JPS6188872A - Method and apparatus for continuous cultivation at high concentration - Google Patents

Method and apparatus for continuous cultivation at high concentration

Info

Publication number
JPS6188872A
JPS6188872A JP59211747A JP21174784A JPS6188872A JP S6188872 A JPS6188872 A JP S6188872A JP 59211747 A JP59211747 A JP 59211747A JP 21174784 A JP21174784 A JP 21174784A JP S6188872 A JPS6188872 A JP S6188872A
Authority
JP
Japan
Prior art keywords
culture
filtration
membrane
tank
circulation circuit
Prior art date
Legal status (The legal status 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 status listed.)
Granted
Application number
JP59211747A
Other languages
Japanese (ja)
Other versions
JPS6363193B2 (en
Inventor
Eitaro Kumazawa
熊沢 栄太郎
Kisao Satou
佐藤 喜砂夫
Yoshitomo Minamide
南出 芳友
Shinichi Takato
高藤 愼一
Kazuo Imon
井門 和夫
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Snow Brand Milk Products Co Ltd
Original Assignee
Snow Brand Milk Products Co Ltd
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 Snow Brand Milk Products Co Ltd filed Critical Snow Brand Milk Products Co Ltd
Priority to JP59211747A priority Critical patent/JPS6188872A/en
Publication of JPS6188872A publication Critical patent/JPS6188872A/en
Publication of JPS6363193B2 publication Critical patent/JPS6363193B2/ja
Granted legal-status Critical Current

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  • Apparatus Associated With Microorganisms And Enzymes (AREA)
  • Micro-Organisms Or Cultivation Processes Thereof (AREA)

Abstract

PURPOSE:To enable the culture of microbial cells in high concentration, by supplying the culture medium to the second filtration membrane in the case of normal flow direction in the circulation circuit, and supplying to the first filtration membrane in the case of reverse flow and continuing the cultivation while removing the metabolic products from the medium. CONSTITUTION:A seed microorganism is inoculated in the culture tank 10, the cultivation is started, and when the concentration of the metabolic product reaches a specific level, the operation of membranes is started. In the case of the normal flow operation shown by the solid arrow, the cell liquid extracted from the culture tank 10 with the pump 37 is passed in the order of the first and the second filtration chambers 11, 12. The first filtration chamber 11 is operated under higher pressure, and the filtrate containing the metabolic product is introduced into the filtration chamber 27, the metabolic product is removed by the RO membrane, and the nutrient component having low molecular weight is returned to the medium supply tank 29. When the system is turned to the reverse-flow operation shown by the dotted arrow by the simultaneous change-over of the three-way solenoid valves 19, 20, 21, 23 and 32, the first filtration chamber is subjected to the back-washing, and is supplied with low-molecular nutrient component.

Description

【発明の詳細な説明】 (産業上の利用分野) 本発明は各種菌体を培養する高濃度培養方法及び装置に
関するものである。
DETAILED DESCRIPTION OF THE INVENTION (Industrial Field of Application) The present invention relates to a high concentration culture method and apparatus for culturing various bacterial cells.

(従来の技術) 一般に各種菌体を培養する場合、その代謝産物により増
殖阻害を受けることが多い。
(Prior Art) Generally, when culturing various types of bacterial cells, their growth is often inhibited by their metabolites.

そこで培養槽とUF膜の濾過膜を組合せて培養中に生産
される代謝産物を除き高濃度培養を達成しようとする試
みは実験室レベルではすでに行われている。
Therefore, attempts have already been made at the laboratory level to remove metabolites produced during culture and achieve high concentration culture by combining a culture tank and a UF membrane filtration membrane.

第2図のものは培養槽(1)と保持タンク(2)との間
に透析槽(3)を設は之等をポンプ(4)を通じて連結
してあり、代謝産物が透析膜を通じて取り除かれるよう
になっていて菌体液は循環するようになっている。
In the one in Figure 2, a dialysis tank (3) is installed between the culture tank (1) and the holding tank (2), which are connected through a pump (4), and metabolites are removed through the dialysis membrane. This allows the bacterial fluid to circulate.

これはJournal the 5ociety Da
iry TecnologyVol 30  NIL 
January 1979に掲載されている。
This is Journal the 5ociety Da
iry Technology Vol 30 NIL
Published in January 1979.

又The Au5tralian Journal o
f Dairy Tecnology−March 1
977には、次のようなことが掲載されている。すなわ
ち、第3図において第1RO膜(5)、第2RO膜(6
)をポンプ(7)と圧力制御弁(8)とを通じて原料タ
ンク(9)に連結してあり、各RO膜で代謝産物を取り
除き低分子栄養成分を回収し、原料タンク(9)に還元
されるようになっている。
Also, The Au5tralian Journal o
f Dairy Technology-March 1
977 contains the following information: That is, in FIG. 3, the first RO film (5) and the second RO film (6)
) is connected to the raw material tank (9) through a pump (7) and a pressure control valve (8), and each RO membrane removes metabolites and recovers low-molecular nutrients, which are then returned to the raw material tank (9). It has become so.

(発明が解決しようとする問題点) 以上のような従来の方法では透析膜より除去された低分
子栄養成分を培養槽に直接供給し、培養槽内の液レベル
を一定に保持し、培養を継続していた。
(Problems to be solved by the invention) In the conventional method as described above, the low molecular weight nutrients removed from the dialysis membrane are directly supplied to the culture tank, the liquid level in the culture tank is maintained constant, and the culture is continued. It was continuing.

しかしこの方法では培養経過に伴い、培養液の高粘度化
、膜面への菌体の目づまりにより液透過速度は急激に低
下し、培養槽内で生成した代謝物を十分に除ききれず、
したがって槽内に代謝物が蓄積され菌体の増殖が抑制さ
れるという欠点がある。
However, with this method, as the culture progresses, the liquid permeation rate rapidly decreases due to the increase in the viscosity of the culture solution and the clogging of the membrane surface with bacterial cells, and the metabolites produced in the culture tank cannot be removed sufficiently.
Therefore, there is a drawback that metabolites are accumulated in the tank and the growth of bacterial cells is suppressed.

(問題点を解決するための手段) したがって本発明の技術的課題は連続的に代謝物を除き
ながら培養を継続させ、連続的に高濃度の菌体を培養さ
せることのできる高濃度連続培養方法及び装置をうろこ
とを目的とするものである。
(Means for Solving the Problems) Therefore, the technical problem of the present invention is a high-concentration continuous culture method capable of continuously cultivating bacterial cells at a high concentration by continuously culturing while removing metabolites. and for measuring equipment.

この技術的課題を解決する本発明の技術的手段は、培養
槽で各種菌体を培養するに当たって、培養中に生産され
る代謝産物を培養槽と共に循環回路を構成する第1.2
濾過膜を通して取り除き、取り除いた代謝産物から低分
子有効成分を回収してこれを新鮮培地と共に前記濾過膜
を通して循環回路に供給するに当たり、循環回路の正流
方向では第2濾過膜に供給し、逆流方向では第1濾過膜
に供給して連続的に高濃度菌液をうるようにしたことと
、培養槽とUF膜等の濾過膜を使用した第1.2濾過室
とを配管装置を介して可逆循環回路を構成する如く連結
し、かつ第1,2濾過室は新鮮培地と代謝産物から回収
した低分子有効成分とを供給できる供給タンクに切換自
在に連結すると共に代謝産物の回収装置に連結し、循環
回路の正流方向では第2濾過室を供給タンクに連結し、
逆流方向では第1濾過室を供給タンクに連結する如く配
管構成されたものである。
The technical means of the present invention to solve this technical problem is that when culturing various types of bacterial cells in a culture tank, the metabolites produced during the culture are collected in the first and second channels, which constitute a circulation circuit together with the culture tank.
In order to recover low-molecular active ingredients from the removed metabolites and supply them to the circulation circuit through the filtration membrane together with a fresh medium, in the forward direction of the circulation circuit, they are supplied to the second filtration membrane, and in the reverse flow direction. In this direction, the first filtration membrane is supplied with a highly concentrated bacterial solution, and the culture tank and the 1.2 filtration chamber, which uses a filtration membrane such as a UF membrane, are connected via piping equipment. The first and second filtration chambers are connected to form a reversible circulation circuit, and the first and second filtration chambers are switchably connected to a supply tank capable of supplying fresh culture medium and low-molecular active ingredients recovered from metabolites, and are also connected to a metabolite recovery device. However, in the forward flow direction of the circulation circuit, the second filtration chamber is connected to the supply tank,
In the reverse flow direction, piping is configured to connect the first filtration chamber to the supply tank.

(発明の効果) 本発明では培養中に生産される代謝産物をUF膜等の濾
過膜を用いて系外に効率的に除去することができ、同伴
して流出する低分子有効成分及び新鮮培地を膜を通して
補充しながら培養し、連続的に高濃度菌液をうろことが
できる。
(Effects of the Invention) In the present invention, metabolites produced during culture can be efficiently removed from the system using a filtration membrane such as a UF membrane, and the low-molecular active ingredients and fresh medium that flow out together It is possible to culture the bacteria while replenishing it through the membrane, and to continuously circulate the highly concentrated bacterial solution.

すなわち、正流運転時には培養槽より菌体液を第1.2
濾過室の順に通過させ、代謝物を含む透過液を高圧側の
第1濾過室より糸外に排出せしめ、一方低圧側となる第
2濾過室には代謝物を除いた低分子栄養物及び新鮮培地
を系外より押込んで培養槽内のタンクレベルを一定に保
持することができる。
In other words, during normal flow operation, the bacterial cell liquid is transferred from the culture tank to the
The permeated liquid containing metabolites is discharged from the first filtration chamber on the high-pressure side to the outside of the thread through the filtration chambers in order, while low-molecular nutrients and fresh culture medium excluding metabolites are passed through the second filtration chamber on the low-pressure side. can be pushed in from outside the system to maintain a constant tank level inside the culture tank.

そして逆流運転時には第1濾過室と第2濾過室の高圧側
と低圧側とが逆転し、これまで透過液を系内から系外に
排出していた第1濾過室では代謝物を除いた低分子栄養
物及び新鮮培地が系外から系内へ押込まれる状態、すな
わち逆′/′jc務状態に入り膜面に付着した菌体等が
除去され、次の正流運転に備えて洗務を兼ねながら低分
子栄養分を補給することができる。
During reverse flow operation, the high-pressure side and low-pressure side of the first and second filtration chambers are reversed, and the first filtration chamber, which previously discharged the permeate from inside the system to the outside, is used to remove metabolites. Molecular nutrients and fresh culture medium are pushed into the system from outside the system, that is, enter a reverse '/'jc state, where bacterial bodies adhering to the membrane surface are removed and washed in preparation for the next forward flow operation. It can replenish low-molecular nutrients while also serving as a supplement.

このように第1.2濾過室の高圧、低圧側を切換えるこ
とにより異なる機能を交互にもたせ透過流速を低下させ
ることなく連続的に代謝物を除きながら培養を継続させ
、連続的に高濃度の菌体を培養することができる。
In this way, by switching between the high pressure and low pressure sides of the 1.2 filtration chamber, different functions can be provided alternately, and the culture can be continued while continuously removing metabolites without reducing the permeation flow rate. The bacterial cells can be cultured.

したがって本発明によれば培養により生成する増殖阻害
物質を除去しながら培養を行うので従来のバッチ式を太
き(上進る高濃度培養が達成できる。
Therefore, according to the present invention, culturing is carried out while removing growth-inhibiting substances produced during culturing, making it possible to achieve higher concentration culturing than the conventional batch method.

又菌体の高濃度化に伴い所定菌体を製造する場合培養タ
ンク容積は大巾に削減できるので小型でコンパクトな設
計ができる。
In addition, when producing a predetermined number of bacterial cells due to an increase in the concentration of bacterial cells, the volume of the culture tank can be greatly reduced, allowing for a small and compact design.

更に又培養により生産される代謝生産物を効果的に除去
しながら培養を行うので所要培地量は115以下に削減
でき、補給する培地は膜を通して系内に入るため培地の
滅菌は不要となり省エネルギーが可能である。
Furthermore, since the culture is carried out while effectively removing the metabolic products produced during the culture, the amount of culture medium required can be reduced to 115 or less, and since the medium to be replenished enters the system through the membrane, sterilization of the medium is not required, resulting in energy savings. It is possible.

(実施例) 以下図面に示す実施例について説明する。(Example) The embodiments shown in the drawings will be described below.

第1図において、(10)は培養槽であり、(11) 
 (12)はフォローファイバーUF膜を使用した第1
.2濾過室である。
In Figure 1, (10) is a culture tank, (11)
(12) is the first method using a follow fiber UF membrane.
.. 2 filtration chambers.

培養槽(10)と第1絶過室(11)とはポンプ(37
)のある管(13)で連結され、第1濾過室(11)と
第2#、、過室(12)とは管(35)で連結されてい
る。
The culture tank (10) and the first overflow chamber (11) are connected to the pump (37).
) are connected by a pipe (13), and the first filter chamber (11) and the second filter chamber (12) are connected by a pipe (35).

第2濾過室(12)は又管(15)を介して培養槽(1
0)に連結され、第1濾過室(11)は管(13)から
バルブ(20)を介して分枝した管(14)で培養槽(
10)に連結されている。
The second filtration chamber (12) is also connected to the culture tank (1) via a pipe (15).
The first filtration chamber (11) is connected to the culture tank (14) via a valve (20) from the pipe (13).
10).

管(13)の途中には以上の外バルブ(19)から分枝
した管(36)がバルブ(21)を介して管(15)に
連結されている。
In the middle of the pipe (13), a pipe (36) branched from the above outer valve (19) is connected to the pipe (15) via a valve (21).

以上のようなことから実線で示す矢印の正流運転時では
培養槽(10)からの菌体液は管(13)から第1濾過
室(11) 、第2濾過室(12)を経て管(15)で
培養槽(10)に還元されるようになっていて1つの循
環回路を構成している。
From the above, during the forward flow operation indicated by the solid line, the bacterial liquid from the culture tank (10) flows from the tube (13) through the first filtration chamber (11) and the second filtration chamber (12). 15) to be returned to the culture tank (10), forming one circulation circuit.

又鎖線で示す矢印の逆流運転時では管(36)から管(
15)を介して第2濾過室(12) 、第1濾過室(1
1)の順に通過し、管(14)で培養槽(10)に還元
されるようになついる。
Also, during reverse flow operation as indicated by the chain line arrow, the flow from the pipe (36) to the pipe (
15) to the second filtration chamber (12) and the first filtration chamber (1
1) and return to the culture tank (10) in the tube (14).

第1.2濾過室には培地供給タンク(29)からポンプ
(31)を通じて培地が管(30)に送られ、バルブ(
32)を介して管(34)  (33)の何れかを通じ
て第1.2濾過室に押込まれるようになっている。
In the 1.2 filtration chamber, the culture medium is sent from the culture medium supply tank (29) to the pipe (30) through the pump (31), and the valve (
32) and into the 1.2 filtration chamber through any of the pipes (34) and (33).

又UF膜より除去された代謝物を含む低分子栄養成分は
第1.2濾過室(11)  (12)から管(22) 
 (40) 、バルブ(23) 、管(24) 、ポン
プ(26)を通じて濾過室(27)に導かれ、ここでR
O膜を通じて代謝物の除去後、管(28)を通じて培地
供給タンク(29)に還元されるようになっている。
In addition, low-molecular nutrients including metabolites removed from the UF membrane are transferred from the 1.2 filtration chambers (11) (12) to the pipes (22).
(40), valve (23), pipe (24) and pump (26) to the filtration chamber (27), where R
After the metabolites are removed through the O membrane, they are returned to the medium supply tank (29) through the pipe (28).

その他管(13)から管(16) 、ポンプ(18)を
通じて濃縮菌液が濃縮菌液回収タンク(17)に回収さ
れるようになっている。
In addition, the concentrated bacterial liquid is collected from the pipe (13) through the pipe (16) and the pump (18) into the concentrated bacterial liquid recovery tank (17).

さて第1図のフローシートにしたがって具体的に説明し
て行くと、まず培養槽(10)で種菌を接種し、培養を
開始する。
Now, to explain in detail according to the flow sheet shown in FIG. 1, first, inoculum is inoculated in a culture tank (10) and culture is started.

数時間後に代謝産物が蓄積され始め、ある濃度に達する
と膜の運転を開始する。
Metabolites begin to accumulate after a few hours, and when a certain concentration is reached, the membrane begins to operate.

実線矢印の正流運転時には培養槽(10)よりポンプ(
37)で引抜いた菌体液を第1.2濾過室の順に通過さ
せる。
During forward flow operation as indicated by the solid line arrow, the pump (
The bacterial cell fluid drawn out in step 37) is passed through the 1.2 filtration chambers in this order.

第1.2濾過室におけるUF膜はフォローファイバーU
F膜からなるものでこの場合第1濾過室(11)が高圧
側となり、代謝物を含む透過液が管(22) 、バルブ
(23) 、管(24) 、ポンプ(26) 、管(2
5)を経て濾過室(27)に導かれここでRO膜で代謝
物が除去され代謝物が除去された低分子栄養成分は管(
28)を通し培地供給タンク(29)に戻され、培地の
有効利用が計れようになっている。
The UF membrane in the 1.2 filtration chamber is the follow fiber U
In this case, the first filtration chamber (11) becomes the high pressure side, and the permeate containing metabolites flows through the tube (22), valve (23), tube (24), pump (26), tube (2).
5) to the filtration chamber (27), where metabolites are removed by an RO membrane, and the low molecular weight nutritional components from which the metabolites have been removed are passed through a tube (27).
28) and returned to the culture medium supply tank (29), so that the culture medium can be used effectively.

一方低圧側となる第2濾過室(12)ではポンプ(31
)により代謝物を除いた低分子栄養物及び新鮮培地がタ
ンク(29)から管(30) 、バルブ(32) 、管
(33)を通じて押込まれ培養槽(1o)内のタンクレ
ベルを一定に保持するように運転される。
On the other hand, in the second filtration chamber (12) on the low pressure side, the pump (31
), low-molecular nutrients and fresh medium from which metabolites have been removed are pushed from the tank (29) through the pipe (30), valve (32), and pipe (33) to maintain a constant tank level in the culture tank (1o). be driven to do so.

以上のような運転状態を継続すると、第1濾過室からの
透過液速度が培養過程で菌数増加に伴う高粘度化、膜の
目づまりにより低下するのでこの時三方電磁バルブ(1
9)  (20)  (21)(23)  (32)を
同時に切換え鎖線矢印で示す逆流運転に入る。
If the above operating conditions are continued, the rate of permeate from the first filtration chamber will decrease due to increased viscosity due to the increase in the number of bacteria during the culture process and clogging of the membrane.
9) Switch (20), (21), (23), and (32) at the same time and enter reverse flow operation as indicated by the chain arrow.

このバルブ切換により高圧側と低圧側とが逆転し、これ
まで透過液を系内から系外に排出していた第1濾過室で
は系外から系内に液が押し込まれる状態すなわち、タン
ク(29) 、ポンプ(31) 、管(30) 、バル
ブ(32) 、管(34)を介して逆洗務状態に入り膜
面に付着した菌体等が除去され、次の正流運転に備えて
洗務を兼ねながら低分子栄養分を補給する。
By switching this valve, the high pressure side and the low pressure side are reversed, and the first filtration chamber, which had previously discharged the permeated liquid from inside the system to the outside, is now in a state where liquid is forced into the system from outside the system, that is, the tank (29 ), the pump (31), the pipe (30), the valve (32), and the pipe (34) enter the backwashing state, and bacteria adhering to the membrane surface are removed and prepared for the next forward flow operation. Replenishes low-molecular nutrients while also doing laundry.

このように第1.2濾過室の高圧、低圧側を切換えるこ
とにより異なる機能を交互にもたせ透過流速を低下させ
ることなく連続的に代謝物を除きながら培養を継続させ
連続的に高濃度の菌体を培養させることができる。
In this way, by switching between the high pressure and low pressure sides of the 1.2 filtration chamber, different functions can be alternately provided, and the culture can be continued while continuously removing metabolites without reducing the permeation flow rate, thereby continuously increasing the concentration of bacteria. The body can be cultured.

実施例のものでは第1.2濾過室を直列に配置したもの
について説明したが並列に配置したものでもよい。
In the embodiment, the first and second filtration chambers are arranged in series, but they may be arranged in parallel.

又実施例のものでは連続式の場合を示しているが二次増
殖期までバッチ式で運転を終了させてもよい。
Further, although the examples show a continuous type operation, the operation may be completed in a batch type until the secondary growth phase.

以下ビヒダス菌の連続高濃度培養の実験について説明す
る。
An experiment of continuous high-concentration culture of Bacterium bifidus will be described below.

10ffi容積培養タンクに酵母エキス1.0%、ペプ
トン1.0 %、バーミエートIf13.0%、肉エキ
ス0,5%、K2HP 040.5%、KH,PO40
,1%アスコルビン酸Na O,1%、水93.8%か
らなる組成の培地61を仕込み、120℃15分間の滅
菌後冷却し、あらかじめ前培養を行ったB 、  L 
ongumの種菌を接種し、33°C±0.5℃の温度
条件で培養を行った。
Yeast extract 1.0%, peptone 1.0%, vermeate If 13.0%, meat extract 0.5%, K2HP 040.5%, KH, PO40 in a 10ffi volumetric culture tank.
, 1% Na O, 1% ascorbate, and 93.8% water were prepared, and the medium 61 was sterilized at 120°C for 15 minutes, cooled, and precultured in advance.
ongum inoculum was inoculated and cultured at a temperature of 33°C±0.5°C.

ここで接種前には培地中にN ガスを通気して培地中に
溶存している02 ガスを除去し、また培養液面上には
N2 ガスを封入して培養中に02ガスとの接触を断っ
た。培養過程における生菌数、乳糖含量及び代謝産物で
あるし一乳酸濃度の変化は第4図に示す。
Before inoculation, N gas is aerated into the culture medium to remove O2 gas dissolved in the culture medium, and N2 gas is sealed above the culture solution surface to prevent contact with O2 gas during culture. I declined. Figure 4 shows changes in the number of viable bacteria, lactose content, and concentration of lactic acid, a metabolite, during the culture process.

初発のPHは6.8であるが培養過程と共にPHは低下
して行き、ある値を越えると増殖が停止するので、本実
験では2N−NH,OHを用いて6.0の定PHコント
ロールを行った。
The initial pH is 6.8, but as the culture progresses, the pH decreases and growth stops when it exceeds a certain value. Therefore, in this experiment, a constant pH of 6.0 was controlled using 2N-NH,OH. went.

培養10時間後には、制限基質となる乳糖濃度は2g/
f以下となり、L−乳酸濃度は10g/j!を越えて増
殖は停止するので、この時期より、本発明で明示した膜
システムの運転を開始する。
After 10 hours of culture, the concentration of lactose, which is the limiting substrate, was 2 g/
f or less, and the L-lactic acid concentration is 10 g/j! Since the growth stops beyond this point, the operation of the membrane system specified in the present invention is started from this period.

実験に使用した膜は、旭化成工業(株)製のフォローフ
ァイバー膜S E P4O10(分画分子量6000、
膜面積0.2 rd、耐熱温度90℃)二本である。膜
内及び配管系内の無菌化は、80℃温水を1時間循環後
、次亜塩素酸液200ppm、浸漬3時間後、無菌水を
用いて次亜塩素酸液を排除し、10jl’培養ダンクと
無菌的に接合し、実験に供した。
The membrane used in the experiment was a follow fiber membrane S E P4O10 (molecular weight cut off 6000,
(Membrane area: 0.2 rd, heat resistant temperature: 90°C). To sterilize the inside of the membrane and piping system, after circulating 80℃ hot water for 1 hour, immersing in 200 ppm hypochlorous acid solution for 3 hours, remove the hypochlorous acid solution using sterile water, and place in a 10jl' culture dunk. They were joined aseptically and used for experiments.

培養開始後10時間よりペリスタポンプを作動させて膜
運転を開始する。正逆の流路切換えはタイマーを用いて
30分間隔で行った。
Ten hours after the start of culture, the peristaltic pump is activated to start membrane operation. The forward and reverse flow path switching was performed at 30 minute intervals using a timer.

供給する培地は酵母エキス1.0%、ペプトン1.0%
、バーミエート粉3.0%、肉エキス0.5%、K2H
P O,0,5%、KH,PO,0,1%、アスコルビ
ン酸Na O,1%、水93.8%組成の培地をあらか
じめ5EP1013膜を通したものを用いた。
The medium to be supplied is yeast extract 1.0% and peptone 1.0%.
, Vermeate flour 3.0%, Meat extract 0.5%, K2H
A medium having a composition of 0.5% PO, 0.1% KH, PO, 1% Na O ascorbate, and 93.8% water was previously passed through a 5EP1013 membrane.

また培養槽液レベルは6ρ一定になるように保、持した
In addition, the culture tank liquid level was kept constant at 6ρ.

その結果、膜を通して透過する液速度は第5.6図に示
すように菌の増殖と共に低下するが、ある目標濃度[1
0”(/ mjり ]に達してから菌液を定常的に排出
しはじめると、液透過速度の低下は認められず、連続的
な菌液の取得が可能であった。
As a result, the rate of liquid permeation through the membrane decreases as the bacteria multiply, as shown in Figure 5.6, but at a certain target concentration [1
When the bacterial liquid started to be discharged steadily after reaching 0'' (/mjri), no decrease in the liquid permeation rate was observed, and it was possible to continuously obtain the bacterial liquid.

定常運転に至るまでの菌の比増殖速度μ(Hr” )は
、−次比増殖速度μ (培8槽のみの運転時)! は0.23 CHr−’ )であり、二次比増殖速度μ
2(膜運転を行いながら培養槽運転を行った時)は0.
10 (Hr  )となる。
The specific growth rate μ (Hr”) of the bacteria until steady operation is reached is the -order specific growth rate μ (when operating only 8 medium tanks)! is 0.23 CHr-'), and the second order specific growth rate μ (Hr'') is 0.23 CHr-') μ
2 (when performing culture tank operation while performing membrane operation) is 0.
10 (Hr).

以上の結果を用いて系内の基質、生菌、生産物の収支を
取る事により、連続運転時の苗生産速度、濃縮菌液抜き
出し速度、基質消費速度、代謝産物速度等の決定を行う
ことができる。
By calculating the balance of substrates, viable bacteria, and products in the system using the above results, the seedling production rate, concentrated bacterial liquid extraction rate, substrate consumption rate, metabolite rate, etc. during continuous operation can be determined. I can do it.

このように−次増殖期間(10時間)、二次増殖期間゛
(8時間)を経た後、菌体濃度10’(−/mε)の液
、0.6 J/HrX36Hr=21.6j!を安定的
に取得する事ができた。
In this way, after passing through the second growth period (10 hours) and the second growth period (8 hours), a solution with a bacterial cell concentration of 10' (-/mε), 0.6 J/HrX36Hr=21.6j! We were able to obtain it stably.

槽内残留液及び配管内液量6.81を回収すると21.
6+ 6.8=28.1となった。この間に使用した培
地量は1001である。また総運転時間は54時間であ
る。
When the remaining liquid in the tank and the amount of liquid in the pipes are 6.81, it is 21.
6+6.8=28.1. The amount of medium used during this period was 1,001. The total operating time is 54 hours.

従来、この種の菌の高濃度培養液の取得には定PH培養
法を用いて菌液を得た後、遠心分離法でスラッジ状菌を
集菌し、分散液を用いて1011(/mjlりの菌液を
得ている。この従来法と、本実験結果を比較すると、 1      1従 来 法1本発明法1比 11培養
槽容積  1 250A   l  10 /  10
.041(培 養 法  1バッチ式 1連続式 1−
11培養時間   116時間 154時間 1311
使用培地量  l  200z   1100 e  
10.5 11取得菌液量  1     1    
1 1110”(−/ml)  l  10j!   
128.4112.8411   換算  I    
 I    1 11菌液/使用培地10.05 (−
)  I O,28415,681111(−)II 以上のように本発明を用いると所定菌数の高濃度液を等
量得る際従来法にくらべて培養槽容積は大巾に縮小でき
、かつ使用培地量も削減できる。
Conventionally, to obtain a high-concentration culture solution of this type of bacteria, a constant pH culture method was used to obtain a bacterial solution, and then the sludge-like bacteria were collected using a centrifugation method, and the dispersion was used to collect 1011 (/mjl). Comparing the results of this experiment with this conventional method, we find that: 1 1 Conventional method 1 Present method 1 Ratio 11 Culture tank volume 1 250 A l 10 / 10
.. 041 (Culture method 1 batch type 1 continuous type 1-
11 Culture time 116 hours 154 hours 1311
Amount of medium used l 200z 1100 e
10.5 11 Obtained bacterial liquid volume 1 1
1 1110” (-/ml) l 10j!
128.4112.8411 Conversion I
I 1 11 bacterial solution/Used medium 10.05 (-
) IO, 28415, 681111 (-) II As described above, when the present invention is used to obtain an equal volume of a highly concentrated solution containing a predetermined number of bacteria, the culture tank volume can be greatly reduced compared to the conventional method, and the culture medium used can be reduced. The amount can also be reduced.

本実験例では透過液中の有効成分の回収は行っていない
が、これを行うと、さらに使用培地量の削減を行うこと
は可能になる。
Although the active ingredients in the permeate were not recovered in this experimental example, if this was done, it would be possible to further reduce the amount of medium used.

なお、本実験例ではビヒダス菌の高濃度培養例を示した
が、好気性菌の培養等にも十分利用できる。
In addition, in this experimental example, a high-concentration culture example of Bifidus bacteria was shown, but it can also be fully used for culturing aerobic bacteria.

又本実験例では分画分子量6000のUF膜を使用した
が除菌効果を有する0、1〜0.45μ程度のマイクロ
フィルターの使用により効果的な代謝物除去が可能であ
る。
Furthermore, although a UF membrane with a molecular weight cutoff of 6000 was used in this experimental example, effective removal of metabolites is possible by using a microfilter with a size of about 0.1 to 0.45 μm, which has a sterilization effect.

【図面の簡単な説明】[Brief explanation of the drawing]

第1図は本発明装置のフローシート 第2.3図は従来の培養装置を示すフローシート 第4図は培養タイムコート図 第5図は膜よりの透過流速変化図 第6図は膜1本当りの透過液流速の経時変化図 第7図は正流運転時におけるUF膜前後の圧力分布図で
ある。 (10)・・・・培養槽 (11)・・・・第1濾過室 (12)・・・・第2濾過室 (29)・・パ・培地供給タンク 培誉時 第6図 培促時間  (+−(r ) 第7図 (入L1)(出口)(入口)     (出D)間  
()T r ) 第5図 膜運転開始から01時間
Figure 1 is a flow sheet of the device of the present invention. Figure 2.3 is a flow sheet of a conventional culture device. Figure 4 is a culture time coat diagram. Figure 5 is a diagram of changes in permeation flow rate through the membrane. Figure 6 is a diagram of one membrane. Fig. 7 is a graph of the pressure distribution before and after the UF membrane during normal flow operation. (10)...Cultivation tank (11)...First filtration chamber (12)...Second filtration chamber (29)...Pa・Medium supply tank When culturing Figure 6 Culture promotion time (+-(r) Figure 7 (Inlet L1) (Exit) (Inlet) (Outlet D)
()Tr) Figure 5: 01 hours from the start of membrane operation

Claims (2)

【特許請求の範囲】[Claims] (1)培養槽で各種菌体を培養するに当たって、培養中
に生産される代謝産物を培養槽と共に循環回路を構成す
る第1、2濾過膜を通して取り除き、取り除いた代謝産
物から低分子有効成分を回収してこれを新鮮培地と共に
前記濾過膜を通して循環回路に供給するに当たり、循環
回路の正流方向では第2濾過膜に供給し、逆流方向では
第1濾過膜に供給して連続的に高濃度菌液をうるように
したことを特徴とする高濃度連続培養方法。
(1) When culturing various types of bacterial cells in a culture tank, metabolites produced during the culture are removed through the first and second filtration membranes, which together with the culture tank constitute a circulation circuit, and low-molecular active ingredients are extracted from the removed metabolites. When collecting and supplying the fresh medium to the circulation circuit through the filtration membrane, in the forward direction of the circulation circuit, it is supplied to the second filtration membrane, and in the reverse direction, it is supplied to the first filtration membrane to continuously maintain high concentration. A high-concentration continuous culture method characterized by making the bacterial solution moist.
(2)培養槽とUF膜等の濾過膜を使用した第1、2濾
過室とを配管装置を介して可逆循環回路を構成する如く
連結し、かつ第1、2濾過室は新鮮培地と代謝産物から
回収した低分子有効成分とを供給できる供給タンクに切
換自在に連結すると共に代謝産物の回収装置に連結し、
循環回路の正流方向では第2濾過室を供給タンクに連結
し、逆流方向では第1濾過室を供給タンクに連結する如
く配管構成されたことを特徴とする高濃度連結培養装置
(2) The culture tank and the first and second filtration chambers using a filtration membrane such as a UF membrane are connected through a piping device to form a reversible circulation circuit, and the first and second filtration chambers are used for fresh culture medium and metabolic switchably connected to a supply tank capable of supplying low-molecular active ingredients recovered from the product, and connected to a metabolic product recovery device;
A high-concentration connected culture device, characterized in that the piping is configured such that the second filtration chamber is connected to the supply tank in the forward flow direction of the circulation circuit, and the first filtration chamber is connected to the supply tank in the reverse flow direction.
JP59211747A 1984-10-09 1984-10-09 Method and apparatus for continuous cultivation at high concentration Granted JPS6188872A (en)

Priority Applications (1)

Application Number Priority Date Filing Date Title
JP59211747A JPS6188872A (en) 1984-10-09 1984-10-09 Method and apparatus for continuous cultivation at high concentration

Applications Claiming Priority (1)

Application Number Priority Date Filing Date Title
JP59211747A JPS6188872A (en) 1984-10-09 1984-10-09 Method and apparatus for continuous cultivation at high concentration

Publications (2)

Publication Number Publication Date
JPS6188872A true JPS6188872A (en) 1986-05-07
JPS6363193B2 JPS6363193B2 (en) 1988-12-06

Family

ID=16610912

Family Applications (1)

Application Number Title Priority Date Filing Date
JP59211747A Granted JPS6188872A (en) 1984-10-09 1984-10-09 Method and apparatus for continuous cultivation at high concentration

Country Status (1)

Country Link
JP (1) JPS6188872A (en)

Cited By (12)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP0269444A2 (en) * 1986-11-26 1988-06-01 Henry B. Kopf Apparatus and method for mass transfer involving biological/pharmaceutical media
EP0336974A1 (en) * 1987-10-01 1989-10-18 Sumitomo Electric Industries, Ltd. Cell propagation apparatus
WO1989012676A1 (en) * 1988-06-21 1989-12-28 Kopf Henry B Culture device and method
JPH0242992A (en) * 1988-08-03 1990-02-13 Tanabe Seiyaku Co Ltd Reaction using biocatalyst and device for said reaction
WO1990002171A1 (en) * 1988-08-31 1990-03-08 Cellco Advanced Bioreactors, Inc. In vitro cell culture reactor
WO1992013940A1 (en) * 1991-02-12 1992-08-20 Synbiotics Corporation Hollow fiber cell propagation
US6388355B2 (en) 1998-06-29 2002-05-14 Mitsubishi Denki Kabushiki Kaisha Motor for an electric power steering assembly
JP2003521877A (en) * 1999-02-05 2003-07-22 プロテイン サイエンシーズ コーポレイション Apparatus and methods for producing and using high density cells and products therefrom
JP2005261342A (en) * 2004-03-19 2005-09-29 Yanmar Co Ltd Plankton culture system
JP2016530893A (en) * 2013-09-16 2016-10-06 ジェンザイム・コーポレーション Method and system for processing cell culture
JP2018521683A (en) * 2015-08-07 2018-08-09 ローディア オペレーションズ Improved production of vanillin by fermentation.
CN113634125A (en) * 2021-10-18 2021-11-12 深圳市路阳农业科技有限公司 Alkaline microorganism culture solution concentration and filtration device

Cited By (17)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP0269444A2 (en) * 1986-11-26 1988-06-01 Henry B. Kopf Apparatus and method for mass transfer involving biological/pharmaceutical media
US6022742A (en) * 1986-11-26 2000-02-08 Kopf; Henry B. Culture device and method
US6048727A (en) * 1986-11-26 2000-04-11 Kopf; Henry B. Apparatus and method for mass transfer involving biological/pharmaceutical media
EP0336974A1 (en) * 1987-10-01 1989-10-18 Sumitomo Electric Industries, Ltd. Cell propagation apparatus
WO1989012676A1 (en) * 1988-06-21 1989-12-28 Kopf Henry B Culture device and method
JPH0242992A (en) * 1988-08-03 1990-02-13 Tanabe Seiyaku Co Ltd Reaction using biocatalyst and device for said reaction
WO1990002171A1 (en) * 1988-08-31 1990-03-08 Cellco Advanced Bioreactors, Inc. In vitro cell culture reactor
WO1992013940A1 (en) * 1991-02-12 1992-08-20 Synbiotics Corporation Hollow fiber cell propagation
US6388355B2 (en) 1998-06-29 2002-05-14 Mitsubishi Denki Kabushiki Kaisha Motor for an electric power steering assembly
JP2003521877A (en) * 1999-02-05 2003-07-22 プロテイン サイエンシーズ コーポレイション Apparatus and methods for producing and using high density cells and products therefrom
JP2005261342A (en) * 2004-03-19 2005-09-29 Yanmar Co Ltd Plankton culture system
JP2016530893A (en) * 2013-09-16 2016-10-06 ジェンザイム・コーポレーション Method and system for processing cell culture
JP2020124204A (en) * 2013-09-16 2020-08-20 ジェンザイム・コーポレーション Method and system for processing cell culture
JP2022050549A (en) * 2013-09-16 2022-03-30 ジェンザイム・コーポレーション Methods and systems for processing cell culture
JP2018521683A (en) * 2015-08-07 2018-08-09 ローディア オペレーションズ Improved production of vanillin by fermentation.
US11060118B2 (en) 2015-08-07 2021-07-13 Rhodia Operations Production of vanillin by fermentation
CN113634125A (en) * 2021-10-18 2021-11-12 深圳市路阳农业科技有限公司 Alkaline microorganism culture solution concentration and filtration device

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