JPS59187784A - Production of hydrogen using clostridium butyricum miyairi - Google Patents

Abstract

PURPOSE:To produce a large amount of hydrogen using Clostridium butyricum Miyairi, by adjusting the dilution ratio in the respective plural tanks, and multiplying only vegetative cells in the tanks. CONSTITUTION:Batchwise cultivation is started in tanks while feeding CO2 from feed ports 7 in the first tanks 2 and the second tank 3. On the arrival at the logarithmic growth phase, a culture medium is fed from a culture medium feeding tank 1 through a pump 8 and a transfer pipe 9 to the first tank 2. A culture fluid is fed from the first tank 2 through a pump 10 and a transfer pipe 11 to the second tank 3 at the same time, and a culture fluid is transferred from the second tank 3 through a pump 12 and a transfer pipe 13 and released to start the continuous cultivation. At this point of time, the feed of CO2 is stopped to change over a cock 14 to the side of a collection pipe 15 for collecting fermented H2 gas. CO2 contained in the H2 gas is passed through an absorption tank 4 filled with a KOH solution and absorbed therein. The resultant H2 gas after removing the CO2 is introduced into an H2-water replacing tank 5.

Description

[Detailed description of the invention] The present invention relates to a method for producing hydrogen using Miyairi bacteria.

-1-,.

Hydrogen is attracting attention as an alternative energy source to replace fossil fuels. It is also used as a chemical raw material for various purposes such as ammonia synthesis and petroleum hydrogenation.

Currently, hydrogen-producing bacteria include hydrogen bacteria, photosynthetic bacteria, and C1.
The genus ostridium is known. Hydrogen bacteria require a gas mixture of oxygen, hydrogen, and carbon dioxide during cultivation, and photosynthetic bacteria require light. Therefore, the culture operation is extremely complicated.

On the other hand, in the case of the genus C1ostridium, during batch culture before continuous culture, it is necessary to perform stirring to completely mix the culture medium and bacteria, and to supply carbon dioxide gas to maintain an anaerobic state. However, during the subsequent cultivation stage, sufficient anaerobic conditions can be maintained by the carbon dioxide produced by the plant itself. Therefore, when collecting hydrogen, there is no need to remove the above-mentioned supply gas, and the culture apparatus can be simplified.

In the present invention, Cl of the genus C1ostridium
ostridium'Il"l
”-butyricum M■YAIf (J (hereinafter referred to as Miyairi bacteria) is used. Miyairi bacteria produce hydrogen as a metabolic product during vegetative cells, so in order to produce hydrogen, only vegetative cells should be grown as much as possible. However, since Miyairi bacteria are spore-forming bacteria, they cannot be kept in the state of vegetative cells for a long time, and spores are formed. It is difficult to make time hydrogen production take place.

As a result of research on methods for obtaining large amounts of hydrogen, the present inventors found that it is necessary to adopt a continuous culture method in order to continuously and sufficiently proliferate vegetative cells. obtained knowledge that it is suitable for obtaining
Based on this, we have arrived at the present invention.

That is, the gist of the present invention is to provide a hydrogen production method using Miyairi bacteria, which uses multiple tanks and adjusts various dilution rates so that only vegetative cells grow as much as possible in the tanks. be.

The Miyairi bacteria targeted by the present invention are as follows.

Bacterial name: Clostridium buty
ricum m1yairi Microorganism Depositary 2 Japan, Agency of Industrial Science and Technology, Institute of Microbial Technology, Microbial Accession Number: No. 1467. The dilution rate used in the present invention is the supply per hour relative to the amount of liquid (β) in the tank. It refers to the amount (4/hr).

For example, when 2 parts of liquid is injected into a tank with a liquid volume of 2β every hour and 28 parts of liquid are discharged every hour, the dilution rate is 1.0/hr. Furthermore, if 1A of liquid is injected into a tank with a liquid volume of 2p per hour, and a liquid of [β] is discharged every hour, the dilution rate is 0.5/hr.
It is.

Miyairi bacteria are spore-forming bacteria, and spore formation is caused by vegetative cells →
This occurs in the order of endophytic cells → mature spores, and hydrogen is produced as a metabolic product during vegetative cells.

In batch culture, spore formation begins over time, and the period of vegetative cells that produce hydrogen is short, so it is extremely inefficient as a method of producing hydrogen.

On the other hand, with continuous culture, a steady state can be maintained by adjusting the dilution rate of the first and second tanks and repeating division by freezing the vegetative cells in each tank. Thereby, hydrogen gas production can be increased.

In addition, Miyairi bacteria can be grown up to 3 to 4 x 108Cel in batch culture.
Although it is possible to obtain only the idle number of bacterial cells of 1/d, in continuous culture, by appropriately selecting the dilution rate of the first tank, 1. Cells of OX 1.09Cel I can be obtained. Therefore, continuous culture can increase hydrogen production.

In the present invention, in order to increase the hydrogen production amount of Miyairi bacteria, continuous culture is performed and the dilution ratio of the first tank and second tank is appropriately selected, thereby culturing Miyairi bacteria in a steady state with many vegetative cells. This led to continuous and efficient production of hydrogen as a metabolite.

A continuous culture device for hydrogen production has, for example, two fermenters. In the first tank, culture corresponding to the mid-logarithmic growth phase of the batch culture is carried out, and in the second tank, culture corresponding to the late logarithmic growth phase is carried out, and in order to prevent spore formation from starting within the residence time in these tanks, Select the dilution rate for each tank. Miyairi bacteria are
In batch culture, spores begin to form 7 to 8 hours after the start of growth, so the sum of the residence times in the first and second tanks is:
Must be within 7-8 hours.

The medium used in this method contains sugars and amino acids.

As a result of examining inorganic salts, etc., we found cornstarch, potato starch as the sugar source, meat decomposition product, pegtone, casamino acid, and an amino acid mixture obtained by hydrolyzing defatted soybean as the amino acid, and Mn 804 as the inorganic salt.
, , Fe 804 and Ca CO 3 , and the above cornstarch, amino acid mixture, and Ca CO 3 are particularly suitable for the growth of vegetative cells.

When using the above medium, the dilution rate of the first tank is 0, 15/h.
r ~0.7/hr, but 0.3~0.6
/hr is preferred, and a dilution rate of 0.15/hr or less may initiate spore formation. Moreover, at 0.7/hr or more, washing out of Miyairi bacteria occurs. Furthermore, the dilution rate of the second tank is determined by the dilution rate of the first tank.

What is shown in the drawings is an apparatus for carrying out the present invention, and the present invention will be explained in more detail with reference to this.

Without stirring, batch culture in the first tank 2 and second tank 3 is started while stirring the insides of both tanks. During this batch culture, carbon dioxide gas is supplied to the first tank and the second tank 3 from the button and carbon dioxide gas supply lower to maintain anaerobic conditions.

Next, when the culture in the first tank 2 and the second tank 3 reaches the logarithmic growth phase, the medium is supplied from the medium supply tank 1 to the first tank 2 through the medium transfer pipe 9 by the pump 8. 1
The culture solution is transferred from the tank 2 to the second tank 3 through the pump 10 and the culture solution transfer tube. Furthermore, the pump 12 from the second tank 3
．． Continuous culture is started by discharging the culture solution through the culture solution transfer tube 13.

The supply of carbon dioxide gas is stopped when continuous culture is started as described above. At that time, the cock 14 is switched to the fermentation hydrogen gas collection pipe 15 side, and the carbon dioxide contained in the fermentation hydrogen gas is removed from the carbon dioxide absorption tank 4 filled with the sN-KOH solution.
It leads to the absorption of carbon dioxide gas. The hydrogen gas from which carbon dioxide gas has been removed is introduced into the hydrogen-water displacement tank 5. For nuclide 5, the amount of hydrogen gas is measured.

In this method, in order to culture Miyairi bacteria efficiently, the culture medium must contain 1.0 to 5.0% cornstarch (100 ml of culture solution contains ItIK, hereinafter abbreviated as W H'chi), and a mixture of amino acids. Liquid 1.0 to 5.0 volume
Those containing 0.5 to 1.5 W/V% of calcium carbonate are preferred.

Each of the above components is suspended in water, heated and sterilized according to a known method to prepare a culture medium.

The pH of the culture solution is near neutral in the steady state of continuous culture, and various temperatures are maintained at 30 to 40°C, for example 37°C.

The culture solution obtained from the second tank is centrifuged to collect spores. The culture solution from which the spores have been separated can be returned to the first tank by a pump through a high temperature tank (not shown) and can be reused.

71 By the method of the present invention, Miyairi bacteria can be efficiently cultured to obtain a large amount of hydrogen.

EXAMPLE Using the apparatus shown in the drawings, Miyairi bacteria were continuously cultured under the following culture conditions.

(1) Various dilution rates 1st tank 0.57 hr 2nd tank 0.2 /hr (2) Batch and continuous culture medium Cornstarch 2 W, Al thiamino acid mixture 2 volumes Calcium carbonate I W/V ti First, add 1β and 2.5 medium to the first tank 2 and second tank 3, respectively.
．． 6 and sterilized at 121° C. for 10 minutes, Miyairi bacteria were inoculated, and at the same time stirring was started using the stirrer 6, carbon dioxide gas was supplied from the carbon dioxide gas supply lower to start batch culture. 1st tank 2. In the second tank 3, the growth of Miyairi bacteria in the tank reaches the 8-log phase of logarithmic growth about 8 hours after inoculation with Miyairi bacteria. Therefore, the medium that has already been sterilized is supplied from the medium supply tank 1 to the first tank 2 at a flow rate of 500 ml, and at the same time, the culture medium is transferred from the first tank 2 to the second tank 3 and from the second tank 3 to the outside of the tank. was transferred at a flow rate of 500 ml/hr to start continuous culture. Next, stop the supply of carbon dioxide gas and turn on cock 1.
Switch 1 to the fermentation gas sampling pipe side. At this time, the fermentation gas passes through the carbon dioxide absorption tank 4 and absorbs carbon dioxide,
It is accumulated in the hydrogen-water replacement tank 5.

The amount of accumulated gas was determined by measuring the pressure of the hydrogen-water tank 5 and converting it into the volume under standard conditions.

The hydrogen gas concentration in the fermentation gas collected as above was measured using gas chromatography (■ Shimadzu Corporation (3C3BT)).
Measured using The results are shown in Table-1. However, the gas capacity is a value converted to standard conditions.

[Brief explanation of drawings]

The drawing shows a schematic diagram of an apparatus for carrying out the invention. 1... Culture medium supply tank, 2... First tank,
3...Second tank, 4...Carbon dioxide absorption tank, 5...Hydrogen-water replacement tank, 6...
- Stirrer, 14...Cook. Procedural amendment (self-effective May 1980, 1911 1, Indication of the case 1982 Patent Application No. 61954 2, Name of the invention Hydrogen production method by Kan Miyairi 3, Person making the amendment Relationship to the case Patent applicant address 4. Agent: 107 Idiots 2 people 5. Subject of amendment: "Detailed Description of the Invention" section of the specification. 6. Contents of amendment: Contents of amendment as shown in the attached sheet (1) Page 8, line 16 of the specification. ~Delete “The culture solution obtained from the second tank can be reused.” in line 20. 443-

Claims (2)

[Claims] (1) A method for producing hydrogen using Miyairi bacteria, which is characterized in that a plurality of tanks are used and the dilution rate of each tank is adjusted so that only vegetative cells grow as much as possible in the tanks. (2) The dilution rate of the first tank is 0.3 to 0.6/hr,
The method according to claim 1, characterized in that the dilution rate of the second tank is determined so that the sum of the residence times of the first tank and the second tank is within 7 to 8 hours.