Method for improving measurement accuracy of microorganism heat production in fermentation vessel

FIELD: medicine.

SUBSTANCE: method involves measuring heat production by variation of heat rate consumed by maintenance of the isothermal mode of a fermentation vessel with intermitted fluid and gas flows through the fermentation vessel with adjusted heat power consumption by mixing the culture fluid. It is combined with eliminating heat power consumption by heat production in the fermentation vessel of the culture fluid mass due to maintaining its initial value at the pre-set accuracy in the environment of the conducted continuous and periodic cultivation processes by equation of energy gain, consumed for each heating cycle of the fermentation vessel, the energy gain value derived by calibration heating of the fermentation vessel filled with the initial culture fluid mass with the heat production value and the maintenance of the culture fluid mass during the microorganism cultivation process are spread out over a period of time.

EFFECT: increased measurement accuracy of continuous and periodic microorganism heat production in the fermentation vessel.

2 cl, 3 dwg

 

The technical field

This invention can be used in biotechnology, biochemistry and technical Microbiology.

Study of the growth and metabolism of microorganisms in the development of biotechnological processes for cleaning the environment, processes, microbiological synthesis of antibiotics, enzymes and to solve other problems is an actively developing area of research in these areas of science and technology. Calorimetry supplements used in these studies, electrochemical and optical methods, being non-specific, non-invasive and highly sensitive method. When it is determined calorimetric method, the time dependence of thermal power (Pt)accompanying these processes, allows to make a conclusion regarding the growth rate, the occurrence of limiting conditions and to provide information about the formation of metabolites [Winkelmann, M., Hüttl, R., and Wolf, G. Application of batch calorimetry for the investigation of microbial activity // Thermochimica Acta. - 2004. - Vol.415 - R-82].

All quantitative calculations in the cultivation of the microorganisms is carried out with the use of accurate data on mass culture liquid (QL) in a fermentation vessel when carrying out these processes [Perth SC. Fundamentals of cultivation of microorganisms and cells. - M.: Mir, 1978. - 332].

The level of technology

Known analog, pozvoliaushie the heat production of the microorganisms, - reaction calorimeter RC1 (Mettler Toledo, Switzerland). This device does not allow for variable thermal contribution in the Pt mixer with a long fermentation process that makes it impossible to use in the main areas of use of fermentation technology [U.Stockar and I.Marison. The use of calorimetry in biotechnology, Advances in Biochemical Engineering / Biotechnology, 1989, vol.40, p.95-136].

Known method, which provides a measurement of thermal power required to maintain the desired temperature of the fermenter under isothermal conditions [Kotelnikov G.V., Moiseyeva S.P. and Krayev V.P. Calorimetric method for adjusting the mass of culture fluid in a bioreactor // Review of Scientific Instruments, 1998, vol.69, p.2137-2140]. When the measured power includes power for heating the bioreactor under isothermal conditions, thermal power, input mixing device, and the heat production from microorganisms. In this method the measured heat capacity depends on the heat of fermentation vessel with the external environment and the magnitude of heat contribution from the mixing of the culture fluid in a fermentation vessel, which cannot be used for measurements of heat production from microorganisms.

The closest proposed to the technical essence and the number of matching characteristics is the way of measuring in a fermentation vessel heat production microorganism is in continuous and batch processes [patent RU No. 2391410, IPC C12Q 3/00]. This method is carried out in a fermentation apparatus which is not provided precision maintaining the setpoint mass QOL when carrying out fermentation processes accompanied by changes aeration, mixing and foaming.

This leads to uncontrolled changes in a fermentation vessel mass QOL and the corresponding change in biomass that generates heat production, which leads to changes in thermal power required to maintain isothermal mode of fermentation vessel. Thus, thermal contribution leads to the error of measurement of heat production, as measuring the heat production of the microorganisms is maintained only by changing the heat output required to maintain isothermal mode of fermentation vessel.

This method does not resolve the contribution to the heat production heat capacity arising from uncontrolled changes in a fermentation vessel mass culture fluid, resulting in an error in the measurement of heat production to tens of percent.

Thus, known methods of measuring heat production microorganisms do not provide the necessary accuracy of measurements of this parameter with change in a fermentation vessel mass culture liquid in the conditions of provodimyh continuous and periodic processes of cultivation.

The objective of the invention is to improve the accuracy of measurements of heat production of microorganisms by eliminating the contribution to the heat production of heat capacity changes in a fermentation vessel mass culture fluid by maintaining its original value with a given accuracy in the conditions of ongoing continuous and periodic processes of cultivation.

Disclosure of inventions

The task is implemented by the claimed method to improve the accuracy of measurements of heat production of microorganisms in a fermentation vessel, which consists in measuring the heat production by the change of heat capacity required to maintain isothermal mode of fermentation vessel at stop streams of liquid and gaseous media through the fermentation vessel, with the introduction of amendments to the contribution of thermal power from the mixing of the culture fluid, thus eliminating the contribution to the heat production of heat capacity changes in a fermentation vessel mass culture fluid by maintaining its original value with a given accuracy in the conditions of ongoing continuous and periodic processes of cultivation on equal increments of energy expended for each heat of fermentation vessel in the process of maintaining the setpoint mass of cultural the second fluid, the incremental value of energy generated by the calibration of the heat of fermentation vessel, filled with the original mass of the culture fluid, the operation of determining heat production and maintain mass QOL in a fermentation vessel in the conditions of ongoing continuous and periodic processes of cultivation of microorganisms spaced in time.

The task is implemented also due to the fact that, according to the proposed method, maintaining the mass culture liquid in the process of cultivation of microorganisms by changing the ducts liquid media through the fermentation vessel.

The task of increasing the accuracy of measurement of heat production by microorganisms is solved in the known fermentation apparatus according to patent RU No. 2391410, which were also the software implementation of the controller mass QOL for implementing the inventive method.

Brief description of drawings

Figure 1 shows the structural diagram of a fermentation apparatus for carrying out the invention. Fermentation apparatus according to figure 1 contains a fermentation vessel 1 with a mixing device 2 connected pneumohydraulic 3 communications to the Executive device 4 equipped with sensors measuring parameters 5 and the heater 6, the connection is local electric utilities 7 with the host computer 8 through the negotiation 9, through which the host computer 8 electric utilities connected actuators 4, wherein the fermentation vessel 1 is installed in a managed thermostatic screen 10 and is provided with an additional mixing device 11 to provide temperature control of fermentation vessel 1 when changing modes of mixing device 2 in the process of measuring the contribution of the heat capacity of the heat production of microorganisms, however, communications are 3 and 7, running in a fermentation vessel 1 are thermal contact with a controlled heating and cooling of the screen 10. Managed thermostat 12 is connected to a managed thermostatic screen 10. The sensors are connected through the negotiation 9 to the input multi-channel analog-to-digital Converter (ADC) 15, which is included in the module Lab-PC+(16)built into the control computer 8. Through the specified device coordination 9 actuators 4 of fermentation vessel 1: Sciclone and Ncseaa connected to the outputs of digital to analog converters CAP and CAP included in the module 16; Sclive and Nset. environment connected to the outputs of the digital-to-analog converters CAP and CAP included in the module Lab-PC+ (17); Clandest, Klodiana and Cloddy of antifoam connected to the outputs: Logvin, Logwin and Logwin; drive PU and drive pudup, the heater is connected to the outputs: schetchik, schetchik, schetchik module PC-TIO-10 (18).

Figure 2 presents curves measured heat capacity over time for various amounts of mass QOL in the fermenter when it software support within a given time interval (tn).

The implementation of the invention

According to figure 2, curve 1 corresponds to the initial mass value QL (heat energy corresponds to the space ACE); curve 2 corresponds to the mass value QL exceeding the value of the original value (the heat energy corresponds to the square ABE); curve 3 corresponds to the value of the mass of QOL, the value of which is less than the original value (the heat energy corresponds to the area of ADE). From these curves it follows that thermal energy curve for 2 more, and for curve 3 is less than the energy for the curve 1. This allows you to use the deviation of thermal energy (area ABC and ACD) at different values of mass QOL in a fermentation vessel to calculate the amount drained QOL to maintain the original value that provides a measurement without having to reach a steady-state values of the power in the heating process and takes a period of time of about 10 sec. It imposes no restrictions on obtaining accurate values of the measured heat production, maintaining the mass QOL and execution is piracy to eliminate thermal contribution mixer conducted in a fermentation vessel fermentation processes.

Figure 3 presents a time chart illustrating an example of a separation in time of measurement of heat production with known measurement of thermal contribution of the mixer in the signal heat production and removal of thermal contribution to the change in mass of QOL in the heat production according to the claimed method, where

ta- interval measuring heat production with a duration of 20 s;

tc- interval measurement of thermal contribution of the mixer with a duration of 20 s;

teinterval eliminate thermal contribution of the mass change of QOL in heat production with a duration of 20 s;

tb, td, tf- time intervals, which is a fermentation process at a given temperature without stopping ducts liquid and gaseous media through the fermentation vessel. On the interval tfcan be advanced by the operator measuring the heat production at intervals similar to the ta;

tgthe time interval of the cycle.

In real processes, the time intervals on the chart can be changed up or down, as the prior art allows you to do this.

A lot of QOL in a fermentation vessel can be maintained by addition or subtraction of its largest energy consumed per unit mass of QOL in a fermentation vessel, i.e. the change of ducts liquid media through the farm is operating the vessel.

When sizing heating the calibration factor is defined as

Δm is the number of merged in a fermentation vessel fluid during calibration;

ΔQ is the energy spent on heating Δm.

The mass is drained from the fermentation vessel liquid mislevis calculated by the formula (2):

where Qn- the energy increase in software support fermentation vessel with a specified number of QOL during the time interval set duration; QNTECthe increment of energy when heating the fermentation vessel in the second and subsequent cycles of heating in the regulatory process mass QOL. Drain QOL is a pump with a rated capacity signal from the control computer.

If (Qn-QNTEC)>0 discharge is not conducted because the number of QOL in a fermentation vessel is less than the specified value. By dispensing in a fermentation vessel nutrients and other components (Qn-QNTEC) changes the sign that entails drain current calculated values of mislev. Maintaining weight QOL is carried out cyclically. However, thanks to the computer control fluid source and the drain of QOL are equal in magnitude. This allows you to adjust the weight of QOL in the fermentation with the court with time intervals of about one hour, because during this time, the accumulation of errors of the metering devices. Calorimetric method provides the necessary accuracy of the mass QOL in a fermentation vessel with an error of about 10 g, which, in comparison with a method of maintaining weight QOL in a fermentation vessel at a level ten times better.

Fermentation apparatus used in the measurement of heat production by the claimed method of ensuring the maintenance of the initial value of mass of QOL, works as follows.

All systems measurement and control in this fermentation apparatus functioning in a normal mode, for obtaining data of figure 1 through the temperature controller fermentation vessel in accordance with the program. The temperature of fermentation vessel 1 is regulated by the temperature sensor signal _t, a signal which is normalized in the device approval 9 and fed to one of the inputs to a multi-channel ADC, which is included in the module Lab-PC+ (16)embedded in the control computer 8. Formed in the managing computer pulse-width signal (PWM) output "Scetchy" module PC-TIO-10 (18)embedded in the computer 8, is fed through a matching device 9 in the heater 6 of fermentation vessel 1, providing automatic temperature regulation in accordance with sastavci, set on the computer 8, the software controller mass QOL, part of the software 19 fermentation apparatus based on LabView, provides the following functions:

software and change the setpoint of the temperature controller fermentation vessel during the heat of fermentation vessel for a fixed interval of time and the cooling of the fermentation vessel to the initial temperature;

- collecting and summing the digital PWM temperature controller during the programme warming;

- calculate the amount of drained fluid and the formation of the control signal to the pump discharge.

These functions are performed as follows.

When the heat source weight of QOL regulator mass QOL calculates the value of Qnwhen subsequent heatings in the same manner calculated QNTEC. For calculation of the calibration coefficient of the fermentation vessel with an initial value of mass QOL merges a known amount of mass QOL Δm, runs the specified program warm-up for the determination of the energy spent on heating Δm, according to the formula ΔQ=(Qn-QNTEC), and calculates the calibration factor Kcaliberby the formula (1). After each of the current warm-up, conducted in cycles, calculated mass is drained from the fermentation vessel liquid is t m islevby the formula (2). Drain QOL is a pump for a fixed time with the changing performance of the control signal output "CAP" module Lab-PC+. The algorithm of operation of the regulator mass QOL involves turning off the pump after draining the mislevan entry in the "CAP" zero.

The regulator mass QOL provides equality Qnand QNTECduring the whole fermentation process, which eliminates the contribution to the heat production of the heat capacity from the mass change of QOL in a fermentation vessel in the conditions of ongoing continuous and periodic processes of cultivation of microorganisms. Define Qnand QNTECposted in time with the operations of measurement and calibration of heat production and are provided with an error less than 0.1%, and the integral control system mass QOL ensures zero error maintain mass QOL. Thus, almost completely eliminates the contribution of the heat capacity of the heat production from changes in a fermentation vessel mass QOL.

Implementation of the claimed method in a fermentation equipment allows researchers to measure the heat production in the ongoing long fermentation processes simple and reliable means.

1. The way to improve the accuracy of measurements of heat production of microorganisms in a fermentation vessel, the conclusion is audica in the measurement of heat production by the change of heat capacity, expended on maintaining the isothermal mode of fermentation vessel at stop streams of liquid and gaseous media through the fermentation vessel, with the introduction of amendments to the contribution of thermal power from the mixing of the culture fluid, wherein eliminate the contribution to the heat production of heat capacity changes in a fermentation vessel mass culture fluid by maintaining its original value with a given accuracy in the conditions of ongoing continuous and periodic processes of cultivation on equal increments of energy expended for each heat of fermentation vessel in the process of maintaining a given value of mass culture fluid, the increment value of the energy obtained during the calibration of the heat of fermentation vessel filled with an initial mass culture fluid while the definition of heat production and maintenance of mass culture fluid within the ongoing process of cultivation of microorganisms spaced in time.

2. The method according to claim 1, characterized in that the maintaining of mass culture liquid in the process of cultivation of microorganisms by changing the ducts liquid media through the fermentation vessel.



 

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