JPH02307599A - Anaerobic digestion - Google Patents

Abstract

PURPOSE:To improve anaerobic digestion efficiency by working bacteria strain having excellent organic substance-decomposing function in acid fermentation process as major bacteria strain. CONSTITUTION:Organic waste is made soluble by acid fermentation using Clorstridium bifermentans as major bacteria strain which converts >=10% of organic substances in a culture into volatile fatty acids with 2-6 of the number of carbon and produces acetic acid >=80wt.% of the total of the fatty acids and a fermentation gas containing >=10vol.% of hydrogen during 2 days anaerobic culture at 37 deg.C in PYG culture fluid and then methane fermentation is carried out using methane bacteria. This solubilizing bacterial solubilizes organic substances at high efficiency and produces acetic acid and hydrogen at extremely high ratio. As a result, organic substances are decomposed efficiently and a gas containing methane in high concentration is obtained.

Description

DETAILED DESCRIPTION OF THE INVENTION [Industrial Application Field] The present invention relates to a method for anaerobic digestion of organic waste using a novel Clostridium microorganism.

[Conventional technology]

There are both aerobic and anaerobic treatment methods for decomposition, or digestion, of organic waste by microbial treatment, but anaerobic microbial treatment is better for decomposition, which is caused by the excessive growth of microorganisms involved in decomposition. It is widely practiced because there are few surplus bacterial cells and it has an energy creation effect by generating methane gas.

Anaerobic digestion processing can be broadly divided into acid fermentation, in which solid organic matter is reduced to a lower molecular weight by facultative anaerobic bacteria and solubilized (liquefied);
There are two decomposition processes: methane fermentation, in which the resulting low molecular weight intermediate products such as alcohols, lower fatty acids, amino acids, etc. are decomposed into methane gas and carbon dioxide gas by methane bacteria, which are unique anaerobic bacteria. The microorganisms involved in the decomposition of organic matter in these processes are the microorganisms present in the digested sludge that is returned as seed sludge, including:
It contains countless microorganisms, from microaerobic bacteria to facultative anaerobes and unique anaerobes, and these microorganisms form an extremely complex bacterial flora. This bacterial flora differs slightly depending on the target of decomposition and the reaction conditions, but until now there has been little detailed knowledge about the anaerobic bacteria that are actually effectively involved in anaerobic digestion.

Therefore, in anaerobic digestion, inoculation with a pure culture of excellent seed bacteria, which is commonly done in the general microbial utilization industry, is not carried out, but instead the sludge that is naturally formed in each digestion tank is used as seed sludge. This is the reality.

[Problem to be solved by the invention]

In the acid fermentation process of anaerobic digestion treatment of organic waste such as sludge, the present invention uses dominant bacterial strains that exhibit an organic matter decomposition ability that is superior to that of the anaerobic bacteria group in the ordinary anaerobically digested sludge. The aim is to make it possible to improve the efficiency of anaerobic digestion by making it act as a gas.

[Means to solve the problem]

In the present invention, when anaerobically cultured in a PYG liquid medium at 37°C for 2 days, more than 10% of the organic matter in the medium has a carbon number of 2 to 6
of volatile fatty acids, and at that time, acetic acid is produced at a ratio of 80% by weight or more based on the total amount of volatile fatty acids produced, and furthermore, a gas containing 10% by volume or more of hydrogen is produced as a fermentation gas. The resulting Clostridium bifermentans
lans (hereinafter referred to as the solubilizing bacteria of the present invention) acts as a dominant bacterial species, organic waste is solubilized by acid fermentation, and further methane fermentation is performed by methane bacteria. It is something to do.

Note that the PYG liquid medium herein means a medium having the following composition, and the details of the culture conditions are as follows.

F'YG liquid medium composition Polypeptone (BBL) 1.0g yeast extract 1.0g glucose
1.Og resazurin sodium (0.1%) 0.4ml 1L-cystine hydrochloride (hydrate) 0.05g distilled water
ioo ugly 1 culture conditions: temperature 3
7°C ± 1°C, anaerobic conditions, static culture. Volatile fatty acids were quantified by gas chromatography on the supernatant obtained by centrifuging the culture medium at 2000G for 30 minutes after the completion of the culture. , acetic acid, propionic acid, isobutyric acid,
Determine butyric acid, isovaleric acid, valeric acid, isocaproic acid and caproic acid.

The solubilizing bacteria of the present invention having the above characteristics were created by the following screening method from anaerobic microorganisms collected by the present inventors from bottom sludge deposited in rivers and sewage channels, compost, landfill soil, etc. It is something that

Screening method: A sterilized washed sludge medium containing no easily soluble nutrients is prepared by the method described below, 10% by weight of a sample of soil or the like is added thereto, and cultured in an anaerobic state for one month. Thereafter, the culture solution is spread on a plate using a sterilized washed sludge agar medium (2% agar added to a sterile washed sludge medium) using the pour-over method, and cultured anaerobically at 37°C. After about a week, a colony that has clearly formed a rear zone (sludge surrounding the colony has been dissolved) on the plate is picked up and expanded onto a new plate similar to the above. Repeat this operation several times (first screening). The strains selected in this way are further subjected to a second screening to select strains with excellent solubilization ability. The selection criteria for the second screening are as follows.

■ Grows in sterile washed sludge media without the need for other nutrient sources.

■ When anaerobic static culture was carried out at 37°C for 20 days in the above medium, 40% of the initial organic matter (loss on ignition at 600°C)
Decomposes the above into gases or volatile substances (lower fatty acids, etc.).

(2) After the above culture, the total concentration of volatile fatty acids dissolved in the medium is 0.2% by weight or more.

■ Acetic acid accounts for 50% of the volatile fatty acids produced above.
occupies more than

(Sterilized washed sludge culture medium preparation method: Surplus sludge collected at the Nibe City sewage treatment plant was sterilized by heating it at 121°C for 15 minutes, 5
Centrifuge at 000G for 10 minutes. The supernatant is discarded, pure water is added to the precipitate, stirred, and centrifuged at 5000G for 10 minutes. After washing with this pure water again,
Add tap water to adjust the organic matter concentration to 1% by weight, and add 121
Heat sterilize in O for 15 minutes to prepare a culture medium.) Seven strains (DYF401, DYF4
05, DYF 451, DYF612, DYF622,
DYF2482 and DYF2553) both exhibited the following mycological properties.

Growth condition Growth on PYG agar medium: Forms milky-white colonies with rough edges Growth on EG blood agar medium: Forms a clear zone (hemolysis) around the colony Behavior towards oxygen: Individuality Anaerobic (0-F test and
Aerobic test using blood agar plate) Heat resistance test (80
'0.10 minutes): Morphology of viable bacterial cells Cell shape and size: Bacillus; 1.2X2-3μm (
PYG agar medium) Presence or absence of cell pleomorphism: Presence or absence of no spore formation: Yes; formed in the center or at the edges (PYG agar medium) Durham staining: Positive Physiological properties Based on the above properties as shown in Table 1, etc. The seven types of anaerobic bacteria mentioned above are t
All were identified as Clostridium pifermentans.

When cultured in a PYG liquid medium, these Clostridium bifermentans exhibited significantly different properties in acetic acid production ability compared to standard strains belonging to the same genus or species. That is, when anaerobically cultured in a PYG liquid medium under the above conditions for 2 days, more than 10% of the organic matter in the medium is converted to volatile fatty acids, and the ratio of acetic acid production to the total volatile fatty acid production at that time (hereinafter referred to as The acetic acid production ratio (referred to as acetic acid production ratio) was 80% by weight or more, but under the same conditions, the acetic acid production ratio in the case of the standard strain was 50% by weight or less (see Example 1 below).

Further, as shown in Table 1, gas production is generally significant, and the produced gas contains hydrogen gas at a high rate of 10% by volume or more (see Table 2).

Table 2 Hydrogen content of fermentation gas (Ma/Ma%) Strain number
PYG liquid medium - sterile washed sludge medium DYF401
18 9DYF405 23
9DYF451 14
11DYF612 13
BDYF622 10 10DYF
2,482 12 9DYF255
3 14 9 Note: Fermentation gas was collected after 2 days of culture in the case of PYG medium, and 10 days after culture in the case of sludge medium, and analyzed by gas chromatography. Note that most of the gases other than hydrogen were carbon dioxide.

The acetic acid production ratio in the PYG liquid medium is shown in Experimental Example 1 below.
As is clear from comparing the results of 2, there is a strong correlation with the acetic acid production ratio in the sterilized washed sludge medium.

Research by Bryal et al. has shown that 70% of the methane produced in anaerobic digestion generally passes through acetic acid, and the remaining 30% passes through hydrogen and carbon dioxide. Methane gasification by anaerobic bacteria progresses faster as the acetation of organic matter progresses, and as the production of hydrogen as the second methane gasification pathway progresses.

Therefore, when cultured anaerobically in a PYG liquid medium for 2 days, more than 10% of the organic activity in the medium was converted to continuous fatty acids, and the acetic acid production ratio at that time was more than 80% by weight, and furthermore, the production of fermentation gas was remarkable. The usefulness of Clostridium pifermentans, which has a high hydrogen production rate, that is, the soluble species of the present invention, as a soluble species in anaerobic digestion was confirmed.

The seven strains of Clostridium vifermentans mentioned above have been deposited with the National Institute of Microbial Technology, and their accession numbers are as follows.

Strain number Accession number DYF401 Microtechnical research institute No. 10497 DY
F 405 Microtechnology Research Institute No. 10498 DYF
451 @1 Research Institute No. 10499 DYF61
2 Microtechnical Research Institute No. 10500 DYF 622
Microtechnology Research Institute No. 10501 DYF2482
Microtechnology Research Institute No. 10502 DYF2553 @1
Kenbokuyori No. 10503 Anaerobic digestion using the soluble compound of the present invention having excellent properties as described above can be carried out in various ways. Typical examples are shown below.

■ When treating something that itself contains anaerobic microorganisms, such as surplus sludge, the soluble specific gravity enriched culture of the present invention can be used as a seed culture in order to make the soluble microorganisms of the present invention act as the dominant bacterial species. Inoculate at high concentration. Alternatively, the waste to be treated is first heat sterilized to eliminate the influence of other competing microorganisms, and then inoculated with the soluble microorganisms of the present invention. The methane bacteria necessary for methane fermentation of the solubilized organic matter may be inoculated at the same time as the soluble foreign material of the present invention, or may be inoculated after completion of solubilization with the solubilized violet of the present invention. Digested sludge extracted from a normal methane fermentation tank can be used as a seed for methane bacteria. The amount used is 5 to 10% of the organic matter in the substrate, regardless of whether or not the substrate sludge is sterilized.
It is enough.

(2) Organic waste such as sludge is charged into a reaction tank filled with the soluble foreign material of the present invention fixed to a carrier by an arbitrary method to solubilize the organic matter. Since immobilization of bacterial cells is effective in maintaining the predominance of soluble dead bacteria of the present invention, the waste to be treated does not need to be sterilized. Methane fermentation may proceed in parallel with solubilization by supplying methane bacteria inoculum to this reaction tank together with the waste to be treated, or the solubilized material may be caused to occur in a separate reaction tank. .

In these methods, the inoculum of the soluble and killed bacteria of the present invention is an aggregate obtained by anaerobically culturing in advance using, for example, meat juice, molasses, pre-settled sludge generated in sewage treatment plants, etc., which have been heat sterilized as a medium. It may be used in the form of a culture. The soluble dead bacteria of the present invention have low motility and tend to settle to the bottom of the culture tank, so by pulling out naturally settled bacteria from the bottom of the tank, an enriched culture with a high bacterial cell concentration can be easily obtained. .

(Effects of the Invention) As mentioned above, when the soluble dead bacteria of the present invention is used for anaerobic digestion of sludge and other organic waste, it solubilizes organic matter at a high rate, and at the same time, it It produces a very high ratio of acetic acid, which is most susceptible to methane, and hydrogen, which is a methane precursor.Therefore, the digestion method of the present invention, in which this bacterium is responsible for the initial acid fermentation process in the anaerobic digestion of sludge, is completely independent of the bacterial species. It can decompose organic matter more efficiently than conventional anaerobic digestion methods, which do not require any countermeasures, and can obtain a large amount of gas with a high methane concentration, contributing to the miniaturization of equipment and energy creation.

In addition, the soluble killed bacteria of the present invention not only has a high acetic acid production ratio as mentioned above, but also because it is Clostridium pipermentans, ■ It is optimal because it survives by forming spores in high-temperature environments. It will not die even if the temperature rises due to an accident when it is used near the temperature (approximately 37 degrees Celsius); ■ Although it is originally an obligate anaerobic bacterium, it will not die even in microaerobic conditions. Therefore, even if there is a chance of air exposure during use, there is no danger of immediate death and it is easy to handle; ■ In a suitable environment, spore formation is low even among Clostridium species, and therefore the metabolic capacity is high (the spore state is a dormant state). It has properties that are extremely advantageous for use in the anaerobic digestion of organic wastes such as sludge, where it is difficult to expect stability of the treated materials or process conditions. The anaerobic digestion method of the present invention using this method has the advantage of showing stable results even if there are fluctuations in the digestion conditions.

Due to the above features, the anaerobic digestion method of the present invention can be widely used to treat surplus sludge generated in sewage treatment plants, as well as various other organic wastes generated in food factories, agricultural, livestock and marine product processing factories, etc. It is advantageous that it can be adopted.

〔Example〕

Hereinafter, the present invention will be explained by showing experimental examples and examples.

Experimental Example 1 Seven types of solubilized tubes of the present invention and three types of standard strains were
Static anaerobic culture was performed at 37°C for 48 hours using a liquid medium. After completion of the culture, the medium was centrifuged at 2000G for 30 minutes, and the volatile fatty acids in the obtained supernatant were quantified by gas chromatography. The results are shown in Table 3.

The three types of standard stocks are as follows.

Standard stock engineering: RIKEN stock C Pifermentans CM1
3H Standard strain ■: Fermentation research stock C, Ace I, Butyricum 1FO
H94B standard strain ■: Fermentation Research Institute strain C. sporogenes 1FO139
50 Experimental Example 2 Anaerobic culture was carried out in the same manner as in Experimental Example 1, except that a sterile washed sludge medium was used instead of the PYG liquid medium and the number of days of culture was changed to 20 days, and then volatile fatty acids were quantified.

The results are shown in Table 4. The concentration of volatile fatty acids in the sterilized washed sludge medium itself was below the detection limit by gas chromatography.

Example 1 Batch-type anaerobic digestion of 100 ml of surplus sludge generated in a sewage treatment plant was carried out in a state in which the solubilizing cylinder of the present invention was administered at a high concentration to become dominant. As the solubilization seed of the present invention, DYF2553 or DYF622 is used, and the inoculum is cultured statically for 3 days in an anaerobic PYG liquid medium at 37°C, centrifuged, and washed once with physiological saline. It was used as As the inoculum for methane bacteria, 5 ml of digested sludge generated at a municipal sewage treatment plant was used.

The treated surplus sludge was collected from a municipal sewage treatment plant and diluted with tap water so that the vS concentration was approximately 1%.

Cultivation was carried out using a batch incubator using a 2001 volume Erlenmeyer flask equipped with a degassing septum at 37°C.
The cells were subjected to anaerobic static culture for 0 days. During the culture period, the amount of gas generated was measured, and after the completion of the culture, the amount of VS remaining in the fermentation residue was examined to determine the vsS decomposition rate.

As a control test, anaerobic digestion was carried out in the same manner as above except that the solubilizing cylinder of the present invention was not added.

The results are shown in Table 5.

Table 5 Note: VS: Vol*Lil* 5oli4s (
6G O℃ ignition loss) VS decomposition rate (%): [Initial VS - remaining VS) X100/Initial VS sludge vs:
Total VS amount of treated surplus sludge and digested sludge added as methane bacteria seed VS: Total VS amount when VS brought in in the form of soluble other countries of the present invention is added to sludge VS Example 2 Geographically targeted surplus sludge Excess sludge was anaerobically digested in the same manner as in Example 1, except that the sludge was sterilized in advance. The results are shown in Table 6.

Table 6 Example 3 Excess sludge was anaerobically digested in the same manner as in Example 1 using a soluble material of the present invention fixed on a ceramic carrier. As a ceramic carrier, Iwao Porcelain Co., Ltd.
60°C immediately before use.
After dry heat sterilization at 0°C, the test bacterial strain enriched culture (cultured statically at 37°C for 2 days in an anaerobic PYG liquid medium) was immersed for 2 days, and the culture was continued while the bacterial cells were added to the carrier. was attached. The carrier to which the bacteria had adhered was washed with physiological saline immediately before use, and was used as a ceramic carrier with immobilized test bacteria. Note that the amount of attached organic matter was measured as vS.

The culture vessel is 200m1 with a degassing septum installed.
A batch incubator with an Erlenmeyer flask was used, and 100+ liters of excess sludge, 50 g of a ceramic carrier with immobilized test bacteria,
Then, 50 ml of digested sludge was added as a methane bacterial inoculum. As a control test, a digestion experiment was conducted in the same manner as above except that 50 g of a ceramic carrier without immobilizing the dead body was added instead of the test bacteria immobilized carrier (Control I), and a digestion experiment was carried out in the same manner as above (Control I), and an anaerobic experiment in which only the surplus sludge was cultured An experiment was conducted to investigate digestion by bacterial groups (control ■).

In the above experiment, the same measurements as in Example 1 were carried out; the results are shown in Tables 7 and 8. In addition, in Table 7, “
The total VSJ is the total VS amount obtained by adding the vS brought in in the form of immobilized bacterial cells to the sludge vs.

Table 7

Claims (2)

[Claims] (1) When anaerobically cultured in a PYG liquid medium at 37°C for 2 days, more than 10% of the organic matter in the medium is converted to volatile fatty acids having 2 to 6 carbon atoms, and at that time, the total amount of volatile fatty acids produced is producing acetic acid at a ratio of 80% by weight or more,
Furthermore, the organic waste is solubilized through acid fermentation in which Clostridium bifermentans, which produces a gas containing 10% by volume or more of hydrogen, acts as a dominant bacterial species, and further methane fermentation by methane bacteria is carried out. An anaerobic digestion method for organic waste characterized by: (2) When anaerobically cultured at 37°C for 2 days in a PYG liquid medium, 10% or more of the organic matter in the medium was converted to volatile fatty acids having 2 to 6 carbon atoms, and at that time, the total amount of volatile fatty acids produced was Organic waste is fed into a reaction tower filled with Clostridium bifermentans fixed on a carrier, which produces acetic acid at a ratio of 80% by weight or more and further produces a gas containing hydrogen at 10% by volume or more as fermentation gas. Solubilizes organic matter by supplying and causing acid fermentation,
A method for anaerobic digestion of organic waste, characterized in that the organic waste is simultaneously or subsequently inoculated with methane bacteria to perform methane fermentation.