WO2002046370A1 - Promoteur pour la culture haute densite d'une bacterie autotrophe - Google Patents

Promoteur pour la culture haute densite d'une bacterie autotrophe Download PDF

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Publication number
WO2002046370A1
WO2002046370A1 PCT/JP2001/010295 JP0110295W WO0246370A1 WO 2002046370 A1 WO2002046370 A1 WO 2002046370A1 JP 0110295 W JP0110295 W JP 0110295W WO 0246370 A1 WO0246370 A1 WO 0246370A1
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WO
WIPO (PCT)
Prior art keywords
culture
bacteria
concentration
autotrophic
promoter
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Application number
PCT/JP2001/010295
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English (en)
Japanese (ja)
Inventor
Satoshi Yoneda
Original Assignee
Bicom Corporation
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Bicom Corporation filed Critical Bicom Corporation
Priority to AU2002224102A priority Critical patent/AU2002224102A1/en
Publication of WO2002046370A1 publication Critical patent/WO2002046370A1/fr

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    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
    • C12N1/38—Chemical stimulation of growth or activity by addition of chemical compounds which are not essential growth factors; Stimulation of growth by removal of a chemical compound
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
    • C12N1/20—Bacteria; Culture media therefor

Definitions

  • the present invention relates to an autotrophic bacterium high-concentration cultivator for culturing autotrophic bacteria at a high concentration (hereinafter simply referred to as “promoting”). It relates also referred) and agents ", more particularly, to accelerators for culturing autotrophic bacteria causing increasingly like medium P H is changed in response to high concentrations in culture toothpick.
  • the conventional culture method is a small scale at the test tube level aiming at pure culture, and the medium in the flask does not suspend in about two months, and the culture method that can be applied industrially is I could not say.
  • pH decreases when nitrification starts in the culture of nitrifying bacteria, for example.
  • the present invention has been made in view of the above circumstances, and an object of the present invention is to provide an accelerator for culturing autotrophic bacteria at a high concentration.
  • the high-concentration autotrophic bacterium cultivation promoter according to claim 1 maintains the pH of a medium that is inclined to an acidic side in the acclimatization process in a predetermined range when the autotrophic bacterium is acclimated for a predetermined period to be cultured at a high concentration.
  • An autotrophic bacterium high-concentration cultivation enhancer that serves as a carbon source together with the bacterium, which is capable of maintaining the pH of the culture medium that exhibits basicity by dissociation and tilts toward the acidic side within a predetermined range, and If two types of properties can be provided by one type of compound, a property that can be a carbon source during the growth of vegetative bacteria, and at least one type of the compound, or two or more types of the above properties can be provided by two or more types of compounds. In the case where the compound can be applied, a mixture of the compound is blended.
  • the high-concentration cultivation agent for autotrophic bacteria according to claim 2 is the accelerator according to claim 1, wherein the mixture is a mixture of the following components (A) and (B). And
  • the accelerator for autotrophic bacterium high concentration culture according to claim 3 is the accelerator according to claim 2, wherein the mixing ratio of the component (A): the component (B) is 1: 4 to 17 in a molar ratio. No.7.
  • the autotrophic bacterium high concentration culture promoter according to claim 4 is the promoter according to claim 2 or 3, wherein the component (A) is sodium carbonate.
  • the component (B) is hydrogencarbonate. It is characterized by being sodium.
  • Figure 2 is a curve showing the concentration of NH 4 —N remaining 4 hours after injection of NH 4 — N (lO Omg / liter), showing the change in NH 4 —N concentration during the nitrification acclimation process of sludge.
  • FIG. The mark in the figure indicates the introduction of NH4-N (100 mg Z liter).
  • FIG. 3 is a graph showing the daily change of the MLSS concentration during the nitrification acclimation process of sludge.
  • FIG. 4 is a graph showing changes in MLSS concentration after acclimation for 8 months by changing the composition ratio of the culture promoter.
  • FIG. 5 is a graph showing changes in MLSS concentration after habituation by changing the type of culture promoter.
  • FIG. 6 is a graph showing the daily change of the MLSS concentration in the process of acclimating sulfur-oxidizing bacteria in sludge.
  • the autotrophic bacterium according to the present invention refers to any bacterium that causes a reaction such that the pH of the medium decreases during cultivation. Specifically, ammonia-oxidizing bacteria (nitrifying bacteria), sulfur-oxidizing bacteria, Examples thereof include carbon monoxide oxidizing bacteria, and a mixed bacterium of two or more of these.
  • the components of the accelerator of the present invention have the property of exhibiting basicity by dissociation and having the property of maintaining the pH of the culture medium inclined toward the acidic side during acclimation within a predetermined range (7.0 to 9.0).
  • the compound is not particularly limited as long as it has two properties, namely, a property that can serve as a carbon source during the growth of the autotrophic bacterium.
  • At least one kind of the compound is a component of the accelerator of the present invention.
  • a mixture of the compounds is a component of the accelerator of the present invention.
  • a mixture of (A) a carbonate which is soluble in water and exhibits basicity by dissociation and (B) a bicarbonate which is soluble in water can be mentioned.
  • hydroxides such as sodium hydroxide and potassium hydroxide, and alkali metal salts such as sodium carbonate.
  • component (B) sodium hydrogen carbonate, magnesium carbonate (4Mg C03 ' Carbonates such as Mg (OH) 2-5H2O).
  • the component (A) is an alkali metal carbonate or an alkaline earth metal carbonate which is soluble in water, such as sodium carbonate, sodium carbonate, potassium carbonate, etc .;
  • the two properties described above are effective when the component is a water-soluble alkaline metal bicarbonate or an alkaline earth metal bicarbonate that is soluble in water, such as sodium bicarbonate or bicarbonate.
  • sodium carbonate and carbonic acid Most preferred is a combination of sodium hydrogen.
  • the activated sludge used in the present invention includes sewage sludge and night soil sludge. These may be freshwater-diluted or seawater-diluted, but if the autotrophic bacteria are cultured using seawater-diluted sludge as a raw material, they are rare. Since a large amount of valuable marine bacteria (salt-resistant bacteria) can be obtained, it is preferable to use activated sludge that has been subjected to seawater dilution.
  • seawater contains marine bacteria that are considered to have higher salt tolerance than freshwater bacteria, their abundance is very small and pure separation is difficult. Its research has lagged behind freshwater bacteria.
  • a large amount of marine bacteria having a high concentration can be obtained by using activated sludge that has been subjected to seawater dilution treatment as described above as a raw material. Marine bacteria have multi-layered cell walls and have strong resistance to various osmotic pressures of treated water and various chemicals that inhibit growth.
  • the culture of nitrifying bacteria contained in the activated sludge is carried out by
  • sludge dewatered filtrate ⁇ anaerobic Nitrification by sludge treatment wastewater such as digestion and desorption solution is required. Therefore, the dissolved oxygen (DO) at this time must be 2 mg Z liter or more. However, for the first time in this experiment, it was found that if the dissolved oxygen concentration was too high, the growth rate would tend to decrease. The details will be described below.
  • dissolved oxygen (DO) concentration S mgZ liter is most preferable.
  • the pH must be 7.0 to 9.0, particularly when using activated sludge diluted with seawater, preferably 7.5 to 8.5, more preferably 7.5 to 7.8. preferable.
  • the growth speed is high in the range of 20 to 40 ° C, and more preferably 25 to 35 ° C.
  • Equation (C) is an equation for the entire nitrifying bacteria.
  • a buffer which has a pH which is inclined toward the acidic side in the culture process and which has a buffering action as a mixture of a normal salt and an acid salt is preferable.
  • a culture promoting agent comprising a combination of sodium carbonate and sodium bicarbonate
  • a carbon source for carbonic assimilation of nitrifying bacteria can be supplied at the same time. The explanation is added below.
  • the sodium carbonate is In some cases, the effect of increasing the pH can be sufficiently recognized.However, the effect of increasing the pH is so large that it cannot be used in a large amount and is not suitable for supplying a sufficient carbon source. is there. On the other hand, when using only sodium hydrogen carbonate, there is no problem in terms of supply as an inorganic carbon source, but a large amount of supply is required in terms of maintaining pH, which is not preferable.
  • a mixture of sodium carbonate and sodium hydrogen carbonate can be suitably used.
  • an aqueous solution of the mixture it became possible to effectively supply an inorganic carbon source for assimilation of carbonic acid in a living body while maintaining a gradually decreasing pH.
  • the mixture ratio of sodium carbonate and sodium hydrogencarbonate in the mixture is preferably 1: 4Z7 to 17/7 in a molar ratio of sodium carbonate: sodium hydrogencarbonate.
  • sodium bicarbonate is 4Z 7 (0.571-) mo 1 Z liter ⁇ 17/7 (2.4 8 8) mo 1 liter
  • An aqueous solution of the mixture is effective.
  • the monitoring of the pH of the culture system may be performed continuously or at predetermined time intervals. It is preferable to use a continuous pH monitoring device such as a pH controller, but the present invention is not limited to this. It is also possible to perform the measurement manually using a pH indicator such as phenol red.
  • the concentration of ammonia in the NH4-N-containing liquid is preferably 100 mg / liter or more and 300 mg / liter or less, and more preferably 200 mg / liter or less.
  • Ammonia is an energy source for the growth of ammonium oxidizing bacteria, which are chemolithotrophic bacteria, by assimilating carbon dioxide. However, excessive amounts of ammonia may inhibit growth and growth.
  • Nitrite oxidizing bacteria contained in activated sludge, like ammonia oxidizing bacteria, are bacteria that take over and oxidize nitrite generated by ammonia oxidizing bacteria. In this case, the initial concentration of ammonia cannot be set unnecessarily high because it is weak to high nitrite concentration. Therefore, the concentration of ammonia
  • sludge treatment waste liquid such as sludge dewatered filtrate or digestion / desorption liquid generated in the water treatment plant.
  • activated sludge is nitrified by sludge treatment wastewater such as sludge dewatered filtrate and digestion desorbent, so that nitrifying bacteria slightly contained in the activated sludge can be cultured at a high concentration.
  • the activated sludge can be reduced to 1/3 to 1/4 in two months, and nitrified sludge having a large specific gravity can be obtained.
  • activated sludge originally contains about 0.35% of nitrifying bacteria.
  • the content of nitrifying bacteria in the activated sludge can be increased by about 10 times (3.5%).
  • other germs cannihilate and die because no external nutrients (feeds) are provided.
  • the amount of activated pollutants is reduced.
  • the germs When the germs almost die, they become hard-to-decompose organic matter called "granules", which are attached to nitric bacteria around the nucleus.
  • nitrifying bacteria sediments in the culture system due to its specific gravity. In order to culture nitrifying bacteria at a high concentration, this good sedimentation property is required. In other words, nitrifying bacteria generally have a low specific gravity and float in pure culture. Therefore, nitrifying bacteria are likely to flow out of the culture system, and high-concentration culture cannot be expected. As a result, the production of nuclei (refractory organic substances) as described above is required for high-concentration culture, but nucleation is not seen in pure culture of nitrifying bacteria, and is not active. Only seen when sludge is used as raw material.
  • the promoter of the present invention enables cultivation of autotrophic bacteria, which has been impossible so far, in a large amount and at a high concentration.
  • Example 1 High concentration culture of nitrifying bacteria [Production of nitrifying activated sludge]
  • Batch culture was performed in a two-day cycle in a fililand draw type culture tank (30 liters) shown in FIG. That is, urine sludge diluted with seawater and anaerobic digestion / elimination liquid (diluted in seawater so that the concentration of NH4-N is 100 mgZ liter) are put into a culture tank, and the temperature in the culture tank is thermostated. So that the temperature becomes 27 ° C with a heater and a heater, and adjust the pH with a pH controller and a culture promoter (0.5 mol / liter Na 2 CO 3 and 1 mol 1 Z liter).
  • the culture was performed by setting the pH to 7.5 to 8.5 with a buffer consisting of NaHCO 3 (when the initial pH was 8.5 or more, dilute sulfuric acid was added. 8.5 or less). The amount of aeration was adjusted with a balloon to adjust the dissolved oxygen (DO) concentration to 4 mg / litre.
  • a buffer consisting of NaHCO 3 (when the initial pH was 8.5 or more, dilute sulfuric acid was added. 8.5 or less).
  • the amount of aeration was adjusted with a balloon to adjust the dissolved oxygen (DO) concentration to 4 mg / litre.
  • the replenisher / desorbent was added to a final concentration of 10 Omg / liter.
  • the aeration was stopped, the sludge was settled for 1 hour, the supernatant was removed, the digestion / desorption solution was added, and the aeration was restarted.
  • S sludge MLS S concentration (g liter) Measure the SV30 and SVI of seawater acclimated nitrifying activated sludge (AMN S), which has been acclimated after a seawater acclimation period of about 60 days, and examine sedimentation characteristics At the same time, the state of floc formation was observed using an optical microscope.
  • APN S seawater acclimated nitrifying activated sludge
  • the graph in Fig. 2 shows the process of acclimating night sludge to seawater (the figure shows the concentration of 100 mg / liter of NH4-N after 4 hours).
  • ammonia and nitrite are separately required as energy substrates, but in a mixed culture system, only ammonia needs to be supplied as an energy substrate.
  • Type of culture promoter Composition
  • High-density cultivation of nitrifying bacteria production of nitrifying activated sludge was performed in the same manner as in Example 2, except that [NaHC03, Na2CO3] was changed.
  • Figure 5 shows the MLSS concentration of AMN S after habituation.

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  • Chemical & Material Sciences (AREA)
  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Engineering & Computer Science (AREA)
  • Organic Chemistry (AREA)
  • Biotechnology (AREA)
  • Zoology (AREA)
  • Wood Science & Technology (AREA)
  • Genetics & Genomics (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Biomedical Technology (AREA)
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  • General Engineering & Computer Science (AREA)
  • General Health & Medical Sciences (AREA)
  • General Chemical & Material Sciences (AREA)
  • Micro-Organisms Or Cultivation Processes Thereof (AREA)
  • Purification Treatments By Anaerobic Or Anaerobic And Aerobic Bacteria Or Animals (AREA)

Abstract

L'invention concerne un promoteur pour la culture haute densité d'une bactérie autotrophe permettant de réguler le pH d'un milieu et variant dans une gamme spécifique au cours de l'acclimatation de la bactérie autotrophe dans une période de culture haute densité déterminée, et faisant office de source de carbone. Ce promoteur comprend un mélange de carbonate de sodium et d'hydrogénocarbonate de sodium, et il présente un rapport de mélange par mol carbonate de sodium: hydrogénocarbonate de sodium de 1: 4/7 à 17/7.
PCT/JP2001/010295 2000-12-08 2001-11-26 Promoteur pour la culture haute densite d'une bacterie autotrophe WO2002046370A1 (fr)

Priority Applications (1)

Application Number Priority Date Filing Date Title
AU2002224102A AU2002224102A1 (en) 2000-12-08 2001-11-26 Promoter for the culture of autrotrophic bacterium at high density

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Application Number Priority Date Filing Date Title
JP2000-375317 2000-12-08
JP2000375317A JP2004283001A (ja) 2000-12-08 2000-12-08 独立栄養細菌を高濃度に培養するための促進剤

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Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2008038076A1 (fr) * 2006-09-27 2008-04-03 Immobiliare G.M. S.R.L Préparations de milieux de culture pour applications industrielles

Families Citing this family (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JP5213153B2 (ja) * 2007-06-13 2013-06-19 学校法人 芝浦工業大学 海水由来微生物による汚染海水浄化方法
JP5039093B2 (ja) * 2009-06-15 2012-10-03 株式会社栄電社 バイオリアクター素子の製造方法

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JPH04197174A (ja) * 1990-11-29 1992-07-16 Mitsubishi Heavy Ind Ltd 硝酸菌培養液及び同菌の高濃度培養方法
JPH04200383A (ja) * 1990-11-30 1992-07-21 Mitsubishi Heavy Ind Ltd 亜硝酸菌培養液及び同菌の高濃度培養方法
JPH0857497A (ja) * 1994-08-24 1996-03-05 Nippon Steel Corp 鉄酸化細菌の増殖方法
JPH11299481A (ja) * 1998-04-17 1999-11-02 Nippon Steel Corp 硫黄酸化細菌の馴養方法および硫黄酸化細菌を用いた排水からの窒素の除去方法

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JPS59232087A (ja) * 1983-06-13 1984-12-26 Mitsubishi Gas Chem Co Inc 嫌気性細菌の培養方法
DE4216357C1 (de) * 1992-05-18 1993-12-09 Nitrochemie Gmbh Abbau stickstoffhaltiger Stoffe mittels eines Mikroorganismus
JPH08252088A (ja) * 1995-03-17 1996-10-01 Nippo Kagaku Kk クロストリジウム属細菌の培養方法
JPH10295285A (ja) * 1997-05-01 1998-11-10 Kurorera Kogyo Kk ワムシ餌料

Patent Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JPH04197174A (ja) * 1990-11-29 1992-07-16 Mitsubishi Heavy Ind Ltd 硝酸菌培養液及び同菌の高濃度培養方法
JPH04200383A (ja) * 1990-11-30 1992-07-21 Mitsubishi Heavy Ind Ltd 亜硝酸菌培養液及び同菌の高濃度培養方法
JPH0857497A (ja) * 1994-08-24 1996-03-05 Nippon Steel Corp 鉄酸化細菌の増殖方法
JPH11299481A (ja) * 1998-04-17 1999-11-02 Nippon Steel Corp 硫黄酸化細菌の馴養方法および硫黄酸化細菌を用いた排水からの窒素の除去方法

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Title
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Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2008038076A1 (fr) * 2006-09-27 2008-04-03 Immobiliare G.M. S.R.L Préparations de milieux de culture pour applications industrielles

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AU2002224102A1 (en) 2002-06-18

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