Fermentation method for producing gibberellic acid with low energy consumption

Abstract

The invention discloses a low-energy-consumption fermentation method for producing gibberellic acid, which does not need stirring during fermentation, and comprises the following steps: S1, taking a seed solution and a fermentation culture solution, the mass of the seed solution being 5-25% of the mass of the fermentation culture solution; s2, adding the seed solution and the fermentation culture solution into a production tank; s3, a target product gibberellic acid biosynthesis reaction is carried out, in the target product gibberellic acid biosynthesis reaction, a carbon source is supplemented in a fed-batch feeding mode, and ammonia water is supplemented to control the PH; in the step S3, the air flow is 1500 to 4600 Nm < 3 > / h, the pH value is 5.0 to 5.2, the pressure is 0.01 to 0.05 MPa, and the reaction time for biosynthesizing the target product gibberellic acid is 8 to 10 days; in the step S3, the material supplementing mode of supplementing the carbon source comprises the steps of firstly supplementing the sugar, then increasing the oil supplementing rate on the basis of maintaining the sugar supplementing rate, and then supplementing the material in a mode that the ratio of the sugar to the oil is gradually reduced, the material supplementing rate is controlled according to dissolved oxygen, when the dissolved oxygen is lower than a tolerance value, the material supplementing rate is reduced, and when the dissolved oxygen is higher than the tolerance value, the material supplementing rate is increased. And the dissolved oxygen in the fermentation process is always kept in a relatively stable interval. According to the method disclosed by the invention, relatively high gibberellic acid yield can be obtained without stirring.

Description

Technical Field

[0001] The invention relates to gibberellic acid production, and in particular to a fermentation method for producing gibberellic acid with low energy consumption. Background Art

[0002] Gibberellic Acid 3 (GA3) is a plant growth regulating hormone produced by Gibberellic fungi. It plays an important regulatory role in plant growth and development and is used in many fields, including agriculture, forestry and brewing, with good economic value and social benefits. The traditional production methods of GA3 mainly include plant extraction, chemical synthesis and microbial fermentation; however, the yield of GA3 extracted from plants is low, it is easy to produce a lot of waste and the cost is high, and the chemical synthesis method is also limited by cumbersome process steps and the resulting pollution; with the rapid development of synthetic biology, microbial production of GA3 has attracted more and more attention.

[0003] At present, GA3 can be produced by fermentation of a variety of fungi, such as Fusarium fujikuroi, Aspergillus niger and Fusarium oxysporum; the same cluster of strains Fusarium moniliforme has a higher yield, but requires solid-state fermentation culture, which is not suitable for industrial production. Therefore, the main application of industrial production of gibberellin GA3 is Fusarium fujikuroi, which has a higher yield and can be fermented in a controlled manner.

[0004] In recent years, due to its wide application in agriculture and horticulture, the GA3 market has significant potential and market demand. Although the industrial production of GA3 has been achieved using Fusarium fusiformis, there are still problems such as low GA3 production level and high cost. The existing GA3 production adopts universal ventilated stirred reactor technology for industrial product production (such as CN201990659U). In GA3 production, energy consumption (air oxygen supply, steam sterilization, stirring power consumption, circulating water cooling) accounts for 20% to 30% of the total product cost, of which stirring accounts for 20% to 40% of the total energy consumption.

[0005] In the existing ventilated stirred reactor technology (such as CN201990659U), in the fermentation tank production stage of GA3, such as CN108285915A, the seed solution, culture solution and feed all enter through the feed port, and the feeding process is: the dissolved oxygen rebounds by 20%, and the automatic flow of glucose and salad oil is turned on, glucose (45%) solution: oil (3.2L:1L), sugar and oil are added at the same time, and the dissolved oxygen control point is 20% to 30%.

[0006] Therefore, how to reduce energy consumption without reducing GA3 production has become an urgent problem that the industry needs to solve.

[0007] The above background technology is for facilitating the understanding of the present invention and is not the known technology disclosed to the general public before the application of the present invention. Summary of the invention

[0008] In view of the above-mentioned defects, the present invention provides a fermentation method for producing gibberellic acid with low energy consumption. In the step of biosynthesizing the target product gibberellic acid, a higher gibberellic acid yield can be obtained without stirring.

[0009] The technical solution is: a fermentation method for producing gibberellic acid with low energy consumption, without stirring during fermentation, comprising the following steps:

[0010] S1, taking seed solution and fermentation culture medium, the volume of seed solution is 5-25% of the volume of fermentation culture medium;

[0011] S2, adding the seed solution and the fermentation culture solution into the production fermentation tank;

[0012] S3, performing an acidogenic reaction, during which a carbon source is supplemented by feeding, and ammonia water is added to control pH;

[0013] In S3, the air flow rate is 1500~4600Nm 3 / h, pH 5.0-5.2, pressure 0.01-0.05MPa, biosynthesis reaction time 8-10 days;

[0014] In S3, the feeding method for supplementing the carbon source is to first feed sugar, then increase the oil feeding rate on the basis of maintaining the sugar feeding rate, and then feed in a manner in which the sugar-oil ratio is gradually reduced. The feeding rate is controlled according to the dissolved oxygen. When the dissolved oxygen is lower than the tolerance value, the feeding rate is reduced. When the dissolved oxygen is higher than the tolerance value, the feeding rate is increased, so that the dissolved oxygen in the fermentation process is always maintained in a relatively stable range; the sugar-oil ratio = sugar solution volume / oil volume.

[0015] Furthermore, the feeding method of supplementing the nitrogen source and the carbon source includes the following stages:

[0016] The first stage: acid production reaction until dissolved oxygen rebounds by 50-80%, then sugar solution is added, the initial rate of sugar solution addition is 20-50L / h, and then gradually increased until the sugar solution addition rate reaches 100-200L / h;

[0017] The second stage: maintain the sugar solution feeding rate, start to feed oil, the initial oil feeding rate is 1-10L / h, and then gradually increase until the oil feeding rate is 30-65L / h;

[0018] The third stage: feed according to the sugar-oil ratio of 4.0-3.0:1, the dissolved oxygen control point is 40%-50%, when the dissolved oxygen is lower than 40-45%, reduce the feeding speed, when the dissolved oxygen is higher than 45-50%, increase the feeding speed;

[0019] The fourth stage: feed according to the sugar-oil ratio of 3.5-3.0:1, the dissolved oxygen control point is 30%-40%, when the dissolved oxygen is lower than 30-35%, reduce the feeding speed, when the dissolved oxygen is higher than 35-40%, increase the feeding speed;

[0020] The fifth stage: feed according to the sugar-oil ratio of 3.0-2.5:1, the dissolved oxygen control point is 20%-30%, when the dissolved oxygen is lower than 20-25%, reduce the feeding speed, when the dissolved oxygen is higher than 25-30%, increase the feeding speed;

[0021] The sixth stage: feed according to the sugar-oil ratio of 2.5-2.0:1, the dissolved oxygen control point is 15%-25%, when the dissolved oxygen is lower than 15-20%, reduce the feeding speed, when the dissolved oxygen is higher than 20-25%, increase the feeding speed;

[0022] The seventh stage: feed according to the sugar-oil ratio of 2.0-1.5:1, the dissolved oxygen control point is 10%-20%, when the dissolved oxygen is lower than 10-15%, reduce the feeding speed, when the dissolved oxygen is higher than 15-20%, increase the feeding speed;

[0023] The eighth stage: feed according to the sugar-oil ratio of 1.5 to 1.0:1, the dissolved oxygen control point is 5% to 15%, when the dissolved oxygen is lower than 5-10%, reduce the feeding speed, when the dissolved oxygen is higher than 10-15%, increase the feeding speed.

[0024] Furthermore, the sugar is glucose, and the oil is salad oil.

[0025] Furthermore, the production tank comprises a tank body, on which a feed port, a discharge port, and a feed replenishment and air passage port are provided, the tank body forms an inner cavity of the tank body, the feed port is provided at the top of the tank body, the discharge port is provided at the bottom of the tank body, the feed replenishment and air passage port is located at the lower part of the tank body, the feed replenishment and air passage port is connected with a main pipe, the main pipe passes through the tank body, a part of the main pipe is located in the inner cavity of the tank body, and the rest of the main pipe is located outside the inner cavity of the tank body, the end of the main pipe located outside the inner cavity of the tank body is connected with an air inlet pipe and a feed replenishment pipe, a feed replenishment valve and a feed replenishment cup are installed on the feed replenishment pipe, an air inlet valve is installed on the air inlet pipe, an inner cylinder is installed in the inner cavity of the tank body, the top of the inner cylinder is provided with an upper opening, the bottom is provided with a lower opening, the inner cylinder forms an inner cavity of the inner cylinder, the main pipe is located in the tank Part of the body cavity passes through the inner cylinder and is located in the inner cavity of the inner cylinder. Its end opening is downward and is connected to the distributor. A first pushing unit is arranged below the bottom of the distributor. More than three horizontal distribution pipes are connected to the side wall of the distributor. The more than three horizontal distribution pipes are evenly distributed along the circumference. More than two second pushing units are arranged above each horizontal distribution pipe. The first pushing unit is located below the bottom of the inner cylinder, and the second pushing unit is located in the inner cavity of the inner cylinder. The first pushing unit is connected to the distributor through a connecting pipe, and the second pushing unit is connected to the horizontal distribution pipe through a connecting pipe. The pushing inlet of the first pushing unit is at the top and the pushing outlet is at the bottom, and the pushing inlet of the second pushing unit is at the bottom and the pushing outlet is at the top.

[0026] Furthermore, the supplementary carbon source and ammonia water are added from a feed pipe.

[0027] Compared with the prior art, the present invention has the following beneficial effects:

[0028] The present invention achieves the purpose of saving energy by eliminating agitation in the production fermentation tank of the present invention, and then improves the yield of the biosynthetic target product gibberellic acid (not lower than the existing ventilation stirring reactor technology) through the unique feeding method of the present invention. BRIEF DESCRIPTION OF THE DRAWINGS

[0029] Figure 1 It is a schematic diagram of the overall structure of the present invention;

[0030] In the figure: 1. tank body, 2. feed port, 3. discharge port, 4. feeding and air passage, 5. main pipe, 51. air inlet pipe, 52. feeding pipe, 53. feeding valve, 54. feeding cup, 55. air inlet valve, 6. inner cylinder, 61. upper opening, 62. lower opening, 7. inner cavity of tank body, 8. inner cavity of inner cylinder, 9. distributor, 10. first pushing unit, 11. horizontal distribution pipe, 12. second pushing unit, 13. connecting pipe. DETAILED DESCRIPTION

[0031] The present invention will be further described below in conjunction with the accompanying drawings.

[0032] In the description of the present invention, it should be noted that, unless otherwise clearly specified and limited, the terms "installed", "connected" and "connected" should be understood in a broad sense, for example, it can be a fixed connection, or it can be an indirect connection through an intermediate medium, it can be the internal connection of two elements or the interaction relationship between two elements. For ordinary technicians in this field, the specific meanings of the above terms in the present invention can be understood according to specific circumstances.

[0033] In the description of the present invention, it should be understood that the terms "upper", "lower", "front", "back", "vertical", "horizontal", "top", "bottom", "inside", "outside" and the like indicate positions or positional relationships based on the positions or positional relationships shown in the drawings, and are only for the convenience of describing the present invention and simplifying the description, rather than indicating or implying that the device or element referred to must have a specific position, be constructed and operated in a specific position, and therefore cannot be understood as limiting the present invention. In the description of the present invention, "plurality" means two or more, unless otherwise precisely and specifically specified.

[0034] The terms "first", "second", "third", "fourth", etc. (if any) in the specification and claims of the present application and the above-mentioned drawings are used to distinguish similar objects, and are not necessarily used to describe a specific order or sequence. It should be understood that the data used in this way can be interchangeable where appropriate, so that the embodiments of the present application described herein can be implemented in an order other than those illustrated or described herein, for example. In addition, the terms "including" and "having" and any of their variations are intended to cover non-exclusive inclusions, for example, a process, method, system, product or device comprising a series of steps or units is not necessarily limited to those steps or units clearly listed, but may include other steps or units that are not clearly listed or inherent to these processes, methods, products or devices.

[0035] In order to make the purpose, technical solution and advantages of the embodiments of the present application clearer, the technical solution in the embodiments of the present application will be clearly and completely described below in conjunction with the drawings in the embodiments of the present application. Obviously, the described embodiments are part of the embodiments of the present application, not all of the embodiments. Based on the embodiments in the present application, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of this application.

[0036] The technical solution of the present invention is described in detail with specific embodiments below. The following specific embodiments can be combined with each other, and the same or similar concepts or processes may not be described in detail in some embodiments.

[0037] Please refer to Figure 1 , Figure 1 It is a schematic diagram of the overall structure of the present invention.

[0038] A low-energy consumption gibberellic acid production tank comprises a tank body 1, on which a feed port 2, a discharge port 3 and a feed and air passage port 4 are provided, the tank body 1 encloses a tank inner cavity 7, the feed port 2 is provided at the top of the tank body 1, the discharge port 3 is provided at the bottom of the tank body 1, the feed and air passage port 4 is located at the lower part of the tank body 1, the feed and air passage port 4 is connected with a main pipe 5, the main pipe 5 passes through the tank body 1, a part of the main pipe 5 is located in the tank inner cavity 7, and the rest of the main pipe 5 is located outside the tank inner cavity 7, the end of the main pipe 5 located outside the tank inner cavity 7 is connected with an air inlet pipe 51 and a feed pipe 52, a feed valve 53 and a feed cup 54 are installed on the feed pipe 52, an air inlet valve 55 is installed on the air inlet pipe 51, an air inlet end of the air inlet pipe 51 is connected with an air compressor (not shown in the figure), a cooling coil (not shown in the figure) and an inner tube 6 are installed in the tank inner cavity 7, the inner tube 6 has an upper opening 61 at the top and an opening 62 at the bottom There is a lower opening 62, and the inner tube 6 forms an inner tube cavity 8. The part of the main pipe 5 located in the inner tube cavity 7 of the tank body passes through the inner tube 6 and is located in the inner tube cavity 8. Its end opening is downward and is connected to the distributor 9. A first pushing unit 10 is arranged below the bottom of the distributor 9. More than three horizontal distribution pipes 11 are connected to the side wall of the distributor 9. The more than three horizontal distribution pipes 11 are evenly distributed along the circumference. More than two second pushing units 12 are arranged above each horizontal distribution pipe 11. The first pushing unit 10 is located below the bottom of the inner tube 6, and the second pushing unit 12 is located in the inner tube cavity 8. The first pushing unit 10 is connected to the distributor 9 through the connecting pipe 13, and the second pushing unit 12 is connected to the horizontal distribution pipe 11 through the connecting pipe 13. The pushing inlet of the first pushing unit 10 is at the top, and the pushing outlet is at the bottom. The pushing inlet of the second pushing unit 12 is at the bottom, and the pushing outlet is at the top.

[0039] The feed valve 53 and the air inlet valve 55 are both electromagnetic valves. The cooling coil and the tank body 1 are the same as those in the prior art. The positions of the feed port 2 and the discharge port 3 are the same as those in the prior art. In the prior art, the cooling coil takes away the heat generated in the reaction to maintain the reaction temperature. The main pipe 5 passes through the feed and air through port 4, and the main pipe 5 and the tank body 1 are fixed by welding at the feed and air through port 4. The bottom of the inner tube 6 is fixed by a support, and the bottom of the support is connected to the inner wall of the bottom of the tank body 1.

[0040] Furthermore, there are eight horizontal distribution pipes 11 , and two second pushing units 12 are installed on each distribution pipe 11 .

[0041] Furthermore, the main bodies of the first pushing unit 10 and the second pushing unit 12 are both cylinders, one end of the cylinder is connected to the pushing inlet, and the other end is connected to the pushing outlet.

[0042] Furthermore, the push inlet and the push outlet are both conical structures, the small ends of the push inlet and the push outlet are connected to the cylinder, and the large ends are far away from the cylinder.

[0043] Furthermore, the diameter of the portion of the connecting pipe 13 located in the push inlet gradually decreases along the gas running direction.

[0044] Furthermore, the pushing outlet of the first pushing unit 10 is adjacent to the discharge port 3 and is located above the discharge port 3 .

[0045] Furthermore, the upper opening 61 is in the shape of an inverted truncated cone.

[0046] Before production, production materials enter the tank body cavity 7 and the inner tube cavity 8 through the feed port 3. During production, compressed gas enters from the air inlet pipe 51 through the main pipe 5, and runs through the first pushing unit 10 and the second pushing unit 12. Among them, the second pushing unit 12 diffuses and flows upward, and circulates from the inside to the outside in a directional manner, forming an inside-up and outside-down. The first pushing unit 10 disturbs and mixes the material retained at the bottom of the tank body cavity 7, and the rising gas drives the material in the retention area to the inner tube cavity 8.

[0047] Example 1 Preparation of Seed Liquid before Fermentation

[0048] The seed liquid before fermentation was prepared according to the process 1.1-1.3 in Example 1 of CN108285915A, wherein the GA3 strain, process and culture medium were exactly the same. The GA3 strain was Gibberella fujikura, and the deposit number was CGMCC No. 23265 (General Microbiological Center of China Microbiological Culture Collection Administration Committee).

[0049] The seed liquid test data is shown in Table 1.

[0050] Table 1 Seed liquid test data before fermentation

[0051] Wet weight PH Mycelium length Mycelium thickness Is the cytoplasm uniform? Whether there are cavitation bubbles Fermentation broth viscosity 17-25％ 4.9-5.5 Longer Thinner More uniform Small amount ++++

[0052] The seed liquid prepared in Example 1 was used to prepare the fermentation liquids of the following Examples 2 to 7 and Comparative Examples 1 to 2.

[0053] In Examples 2 to 7 and Comparative Examples 1 to 2, the concentration and dosage of the seed solution are equal.

[0054] Example 2

[0055] This embodiment adopts Figure 1 The low-energy consumption gibberellic acid production tank of the present invention has a volume of the inner cavity 7 of the tank body of 156 tons (measured in terms of water volume), the axis of the inner cylinder 6 is the same as the axis of the tank body 1, the diameter of the inner cylinder 6 is 4 / 5 of the inner diameter of the tank body 1, and the height is 0.7 of the height of the tank body 1. The distance between the end of the other end of the horizontal distribution pipe 11 and the inner wall of the inner cylinder 6 is 1 / 16 of the inner radius of the inner cylinder 6, and the distance between the outer wall of the bottom of the inner cylinder 6 and the discharge port is approximately 1 / 4 of the inner radius of the tank body 1. The tank body 1 has a height of 12m and a diameter of 4m.

[0056] A low-energy consumption gibberellic acid fermentation method comprises the following steps:

[0057] S1, taking the seed solution and fermentation culture medium of Example 1, the mass of the seed solution is 10% of the mass of the fermentation culture medium.

[0058] S2, adding the seed liquid and fermentation culture liquid into the tank body 1 from the feed port 2.

[0059] S3, turn on the heating device ( Figure 1 (not shown) and opening the air inlet valve 55 to carry out acid production reaction. During the acid production reaction, nitrogen source and carbon source are supplemented and pH is controlled by adding ammonia water. The biosynthesis reaction time is 198h.

[0060] Air flow rate is 2200Nm 3 / h, the operating tank pressure is controlled at 0.035MPa, the temperature is controlled at 29±0.2℃, the pH is 5.0~5.2, and the feed and ammonia water are added from the feed pipe 52 (when feeding, open the feed valve 53).

[0061] After the fermentation was completed, samples were taken for testing. The results are shown in Table 3.

[0062] In S1, the fermentation medium and the amount added are as shown in Table 2:

[0063] Table 2 Fermentation broth and addition amount

[0064]

[0065]

[0066] In S3, the feeding method is as follows:

[0067] The first stage: Acid production reaction occurs until the dissolved oxygen rebounds by 60%, and then glucose (45%) solution is added. The initial rate of glucose solution addition is 30L / h, and then it is gradually increased until the glucose solution addition rate reaches 130L / h. The glucose solution addition rate is maintained unchanged. This stage is about 3-4h.

[0068] The second stage: start to add oil, the initial rate of oil addition is 2L / h, and then gradually increase until the oil addition rate reaches 42L / h. During this stage, the glucose solution addition rate is constant at 130L / h, and this stage lasts for about 40-45h.

[0069] The third stage: feed according to the sugar-oil ratio (sugar solution volume / oil volume) of 3.2:1, the dissolved oxygen control point is 45%, when the dissolved oxygen is lower than 45%, reduce the feeding speed, when the dissolved oxygen is higher than 45%, increase the feeding speed, this stage is 24h;

[0070] The fourth stage: feed according to the sugar-oil ratio (sugar solution volume / oil volume) of 2.7:1, the dissolved oxygen control point is 35%, when the dissolved oxygen is lower than 35%, reduce the feeding speed, when the dissolved oxygen is higher than 35%, increase the feeding speed, this stage is 24h;

[0071] The fifth stage: feed according to the sugar-oil ratio (sugar solution volume / oil volume) of 2.5:1, the dissolved oxygen control point is 25%, when the dissolved oxygen is lower than 25%, reduce the feeding speed, when the dissolved oxygen is higher than 25%, increase the feeding speed, this stage is 6h;

[0072] The sixth stage: feed according to the sugar-oil ratio (sugar solution volume / oil volume) of 2.0:1, the dissolved oxygen control point is 20%, when the dissolved oxygen is lower than 20%, reduce the feeding rate, when the dissolved oxygen is higher than 20%, increase the feeding rate, this stage is 6h.

[0073] The seventh stage: feed according to the sugar-oil ratio of 1.5:1 (sugar solution volume / oil volume). The dissolved oxygen control point is 15%. When the dissolved oxygen is lower than 15%, reduce the feeding rate. When the dissolved oxygen is higher than 15%, increase the feeding rate. This stage lasts for 6 hours.

[0074] The eighth stage: feed according to the sugar-oil ratio of 1.0:1 (sugar solution volume / oil volume), the dissolved oxygen control point is 10%, when the dissolved oxygen is lower than 10%, reduce the feeding speed, when the dissolved oxygen is higher than 10%, increase the feeding speed. This stage lasts until the end of production (i.e. tank loading).

[0075] Example 3

[0076] The difference between this embodiment and embodiment 2 is only the feeding method. The feeding method of this embodiment is:

[0077] The first stage: Acid production reaction occurs until the dissolved oxygen rebounds by 60%, and then glucose (45%) solution is added. The initial rate of glucose solution addition is 30L / h, and then it is gradually increased until the glucose solution addition rate reaches 130L / h. The glucose solution addition rate is maintained unchanged. This stage is about 3-4h.

[0078] The second stage: start to add oil, the initial rate of oil addition is 2L / h, and then gradually increase until the oil addition rate reaches 42L / h. During this stage, the glucose solution addition rate is constant at 130L / h, and this stage lasts for about 40-45h.

[0079] The third stage: feed according to the sugar-oil ratio (sugar solution volume / oil volume) of 3.2:1, the dissolved oxygen control point is 45%, when the dissolved oxygen is lower than 45%, reduce the feeding speed, when the dissolved oxygen is higher than 45%, increase the feeding speed, this stage is 24h;

[0080] The fourth stage: feed according to the sugar-oil ratio (sugar solution volume / oil volume) of 2.7:1, the dissolved oxygen control point is 35%, when the dissolved oxygen is lower than 35%, reduce the feeding speed, when the dissolved oxygen is higher than 35%, increase the feeding speed, this stage is 24h;

[0081] The fifth stage: feed according to the sugar-oil ratio (sugar solution volume / oil volume) of 2.5:1, the dissolved oxygen control point is 25%, when the dissolved oxygen is lower than 25%, reduce the feeding speed, when the dissolved oxygen is higher than 25%, increase the feeding speed, this stage is 6h;

[0082] The sixth stage: feed according to the sugar-oil ratio (sugar solution volume / oil volume) of 2.0:1, the dissolved oxygen control point is 20%, when the dissolved oxygen is lower than 20%, reduce the feeding rate, when the dissolved oxygen is higher than 20%, increase the feeding rate, this stage is 6h.

[0083] The seventh stage: feed according to the sugar-oil ratio of 1.5:1 (sugar solution volume / oil volume), the dissolved oxygen control point is 15%, when the dissolved oxygen is lower than 15%, reduce the feeding speed, when the dissolved oxygen is higher than 15%, increase the feeding speed. This stage lasts until the end of production (i.e. tank loading).

[0084] After the fermentation was completed, samples were taken for testing. The results are shown in Table 3.

[0085] Example 4

[0086] The difference between this embodiment and embodiment 2 is only the feeding method. The feeding method of this embodiment is:

[0087] The first stage: Acid production reaction occurs until the dissolved oxygen rebounds by 60%, and then glucose (45%) solution is added. The initial rate of glucose solution addition is 30L / h, and then it is gradually increased until the glucose solution addition rate reaches 130L / h. The glucose solution addition rate is maintained unchanged. This stage is about 3-4h.

[0088] The second stage: start to add oil, the initial rate of oil addition is 2L / h, and then gradually increase until the oil addition rate reaches 42L / h. During this stage, the glucose solution addition rate is constant at 130L / h, and this stage lasts for about 40-45h.

[0089] The third stage: feed according to the sugar-oil ratio (sugar solution volume / oil volume) of 3.2:1, the dissolved oxygen control point is 45%, when the dissolved oxygen is lower than 45%, reduce the feeding speed, when the dissolved oxygen is higher than 45%, increase the feeding speed, this stage is 24h;

[0090] The fourth stage: feed according to the sugar-oil ratio (sugar solution volume / oil volume) of 2.7:1, the dissolved oxygen control point is 35%, when the dissolved oxygen is lower than 35%, reduce the feeding speed, when the dissolved oxygen is higher than 35%, increase the feeding speed, this stage is 24h;

[0091] The fifth stage: feed according to the sugar-oil ratio (sugar solution volume / oil volume) of 2.5:1, the dissolved oxygen control point is 25%, when the dissolved oxygen is lower than 25%, reduce the feeding speed, when the dissolved oxygen is higher than 25%, increase the feeding speed, this stage is 6h;

[0092] The sixth stage: feed according to the sugar-oil ratio of 2.0:1 (sugar solution volume / oil volume), the dissolved oxygen control point is 20%, when the dissolved oxygen is lower than 20%, reduce the feeding speed, when the dissolved oxygen is higher than 20%, increase the feeding speed. This stage lasts until the end of production (i.e. tank loading).

[0093] Example 5

[0094] The difference between this embodiment and embodiment 2 is only the feeding method. The feeding method of this embodiment is:

[0095] The first stage: Acid production reaction occurs until the dissolved oxygen rebounds by 60%, and then glucose (45%) solution is added. The initial rate of glucose solution addition is 30L / h, and then it is gradually increased until the glucose solution addition rate reaches 130L / h. The glucose solution addition rate is maintained unchanged. This stage is about 3-4h.

[0096] The second stage: start to add oil, the initial rate of oil addition is 2L / h, and then gradually increase until the oil addition rate reaches 42L / h. During this stage, the glucose solution addition rate is constant at 130L / h, and this stage lasts for about 40-45h.

[0097] The third stage: feed according to the sugar-oil ratio (sugar solution volume / oil volume) of 3.2:1, the dissolved oxygen control point is 45%, when the dissolved oxygen is lower than 45%, reduce the feeding speed, when the dissolved oxygen is higher than 45%, increase the feeding speed, this stage is 24h;

[0098] The fourth stage: feed according to the sugar-oil ratio (sugar solution volume / oil volume) of 2.7:1, the dissolved oxygen control point is 35%, when the dissolved oxygen is lower than 35%, reduce the feeding speed, when the dissolved oxygen is higher than 35%, increase the feeding speed, this stage is 24h;

[0099] The fifth stage: feed according to the sugar-oil ratio of 2.5:1 (sugar solution volume / oil volume). The dissolved oxygen control point is 25%. When the dissolved oxygen is lower than 25%, the feeding rate is reduced. When the dissolved oxygen is higher than 25%, the feeding rate is increased. This stage lasts until the end of production (i.e., tank loading).

[0100] Example 6

[0101] The difference between this embodiment and embodiment 2 is that the feeding method adopts the feeding method in embodiment 1 of CN108285915B, that is, the feeding method is: when the dissolved oxygen rebounds by 20%, the automatic flow addition of glucose and salad oil is turned on for feeding, glucose (45%) solution: oil (3.2L:1L), the dissolved oxygen control point is 20% to 30%, when the dissolved oxygen is lower than 20%, the feeding speed is reduced, and when the dissolved oxygen is higher than 30%, the feeding speed is increased.

[0102] After the fermentation was completed, samples were taken for testing. The results are shown in Table 3.

[0103] Example 7

[0104] The only difference between this embodiment and embodiment 2 is that the supplementary material is added from the feed inlet 2.

[0105] After the fermentation was completed, samples were taken for testing. The results are shown in Table 3.

[0106] Comparative Example 1

[0107] The difference between this comparative example and Example 2 is that the supplementary material is added from the feed inlet 2, and the supplementary material method is the same as the supplementary material method in Example 1 of CN108285915A.

[0108] After the fermentation was completed, samples were taken for testing. The results are shown in Table 3.

[0109] Comparative Example 2

[0110] Compared with Example 2, this comparative example has the following differences:

[0111] 1. The production tank adopts the production tank of CN201990659U. The tank body, height, diameter, volume, heating and cooling system of the production tank are the same as those of the production tank of the present invention. The only difference is that the CN201990659U is a stirring type with an agitator installed inside, while the present invention has an inner cylinder 6 and supporting components such as a first pushing unit 10, etc.

[0112] 2. The stirring speed of this comparative example is: 150-200rpm, and the motor power is 500kw.h.

[0113] 3. The supplementary material is added from the feed inlet, and the supplementary material method is the same as the supplementary material method in Example 1 of CN108285915A.

[0114] After the fermentation was completed, samples were taken for testing. The results are shown in Table 3.

[0115] Table 3 Fermentation broth test data

[0116]

[0117] The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. For those skilled in the art, the present invention may have various modifications and variations. Any modification, equivalent replacement, improvement, etc. made within the spirit and principle of the present invention shall be included in the protection scope of the present invention.

Claims

1. A fermentation method for producing gibberellic acid with low energy consumption, characterized in that: Fermentation is performed without stirring and includes the following steps: S1, taking seed solution and fermentation culture medium, the volume of seed solution is 5-25% of the volume of fermentation culture medium; S2, adding the seed solution and the fermentation culture solution into the production fermentation tank; S3, performing an acidogenic reaction, during which a carbon source is supplemented by feeding, and ammonia water is added to control pH; In S3, the air flow rate is 1500~4600Nm 3 / h, pH 5.0-5.2, pressure 0.01-0.05MPa, biosynthesis reaction time 8-10 days; In S3, the feeding method for supplementing the carbon source is to first feed sugar, then increase the oil feeding rate on the basis of maintaining the sugar feeding rate, and then feed in a manner in which the sugar-oil ratio is gradually reduced. The feeding rate is controlled according to the dissolved oxygen. When the dissolved oxygen is lower than the tolerance value, the feeding rate is reduced. When the dissolved oxygen is higher than the tolerance value, the feeding rate is increased, so that the dissolved oxygen in the fermentation process is always maintained in a relatively stable range; the sugar-oil ratio = sugar solution volume / oil volume.

2. The fermentation method for producing gibberellic acid with low energy consumption according to claim 1, characterized in that: The feeding method of the supplementary carbon source comprises the following stages: The first stage: acid production reaction until dissolved oxygen rebounds by 50-80%, then sugar solution is added, the initial rate of sugar solution addition is 20-50L / h, and then gradually increased until the sugar solution addition rate reaches 100-200L / h; The second stage: maintain the sugar solution feeding rate, start to feed oil, the initial oil feeding rate is 1-10L / h, and then gradually increase until the oil feeding rate is 30-65L / h; The third stage: feed according to the sugar-oil ratio of 4.0-3.0:1, the dissolved oxygen control point is 40%-50%, when the dissolved oxygen is lower than 40-45%, reduce the feeding speed, when the dissolved oxygen is higher than 45-50%, increase the feeding speed; The fourth stage: feed according to the sugar-oil ratio of 3.5-3.0:1, the dissolved oxygen control point is 30%-40%, when the dissolved oxygen is lower than 30-35%, reduce the feeding speed, when the dissolved oxygen is higher than 35-40%, increase the feeding speed; The fifth stage: feed according to the sugar-oil ratio of 3.0-2.5:1, the dissolved oxygen control point is 20%-30%, when the dissolved oxygen is lower than 20-25%, reduce the feeding speed, when the dissolved oxygen is higher than 25-30%, increase the feeding speed; The sixth stage: feed according to the sugar-oil ratio of 2.5-2.0:1, the dissolved oxygen control point is 15%-25%, when the dissolved oxygen is lower than 15-20%, reduce the feeding speed, when the dissolved oxygen is higher than 20-25%, increase the feeding speed; The seventh stage: feed according to the sugar-oil ratio of 2.0-1.5:1, the dissolved oxygen control point is 10%-20%, when the dissolved oxygen is lower than 10-15%, reduce the feeding speed, when the dissolved oxygen is higher than 15-20%, increase the feeding speed; The eighth stage: feed according to the sugar-oil ratio of 1.5 to 1.0:1, the dissolved oxygen control point is 5% to 15%, when the dissolved oxygen is lower than 5-10%, reduce the feeding speed, when the dissolved oxygen is higher than 10-15%, increase the feeding speed.

3. The fermentation method for producing gibberellic acid with low energy consumption according to any one of claims 1 or 2, characterized in that: The sugar is glucose, and the oil is salad oil.

4. The fermentation method for producing gibberellic acid with low energy consumption according to any one of claims 1 to 3, characterized in that: The production fermentation tank comprises a tank body, on which a feed inlet, a feed outlet, and a feed supplement and air passage are provided, the tank body forms an inner cavity of the tank body, the feed inlet is provided at the top of the tank body, the feed outlet is provided at the bottom of the tank body, the feed supplement and air passage are located at the lower part of the tank body, the feed supplement and air passage are connected with a main pipe, the main pipe passes through the tank body, a part of the main pipe is located in the inner cavity of the tank body, and the rest of the main pipe is located outside the inner cavity of the tank body, the end of the main pipe located outside the inner cavity of the tank body is connected with an air inlet pipe and a feed supplement pipe, a feed supplement valve and a feed supplement cup are installed on the feed supplement pipe, an air inlet valve is installed on the air inlet pipe, an inner cylinder is installed in the inner cavity of the tank body, the top of the inner cylinder is provided with an upper opening, the bottom is provided with a lower opening, the inner cylinder forms an inner cavity of the inner cylinder, the main pipe is located in the tank body Part of the cavity passes through the inner cylinder and is located in the inner cavity of the inner cylinder. Its end opening is downward and is connected to the distributor. A first pushing unit is arranged below the bottom of the distributor. More than three horizontal distribution pipes are connected to the side wall of the distributor. The more than three horizontal distribution pipes are evenly distributed along the circumference. More than two second pushing units are arranged above each horizontal distribution pipe. The first pushing unit is located below the bottom of the inner cylinder, and the second pushing unit is located in the inner cavity of the inner cylinder. The first pushing unit is connected to the distributor through a connecting pipe, and the second pushing unit is connected to the horizontal distribution pipe through a connecting pipe. The pushing inlet of the first pushing unit is at the top and the pushing outlet is at the bottom, and the pushing inlet of the second pushing unit is at the bottom and the pushing outlet is at the top.

5. The fermentation method for producing gibberellic acid with low energy consumption according to claim 4, characterized in that: The supplementary carbon source and ammonia water are added from the feed pipe.

6. The fermentation method for producing gibberellic acid with low energy consumption according to any one of claims 1 to 5, characterized in that: The fermentation broth comprises: Corn gluten meal 10-40 g / L; Potassium dihydrogen phosphate 1~10g / L; Small molecule organic carbon source 5-30g / L; Vegetable oil 0.5-5 g / L; Magnesium sulfate 0.5-2 g / L; Ammonium sulfate 0.5-2 g / L; and Trace elements 0.1~1g / L.

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