JP2004283001A - Promotor for culturing autotrophic bacterium at high concentration - Google Patents
Promotor for culturing autotrophic bacterium at high concentration Download PDFInfo
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- JP2004283001A JP2004283001A JP2000375317A JP2000375317A JP2004283001A JP 2004283001 A JP2004283001 A JP 2004283001A JP 2000375317 A JP2000375317 A JP 2000375317A JP 2000375317 A JP2000375317 A JP 2000375317A JP 2004283001 A JP2004283001 A JP 2004283001A
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- 241000894006 Bacteria Species 0.000 title claims abstract description 83
- 230000001651 autotrophic effect Effects 0.000 title claims abstract description 31
- 238000012258 culturing Methods 0.000 title abstract description 11
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- UIIMBOGNXHQVGW-UHFFFAOYSA-M Sodium bicarbonate Chemical compound [Na+].OC([O-])=O UIIMBOGNXHQVGW-UHFFFAOYSA-M 0.000 claims abstract description 26
- 239000000203 mixture Substances 0.000 claims abstract description 18
- OKTJSMMVPCPJKN-UHFFFAOYSA-N Carbon Chemical compound [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 claims abstract description 17
- 229910052799 carbon Inorganic materials 0.000 claims abstract description 17
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- 238000000034 method Methods 0.000 claims abstract description 13
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- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 10
- 239000003513 alkali Substances 0.000 claims description 9
- -1 alkaline earth metal carbonate Chemical class 0.000 claims description 7
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- 229910000028 potassium bicarbonate Inorganic materials 0.000 description 1
- 235000015497 potassium bicarbonate Nutrition 0.000 description 1
- 239000011736 potassium bicarbonate Substances 0.000 description 1
- 229910000027 potassium carbonate Inorganic materials 0.000 description 1
- 235000011181 potassium carbonates Nutrition 0.000 description 1
- TYJJADVDDVDEDZ-UHFFFAOYSA-M potassium hydrogencarbonate Chemical compound [K+].OC([O-])=O TYJJADVDDVDEDZ-UHFFFAOYSA-M 0.000 description 1
- VLYFRFHWUBBLRR-UHFFFAOYSA-L potassium;sodium;carbonate Chemical compound [Na+].[K+].[O-]C([O-])=O VLYFRFHWUBBLRR-UHFFFAOYSA-L 0.000 description 1
- 230000003252 repetitive effect Effects 0.000 description 1
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- 150000003839 salts Chemical class 0.000 description 1
- 238000000926 separation method Methods 0.000 description 1
- KKCBUQHMOMHUOY-UHFFFAOYSA-N sodium oxide Chemical compound [O-2].[Na+].[Na+] KKCBUQHMOMHUOY-UHFFFAOYSA-N 0.000 description 1
- 229910001948 sodium oxide Inorganic materials 0.000 description 1
- 229910000031 sodium sesquicarbonate Inorganic materials 0.000 description 1
- 235000018341 sodium sesquicarbonate Nutrition 0.000 description 1
- 229910052938 sodium sulfate Inorganic materials 0.000 description 1
- 235000011152 sodium sulphate Nutrition 0.000 description 1
- 239000000243 solution Substances 0.000 description 1
- 239000006228 supernatant Substances 0.000 description 1
- 238000003786 synthesis reaction Methods 0.000 description 1
- PFMBRNMAXCVTIV-UHFFFAOYSA-K trisodium hydrogen carbonate Chemical compound [Na+].[Na+].[Na+].OC([O-])=O.OC([O-])=O.OC([O-])=O PFMBRNMAXCVTIV-UHFFFAOYSA-K 0.000 description 1
- WCTAGTRAWPDFQO-UHFFFAOYSA-K trisodium;hydrogen carbonate;carbonate Chemical compound [Na+].[Na+].[Na+].OC([O-])=O.[O-]C([O-])=O WCTAGTRAWPDFQO-UHFFFAOYSA-K 0.000 description 1
- 210000002700 urine Anatomy 0.000 description 1
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- 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
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- 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
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Abstract
Description
【０００１】
【発明の実施の形態】
本発明は、独立栄養細菌を高濃度に培養するための独立栄養細菌高濃度培養促進剤（以下、単に「促進剤」ともいう）に関し、詳しくは、培養時において次第に培地のｐＨが変化するような反応をおこす独立栄養細菌を高濃度に培養するための促進剤に関する。
【０００２】
【従来の技術と発明が解決しようとする課題】
アンモニア酸化細菌（硝化細菌）や硫黄酸化細菌などの独立栄養細菌は、増殖速度が遅く、コロニーを作って生活しないことに起因して、その存在が確認されて百年以上経た今日まで、工業的に大量に高濃度培養することに成功したという報告はない。
【０００３】
すなわち、従来の培養方法は、純粋培養を目指す試験管レベルの小規模のものであり、２ヶ月程ではフラスコ内の培地が懸濁しない程度であり、工業的に応用できる培養方法とは言えなかった。
【０００４】
説明を加えると、例えば硝化細菌の培養において硝化が始まると、ｐＨが低下するわけであるが、従来、このｐＨを効果的に上げる方法が分からなかった。
【０００５】
一方、硝化に伴って炭素源が減少していくわけであるが、従来、炭素源の供給として二酸化炭素を使用していた。二酸化炭素の供給により確かに炭素源の枯渇を防ぐことができるが、前述したｐＨの低下がさらに進み、延いては硝化細菌の活動が停止し、これが細菌増殖の限界であった。
【０００６】
［発明の目的］
本発明は上記の実情に鑑みてなされたものであり、その目的は、独立栄養細菌を高濃度に培養するための促進剤を提供するところにある。
【０００７】
【課題を解決するための手段】
請求項１に記載の独立栄養細菌高濃度培養促進剤は、独立栄養細菌を高濃度に培養すべく所定期間馴養する際、当該馴養過程において酸性側に傾く培地のｐＨを所定範囲に維持させると共に炭素源となる独立栄養細菌高濃度培養促進剤であって、解離することにより塩基性を呈して前記酸性側に傾く培地のｐＨを所定範囲に維持させることのできる性質と、前記独立栄養細菌の増殖時の炭素源となりうる性質との２つの性質を、１種類の化合物で以て付与できる場合は当該化合物の少なくとも１種、または上記２つの性質を２種以上の化合物で以て付与できる場合は、当該化合物の少なくとも２種が混合された混合物が配合されてなるものである。
【０００８】
請求項２に記載の独立栄養細菌高濃度培養促進剤は、請求項１に記載の促進剤において、前記した混合物が、下記（Ａ）成分と（Ｂ）成分との混合物であることを特徴とする。
【０００９】
（Ａ）水に可溶で、解離することにより塩基性を示す塩基性物質。
【００１０】
（Ｂ）水に可溶性を示す炭酸塩。
【００１１】
請求項３に記載の独立栄養細菌高濃度培養促進剤は、請求項２に記載の促進剤において、前記（Ａ）成分が炭酸アルカリ塩又は炭酸アルカリ土類金属塩であり、前記（Ｂ）成分が炭酸水素アルカリ塩又は炭酸水素アルカリ土類金属塩であることを特徴とする。
【００１２】
請求項４に記載の独立栄養細菌高濃度培養促進剤は、請求項２または３に記載の促進剤において、前記（Ａ）成分：（Ｂ）成分の混合割合がモル比で４〜７：４〜８であることを特徴とする。
【００１３】
請求項５に記載の独立栄養細菌高濃度培養促進剤は、請求項２〜４のいずれか１項に記載の促進剤において、前記（Ａ）成分が、炭酸ナトリウムであり、前記（Ｂ）成分が、炭酸水素ナトリウムであることを特徴とする。
【００１４】
【発明の実施の形態】
独立栄養細菌
本発明における独立栄養細菌とは、培養時において培地のｐＨが下がっていくような反応を起こす細菌の全てをいい、具体的には、アンモニア酸化細菌（硝化細菌）、硫黄酸化細菌、一酸化炭素酸化細菌など、及びこれら２つ以上の混合菌などが挙げられる。
【００１５】
これらの細菌は、例えば、活性汚泥、海水中等に含まれているが、どちらかと言えば、細菌がより多く含有するという点で、活性汚泥を用いる方が好適である。
【００１６】
促進剤成分
本発明の促進剤の成分としては、解離することにより塩基性を呈し、馴養中に酸性側に傾く培地のｐＨを所定範囲（７．０〜９．０）に維持させることのできる性質と、前記独立栄養細菌の増殖時の炭素源となりうる性質との２つの性質を備えている化合物であれば、特に限定されるものではない。
【００１７】
前記２つの性質を１種類の化合物で以て付与できる場合は当該化合物の少なくとも１種が本発明の促進剤の成分となる。
【００１８】
その具体例としては、二炭酸水素三ナトリウム（＝セスキ炭酸ナトリウム）（Ｎａ２ＣＯ３・ＮａＨＣＯ３・２Ｈ２Ｏ）や、この化合物におけるアルカリが、カリウムやマグネシウムなどに一部あるいは全部置換された化合物などが挙げられる。
【００１９】
また、上記２つの性質を２種以上の化合物で以て付与できる場合は、当該化合物の少なくとも２種が混合された混合物が本発明の促進剤の成分となる。例えば、（Ａ）水に可溶性を示し解離することにより塩基性を示すアルカリと、（Ｂ）水に可溶性を示す炭酸塩との混合物が挙げられ、具体的には、（Ａ）成分として、水酸化ナトリウム、水酸化カリウムなどの水酸化物や、炭酸ナトリウム、硫酸ナトリウムなどのアルカリ塩が挙げられ、（Ｂ）成分としては、炭酸水素ナトリウム、炭酸水酸化マグネシウム（４ＭｇＣＯ３・Ｍｇ（ＯＨ）２・５Ｈ２Ｏ）などの炭酸塩が挙げられる。
【００２０】
なかでも、前記（Ａ）成分が、炭酸ナトリウム、炭酸ナトリウムカリウム、炭酸カリウム等の、水に可溶性を示す炭酸アルカリ塩又は炭酸アルカリ土類金属塩であり、前記（Ｂ）成分が、炭酸水素ナトリウム、炭酸水素カリウム等の、水に可溶性を示す炭酸水素アルカリ塩又は炭酸水素アルカリ土類金属塩であることが、前述した２つの性質を効果的に付与できるという点で好ましく、なかでも、炭酸ナトリウムと炭酸水素ナトリウムの組合せが最も好ましい。
【００２１】
活性汚泥
本発明に使用する活性汚泥は、下水汚泥やし尿汚泥が挙げられる。これらは、淡水希釈処理されたものであってもよく、あるいは海水希釈処理されたものであっても構わないが、海水希釈汚泥を原料として当該独立栄養細菌を培養すれば、希少価値とされる海洋性の細菌（耐塩性の細菌）が大量に得られるので、海水希釈処理された活性汚泥を使用することが好適である。
【００２２】
説明を加えると、天然の海水中には淡水性細菌よりも高い耐塩性を有すると考えられる海洋性の細菌が存在するが、その存在量は非常に少なく、純粋分離が困難であるため、淡水性細菌に比べてその研究は遅れている。しかしながら、本発明の培養方法であれば、前述したように海水希釈処理された活性汚泥を原料とすることにより、高い濃度の海洋性細菌を大量に得ることができる。海洋性細菌は、多層の細胞壁を備え、処理水の浸透圧変化や生育を阻害する種々の化学物質に対して強力な耐性を持つ。
【００２３】
硝化細菌の培養（馴養）条件
活性汚泥に含まれる硝化細菌の培養は、当該活性汚泥を、所定期間（例えば、１ヶ月、２ヶ月あるいは３ヶ月）、汚泥脱水濾液や（嫌気性）消化脱離液などの汚泥処理廃液により硝化馴養するわけであるが、この硝化馴養は好気的に行なう必要があるため、この際の溶存酸素（ＤＯ）を２ｍｇ／リットル以上とする必要がある。しかしながら、溶存酸素濃度を無闇に高くし過ぎると逆に増殖スピードが低下する傾向になることが今回の実験で初めて分かった。以下詳述する。
【００２４】
硝化細菌による硝化スピードは、溶存酸素が高ければ高いほど速くなるので、硝化馴養集積にあっても同様、高いほど速く進むであろうと考えられていたが、あにはからんや、活性汚泥を原料とした硝化細菌の馴養集積は、溶存酸素（ＤＯ）５ｍｇ／リットルを超えるところぐらいからそのスピードが低下することが分かった。なお、溶存酸素（ＤＯ）濃度２〜４ｍｇ／リットルが最も好ましい。
【００２５】
また、ｐＨは７．０〜９．０である必要があり、（特に海水希釈の活性汚泥を使用する場合）７．５〜８．５が好ましく、７．５〜７．８が更に好ましい。
【００２６】
培養温度に関しては、２０〜４０℃の範囲であれば増殖スピードが速く、２５〜３５℃であればさらに好ましい。
【００２７】
なお、培養の過程において、ｐＨが低下するとともに、アルカリ度が減少する。すなわち、アンモニア酸化細菌によるＮＨ４ ＋のＮＯ２ −への酸化、亜硝酸酸化細菌によるＮＯ２ −のＮＯ３ −への酸化は次の（Ａ）（Ｂ）の２式で示される。なお、式（Ｃ）は、硝化細菌全体としての式である。
【００２８】
【化１】
これらより、ＮＨ４−ＮをＮＯ３−Ｎにまで酸化するのに４．５７ｍｇＯ２／ｍｇＮＨ４−Ｎの酸素を必要とし、硝化反応の進行に伴い水素イオンが放出されるため、培養系のｐＨが低下するとともにアルカリ度が減少することが分かる。ｐＨの低下に伴い培養速度が減少するため、緩衝液などを使ってｐＨを所定値に保持しなければ、従来法と同様、微生物の活動が停止してしまう。
【００３０】
そこで本発明では、培養過程において酸性側に傾くｐＨを、非水素化物と水素化物とを混合物にして緩衝作用を備えたものが好適であると考え、具体的な化合物を見い出すべく、多数の化合物によるトライ・アンド・エラーを重ねた結果、炭酸ナトリウムと炭酸水素ナトリウムとの組み合わせよりなる培養促進剤の投入によってｐＨを復帰させることが最も好適であることを見い出した。
【００３１】
一般に、細菌細胞の合成反応が次式で表現できることが知られている。
【００３２】
【化２】
これを、上記した混合培養系（Ｃ）の生化学反応式に適用すると、およそ次のようになる。
【００３４】
【化３】
上式から明らかなように、硝化細菌の培養には、エネルギー基質のアンモニウムイオンと比較しても多量の炭素源が必要である。
【００３６】
前述したように、炭酸ナトリウムと炭酸水素ナトリウムとの組み合わせよりなる培養促進剤を供給することにより、硝化細菌の炭酸同化のための炭素源を同時に供給することができる。以下、説明を加える。
【００３７】
炭酸ナトリウムのみを用いると、当該炭酸ナトリウムが強アルカリであることからしても、低下するｐＨを上げる効果は充分に認められるが、ｐＨ上昇の効果が大きいために多量には使用できず、充分な炭素源を供給するには不向きな点がある。他方、炭酸水素ナトリウムのみを用いる場合には、無機炭素源としての供給という点では問題がないものの、ｐＨを保持するという点では大量の供給が必要となり好ましくない。
【００３８】
このような長短所に鑑み、炭酸ナトリウムと炭酸水素ナトリウムとの混合物が好適に利用できる。当該混合物の水溶液を用いることによって、漸次低下していくｐＨを一定に保ちつつ、生体の炭酸同化のための無機炭素源を有効に供給することが可能となった。
【００３９】
前記混合物における炭酸ナトリウムと炭酸水素ナトリウムの配合割合としては、炭酸ナトリウム：炭酸水素ナトリウムが、モル比で４〜７：４〜８（１：４／７〜２）であることが好適であり、具体的には、炭酸ナトリウム０．４〜０．７（ｍｏｌ／リットル）と炭酸水素ナトリウム０．４〜０．８（ｍｏｌ／リットル）の混合水溶液が効果的である。
【００４０】
なお、培養系のｐＨの監視は、連続的に行なってもよいし、所定時間ごとに行なってもよい。ｐＨコントローラー等の連続ｐＨ監視装置を利用することが好ましいが、これに限らず、フェノールレッド等のｐＨ指示薬を利用して手作業で行なうことも可能である。
【００４１】
ＮＨ４−Ｎ含有液におけるアンモニアの濃度は１００ｍｇ／リットル以上であって３００ｍｇ／リットル以下、さらには２００ｍｇ／リットル以下に抑えることが好ましい。アンモニアは、化学独立栄養細菌であるアンモニア酸化細菌が炭酸同化を行って生育する際のエネルギー源であるが、過剰にあると、むしろ生育・増殖の阻害になる場合がある。また、アンモニア酸化細菌と同様に活性汚泥に含まれる亜硝酸酸化細菌は、アンモニア酸化細菌によって生成した亜硝酸を引き継いで酸化する細菌であるという割りには高い亜硝酸濃度に弱いため、アンモニアの初期濃度を無闇に高く設定することはできない。従って、アンモニアの濃度が３００ｍｇ／リットルを超える場合には、適宜、海水あるいは淡水などで希釈することが好適である。
【００４２】
なお、ＮＨ４−Ｎ含有液として、水処理場内にて発生する汚泥脱水濾液や消化脱離液などの汚泥処理廃液を利用することが好ましい。
【００４３】
上記した培養条件で、活性汚泥を、汚泥脱水濾液や消化脱離液などの汚泥処理廃液により硝化馴養することにより当該活性汚泥にわずかに含まれる硝化細菌を高濃度に培養することができるわけであるが、これに加え、本発明によれば、活性汚泥を２ヶ月で１／３〜１／４に減容することができ、かつ比重の大きい硝化汚泥を得ることができる。
【００４４】
すなわち、そもそも活性汚泥中には硝化細菌が０．３５％程度含有するといわれている。このような活性汚泥を原料としてＮＨ４−Ｎ含有液により約２ヶ月間、活性汚泥を馴養集積することにより、当該活性汚泥中の硝化細菌の含有率が、約１０倍（３．５％）に増加する。その過程において、他の雑菌は、外部から栄養源（エサ）が与えられないことから共食いし死滅していく。そしてこの結果として、活性汚泥が減量する（減容される）。
【００４５】
雑菌がほとんど死に絶えると、“グラニュー（粒）”と呼ばれる比重の大きい難分解性有機物となり、これを核として周囲に硝化細菌が取り付く。硝化細菌が取り付いた難分解性有機物は、比重の大きさから培養系において沈降する。硝化細菌を高濃度に培養するには、この沈降性の良さが必要となる。すなわち、一般的にいって硝化細菌は比重が軽く、純粋培養では浮遊してしまう。そのため、硝化細菌は培養系から流れ出てしまう可能性が高く、高濃度培養は期待できない。これにより、高濃度の培養には、上記したような核（難分解性有機物）の生成が必要になるわけであるが、核の生成は、硝化細菌の純粋培養では見られず、活性汚泥を原料としたときにのみ見られる。
【００４６】
【実施例】
以下、本発明の一実施例を挙げて説明するが、本発明はこれによって限定するものではない。
【００４７】
実施例１（硝化細菌の高濃度培養［硝化活性汚泥の製造］）
図１に示すｆｉｌｌ ａｎｄ ｄｒａｗ式培養槽（３０リットル）で２日サイクルの回分培養を行った。すなわち、海水希釈し尿汚泥、及び嫌気性消化脱離液（ＮＨ４−Ｎの濃度が１００ｍｇ／リットルとなるように海水希釈されている）を培養槽に入れ、培養槽内温度をサーモスタットとヒータで２７℃となるように、またｐＨをｐＨコントローラーおよび培養促進剤（１ＮのＮａＨＣＯ３、および０．５ＮのＮａ２ＣＯ３からなる緩衝剤）により７．５〜８．５に保つように設定して培養を行った（初期のｐＨが８．５以上の時は、希硫酸を加えて８．５以下に調整する）。また、溶存酸素（ＤＯ）濃度が４ｍｇ／リットルになるように散気球で曝気量を調節した。
【００４８】
曝気開始１日後に終濃度が１００ｍｇ／リットルとなるように再び消化脱離液を添加した。また、２日目には曝気を止め、１時間汚泥を沈殿させ上澄液を除去した後、消化脱離液を投入し、曝気を再開するという運転を繰り返した。
【００４９】
元の海水希釈し尿汚泥の塩分濃度が海水比８０％にあたるため、馴養は海水比８０％から開始し、１００ｍｇ／リットルのＮＨ４−Ｎが培養１日後に完全にＮＯ３−Ｎに硝化されるようになった段階で海水比を１００％に上げた。
【００５０】
曝気開始後の数時間はＮＨ４−Ｎ濃度が直線的に減少するため、曝気開始から、０，１，２，３，４時間後の残存ＮＨ４−Ｎ濃度を測定し、ＮＨ４−Ｎ濃度が直線的に変化する区間の傾きから変化速度を求め、これをＭＬＳＳ濃度で除した値を硝化速度とした（下記式参照）。
【００５１】
【数１】
約６０日間の海水馴養期間を経て馴養が完了した海水馴養硝化活性汚泥（ＡＭＮＳ）のＳＶ３０、ＳＶＩを測定し、沈降特性を調べるとともに、光学顕微鏡を用いてフロック形成状況を観察した。
【００５３】
図２のグラフに、し尿汚泥の海水馴養過程を示す（図には、濃度１００ｍｇ／リットルのＮＨ４−Ｎの４時間後の濃度を示した）。
【００５４】
図２のグラフから、馴養開始２ヶ月後には、海水比１００％で１００ｍｇ／リットルのＮＨ４−Ｎを４時間でほぼ完全に硝化できるＡＭＮＳを調製することができることが分かる。無機炭素源の不足による硝化活性汚泥の損失を防ぐべく、ＮａＨＣＯ３とＮａ２ＣＯ３を組み合わせた無機炭素源によるｐＨ調整を採用したが、これにより、馴養２ヶ月後には、図３に示すように、ＡＭＮＳのＭＬＳＳ濃度を馴養前と比べ２倍に増やすことができた。
【００５５】
ＡＭＮＳにおける硝化細菌の硝化速度を下記表に示す。
【００５６】
【表１】
【００５７】
培養槽を静置させると、細菌フロックが確認でき、比重が海水より重たいため大半の細菌フロックが沈澱する。これは、アンモニア酸化細菌と亜硝酸酸化細菌の各菌それぞれの純粋培養では見られないが、活性汚泥を原料とした混合培養において現れるものである。ＡＭＮＳのフロックを顕微鏡で観測した結果、汚泥は直径５０〜１００μｍのフロックからなることが分かった。また、ＡＭＮＳを走査型電子顕微鏡（ＳＥＭ）で観測した結果、汚泥フロック内部に２０〜１００μｍの糸状菌や粘着質物からなるグラニュールが含まれていることが分かった。そして、ＡＭＮＳのＳＶ３０、ＳＶＩを求めたところ、上記表１に併記した通り、９％、４２．６となり、沈降性に優れていることが分かった。これにより、海水希釈のし尿処理場汚泥から、海水中で（もちろん、淡水中でも）高い活性を持つ硝化活性汚泥を極めて短期間で大量生産できることが明らかとなった。
【００５８】
アンモニア酸化細菌と亜硝酸酸化細菌の個々の純粋培養と比較して、活性汚泥を原料とした混合培養の長所は次のとおりである。
【００５９】
純粋培養系では、エネルギー基質としてそれぞれアンモニア、亜硝酸が別個に必要であるが、混合培養系ではエネルギー基質としてアンモニアだけ供給すれば済む。
【００６０】
また、純粋培養系では、菌体を高濃度に増殖することが極めて困難であるが、混合培養系では培地を懸濁させる程度まで増殖することが容易である。これは、アンモニア酸化細菌や亜硝酸酸化細菌が同族内などではコロニーやフロックを形成しないで、浮遊生活を送る生態に起因すると推察される。
【００６１】
なお、このようにして得た海洋性硝化細菌は工業技術院生命工学工業技術研究所に寄託されている（寄託番号：ＦＥＲＭ ＢＰ−７１５０，識別表示：ＢＩＣＯＭ Ｎｉｔｒｉｆｙｉｎｇ Ｂａｃｔｅｒｉａ ＳＷＡＱ ＳＰ−７８）。
【００６２】
実施例２（促進剤の組成比を変えた馴養実験）
上記した実施例１の馴養実験を更に引き続き６ヶ月（合計８ヶ月）行なって、馴養後のＡＭＮＳのＭＬＳＳ濃度を測定した。結果を図４に示す。
【００６３】
培養促進剤の配合比（Ｎａ２ＣＯ３（０．５Ｎ）、ＮａＨＣＯ３（１Ｎ））を変えて同様の馴養実験を８ヶ月行ない、ＭＬＳＳ濃度がどれほど上昇するかを測定した。結果を、図４に併記する。
【００６４】
実施例３（促進剤の組成を変えた馴養実験）
培養促進剤の種類（組成）［ＮａＨＣＯ３、Ｎａ２ＣＯ３］を変えたという以外は、実施例２と同様に、硝化細菌の高濃度培養［硝化活性汚泥の製造］を行なった。馴養後のＡＭＮＳのＭＬＳＳ濃度を図５に示す。
【００６５】
実施例４（硫黄酸化細菌の高濃度培養）
基本的には、上記の硝化細菌の高濃度培養と同じ培養方法に従えばよいので、以下では簡単に述べる。
【００６６】
５リットルのビーカーに下水汚泥処理場返送汚泥を、ＭＬＳＳ濃度が１３００ｍｇ／Ｌになるように入れ、下記［表２］の硫黄酸化細菌培地で１日サイクルの反復半回分制限操作法（ＣＦ−ＲＯＭ法）にて硫黄酸化汚泥の馴養を行なった。培養温度はヒーターとサーモスタットで２５〜２７℃に保温し、ｐＨは、ＮａＨＣＯ３とＮａ２ＣＯ３の混合液（モル比でＮａＨＣＯ３：Ｎａ２ＣＯ３＝１．０：０．５）で７．０〜７．８の範囲内に調整した。
【００６７】
【表２】
時間の経過とともに増加するＭＬＳＳ濃度を図６に示す。
【００６９】
【発明の効果】
本発明の促進剤により、従来は不可能とされてきた独立栄養細菌を大量に、かつ高濃度に培養することができる。
【図面の簡単な説明】
【図１】汚泥硝化馴養装置の一例を示した略示説明図である。
【図２】汚泥の硝化馴養過程においてＮＨ４−Ｎ濃度の変化を示したグラフ図である。
【図３】汚泥の硝化馴養過程においてＭＬＳＳ濃度の経日的変化を示したグラフ図である。
【図４】培養促進剤の組成配合の割合を変えて８ヶ月間、馴養を行なったＭＬＳＳ濃度の変化を示すグラフ図である。
【図５】培養促進剤の種類を変えて馴養を行なったＭＬＳＳ濃度の変化を示すグラフ図である。
【図６】汚泥中の硫黄酸化細菌の馴養過程においてＭＬＳＳ濃度の経日的変化を示したグラフ図である。[0001]
BEST MODE FOR CARRYING OUT THE INVENTION
The present invention relates to an autotrophic bacterium high-concentration culture promoter for culturing autotrophic bacteria at a high concentration (hereinafter, also simply referred to as “promoter”). The present invention relates to an accelerator for culturing autotrophic bacteria that cause a severe reaction at a high concentration.
[0002]
[Prior Art and Problems to be Solved by the Invention]
Autotrophic bacteria, such as ammonia-oxidizing bacteria (nitrifying bacteria) and sulfur-oxidizing bacteria, have a slow growth rate and do not live as colonies. There is no report that high-density cultivation was successful in large quantities.
[0003]
That is, the conventional culture method is a small-scale test tube-level one aiming at pure culture, and the medium in the flask is not suspended in about two months, and cannot be said to be an industrially applicable culture method. Was.
[0004]
In addition, for example, when nitrification starts in the culture of nitrifying bacteria, the pH decreases. However, conventionally, there has been no known method for effectively increasing the pH.
[0005]
On the other hand, although the carbon source decreases with nitrification, carbon dioxide has conventionally been used as a carbon source supply. Although the supply of carbon dioxide can certainly prevent the depletion of the carbon source, the above-mentioned lowering of the pH has further progressed, and the activity of nitrifying bacteria has stopped, and this has been the limit of bacterial growth.
[0006]
[Object of the invention]
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.
[0007]
[Means for Solving the Problems]
The autotrophic bacterium high concentration cultivation promoter according to
[0008]
An autotrophic bacterium high-concentration culture promoter according to
[0009]
(A) A basic substance that is soluble in water and shows basicity when dissociated.
[0010]
(B) Carbonates soluble in water.
[0011]
The autotrophic bacterium high concentration culture promoter according to
[0012]
The autotrophic bacterium high-concentration culture promoter according to
[0013]
An autotrophic bacterium high-concentration culture promoter according to
[0014]
BEST MODE FOR CARRYING OUT THE INVENTION
Autotrophic bacteria The autotrophic bacteria in the present invention refer to all bacteria that cause a reaction such that the pH of the medium decreases during culturing, and specifically, ammonia-oxidizing bacteria (nitrifying bacteria), Examples include sulfur oxidizing bacteria, carbon monoxide oxidizing bacteria, and a mixture of two or more thereof.
[0015]
These bacteria are contained, for example, in activated sludge, seawater and the like, but if anything, it is preferable to use activated sludge in that it contains more bacteria.
[0016]
Accelerator component The accelerating agent component of the present invention maintains the pH of a medium which exhibits basicity by dissociation and tilts 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 is a compound having two properties, that is, a property that can be caused and a property that can be a carbon source during the growth of the autotrophic bacterium.
[0017]
When the above two properties can be imparted by one kind of compound, at least one kind of the compound becomes a component of the accelerator of the present invention.
[0018]
Specific examples thereof include trisodium bicarbonate (= sodium sesquicarbonate) (Na 2 CO 3 .NaHCO 3 .2H 2 O), and alkali or potassium in this compound is partially or entirely replaced by potassium, magnesium, or the like. And the like.
[0019]
When the above two properties can be imparted by two or more compounds, a mixture of at least two of the compounds is a component of the accelerator of the present invention. For example, a mixture of (A) an alkali which is soluble in water and exhibits basicity by dissociation and (B) a carbonate which is soluble in water can be mentioned. Specifically, as the component (A), water is used. Examples thereof include hydroxides such as sodium oxide and potassium hydroxide, and alkali salts such as sodium carbonate and sodium sulfate. As the component (B), sodium hydrogen carbonate and magnesium hydroxide (4MgCO 3 .Mg (OH) 2 .5H 2 O).
[0020]
Among them, the component (A) is an alkali carbonate or an alkaline earth metal salt which is soluble in water, such as sodium carbonate, sodium potassium carbonate and potassium carbonate, and the component (B) is sodium hydrogencarbonate. , Potassium bicarbonate and the like are preferably water-soluble alkali hydrogen carbonate or alkaline earth metal bicarbonate in that the two properties described above can be effectively imparted. And sodium bicarbonate are most preferred.
[0021]
Activated sludge The activated sludge used in the present invention includes sewage sludge and human waste sludge. These may be those that have been subjected to fresh water dilution treatment, or may be those that have been subjected to sea water dilution treatment.However, if the autotrophic bacteria are cultured using seawater dilution sludge as a raw material, they are considered to be rare. Since a large amount of marine bacteria (salt-resistant bacteria) can be obtained, it is preferable to use activated sludge that has been subjected to a seawater dilution treatment.
[0022]
To add an explanation, natural seawater contains marine bacteria that are considered to have higher salt tolerance than freshwater bacteria, but their abundance is very small and pure separation is difficult. Its research has been delayed compared to bacterium. However, according to the culture method of the present invention, a large amount of marine bacteria with a high concentration can be obtained by using activated sludge that has been subjected to seawater dilution treatment as described above. Marine bacteria have multi-layered cell walls and have strong resistance to various chemical substances that inhibit osmotic pressure change and growth of treated water.
[0023]
Culture (cultivation) conditions of nitrifying bacteria Culture of nitrifying bacteria contained in the activated sludge is performed by subjecting the activated sludge to a sludge dewatering filtrate or (anaerobic) for a predetermined period (for example, 1 month, 2 months or 3 months). ) Nitrification is performed by sludge treatment waste liquid such as digestion and desorption liquor. Since nitrification must be performed aerobically, the dissolved oxygen (DO) must be 2 mg / liter or more. is there. However, for the first time in this experiment, it was found that if the dissolved oxygen concentration was too high, the growth speed would tend to decrease. The details will be described below.
[0024]
The rate of nitrification by nitrifying bacteria is higher as the dissolved oxygen is higher, so it was thought that the higher the dissolved oxygen, the faster the nitrification accumulates. It was found that the speed of acclimatization and accumulation of nitrifying bacteria used as a raw material was reduced at a place where the dissolved oxygen (DO) exceeded 5 mg / liter. Note that the dissolved oxygen (DO) concentration is most preferably 2 to 4 mg / liter.
[0025]
In addition, the pH needs to be 7.0 to 9.0 (especially when activated sludge diluted with seawater is used), preferably 7.5 to 8.5, more preferably 7.5 to 7.8.
[0026]
Regarding the cultivation temperature, the growth speed is high in the range of 20 to 40 ° C, and more preferably 25 to 35 ° C.
[0027]
In the course of the culture, the alkalinity decreases as the pH decreases. That is, the oxidation of NH 4 + to NO 2 − by ammonia oxidizing bacteria and the oxidation of NO 2 − to NO 3 − by nitrite oxidizing bacteria are represented by the following two equations (A) and (B). Expression (C) is an expression for the entire nitrifying bacteria.
[0028]
Embedded image
From these, 4.57 mg O 2 / mg NH 4 —N oxygen is required to oxidize NH 4 —N to NO 3 —N, and hydrogen ions are released with the progress of the nitrification reaction. It can be seen that the alkalinity decreases as the pH decreases. Since the cultivation rate decreases as the pH decreases, unless the pH is maintained at a predetermined value using a buffer or the like, the activity of the microorganism stops as in the conventional method.
[0030]
Therefore, in the present invention, it is considered that a compound having a buffering action as a mixture of a non-hydride and a hydride is preferable in terms of the pH that tilts toward the acidic side during the culture process, As a result of repeated trial-and-error tests, it has been found that it is most preferable to restore the pH by adding a culture promoter composed of a combination of sodium carbonate and sodium bicarbonate.
[0031]
Generally, it is known that the synthesis reaction of bacterial cells can be expressed by the following equation.
[0032]
Embedded image
When this is applied to the biochemical reaction formula of the mixed culture system (C) described above, it is approximately as follows.
[0034]
Embedded image
As is clear from the above formula, culturing nitrifying bacteria requires a large amount of carbon source as compared with ammonium ion as an energy substrate.
[0036]
As described above, by supplying a culture promoter comprising a combination of sodium carbonate and sodium bicarbonate, a carbon source for carbonic assimilation of nitrifying bacteria can be simultaneously supplied. Hereinafter, an explanation will be added.
[0037]
When only sodium carbonate is used, the effect of increasing the decreasing pH is sufficiently recognized even from the fact that the sodium carbonate is a strong alkali, but the effect of increasing the pH is large, so that it cannot be used in a large amount. There are some points that are not suitable for supplying a good carbon source. On the other hand, when only sodium hydrogen carbonate is used, 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.
[0038]
In view of such advantages and disadvantages, a mixture of sodium carbonate and sodium hydrogen carbonate can be suitably used. By using an aqueous solution of the mixture, it has become possible to effectively supply an inorganic carbon source for carbonic acid assimilation of a living body while keeping a gradually decreasing pH constant.
[0039]
As a blending ratio of sodium carbonate and sodium hydrogen carbonate in the mixture, it is preferable that the molar ratio of sodium carbonate: sodium hydrogen carbonate is 4 to 7: 4 to 8 (1: 4/7 to 2), Specifically, a mixed aqueous solution of sodium carbonate 0.4 to 0.7 (mol / liter) and sodium hydrogen carbonate 0.4 to 0.8 (mol / liter) is effective.
[0040]
The monitoring of the pH of the culture system may be performed continuously or may be performed 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, and it is also possible to perform the measurement manually using a pH indicator such as phenol red.
[0041]
The concentration of ammonia in the NH 4 —N-containing liquid is preferably 100 mg / L or more and 300 mg / L or less, and more preferably 200 mg / L or less. Ammonia is an energy source for the growth of ammonium oxidizing bacteria, which are chemolithotrophic bacteria, by performing carbon assimilation. However, if it is excessive, it may rather hinder growth and growth. In addition, nitrite oxidizing bacteria contained in activated sludge, like ammonia oxidizing bacteria, are weak against high nitrite concentration, because they are bacteria that take over and oxidize nitrite generated by ammonia oxidizing bacteria. The concentration cannot be set unnecessarily high. Therefore, when the concentration of ammonia exceeds 300 mg / liter, it is preferable to appropriately dilute it with seawater or fresh water.
[0042]
In addition, it is preferable to use sludge treatment waste liquid such as sludge dehydration filtrate or digestion / desorption liquid generated in the water treatment plant as the NH 4 —N-containing liquid.
[0043]
Under the above culture conditions, the activated sludge can be cultured at a high concentration of nitrifying bacteria slightly contained in the activated sludge by nitrifying and acclimating the sludge with a sludge treatment waste liquid such as a sludge dewatering filtrate or a digestion / desorption liquid. However, in addition to this, according to the present invention, the activated sludge can be reduced in volume to 1/3 to 1/4 in two months, and nitrified sludge having a large specific gravity can be obtained.
[0044]
That is, it is said that activated sludge contains nitrifying bacteria at about 0.35% in the first place. By accumulating the activated sludge using such activated sludge as a raw material with an NH 4 —N-containing liquid for about 2 months, the content of nitrifying bacteria in the activated sludge is increased by about 10 times (3.5%). To increase. In the process, other germs cannihilate and die because no external nutrients (feeds) are provided. As a result, the amount of activated sludge is reduced (volume is reduced).
[0045]
When the germs almost die, they become hard-to-decompose organic matter having a large specific gravity called "granules", and nitric bacteria attach to the surroundings using the nucleus as a core. The hardly decomposable organic matter attached to the nitrifying bacteria precipitates in the culture system due to its specific gravity. In order to culture nitrifying bacteria at a high concentration, good sedimentation is required. That is, 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-density culture cannot be expected. As a result, the production of nuclei (refractory organic substances) as described above is necessary for high-concentration culture, but the production of nuclei is not seen in pure culture of nitrifying bacteria, and activated sludge is produced. Only found when used as a raw material.
[0046]
【Example】
Hereinafter, an example of the present invention will be described, but the present invention is not limited thereto.
[0047]
Example 1 (High concentration culture of nitrifying bacteria [production of nitrifying activated sludge])
Batch culture in a two-day cycle was performed in a fill and draw type culture tank (30 liters) shown in FIG. That is, urine sludge diluted with seawater, and an anaerobic digestion / elimination liquid (diluted in seawater so that the concentration of NH 4 -N becomes 100 mg / liter) are put into a culture tank, and the temperature in the culture tank is adjusted with a thermostat and a heater. The temperature was set at 27 ° C. and the pH was maintained at 7.5 to 8.5 by a pH controller and a culture promoter (a buffer consisting of 1N NaHCO 3 and 0.5N Na 2 CO 3 ). (When the initial pH was 8.5 or more, the pH was adjusted to 8.5 or less by adding dilute sulfuric acid). Further, the amount of aeration was adjusted with a balloon so that the dissolved oxygen (DO) concentration became 4 mg / liter.
[0048]
One day after the start of aeration, the digestion / elimination liquid was added again so that the final concentration became 100 mg / liter. On the second day, the aeration was stopped, the sludge was settled for 1 hour, the supernatant was removed, the digestion / desorption solution was added, and the operation of restarting the aeration was repeated.
[0049]
Since the salinity of the original seawater diluted excrement sludge hits the 80% seawater ratio, acclimatization starts at 80% seawater ratio, NH 4 -N of 100mg / liter is nitrification completely NO 3 -N after one day culture At this stage, the seawater ratio was increased to 100%.
[0050]
Since the NH 4 —N concentration decreases linearly for several hours after the start of aeration, the remaining NH 4 —
[0051]
(Equation 1)
SV30 and SVI of the seawater acclimated nitrifying activated sludge (AMNS), which had been acclimatized after a seawater acclimation period of about 60 days, were measured, sedimentation characteristics were examined, and the state of floc formation was observed using an optical microscope.
[0053]
The graph of FIG. 2 shows the process of acclimating human sludge to seawater (the figure shows the concentration of NH 4 —N at a concentration of 100 mg / liter after 4 hours).
[0054]
From the graph of FIG. 2, it can be seen that two months after the start of acclimation, AMNS capable of almost completely nitrifying 100 mg / liter of NH 4 —N at a seawater ratio of 100% in 4 hours can be prepared. In order to prevent the loss of nitrification activated sludge due to the shortage of the inorganic carbon source, pH adjustment by the inorganic carbon source in which NaHCO 3 and Na 2 CO 3 were combined was adopted. In addition, the MLSS concentration of AMNS could be increased twice as much as before the acclimation.
[0055]
The nitrification rate of nitrifying bacteria in AMNS is shown in the table below.
[0056]
[Table 1]
[0057]
When the culture tank is allowed to stand, bacterial flocs can be confirmed, and most of the bacterial flocs precipitate because the specific gravity is heavier than seawater. This is not seen in the pure culture of each of the ammonia-oxidizing bacteria and the nitrite-oxidizing bacteria, but appears in the mixed culture using activated sludge as a raw material. As a result of observing the AMNS floc with a microscope, it was found that the sludge was composed of flocs having a diameter of 50 to 100 µm. Further, as a result of observing the AMNS with a scanning electron microscope (SEM), it was found that the sludge flocs contained 20 to 100 μm granules made of filamentous fungi or sticky substances. And when SV30 and SVI of AMNS were calculated | required, it was set to 9% and 42.6 as described in Table 1, and it turned out that it is excellent in sedimentation property. Thus, it was revealed that nitrification-activated activated sludge having high activity in seawater (of course, even in freshwater) can be mass-produced in a very short period of time from seawater-diluted human waste treatment plant sludge.
[0058]
The advantages of the mixed culture using activated sludge as a raw material as compared with the individual pure cultures of ammonia oxidizing bacteria and nitrite oxidizing bacteria are as follows.
[0059]
In a pure culture system, 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.
[0060]
In a pure culture system, it is extremely difficult to grow cells at a high concentration, but in a mixed culture system, it is easy to grow to the extent that the medium is suspended. This is presumed to be due to the ecology of ammonia floating bacteria and nitrite oxidizing bacteria not forming colonies and flocs in their cognate and living in a floating life.
[0061]
The marine nitrifying bacteria thus obtained have been deposited with the National Institute of Bioscience and Human-Technology, National Institute of Advanced Industrial Science and Technology (Deposit No .: FERM BP-7150, identification: BICOM Nitrifying Bacteria SWAQ SP-78).
[0062]
Example 2 (acclimation experiment in which the composition ratio of the accelerator was changed)
The acclimatization experiment of Example 1 described above was further continued for 6 months (total 8 months), and the MLSS concentration of AMNS after acclimation was measured. FIG. 4 shows the results.
[0063]
The same habituation experiment was performed for 8 months while changing the mixing ratio of the culture promoter (Na 2 CO 3 (0.5N), NaHCO 3 (1N)) to measure how much the MLSS concentration increased. The results are also shown in FIG.
[0064]
Example 3 (acclimation experiment in which the composition of the accelerator was changed)
Except that the type (composition) of the culture promoter (NaHCO 3 , Na 2 CO 3 ) was changed, high-density cultivation of nitrifying bacteria [production of nitrifying activated sludge] was performed in the same manner as in Example 2. FIG. 5 shows the MLSS concentration of AMNS after acclimation.
[0065]
Example 4 (High concentration culture of sulfur oxidizing bacteria)
Basically, the same culturing method as in the above-described high-density culturing of nitrifying bacteria may be used, and therefore, a brief description will be given below.
[0066]
The sludge returned from the sewage sludge treatment plant is put into a 5 liter beaker so that the MLSS concentration becomes 1300 mg / L, and a one-day cycle repetitive half-batch limiting operation method (CF-ROM) is performed using a sulfur-oxidizing bacterial medium shown in [Table 2] below. Method) to acclimatize the sulfur oxidation sludge. The culture temperature was maintained at 25 to 27 ° C. with a heater and a thermostat, and the pH was 7 with a mixed solution of NaHCO 3 and Na 2 CO 3 (NaHCO 3 : Na 2 CO 3 = 1.0: 0.5 in molar ratio). It adjusted within the range of 0.0-7.8.
[0067]
[Table 2]
FIG. 6 shows the MLSS concentration increasing with time.
[0069]
【The invention's effect】
The promoter of the present invention enables cultivation of autotrophic bacteria, which have been impossible so far, in large amounts and at high concentrations.
[Brief description of the drawings]
FIG. 1 is a schematic explanatory view showing an example of a sludge nitrification acclimation apparatus.
FIG. 2 is a graph showing a change in NH 4 —N concentration in a nitrification acclimation process of sludge.
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 habituation for 8 months by changing the composition ratio of the culture promoter.
FIG. 5 is a graph showing the change in MLSS concentration after habituation was performed by changing the type of culture promoter.
FIG. 6 is a graph showing the daily change of MLSS concentration in the process of acclimating sulfur oxidizing bacteria in sludge.
Claims (5)
解離することにより塩基性を呈して前記酸性側に傾く培地のｐＨを所定範囲に維持させることのできる性質と、前記独立栄養細菌の増殖時の炭素源となりうる性質との２つの性質を、１種類の化合物で以て付与できる場合は当該化合物の少なくとも１種、または上記２つの性質を２種以上の化合物で以て付与できる場合は、当該化合物の少なくとも２種が混合された混合物
が配合されてなることを特徴とする独立栄養細菌高濃度培養促進剤。An autotrophic bacterium high-concentration cultivation promoter that becomes a carbon source while maintaining the pH of the medium that tilts to the acidic side in the acclimatization process in a predetermined range when acclimatizing the autotrophic bacterium at a high concentration for a predetermined period,
The two properties of a medium that exhibits basicity by dissociation and can maintain the pH of the medium inclined toward the acidic side within a predetermined range, and a property that can serve as a carbon source during growth of the autotrophic bacterium are as follows. In the case of being able to be provided by two kinds of compounds, at least one kind of the compound is used, or in the case where the above two properties can be provided by two or more kinds of compounds, a mixture of at least two kinds of the compound is mixed. A high-concentration autotrophic bacterium cultivation promoter, comprising:
（Ａ）水に可溶で、解離することにより塩基性を示す塩基性物質。
（Ｂ）水に可溶性を示す炭酸塩。The said mixture is a mixture of the following (A) component and (B) component, The autotrophic bacterium high concentration culture promoter of Claim 1 characterized by the above-mentioned.
(A) A basic substance that is soluble in water and shows basicity when dissociated.
(B) Carbonates soluble in water.
前記（Ｂ）成分が炭酸水素アルカリ塩又は炭酸水素アルカリ土類金属塩であることを特徴とする請求項２に記載の独立栄養細菌高濃度培養促進剤。The component (A) is an alkali carbonate or an alkaline earth metal carbonate,
3. The high-concentration autotrophic bacterium cultivation promoter according to claim 2, wherein the component (B) is an alkali hydrogen carbonate or an alkaline earth metal bicarbonate.
前記（Ｂ）成分が、炭酸水素ナトリウムである
ことを特徴とする請求項２〜４のいずれか１項に記載の独立栄養細菌高濃度培養促進剤。The component (A) is sodium carbonate,
The said (B) component is sodium hydrogen carbonate, The autotrophic bacterium high concentration culture promoter of any one of Claims 2-4 characterized by the above-mentioned.
Priority Applications (3)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP2000375317A JP2004283001A (en) | 2000-12-08 | 2000-12-08 | Promotor for culturing autotrophic bacterium at high concentration |
| PCT/JP2001/010295 WO2002046370A1 (en) | 2000-12-08 | 2001-11-26 | Promoter for the culture of autrotrophic bacterium at high density |
| AU2002224102A AU2002224102A1 (en) | 2000-12-08 | 2001-11-26 | Promoter for the culture of autrotrophic bacterium at high density |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP2000375317A JP2004283001A (en) | 2000-12-08 | 2000-12-08 | Promotor for culturing autotrophic bacterium at high concentration |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| JP2004283001A true JP2004283001A (en) | 2004-10-14 |
Family
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| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| JP2000375317A Pending JP2004283001A (en) | 2000-12-08 | 2000-12-08 | Promotor for culturing autotrophic bacterium at high concentration |
Country Status (3)
| Country | Link |
|---|---|
| JP (1) | JP2004283001A (en) |
| AU (1) | AU2002224102A1 (en) |
| WO (1) | WO2002046370A1 (en) |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JP2008307459A (en) * | 2007-06-13 | 2008-12-25 | Shibaura Institute Of Technology | Purification method of contaminated seawater by seawater-derived microorganisms |
| JP2010284617A (en) * | 2009-06-15 | 2010-12-24 | Eidensha:Kk | Bioreactor element, method for producing the same and method for using the same |
Families Citing this family (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| ITMI20061843A1 (en) * | 2006-09-27 | 2008-03-28 | Immobiliare G M S R L | FORMULATIONS OF MEDIUM CULTURALS SUITABLE FOR INDUSTRIAL APPLICATION |
Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPS59232087A (en) * | 1983-06-13 | 1984-12-26 | Mitsubishi Gas Chem Co Inc | Cultivation of anaerobic bacteria |
| DE4216357C1 (en) * | 1992-05-18 | 1993-12-09 | Nitrochemie Gmbh | New denitrifying bacterium - useful for degrading explosive nitrate ester(s) after alkaline hydrolysis |
| JPH08252088A (en) * | 1995-03-17 | 1996-10-01 | Nippo Kagaku Kk | Method for culturing bacteria in clostridium |
| JPH10295285A (en) * | 1997-05-01 | 1998-11-10 | Kurorera Kogyo Kk | Rotatoria feed |
Family Cites Families (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JP2989257B2 (en) * | 1990-11-29 | 1999-12-13 | 三菱重工業株式会社 | Nitric acid bacteria culture solution and high concentration culture method of the same |
| JP2989258B2 (en) * | 1990-11-30 | 1999-12-13 | 三菱重工業株式会社 | Nitrite bacteria culture solution and high concentration culture method of the bacteria |
| JP3136458B2 (en) * | 1994-08-24 | 2001-02-19 | 新日本製鐵株式会社 | How to grow iron-oxidizing bacteria |
| JP3749617B2 (en) * | 1998-04-17 | 2006-03-01 | 新日本製鐵株式会社 | Method of acclimatizing sulfur-oxidizing bacteria and method of removing nitrogen from wastewater using sulfur-oxidizing bacteria |
-
2000
- 2000-12-08 JP JP2000375317A patent/JP2004283001A/en active Pending
-
2001
- 2001-11-26 WO PCT/JP2001/010295 patent/WO2002046370A1/en not_active Ceased
- 2001-11-26 AU AU2002224102A patent/AU2002224102A1/en not_active Abandoned
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPS59232087A (en) * | 1983-06-13 | 1984-12-26 | Mitsubishi Gas Chem Co Inc | Cultivation of anaerobic bacteria |
| DE4216357C1 (en) * | 1992-05-18 | 1993-12-09 | Nitrochemie Gmbh | New denitrifying bacterium - useful for degrading explosive nitrate ester(s) after alkaline hydrolysis |
| JPH08252088A (en) * | 1995-03-17 | 1996-10-01 | Nippo Kagaku Kk | Method for culturing bacteria in clostridium |
| JPH10295285A (en) * | 1997-05-01 | 1998-11-10 | Kurorera Kogyo Kk | Rotatoria feed |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JP2008307459A (en) * | 2007-06-13 | 2008-12-25 | Shibaura Institute Of Technology | Purification method of contaminated seawater by seawater-derived microorganisms |
| JP2010284617A (en) * | 2009-06-15 | 2010-12-24 | Eidensha:Kk | Bioreactor element, method for producing the same and method for using the same |
Also Published As
| Publication number | Publication date |
|---|---|
| WO2002046370A1 (en) | 2002-06-13 |
| AU2002224102A1 (en) | 2002-06-18 |
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