KR101459022B1 - Semicontinuous cultivation system for photo organism and the method thereof - Google Patents

Abstract

The present invention relates to a semi-continuous culture system for photobiological culture and a method of culturing the same. More specifically, the present invention provides two reactors to initially grow photobioreactors in a primary reactor, and when the initial growth is completed, The remainder is transferred to the second reactor and the inducer medium is supplied to the second reactor so that the remainder of the growth is completed in the second reactor. In the first reactor, the growth medium is supplied as the amount of the culture solution, To a semi-continuous culture system for photobiological culture and a culture method thereof.

Description

TECHNICAL FIELD [0001] The present invention relates to a semi-continuous culture system for photobiological culture,

The present invention relates to a semi-continuous culture system for photobiological culture and a method of culturing the same. More specifically, the present invention provides two reactors to initially grow photobioreactors in a primary reactor, and when the initial growth is completed, The remainder is transferred to the second reactor and the inducer medium is supplied to the second reactor so that the remainder of the growth is completed in the second reactor. In the first reactor, the growth medium is supplied as the amount of the culture solution, To a semi-continuous culture system for photobiological culture and a culture method thereof.

Global warming caused by greenhouse gas emissions caused by the use of fossil fuels is causing climate change and global environmental change, threatening the survival of all life forms, including humans. Therefore, many researches and developments have been made to reduce carbon dioxide, and as a part thereof, researches on the field of biochemical conversion of carbon dioxide have been actively carried out.

As a photosynthetic organism for biological conversion treatment of carbon dioxide, microalgae have been actively studied. Like other photosynthetic creatures, microalgae, which are phytoplankton, act as the energy source of the sun, and have the property of growing by carrying out a photosynthesis action to immobilize carbon dioxide.

The reason why microalgae are attracted by the means of immobilizing carbon dioxide is that the energy required to recover carbon dioxide is very small because the plant can utilize solar energy as a main energy source just as it absorbs carbon dioxide. Therefore, since the amount of carbon dioxide generated by the operation of the carbon dioxide immobilization process is small, the removal efficiency is high in view of the carbon dioxide resin.

Second, it has a very low CO2 footprint due to its high CO2 immobilization rate. According to a study by Tokyo Electric Power Research Institute, the rate of CO2 fixation of microalgae was 2.8 times higher than that of the fastest growing sugarcane and 15 times higher than that of the most common species of pine trees in Korea.

In addition, since carbon dioxide can be directly immobilized from the combustion gas, there is an advantage that a separation and concentration process for carbon dioxide is not necessary. In addition, the microalgae produced during the carbon dioxide immobilization process contain various useful substances and can be used as biological products.

However, when the process of immobilizing carbon dioxide using microalgae is carried out by a bioreactor applied in a practical industry, the supply of light energy necessary for energy saving and microalgae growth is not easy due to high consumption of electric energy There is a problem.

In general, a photosynthetic biological culture apparatus for the purpose of immobilization of carbon dioxide can be divided into an open type culture apparatus for large-scale outdoor cultivation and a closed-type photosynthetic bioreactor having a small volume. In the case of open outdoor mass culture apparatus, it was mainly used in Germany, Japan, USA, etc. in the form of a lake or a large pond.

However, such a pond-type open outdoor mass culture device is expensive and requires a large amount of energy to be consumed during continuous stirring, and it is difficult to prevent contamination and to separate / refine cultured microalgae. In addition, in the case of a mass culture apparatus, since the effective light transmission is not generally performed to the inside, the growth rate of the photosynthetic organism is slow and the growth yield is low.

The closed-type photosynthetic bioreactor currently developed includes an agitated reactor, a plate reactor, a tubular reactor, a column reactor, and a polymer film reactor.

However, the culture method using such a reactor has a problem in that when cultivating a photobiological organ showing cell growth and secondary metabolite induction, productivity is low due to a long initial induction period in a batch culture or a fed-batch culture And the content of the secondary metabolites contained in the recovered organisms is unstable in the continuous culture.

Korean Patent Publication No. 10-2011-0085428 Korean Patent Registration No. 10-0490641

SUMMARY OF THE INVENTION The present invention has been accomplished in order to solve the above-mentioned problems, and it is an object of the present invention to provide a method for preparing a bioreactor, which comprises two reactors to initially grow photobioregions in a first reactor, And at the same time, an inducer medium is supplied to the second reactor to complete the remainder of the growth in the second reactor. In the first reactor, the growth medium is supplied as much as the amount of the culture medium to be continuously operated, Can improve the productivity and can provide an environment suitable for the induction unit depending on the purpose in the secondary reactor so that the amount of conversion to the objective product can be increased and productivity can be improved so that commercialization through mass culture is possible A semi-continuous culture system for biological culture and a culture method thereof are provided. .

It is another object of the present invention to provide a semi-continuous culture system for photobiological culture and a method for culturing the same, which can operate a smaller number of reactors than a conventional batch culture and thus reduce operating costs due to a reduced reactor consumption.

According to an aspect of the present invention,

A first reactor in which the inoculated organisms are firstly grown using the culture medium stored therein and the carbon dioxide introduced into the culture medium; A second reactor in which when the primary growth of the photobiestheses is completed from the primary reactor and is discharged together with the culture liquid, the secondary reactor is supplied with the same and the secondary growth of the photobiestheses is performed using the carbon dioxide introduced into the primary reactor; A storage tank for storing a culture solution in which the secondary growth of the photobiological organism is completed in the secondary reactor; A growth reactor culture tank for supplying a growth reactor culture medium, which is a culture solution, to the primary reactor; An inducer culture tank for supplying an induction culture medium as a culture medium to the second reactor; A culture liquid transfer pump installed in association with the primary reactor, the secondary reactor, and the storage tank and rotating in the normal or reverse direction according to an external operation to transfer the culture liquid; And feeding the culture liquid to the secondary reactor by controlling the culture liquid transfer pump when the primary growth of the primary reactor is completed and controlling the culture liquid transfer pump when the secondary growth in the secondary reactor is completed, And a controller for discharging the refrigerant to the storage tank.

Here, a culture liquid transfer line is provided between the first reactor and the second reactor so as to be spaced apart from the inner bottom surface of the first reactor so as to be transferred to the second reactor while remaining a part of the culture liquid of the first reactor. And is installed to the inner bottom surface of the secondary reactor so as to transfer all the culture liquid of the secondary reactor to the storage tank.

Here, the culture liquid remaining in the primary reactor is 10 to 50% by volume of the total culture liquid.

Here again, the primary reactor is grown inoculation only at the initial time of the photobioreactor, and subsequently grows continuously in the culture medium.

Here, the first reactor is provided with a nitrogen sensor to measure the nitrogen concentration of the culture liquid.

Here, the second reactor is provided with an optical sensor for measuring the optical density of the culture liquid.

In this case, the controller measures the nitrogen concentration of the culture liquid through the nitrogen sensor of the first reactor. When the nitrogen concentration becomes equal to the preset concentration, the culture liquid transfer pump is operated to transfer a part of the culture liquid to the second reactor When the increase in the optical density of the culture solution becomes equal to the predetermined increase amount through the photosensor of the secondary reactor, the culture solution transfer pump is operated to discharge the culture solution to the storage tank.

Here, the controller may further include a step of partially discharging the culture fluid of the first reactor to the second reactor, then re-supplying the growth medium of the grower discharge tank to the first reactor by the discharge amount, After discharging to the storage tank, the inducer medium of the induction device discharge tank is re-supplied to the secondary reactor by the discharge amount.

Here, the growth medium of the growth medium and the inductor medium of the inductor medium tank may be the same or different from each other.

Here, the first reactor and the second reactor are any one of an agitation type reactor, a plate type reactor, a tubular reactor, a column type reactor, and a polymer film type reactor having the same capacity.

According to another aspect of the present invention,

When the nitrogen concentration in the culture solution of the first reactor is measured and the nitrogen concentration becomes equal to the predetermined concentration, the culture solution is added to the culture solution To a secondary reactor; A continuous primary growth step in which the growth medium is continuously fed to the primary reactor by the amount of the culture medium discharge when the transfer of the culture liquid from the primary reactor to the secondary reactor is completed; When a culture medium containing the first-grown photobioreactor of the first reactor is supplied to the second reactor, a predetermined amount of the inducer medium is supplied to the inside of the second reactor by the remaining amount of the culture medium of the first reactor, A secondary growth step for secondary growth; And a discharging step of measuring the optical density of the culture solution of the secondary reactor and discharging the culture solution when the increase amount of the optical density becomes equal to the predetermined increase amount.

Here, the culture liquid remaining in the primary reactor is 10 to 50% by volume of the total culture liquid.

Herein, the growth medium of the grower culture tank and the culture medium of the inducer of the induction medium culture tank may be the same or different from each other.

Here, the first reactor and the second reactor are any one of an agitation type reactor, a plate type reactor, a tubular reactor, a column type reactor, and a polymer film type reactor having the same capacity.

According to the semi-continuous culture system for photobiological culture of the present invention and the method for culturing the same, two reactors are provided to initially grow the photobioregions in the first reactor, and when the initial growth is completed, The remainder is transferred to the second reactor and the inducer medium is supplied to the second reactor so that the remainder of the growth is completed in the second reactor. In the first reactor, the growth medium is supplied as the amount of the culture solution, It is possible to improve the productivity as a whole by relatively reducing the incubation period and to provide an environment suitable for the induction machine depending on the purpose in the second reactor so that the conversion amount to the target product can be increased and the productivity can be improved Commercialization through mass culture can be achieved.

Further, according to the present invention, since a smaller number of reactors can be operated than conventional batch culture, the operation cost can be reduced because the reactor consumption is small.

BRIEF DESCRIPTION OF THE DRAWINGS FIG. 1 is a schematic diagram showing a configuration of a semi-continuous culture system for photobiological culture according to the present invention; FIG.
FIG. 2 is a process diagram for explaining a semi-continuous culture method for culturing an organism according to the present invention.
3 is a graph comparing the number of cells and incubation period of a semi-continuous culture system for photobiological culture according to the present invention and a batch type culture apparatus (batch) as a comparative example.
FIG. 4 is a graph comparing the growth curves of a semi-continuous culture system (semi-continuous) for photobiological culture according to the present invention and a batch type culture apparatus (comparative example).
FIG. 5 is a graph comparing the productivity of fatty acids converted into biofuels by a semi-continuous culture system for photobiological culture according to the present invention and a batch type culture apparatus (comparative example).

DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS Hereinafter, the construction of a semi-continuous culture system for photobiological culture according to the present invention will be described in detail with reference to the accompanying drawings.

In the following description of the present invention, detailed description of known functions and configurations incorporated herein will be omitted when it may make the subject matter of the present invention rather unclear. The following terms are defined in consideration of the functions of the present invention, and these may be changed according to the intention of the user, the operator, or the like. Therefore, the definition should be based on the contents throughout this specification.

BRIEF DESCRIPTION OF THE DRAWINGS FIG. 1 is a schematic diagram showing a configuration of a semi-continuous culture system for photobiological culture according to the present invention; FIG.

1, a semi-continuous culture system 100 for culturing a bio-organism according to the present invention comprises a primary reactor 110, a secondary reactor 120, a culture liquid transfer line 130, A growth medium discharge tank 150, an induction machine discharge tank 160, a culture liquid transfer pump 170,

First, the first reactor 110 grows the inoculated organism first by using the culture medium stored therein and carbon dioxide introduced into the inside. Here, the primary reactor 110 grows continuously after being inoculated only at the initial one time, and then remains in the culture medium. Here, the first reactor 110 is provided with a nitrogen sensor 111 for measuring the nitrogen concentration of the culture liquid to confirm whether the first growth is possible. Here, the first reactor 110 is any one of an agitating reactor, a plate reactor, a tubular reactor, a column reactor, and a polymer film reactor.

When the primary growth of the photobioregion is completed and the secondary reactor 120 is discharged together with the culture solution from the primary reactor 110, the photoreactor is secondarily grown using the carbon dioxide injected into the secondary reactor 120. Here, the second reactor 120 is provided with an optical sensor 121 for measuring the optical density of the culture liquid to confirm whether or not the second growth occurs. Here, the second reactor 120 may be any one of an agitating reactor, a plate reactor, a tubular reactor, a column reactor, and a polymer film reactor having the same capacity as that of the first reactor 110, The same reactor as the first reactor 110 is used.

The culture liquid transfer line 130 is installed between the first reactor 110 and the second reactor 120. One side of the culture solution transfer line 130 is transferred to the second reactor 120 while remaining a part of the culture solution of the first reactor 110. [ And the other side of the second reactor 120 is connected to the inside of the second reactor 120 so as to transfer all of the culture solution of the second reactor 120 to the storage tank 140 to be described later. It is installed to the bottom. In this case, the culture solution remaining in the first reactor 110 by the culture solution transfer line 130 is preferably 10 to 50 volume of the total culture solution so that the first- %.

In addition, the storage tank 140 stores the culture liquid in which the secondary growth of the photobioregings is completed in the secondary reactor 120.

In addition, the growth medium tank 150 supplies the growth medium 151, which is a culture medium, to the first reactor 110.

On the other hand, the inducer feed tank 160 supplies the inducer feed medium 161, which is a culture medium, to the second reactor 120. Here, the inducer culture medium 161 of the inducer culture medium tank 160 is composed of the same or different components as the growth medium of the growth medium culture tank 150.

The culture liquid transfer pump 170 is installed in association with the first reactor 110, the second reactor 120, and the storage tank 140, and is rotated in the forward and reverse directions according to the external operation to transfer the culture liquid. At this time, the culture liquid transfer pump 170 is preferably a decompression pump.

The controller 180 controls the culture liquid transfer pump 170 to supply a part of the culture liquid to the second reactor 120 and to supply the second liquid to the second reactor 120 in the second reactor 120. [ When the tea growth is completed, the culture liquid transfer pump 170 is controlled to discharge the culture liquid to the storage tank 140. Here, the controller 180 measures the nitrogen concentration of the culture liquid through the nitrogen sensor 111 of the first reactor 110 and operates the culture liquid transfer pump 170 when the nitrogen concentration becomes equal to the predetermined concentration, Is transferred to the secondary reactor 120 and the amount of increase in the optical density of the culture liquid through the photosensor 121 of the secondary reactor 120 becomes equal to a predetermined increase amount, the culture liquid transfer pump 170 is operated to store the culture liquid And discharges them all to the tank 140. Hereinafter, the controller 180 may further include a step of partially discharging the culture fluid of the first reactor 110 to the second reactor 120, and then discharging the growth medium 151 of the grower discharge tank 150 to the first reactor 110 The culture medium of the second reactor 120 is discharged to the storage tank 140 and then the inducer medium 161 of the inducer discharge tank 160 is re-supplied to the second reactor 120 by the discharge amount . At this time, it is preferable that the predetermined nitrogen concentration is "0 ", and the increase amount of the optical density is" 0 ".

On the other hand, the semi-continuous culture system 100 for photobiological culture according to the present invention comprises an electromagnetic valve (V) for varying the flow path under the control of the controller (180) on the piping line connecting the respective components, a flow meter ), And the like, and may further include a pump as needed.

Hereinafter, a semi-continuous culture method for culturing an organism according to the present invention will be described in detail with reference to the accompanying drawings.

FIG. 2 is a process diagram for explaining a semi-continuous culture method for culturing an organism according to the present invention.

Referring to FIG. 2, the semi-continuous culture method for cultivation of an organism according to the present invention includes a primary growth step, a continuous primary growth step, a secondary growth step and a discharge step.

"Primary Growth Process-S100"

The controller 180 supplies a certain amount of the growth medium 151 of the growth medium discharge tank 150 to the first reactor 110 while the administrator inoculates the photobiore into the first reactor 110, First, grow.

The controller 180 measures the nitrogen concentration of the culture liquid in the first reactor 110 in real time and controls the culture liquid transfer pump 170 when the nitrogen concentration "0 " 130) to the second reactor (120).

&Quot; Continuous primary growth step-S110 "

When the transfer of the culture liquid from the first reactor 110 to the second reactor 120 is completed, the controller 180 causes the first reactor 110 to regenerate the growth medium 151 of the growth medium discharge tank 150 And then grows the photobioregions continuously. Then, the controller 180 measures the nitrogen concentration of the culture liquid in the first reactor 110 in real time and controls the culture liquid transfer pump 170 when the nitrogen concentration "0 " ) To the second reactor (120).

&Quot; Secondary growth process-S120 "

The controller 180 supplies the culture solution containing the first grown cultivar of the first reactor 110 to the second reactor 120 by the amount of the culture solution remaining in the first reactor 110, The inducer culture medium 161 of the inducer culture tank 160 is supplied to the inside of the induction device culture tank 160, and carbon dioxide is supplied to grow the culture medium.

&Quot; Drain process-S130 "

In this state, the controller 180 measures the optical density of the culture liquid in the second reactor 120 in real time, and if the optical density value is no longer increased after the secondary growth is completed, the controller 180 controls the culture liquid transfer pump 170, The culture solution of the secondary reactor 120 is discharged to the storage tank 140 through the transfer line 130 and stored.

&Quot; Example &

As a first reactor, a second reactor, and a comparative reactor of the present invention, a polymer film type photobioreactor (the polymer film type photobioreactor disclosed in Korean Patent No. 10-1148194 and a description thereof is omitted) was used.

Neochloris oleoabundans # 1185 (The University of Texas at Austin) was used as a photobioreactor . Neoclose Oligo abundance is a species of freshwater species microalgae, which has the difficult conditions to cultivate among photosynthetic organisms, but it has a high photosynthetic efficiency and lipid-rich characteristics, and is a species capable of producing the next generation biofuels.

Neo-chloris oligo abundance was cultured in TAP-C medium for growth period and TAP-N medium was used for induction period. The components of the TAP-C medium are shown in Table 1 below, and the components of the TAP-N medium are shown in Table 2 below.

The neoclose oligubendans were cultured under the conditions shown in Table 3 below, and cultured under the same conditions until the light density, the gas flow rate, the gas concentration, and the distribution of the medium to avoid the influence of other conditions.

FIG. 3 is a graph comparing cell numbers and incubation periods of a semi-continuous culture system (semi-continuous) for photobiological culture according to the present invention and a batch type culture apparatus (comparative example) A graph comparing the growth curves of the semi-continuous culture system for photobiological culture (semi-cons.) And the batch culture apparatus (batch) for comparative example.

In this embodiment, 70% of the microalgae were transferred to the induction reactor, and the remaining 30% of the growth reactor was fed with 1.43-fold concentration of fresh growth medium and 1-L of concentrated induction medium concentrated 15 times in the induction machine.

In this example, a part of samples were taken at each measurement, and the number of cells per unit volume of microalgae was measured to calculate the total number of cells. The Y-axis values are expressed as natural logarithm values. As shown in FIG. 3, the total number of cells in the batch culture was about 20.82 as a natural logarithmic value, and about 21.82 in the semi-continuous culture, indicating that about 1.9 million,000 cells were proliferated.

In contrast to the batch culture, in the semi-continuous culture system 100 for photobiological culture according to the present invention, 5% of carbon dioxide used as a carbon source was reduced by 2.3% after passing through a growth culture incubator, Was removed.

In this example, a portion of each measurement was taken for fatty acid production and the dry weight and fatty acid content of the microalgae were measured.

FIG. 5 is a graph comparing the productivity of fatty acids converted into biofuels by a semi-continuous culture system for photobiological culture according to the present invention and a batch type culture apparatus (comparative example).

This result also by calculating the productivity between the two fatty acid groups culture through 5, the batch 13.413 mgL ^-1 d ^-1, in semi-continuous culture was found to be 22.685 mgL ^-1 d ^-1.

While the present invention has been particularly shown and described with reference to exemplary embodiments thereof, it is to be understood that the invention is not limited to the disclosed exemplary embodiments. It is to be understood, however, that the invention is not to be limited to the specific forms thereof, which are to be considered as being limited to the specific embodiments, but on the contrary, the intention is to cover all modifications, equivalents, and alternatives falling within the spirit and scope of the invention as defined by the appended claims. .

110: primary reactor 120: secondary reactor
130: Culture medium transfer line 140: Storage tank
150: Growth medium discharge tank 160: Induction machine discharge tank
170: Culture medium transfer pump 180: Controller

Claims (14)

The inoculated organisms are firstly grown using the culture medium stored therein and the carbon dioxide introduced into the culture medium, but the organisms are inoculated only once at the initial stage, and then continuously grown in the culture medium. A primary reactor in which 50% by volume of culture solution remains;
A second reactor in which when the primary growth of the photobiestheses is completed from the primary reactor and is discharged together with the culture liquid, the secondary reactor is supplied with the same and the secondary growth of the photobiestheses is performed using the carbon dioxide introduced into the primary reactor;
A storage tank for storing a culture solution in which the secondary growth of the photobiological organism is completed in the secondary reactor;
A growth reactor culture tank for supplying a growth reactor culture medium, which is a culture solution, to the primary reactor;
An inducer culture tank for supplying an induction culture medium as a culture medium to the second reactor;
A culture liquid transfer pump installed in association with the primary reactor, the secondary reactor, and the storage tank and rotated in the forward and reverse directions according to an external operation to transfer the culture liquid;
The second reactor is installed so as to be spaced apart from the inner bottom surface of the first reactor so as to be transferred to the second reactor while remaining a part of the culture solution of the first reactor and to transfer all of the culture solution of the second reactor to the storage tank, A culture liquid transfer line installed up to an inner bottom surface of the reactor; And
The nitrogen concentration of the culture liquid is measured through the nitrogen sensor of the first reactor. When the nitrogen concentration becomes equal to the predetermined concentration, the culture liquid transfer pump is operated to transfer a part of the culture liquid to the secondary reactor, When the increase amount of the optical density of the culture liquid becomes equal to the predetermined increase amount through the sensor, the culture liquid transfer pump is operated to discharge the culture liquid to the storage tank, the culture liquid of the first reactor is partially discharged to the second reactor, The culture medium of the second reactor is discharged to the storage tank and then the culture medium of the inducer of the induction medium discharge tank is supplied to the secondary reactor as much as the discharge amount of the second reactor Wherein the controller comprises a controller for supplying the culture medium to the culture medium. delete delete delete The method according to claim 1,
The first reactor may comprise:
A semi-continuous culture system for photobiological culture, characterized in that a nitrogen sensor is provided to measure the nitrogen concentration of the culture. 6. The method of claim 5,
The second reactor may comprise:
A semi-continuous culture system for photobiological culture, characterized in that an optical sensor is provided to measure the optical density of the culture. delete delete The method according to claim 1,
The grower medium of the growth medium tank and the inducer medium of the induction medium medium tank,
Wherein the culture medium is composed of the same or different components. The method according to claim 1,
The first reactor and the second reactor may be,
Continuous culture system for photobiological culture, characterized in that it is any one of an agitation type reactor, a plate type reactor, a tubular reactor, a column type reactor and a polymer film type reactor having the same capacity. The first reactor is supplied with a certain amount of a growth medium, followed by inoculation with the organism, carbon dioxide is added to the first reactor, and the nitrogen concentration of the culture solution in the first reactor is measured. When the nitrogen concentration becomes equal to the preset concentration A primary growth step of retaining 10 to 50% by volume of the total culture liquid in the primary reactor and transferring the remainder to the secondary reactor;
A continuous primary growth step in which the growth medium is continuously fed to the primary reactor by the amount of the culture medium discharge when the transfer of the culture liquid from the primary reactor to the secondary reactor is completed;
When a culture medium containing the first-grown photobioreactor of the first reactor is supplied to the second reactor, a predetermined amount of the inducer medium is supplied to the inside of the second reactor by the remaining amount of the culture medium of the first reactor, A secondary growth step for secondary growth; And
And a discharging step of discharging and storing the culture liquid when the optical density of the culture liquid of the secondary reactor is measured and the increase amount of the optical density becomes equal to the predetermined increase amount. delete 12. The method of claim 11,
The grower medium of the growth medium tank and the inducer medium of the induction medium medium tank,
Wherein the culture medium is composed of the same or different components. 12. The method of claim 11,
The first reactor and the second reactor may be,
A semi-continuous culture method for photobiological culture, characterized in that it is any one of an agitated reactor having the same capacity, a plate reactor, a tubular reactor, a column reactor, and a polymer film reactor.

Images