Sterilization method and system for large bioreactor

Abstract

The invention relates to the technical field of industrial equipment sterilization, in particular to a sterilization method and system for a large bioreactor. The sterilization method comprises the following steps: carrying out at least one round of rinsing and gaseous sterilization on the large bioreactor to finish sterilization of the large bioreactor, wherein the rinsing time ranges from 30 min to 120 min, and the gaseous sterilization time ranges from 10 min to 60 min; a rinsing solution for rinsing comprises a rinsing solution at a preset temperature, and a sterilization medium for gaseous sterilization comprises sterilization gas; according to the sterilization method, at least one round of rinsing is conducted on the large bioreactor through rinsing liquid with the preset temperature, at least one round of gaseous sterilization is conducted through sterilization gas, infectious microbes in dead corners of the large bioreactor can be effectively killed, and the sterilization method only needs the mode of combining rinsing and gaseous sterilization, so that the sterilization efficiency is improved. The whole sterilization method is simple to operate and has a good sterilization effect on a large bioreactor.

Description

Technical Field

[0001] The present application relates to the technical field of industrial equipment sterilization, and in particular to a sterilization method and system for a large bioreactor. Background Art

[0002] The technology of using CO as a carbon source to ferment alcohols is an advanced carbon utilization and carbon emission reduction technology. This technology uses CO-containing gas produced by steel, metallurgy, petrochemical refining, and agricultural and forestry waste as raw materials, and produces high value-added products such as ethanol through biological fermentation processes. However, in the biological fermentation process, due to the large size of the bioreactor, it is difficult to carry out effective sterilization treatment, which leads to the frequent contamination of these large bioreactors during operation. In the case of the presence of a large number of miscellaneous bacteria in these large bioreactors, the growth and reproduction of these miscellaneous bacteria will utilize a large amount of carbon sources, thereby reducing the production of target alcohol products and causing a great waste of production materials.

[0003] At present, the sterilization technology for large bioreactors is difficult to completely kill bacteria, and most large bioreactors are equipped with ceramic membrane systems. In order to avoid damage to the ceramic membrane system during sterilization, it needs to be disassembled for sterilization, which undoubtedly increases the complexity of the sterilization operation.

[0004] At present, the sterilization technologies for large-scale bioreactors mainly include: (1) Cleaning in Place (CIP) process, the core principle of which is to spray CIP spray balls in devices at different positions in these large-scale bioreactors in advance, and then use CIP alkaline solution with a temperature of 80°C to 85°C and a mass concentration of 2% to 3% to sterilize the large-scale bioreactor; (2) High-pressure and high-temperature steam closed disinfection, the core of which is to use 6bar to 8bar high-temperature steam to seal and sterilize the bioreactor. Summary of the invention

[0005] The present application provides a sterilization method and system for a large bioreactor to solve the following technical problem: how to improve the sterilization effect of a large bioreactor under simple operation.

[0006] In a first aspect, the present application provides a sterilization method for a large bioreactor, the sterilization method comprising:

[0007] subjecting the large-scale bioreactor to at least one round of rinsing and gas sterilization to complete sterilization of the large-scale bioreactor;

[0008] Wherein, the rinsing time is 30min to 120min, and the gas sterilization time is 10min to 120min;

[0009] The rinsing liquid for rinsing includes a rinsing liquid with a preset temperature, and the sterilization medium for gaseous sterilization includes a sterilization gas.

[0010] Optionally, the added volume V1 of the rinse solution and the volume V2 of the large bioreactor satisfy the relationship: V1:V2=(2-10):100.

[0011] Optionally, the preset temperature is 50°C to 90°C.

[0012] Optionally, the weight m1 of the sterilizing gas and the volume V2 of the large bioreactor satisfy the relationship: m1:V2=(10:1)-(200:1), wherein the unit of m1 is mg and the unit of V2 is m 3 .

[0013] Optionally, the sterilizing gas includes ozone and / or chlorine dioxide.

[0014] Optionally, when the sterilizing gas includes ozone, the time of the gaseous sterilization is 20 min to 120 min, and the weight m1 of the sterilizing gas and the volume V1 of the large bioreactor satisfy the relationship: m1:V2=(10:1)~(150:1), wherein the unit of m1 is mg and the unit of V2 is m 3 ; and / or

[0015] In the case where the sterilizing gas includes chlorine dioxide, the time of the gaseous sterilization is 10 minutes to 120 minutes, and the weight m1 of the sterilizing gas and the volume V2 of the large bioreactor satisfy the relationship: m1:V2=(20:1)~(200:1), wherein the unit of m1 is mg and the unit of V2 is m 3 .

[0016] Optionally, the large bioreactor is subjected to at least one round of rinsing and gas sterilization to complete the sterilization of the large bioreactor, comprising the steps of:

[0017] Using a rinsing liquid at a preset temperature to perform a first round of rinsing on the large-scale bioreactor to obtain a primary rinsed bioreactor;

[0018] Using a sterilizing gas to sterilize the bioreactor in a gaseous state to obtain a secondary sterilized bioreactor;

[0019] The secondary sterilization bioreactor is vented, and then the vented secondary sterilization bioreactor is rinsed for a second round using the rinsing liquid at the preset temperature to complete the sterilization of the large bioreactor, wherein the endpoint pH of the second round of rinsing is 8.0-9.0.

[0020] Optionally, the endpoint sterilizing gas concentration of the exhaust gas is less than 10 ppm.

[0021] In a second aspect, the present application provides a sterilization system for a large bioreactor, the sterilization system being adapted to the sterilization method described in the first aspect, the sterilization system being connected to a circulation section of the large bioreactor, and the sterilization system comprising:

[0022] The sterilization treatment part includes a rinsing liquid storage tank, a rinsing liquid delivery pump and a sterilization gas generating device, wherein the liquid outlet of the rinsing liquid storage tank is connected to the liquid inlet of the rinsing liquid delivery pump;

[0023] The control unit includes a metering flow meter and a flow regulating valve. The liquid outlet of the rinse liquid delivery pump is connected to the liquid inlet of the metering flow meter, the liquid outlet of the sterilization gas generating device is connected to the gas inlet of the metering flow meter, the discharge port of the metering flow meter is connected to the feed port of the flow regulating valve, and the discharge port of the flow regulating valve is connected to at least one sterilization feed port of the large bioreactor.

[0024] Optionally, the control unit further includes a sterilizing medium concentration detector, a pH detector, a controller and a discharge valve, the feed port of the discharge valve is connected to the discharge end of the large bioreactor, the sterilizing medium concentration detector and the pH detector are arranged at the discharge end of the large bioreactor, and the sterilizing medium concentration detector and the pH detector are arranged at the feed end of the discharge valve;

[0025] The controller is connected to the metering flow meter, the flow regulating valve, the sterilization medium concentration detector, the pH detector and the discharge valve through electrical signals respectively.

[0026] The above technical solution provided by the embodiment of the present application has the following advantages compared with the prior art:

[0027] The embodiment of the present application provides a sterilization method for a large bioreactor, which uses a rinsing liquid with a preset temperature to perform at least one round of rinsing on the large bioreactor, and the rinsing time is 30 minutes to 120 minutes. The bacteria attached to the large bioreactor are flushed out by rinsing. At the same time, the rinsing liquid introduced by the rinsing can be used as a solvent for the sterilizing gas to increase the contact area between the sterilizing gas and the bacteria, and effectively kill the bacteria in the dead corners of the rinsing of the large bioreactor, thereby improving the sterilization effect of gaseous sterilization; in addition, the sterilizing gas is used for at least one round of gaseous sterilization, and the gaseous sterilization time is 10 minutes to 60 minutes. Through the gaseous sterilization method, the bacteria in the dead corners of the rinsing of the large bioreactor can be effectively killed. In addition, the sterilizing gas can cooperate with the rinsing liquid remaining in the large bioreactor during the rinsing stage to form sterilizing droplets, so as to further effectively kill the bacteria in the dead corners of the rinsing of the large bioreactor; in addition, the sterilization method only requires a combination of rinsing and gaseous sterilization, and the overall sterilization method is simple to operate and has a good sterilization effect on large bioreactors. BRIEF DESCRIPTION OF THE DRAWINGS

[0028] The accompanying drawings, which are incorporated in and constitute a part of this specification, illustrate embodiments consistent with the present application and, together with the description, serve to explain the principles of the present application.

[0029] In order to more clearly illustrate the embodiments of the present application or the technical solutions in the prior art, the drawings required for use in the embodiments or the description of the prior art will be briefly introduced below. Obviously, for ordinary technicians in this field, other drawings can be obtained based on these drawings without paying any creative labor.

[0030] Figure 1 A schematic flow chart of a sterilization method for a large bioreactor provided in an embodiment of the present application;

[0031] Figure 2 A detailed schematic diagram of a sterilization method for a large bioreactor provided in an embodiment of the present application;

[0032] Figure 3 A structural logic diagram of a sterilization system of a large bioreactor provided in an embodiment of the present application;

[0033] Figure 4 A schematic diagram of the structure of a sterilization system of a large bioreactor provided in an embodiment of the present application;

[0034] Among them, 1-large bioreactor, 2-rinsing liquid storage tank, 3-rinsing liquid delivery pump, 4-sterilizing gas generating device, 5-metering flow meter, 6-flow regulating valve, 7-sterilizing medium concentration detector, 8-pH detector, 9-controller, 10-discharge valve. DETAILED DESCRIPTION

[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 making creative work are within the scope of protection of this application.

[0036] Various embodiments of the present application may be presented in the form of a range; it should be understood that the description in the form of a range is only for convenience and brevity and should not be understood as a rigid limitation on the scope of the present application; therefore, it should be considered that the range description has specifically disclosed all possible sub-ranges and single numerical values ​​within the range; for example, it should be considered that the range description from 1 to 6 has specifically disclosed sub-ranges, such as from 1 to 3, from 1 to 4, from 1 to 5, from 2 to 4, from 2 to 6, from 3 to 6, etc., as well as single numbers within the range, such as 1, 2, 3, 4, 5 and 6, which applies regardless of the range; in addition, whenever a numerical range is indicated in this document, it is meant to include any cited numbers (fractions or integers) within the indicated range.

[0037] In this document, the terms including "including" and "including" mean "including but not limited to". Relational terms such as "first" and "second" are only used to distinguish one entity or operation from another entity or operation, and do not necessarily require or imply any such actual relationship or order between these entities or operations. "And / or" describes the association relationship of associated objects, indicating that there can be three relationships. For example, A and / or B can mean: A exists alone, A and B exist at the same time, and B exists alone; where A and B can be singular or plural. "At least one" means one or more, and "plurality" means two or more; "at least one", "at least one of the following" or similar expressions refer to any combination of these items, including any combination of single or plural items; for example, "at least one of a, b, or c", or "at least one of a, b, and c" can all mean: a, b, c, ab (i.e., a and b), ac, bc, or abc, where a, b, and c can be single or multiple. Unless otherwise specified, various raw materials, reagents, instruments and equipment used in this application can be purchased from the market or prepared by existing methods.

[0038] It should be noted that, with respect to the prior art (1) described in the background art, the main processing flow of the cleaning in place (CIP) process in the prior art (1) is: 1) System isolation: stop the inflow and discharge of all materials in the system, and confirm that all isolation valves are in a closed state. Close the isolation valve of the raw gas and the hand valve on the dosing line, and at the same time close the isolation valve on the clear liquid line of the ceramic membrane system; 2) Nitrogen replacement: connect the system to nitrogen, and then start to feed nitrogen. Use the method of increasing and decreasing pressure to replace the gas in the tank until the gas analysis shows that CO is zero; 3) Fermentation liquid emptying: the mash is discharged to the mash tank through the mash pipeline or the membrane circulation pump; 4) System emptying: open all low-point emptying valves to empty all dead corner liquid, and fully empty the membrane filtration and related system pipelines; 5) Pre-rinsing the fermentation tank and ceramic membrane: heat the rinse water to 50°C, open the bioreactor CIP, and allow the liquid to enter the bioreactor. Take samples of the rinse water regularly for observation. If the water is turbid, discharge it into the sewage treatment and introduce fresh rinse water at the same time. The membrane filtration system is cleaned together with the fermentation tank system. After rinsing, the wastewater should be discharged through the bioreactor membrane system circulation pump, and the bioreactor has the CIP cleaning conditions; 5) CIP cleaning: prepare fresh CIP solution (sodium hydroxide with a mass concentration of 2% to 3%) in the CIP tank; start the CIP pump and heat it to 80℃ to 85℃ through heating treatment; introduce CIP liquid into the bioreactor to allow CIP to enter the fermentation tank, and at the same time, the bioreactor establishes a certain liquid level; the CIP cleaning of the membrane filtration system is cleaned together with the fermentation tank system; when the CIP cleaning is completed, the wastewater can be sent to different destinations through the membrane circulation pump; 6) Rinse after CIP cleaning: repeat the pre-rinsing steps of the fermentation tank above. Until it is confirmed that the pH value of the water after rinsing is close to that of clean water (pH value is about 8.0 to 9.0), the pH value of the water stored in the rinse water system must be close to the clean water value before it can be discharged. The inventors found that the prior art (1) has the following defects: 1) The CIP in-situ cleaning system using sodium hydroxide with a mass concentration of 2% to 3% as the medium has limited effect on killing bacteria, and some bacteria cannot be completely killed; 2) For large bioreactors, in-situ CIP cannot achieve immersion killing, and the in-situ CIP system cannot achieve 100% full coverage of blind ends and dead corners. 3) The in-situ CIP system has not yet been able to kill the various medium pipes entering the bioreactor.

[0039] With respect to the prior art (2) described in the background technology, the main process of the closed sterilization in the prior art (2) is: continuously introduce 6bar to 8bar high-temperature steam into the large bioreactor, wait until the high-temperature steam in the large bioreactor reaches a certain pressure and stabilizes, continuously replenish the high-temperature steam until the pressure of the large bioreactor is constant, and then perform high-temperature sterilization for 2h to 3h, and then release the pressure and cool down to complete the sterilization. However, the inventors found that the prior art (2) has the following defects: 1) In the process of using steam sterilization technology, the bioreactor system needs to be sealed, and changes in the external environment, such as rain, snow, etc., will cause the surface temperature of the large bioreactor to drop suddenly, resulting in negative pressure in the bioreactor system and damage to the equipment; 2) The ceramic membrane system cannot achieve steam high-temperature sterilization due to special material limitations.

[0040] Figure 1 The schematic diagram of the sterilization process of a large bioreactor provided in the embodiment of the present application is exemplarily shown;

[0041] like Figure 1 As shown, the present application embodiment provides a sterilization method for a large bioreactor, the sterilization method comprising:

[0042] S1. The large bioreactor is subjected to at least one round of rinsing and gas sterilization to complete the sterilization of the large bioreactor;

[0043] Wherein, the rinsing time is 30min to 120min, and the gas sterilization time is 10min to 120min;

[0044] The rinsing liquid for rinsing includes a rinsing liquid with a preset temperature, and the sterilization medium for gaseous sterilization includes a sterilization gas.

[0045] It should be noted that the rinsing time can be 30 min, 40 min, 50 min, 60 min, 70 min, 80 min, 90 min, 100 min, 110 min or 120 min.

[0046] It should be noted that the time for gaseous sterilization can be 10 min, 20 min, 30 min, 40 min, 50 min, 60 min, 70 min, 80 min, 90 min, 100 min, 110 min or 120 min.

[0047] It should be noted that in order to achieve the best sterilization effect and ensure the safety of large-scale bioreactors, rinsing can be used to flush out residual sterilization gas after gaseous sterilization.

[0048] It should be noted that the sterilization method can be applied not only to conventional areas of large-scale bioreactors, but also to ceramic membrane filtration systems of large-scale bioreactors.

[0049] It should be noted that the sterilization method is based on a large bioreactor with a CIP cleaning system.

[0050] In some optional embodiments, the added volume V1 of the rinse solution and the volume V2 of the large bioreactor satisfy the relationship: V1:V2=(2-10):100;

[0051] In these embodiments, the added volume V1 of the rinsing liquid and the volume V2 of the large bioreactor can satisfy the relationship: V1:V2=(2~10):100, so that there is a sufficient amount of rinsing liquid in the large bioreactor during the rinsing stage, so as to effectively kill and flush out the bacteria attached to the large bioreactor. At the same time, a large amount of droplets can remain after the sufficient amount of rinsing liquid is emptied. These droplets can be used as solvents for the sterilization gas to increase the contact area between the sterilization gas and the bacteria, thereby improving the sterilization effect of gaseous sterilization.

[0052] The added volume V1 of the rinse solution and the volume V2 of the large bioreactor satisfy the relationship: V1:V2=2:100, 3:100, 4:100, 5:100, 6:100, 7:100, 8:100, 9:100 or 10:100.

[0053] In some optional embodiments, the preset temperature is 50°C to 90°C;

[0054] In these embodiments, the preset temperature can be 50°C to 90°C, which can cause the rinsing liquid to have a sufficiently high temperature. The rinsing liquid of sufficient temperature can effectively kill and flush out the miscellaneous bacteria in the large bioreactor. In addition, the residual rinsing liquid of sufficient temperature can quickly dissolve the sterilization gas to increase the contact area between the sterilization gas and the miscellaneous bacteria, thereby improving the sterilization effect of gaseous sterilization.

[0055] It should be noted that the preset temperature can be 50°C, 55°C, 60°C, 65°C, 70°C, 75°C, 80°C, 85°C or 90°C.

[0056] In some optional embodiments, the weight m1 of the sterilizing gas and the volume V2 of the large bioreactor satisfy the relationship: m1:V2=(10:1)-(200:1), wherein the unit of m1 is mg and the unit of V2 is m 3 ;

[0057] In these embodiments, the weight m1 of the sterilizing gas and the volume V2 of the large bioreactor can satisfy the relationship: m1:V2=(10:1)~(200:1), so that the gaseous sterilization stage has sufficient sterilizing gas to effectively kill the miscellaneous bacteria in the dead corners of the large bioreactor during rinsing.

[0058] The weight m1 of the sterilizing gas and the volume V2 of the large bioreactor can satisfy the relationship: m1:V2=10:1, 20:1, 30:1, 40:1, 50:1, 60:1, 70:1, 80:1, 90:1, 100:1, 110:1, 120:1, 130:1, 140:1, 150:1, 160:1, 170:1, 180:1, 190:1 or 200:1.

[0059] In some optional embodiments, the sterilizing gas includes ozone and / or chlorine dioxide;

[0060] In some optional embodiments, when the sterilizing gas includes ozone, the gaseous sterilization time is 20 min to 120 min, and the weight m1 of the sterilizing gas and the volume V1 of the large bioreactor satisfy the relationship: m1:V2=(10:1)~(150:1), wherein the unit of m1 is mg and the unit of V2 is m 3 ; and / or

[0061] In the case where the sterilizing gas includes chlorine dioxide, the time of the gaseous sterilization is 10 minutes to 120 minutes, and the weight m1 of the sterilizing gas and the volume V2 of the large bioreactor satisfy the relationship: m1:V2=(20:1)~(200:1), wherein the unit of m1 is mg and the unit of V2 is m 3 ;

[0062] In these embodiments, when the sterilization gas includes ozone, the gaseous sterilization time can be 20 minutes to 120 minutes, and the weight m1 of the sterilization gas and the volume V1 of the large bioreactor can satisfy the relationship: m1:V2=(10:1)~(150:1); in addition, when the sterilization gas includes chlorine dioxide, the gaseous sterilization time can be 10 minutes to 60 minutes, and the weight m1 of the sterilization gas and the volume V2 of the large bioreactor can satisfy the relationship: m1:V2=(20:1)~(200:1); by using different gaseous sterilization times and different weights of sterilization gases for different types of sterilization gases, gaseous sterilization can be fully carried out to effectively kill the miscellaneous bacteria in the dead corners of rinsing of the large bioreactor.

[0063] When the sterilization gas includes ozone, the time of the gaseous sterilization can be 20 min, 30 min, 40 min, 50 min, 60 min, 70 min, 80 min, 90 min, 100 min, 110 min or 120 min.

[0064] The weight m1 of the sterilizing gas and the volume V1 of the large bioreactor may satisfy the relationship: m1:V2=10:1, 20:1, 30:1, 40:1, 50:1, 60:1, 70:1, 80:1, 90:1, 100:1, 110:1, 120:1, 130:1, 140:1 or 150:1.

[0065] When the sterilizing gas includes chlorine dioxide, the time for the gaseous sterilization can be 10 min, 20 min, 30 min, 40 min, 50 min, 60 min, 70 min, 80 min, 90 min, 100 min, 110 min or 120 min.

[0066] The weight m1 of the sterilizing gas and the volume V1 of the large bioreactor can satisfy the relationship: m1:V2=20:1, 30:1, 40:1, 50:1, 60:1, 70:1, 80:1, 90:1, 100:1, 110:1, 120:1, 130:1, 140:1, 150:1, 160:1, 170:1, 180:1, 190:1 or 200:1.

[0067] Figure 1 A schematic diagram of a sterilization method for a large bioreactor provided in an embodiment of the present application is shown exemplarily;

[0068] In some optional embodiments, such as Figure 2 As shown, the large bioreactor is subjected to at least one round of rinsing and gas sterilization to complete the sterilization of the large bioreactor, comprising the steps of:

[0069] S101. The large bioreactor is subjected to a first round of rinsing with a rinsing liquid at a preset temperature to obtain a primary rinsed bioreactor;

[0070] S102. Using a sterilizing gas to sterilize the bioreactor in a gaseous state to obtain a secondary sterilized bioreactor;

[0071] S103. Exhaust the secondary sterilization bioreactor, and then use the preset temperature rinsing liquid to perform a second round of rinsing on the exhausted secondary sterilization bioreactor to complete the sterilization of the large bioreactor, wherein the endpoint pH of the second round of rinsing is 8.0-9.0.

[0072] In these embodiments, a first round of rinsing is performed using a rinse liquid at a preset temperature, and the bacteria can be preliminarily killed by the rinse liquid at a higher temperature. In addition, a sterilizing gas is used for gaseous sterilization, which can effectively kill bacteria in dead corners of rinsing in a large bioreactor. A second round of rinsing is then performed using a rinse liquid at a preset temperature, and the bacteria residues killed in the bioreactor after the secondary sterilization after exhaust can be flushed out, thereby effectively killing bacteria in the large bioreactor. In addition, the endpoint pH of the second round of rinsing can be 8.0-9.0, which can ensure that the sterilizing gas and other acidic harmful substances are not dissolved in the rinsing liquid after multiple rinsings in the second round, thereby obtaining a low-harm or even harmless discharged rinse liquid.

[0073] The endpoint pH of the second rinse may be 8.0, 8.1, 8.2, 8.3, 8.4, 8.5, 8.6, 8.7, 8.8, 8.9 or 9.0.

[0074] In some optional embodiments, the endpoint sterilizing gas concentration of the exhaust gas is less than 10 ppm;

[0075] In these embodiments, the terminal sterilizing gas concentration of the exhaust gas may be less than 10 ppm, which means that the sterilizing gas in the large bioreactor can be effectively discharged through the exhaust gas, facilitating the subsequent second round of rinsing.

[0076] Figure 3 The structure logic diagram of a sterilization system of a large bioreactor provided in an embodiment of the present application is exemplarily shown;

[0077] Figure 4 The schematic diagram of the structure of a sterilization system of a large bioreactor provided in an embodiment of the present application is exemplarily shown;

[0078] Based on a general inventive concept, such as Figure 3 and Figure 4 As shown, the embodiment of the present application provides a sterilization system for a large bioreactor, the sterilization system is adapted to the sterilization method, the sterilization system is connected to the circulation section of the large bioreactor 1, and the sterilization system includes:

[0079] The sterilization treatment part includes a rinsing liquid storage tank 2, a rinsing liquid delivery pump 3 and a sterilization gas generating device 4, wherein the liquid outlet of the rinsing liquid storage tank 2 is connected to the liquid inlet of the rinsing liquid delivery pump 3;

[0080] The control part includes a metering flowmeter 5 and a flow regulating valve 6. The liquid outlet of the rinse liquid delivery pump 3 is connected to the liquid inlet of the metering flowmeter 5, the liquid outlet of the sterilization gas generating device 4 is connected to the gas inlet of the metering flowmeter 5, the discharge port of the metering flowmeter 5 is connected to the feed port of the flow regulating valve 6, and the discharge port of the flow regulating valve 6 is connected to at least one sterilization feed port of the large bioreactor 1.

[0081] The sterilization system is implemented based on the above-mentioned sterilization method. The specific steps of the sterilization method can refer to the above-mentioned embodiment. Since the sterilization system adopts part or all of the technical solutions of the above-mentioned embodiment, it has at least all the beneficial effects brought by the technical solutions of the above-mentioned embodiment, which will not be repeated here one by one.

[0082] It should be noted that the circulation section of the large bioreactor 1 has multiple liquid inlets to meet the liquid inlet requirements of the rinsing liquid at different stages.

[0083] It should be noted that the metering flow meter 5 and the flow regulating valve 6 cooperate with each other to adjust the added weight of the rinsing liquid and the sterilizing gas.

[0084] In some optional embodiments, the control unit further includes a sterilizing medium concentration detector 7, a pH detector 8, a controller 9 and a discharge valve 10, the feed port of the discharge valve 10 is connected to the discharge end of the large bioreactor 1, the sterilizing medium concentration detector 7 and the pH detector 8 are arranged at the discharge end of the large bioreactor 1, and the sterilizing medium concentration detector 7 and the pH detector 8 are arranged at the feed end of the discharge valve 10;

[0085] The controller 9 is connected to the metering flow meter 5, the flow regulating valve 6, the sterilizing medium concentration detector 7, the pH detector 8 and the discharge valve 10 respectively through electrical signals;

[0086] In these embodiments, the control unit may further include a sterilizing medium concentration detector 7, a pH detector 8, a controller 9 and a discharge valve 10. The controller 9 collects data from the sterilizing medium concentration detector 7 and the metering flowmeter 5, and can adjust the opening and closing degree of the flow regulating valve 6 to adjust the amount of rinse liquid or sterilizing gas added. In addition, the controller 9 collects data from the pH detector 8 to clarify the discharge timing of the discharge valve 10. When the pH measured by the pH detector 8 is 8.0 to 9.0, the discharge valve 10 can discharge low-harm or even harmless rinse liquid.

[0087] It should be noted that the sterilization medium concentration detector 7 can be set at multiple locations in the large bioreactor 1, for example, the sterilization medium concentration detector 7 can be set at the circulation section of the large bioreactor 1, the bottom liquid outlet, and the liquid outlet of the ceramic membrane system outside the large bioreactor 1.

[0088] The present application is further described below in conjunction with specific examples. The experimental methods in the following examples that do not specify specific conditions are usually measured according to industry standards; if there are no corresponding industry standards, they are carried out according to common international standards, conventional conditions, or conditions recommended by the manufacturer.

[0089] Example 1

[0090] like Figure 2 As shown, a sterilization method for a large bioreactor comprises:

[0091] S101. The large bioreactor is rinsed with a rinse solution at a preset temperature for the first round to obtain a primary rinsed bioreactor;

[0092] S102. Using a sterilizing gas to sterilize the bioreactor in a gaseous state to obtain a secondary sterilized bioreactor;

[0093] S103. Exhaust the secondary sterilization bioreactor, and then use a rinsing liquid with a preset temperature to perform a second round of rinsing on the exhausted secondary sterilization bioreactor to complete the sterilization of the large bioreactor, wherein the endpoint pH of the second round of rinsing is 8.0-9.0.

[0094] The first rinse time is 60 minutes, and the second rinse time is 60 minutes.

[0095] The added volume V1 of the rinse solution and the volume V2 of the large bioreactor satisfy the relationship: V1:V2=5:100.

[0096] The preset temperature is 80℃.

[0097] Sterilizing gases include ozone.

[0098] When the sterilizing gas includes ozone, the gaseous sterilization time is 80 minutes. The weight m1 of the sterilizing gas and the volume V1 of the large bioreactor satisfy the relationship: m1:V2=100:1, where the unit of m1 is mg and the unit of V2 is m 3 .

[0099] like Figure 3 As shown, a sterilization system of a large bioreactor, the sterilization system is connected to the circulation section of the large bioreactor 1, and includes:

[0100] The sterilization treatment part includes a rinsing liquid storage tank 2, a rinsing liquid delivery pump 3 and a sterilization gas generating device 4, wherein the liquid outlet of the rinsing liquid storage tank 2 is connected to the liquid inlet of the rinsing liquid delivery pump 3;

[0101] The control part includes a metering flow meter 5, a flow regulating valve 6, a sterilizing medium concentration detector 7, a pH detector 8, a controller 9 and a discharge valve 10. The liquid outlet of the rinsing liquid delivery pump 3 is connected to the liquid inlet of the metering flow meter 5, the liquid outlet of the sterilizing gas generating device 4 is connected to the gas inlet of the metering flow meter 5, the material outlet of the metering flow meter 5 is connected to the material inlet of the flow regulating valve 6, and the material outlet of the flow regulating valve 6 is connected to at least one sterilizing material inlet of the large bioreactor 1;

[0102] The sterilizing medium concentration detector 7 and the pH detector 8 are arranged at the discharge end of the large bioreactor 1, and the sterilizing medium concentration detector 7 and the pH detector 8 are arranged at the feed end of the discharge valve 10;

[0103] The controller 9 is connected to the metering flow meter 5, the flow regulating valve 6, the sterilizing medium concentration detector 7, the pH detector 8 and the discharge valve 10 through electrical signals.

[0104] Example 2

[0105] Based on the contents disclosed in Example 1, the following modifications are further made:

[0106] The first rinsing time is 30 minutes, and the second rinsing time is 120 minutes.

[0107] The added volume V1 of the rinse solution and the volume V2 of the large bioreactor satisfy the relationship: V1:V2=2:100.

[0108] The preset temperature is 60℃.

[0109] When the sterilizing gas includes ozone, the gaseous sterilization time is 120 minutes. The weight m1 of the sterilizing gas and the volume V1 of the large bioreactor satisfy the relationship: m1:V2=10:1, where the unit of m1 is mg and the unit of V2 is m 3 .

[0110] Example 3

[0111] Based on the contents disclosed in Example 1, the following modifications are further made:

[0112] The first rinsing time is 100 minutes, and the second rinsing time is 50 minutes.

[0113] The added volume V1 of the rinse solution and the volume V2 of the large bioreactor satisfy the relationship: V1:V2=10:100.

[0114] The preset temperature is 90℃.

[0115] When the sterilizing gas includes ozone, the gaseous sterilization time is 20 minutes. The weight m1 of the sterilizing gas and the volume V1 of the large bioreactor satisfy the relationship: m1:V2=150:1, where the unit of m1 is mg and the unit of V2 is m 3 .

[0116] Example 4

[0117] Based on the contents disclosed in Example 1, the following modifications are further made:

[0118] Sterilizing gases include chlorine dioxide.

[0119] When the sterilizing gas includes chlorine dioxide, the gaseous sterilization time is 20 minutes, and the weight m1 of the sterilizing gas and the volume V2 of the large bioreactor satisfy the relationship: m1:V2=100:1, where the unit of m1 is mg and the unit of V2 is m 3 .

[0120] Comparative Example 1

[0121] Based on the contents disclosed in Example 1, the following modifications are further made:

[0122] No sterilizing gas is used.

[0123] Comparative Example 2

[0124] Based on the contents disclosed in Example 1, the following modifications are further made:

[0125] No second rinse cycle is used.

[0126] Comparative Example 3

[0127] Based on the contents disclosed in Example 1, the following modifications are further made:

[0128] The time for the first rinsing round is 10 minutes, the time for the second rinsing round is 10 minutes, and the time for gas sterilization is 5 minutes.

[0129] Comparative Example 4

[0130] Based on the contents disclosed in Example 1, the following modifications are further made:

[0131] The time for the first rinsing round is 200 minutes, the time for the second rinsing round is 200 minutes, and the time for gas sterilization is 120 minutes.

[0132] Comparative Example 5

[0133] Based on the contents disclosed in Example 1, the following modifications are further made:

[0134] The preset temperature is 35℃.

[0135] Comparative Example 6

[0136] Based on the contents disclosed in Example 1, the following modifications are further made:

[0137] The preset temperature is 100°C.

[0138] Related experiments and effect data:

[0139] The sterilization methods disclosed in the embodiments and comparative examples were used to treat multiple groups of large bioreactors 1 or ceramic membrane systems outside thereof under the same fermentation conditions, and the rinse water from the final second round of rinsing was collected. The rinse water was inoculated into purified water with a total colony count of ≤1 cfu / g at the same inoculum amount, and the total colony count after 72 hours was counted. The results are shown in Table 1.

[0140] Table 1 The total number of colonies after inoculation and cultivation of rinse water treated in each embodiment and comparative example

[0141]

[0142]

[0143] As can be seen from Table 1, the sterilization method for a large bioreactor provided in the embodiment of the present application can effectively reduce the number of miscellaneous bacteria in the large bioreactor 1 or the ceramic membrane system outside the large bioreactor 1 to 20 cfu / g, which shows that the sterilization method has a good sterilization effect. In addition, it can be seen from Comparative Examples 4 and 6 that although the sterilization effects of the sterilization methods disclosed in Comparative Examples 4 and 6 are close to those of the embodiments, Comparative Example 4 uses a longer rinsing time and gaseous sterilization time, which undoubtedly increases the operating cycle of the equipment, thereby increasing the energy consumption of the overall sterilization method; and Comparative Example 6 uses a higher preset temperature rinse liquid, which will increase the heat loss of the equipment of the overall sterilization system, thereby increasing the energy consumption of the overall sterilization method.

[0144] In summary, the embodiment of the present application provides a sterilization method for a large bioreactor, which only requires a combination of rinsing and gaseous sterilization. The overall sterilization method is simple to operate and has a good sterilization effect on large bioreactors.

[0145] In addition, the sterilization method for a large bioreactor provided in the embodiment of the present application has a good sterilization effect, can reduce the waste of carbon resources caused by contaminating bacteria in the fermentation stage of the large bioreactor, and can improve the operating efficiency of the large bioreactor and the corresponding fermentation product yield.

[0146] In addition, the sterilization method of a large bioreactor provided in the embodiment of the present application can fully sterilize the ceramic membrane of the large bioreactor to reduce the risk of contamination of the large bioreactor caused by incomplete sterilization of the ceramic membrane.

[0147] In addition, a sterilization method for a large bioreactor provided in an embodiment of the present application can perform fumigation sterilization on a large bioreactor without dead ends through sterilizing gases such as ozone or chlorine dioxide gas, and will not affect the target bacteria in the large bioreactor.

[0148] The above is only a specific implementation of the present application, so that those skilled in the art can understand or implement the present application. It will be apparent to those skilled in the art that various modifications to these embodiments are possible, and the general principles defined in the present application can be implemented in other embodiments without departing from the spirit or scope of the present application. Therefore, the present application will not be limited to these embodiments shown in the present application, but will conform to the widest scope consistent with the principles and novel features applied for by the present application.

Claims

1. A method for sterilizing a large bioreactor, characterized in that: The sterilization method comprises: subjecting the large-scale bioreactor to at least one round of rinsing and gas sterilization to complete sterilization of the large-scale bioreactor; Wherein, the rinsing time is 30min to 120min, and the gas sterilization time is 10min to 120min; The rinsing liquid for rinsing includes a rinsing liquid with a preset temperature, and the sterilization medium for gaseous sterilization includes a sterilization gas.

2. The sterilization method according to claim 1, characterized in that: The added volume V1 of the rinse liquid and the volume V2 of the large bioreactor satisfy the relationship: V1:V2=(2-10):

100.

3. The sterilization method according to claim 1, characterized in that: The preset temperature is 50°C to 90°C.

4. The sterilization method according to claim 1, characterized in that: The weight m1 of the sterilizing gas and the volume V2 of the large bioreactor satisfy the relationship: m1:V2=(10:1)-(200:1), where the unit of m1 is mg and the unit of V2 is m 3 .

5. The sterilization method according to claim 1, characterized in that: The sterilizing gas includes ozone and / or chlorine dioxide.

6. The sterilization method according to claim 5, characterized in that: When the sterilizing gas includes ozone, the time of the gaseous sterilization is 20 minutes to 120 minutes, and the weight m1 of the sterilizing gas and the volume V1 of the large bioreactor satisfy the relationship: m1:V2=(10:1)~(150:1), wherein the unit of m1 is mg and the unit of V2 is m 3 ; and / or In the case where the sterilizing gas includes chlorine dioxide, the time of the gaseous sterilization is 10 minutes to 120 minutes, and the weight m1 of the sterilizing gas and the volume V2 of the large bioreactor satisfy the relationship: m1:V2=(20:1)~(200:1), wherein the unit of m1 is mg and the unit of V2 is m 3 .

7. The sterilization method according to claim 1, characterized in that: The large-scale bioreactor is subjected to at least one round of rinsing and gas sterilization to complete the sterilization of the large-scale bioreactor, comprising the steps of: Using a rinsing liquid at a preset temperature to perform a first round of rinsing on the large-scale bioreactor to obtain a primary rinsed bioreactor; Using a sterilizing gas to sterilize the bioreactor in a gaseous state to obtain a secondary sterilized bioreactor; The secondary sterilization bioreactor is vented, and then the vented secondary sterilization bioreactor is rinsed for a second round using the rinsing liquid at the preset temperature to complete the sterilization of the large bioreactor, wherein the endpoint pH of the second round of rinsing is 8.0-9.

0.

8. The sterilization method according to claim 7, characterized in that: The endpoint sterilizing gas concentration of the exhaust gas is less than 10 ppm.

9. A sterilization system for a large bioreactor, characterized in that: The sterilization system is adapted to the sterilization method according to any one of claims 1 to 7, the sterilization system is connected to the circulation section of the large bioreactor, and the sterilization system comprises: The sterilization system is connected to the circulation section of the large bioreactor (1), and the sterilization system comprises: The sterilization treatment section comprises a rinsing liquid storage tank (2), a rinsing liquid delivery pump (3) and a sterilization gas generating device (4), wherein the liquid outlet of the rinsing liquid storage tank (2) is connected to the liquid inlet of the rinsing liquid delivery pump (3); The control unit includes a metering flow meter (5) and a flow regulating valve (6); the liquid outlet of the rinse liquid delivery pump (3) is connected to the liquid inlet of the metering flow meter (5); the liquid outlet of the sterilizing gas generating device (4) is connected to the gas inlet of the metering flow meter (5); the material outlet of the metering flow meter (5) is connected to the material inlet of the flow regulating valve (6); and the material outlet of the flow regulating valve (6) is connected to at least one sterilization material inlet of the large bioreactor (1).

10. The sterilization system according to claim 9, characterized in that: The control unit further comprises a sterilizing medium concentration detector (7), a pH detector (8), a controller (9) and a discharge valve (10); the feed port of the discharge valve (10) is connected to the discharge end of the large bioreactor (1); the sterilizing medium concentration detector (7) and the pH detector (8) are arranged at the discharge end of the large bioreactor (1); and the sterilizing medium concentration detector (7) and the pH detector (8) are arranged at the feed end of the discharge valve (10); The controller (9) is connected to the metering flow meter (5), the flow regulating valve (6), the sterilizing medium concentration detector (7), the pH detector (8) and the discharge valve (10) respectively through electrical signals.

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