JPH11243946A - Mass-culture of photosynthetic bacteria - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide a method for the pure culture of photosynthetic bacteria in a mass capable of stably performing the pure culture of photosynthetic bacteria in a mass at a relatively low cost without using a pressure vessel and a boiler. SOLUTION: A liquid medium is poured into a closeable vessel made of a fiber-reinforced plastic(FRP). The liquid medium in the vessel is intermittently sterilized by boiling under agitation and a seed of photosynthetic bacteria is charged into the liquid medium in the vessel. The vessel is closed and the liquid medium containing the seed is cultured under agitation. The photosynthesis can be carried out in high efficiency without using an observation window or a light source since an FRP vessel has a light-transmitting property. The sundry germs in the medium are sterilized by the intermittent sterilization to enable the culture of photosynthetic bacteria.

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

TECHNICAL FIELD The present invention relates to a method for culturing a large amount of photosynthetic bacteria.

[0002]

2. Description of the Related Art As a method for purely cultivating only photosynthetic bacteria in a large amount, a method using a pressure-resistant container such as a fermentation tank is most common. In this method, first, a liquid medium is put into a container body of a pressure-resistant container having a double-wall structure for about 7 to 8 minutes, and stirring and aeration are performed. Then, live steam is blown from below the tank together with the air, and the inside of the container body is maintained at 120 to 130 ° C. and 1.5 to 2 atm.
The heating and pressurizing state is continued for about 40 to 60 minutes to sterilize the inside of the container body containing the liquid medium. Next, cooling water is injected between the inner and outer walls of the pressure-resistant container to cool the liquid medium, and then a seed bacterium is introduced. The inside of the container body is maintained at about 30 ° C., and the culture is started while performing aeration and stirring. After about 4 days to 1 week, the culture is completed.

[0003]

SUMMARY OF THE INVENTION In the case of a conventional culture method,
Expensive equipment such as pressure-resistant containers, boilers, and coolers are required. Since the temperature of the surrounding environment of the culture device rises, appropriate ventilation equipment is also required. In addition, a professional operator who handles pressure vessels and boilers must be stationed. As a result, running costs as well as initial investment tend to increase.

The pressure-resistant container is made of a material (for example,
Stainless steel SUS304 Because it does not have translucency, culture
To cause or promote photosynthesis by illuminating bacteria
Is provided with a pressure-resistant floodlight window on the tank wall
Alternatively, the light source must be arranged in the container body.

An object of the present invention is to provide a method for large-scale pure cultivation of photosynthetic bacteria, which can stably carry out large-scale pure culture of photosynthetic bacteria relatively inexpensively without using a pressure-resistant container or a boiler. is there.

[0006] Another object of the present invention is to provide a method for large-scale pure culture of photosynthetic bacteria, which enables efficient photosynthesis without forming a light-projecting window or disposing a light source in the apparatus main body. is there.

It is a further object of the present invention to provide a method for large-scale pure cultivation of photosynthetic bacteria which can be carried out without the need for an expert manager of equipment.

[0008]

The present invention has the following configuration to achieve the above object. That is, in the method of the present invention, first, a liquid medium is poured into a sealable container molded of fiber reinforced plastic (FRP). Next, after the liquid medium in the container is stirred and intermittently sterilized by heating and boiling, a seed of a photosynthetic bacterium is put into the container in the liquid medium. Then, the container is sealed, and the culture is performed while stirring the liquid medium containing the inoculum. The photosynthetic bacterium to be cultured is, for example, Rhodobacter capsulata.

[0009] The container is provided with an inlet for the liquid medium and the inoculum at the upper part, and an outlet at the lower part, and each of the ports is formed so as to be sealable. The liquid medium is injected into the container for about 8 to 9 minutes. Heat sterilization employs intermittent sterilization in which heating boiling and standing cooling are repeated. The intermittent sterilization is preferably repeated about three cycles with heating and boiling and cooling as one cycle. It is desirable that stirring during intermittent sterilization be performed continuously, and stirring after introduction of the inoculum be performed intermittently. In addition, if the heating is performed by a heater heating means disposed in the container, it is easy to control.

[0010]

DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS The present invention will be described below in detail based on illustrated embodiments. FIG. 1 is a block diagram showing steps of a method according to one embodiment of the present invention. First, a liquid medium is prepared. The composition of the liquid medium is appropriately selected according to the type of photosynthetic bacteria to be cultured. This liquid medium is poured into the container 1 as a culture tank for about 8 to 9 minutes.

The container is made of FRP (fiber reinforced plastic)
And has a cylindrical shape without a window as shown in FIG. F
Since the RP is easy to process, for example, a container having a capacity of about 300 liters to about 10 t can be manufactured relatively inexpensively. A lid 2 is attached to the upper part of the container so as to be freely opened and closed. The lid 2 is provided with a sealable exhaust port 3 and an inoculation port 4. Although not shown, the exhaust port 3 is provided with a filter material capable of passing steam and preventing passage of various bacteria as much as possible.

At the bottom of the container, a sheath heater 5 for heating and boiling the liquid medium is provided. Seeds heater 5
Is turned ON / OFF by the control circuit 5a. A plate heater may be used instead of the sheath heater. Also,
Ventilation and stirring blades 6 are provided in the container. The shaft 6a of the stirring blade 6 is connected to a motor 7 on the lid. The motor 7 is timely driven by a timer circuit (not shown), and rotates the stirring blade 6 via the shaft 6a to stir the contents of the container. Shaft 6a leading to stirring blade
Has a hollow inside to allow forced ventilation from outside the container. A filter for capturing various bacteria is provided in an air passage leading to the shaft 6a. The body of the container 1 is covered with a jacket 8 made of FRP for forcibly cooling the contents. The cooling jacket 8 has a cooling water inlet 8a at a lower part and a cooling water outlet 8b at an upper part.

The liquid medium filled in the container is supplied to the exhaust port 3
Sterilized intermittently with the open. Intermittent sterilization is performed by repeated heating and boiling. The heating of the liquid medium is performed by operating the sheath heater 5 described above. To give a specific example, first, the liquid medium is reduced to 1 by the sheath heater 5.
Heat to 00 ° C. and boil for about 3 hours. Since the steam accompanying the boiling is discharged out of the vessel through the exhaust port 3,
The pressure inside the container does not increase. After heating and boiling for a predetermined time,
The heater 5 is turned off once. At the same time, water flows in from the inlet 8a of the jacket 8 and circulates through the jacket to discharge the absorbed heat from the outlet 8b. Thereby, the liquid medium in the container is rapidly cooled. The liquid medium rapidly cooled to about 30 ° C. is kept at a constant temperature overnight while maintaining the vicinity of the temperature (about 30 ° C.). The motor 7 is continuously driven during a series of steps of heating and cooling. The liquid culture medium is constantly stirred by the stirring blades 6 receiving the driving force of the motor, and is aerated by the gas flowing in from the lower end of the shaft. Note that the cooling means may be means for circulating cooling water in a pipe arranged in the container.

After standing at a constant temperature, boiling with a heater, rapid cooling, and standing at a constant temperature are further repeated once, and thereafter, re-boiling and re-cooling are performed by a heater. By the first boiling, the germinated bacteria contained in the liquid medium are killed. During the first incubation, the spore-form bacteria contained in the liquid medium germinate. The germinated bacteria are similarly killed by the second boil. Even if bacteria that did not germinate in the first incubation may germinate in the second incubation, the bacteria die in the third boiling. By repeating boiling and standing twice, the liquid medium can be sterilized as much as possible. However, it is desirable to perform three intermittent sterilizations just in case.

A seed medium is introduced into the liquid medium after sterilization.
The amount of the inoculum is in the range of 1 to 20% of the liquid medium,
% Is desirable. After the introduction of the inoculum, the culture is performed while the stirring blade 6 is intermittently rotated. Stirring is 50-100 rpm
It suffices to run for about 10 minutes a day at a rotation speed of. Frequent agitation is preferably used because dissolved oxygen in the contents of the container (mixture of photosynthetic bacteria and liquid medium) tends to cause growth of bacteria other than photosynthetic bacteria. In addition, depending on the type of photosynthetic bacteria, nitrogen instead of air is ventilated from the shaft 6a.

The culturing period is about 5 to 10 days. During this culture period, the photosynthetic bacteria in the container contents grow preferentially because the liquid medium is sterilized as described above. Moreover, since the container is molded by FRP,
Unless you choose a pigment with poor translucency for the pigment,
Light is applied to the contents of the container, and photosynthesis by the photosynthetic bacteria is more actively and efficiently performed.

Examples Hereinafter, photosynthetic bacteria (
_{Rhodobacter capsulata})
Show. In a 10-ton culture tank, add sodium propionate 0.1%.
1%, monopotassium phosphate 0.5%, dipotassium phosphate
0.06%, ammonium sulfate 0.1%, magnesium sulfate
Heptahydrate 0.02%, sodium chloride 0.02%
%, Calcium chloride dihydrate 0.05%, Bacto
Paste extract (_Difco) 0.01% vitamin solution (H
Amine hydrochloride 50mg, niacin 50mg, para-amino
30 mg of benzoic acid, 10 mg of pyridoxyhydrochloride, and pichi
ON 5mg dissolved in distilled water 100ml) 1ml
/_liter, Trace element solution (EDTA-2Na 1000m
g, FeCl2 6H2 O 2000mg, ZnCl2  1
00 mg, MnCl2 ・ 4H2 O 100mg, H3 BO3
  100 mg, CoCl2 ・ 2H2 O 100 mg, Na
Two MoO4 ・ 2H2 O 20mg, CuCl2 ・ 2H2 O
10 mg, NiCl2 ・ 6H2 O10mg and Na2 Se
O3  5m dissolved in 1 liter of distilled water) 1m
l /^liter9000 liters of medium containing
Intermittent sterilization at 3 ° C. for 3 hours was repeated three times. Medium at 30 ° C
After cooling to another culture tank^{Rh
odopbacter capsulata}3g wet weight /^literSeed concentration 9
00^literWas aseptically inoculated. 10 minutes at 100 rpm
After stirring, the stirring was stopped and culture was performed. Stir for one day
Perform 10 minutes each, culture for 10 days, and cultivate 34 kg wet weight bacteria.
I got a body.

[0018]

According to the present invention, the conditions necessary for the growth of photosynthetic bacteria in a liquid culture medium are adjusted by performing intermittent sterilization by heating and boiling and standing at a constant temperature using a container made of FRP. Large-scale pure culture of photosynthetic bacteria can be performed stably at low cost without using equipment such as a pressure-resistant container and a boiler.

Further, according to the present invention, since the container is made of FRP having translucency, the photosynthetic bacteria in the container can be efficiently photosynthesized without providing a light-projecting window or a light source in the container. be able to.

Further, according to the present invention, a large amount of photosynthetic bacteria can be easily and purely cultured without the need for an expert manager of equipment such as a pressure vessel and a boiler.

[Brief description of the drawings]

FIG. 1 is a block diagram showing steps of a method according to one embodiment of the present invention.

FIG. 2 is a front view showing a container used in the method of FIG. 1 in a partial cross section.

[Explanation of symbols]

 DESCRIPTION OF SYMBOLS 1 ... Container 3 ... Discharge port 4 ... Inoculation port 5 ... Heater 6 ... Stirring blade 8 ... ·Jacket

 ──────────────────────────────────────────────────続 き Continuing from the front page (72) Inventor Tatsuharu Kobayashi 2904 Shinjuku, Oaza, Matsumoto, Nagano Pref.

Claims (7)

[Claims] 1. A liquid medium is poured into a sealable container molded of fiber reinforced plastic (FRP), and the liquid medium in the container is stirred and intermittently sterilized by heating and boiling. A mass culture method for photosynthetic bacteria, in which a bacterial inoculum is charged, the container is closed, and the liquid medium containing the inoculum is cultured with stirring. 2. The method according to claim 1, wherein the photosynthetic bacterium is Rhodobacter capsulata. 3. The method according to claim 1, wherein the liquid medium is injected into the container for 8 to 9 minutes. 4. The method according to claim 1, wherein the stirring during the intermittent sterilization is continuous stirring. 5. The method according to claim 1, wherein said intermittent sterilization comprises three cycles of heating boiling and cooling with heating boiling and standing cooling as one cycle. 6. The method according to claim 1, wherein the stirring during the culture is intermittent stirring. 7. The method according to claim 1, wherein said heating is performed by heater heating means.