CN115418339B - Method for improving fermentation activity of monascus spores - Google Patents
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Abstract
The invention provides a method for improving fermentation activity of monascus spores, and belongs to the technical field of microorganisms. Specifically, inoculating monascus spore suspension into a monascus liquid culture medium with an inoculum size of 2-5%, carrying out shaking culture for 30+/-5 min at 43-47 ℃, transferring to 21-27 ℃ for continuous culture for 2-14 h, transferring the culture solution to 32+/-1 ℃ for continuous culture for 40-60 h, and obtaining the monascus spore culture solution with enhanced fermentation activity, wherein the activity of the obtained monascus spores is obviously improved. The method can obviously improve the quantity and quality of the fungus balls in the monascus spore culture solution, thereby starting the fermentation process of monascus in long-shaped rice more quickly, and obviously shortening the fermentation period while improving the color value/citrinin ratio of the prepared monascus, so that the production efficiency of the monascus is obviously improved, and the aim of quickly preparing high-quality monascus is fulfilled.
Description
Technical Field
The invention belongs to the technical field of microbial culture, and particularly relates to a method for improving fermentation activity of monascus spores.
Background
Monascus has been widely used in products of various industries in China from ancient times, and is currently the only microorganism allowed to be used for producing natural edible pigments, and plays an important role in microbial resources in China. Red rice is obtained by inoculating Monascus purpureus on long-shaped rice and performing solid state fermentation. In the past, the application and research of monascus red pigment are more and more, but with the progress of living standard, the diversified requirements of people on food are more and more. As can be seen from the types of foods sold in the market, the demand of yellow pigment is increasing, and foods such as bread, biscuits, cakes, beverages, edible oil, sauce, meat, instant noodles and the like are mainly yellow.
At present, monascus yellow pigment has high economic benefit and wide application range, and becomes a research hotspot at home and abroad. The monascus conidium is easy to process and can be stored for a long time stably, and the monascus conidium serving as seeds for preparing the monascus rice is beneficial to industrial production of monascus rice products. However, in the process of fermenting and producing red yeast rice, the fermentation period of the red yeast rice is longer due to lower fermentation activity of the red yeast spore culture solution, and the operation time and the operation cost of fermentation equipment are higher. In order to solve the technical problem, the invention provides the monascus spore seed production method which can obviously improve the quantity and quality of the fungus balls in the monascus spore culture solution, so that the fermentation process of monascus in long-shaped rice can be started more quickly, the prepared monascus color value/citrinin ratio is improved, and the fermentation period is obviously shortened, thereby achieving the aim of obviously improving the production efficiency of monascus rice. The method is simple, convenient and effective, can be used for rapidly preparing the high-quality red yeast rice, and has higher practical application value.
Disclosure of Invention
The invention provides a method for improving the fermentation activity of monascus spores, which aims to solve the technical problems of low fermentation activity of monascus spores, long fermentation period of the prepared monascus rice, non-ideal various fermentation indexes and higher operation time and operation cost of fermentation equipment.
The method for improving the fermentation activity of monascus spores comprises the following steps:
(1) Inoculating monascus to monascus solid culture medium, placing on a flat plate at 32+/-1 ℃ for culturing 6-8 d, flushing monascus spores from the flat plate after culturing, filtering by a cell filter, diluting and mixing uniformly to obtain the monascus solid culture medium with the concentration of 4.83-6.0x10 6 individual/mL monascus spore suspension;
(2) Inoculating monascus spore suspension with an inoculum size of 2-5% into a monascus liquid culture medium, performing shake culture for 30+/-5 min at the temperature of 43-47 ℃, and transferring to the temperature of 21-27 ℃ for continuous shake culture for 2-14 h;
(3) Transferring the culture solution into the culture solution, and continuously carrying out shaking culture for 40-60 hours at the temperature of 32+/-1 ℃ to obtain the monascus spore culture solution with enhanced fermentation activity.
Wherein the cell filter is a cell filter with a filter diameter of 40 μm.
Wherein the shaking culture is 120-180 r/min.
Wherein, the monascus spores are washed out from the flat plate, specifically, physiological saline containing 0.05-0.15% Tween 80 is put on the flat plate, spores on the flat plate are scraped by a coating rod, and then the monascus spores are washed out.
Wherein, the formula of the monascus solid culture medium comprises the following components in percentage by mass and volume: 50-65 g/L of glucose, 20-25 g/L of peptone, 25-35 g/L of soluble starch and 15-20 g/L of agar.
Wherein, the formula of the monascus liquid culture medium comprises the following components in percentage by mass and volume: 18-25 g/L of peptone and 60-65 g/L of soluble starch.
Advantageous effects
The current common method for culturing monascus spores is to add spores into a liquid culture medium for shaking culture of 36-48 h, so that the cultured fungus balls are larger and the specific surface area is smaller, and if the fungus balls in the form are directly added into rice for fermenting monascus rice, the contact area between mycelium and rice is smaller, so that the fermentation time of monascus rice is longer, and the running time and running cost of fermentation equipment are increased. Therefore, the invention provides that the monascus spores are firstly cultured for a short time at 45 ℃, then are transferred into a culture medium at 25 ℃ for 8h, the monascus spores cultured under the severe temperature difference can form a form of small-sized mycelia with more mycelia branching, the amylase secreted by monascus in a short mycelia form can have higher enzyme activity, sufficient nutrients can be provided for the growth of the mycelia in the initial stage of fermentation, and amylase secreted by mycelia can produce a large amount of glucose, and ethanol can be produced when the glucose is excessive, and the ethanol can stimulate pigment production, so that the pigment yield is improved, the fermentation period is shortened, and conditions are provided for rapidly preparing the citrinin-rich red yeast rice with high color value and low color value.
Industrial production is not only focused on the yield, but also considers the time cost of production. Through researches, in the process of treating monascus spores at high and low temperatures, the production efficiency of monascus beige pigment can be obviously improved only under the temperature condition provided by the invention. The red yeast rice prepared by culturing monascus spores by adopting the high-low temperature method provided by the invention can reach the highest pigment production efficiency 665.2U g at 11 th 11 d -1 And/d, the ratio of Huang Sejia/citrinin can reach 56.3. Whereas the control (32 ℃ C. Constant temperature method) reaches the highest pigment production efficiency 449.2U.g only at 13 th 13 d -1 /d, and at this point the Huang Sejia/citrinin ratio was 39.6; in contrast, the pigment production efficiency at 11. 11 d was only 413.5U g -1 And/d, at this time, the ratio of Huang Sejia/citrinin is only 39.0. Therefore, the red yeast rice prepared by the monascus spore seed production method provided by the invention can end fermentation at 11 th d, compared with a control (traditional constant temperature culture method), the fermentation time can be shortened by 2 d, and the prepared red Qu Mihuang color value/citrinin ratio is improved by 42.2% compared with the control. Therefore, the seed production method provided by the invention can achieve the purpose of rapidly preparing high-quality red yeast rice on the basis of improving the quantity and quality of the monascus balls. At present, the inventor has not inquired about the related published report of utilizing high and low temperature culture fungal spores to improve spore fermentation activity.
Description of the drawings:
FIG. 1 shows the effect of a short-time culture at 45℃on the morphology of the pellet by transferring Monascus spores to low-temperature culture (21℃at 23℃at 25℃at 27℃in this order from left to right);
FIG. 2 shows the effect of the transformation of Aspergillus candidus spores to mycelium morphology (21 ℃,23 ℃,25 ℃,27 ℃ in this order from left to right) after a brief incubation at 45 ℃;
FIG. 3 is a comparison of amylase activity of Monascus spore culture broth after a brief incubation at 45℃and a transition to different low temperature cultures;
FIG. 4 shows the effect of Monascus spores on fermentation of red yeast rice at 45℃after brief incubation and at different low temperatures;
FIG. 5 is the effect of high and low temperature culture of Monascus spores on red Qu Mihuang color scale;
FIG. 6 is the effect of high and low temperature culture of Monascus spores on the production efficiency of red Qu Mise hormone;
FIG. 7 shows the effect of high and low temperature culture of Monascus spores on the content of red Qu Mijie mycin.
Detailed Description
The present invention is further illustrated below with reference to specific examples, which are to be construed as merely illustrative of the invention and not limiting of its scope, as various equivalent modifications to the invention will fall within the scope of the claims appended hereto, after reading the invention.
In all examples the colour number of the finished product was measured according to the method of national standard GB 1886.19-2015.
1. The method for measuring amylase activity of monascus spore culture solution comprises the following steps: centrifuging the monascus spore culture solution at the temperature of 4 ℃ and the speed of 8000 r/min for 10 min, wherein the supernatant obtained by centrifugation is amylase crude enzyme solution; taking 5 50 tubes mL with plug scale centrifuge tubes, numbering, adding 1 mg/mL glucose control solution 0, 0.4, 0.6, 0.8 and 1.0 mL respectively, fixing the volume to 2 mL by distilled water, shaking uniformly, adding 2 mL DNS reagent into each tube, and placing in a boiling water bath for color development for 5 min. Immediately after removal, the solution was cooled with water and the volume was set to 20 mL. Measuring the absorbance of the rest tubes at 540 and nm wavelength by taking 0 tube without glucose as a blank, taking absorbance A as an ordinate and glucose mass m as an abscissa, drawing a standard curve for measuring glucose by a DNS method, and calculating the amylase activity in a sample according to a linear regression equation; 2 mL2% (W/V) soluble starch suspension and 2 mL pH5.0 0.2 mol/L phosphate buffer solution are added into a 50 mL centrifuge tube, the mixed solution is preheated for 10 min in advance in a water bath environment at 40 ℃,1 mL crude enzyme solution is added, the temperature is 40 ℃, 160 r/min is used for reacting 1 h, 2 ml of LDNS reagent is added for stopping the reaction, and the mixed solution is continuously shaken in the reaction process. The mixture was boiled in a boiling water bath for 5min, immediately after removal, cooled with water and distilled water was added to a volume of 20 mL. The control group was 1 mL inactivated crude enzyme solution to which 2 ml dns reagent was added, and the subsequent operations were the same. The absorbance of the solution was measured with a spectrophotometer at a wavelength of 540 nm. The resulting reducing sugar content was calculated with reference to a glucose standard curve, and the enzyme activity of the amylase produced by monascus was calculated. 1 mL crude enzyme solution is subjected to enzymolysis to generate 1 mg glucose which is defined as an enzyme activity unit U/mL in 1 h under the condition of pH5.0 at 40 ℃, and the enzyme activity calculation formula is as follows:
enzyme activity (U/mL) =m×n/t
m-the absorbance is carried into a standard curve to calculate the amount of reducing sugar, mg;
n is the dilution multiple of the enzyme solution;
t-reaction time.
2. The preparation method of the red yeast rice comprises the following steps: the pH of the water was adjusted to 3.5 with HCl, and after immersing 30 g long-grained nonglutinous rice 10 h in 100 mL water, it was filtered with 2 layers of gauze, drained to a constant weight of 43.5 g, and then sealed with 6 layers of gauze and two layers of kraft paper. Sterilizing at 121deg.C for 20 min. After sterilization, the rice grains are evenly separated in an ultra-clean workbench by stirring when the rice grains are hot. After the rice is cooled to room temperature, 2 mL monascus spore culture solution is inoculated, the monascus spores are cultured at 32 ℃, each group is subjected to three repeated tests, water is added once every 24 h and the monascus is turned over, water 2 mL is added from 1 st to 2 nd d, water 4 mL is added from 3 rd d, and the monascus rice is cultured until 11 th d is obtained, so that Huang Sejia and citrinin contents of the monascus rice are measured.
3. The method for measuring monascus pigment comprises the following steps: and (3) after the red yeast rice is fermented, drying the red yeast rice at 60 ℃ for 48 h, crushing the dried red yeast rice by a crushing grinder, sieving the crushed red yeast rice by a 60-mesh sieve, and collecting the red yeast rice for later use. The red rice flour 0.2 g which is evenly mixed after being crushed is weighed, dissolved by 100 mL of 70% ethanol solution and transferred into a 100 mL volumetric flask, and the volume is fixed to the scale mark. Placing in a constant-temperature water bath at 60 ℃ for 1 h, taking out, cooling to room temperature, continuously adding 70% ethanol solution to the scale mark, shaking uniformly, and filtering with filter paper. The absorbance A of the sample was measured at a wavelength of 410 nm using a 70% ethanol solution as a reference. The calculation formula of the yellow valence X (U/g) of the monascus pigment extract is as follows:
X=A×100/m×b
wherein: a-absorbance of the sample;
m-the mass of the sample in grams (g) is weighed;
b-dilution factor.
4. The method for measuring citrinin comprises the following steps: weighing red rice flour 2.0 g which is crushed and uniformly mixed into a 50 mL centrifuge tube, adding 8 ml of 70% methanol solution for dissolution, shaking uniformly by vortex shaking for 3 min, using ultrasonic shaking for 30 min for dissolution assistance, centrifuging for 10 min at 8000 r/min, collecting supernatant, continuously adding 8 ml of 70% methanol into the precipitate, repeating the above operation once, enriching the supernatant obtained by the two steps, and drying at 60 ℃ for 3 d. After the drying is finished, adding 20 mL of methanol solution with pH of 1.5 into a centrifuge tube for re-dissolution, uniformly stirring after the dissolution is finished, diluting 600 times and 700 times, measuring fluorescence values of the solvent under the conditions of excitation wavelength λex=330 nm, emission wavelength λem=485 nm and slit width Ex=10 nm and Em=5 nm respectively by using a fluorescence spectrophotometer, and bringing the fluorescence values into a citrinin standard curve equation y=1865.4x+2.0566 (R2=0.999) to obtain citrinin content.
Example 1
Based on the above measurement method, the examples compare the changes of monascus fermentation indexes after high and low temperature treatment under the same conditions, and the specific method is as follows:
1) The preparation method of the monascus spore suspension comprises the following steps: inoculating Monascus purpureus to Monascus solid culture medium by three-point method, culturing at 32deg.C for 7 d, collecting appropriate amount of physiological saline containing 0.1% Tween 80, scraping spores on the plate with coating rod, filtering the eluted liquid with 40 μm cell filter, and adjusting spore concentration to (5.0+ -0.17) ×10 6 Obtaining monascus spore suspension by each mL; wherein, the formula of the monascus solid culture medium is as follows: each liter of water contains glucose 60g, peptone 20g, soluble starch 30 g and agar 18g.
2) The preparation method of the monascus spore culture solution comprises the following steps: the concentration was set at (5.0.+ -. 0.17). Times.10 6 Inoculating the monascus spore suspension with the inoculum size of 3% into a monascus liquid culture medium for culturing to obtain the monascus spore culture liquid, wherein the monascus liquid culture medium comprises 20-g of peptone and 60-g of soluble starch in each liter of water.
Experiment 1
The concentration was set at (5.0.+ -. 0.17). Times.10 6 Inoculating Monascus spore suspension at 3% inoculum size into the Monascus liquid culture medium, shake culturing at 45deg.C for 30 min at 150 r/min, transferring to 21 deg.C, 23 deg.C, 25 deg.C, and 27 deg.C for 6 h, shake culturing at 32deg.C for 48 h, and observing the forms of fungus balls and mycelia.
Experiment 2
The concentration was set at (5.0.+ -. 0.17). Times.10 6 Inoculating the monascus spore suspension with the volume of 3% into a monascus liquid culture medium, performing shake culture at 45 ℃ for 30 min at 150 r/min, and then transferring the monascus spore suspension into 23 ℃ and 25 ℃ for continuous shake culture for 2-14 h. Samples were taken every 2 hours to determine amylase activity of monascus spore culture broth.
Experiment 3
The concentration was set at (5.0.+ -. 0.17). Times.10 6 Inoculating Monascus spore suspension with 3% inoculum size into Monascus liquid culture medium, shake culturing at 45deg.C for 30 min at 150 r/min, shake culturing at 23deg.C for 12 h and 25 deg.C for 8 hr, and shake culturing at 32deg.C for 48 h. Red rice was prepared according to the above method, and the red rice cooled to room temperature was inoculated with the different treated red koji spore culture solutions, respectively, and cultured to 11 th d to determine the Huang Sejia and citrinin content of red rice.
Experiment 4
The concentration was set at (5.0.+ -. 0.17). Times.10 6 Inoculating Monascus spore suspension at 3% inoculum size into Monascus liquid culture medium, shake culturing at 45deg.C for 30 min at 150 r/min, shake culturing at 25deg.C for 8h, and shake culturing at 32deg.C until 48 h. Red rice was prepared according to the above method, and red koji spore culture solution was inoculated into the long-grained nonglutinous rice cooled to room temperature. Samples were taken daily from 9 th d, and Huang Sejia and citrinin contents of red yeast rice 9-16 th d were measured to calculate the production efficiency of pigment (production efficiency=pigment yield/fermentation days).
Comparative experiments
The monascus spores are cultured by adopting a traditional constant temperature method, and the specific method is as follows:
the concentration was set at (5.0.+ -. 0.17). Times.10 6 The Monascus spore suspension was inoculated into Monascus liquid medium at an inoculum size of 3%, and cultured with shaking at 32deg.C for 48 h. Red rice is prepared according to the method, and the red aspergillus spore culture solution is inoculated into the long-shaped rice cooled to room temperature and cultured by a traditional constant temperature method. Samples were taken daily from 9 th d, and Huang Sejia and citrinin contents of red yeast rice 9-16 th d were measured to calculate the production efficiency of pigment (production efficiency=pigment yield/fermentation days).
From the results of the fungus ball and hypha morphology in experiment 1, it can be seen that the effect of the transition from the short-time culture at 45 ℃ to the low-temperature culture on the fungus ball and hypha morphology in the monascus spore culture solution is remarkable. As shown in FIG. 1, the size of the fungus balls in the monascus spore culture solution of the 23 ℃ and 25 ℃ experimental groups is smaller than that of the monascus spore culture solution of the 21 ℃ and 27 ℃ experimental groups, and the fungus balls are not linearly related to the low temperature, and the formation of the fungus balls is not favored by the too large or too small temperature difference of the high and low temperature combination. In FIG. 1, 21 ℃,23 ℃,25 ℃,27 ℃ are sequentially carried out from left to right. The edge of the fungus ball is subjected to microscopic examination, as shown in fig. 2, the edge of the fungus ball of the test group at 25 ℃ and 23 ℃ has more hyphae and more branches, the edge of the fungus ball of the test group at 21 ℃ has longer hyphae but fewer branches, the edge of the fungus ball of the test group at 27 ℃ has shorter hyphae and fewer branches, and the hyphae are easy to wind into big balls when fewer branches of the hyphae are wound, so that the high-low temperature combination of monascus spore culture is preliminarily determined to be 45 ℃ +23 ℃ and 45 ℃ +25 ℃.
From the experimental results of the amylase activity of the monascus spore culture solution in experiment 2, the influence of the short-time culture at 45 ℃ on the amylase activity of the monascus spore culture solution by the low-temperature culture is obvious. As shown in FIG. 3, the amylase activity of the monascus spore culture fluid of the 25 ℃ experimental group is higher than that of the 23 ℃ experimental group, the enzyme activity of the monascus spore culture fluid at 25 ℃ can reach a peak value of 9.9+/-0.3U/mL at 8h, and the enzyme activity of the monascus spore culture fluid at 23 ℃ reaches a peak value of 7.3+/-0.5U/mL at 12 h, so that the low-temperature treatment condition of the monascus spores is determined to be 8h at 25 ℃ and 12 h at 23 ℃ by taking the amylase activity of the monascus culture fluid as an index.
From Huang Sejia and citrinin contents of red yeast rice in experiment 3 and comparison experiments, it can be seen that the effect of transferring the red yeast rice spores to low-temperature culture after short-time culture at 45 ℃ on Huang Sejia and citrinin of the prepared red yeast rice is remarkable. As shown in FIG. 4, the experimental group with the color value of 45 ℃ plus 25 ℃ is the highest, and the experimental group with the color value of 45 ℃ plus 23 ℃ is the next, and the combination of the two groups at high and low temperatures can obviously improve the ratio of red Qu Mihuang color value to citrinin, but the improvement range of the experimental group with the color value of 45 ℃ plus 23 ℃ is obviously lower than that of the experimental group with the color value of 45 ℃ plus 25 ℃. In combination with the previous experimental results, amylase activity in monascus spore broth cultured at 23 ℃ was significantly lower than that cultured at 25 ℃. It can be seen that too low a temperature may inhibit the growth of spores, thereby affecting monascus production. Therefore, the conditions of short-time culture at 45 ℃ and low-temperature culture are finally determined to be 8h at 25 ℃ by taking the monascus mycelium form, the spore culture solution amylase activity and the yellow valence/citrinin of red rice as indexes into comprehensive consideration.
From Huang Sejia and citrinin contents of red yeast rice in experiment 4 and a comparison experiment, it can be seen that the quality and the production efficiency of red yeast rice are obviously affected by the monascus spores which are cultured for 8 hours at the temperature of 25 ℃ after being cultured for a short time at the temperature of 45 ℃. As shown in FIG. 5, the yield of the yellow pigment of the experimental group which is cultured for a short time at 45 ℃ and then cultured for 8h at 25 ℃ is obviously higher than that of the control group, and the highest yield of the yellow pigment can reach 8157.5 +/-74.3U/g, and is improved by 31.34% compared with that of the control group, so that the monascus spores prepared by the method can obviously improve the yield of the yellow pigment of the monascus rice.
The rapid development of fermentation enterprises is not only based on the excellent quality of the products, but also focused on improving the production efficiency, so as to reduce the cost as much as possible to improve the profit value of the enterprises, and therefore, for the production of red yeast rice, improving the pigment production efficiency is an important goal, namely, improving the effective output of pigment in unit time as much as possible. As shown in FIG. 6, the production efficiency of yellow pigment is significantly higher in the experimental group from 45℃short-time culture to 25℃culture 8h than in the control group, and the production efficiency can reach the peak at 11 th d (665.2U. G) -1 D), while the control group reaches the highest production efficiency 449.2U g at 13 th 13 d -1 And/d, the highest production efficiency of the yellow pigment of the high-low temperature experiment group is improved by 48.06 percent compared with that of the control group, and the spores after high-low temperature culture can be seen
The production efficiency of the monascus yellow pigment is improved.
As shown in FIG. 7, in the fermentation process, the yield of the citrinin is higher in the experimental group from 9 to 13 d red Qu Mijie to the low temperature culture 8h after the short-time culture at 45 ℃ than in the control group, and is lower in the experimental group from 14 to d, the metabolism rate of the monascus is possibly reduced, and thus the generation amount of the citrinin is reduced. As can be seen from FIG. 6, in 11 th d of red yeast rice fermentation, the pigment production efficiency of the high and low temperature experiment group is 160.85% of that of the control group, and at this time (11 th d), although the citrinin content is higher than that of the control group, the color value is obviously higher than that of the control group (FIG. 5), so that the Huang Sejia/citrinin ratio can reach 56.3, and the pigment production efficiency is improved by 44.3% compared with that of the control group. Therefore, the monascus spores prepared by the method can obviously shorten the fermentation period while improving the yellow value/citrinin ratio and the highest production efficiency, and the method is more suitable for industrially producing high-quality high-color-value low-citrinin monascus rice.
Example 2
A method for improving the fermentation activity of monascus spores comprises the following steps:
(1) inoculating Monascus purpureus to solid culture medium by three-point method, culturing at 32+ -1deg.C for 6d, collecting physiological saline containing 0.05% Tween 80, scraping spores on the plate with coating rod, filtering the eluted liquid with 40 μm cell filter, and adjusting spore concentration to 6.0X10 6 Obtaining monascus spore suspension after each mL.
(2) The concentration was set at 6.0X10 6 Inoculating 2% of the Monascus spore suspension to the Monascus liquid culture medium for culturing to obtain Monascus spore culture solution, shake culturing at 43deg.C and 120r/min for 30+ -5 min, and shake culturing at 25deg.C for 8 hr.
(3) After the culture is finished, transferring the culture solution into the culture solution at the temperature of 32+/-1 ℃ for continuous shaking culture for 40 hours, and obtaining the monascus spore culture solution with enhanced fermentation activity.
At 25 ℃, the enzyme activity of the monascus spore culture solution of the embodiment can reach a peak value of 6.7U/mL at the 8 th h, and after monascus rice is prepared by using the monascus spores with enhanced fermentation activity prepared by the embodiment, the yellow pigment content is increased by 18.61%.
Wherein, the formula of the monascus solid culture medium is as follows: each liter of water contains 50 g g g glucose, 25g peptone g g soluble starch 25g g g agar; the monascus liquid culture medium has the formula that each liter of water contains 25g g of peptone and 65g of soluble starch.
Example 3
A method for improving the fermentation activity of monascus spores comprises the following steps:
(1) inoculating Monascus purpureus to solid culture medium by three-point method, culturing at 32+ -1deg.C for 8d, collecting physiological saline containing 0.15% Tween 80, scraping spores on the plate with coating rod, filtering the eluted liquid with 40 μm cell filter, and adjusting spore concentration to 5.5X10 6 The red yeast rice is obtained by one/mLMould spore suspension.
(2) The concentration was 5.5X10 6 Inoculating the monascus spore suspension with the inoculum size of 5% into the monascus liquid culture medium for culture to obtain the monascus spore culture liquid, carrying out shake culture for 30+/-5 min at the temperature of 47 ℃ and 180r/min, and then transferring the monascus spore suspension into the monascus liquid culture medium for continuous shake culture at the temperature of 25 ℃ and 180r/min for 8h.
(3) After the culture is finished, transferring the culture solution into a temperature of 32+/-1 ℃ for continuous shaking culture for 60 hours to obtain the fermentation activity
Monascus spore culture solution with enhanced properties.
At 25 ℃, the enzyme activity of the monascus spore culture solution of the embodiment can reach 4.5U/mL peak value at the 8 th h, and after monascus rice is prepared by using the monascus spores with enhanced fermentation activity prepared by the embodiment, the yellow pigment content is increased by 12.10%.
Wherein, the formula of the monascus solid culture medium is as follows: each liter of water contains 65 to g g of glucose, 22 to g g of peptone, 35 to g g of soluble starch and 20g of agar; the monascus liquid culture medium has the formula that each liter of water contains 18g g of peptone and 60g of soluble starch.
Claims (5)
1. A method for improving the fermentation activity of monascus spores is characterized by comprising the following steps: the method for improving the fermentation activity of monascus spores comprises the following steps:
(1) Inoculating Monascus purpureus to Monascus purpureus solid culture medium, culturing at 32+ -1deg.C on plate for 6-8 d, washing Monascus purpureus spores from plate, filtering with cell filter, diluting, and mixing uniformly to obtain concentration of (4.83-6.0) ×10 6 individual/mL monascus spore suspension;
(2) Inoculating monascus spore suspension with an inoculum size of 2-5% into a monascus liquid culture medium, performing shake culture for 30+/-5 min at the temperature of 43-47 ℃, transferring to the temperature of 23-25 ℃ and continuing shake culture for 8-12 h to obtain a culture solution;
(3) Transferring the culture solution into a culture solution with the fermentation activity enhanced, and continuously carrying out shaking culture for 40-60 hours at the temperature of 32+/-1 ℃ to obtain an monascus spore culture solution with the fermentation activity enhanced;
wherein, the shaking culture in the step (2) and the step (3) is 120-180 r/min.
2. The method for improving the fermentation activity of monascus spores according to claim 1, wherein: the cell filter is a cell filter with a filter diameter of 40 μm.
3. The method for improving the fermentation activity of monascus spores according to claim 1, wherein: washing monascus spores from a flat plate, specifically, putting physiological saline containing 0.05-0.15% Tween 80 on the flat plate, scraping spores on the flat plate by using a coating rod, and washing monascus spores.
4. The method for improving the fermentation activity of monascus spores according to claim 1, wherein: the monascus solid culture medium comprises the following components in percentage by mass and volume: 50-65 g/L of glucose, 20-25 g/L of peptone, 25-35 g/L of soluble starch and 15-20 g/L of agar.
5. The method for improving the fermentation activity of monascus spores according to claim 1, wherein: the monascus liquid culture medium comprises the following components in percentage by mass and volume: 18-25 g/L of peptone and 60-65 g/L of soluble starch.
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