CN104311348B - The Pleurotus eryngii liquid fermentation medium improved and the method utilizing its cultivation pleurotus eryngii liquid strain - Google Patents
The Pleurotus eryngii liquid fermentation medium improved and the method utilizing its cultivation pleurotus eryngii liquid strain Download PDFInfo
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- CN104311348B CN104311348B CN201410568173.3A CN201410568173A CN104311348B CN 104311348 B CN104311348 B CN 104311348B CN 201410568173 A CN201410568173 A CN 201410568173A CN 104311348 B CN104311348 B CN 104311348B
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- C—CHEMISTRY; METALLURGY
- C05—FERTILISERS; MANUFACTURE THEREOF
- C05G—MIXTURES OF FERTILISERS COVERED INDIVIDUALLY BY DIFFERENT SUBCLASSES OF CLASS C05; MIXTURES OF ONE OR MORE FERTILISERS WITH MATERIALS NOT HAVING A SPECIFIC FERTILISING ACTIVITY, e.g. PESTICIDES, SOIL-CONDITIONERS, WETTING AGENTS; FERTILISERS CHARACTERISED BY THEIR FORM
- C05G3/00—Mixtures of one or more fertilisers with additives not having a specially fertilising activity
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- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01G—HORTICULTURE; CULTIVATION OF VEGETABLES, FLOWERS, RICE, FRUIT, VINES, HOPS OR SEAWEED; FORESTRY; WATERING
- A01G18/00—Cultivation of mushrooms
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- C—CHEMISTRY; METALLURGY
- C05—FERTILISERS; MANUFACTURE THEREOF
- C05B—PHOSPHATIC FERTILISERS
- C05B17/00—Other phosphatic fertilisers, e.g. soft rock phosphates, bone meal
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Abstract
The present invention relates to a kind of microbiological culture media and cultural method, particularly relate to the Pleurotus eryngii liquid fermentation medium of improvement and utilize its method cultivating pleurotus eryngii liquid strain. Select the glucose of effective dose, analysis for soybean powder, Semen Maydis powder, potassium dihydrogen phosphate, magnesium sulfate, vitamin B1, gelatin, agar or its mixture, the fermentation medium of the liquid spawn of sterilized water preparation Pleurotus eryngii, prepare pleurotus eryngii liquid strain through shaken cultivation. The invention solves the production easy infection miscellaneous bacteria caused during existing Pleurotus eryngii industrial produces due to liquid spawn quality unstable (bacterium solution mycelium pellet, mycelium fragment are relatively big, bacteria suspension is uneven), growth efficiency is low, it is unsuitable for the problems such as batch production application, there is each Component Source of culture medium easy, pleurotus eryngii liquid strain mycelium pellet diameter prepared by this method is used substantially to diminish, bacterium solution density improves, and even suspension, after inoculation solid medium, growth vigor is better than traditional method liquid spawn, fast growth, the advantages such as mycelia is sturdy.
Description
Technical field
The present invention relates to a kind of microbiological culture media and cultural method, particularly relate to the Pleurotus eryngii liquid fermentation medium of improvement and utilize its method cultivating pleurotus eryngii liquid strain.
Background technology
Pleurotus eryngii is the nutrition and health care food deeply liked by people, in recent years due to the fast development of industrial cultivation technique, for shortening strain manufacturing cycle, improve strain quality and production efficiency, domestic and international manufacturer research and development liquid spawn is for substituting during current Pleurotus eryngii produces the solid spawn commonly used, and starts application in actual production.
Pleurotus eryngii liquid strain is to produce great-hearted Pleurotus eryngii mycelium by liquid fermentation process. at present, domestic quality and the quality not yet promulgating that unified national standard carrys out specification pleurotus eryngii liquid strain, but people sum up application effect and have worked out some endemicity standards in actual application, such as Liaoning Province's provincial standard-" DB21/T1693-2008 edible fungi liquid strain production technology regulation " etc., clear stipulaties in these standards, qualified edible fungi liquid strain should " visible distinctive hypha form, spherical and plexi mycelium is distributed in a large number, mycelia is sturdy, in mycelia, protoplasm is evenly distributed " " bacterium solution is thickness slightly, there is a large amount of lamellar or spherical mycelium suspended, it is evenly distributed ", that is excellent liquid spawn mycelium pellet density is high, diameter is little, it is evenly distributed,On the other hand, in production application, pleurotus eryngii liquid strain is to carry out mechanical inoculation operation by production line inoculation device, owing to inoculation device spout is narrow, for preventing bacterium solution blocking pipeline, also need to the pretreating process before increasing liquid-spawn inoculation (method being generally adopted mechanical activation comminution makes the mycelium pellet diameter in bacterium solution diminish, and makes bacterium solution become uniformly).
Therefore it is applicant's understanding that the physical characteristics such as the size of Pleurotus eryngii liquid fermentation mycelium pellet, density, the uniformity are the crucial Con trolling index of high-quality liquid spawn.
The domestic research report about Pleurotus eryngii liquid culture and patented technology concentrate on the optimization aspect of fermentation technology substantially, to improving the research of zymocyte pompon physical characteristic and patent mainly relies on and adds bead, little spring in the method for mechanical activation comminution or liquid medium within and reduce fermented hypha bulb diameter by breaing up the mode of mycelium pellet, increase density and the uniformity. The method complex process of mechanical activation comminution, very easily bacteria infection causes cultivating unsuccessfully; The method adding bead and little spring is only applicable to the small-scale production of shake flask fermentation, is applied to batch production fermentation tank and produces operable hardly.
Goal of the invention
It is an object of the invention to provide the pleurotus eryngii liquid strain fermentation medium of a kind of improvement and utilize its method cultivating pleurotus eryngii liquid strain. Mycelium pellet diameter can be effectively reduced, increase mycelium pellet density.
The overall technology design of the present invention is:
The pleurotus eryngii liquid strain fermentation medium of improvement, this fermentation medium adopts the component composition of following weight/mass percentage composition: glucose 1%-2%; Analysis for soybean powder 1%-2%; Semen Maydis powder 2%-3%; Potassium dihydrogen phosphate 0.05%-0.2%; Magnesium sulfate 0.01%-0.05%; Vitamin B10.02g/l-0.05g/l, a kind of in gelatin, agar or 0.1 gram/100ml-0.3 gram/100ml of mixture of the two, surplus is sterilized water, and pH is natural.
Utilizing the method that above-mentioned fermentation medium cultivates pleurotus eryngii liquid strain, be inoculated in the culture medium of claim 1 by pleurotus eryngii quel strains and carry out cultivation make pleurotus eryngii liquid strain, wherein condition of culture is temperature is 25 �� 3 DEG C, and rotating speed is 180-200r/m; 6-8d is cultivated in shaking table concussion; Described pleurotus eryngii quel strains selects the one in Pleurotus eryngii (Pleurotuseryngii) CICC14011, Pleurotus eryngii (Pleurotuseryngii) ACCC51678, Pleurotus eryngii (Pleurotuseryngii) ACCC51330.
In the present invention, the assay method of all technical is as follows:
1, Pleurotus eryngii mycelium pellet density quantification method: uniformly take 1ml culture fluid, physiological saline solution is diluted to 10ml, mixing, takes 1mL diluent and is placed in plate, launches. Black background paper, counting is served as a contrast under plate.
2, Pleurotus eryngii mycelium pellet dry weight method (Biomass): uniformly take 100ml culture fluid, filters through 80 eye mesh screens, washs with physiological saline solution, clarifies to cleaning mixture. Mycelium pellet is placed on the qualitative filter paper of constant weight, 60 DEG C dry to constant weight after weigh.
3, mycelium pellet measuring diameter: take mycelium pellet 20 at random, be arranged in a straight line, surveys its length with slide gauge, calculates the average diameter (cm) obtaining 20 mycelium pellets, repeats 3 times.
For determining thickening agent kind, applicant carried out following experiment:
One, the choosing and impact that Pleurotus eryngii mycelium pellet is grown by thickening agent of thickening agent
(1) impact that Pleurotus eryngii mycelium pellet is grown by xanthan gum, hydroxyethyl cellulose and methylcellulose
Experiments show that, add xanthan gum, hydroxyethyl cellulose or methylcellulose in the medium, it is suppressed that the growth of Pleurotus eryngii mycelium pellet.
Inspection information: xanthan gum is a kind of unit cell polysaccharide produced by Rhodopseudomonas; Research about edible fungi bacterial disease in recent years shows, major part pathogenic bacterium are Rhodopseudomonas, therefore the growth of edible fungi is also had inhibitory action by its metabolite. Hydroxy combining on methylcellulose, hydroxyethyl cellulose meeting and nutrient substance, affects mycelia absorbing fluid medium Middle nutrition material.
(2) impact that Pleurotus eryngii mycelium pellet is grown by starch and hydroxymethyl cellulose
Experiments show that (as shown in Figure 1-2), add starch or hydroxymethyl cellulose in the medium, contrast with blank group, mycelium pellet growth is not significantly affected.
(3) impact that Pleurotus eryngii mycelium pellet is grown by sodium alginate
Experiments show that (as shown in Figure 3), with the addition of after sodium alginate in liquid medium within, the diameter of Pleurotus eryngii mycelium pellet is significantly greater than blank group, does not meet and produces the requirement that Pleurotus eryngii mycelium pellet diameter is wanted little.
(4) impact that Pleurotus eryngii mycelium pellet is grown by agar and gelatin
Investigating the agar adding variable concentrations in the medium, the growing state of Pleurotus eryngii mycelium pellet, result is as shown in Figure 4. When in fluid medium, the content of agar is more than 0.4%, fluid medium solidifies, it is impossible to carry out shaking table cultivation. When the agar content in fluid medium is 0.3%, mycelium pellet diameter is minimum, for 2.41mm, compared with blank group, reduces about 18.9%, and now, the density of mycelium pellet also reaches maximum, is 3.14 �� 104Individual/L, compared with blank group, improves 57.9%. Now mycelium pellet dry weight is also higher, for 6.14g/100mL.
Investigate in liquid medium within the gelatin adding different content, the growing state of Pleurotus eryngii mycelium pellet, as shown in Figure 5. When gelatin concentration is when 0.2%, mycelium pellet diameter reaches minimum, for 2.54mm, compared with blank group, reduces about 14.5%, and now, mycelium pellet density, also close to peak value, is 3.03 �� 104Individual/L, contrasts with blank group, improves 52.3%. Mycelium pellet dry weight reaches maximum, 7.93g/100mL.
(5) contrast of 0.3% agar, 0.2% gelatin and blank group is added
Result according to above-mentioned (4th) article, when the thickening agent added is 0.3% agar or when being 0.2% gelatin, the Pleurotus eryngii mycelium pellet obtained meets the initial requirement of experiment most, two kinds of concentration acquired results and blank group are contrasted, as shown in Figure 6, mycelium pellet diameter, both less than blank group; Mycelium pellet density, both more than blank group; And both difference on diameter and density are little. And in mycelium pellet dry weight, 0.2% gelatin group is better than blank group, blank group is better than 0.3% agar group.
(6) 0.3% agar group and 0.2% gelatin group Pleurotus eryngii mycelium pellet are carried out vigor measurement
According to experiment the data obtained, make Fig. 7, Fig. 8. Added 0.3% agar or 0.2% gelatin gained mycelium pellet mycelia day growth rate linear equation on plate in the medium: y=0.5443x+1.3643, R2=0.9088 (adding 0.3% agar); Y=0.5411x+1.0257, R2=0.9541 (adding 0.2% gelatin). By contrasting, it has been found that 0.3% agar group mycelium pellet vigor is slightly better than 0.2% gelatin group mycelium pellet vigor. And both mycelium pellet vigor are better than matched group.
Two, the liquid spawn hypha form that the pleurotus eryngii liquid strain utilizing present patent application method to obtain and traditional method obtain has been contrasted by applicant, and result is as follows.
As it is shown in figure 9, wherein compared with matched group (left figure), utilize the liquid spawn mycelium pellet diameter that the present processes obtains substantially to diminish, bacterium solution density improves, and even suspension.
Three, the mycelium of applicant's pleurotus eryngii liquid strain to utilizing the present processes to obtain has carried out microscopic observation, and result is as follows.
Improved culture medium fermented hypha form is spherical or lamellar; Observing under 400 power microscopes, mycelia is sturdy, and hyphal cell protoplasm fills, uniformly (Figure 10).
Four, the mycelial growth vigor after applicant's pleurotus eryngii liquid strain to utilizing the present processes to obtain and conventional liq strain inoculation solid medium has contrasted, and result is as follows.
After using pleurotus eryngii liquid strain inoculation solid culture medium prepared by conventional medium, mycelial growth relatively slow (white portion is mycelial growth overlay area, less); After using the medium preparing liquid-spawn inoculation solid culture medium after improvement, mycelial growth grows fast (white portion is mycelial growth overlay area, more), illustrates that liquid spawn prepared by improved culture medium flushes, has using value.
Substantive distinguishing features and the notable technological progress of acquirement that the present invention possesses are in that:
The each Component Source of culture medium in the present invention is easy, and after adding gelatin or agar, mycelium pellet diameter substantially diminishes, and bacterium solution density improves, and even suspension. Using pleurotus eryngii liquid strain prepared by this method healthy and strong, after inoculation solid medium, growth vigor is better than traditional method liquid spawn, fast growth, and mycelia is sturdy, and hyphal cell protoplasm is full uniformly.
Accompanying drawing explanation
Fig. 1 is that different starch concentration affects schematic diagram to what Pleurotus eryngii mycelium pellet grew.
As it is shown in figure 1, add starch in the medium, contrast with blank group, mycelium pellet growth is not significantly affected.
Fig. 2 is that different hydroxymethyl cellulose concentration affects schematic diagram to what Pleurotus eryngii mycelium pellet grew.
As in figure 2 it is shown, add hydroxymethyl cellulose in the medium, contrast with blank group, mycelium pellet growth is not significantly affected.
Fig. 3 is that the mycelia growing of pleurotus eryngii is affected schematic diagram by different sodium alginate concentration.
As it is shown on figure 3, with the addition of in liquid medium within after sodium alginate, the diameter of Pleurotus eryngii mycelium pellet is significantly greater than blank group, does not meet and produces the requirement that Pleurotus eryngii mycelium pellet diameter is wanted little.
Fig. 4 is that the mycelia growing of pleurotus eryngii is affected schematic diagram by different agar concentration.
As shown in Figure 4, when in fluid medium, the content of agar is more than 0.4%, fluid medium solidifies, it is impossible to carry out shaking table cultivation. ) when the lightweight content in fluid medium is 0.3%, mycelium pellet diameter is minimum, for 2.41mm, compared with blank group, reduce about 18.9%, and now, the density of mycelium pellet also reaches maximum, is 3.14 �� 104Individual/l, compared with blank group, improves 57.9%. Now mycelium pellet dry weight is also higher, for 6.14g/100ml.
Fig. 5 is that the mycelia growing of pleurotus eryngii is affected schematic diagram by different gelatin concentration.
As it is shown in figure 5, when gelatin concentration is when 0.2%, mycelium pellet diameter reaches minimum, for 2.54mm, compared with blank group, reducing about 14.5%, now, mycelium pellet density, also close to peak value, is 3.03 �� 104Individual/l, contrasts with blank group, improves 52.3%. Mycelium pellet dry weight reaches maximum, 7.93g/100ml.
Fig. 6 is the Pleurotus eryngii mycelia and the blank group contrast schematic diagram that add 0.2% gelatin, 0.3% agar cultivation.
As shown in Figure 6, mycelium pellet diameter, both less than blank group;Mycelium pellet density, both more than blank group; And both difference on diameter and density are little. And in mycelium pellet dry weight, 0.2% gelatin group is better than blank group, blank group is better than 0.3% agar group.
Fig. 7 is 0.3% agar mycelium pellet day growth rate diagram.
Fig. 8 is 0.2% gelatin mycelium pellet day growth rate diagram.
According to experiment the data obtained, make Fig. 7, Fig. 8. Added 0.3% agar or 0.2% gelatin gained mycelium pellet mycelia day growth rate linear equation on plate in the medium: y=0.5443x+1.3643, R2=0.9088 (adding 0.3% agar); Y=0.5411x+1.0257, R2=0.9541 (adding 0.2% gelatin). By contrasting, it has been found that 0.3% agar group mycelium pellet vigor is slightly better than 0.2% gelatin group mycelium pellet vigor. And both mycelium pellet vigor are better than Yuanping City's plate switching strain.
Fig. 9 is liquid spawn mycelium morphology (density and the diameter) schematic diagram that the pleurotus eryngii liquid strain utilizing present patent application method to obtain obtains with traditional method.
As it is shown in figure 9, wherein compared with matched group (left figure), utilize the liquid spawn mycelium pellet diameter that the present processes obtains substantially to diminish, bacterium solution density improves, and even suspension.
Figure 10 is the pleurotus eryngii liquid strain mycelium microscopic morphology obtained and the mycelia microstructure schematic diagram of the method utilizing the present invention.
Wherein Figure 10 left side is the pleurotus eryngii liquid strain mycelium microscopic morphology schematic diagram obtained of the method utilizing the present invention; Figure 10 right side is the pleurotus eryngii liquid strain mycelia microstructure schematic diagram obtained utilizing microscope to observe the method adopting the present invention.
As shown in Figure 10 left side, improved culture medium fermented hypha volume morphing is spherical or lamellar; As shown in Figure 10 right side, observing under 400 power microscopes, mycelia is sturdy, and hyphal cell protoplasm fills, uniformly.
Figure 11 is that the pleurotus eryngii liquid strain adopting the inventive method to prepare inoculates the mycelial growth contrast situation schematic diagram after solid medium with conventional liq strain.
As Figure 11 is left, after using pleurotus eryngii liquid strain inoculation solid culture medium prepared by conventional medium, mycelial growth relatively slow (white portion is mycelial growth overlay area, less); As Figure 11 is right, after using the medium preparing liquid-spawn inoculation solid culture medium after improvement, mycelial growth grows fast (white portion is mycelial growth overlay area, more), illustrate that liquid spawn prepared by improved culture medium flushes, have actual application value.
Detailed description of the invention
Below in conjunction with embodiment, the present invention is described further; but it is not as a limitation of the invention; the content that protection scope of the present invention is recorded with claim is as the criterion, and any equivalent technical elements made according to description is replaced, all without departing from protection scope of the present invention
Embodiment 1
The pleurotus eryngii liquid strain fermentation medium of improvement, this fermentation medium adopts the component composition of following weight/mass percentage composition: glucose 1%; Analysis for soybean powder 1%; Semen Maydis powder 2%; Potassium dihydrogen phosphate 0.05%; Magnesium sulfate 0.01%; Vitamin B10.02g/l, 0.1 gram/100ml of gelatin, surplus is sterilized water, and pH is natural.
Utilizing the method that above-mentioned fermentation medium cultivates pleurotus eryngii liquid strain, condition of culture is temperature is 25 �� 3 DEG C, and rotating speed is 180-200r/m; 6-8d is cultivated in shaking table concussion.
Described pleurotus eryngii quel strains selects the one in Pleurotus eryngii (Pleurotuseryngii) CICC14011, Pleurotus eryngii (Pleurotuseryngii) ACCC51678, Pleurotus eryngii (Pleurotuseryngii) ACCC51330.
Embodiment 2
The pleurotus eryngii liquid strain fermentation medium of improvement, this fermentation medium adopts the component composition of following weight/mass percentage composition: glucose 2%; Analysis for soybean powder 2%; Semen Maydis powder 3%; Potassium dihydrogen phosphate 0.2%; Magnesium sulfate 0.05%; Vitamin B10.05g/l, 0.3 gram/100ml of agar, surplus is sterilized water, and pH is natural.
All the other contents are with embodiment 1.
Embodiment 3
The pleurotus eryngii liquid strain fermentation medium of improvement, this fermentation medium adopts the component composition of following weight/mass percentage composition: glucose 2%; Analysis for soybean powder 1%; Semen Maydis powder 2%; Potassium dihydrogen phosphate 0.2%; Magnesium sulfate 0.05%; Vitamin B10.02g/l, gelatin, agar 0.3 gram/100ml of mixture, surplus is sterilized water, and pH is natural.
All the other contents are with embodiment 1.
Embodiment 4
The pleurotus eryngii liquid strain fermentation medium of improvement, this fermentation medium adopts the component composition of following weight/mass percentage composition: glucose 1.5%; Analysis for soybean powder 1.5%; Semen Maydis powder 2.3%; Potassium dihydrogen phosphate 0.1%; Magnesium sulfate 0.04%; Vitamin B10.03g/l, 0.2 gram/100ml of gelatin, surplus is sterilized water, and pH is natural.
All the other contents are with embodiment 1.
Embodiment 5
The pleurotus eryngii liquid strain fermentation medium of improvement, this fermentation medium adopts the component composition of following weight/mass percentage composition: glucose 2%; Analysis for soybean powder 1.5%; Semen Maydis powder 2.5%; Potassium dihydrogen phosphate 0.1%; Magnesium sulfate 0.04%; Vitamin B10.04g/l, 0.25 gram/100ml of agar, surplus is sterilized water, and pH is natural.
All the other contents are with embodiment 1.
Claims (2)
1. the pleurotus eryngii liquid strain fermentation medium of improvement, it is characterised in that this fermentation medium adopts the component composition of following weight/mass percentage composition: glucose 1%-2%; Analysis for soybean powder 1%-2%; Semen Maydis powder 2%-3%; Potassium dihydrogen phosphate 0.05%-0.2%; Magnesium sulfate 0.01%-0.05%; Vitamin B10.02g/l-0.05g/l, a kind of in gelatin, agar or 0.1 gram/100ml-0.3 gram/100ml of mixture of the two, surplus is sterilized water, and pH is natural.
2. utilize the method that fermentation medium as claimed in claim 1 cultivates pleurotus eryngii liquid strain, it is characterized in that being inoculated in the culture medium of claim 1 by pleurotus eryngii quel strains and carry out cultivation making pleurotus eryngii liquid strain, wherein condition of culture is temperature is 25 �� 3 DEG C, and rotating speed is 180-200r/m; 6-8d is cultivated in shaking table concussion; Described pleurotus eryngii quel strains selects the one in Pleurotus eryngii (Pleurotuseryngii) CICC14011, Pleurotus eryngii (Pleurotuseryngii) ACCC51678, Pleurotus eryngii (Pleurotuseryngii) ACCC51330.
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| CN105009931A (en) * | 2015-04-13 | 2015-11-04 | 鲁东大学 | Preparation of liquid strain for pleurotus eryngii and research method of culture technique of high-quality high-yield pleurotus eryngii through liquid strain |
| CN104844354B (en) * | 2015-04-29 | 2017-11-14 | 天津农学院 | A kind of high density pleurotus eryngii liquid strain fermentation medium |
| CN106386173A (en) * | 2016-09-18 | 2017-02-15 | 胡云龙 | Pleurotus eryngii high-yield cultivating method |
| CN107736184A (en) * | 2017-11-07 | 2018-02-27 | 江苏久禾生物科技发展有限公司 | A kind of Pleurotus eryngii industrial cultivation method based on high activity liquid spawn |
| CN108147875A (en) * | 2018-03-07 | 2018-06-12 | 常熟理工学院 | A kind of factory culture pleurotus eryngii cultivating material |
| CN109526565A (en) * | 2018-12-13 | 2019-03-29 | 成都中延榕珍菌业有限公司 | A kind of Pleurotus eryngii liquid fermentation medium and strain cultivation method and method for planting almond abalone mushroom |
| CN113186106A (en) * | 2021-03-31 | 2021-07-30 | 盐城工学院 | Pleurotus eryngii liquid culture medium and application thereof |
| CN113973646A (en) * | 2021-07-28 | 2022-01-28 | 盐城工学院 | Culture medium and culture method for improving yield of pleurotus eryngii |
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