhetate sweeting agent with preparation.Chinese patent CN03150648.8 water-soluble bee glue and extracting method thereof utilize the isolating temperature of solubilising, ultrasonic technique and control, heat-up time, and the activeconstituents in the propolis is extracted.The preparation technology and the application of Chinese patent CN200510049326.4 propolis solution with after ethanol-water dissolution, distill out ethanol, make water-soluble bee glue.
Summary of the invention
The present invention is in order to solve the relatively poor shortcoming of existing plant culture antimicrbial power, to utilize the harmless antimicrobial characteristic of propolis Nantural non-toxic, increasing the water-soluble of propolis.
The principle of invention: plant tissue culture is meant that under aseptic condition the technology of utilizing artificial medium that organ, tissue, cell and the protoplastis of plant are cultivated is one of core technology of modern biotechnology.Plant tissue culture technique started from for 20 beginnings of the century, and over more than 90 year, through many investigators' hard exploration, this technology is gradually improved, and fully demonstrates its superiority in scientific research and modernized agriculture production.It has can breed improved seeds fast, improve quality, save the labour, make full use of the space, characteristics such as natural condition restriction are produced, are not subjected in annual test.The tissue cultured seedling volume of its cultivation is little, and being easy to carry exchanges with resource.At present in the world, plant tissue culture technique has been jumped by the laboratory study stage one in many countries and regions and has become extensive industrial production method in batch.This technology is considered to a very potential new and high technology promoting to have obtained huge economy, society and ecological benefits aspect the agricultural modernization." the quick breeding of good clone and the commercialization of test tube kind are one of present plant tissue culture and cell cultures main flows on using ".One of urgent problem was that tissue cultured seedling is to cultivate in totally enclosed gnotobasis during batch production was produced, and schedule of operation is numerous and diverse, and incubation time is longer, and each link all might increase the opportunities for contamination of assorted bacterium, expended great amount of manpower and material resources, financial resources.
Natural propolis is a kind of ten minutes valuable animal natural resources, propolis and activeconstituents thereof are water-soluble hardly, major part is dissolved in ethanol, also be dissolved in turps, very easily be dissolved in ether, chloroform and acetone, benzene, but, these organic solvents are deleterious to plant-growth, can not be with an organic solvent in plant tissue culture, therefore, use ethanol, use the cycloheptaamylose inclusion again as the organic solvent extraction effective bee glue components, dry the place to go alcohol solvent, can obtain the propolis that is dissolved in boiling water fully of organic solvent-free.Propolis has the effect of resisting pathogenic microbes, honeybee can live in dark, warm wet beehive, avoid the infection of the various germs of nature, major cause is exactly because the polyphenolic compound content height in the propolis, tangible anti-microbial activity is arranged, to numerous bacteriums, fungi, viral simultaneously inhibited, be a kind of natural broad-spectrum antimicrobial material.
Cycloheptaamylose can increase the water solubility of medicine, mechanism is to utilize special structure of cycloheptaamylose and character, with water-fast effective bee glue components bag and, thereby effective bee glue components is dissolved with the dissolving of cycloheptaamylose, increase the water solubility of effective bee glue components.
One of the object of the invention provides a kind of propolis-containing fungistat plant medium composition.Its moiety and proportioning are: propolis inclusion compound weight (gram): plant culturing minimum medium volume (milliliter) is 1-3: 100; Described propolis inclusion compound is made up of propolis ethanol jelly and cycloheptaamylose, and the weight ratio of propolis ethanol jelly and cycloheptaamylose is 1: 2.7.
Described plant culturing minimum medium (MS substratum), be that Murashige and Skoog cultivated design in 1962 for tobacco cell, be characterized in that inorganic salt and ionic concn are higher, be more stable ionic equilibrium solution, its nitrate content height, the quantity and the ratio of its nutrient are suitable, can satisfy the nutrition and the physiological requirements of vegetable cell, thereby the scope of application is wider, and most plant tissue culture are bred with it fast as minimum medium.
Two of the object of the invention provides a kind of propolis-containing fungistat plant culture preparation method, and its step and condition are as follows:
(1) propolis at 80 ℃ of heating in water bath paraffin removals, be the propolis raw material of 95% dissolve with ethanol paraffin removal with the concentration (v/v) of 3 times of the quality of the propolis of paraffin removal, filter, get supernatant liquor, discard residue, obtain propolis ethanol jelly.
(2) by proportioning propolis ethanol jelly and cycloheptaamylose thorough mixing, brute force rolls inclusion, drying, pulverizes, sieving obtains the propolis inclusion compound; With propolis inclusion compound and plant culturing minimum medium thorough mixing, can obtain a kind of propolis-containing fungistat plant medium composition by proportioning.
A kind of propolis-containing fungistat plant medium composition fungistatic effect time length is longer, reaches more than 45% in the surviving rate of two weeks endopolyploid tawny daylily good luck microbiological contamination seedling.It can effectively reduce or prevent the microbial contamination of substratum in the plant tissue culture course, and the plant tissue normal growth is grown.The composition of the propolis-containing fungistat plant culture that method of the present invention makes, it had both preserved the effective antibiotic composition of propolis, and it is water-soluble to the propolis antimicrobial component to increase cycloheptaamylose again.This propolis inclusion compound can be dissolved in boiling water fully.
Embodiment:
Embodiment 1
(1) paraffin removal propolis raw material (hair glue) 100g, heating in water bath is removed beeswax, with 200ml concentration is 95% ethanol (v/v) dissolving, filter, get supernatant liquor, use 95% ethanol (v/v) of 1 times of propolis throw out volume to extract supernatant liquor again, merge supernatant liquor, discard residue, 100ml alcohol solvent, the 200g propolis ethanol jelly that obtains purifying are reclaimed in evaporation.
(2) with the propolis ethanol jelly and the cycloheptaamylose that obtain, be 1: 2.7 thorough mixing by weight, be that 200g propolis ethanol jelly and 540 gram cycloheptaamyloses brute forces stir or roll inclusion, hot-air seasoning, pulverize, sieve (the order number is the 70-100 order) can obtain propolis inclusion compound dry powder 60 grams.This propolis inclusion compound can be dissolved in boiling water fully.
Embodiment 2
(1) preparation of propolis ethanol jelly is with embodiment 1;
(2) the propolis inclusion compound of 2 grams and 100 milliliters plant culturing minimum medium thorough mixing, all the other conditions can obtain a kind of propolis-containing fungistat plant medium composition with embodiment 1.This propolis inclusion compound can be dissolved in boiling water fully.
Embodiment 3
(1) preparation of propolis ethanol jelly is with embodiment 1;
(2) the propolis inclusion compound of 3 grams and 100 milliliters plant culturing minimum medium thorough mixing, all the other conditions can obtain a kind of propolis-containing fungistat plant medium composition with embodiment 1.This propolis inclusion compound can be dissolved in boiling water fully.
Embodiment 4 different sorts fungistats and the antibacterial Comparative Examples of propolis-containing fungistat plant medium composition
Tawny daylily (Hemerocallis fulva L.) is the Liliaceae day lily.Modern tawny daylily is the cross-fertilize seed of polyploid, and it merges the proterties of 15 initial specieses, makes pattern flower shape very abundant, and color has almost comprised the look of seven coloured silks of sunlight except that pure white, pure blue.And advantages such as modern tawny daylily tool is cold-resistant, adaptability is strong, leafage attitude grace, florescence length are the desirable flowers of flower garden, flower bed.But because of its heterozygosity highly, the planting seed breeding is difficult to keep good character.Use division propagation, indifferent because of sprout tillers again, breeding coefficient is lower, also is difficult for promoting.If breed fast with tissue culture method, then can reach and not only keep the purpose of good character but also large quantities of breeding, be the popularization blaze the trail of polyploid hybrid tawny daylily.In polyploid good luck tissue rapid propagation system, the media surface of some test-tube plantlet has the growth of cloud contaminated bacteria, and the contaminated bacteria kind has 2 kinds to be gram negative bacillus, and a kind is Gram-positive bacillus, and this contaminated bacteria has reduced the seedling rate of tissue culture.Inoculate after with the sterilization of 2% clorox on the substratum of bacteriostatic agent or else and still have the microbiological contamination phenomenon polluting seedling, it is better as the fungistat effect of plant culture to use Sodium Benzoate and potassium sorbate always.This experiment treatment process: adopt different concns chemistry fungistat and propolis inclusion compound configuration substratum, be used to plant with the pollution seedling of 2% clorox sterilization after 5 minutes, drug study is designed to water blank+MS and carries out bacteriostatic experiment for contrast, 1%-3% propolis inclusion compound+MS, 2 ‰ potassium sorbate+MS and 1 ‰ Sodium Benzoates+MS.In Bechtop, with the microbiological contamination seedling with aseptic water washing three times, 2% clorox sterilization 5 minutes, again with sterilized water from giving a baby a bath on the third day after its birth time, the aseptic filter paper suck dry moisture is seeded in material in the various substratum of experiment usefulness.Observations after 2 weeks, record, the surviving rate of calculating test-tube plantlet.
Table 1 different sorts fungistat and the antibacterial contrast of propolis-containing fungistat plant medium composition
| Kind and concentration (MS+ processing) | The quantity (1 seedling/test tube) of inoculation test tube | Survive quantity (1 seedling/test tube) during 2 weeks | Surviving rate (%) |
| The water blank | 30 | ?8 | ?27 |
| 1% propolis inclusion compound | 30 | ?14 | ?47 |
| 2% propolis inclusion compound | 30 | ?15 | ?50 |
| 3% propolis inclusion compound | 30 | ?13 | ?43 |
| 2 ‰ potassium sorbate | 30 | ?18 | ?60 |
| 1 ‰ Sodium Benzoates | 30 | ?15 | ?50 |
The result shows: 2 ‰ potassium sorbate+MS effect is best, and surviving rate reaches 60%; 1 ‰ Sodium Benzoates+MS takes second place, and surviving rate reaches 50%; 2% propolis-containing fungistat plant medium composition surviving rate reaches 50%; The surviving rate of 1% propolis-containing fungistat plant medium composition and 3% propolis-containing fungistat plant medium composition is respectively 47% and 43%; Water blank MS surviving rate only is 27%; Illustrate that propolis inclusion compound, potassium sorbate and Sodium Benzoate all have the effect of obvious inhibition substratum bacterium.Do not see in the experiment that growth has detrimentally affect to 1%-3% propolis-containing fungistat plant medium composition to tissue cultured seedling.
Claims (2)
1, a kind of composition of propolis-containing fungistat plant culture is characterized in that, its moiety and proportioning are: propolis inclusion compound weight (gram): plant culturing minimum medium volume (milliliter) is 1-3: 100; Described propolis inclusion compound is made up of propolis ethanol jelly and cycloheptaamylose, and the weight ratio of propolis ethanol jelly and cycloheptaamylose is 1: 2.7.
2, the preparation method of a kind of propolis-containing fungistat plant culture as claimed in claim 1 is characterized in that its step and condition are as follows:
(1) propolis at 80 ℃ of heating in water bath paraffin removals, be 95% dissolve with ethanol paraffin removal propolis with the concentration (v/v) of 3 times of the quality of paraffin removal propolis, filter, get supernatant liquor, discard residue, obtain propolis ethanol jelly;
(2) by proportioning propolis ethanol jelly and cycloheptaamylose thorough mixing, brute force rolls inclusion, drying, pulverizes, sieving obtains the propolis inclusion compound; With propolis inclusion compound and plant culturing minimum medium thorough mixing, can obtain a kind of propolis-containing fungistat plant medium composition by proportioning.
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| CNB2006101721934A CN100497599C (en) | 2006-12-30 | 2006-12-30 | Plant medium composition containing bee glue bacteriostatic agent and preparation process thereof |
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Cited By (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| MD191Z5 (en) * | 2010-01-21 | 2010-11-30 | Государственный Медицинский И Фармацевтический Университет "Nicolae Testemitanu" Республики Молдова | Nutrient medium for the cultivation of cells of animal origin |
| CN105130638A (en) * | 2015-08-27 | 2015-12-09 | 马鞍山市安康菌业有限公司 | Efficient oudemansiella radicata culture medium with propolis being antibacterial and nutrient substances and preparation method therefor |
| CN105237121A (en) * | 2015-08-27 | 2016-01-13 | 马鞍山市安康菌业有限公司 | High-effective dendrobium officinale culture medium employing natural bacteria inhibiting substances and preparation method thereof |
| CN108624513A (en) * | 2018-06-15 | 2018-10-09 | 成都本则生科技有限公司 | A kind of method of High Density Cultivation D-pantoyl lactone hydrolase producing strains and application |
| CN109198531A (en) * | 2018-09-14 | 2019-01-15 | 杭州和蜂园保健品有限公司 | A kind of propolis jelly and preparation method thereof |
| CN109548862A (en) * | 2018-12-29 | 2019-04-02 | 成都市农林科学院 | A kind of slow-release compound preservative and preparation method thereof fresh-keeping for hickory chick diseases prevention |
Family Cites Families (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN100500164C (en) * | 2005-03-10 | 2009-06-17 | 浙江省中医药研究院 | Process for preparing bee glue water solution and use |
| CN1879656B (en) * | 2006-04-27 | 2010-04-21 | 魏春华 | Solid water soluble propolis composition and preparation method thereof |
-
2006
- 2006-12-30 CN CNB2006101721934A patent/CN100497599C/en not_active Expired - Fee Related
Cited By (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| MD191Z5 (en) * | 2010-01-21 | 2010-11-30 | Государственный Медицинский И Фармацевтический Университет "Nicolae Testemitanu" Республики Молдова | Nutrient medium for the cultivation of cells of animal origin |
| CN105130638A (en) * | 2015-08-27 | 2015-12-09 | 马鞍山市安康菌业有限公司 | Efficient oudemansiella radicata culture medium with propolis being antibacterial and nutrient substances and preparation method therefor |
| CN105237121A (en) * | 2015-08-27 | 2016-01-13 | 马鞍山市安康菌业有限公司 | High-effective dendrobium officinale culture medium employing natural bacteria inhibiting substances and preparation method thereof |
| CN108624513A (en) * | 2018-06-15 | 2018-10-09 | 成都本则生科技有限公司 | A kind of method of High Density Cultivation D-pantoyl lactone hydrolase producing strains and application |
| CN108624513B (en) * | 2018-06-15 | 2020-11-03 | 成都本则生科技有限公司 | Method for high-density culture of D-pantolactone hydrolase producing strain and application |
| CN109198531A (en) * | 2018-09-14 | 2019-01-15 | 杭州和蜂园保健品有限公司 | A kind of propolis jelly and preparation method thereof |
| CN109548862A (en) * | 2018-12-29 | 2019-04-02 | 成都市农林科学院 | A kind of slow-release compound preservative and preparation method thereof fresh-keeping for hickory chick diseases prevention |
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| CN100497599C (en) | 2009-06-10 |
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