Method for producing monascus by utilizing highland barley
Through the purification of Aspergillus rosary and the puffing treatment of highland barley, the fermentation process of highland barley red rosary is optimized, and the problems of nutrient loss and high process complexity are solved, and high-quality and high-value-added barley red rosary production is achieved.
Patent Information
- Application Number
- CN202510844192.2
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2025-06-23
- Publication Date
- 2025-09-05
AI Technical Summary
There are problems of large loss of nutrients and high process complexity in the existing high barley red citrus production process, which leads to unstable product quality and affects market competitiveness.
The commercially available red chorus raw materials are used for purification and culture. Through the puffing treatment and optimization of fermentation process of barley, the puffing barley is used as raw material for culture of Aspergillus rosy, including activation, purification and expansion culture of Aspergillus rosy, and the fermentation conditions are controlled to improve the adaptability and growth effect of Aspergillus rosy.
The nutrients in barley are effectively retained, the production process is simplified, the quality and production efficiency of red citrus are improved, the risk of pollution is reduced, and high value-added barley red citrus products are realized.
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Figure CN120591352A_ABST
Abstract
Description
Technical Field
[0001] The present invention relates to the technical field of red yeast rice preparation, in particular to a method for producing red yeast rice using highland barley. Background Art
[0002] Highland barley, a cold-resistant naked-grain barley unique to the Qinghai-Tibet Plateau, is rich in functional ingredients such as β-glucan, polyphenols and selenium, and has unique nutritional advantages of low glycemic index and high antioxidant capacity. However, current product forms are mostly limited to traditional highland barley wine and tsampa, and there is an urgent need to develop functional ingredient stabilization technology and high-value utilization solutions for by-products.
[0003] The development of highland barley red yeast rice products is of great significance to the development of the big health industry. As the main production area of highland barley, Qinghai has unique resource advantages. Using highland barley to produce highland barley red yeast rice through technological innovation can not only increase the added value of highland barley, but also help to break through the problems of large loss of highland barley nutrients and high process complexity in the traditional process of highland barley red yeast rice, and provide more competitive product production processes for the functional food market.
[0004] Although the development of highland barley red yeast rice products has made certain progress in recent years, there are still the following deficiencies in their practical application:
[0005] First, there is a large loss of nutrients: According to existing technology, traditional highland barley processing usually requires deep peeling or crushing to improve the success rate of Monascus inoculation and fermentation efficiency; however, this pretreatment method inevitably leads to a large loss of functional ingredients rich in highland barley skin, thereby weakening the nutritional value and functionality of the final product.
[0006] Second, the process is highly complex and susceptible to contamination during production. Some patents propose combining liquid fermentation with solid-state fermentation to produce red highland barley yeast. This requires complex equipment and complicated procedures, and requires high-scale production and cost control. These complex process conditions can lead to unstable product quality, impacting market competitiveness.
[0007] In summary, the development and utilization of highland barley red yeast rice still faces the problems of insufficient strain adaptability, large loss of nutrients, and high process complexity; the existence of these problems not only restricts the technological progress of the highland barley red yeast rice industry, but also hinders the widespread application of highland barley red yeast rice in the big health industry; therefore, an innovative method is urgently needed to achieve high-quality and high-value-added development of highland barley red yeast rice products by optimizing strain selection, improving raw material pretreatment and fermentation technology. Summary of the Invention
[0008] In order to effectively solve the problem of nutrient loss caused by deep peeling or crushing in the current traditional process, the present invention provides a method for producing highland barley red yeast rice by purifying and culturing Monascus purpureus from commercially available red yeast rice raw materials and using puffed highland barley as follows:
[0009] A method for producing red yeast rice from highland barley comprises the following steps: cleaning red yeast rice raw materials, activating, purifying and propagating monascus; and culturing highland barley red yeast rice from highland barley raw materials, comprising: selecting and cleaning highland barley, then puffing the cleaned highland barley, sterilizing the puffed highland barley, and finally culturing highland barley red yeast rice from the propagated monascus.
[0010] The red yeast rice raw material is cleaned; the commercial red yeast rice raw material is used as the source of the strain, 10g of the red yeast rice raw material is weighed, sterilized with 75% alcohol, and the alcohol is filtered out after sterilization; then washed with sterile water 5 times and the water is dried;
[0011] The red yeast rice is activated; the cleaned red yeast rice raw material is added to the seed activation liquid and immersed; then placed in a constant temperature and humidity incubator, the temperature and humidity are set to: temperature 35 ° C, humidity 70%, and after culturing for 12 hours, when a velvety bacterial film is formed on the surface of the red yeast rice raw material, the red yeast rice activation is completed;
[0012] Purification of the Monascus purificans;
[0013] Take one activated red yeast rice raw material, crush it in a 1.5ml sterile centrifuge tube, add 1ml sterile water to dilute it, and use a sterile disposable pipette to draw a small amount and inoculate it on the solid culture medium. The solid culture medium is placed in a constant temperature and humidity incubator, and the temperature and humidity are set to: temperature 35°C, humidity 70%, and culture for 8 hours. When red yeast rice colonies grow on the solid culture medium, pick the pure color part and transfer it to a new culture medium. Repeat several times until the grown colonies have a regular shape and uniform color, and the purification of red yeast rice is completed.
[0014] The Monascus propagation culture: a small number of colonies of the Monascus purified in step 3 are picked up with tweezers in a sterile environment and inoculated into a liquid seed culture solution in a conical flask for propagation culture; when the color of the seed culture solution gradually changes from clear to orange-yellow, the Monascus propagation culture is completed;
[0015] The seed activation liquid is prepared from 75% alcohol and maltose aqueous solution, wherein the alcohol dosage is 3 ml and the maltose dosage is 2 g for every 100 ml of the seed activation liquid.
[0016] The solid culture medium is prepared by mixing 70 grams of glucose, 15 grams of beef extract, 2 grams of NaNO3, 0.5 grams of MgSO4·7H2O, 1.5 grams of KH2PO4, and 2 grams of agar powder into 1000 ml of sterile water and stirring evenly. The mixture is heated to 121° C., sterilized under high pressure for 30 minutes, and then dispensed into petri dishes and cooled and solidified to form a solid culture medium.
[0017] The seed culture medium is prepared by mixing 70 grams of glucose, 15 grams of beef extract, 2 grams of NaNO3, 0.5 grams of MgSO4·7H2O, and 1.5 grams of KH2PO4 into 1000 ml of sterile water and stirring evenly. The mixture is then divided into 250 ml conical flasks and sterilized under high pressure at 0.1 MPa and 121°C for 30 minutes to complete the preparation of the seed culture medium.
[0018] The specific culture conditions for the Monascus propagation culture are: placing the inoculated seed culture solution at a temperature of 35° C. and a humidity of 70% for culture; and culturing for 72 hours until a large number of Monascus fungi visible to the naked eye are generated in the seed culture solution.
[0019] The selection and cleaning of the highland barley are as follows: weighing 50g of highland barley and putting it into a conical flask, adding sterile water and quickly washing it 5 times, removing impurities on the surface of the highland barley, draining the water and placing it in a sterile dry place for 1-2h, until the surface of the highland barley in the conical flask is basically dry.
[0020] The puffing of highland barley is as follows: the dried highland barley is put into a puffing device, and the highland barley is heated to break the skin and puff.
[0021] The sterilization of the puffed barley is as follows: the puffed barley is placed in a high-pressure steam sterilization device, sterilized under high pressure for 30 minutes at 0.1 MPa and 121° C., and then allowed to stand until it reaches room temperature.
[0022] The highland barley red yeast rice is cultivated as follows: in a sterile environment, 0-4 ml of acetic acid with a pH of 4 and 2-4 ml of 75% alcohol are added to puffed highland barley, 10 ml of the bacterial solution cultivated in step 4 is measured with a graduated cylinder, and the mixture is evenly mixed; the bottle is sealed, and the mixture is placed in a constant temperature and humidity incubator for cultivation, with the temperature controlled at 25-40 DEG C and the humidity at 70%; the cultivation is carried out for 96-192 hours until the highland barley red yeast rice is cultivated; the highland barley red yeast rice is moved to a drying oven for drying, with the drying temperature not exceeding 50 DEG C; and the highland barley red yeast rice is dried until the humidity is below 25%, thereby obtaining the highland barley red yeast rice product.
[0023] The advantages and beneficial effects of the present invention are:
[0024] This method utilizes commercially available red yeast rice raw materials to independently isolate and purify the strain, which demonstrates good adaptability through cultivation using highland barley as the raw material. It also effectively addresses the nutrient loss problem associated with traditional processes, which require deep peeling or crushing. Physical puffing allows the barley skin to crack without peeling, effectively preserving the beneficial active ingredients in the barley. This also alters the barley's nutritional composition, effectively promoting the growth of the Monascus fungus. Furthermore, when using the puffed barley for cultivation of the red yeast rice, the mycelium grows abundantly both on the surface and within the barley, resulting in high-quality red yeast rice produced in a shorter production time. BRIEF DESCRIPTION OF THE DRAWINGS
[0025] Figure 1 is a flow chart of the present invention;
[0026] Figure 2 This is a schematic diagram of bacteria growing in a conical flask during the culture of highland barley red yeast rice in Comparative Example 1 of the present invention;
[0027] Figure 3 This is a schematic diagram of miscellaneous bacteria and Monascus growing in a conical flask during the culture of highland barley red yeast rice in Comparative Example 2 of the present invention;
[0028] Figure 4 This is a schematic diagram of miscellaneous bacteria and Monascus growing in a conical flask during the culture of highland barley red yeast rice in Comparative Example 3 of the present invention;
[0029] Figure 5 This is a schematic diagram of Monascus growing in a conical flask during the culture of highland barley red yeast rice in Comparative Example 4 of the present invention;
[0030] Figure 6 Schematic diagram of the growth of purified Monascus colonies on a solid culture medium;
[0031] Figure 7 Schematic diagram of the completion of the expansion culture of Monascus purpurogenum in seed culture of the present invention;
[0032] Figure 8 Schematic diagram of the growth of Monascus purpureus on unexpanded highland barley for 72 hours;
[0033] Figure 9 This is a schematic diagram of the growth of Monascus purpureus in the present invention after being cultured on puffed highland barley for 72 hours. DETAILED DESCRIPTION
[0034] The following will systematically describe the technical solutions in the embodiments of the present invention in conjunction with the comparative examples and examples of the present invention. These comparative examples and examples are intended only to illustrate the present invention and are not intended to limit the scope of the present invention. The experimental methods in the following comparative examples and examples where specific conditions are not specified are generally based on conventional conditions.
[0035] like Figure 1-9As shown, the present invention will illustrate the above steps one by one through 4 comparative examples and 4 embodiments.
[0036] Comparative Example 1
[0037] The commercially available red yeast rice raw material is used as the strain source, and an appropriate amount of the red yeast rice raw material is weighed. In step 1, highland barley is used as the raw material to produce highland barley red yeast rice.
[0038] The method for step 1 is:
[0039] a selection and cleaning of highland barley: Weigh 50g highland barley into a conical flask, add sterile water and quickly wash 5 times, remove impurities from the surface of highland barley, drain the water and place in a sterile dry place, until the surface of highland barley in the conical flask is basically dry;
[0040] b. Sterilization of highland barley: Place highland barley in a high-pressure steam sterilizer, sterilize under high pressure at 0.1MPa and 121°C for 30min, and let stand to room temperature;
[0041] c. Culture of highland barley red yeast rice: In a sterile environment, 10g of red yeast rice raw material was added to the highland barley culture medium, mixed evenly, sealed, and placed in an incubator for culture, controlling the temperature at 35 ℃ and the humidity at 70%; contamination occurred after 24h of culture; Figure 2 As shown, various colored bacteria appeared in the highland barley culture medium, and no clear Monascus colonies could be observed.
[0042] Comparative Example 2
[0043] According to Comparative Example 1, the red yeast rice raw material was soaked and disinfected in 75% alcohol before use.
[0044] Step 1: Cleaning the red yeast rice;
[0045] Use commercially available red yeast rice as the source of the strain, weigh an appropriate amount of red yeast rice, soak it in 75% alcohol for sterilization, filter out the alcohol after sterilization, then wash it with sterile water 5 times and drain it dry.
[0046] Step 2 utilizes highland barley as raw material to produce highland barley red yeast rice, and its method is:
[0047] a selection and cleaning of highland barley: Weigh 50g highland barley into a conical flask, add sterile water and quickly wash 5 times, remove impurities from the surface of highland barley, drain the water and place in a sterile dry place, until the surface of highland barley in the conical flask is basically dry;
[0048] b. Sterilization of highland barley: Place highland barley in a high-pressure steam sterilizer, sterilize under high pressure at 0.1MPa and 121°C for 30min, and let it stand to room temperature;
[0049] c. Culture of highland barley red yeast rice: In a sterile environment, 10g of cleaned red yeast rice raw material was added to the highland barley culture medium, mixed well, sealed, and placed in an incubator for culture, controlling the temperature at 35 ℃ and the humidity at 70%; contamination occurred after 72h of culture; Figure 3 As shown, more than two colors of bacteria appeared in the highland barley culture medium, and obvious Monascus colonies were observed.
[0050] Comparative Example 3
[0051] According to Comparative Examples 1 and 2, the red yeast rice raw material is immersed in 75% alcohol for disinfection and then activated and cultured before use.
[0052] Step 1: Cleaning the red yeast rice;
[0053] Use commercially available red yeast rice as the source of the strain, weigh an appropriate amount of red yeast rice, soak it in 75% alcohol for sterilization, filter out the alcohol after sterilization, then wash it with sterile water 5 times and drain it dry.
[0054] Step 2: activating Monascus;
[0055] The cleaned red yeast rice is immersed in the seed activation solution; then placed in an incubator and cultured for 12 hours to complete the red yeast rice activation;
[0056] Step 3 utilizes highland barley as raw material to produce highland barley red yeast rice, and its method is:
[0057] a selection and cleaning of highland barley: Weigh 50g highland barley into a conical flask, add sterile water and quickly wash 5 times, remove impurities from the surface of highland barley, drain the water and place in a sterile dry place, until the surface of highland barley in the conical flask is basically dry;
[0058] b. Sterilization of highland barley: Place highland barley in a high-pressure steam sterilizer, sterilize under high pressure at 0.1MPa and 121°C for 30min, and let it stand to room temperature;
[0059] c. Cultivation of highland barley red yeast rice: In a sterile environment, 10g of cleaned and activated red yeast rice raw materials were added to the highland barley culture medium, mixed well, sealed, and placed in an incubator for cultivation at a controlled temperature of 35 ℃ and a humidity of 70%; contamination occurred after 168h of culture; Figure 4 As shown, more than two colors of bacteria appeared in the highland barley culture medium, and more obvious Monascus colonies were observed.
[0060] Comparative Example 4
[0061] According to Comparative Examples 1, 2, and 3, the red yeast rice raw material is immersed in 75% alcohol for disinfection, then activated and cultured, and finally purified and expanded for use.
[0062] Step 1: Cleaning the red yeast rice;
[0063] Use commercially available red yeast rice as the source of the strain, weigh an appropriate amount of red yeast rice, soak it in 75% alcohol for sterilization, filter out the alcohol after sterilization, then wash it with sterile water 5 times and drain it dry.
[0064] Step 2: activating Monascus;
[0065] The cleaned red yeast rice is immersed in the seed activation solution; then placed in an incubator and cultured for 12 hours to complete the red yeast rice activation;
[0066] Step 3: purification of Monascus;
[0067] Take one activated red yeast rice raw material, crush it in a 1.5ml sterile centrifuge tube, add 1ml sterile water to dilute it, and use a sterile disposable pipette to draw a small amount and inoculate it on the solid culture medium. The solid culture medium is placed in a constant temperature and humidity incubator, and the temperature and humidity are set to: temperature 35°C, humidity 70%, and culture for 8 hours. When red yeast rice colonies grow on the solid culture medium, pick the pure color part and transfer it to a new culture medium. Repeat several times until the grown colonies have a regular shape and uniform color, and the purification of red yeast rice is completed.
[0068] Step 4: Monascus propagation culture:
[0069] In a sterile environment, a small number of colonies of the Monascus purified in step 3 were picked up with tweezers and inoculated into a liquid seed culture solution placed in a conical flask for propagation culture; when the color of the seed culture solution gradually changed from clear to orange-yellow, the propagation culture of the Monascus was completed;
[0070] Step 5 utilizes highland barley as raw material to produce highland barley red yeast rice, and its method is:
[0071] a selection and cleaning of highland barley: Weigh 50g highland barley into a conical flask, add sterile water and quickly wash 5 times, remove impurities from the surface of highland barley, drain the water and place in a sterile dry place, until the surface of highland barley in the conical flask is basically dry;
[0072] b. Sterilization of highland barley: Place highland barley in a high-pressure steam sterilizer, sterilize under high pressure at 0.1MPa and 121°C for 30min, and let it stand to room temperature;
[0073] C. Cultivation of highland barley red yeast rice: In a sterile environment, 2 ml of acetic acid having a pH of 4 was added to the highland barley, 3 ml of 75% alcohol was added, and 10 ml of the cultured bacteria solution in step 4 was measured with a graduated cylinder and mixed uniformly; the bottle was sealed and placed in a constant temperature and humidity incubator for cultivation, controlling the temperature at 35 ° C and the humidity at 70%; after cultivation for 192 h, the highland barley red yeast rice culture was completed; such as Figure 8As shown, no bacteria were observed, and the product was moved to a drying box for drying at a temperature not exceeding 50°C. The product was dried until the humidity was below 25%, which was the finished product of highland barley red yeast rice.
[0074] According to the comparison of Comparative Examples 1, 2, and 3 with Comparative Example 4, it can be clearly concluded that if the red yeast rice raw material is not cleaned, activated, purified, and expanded, it is used as a strain for the culture of highland barley red yeast rice. As a result, the highland barley culture medium based on highland barley is more or less contaminated by miscellaneous bacteria during the process of culturing highland barley red yeast rice. This shows that the purification step of Monascus in the present invention plays a decisive role in the success or failure of the final culture of highland barley red yeast rice.
[0075] Comparative Example 4 successfully cultivated a highland barley red yeast rice product; the following four examples are based on Comparative Example 4, and different parameters in the cultivation process of Comparative Example 4 are adjusted to obtain the optimal cultivation conditions for highland barley red yeast rice.
[0076] In Example 1, highland barley is puffed, in Example 2, different temperatures are set, in Example 3, different acetic acid levels are set, and in Example 4, different alcohol contents are set.
[0077] Example 1:
[0078] In this embodiment, highland barley is puffed to prepare highland barley solid culture medium for highland barley red yeast rice culture.
[0079] Step 1: Cleaning the red yeast rice;
[0080] Using commercially available red yeast rice as the source of the strain, weigh 10g of red yeast rice, sterilize it with 75% alcohol, and filter out the alcohol after sterilization; then wash it with sterile water 5 times and drain it dry;
[0081] Step 2: activating Monascus;
[0082] The cleaned red yeast rice is immersed in the seed activation solution; then placed in an incubator and cultured for 12 hours. When a velvety bacterial film forms on the surface of the red yeast rice, the red yeast rice activation is complete.
[0083] Step 3: purification of Monascus;
[0084] Take 1 activated red yeast rice raw material, crush it in a 1.5ml sterile centrifuge tube, add 1ml sterile water to dilute it, and use a sterile disposable pipette to take a small amount and inoculate it on the solid culture medium for 8 hours. After the red yeast colonies grow on the solid culture medium, pick the pure color part and transfer it to the new culture medium. Repeat several times until the colonies finally grow with regular shape and uniform color, and the purification of red yeast is completed. Figure 6 The figure shows the final effect of the purification of Monascus on solid culture medium.
[0085] Step 4: Monascus propagation culture:
[0086] In a sterile environment, a few colonies of the Monascus purified in step 3 were picked up with tweezers and inoculated into the liquid seed culture medium placed in a conical flask for expansion culture; Figure 7 As shown, the color of the seed culture solution gradually changes from clear to orange-yellow, and the propagation culture of Monascus is completed;
[0087] In step 5, the puffed highland barley and the unpuffed highland barley are used as raw materials to produce highland barley red yeast rice.
[0088] The seed activation solution in step 2 is prepared from 75% alcohol and maltose aqueous solution, wherein the amount of 75% alcohol and maltose is 3 ml and 2 g per 100 ml of the seed activation solution.
[0089] The incubator in step 2 is a constant temperature and humidity incubator, and its incubation temperature and humidity conditions are set as: temperature 35° C., humidity 70%.
[0090] The solid culture medium of step 3 is prepared by mixing 70 g of glucose, 15 g of beef extract, 2 g of NaNO3, 0.5 g of MgSO4·7H2O, 1.5 g of KH2PO4, and 2 g of agar powder into 1000 ml of sterile water and stirring evenly. The mixture is heated to 121° C., sterilized under high pressure for 30 min, and then dispensed into petri dishes to cool and solidify to form a solid culture medium.
[0091] The preparation method of the seed culture medium described in step 4 is as follows: 70 grams of glucose, 15 grams of beef extract, 32 grams of NaNO, 0.5 grams of MgSO4·7H2O, and 1.5 grams of KH2PO4 are mixed with 1000 ml of sterile water and stirred evenly, and then the mixture is divided into 250 ml conical flasks, wrapped with 8 layers of clean gauze, and sterilized under high pressure at 0.1 MPa and 121°C for 30 minutes to complete the preparation of the seed culture medium.
[0092] The specific culture conditions for the Monascus propagation culture are: placing the inoculated seed culture solution at a temperature of 35° C. and a humidity of 70% for culture, and observing regularly; and culturing for 72 hours until a large number of Monascus fungi visible to the naked eye are generated in the seed culture solution.
[0093] Step 5 utilizes highland barley as raw material to produce highland barley red yeast rice, and its method is:
[0094] a selection and cleaning of highland barley: Weigh 50g highland barley into a conical flask, add sterile water and quickly wash 5 times, remove impurities from the surface of highland barley, drain the water and place in a sterile dry place, until the surface of highland barley in the conical flask is basically dry;
[0095] b. Extrusion of highland barley: After drying, the highland barley is placed in the extrusion equipment, and the highland barley is heated and the skin is broken and expanded;
[0096] c. Sterilization of puffed barley: After puffing, the barley was placed in a high-pressure steam sterilizer at 0.1MPa and 121°C for 30min and then allowed to stand at room temperature;
[0097] d. the production of highland barley red yeast rice: the production of highland barley red yeast rice: in a sterile environment, add acetic acid 2ml that pH is 4 respectively to culture medium, add 75% alcohol 3ml, measure 10ml of bacterial liquid cultivated in step 4 with graduated cylinder, after mixing; Seal bottle mouth, be placed in incubator and cultivate, control temperature 35 ℃, humidity 70%; Cultivate 72h, highland barley red yeast rice is cultivated; Move to drying oven and dry, and baking temperature is no more than 50 ℃; Drying to humidity is below 25%, is highland barley red yeast rice finished product.
[0098] The unexpanded highland barley method in step 5 is as follows:
[0099] a selection and cleaning of highland barley: Weigh 50g highland barley into a conical flask, add sterile water and quickly wash 5 times, remove impurities from the surface of highland barley, drain the water and place in a sterile dry place, until the surface of highland barley in the conical flask is basically dry;
[0100] b. Sterilization of unexpanded barley: Place the barley in a high-pressure steam sterilizer at 0.1MPa and 121°C for 30min after autoclaving and allow to stand at room temperature;
[0101] c. production of highland barley red yeast rice: in a sterile environment, 2ml of acetic acid with a pH of 4 was added to the culture medium, 3ml of 75% alcohol was added, 10ml of the cultured bacteria solution in step 4 was measured with a graduated cylinder, and mixed uniformly; the bottle was sealed and placed in an incubator for cultivation, with a controlled temperature of 35°C and a humidity of 70%; the highland barley red yeast rice was cultivated for 96h; the highland barley red yeast rice was moved to a drying oven for drying, and the drying temperature did not exceed 50°C; and the highland barley red yeast rice was dried to a humidity of less than 25%, which was the finished product.
[0102] Comparison table of red yeast rice production when barley is puffed and unpuffed:
[0103]
[0104] As shown in the table above, the time for the culture medium to change color using unexpanded highland barley is more than double that using expanded highland barley, and the shortest time using expanded highland barley is 96h; In addition, the test results show that Figure 8 and 9 This process produces purer red yeast rice.
[0105] Example 2
[0106] According to Example 1, in this embodiment, highland barley seeds are puffed and highland barley red yeast rice is cultured.
[0107] Step 1: Cleaning the red yeast rice;
[0108] Using commercially available red yeast rice as the source of the strain, weigh 10g of red yeast rice, sterilize it with 75% alcohol, and filter out the alcohol after sterilization; then wash it with sterile water 5 times and drain it dry;
[0109] Step 2: activating Monascus;
[0110] The cleaned red yeast rice is immersed in the seed activation solution; then placed in an incubator and cultured for 12 hours to complete the red yeast rice activation;
[0111] Step 3: purification of Monascus;
[0112] Take one grain of activated red yeast rice, crush it in a 1.5ml sterile centrifuge tube, add 1ml of sterile water to dilute it, and use a sterile disposable pipette to take a small amount and inoculate it onto a solid culture medium for 8 hours. After the red yeast rice colonies grow on the solid culture medium, pick out the pure color part and transfer it to a new culture medium. Repeat this process several times until the grown colonies have regular shape and uniform color, and the purification of red yeast rice is completed.
[0113] Step 4: Monascus propagation culture:
[0114] In a sterile environment, a small amount of the Monascus purified in step 3 is picked up with tweezers and inoculated into a liquid seed culture solution placed in a conical flask for propagation culture; the color of the seed culture solution gradually changes from clear to orange-yellow, and the propagation culture of the Monascus is completed;
[0115] The seed activation solution in step 2 is prepared from an alcohol and maltose aqueous solution, wherein 3 ml of 75% alcohol and 2 g of maltose are used per 100 ml of the seed activation solution.
[0116] The incubator in step 2 is a constant temperature and humidity incubator, and its incubation temperature and humidity conditions are set as: temperature 35° C., humidity 70%.
[0117] The solid culture medium of step 3 is prepared by mixing 70 g of glucose, 15 g of beef extract, 2 g of NaNO3, 0.5 g of MgSO4·7H2O, 1.5 g of KH2PO4, and 2 g of agar powder into 1000 ml of sterile water and stirring evenly. The mixture is heated to 121° C., sterilized under high pressure for 30 min, and then dispensed into petri dishes and cooled and solidified to form a solid culture medium.
[0118] The seed culture medium described in step 4 is prepared by mixing 70 grams of glucose, 15 grams of beef extract, 2 grams of NaNO3, 0.5 grams of MgSO4·7H2O, and 1.5 grams of KH2PO4 into 1000 ml of sterile water and stirring evenly. The mixture is then divided into 250 ml conical flasks, wrapped with 8 layers of clean gauze, and sterilized under high pressure at 0.1 MPa and 121°C for 30 minutes to complete the preparation of the seed culture medium.
[0119] The specific culture conditions for the propagation of Monascus are: placing the inoculated seed culture solution at a temperature of 35°C and a humidity of 70%, and observing it regularly; after culturing for 72 hours, a large number of Monascus fungi visible to the naked eye are generated in the seed culture solution.
[0120] Step 5 utilizes highland barley as raw material to produce highland barley red yeast rice, and its method is:
[0121] a selection and cleaning of highland barley: Weigh 50g highland barley into a conical flask, add sterile water and quickly wash 5 times, remove impurities from the surface of highland barley, drain the water and place in a sterile dry place, until the surface of highland barley in the conical flask is basically dry;
[0122] b. Extrusion of highland barley: After drying, the highland barley is placed in the extrusion equipment, and the highland barley is heated and the skin is broken and expanded;
[0123] c. Sterilization of puffed barley: After puffing, the barley was placed in a high-pressure steam sterilizer at 0.1MPa and 121°C for 30min and then allowed to stand at room temperature;
[0124] D. the production of highland barley red yeast rice: in a sterile environment, add acetic acid 2ml that pH is 4 respectively to culture medium, add 75% alcohol 3ml, measure 10ml of bacterial liquid cultivated in step 4 with graduated cylinder, after mixing; Seal bottle mouth, be placed in incubator and cultivate, control temperature 25, 30, 35, 40 ℃, humidity 70%; Cultivate 96-192h, highland barley red yeast rice is cultivated; Move to drying oven and carry out drying, and baking temperature is no more than 50 ℃; Drying to humidity is below 25%, is highland barley red yeast rice finished product.
[0125] Table 1 Comparison of the time taken to culture highland barley red yeast rice under different culture temperature conditions with the same amount of alcohol and acetic acid:
[0126]
[0127]
[0128] It can be concluded from the above table that, except for the different cultivation time, there is no obvious difference in the final results of each stage of cultivation.
[0129] Example 3
[0130] According to Examples 1 and 2, the highland barley seeds were puffed and cultured at a temperature of 35° C. to culture highland barley red yeast rice.
[0131] Step 1: Cleaning the red yeast rice;
[0132] Use commercially available red yeast rice as the source of the strain, weigh an appropriate amount of red yeast rice, sterilize it with 75% alcohol, filter out the alcohol after sterilization; then wash it with sterile water 5 times and drain it dry;
[0133] Step 2: activating Monascus;
[0134] The cleaned red yeast rice is immersed in the seed activation solution; then placed in an incubator and cultured for 12 hours to complete the red yeast rice activation;
[0135] Step 3: purification of Monascus;
[0136] Take one grain of activated red yeast rice, crush it in a 1.5ml sterile centrifuge tube, add 1ml of sterile water to dilute it, and use a sterile disposable pipette to take a small amount and inoculate it onto a solid culture medium for 8 hours. After the red yeast rice colonies grow on the solid culture medium, pick out the pure color part and transfer it to a new culture medium. Repeat this process several times until the grown colonies have regular shape and uniform color, and the purification of red yeast rice is completed.
[0137] Step 4: Monascus propagation culture:
[0138] In a sterile environment, a small amount of the Monascus purified in step 3 is picked up with tweezers and inoculated into a liquid seed culture solution placed in a conical flask for propagation culture; the color of the seed culture solution gradually changes from clear to orange-yellow, and the propagation culture of the Monascus is completed;
[0139] The seed activation solution in step 2 is prepared from 75% alcohol and maltose aqueous solution, wherein the amount of 75% alcohol is 3 ml and the amount of maltose is 2 g per 100 ml of seed activation solution.
[0140] The incubator in step 2 is a constant temperature and humidity incubator, and its incubation temperature and humidity conditions are set as: temperature 35° C., humidity 70%.
[0141] The solid culture medium of step 3 is prepared by mixing 70 g of glucose, 15 g of beef extract, 2 g of NaNO3, 0.5 g of MgSO4·7H2O, 1.5 g of KH2PO4, and 2 g of agar powder into 1000 ml of sterile water and stirring evenly. The mixture is heated to 121° C., sterilized under high pressure for 30 min, and then dispensed into petri dishes and cooled and solidified to form a solid culture medium.
[0142] The seed culture medium described in step 4 is prepared by mixing 70 grams of glucose, 15 grams of beef extract, 2 grams of NaNO3, 0.5 grams of MgSO4·7H2O, and 1.5 grams of KH2PO4 into 1000 ml of sterile water and stirring evenly. The mixture is then divided into 250 ml conical flasks, wrapped with 8 layers of clean gauze, and sterilized under high pressure at 0.1 MPa and 121°C for 30 minutes to complete the preparation of the seed culture medium.
[0143] The specific culture conditions for the propagation of Monascus are: placing the inoculated seed culture solution at a temperature of 35°C and a humidity of 70%, and observing it regularly; after culturing for 72 hours, a large number of Monascus fungi visible to the naked eye are generated in the seed culture solution.
[0144] Step 5 utilizes highland barley as raw material to produce highland barley red yeast rice, and its method is:
[0145] a selection and cleaning of highland barley: Weigh 50g highland barley into a conical flask, add sterile water and quickly wash 5 times, remove impurities from the surface of highland barley, drain the water and place in a sterile dry place, until the surface of highland barley in the conical flask is basically dry;
[0146] b. Extrusion of highland barley: After drying, the highland barley is placed in the extrusion equipment, and the highland barley is heated and the skin is broken and expanded;
[0147] c. Sterilization of puffed barley: After puffing, the barley was placed in a high-pressure steam sterilizer at 0.1MPa and 121°C for 30min and then allowed to stand at room temperature;
[0148] d. the production of highland barley red yeast rice: in a sterile environment, add acetic acid 0,1,2,3,4ml that pH is 4 respectively to culture medium, add 75% alcohol 2ml, measure 10ml of bacterial liquid cultivated in step 4 with graduated cylinder, after mixing; Seal bottle mouth, be placed in incubator and cultivate, control temperature 35 ℃, humidity 70%; Cultivate 92-120h, highland barley red yeast rice is cultivated; Move to drying oven and carry out drying, and baking temperature is no more than 50 ℃; Drying to humidity is below 25%, is highland barley red yeast rice finished product.
[0149] Table 2 Comparison of the time taken after adding different amounts of acetic acid under the same alcohol dosage and the same culture temperature conditions:
[0150]
[0151] It can be concluded from the above table that Monascus has the characteristics of resistance to acetic acid and alcohol. From the experimental results, the growth time is the same when the amount of acetic acid added in step 5-d is 1-2 ml / 50 g. However, considering that acetic acid has the effect of inhibiting the growth of miscellaneous bacteria, the sterility requirements of the culture environment can be reduced and the production equipment can be simplified. Under the condition of not significantly affecting the growth of Monascus, the amount of acetic acid added is preferably 2 ml / 50 g.
[0152] Example 4
[0153] According to Examples 1, 2, and 3, the highland barley seeds were puffed, and the culture temperature was 35° C., and 2 ml of acetic acid with a pH of 4 was used to culture highland barley red yeast rice.
[0154] Step 1: Cleaning the red yeast rice;
[0155] Use commercially available red yeast rice as the source of the strain, weigh an appropriate amount of red yeast rice, sterilize it with 75% alcohol, filter out the alcohol after sterilization; then wash it with sterile water 5 times and drain it dry;
[0156] Step 2: activating Monascus;
[0157] The cleaned red yeast rice is immersed in the seed activation solution; then placed in an incubator and cultured for 12 hours to complete the red yeast rice activation;
[0158] Step 3: purification of Monascus;
[0159] Take one grain of activated red yeast rice, crush it in a 1.5ml sterile centrifuge tube, add 1ml of sterile water to dilute it, and use a sterile disposable pipette to take a small amount and inoculate it onto a solid culture medium for 8 hours. After the red yeast rice colonies grow on the solid culture medium, pick out the pure color part and transfer it to a new culture medium. Repeat this process several times until the grown colonies have regular shape and uniform color, and the purification of red yeast rice is completed.
[0160] Step 4: Monascus propagation culture:
[0161] In a sterile environment, a small amount of the Monascus purified in step 3 is picked up with tweezers and inoculated into a liquid seed culture solution placed in a conical flask for propagation culture; the color of the seed culture solution gradually changes from clear to orange-yellow, and the propagation culture of the Monascus is completed;
[0162] The seed activation solution in step 2 is prepared from 75% alcohol and maltose aqueous solution, wherein the amount of 75% alcohol is 3 ml and the amount of maltose is 2 g per 100 ml of seed activation solution.
[0163] The incubator in step 2 is a constant temperature and humidity incubator, and its incubation temperature and humidity conditions are set as: temperature 35° C., humidity 70%.
[0164] The solid culture medium of step 3 is prepared by mixing 70 g of glucose, 15 g of beef extract, 2 g of NaNO3, 0.5 g of MgSO4·7H2O, 1.5 g of KH2PO4, and 2 g of agar powder into 1000 ml of sterile water and stirring evenly. The mixture is heated to 121° C., sterilized under high pressure for 30 min, and then dispensed into petri dishes and cooled and solidified to form a solid culture medium.
[0165] The seed culture medium described in step 4 is prepared by mixing 70 grams of glucose, 15 grams of beef extract, 2 grams of NaNO3, 0.5 grams of MgSO4·7H2O, and 1.5 grams of KH2PO4 into 1000 ml of sterile water and stirring evenly. The mixture is then divided into 250 ml conical flasks, wrapped with 8 layers of clean gauze, and sterilized under high pressure at 0.1 MPa and 121°C for 30 minutes to complete the preparation of the seed culture medium.
[0166] The specific culture conditions for the propagation of Monascus are: placing the inoculated seed culture solution at a temperature of 35°C and a humidity of 70%, and observing it regularly; after culturing for 72 hours, a large number of Monascus fungi visible to the naked eye are generated in the seed culture solution.
[0167] Step 5 utilizes highland barley as raw material to produce highland barley red yeast rice, and its method is:
[0168] a selection and cleaning of highland barley: Weigh 50g highland barley into a conical flask, add sterile water and quickly wash 5 times, remove impurities from the surface of highland barley, drain the water and place in a sterile dry place, until the surface of highland barley in the conical flask is basically dry;
[0169] b. Extrusion of highland barley: After drying, the highland barley is placed in the extrusion equipment, and the highland barley is heated and the skin is broken and expanded;
[0170] c. Sterilization of puffed barley: After puffing, the barley was placed in a high-pressure steam sterilizer at 0.1MPa and 121°C for 30min and then allowed to stand at room temperature;
[0171] d. the production of highland barley red yeast rice: in a sterile environment, in expanded highland barley, add acetic acid 2ml that pH is 4 respectively, add 75% alcohol 0,1,2,3,4ml, measure 10ml of the bacterial liquid cultivated in step 4 with a graduated cylinder, after mixing; seal the bottle mouth, be placed in an incubator and cultivate, control temperature 35 ℃, humidity 70%; cultivate 92-136h, highland barley red yeast rice is cultivated; move to a drying oven and dry, and the drying temperature is no more than 50 ℃; drying to a humidity of below 25% is the highland barley red yeast rice finished product.
[0172] Table 3 Comparison of the time taken after adding different amounts of alcohol under the same acetic acid dosage and the same culture temperature conditions:
[0173]
[0174] It can be concluded from the above table that Monascus is resistant to acetic acid and alcohol. From the experimental results, the growth time differed by 12 hours when the amount of acetic acid added in step 5-d was 0, 1, and 2 ml / 50 g. However, considering that acetic acid has the effect of inhibiting the growth of miscellaneous bacteria, the sterility requirements of the culture environment can be reduced and the production equipment can be simplified. Under the condition that it does not significantly affect the growth of Monascus, the amount of alcohol added is set to 2 ml / 50 g.
[0175] From the above embodiments 1, 2, 3, and 4, it can be concluded that when producing highland barley red yeast rice, 2 ml of acetic acid with a pH of 4 and 3 ml of 75% alcohol are added to the puffed highland barley, and when the incubator temperature is set to 35° C., the highland barley red yeast rice incubation time is the shortest, and there is no difference in product phase, which is the optimal embodiment.
[0176] The above shows and describes the basic principles and main features of the present invention and the advantages of the present invention; those skilled in the art should understand that the present invention is not limited to the above embodiments, and the above embodiments and descriptions are only for illustrating the principles of the present invention. Without departing from the spirit and scope of the present invention, the present invention may have various changes and improvements, and these changes and improvements fall within the scope of the present invention to be protected; the scope of protection claimed in the present invention is defined by the attached claims and their equivalents.
Claims
1. A method for producing red yeast rice using highland barley, characterized in that: Including red yeast rice cleaning, red yeast rice activation, purification and expansion culture; And using highland barley as raw material to cultivate highland barley red yeast rice: including: selecting and cleaning highland barley, then puffing the cleaned highland barley, sterilizing the puffed highland barley, and finally using the expanded and cultured Monascus to cultivate highland barley red yeast rice.
2. A method for producing red yeast rice using highland barley according to claim 1, characterized in that: The red yeast rice raw material is cleaned; the commercial red yeast rice raw material is used as the source of the strain, 10g of the red yeast rice raw material is weighed, sterilized with 75% alcohol, and the alcohol is filtered out after sterilization; then washed with sterile water 5 times and the water is dried; The red yeast rice is activated; the cleaned red yeast rice raw material is added to the seed activation liquid and immersed; then placed in a constant temperature and humidity incubator, the temperature and humidity are set to: temperature 35 ° C, humidity 70%, and after culturing for 12 hours, when a velvety bacterial film is formed on the surface of the red yeast rice raw material, the red yeast rice activation is completed; Purification of the Monascus purificans; Take one activated red yeast rice raw material, crush it in a 1.5ml sterile centrifuge tube, add 1ml sterile water to dilute it, and use a sterile disposable pipette to draw a small amount and inoculate it on the solid culture medium. The solid culture medium is placed in a constant temperature and humidity incubator, and the temperature and humidity are set to: temperature 35°C, humidity 70%, and culture for 8 hours. When red yeast rice colonies grow on the solid culture medium, pick the pure color part and transfer it to a new culture medium. Repeat several times until the grown colonies have a regular shape and uniform color, and the purification of red yeast rice is completed. The Monascus propagation culture is as follows: a small number of colonies of the Monascus purified in step 3 are picked up with tweezers in a sterile environment and inoculated into a liquid seed culture solution in a conical flask for propagation culture; when the color of the seed culture solution gradually changes from clear to orange-yellow, the Monascus propagation culture is completed.
3. A method for producing red yeast rice using highland barley according to claim 2, characterized in that: The seed activation liquid is prepared from 75% alcohol and maltose aqueous solution, wherein the alcohol dosage is 3 ml and the maltose dosage is 2 g for every 100 ml of the seed activation liquid.
4. A method for producing red yeast rice using highland barley according to claim 2, characterized in that: The solid culture medium is prepared by mixing 70 grams of glucose, 15 grams of beef extract, 2 grams of NaNO3, 0.5 grams of MgSO4·7H2O, 1.5 grams of KH2PO4, and 2 grams of agar powder into 1000 ml of sterile water and stirring evenly. The mixture is heated to 121° C., sterilized under high pressure for 30 minutes, and then dispensed into petri dishes and cooled and solidified to form a solid culture medium.
5. A method for producing red yeast rice using highland barley according to claim 2, characterized in that: The seed culture medium is prepared by mixing 70 grams of glucose, 15 grams of beef extract, 2 grams of NaNO3, 0.5 grams of MgSO4·7H2O, and 1.5 grams of KH2PO4 into 1000 ml of sterile water and stirring evenly. The mixture is then divided into 250 ml conical flasks and sterilized under high pressure at 0.1 MPa and 121°C for 30 minutes to complete the preparation of the seed culture medium.
6. A method for producing red yeast rice using highland barley according to claim 2, characterized in that: The specific culture conditions for the Monascus propagation culture are: placing the inoculated seed culture solution at a temperature of 35° C. and a humidity of 70% for culture; and culturing for 72 hours until a large number of Monascus fungi visible to the naked eye are generated in the seed culture solution.
7. The method for producing red yeast rice using highland barley according to claim 1, wherein: The selection and cleaning of the highland barley are as follows: weighing 50g of highland barley and putting it into a conical flask, adding sterile water and quickly washing it 5 times, removing impurities on the surface of the highland barley, draining the water and placing it in a sterile dry place for 1-2h, until the surface of the highland barley in the conical flask is basically dry.
8. The method for producing red yeast rice using highland barley according to claim 7, wherein: The puffing of the highland barley is as follows: the dried highland barley is put into a puffing device, and the highland barley is heated to break the skin and puff.
9. The method for producing red yeast rice using highland barley according to claim 8, wherein: The sterilization of the puffed barley is to put the puffed barley into a high-pressure steam sterilization device, sterilize it under high pressure for 30 minutes at 0.1 MPa and 121° C., and then let it stand until it reaches room temperature.
10. The method for producing red yeast rice using highland barley according to claim 9, wherein: Cultivation of highland barley red yeast rice: in a sterile environment, respectively adding 0-4 ml of acetic acid with a pH of 4 and 2-4 ml of 75% alcohol to puffed highland barley, and measuring 10 ml of the bacterial solution cultured in step 4 with a graduated cylinder, and uniformly mixing the mixture; sealing the bottle mouth, placing the mixture in a constant temperature and humidity incubator for cultivation, controlling the temperature at 25-40 DEG C and the humidity at 70%; culturing the mixture for 96-192 hours until the highland barley red yeast rice is cultivated; moving the mixture to a drying oven for drying, with the drying temperature not exceeding 50 DEG C; and drying the mixture until the humidity is below 25% to obtain the highland barley red yeast rice finished product.