JPS61274707A - Regenerated porous membrane for separating bacteria - Google Patents
Regenerated porous membrane for separating bacteriaInfo
- Publication number
- JPS61274707A JPS61274707A JP11680585A JP11680585A JPS61274707A JP S61274707 A JPS61274707 A JP S61274707A JP 11680585 A JP11680585 A JP 11680585A JP 11680585 A JP11680585 A JP 11680585A JP S61274707 A JPS61274707 A JP S61274707A
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- Prior art keywords
- average pore
- porous membrane
- pore size
- cellulose
- minimum
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Abstract
Description
【発明の詳細な説明】
〔（イ）技術分野〕
本発明は、水溶液中に分散している微生物粒子を分離除
去する多孔膜に関する。さらに詳しくは、タンパク質や
電解質を溶解する水溶液中に分散した、直径０．０２μ
ｍ以−Ｌのうイルスあるいはリケッチャ、クラミジア、
マイコプラズマ等を含めた細菌を分離除去し、あるいは
微生物粒子を含む水溶液よりタンパク質を分離濃縮する
のに特に適した多孔膜に関する。Detailed Description of the Invention [(a) Technical Field] The present invention relates to a porous membrane that separates and removes microbial particles dispersed in an aqueous solution. More specifically, it is 0.02μ in diameter dispersed in an aqueous solution that dissolves proteins and electrolytes.
M-L virus or Rickettsia, chlamydia,
The present invention relates to a porous membrane particularly suitable for separating and removing bacteria including mycoplasma, or for separating and concentrating proteins from an aqueous solution containing microbial particles.
人間を含めた動物の血液より血球成分を除去して得られ
る面漿、あるいは注射液や医薬品、生化学工業用に利用
される溶媒や化学薬品の水溶液中には微生物が存在して
はならない。特に、滅菌処理が不可能な溶液系（たとえ
ば血液凝固因子を含む溶液系）では、微生物の混入を徹
底的に防止しなくてはならない。しかしながら、完全に
防止することはできず、従って、最終製品から、あるい
は製造工程中において混入微生物を除去せざるを得ない
。一方、生物化学工業分野、特に遺伝子Ｔ学で製造され
るホルモン、インターフェロンなど、あるいは抗生物質
などの高温での滅菌が不可能な医薬品より微生物を除去
する必要性も近年増大しつつある。Microorganisms must not be present in plasma obtained by removing blood cell components from the blood of animals including humans, or in aqueous solutions of solvents and chemicals used for injections, pharmaceuticals, and the biochemical industry. In particular, in solution systems that cannot be sterilized (for example, solution systems containing blood coagulation factors), contamination with microorganisms must be thoroughly prevented. However, it cannot be completely prevented, and therefore contaminating microorganisms must be removed from the final product or during the manufacturing process. On the other hand, in recent years there has been an increasing need to remove microorganisms from the biochemical industry, particularly from hormones and interferons produced in gene T science, and from pharmaceuticals such as antibiotics that cannot be sterilized at high temperatures.
従来、水溶液中に分散した微生物粒子を除去ずる方法と
して、ゲル濾過法、遠心分離法、吸着分離法、沈澱法、
限外濾過法が利用されている。ケル濾過法は、ゲル濾過
に用いられる溶媒により目的物質が稀釈され、そのため
工業的に適用するのが困難である。遠心分離法は、微生
物粒子の直径が１０μｍ以上でかつ、水溶液の粘度が小
さい場合にのみ適用できる。吸着分離法は、特定の少量
の微生物粒子の除去に利用できるが、多数の微生物粒子
が多量に分散している水溶液にはこの方法は適用されな
い。沈澱法は比較的多量の水溶液の処理に利用できるが
、この方法単独では微生物粒子を完全に除去することは
不可能で、一般には抽出処理と組合わせて利用される。Conventionally, methods for removing microbial particles dispersed in an aqueous solution include gel filtration, centrifugation, adsorption separation, precipitation,
Ultrafiltration is used. In the gel filtration method, the target substance is diluted by the solvent used in gel filtration, and therefore it is difficult to apply it industrially. The centrifugation method can be applied only when the microbial particles have a diameter of 10 μm or more and the aqueous solution has a low viscosity. Adsorption separation methods can be used to remove specific small amounts of microbial particles, but this method is not applicable to aqueous solutions in which large numbers of microbial particles are dispersed in large quantities. Although the precipitation method can be used to treat relatively large amounts of aqueous solutions, it is not possible to completely remove microbial particles using this method alone, and it is generally used in combination with an extraction process.
しかしながら、たとえ両者を組合わセでも、微生物粒子
を完全に除去することは困難であり、また工程が一般に
複雑であり、それぞれの工程で微生物粒子が混入する恐
れがある。限外濾過法は、あらゆる微生物粒子の除去法
としてその将来性が期待されているにもかかわらず、は
とんど工業的には利用されていない。その理由は下記の
とおりである。直径０．０２〜０．４μｍの微生物粒子
を除去しようとすると、限外濾過速度が極端に小さくな
る。また、微生物粒子を分散した水溶液中には、一般に
タンパク質等の高分子量の吸着性物質が溶解しているた
め、限外濾過に利用される膜表面に、濾過中にタンパク
質が吸着し、濾過速度が経時的に減少する。さらに、こ
の際、微生物粒子のみならず、目的物質であるタンパク
質も膜で捕捉され濾液中に回収されるタンパク質量が低
下する。However, even if the two are combined, it is difficult to completely remove microbial particles, and the steps are generally complicated, so there is a risk that microbial particles may be mixed in each step. Although ultrafiltration is a promising method for removing all kinds of microbial particles, it is hardly used industrially. The reason is as follows. When attempting to remove microbial particles with a diameter of 0.02 to 0.4 μm, the ultrafiltration rate becomes extremely low. In addition, since high-molecular-weight adsorbent substances such as proteins are generally dissolved in an aqueous solution in which microbial particles are dispersed, proteins are adsorbed to the surface of the membrane used for ultrafiltration during filtration, which increases the filtration rate. decreases over time. Furthermore, at this time, not only the microbial particles but also the target substance protein is captured by the membrane, reducing the amount of protein recovered in the filtrate.
（（Ａ）発明が解決しようとする問題点〕本発明の目的
は、上述の限外濾過法の欠点を除去し、生体系へ安全に
適用可能で、かつ適用後の処理が簡拳な、微生物粒子の
除去に適した多孔膜、ならびに、微生物粒子を混入する
タンパク質水溶液よりタンパク質を微生物粒子と分離し
濃縮するのに適した多孔膜を提供するにある。((A) Problems to be Solved by the Invention) The purpose of the present invention is to eliminate the drawbacks of the above-mentioned ultrafiltration method, to provide a system that can be safely applied to biological systems and that requires simple processing after application. It is an object of the present invention to provide a porous membrane suitable for removing microbial particles, and a porous membrane suitable for separating and concentrating proteins from microbial particles from an aqueous protein solution containing microbial particles.
〔（ニ）問題点を解決するための手段〕上記問題点を解
決する本発明の微生物粒子分離用セルロース多孔膜は、
極小平均孔径が０．０２μｍ〜０．２μｍであり、かつ
分離すべき微生物粒子の最小の直径の１．０倍以下、０
．３倍以上であり、また、４次の平均孔半径（後述）〒
４と３次の平均孔半径（後述）手桶Ｔ３との比ｒ４／ｒ
３カ月、３以下ある孔径分布を有する銅安法セルロース
多孔膜であって、極小面内空孔率が１０％以上で、１つ
膜表面の平均孔径と極小平均孔径との比が４以上である
ことを特徴とする。[(d) Means for solving the problems] The cellulose porous membrane for separating microorganism particles of the present invention that solves the above problems has the following features:
The minimum average pore diameter is 0.02 μm to 0.2 μm, and 1.0 times or less of the minimum diameter of the microbial particles to be separated, 0
．． It is 3 times or more, and the average pore radius of the 4th order (described later)
Ratio r4/r of 4 and 3rd order average hole radius (described later) T3
A copper ammonium cellulose porous membrane having a pore size distribution of 3 or less, with a minimum in-plane porosity of 10% or more and a ratio of the average pore size of one membrane surface to the minimum average pore size of 4 or more. characterized by something.
ここで「多孔膜」とは、いわゆる平面状多孔膜およびチ
ューブ状多孔膜のいずれであってもよいが、直径１ｍｍ
以下の中空糸状膜を含まない。多孔膜は一般に膜表面に
平行な多数の層の積重ねとして近伯される。この層内で
の平均孔径は、それぞれの層で異なる。それらの平均孔
径のうち、最も小さい平均孔径を極小平均孔径と定義す
る。また、極小平均孔径を与える層内での空孔率を極小
面内空孔率と定義する。Here, the "porous membrane" may be either a so-called planar porous membrane or a tubular porous membrane, with a diameter of 1 mm.
Does not include the following hollow fiber membranes. Porous membranes are generally described as a stack of multiple layers parallel to the membrane surface. The average pore size within this layer is different for each layer. Among those average pore diameters, the smallest average pore diameter is defined as the minimal average pore diameter. Further, the porosity within the layer that provides the minimum average pore diameter is defined as the minimum in-plane porosity.
本発明の重要な第１の特徴は、銅安法セルロース（以下
、「銅安セルロース」と略称する）を素材高分子として
採用する点にある。タンパク質と高分子素材との吸着性
に関する相関性を検討した結果、−Ｃ的には親水性素材
はど、タンパク質の吸着性が小さい。The first important feature of the present invention is that ammonium-produced cellulose (hereinafter abbreviated as "ammonium cellulose") is employed as the material polymer. As a result of examining the correlation between protein and polymer material adsorption, it was found that hydrophilic materials have lower protein adsorption in terms of -C.
ここで１−セルロース−１とは、いわゆる再生セルロー
スを意味する。再生セルロースより構成される多孔膜は
、セルロース誘導体（通常、セルロースアセテート）多
孔膜をケン化処理するか、あるいは銅安セルロースまた
はビスコース法セルロース原液より直接、流延成膜（あ
るいは紡口を用いたチューブ成膜）した多孔膜として作
製される。Here, 1-cellulose-1 means so-called regenerated cellulose. Porous membranes composed of regenerated cellulose are produced by saponifying porous membranes of cellulose derivatives (usually cellulose acetate), or by casting directly from ammonium cellulose or viscose cellulose stock solution (or using a spinneret). It is produced as a porous membrane using tube deposition (tube deposition).
−上記の３種の再生セルロース多孔膜間でタンパク質の
吸着性の差を比較検討した結果、セルロース誘導体多孔
膜をケン化処理したものが最も吸着性が大きく、残２者
間の差は小さい。銅安セルロースは、ビスコース法セル
ロースに比較して、７０℃の蒸留水中（１時間）への溶
解量が少な（、生体への安全性、力学的強靭性において
優れ、しかも同一の空孔率で比較した際、限外濾過速度
が大きい。また、蒸気滅菌処理に伴なら平均孔径の変化
が小さい。- As a result of comparing and examining the difference in protein adsorption properties among the above three types of regenerated cellulose porous membranes, the cellulose derivative porous membrane subjected to saponification treatment has the highest adsorption property, and the difference between the remaining two is small. Copper ammonium cellulose has a lower dissolution amount in distilled water (1 hour) at 70°C than viscose cellulose (it is superior in biological safety and mechanical toughness, and has the same porosity). When compared, the ultrafiltration rate is high. Also, when compared with steam sterilization, the change in average pore diameter is small.
銅安セルロースのセルロースの粘度平均分子ｉｔは７Ｘ
１０’以−１−が好ましく、また０、ｌ Ｎ ＮａＯＨ
水溶液中へ熔解する成分が少なければ少ないほど望まし
い。４０℃、４８時間０．１ Ｎ Ｎａ０１１水溶液中
に浸漬した際、この溶解分力用Ｏｐｐｍ以下であれば、
この多孔膜は血詣中より微生物を除去するのに最適であ
る。The viscosity average molecule of copper ammonium cellulose is 7X
10' or more -1- is preferable, and 0, l N NaOH
It is desirable that fewer components dissolve into the aqueous solution. When immersed in a 0.1 N Na011 aqueous solution at 40°C for 48 hours, if the melting force Oppm is below this,
This porous membrane is ideal for removing microorganisms from blood samples.
上述のようなセルロースからなる多孔膜を作製するには
、高純度セルロース原料を用いて銅安セルロースを作製
するかあるいは多孔膜を作製後に０、 Ｉ Ｎ ＮａＯ
Ｈ水溶液で７２時間以上４５℃で洗滌処理すれば良い。To produce a porous membrane made of cellulose as described above, copper ammonium cellulose is produced using a high-purity cellulose raw material, or 0, I N NaO is prepared after producing the porous membrane.
What is necessary is to perform washing treatment with an H aqueous solution at 45° C. for 72 hours or more.
高純度セルロース原料を用いれば、上記溶解量が著しく
減少するので、より好ましい。It is more preferable to use a high-purity cellulose raw material because the above-mentioned amount of dissolution is significantly reduced.
ここで「高純度セルロース原料」とは、αセルロース含
有率が９５重量％以上で重合度５００以トの木綿リンタ
ーおよび木材パルプを指す。これらの原料について、ブ
リーチング、洗務工程中での分解および酸化を防ＩＩ−
シつつ、不純物の混入をさけるために、常に精製された
水を用いる。The term "high-purity cellulose raw material" as used herein refers to cotton linters and wood pulps having an α-cellulose content of 95% by weight or more and a degree of polymerization of 500 or more. These raw materials are protected against decomposition and oxidation during bleaching and washing processes.
However, always use purified water to avoid contamination with impurities.
本発明の多孔膜の第２の特徴は、極小平均孔径（後記の
ように測定される４次の極小平均孔径２〒−４を本発明
では極小平均孔径と略称する。）が０．０２〜０．２μ
ｍ、極小面内空孔率力月Ｏ％以上、４次の平均孔半径７
４と３次の平均孔半径〒３との比がｒｓｌ〒３≦１．３
の多孔膜を用いる点にある。The second feature of the porous membrane of the present invention is that the minimum average pore diameter (the 4th order minimum average pore diameter 2〒-4 measured as described below is abbreviated as the minimum average pore diameter in the present invention) is 0.02 to 0.2μ
m, minimum in-plane porosity 0% or more, 4th order average pore radius 7
The ratio of 4th and 3rd order average hole radius 〒3 is rsl 〒3≦1.3
The point is that a porous membrane is used.
本発明の多孔膜は、膜断面および膜表面を電子顕微鏡で
観察した際、断面および膜表面のいずれの面においても
、０．０２．ｌｒｍ以−ヒの孔が認められる膜である。The porous membrane of the present invention has a cross section and a membrane surface of 0.02% in both the cross section and membrane surface when observed with an electron microscope. This is a membrane in which pores larger than lrm are observed.
極小平均孔径が０．０２μｍ未満であると限外濾過速度
が極端に小さく、また水溶液中に溶解しているタンパク
質の濾液中への回収率が著しく低下する。たとえば、血
清アルブミンでは濾液中への回収率が５％未満となる。If the minimum average pore diameter is less than 0.02 μm, the ultrafiltration rate will be extremely low, and the recovery rate of proteins dissolved in the aqueous solution into the filtrate will be significantly reduced. For example, serum albumin has a recovery rate of less than 5% in the filtrate.
したがって、極小平均孔径は微生物粒子が除去可能な範
囲で大きければ大きいほど良いが、限外濾過時に、被濾
過焼体の供給回路からの微生物粒子の汚染を避けるため
に、０．２μｍ以下であることが必要である。Therefore, the minimum average pore diameter should be as large as possible to remove microbial particles, but it should be 0.2 μm or less in order to avoid contamination of microbial particles from the supply circuit of the sintered body during ultrafiltration. It is necessary.
極小面内空孔率は１０％以」＝必要である。１０％未満
では限外濾過速度は急激に低下する。好ましくは２０％
以上である。限外濾過速度に及ぼす面内空孔率の影響は
、１０％未満では極小面内空孔率の約５乗、１０〜３０
％では約２乗、３０％を超えると約１乗に仕例して限外
濾過速度は増加する。Minimum in-plane porosity of 10% or more is required. If it is less than 10%, the ultrafiltration rate decreases rapidly. Preferably 20%
That's all. The influence of the in-plane porosity on the ultrafiltration rate is approximately 5th power of the minimum in-plane porosity when it is less than 10%, which is 10 to 30
%, the ultrafiltration rate increases to the second power, and when it exceeds 30%, the ultrafiltration rate increases to the first power.
一方、極小面内空孔率が６０％を越えると、多孔膜の力
学的性質は著しく低下し、ピンホール等の欠損部が生じ
たり、多孔膜を構成するセルロース分子が、濾液中ある
いは被濾過液中に脱落分散する恐れがある。On the other hand, when the minimal in-plane porosity exceeds 60%, the mechanical properties of the porous membrane deteriorate significantly, causing defects such as pinholes, and cellulose molecules constituting the porous membrane may be present in the filtrate or filtered material. There is a risk of it falling off and dispersing into the liquid.
同一の極小平均孔径および極小面内空孔率で、限外濾過
速度に及ぼす孔径分布の影響を検討した結果、〒４７〒
３の値が小さいほど限外濾過速度が大きくなる。しかも
、７４７〒３≦１．３となると微生物粒子の直径の１．
０倍の極小平均孔径の多孔膜で、被濾過流体を静止下で
限外濾過（以下、「垂直濾過」と呼ぶことがある。）し
ても、濾液中には微生物粒子はほとんど観測できない。As a result of examining the influence of pore size distribution on ultrafiltration rate with the same minimum average pore diameter and minimum in-plane porosity, we found that
The smaller the value of 3, the higher the ultrafiltration rate. Moreover, when 747〒3≦1.3, 1.3 times the diameter of the microorganism particle.
Even when a fluid to be filtered is subjected to ultrafiltration under static conditions (hereinafter sometimes referred to as "vertical filtration") using a porous membrane with an extremely small average pore size of 0 times, almost no microbial particles can be observed in the filtrate.
孔径分布の孔径の大きい部分でのすそひきをなくすれば
、〒４７〒３を急速に小さくすることができる。By eliminating the skirting in the large pore diameter portion of the pore size distribution, 〒47〒3 can be rapidly reduced.
（ｌＯ）
〒４／７３≦１．１５で、かつ平均孔径が分離すべき微
生物粒子の最小の直径の０．９倍以下であれば、垂直濾
過で得られる濾液中に微生物が漏出することはない。(lO) If 4/73≦1.15 and the average pore size is 0.9 times or less the minimum diameter of the microbial particles to be separated, microorganisms will not leak into the filtrate obtained by vertical filtration. do not have.
本発明の多孔膜の第３の特徴は、膜表面の平均孔径と極
小平均孔径との比が４以上である点にある。同一素材高
分子で同一の極小平均孔径の多孔膜で限外濾過速度に及
ばず多孔膜断面構造の影響を検討した結果、膜表面（膜
の表裏両面のうち平均孔径が大きなほうの表面）の平均
孔径と極小平均孔径の比が大きくなればなるほど、限外
濾過速度は上昇する。この上昇度は４以」二では特に著
しい。また、膜面積１ ｃＪ当り、一定圧力下での垂直
濾過で濾過速度が、濾過初期の値の６割に減少するまで
の時間は、上記の比が４以上では極端に大きくなる。特
に単位膜面積当りに濾過すべき被濾過流体量が多い場合
、例えば、多量の溶液中に少量の微生物粒子が分散した
系からの微生物粒子の除去、あるいは該粒子の濃縮の際
には、−１−配圧が４以上であることが必須である。A third feature of the porous membrane of the present invention is that the ratio of the average pore diameter of the membrane surface to the minimum average pore diameter is 4 or more. As a result of examining the influence of the cross-sectional structure of the porous membrane, the ultrafiltration rate was not affected by porous membranes made from the same polymer material and having the same minimum average pore diameter. The larger the ratio of the average pore size to the minimum average pore size, the higher the ultrafiltration rate. This degree of increase is particularly remarkable in cases of 4 and above. Further, the time required for the filtration rate to decrease to 60% of the initial value of filtration in vertical filtration under a constant pressure per 1 cJ of membrane area becomes extremely long when the above ratio is 4 or more. In particular, when the amount of fluid to be filtered per unit membrane area is large, for example, when removing microbial particles from a system in which a small amount of microbial particles are dispersed in a large amount of solution, or concentrating the particles, - 1-It is essential that the pressure distribution is 4 or more.
膜厚をｄとすると、極小平均孔径を与える層が膜表面か
ら０．９ ｄ〜０．１ｄの間にあれば、同一の極小面内
空孔率と極小平均孔径を有する膜の限外濾過速度は大き
くなり、また濾過速度の経時的な変化も少ない。中空繊
維状の膜と比較して、平面状の膜では、温和な状況下で
の限外濾過が可能であり、単時間での濾過および膜表面
上に濃縮された微生物粒子の回収はより容易である。When the membrane thickness is d, if the layer giving the minimum average pore diameter is between 0.9 d and 0.1 d from the membrane surface, the ultrafiltration of the membrane with the same minimum in-plane porosity and minimum average pore diameter is possible. The speed increases, and the filtration rate changes less over time. Compared to hollow fiber membranes, planar membranes allow ultrafiltration under mild conditions, and easier filtration in a single time and recovery of microbial particles concentrated on the membrane surface. It is.
多孔膜の膜厚は薄ければ薄いほど、一般には濾過速度が
大きくなるので好ましい。しかしながら、壁厚が１０μ
ｍ未満になると、多孔膜にはピンホールが多発し、微生
物粒子が濾液中にもれ出てくる。膜厚が１０μｍで何故
ピンホール発生頻度が急激に変化するかは現在不明であ
る。おそらく、多孔膜の製法と密接に関連しているもの
と考えられる。膜厚が５００μｍを越えると、被濾過流
体中のタンパク質の吸着量が増大する。極小面内空孔率
が大きくなれば膜厚をより大きく設計するのが良い。微
生物粒子を分散した水溶液の粘度は一般には大きい。そ
のため多孔膜間を流れる被濾過流体の流れ厚さは２００
μｍ以上、２ｍｍ以下であることが望ましい。被濾過流
体中に分散した微生物粒子量が多い場合、被濾過流体の
膜表面−Ｅの流れ速度を大きくすることが望ましい。短
時間のｉＩ−過には垂直濾過が望ましい。The thinner the porous membrane is, the higher the filtration rate is, so it is preferable. However, the wall thickness is 10μ
If it is less than m, the porous membrane will have many pinholes and microbial particles will leak into the filtrate. It is currently unknown why the frequency of pinhole occurrence changes rapidly when the film thickness is 10 μm. This is probably closely related to the manufacturing method of the porous membrane. When the film thickness exceeds 500 μm, the amount of protein adsorbed in the fluid to be filtered increases. As the minimum in-plane porosity increases, it is better to design the film thickness to be larger. The viscosity of an aqueous solution in which microbial particles are dispersed is generally high. Therefore, the flow thickness of the fluid to be filtered flowing between the porous membranes is 200 mm.
It is desirable that the thickness is not less than μm and not more than 2 mm. When the amount of microbial particles dispersed in the fluid to be filtered is large, it is desirable to increase the flow rate of the fluid to be filtered across the membrane surface -E. Vertical filtration is preferred for short-term iI-filtration.
本発明の多孔膜の第４の特徴は、分離すべき微生物粒子
の直径の最小の粒子の粒子径の１．０倍以下、０．３倍
以上の平均孔径を持つ点にある。本発明では、銅安セル
ロースを素材に用いるため、水溶液中のタンパク質の吸
着が、他の素材にくらべて極端に少なく、限外濾過過程
中の目づまり現象は少ない。そのため、平均孔径は分離
すべき微生物粒子の最小の粒子の粒子径の１．０倍以下
であることが必要である。１．０倍を越えると、濾液中
にわずかであるが微粒子が漏出する。ただし、粒子が球
状でない場合は楕円体で返信し、直径と短径との平均を
粒子直径と定義する。濾過速度を大きくするために該粒
子の直径の０．３倍以上の平均孔径に設計する必要があ
る。〒、／〒３≦１．１５が満足されているならば、極
小平均孔径が該粒子の直径の０．９倍以下、０．５倍以
上の範囲内で設計すれば、濾液中には微生物粒子が皆無
でかつ十分な濾過速度を実現することができる。A fourth feature of the porous membrane of the present invention is that it has an average pore diameter that is 1.0 times or less and 0.3 times or more the diameter of the smallest microorganism particle to be separated. In the present invention, since copper ammonium cellulose is used as a material, adsorption of proteins in an aqueous solution is extremely small compared to other materials, and clogging phenomenon during the ultrafiltration process is small. Therefore, the average pore size needs to be 1.0 times or less the particle size of the smallest microorganism particles to be separated. If it exceeds 1.0 times, a small amount of fine particles will leak into the filtrate. However, if the particle is not spherical, it is returned as an ellipsoid, and the average of the diameter and the short axis is defined as the particle diameter. In order to increase the filtration rate, it is necessary to design the average pore size to be 0.3 times or more the diameter of the particles. If 〒,/〒3≦1.15 is satisfied, if the minimum average pore diameter is designed within the range of 0.9 times or less and 0.5 times or more of the particle diameter, microorganisms will be present in the filtrate. It is possible to achieve a sufficient filtration rate with no particles at all.
再生セルロースは水溶液中で一般には膨潤する。Regenerated cellulose generally swells in aqueous solution.
膨潤によってセルロース多孔膜が変形し、そのため多孔
膜面上での目づまりが起こることがある。Swelling deforms the cellulose porous membrane, which may cause clogging on the porous membrane surface.
これを防くには、多孔膜を構成するセルロース分子鎖の
面内配向度が６０％以上であることが好ましい。多孔膜
平面内でのセルロース分子鎖の配向は事実上存在しない
ことが望ましい。また面内配向度が大きくなりすぎると
膜厚方向での膨潤時の変形、および膜平面方向での収縮
が起こるため、面内配向度が８０％以下であることが好
ましい。To prevent this, it is preferable that the degree of in-plane orientation of the cellulose molecular chains constituting the porous membrane is 60% or more. It is desirable that there is virtually no orientation of cellulose molecular chains within the plane of the porous membrane. Further, if the degree of in-plane orientation becomes too large, deformation during swelling in the film thickness direction and contraction in the plane direction of the film will occur, so it is preferable that the degree of in-plane orientation is 80% or less.
本発明の多孔膜は、水溶液中に分散する微生物の分離除
去または微生物の分離濃縮、あるいは、水溶液中のタン
パク質の精製に利用できる。水溶液中に分散せる除去す
べき微生物の例としては、酵母、細菌、マイコプラズマ
、ウィルス等が挙げられる。本発明の多孔膜が好適に利
用されるのは微生物粒子の直径が１μｍ以下のリケッチ
ャやつィルス粒子の除去用、特にタンパク質でｌ） Ｎ
ＡまたはＲＮ Ａの周囲を囲まれたウィルス粒子の除
去に最適である。The porous membrane of the present invention can be used to separate and remove microorganisms dispersed in an aqueous solution, to separate and concentrate microorganisms, or to purify proteins in an aqueous solution. Examples of microorganisms to be removed that are dispersed in an aqueous solution include yeast, bacteria, mycoplasma, viruses, and the like. The porous membrane of the present invention is suitably used for the removal of rickettsia and virus particles with microbial particles having a diameter of 1 μm or less, especially proteins.
Ideal for removing virus particles surrounded by A or RNA A.
本発明の多孔膜の極小平均孔径（２ｒ、）、極小面内空
孔率、１−４／−〒−３および面内配向度はそれぞれ以
下の方法で決定された。The minimum average pore diameter (2r), minimum in-plane porosity, 1-4/-〒-3, and degree of in-plane orientation of the porous membrane of the present invention were determined by the following methods.
〈極小平均孔径〉、＜極小面内空孔率〉および〈〒４７
〒３〉
多孔膜の膜厚部の断面を走査型電子顕微鏡で観察し、孔
径の最小部分を膜厚（ｄｉに対して誤差１０％以内の範
囲で決定する。この最小な部分を通って多孔膜の膜表面
に平行に厚さ約０，１μｍの超薄切片を作成し、この切
片の電子顕黴鏡写貫をとる。<Minimum average pore diameter>, <Minimum in-plane porosity> and <〒47
〒3〉 Observe the cross section of the thick part of the porous membrane with a scanning electron microscope, and determine the minimum part of the pore diameter within a range of 10% error with respect to the film thickness (di). An ultrathin section with a thickness of about 0.1 μm is prepared parallel to the membrane surface of the membrane, and an electron microscopy copy of this section is taken.
注目する切片のｌｃＪ当りの孔半径が（ｊｌ〜（ｒ＋ｄ
ｒ）に存在する孔の数をＮｆｒｌｄｒと表示する。３次
および４次の極小平均孔径（それぞれ〒３および７４）
および極小面内空孔率Ｐｒは次式で定義される。The hole radius per lcJ of the section of interest is (jl~(r+d
The number of holes present in r) is denoted as Nfrldr. Tertiary and quaternary minimal average pore diameters (3 and 74, respectively)
and the minimum in-plane porosity Pr are defined by the following equation.
＝
＋１１ｆ ｒ’Ｎ（ｒ）ｄｒ
ｒ４／〒３は＋１１ 、 （２１式で算出された〒３
、〒４から直接計算される。極小平均孔径は２〒４で与
えられる。孔径分布関数Ｎ１ｒ）は以下のように超薄切
片の電子顕微鏡写真より定める。=
+11f r'N(r)dr r4/〒3 is +11, (〒3 calculated by formula 21
, calculated directly from 〒4. The minimum average pore diameter is given by 2〒4. The pore size distribution function N1r) is determined from an electron micrograph of an ultrathin section as follows.
すなわち、孔径分布を評価したい部分の走査型電子顕微
鏡写真を適当な大きさくたとえば２０ｃｍｘ２０ｅｍ）
に拡大焼付けし、得られた写真上に等間隔にテストライ
ン（直線）を２０本描く。おのおのの直線は多数の孔を
横切る。孔を横切った際の孔内に存在する直線の長さを
測定し、この長さの頻度分布関数を用いて、たとえばス
テレオロジ（たとえば諏訪紀夫著″定量形態学″岩波書
店）の方法でＮ ｔｒ＋を定める。That is, take a scanning electron micrograph of the part where you want to evaluate the pore size distribution and cut it to an appropriate size (for example, 20 cm x 20 em).
20 test lines (straight lines) were drawn at equal intervals on the resulting photograph. Each straight line crosses a number of holes. The length of the straight line existing in the hole when it crosses the hole is measured, and the frequency distribution function of this length is used to determine the N Define tr+.
く面内配向度〉
理学電機社製Ｘ線発生装置（ＲＵ−２００Ｐい、ゴニオ
メータ−（ＳＧ−９Ｒ） 、計数管にはシンチレーショ
ンカウンター、計数部には波高分析器（ＰＨＡ）を用い
、ニッケルフィルターで単色化したＣｕ −Ｋａ線（波
長λ−１，５４２人）で対称透過法により測定した。多
孔膜の表裏面を積重ねて、平行に束ねて、Ｘ線を膜面に
平行な方向に入射するように、理学電機社製の繊維試料
測定装置（回転試料台）に固定した。スキャニング速度
４°／分、チャート速度／Ｃｍ／分、タイムコンスタン
ト１〜２秒、ダイパージェットスリット２ｍｍφ、シシ
ービングスリット縦幅１．９ ｍｍ Ｘ横幅３．５ｍｍ
、ｉｉ度３０℃、相対温度５０％の条件下で、一定回折
角度（回折角２θ−２１，５°−（００２）面の回折角
に対応）で子午線から赤道線を経て再び子午線に至る１
８０°の方位角方向のＸ線回折強度曲線を測定した。こ
の曲線の半価幅Ｈ（度）を読取り、この値を（４）式に
代入すれば、面内配向度が算出できる。In-plane orientation> Rigaku Denki X-ray generator (RU-200P), goniometer (SG-9R), scintillation counter for the counter, pulse height analyzer (PHA) for the counting section, nickel filter The measurements were carried out using the symmetrical transmission method using monochromatic Cu-Ka rays (wavelength λ - 1,542).The front and back surfaces of the porous membranes were stacked and bundled in parallel, and the X-rays were incident in the direction parallel to the membrane surface. It was fixed to a fiber sample measuring device (rotating sample stand) manufactured by Rigaku Denki Co., Ltd. Scanning speed 4°/min, chart speed/Cm/min, time constant 1 to 2 seconds, diameter jet slit 2 mmφ, sissieving Slit length 1.9 mm x width 3.5 mm
, ii under the conditions of 30°C and 50% relative temperature, from the meridian to the equator line and back to the meridian at a constant diffraction angle (corresponding to the diffraction angle of the 2θ-21,5°-(002) plane).
The X-ray diffraction intensity curve in the 80° azimuthal direction was measured. By reading the half width H (degrees) of this curve and substituting this value into equation (4), the degree of in-plane orientation can be calculated.
面内配向度（％）＝（１８０−Ｈ）／１８０Ｘ１００
（４１〔（ホ）発明の効果〕
本発明の多孔膜は、生体への安全性および力学的強靭性
に優れ、水溶液中に分散する微生物の分離除去または微
生物の分離濃縮、あるいは、水溶液中のタンパク質の精
製に有利に適用できる。In-plane orientation degree (%) = (180-H)/180X100
(41 [(E) Effects of the Invention] The porous membrane of the present invention has excellent biological safety and mechanical toughness, and is capable of separating and removing microorganisms dispersed in an aqueous solution, separating and concentrating microorganisms, or It can be advantageously applied to protein purification.
以下、実施例について本発明を具体的に説明する。 The present invention will be specifically described below with reference to Examples.
実施例１
セルロース・リンター（αセルロース含有率９６％以上
、平均分子量２．６ Ｘ １０５）を公知の方法で調製
した銅アンモニア溶液中に６重量％の濃度で溶解した。Example 1 Cellulose linter (α-cellulose content 96% or more, average molecular weight 2.6×105) was dissolved at a concentration of 6% by weight in a copper ammonia solution prepared by a known method.
その後、ガラス板上に厚さ２５０μｍのアプリケーター
で流延し、直ちにアセトン／アンモニア／水（４５／
０．９ ／１００重量比）の混合液（２０℃）に３０分
間浸漬後、２０℃で２重量％の硫酸水溶液に１５分間浸
漬し、水洗した。その後、方形の枠に膜の周辺を固定し
、アセトン中に１５分間浸漬して膜中の水分をアセトン
で置換し、３０°Ｃで゛真空乾燥した。かくして得られ
た多孔膜の特性を第１表にまとめて示す。なお、極小平
均孔径を与える層は膜表面から１．Ｏｄ（裏面）に位置
していた。Then, it was cast onto a glass plate using a 250 μm thick applicator, and immediately acetone/ammonia/water (45/
0.9/100 weight ratio) for 30 minutes, then immersed in a 2% by weight sulfuric acid aqueous solution at 20°C for 15 minutes, and washed with water. Thereafter, the periphery of the membrane was fixed in a rectangular frame, immersed in acetone for 15 minutes to replace the moisture in the membrane with acetone, and vacuum dried at 30°C. The properties of the porous membrane thus obtained are summarized in Table 1. Note that the layer that provides the minimum average pore diameter is 1.0 mm from the membrane surface. It was located on Od (back side).
第１表 多孔膜の特性
０．１０．ｕｎ １２０μｍ ３８％ ７２％ ６
．２ １．１２上記多孔膜を金属製濾過器４７ｍ
ｍφに装着した。Table 1 Characteristics of porous membrane 0.10. un 120μm 38% 72% 6
．． 2 1.12 Pass the above porous membrane through a 47m metal filter.
It was attached to mφ.
生理的食塩水に卵白アルブミンを５ ｇ／ｄｐで熔解し
、さらに該液中にタバコモザイクうイルス（粒子径０．
１５８μｍ）を１０６個／ｍ７！の割合で分散させた。Ovalbumin was dissolved at 5 g/dp in physiological saline, and tobacco mosaic virus (particle size: 0.5 g/dp) was dissolved in the solution.
158μm) 106 pieces/m7! Distributed at a ratio of
該液を一ヒ述の濾過器で垂直濾過した。The liquid was vertically filtered using the filter described above.
濾液中のウィルス粒子数を電子顕微鏡法で測定した。そ
の結果、ウィルス粒子阻止率１００％、アルブミン阻止
率３％で、限外濾過速度の経時変化は濾過量Ｂでは濾過
初期の値の６０％であった。The number of virus particles in the filtrate was determined by electron microscopy. As a result, the virus particle inhibition rate was 100%, the albumin inhibition rate was 3%, and the change in ultrafiltration rate over time was 60% of the value at the initial stage of filtration at filtration amount B.
実施例２
実施例１と同様の方法でセルロース濃度６重量％の銅ア
ンモニア溶液を調製した。スリット幅３００μｍの直線
状のスリットより該溶液を、アセトン／アンモニア／水
（４５／ ０．９ ／１００重鼠仕）混合液中に紡出さ
せ、得られたシート状物の両端部を固定し、定長幅で２
０℃の該混合液中に１０分間浸漬し、引き続き２０°Ｃ
で２重量％の硫酸水溶液中で再生処理する。その後、連
続的に２０℃のアセトン中に浸漬して膜中の水をアセト
ンで置換する。この際、該膜の横幅が変化しないように
、シートの両端は把持しておいた。アセトンは空気中で
多孔膜から除去された。かくして得られた多孔膜の特性
を第２表にまとめて示す。なお、極小平均孔径を与える
層は膜表面から０．５ｄに位置していた。Example 2 A cuprammonium solution having a cellulose concentration of 6% by weight was prepared in the same manner as in Example 1. The solution was spun into a mixture of acetone/ammonia/water (45/0.9/100) through a linear slit with a slit width of 300 μm, and both ends of the resulting sheet were fixed. , 2 with constant length and width
Immerse in the mixture at 0°C for 10 minutes and then at 20°C.
The sample is regenerated in a 2% by weight aqueous sulfuric acid solution. Thereafter, the membrane is continuously immersed in acetone at 20° C. to replace water in the membrane with acetone. At this time, both ends of the sheet were held so that the width of the film did not change. Acetone was removed from the porous membrane in air. The properties of the porous membrane thus obtained are summarized in Table 2. Note that the layer providing the minimum average pore diameter was located 0.5 d from the membrane surface.
第２表 多孔膜の特性
０．１３μｍ １５０．ｃ＋ｍ ４２％ ７６％
５．６ １．１３実施例１と同様に生理的食塩水
に卵白アルブミンを溶解させ、タバコモザイクウィルス
を分散させた液を用いて、垂直濾過法で限外濾過速度の
経時変化および濾液の組成を測定した。濾過量１．４７
！で濾過初期の濾過速度の６０％となった。限外濾過速
度は実施例１の膜と比較して（同一の負荷圧、同一の有
効濾過面積下での比較）、約１．８倍であった。ウィル
ス阻止率１００％、アルブミン阻止率２％であった。Table 2 Characteristics of porous membrane 0.13μm 150. c+m 42% 76%
5.6 1.13 Similar to Example 1, ovalbumin was dissolved in physiological saline and tobacco mosaic virus was dispersed in the solution, and the ultrafiltration rate was measured over time and the composition of the filtrate using the vertical filtration method. was measured. Filtration amount 1.47
! The filtration rate was 60% of the initial filtration rate. The ultrafiltration rate was about 1.8 times higher than that of the membrane of Example 1 (comparison under the same load pressure and the same effective filtration area). The virus inhibition rate was 100% and the albumin inhibition rate was 2%.
Claims (1)
孔率が１０％以上の銅安法セルロース多孔膜において、
膜表面の平均孔径と極小平均孔径との比が４以上で、か
つ該極小平均孔径が分離すべき微生物粒子の最小の粒子
の粒子直径の１．０倍以下、０．３倍以上で、また４次
の平均孔半径＠ｒ＠＿４と３次の平均孔半径＠ｒ＠＿３
との比＠ｒ＠＿４／＠ｒ＠＿４が１．３以下である孔径
分布を有することを特徴とする微生物粒子分離用セルロ
ース多孔膜。 ２、銅安セルロース用セルロース原料として高純度セル
ロースである特許請求の範囲第１項記載のセルロース多
孔膜。 ３、極小面内空孔率が２０％以上、６０％以下であり、
多孔膜を構成するセルロース分子類の面内配向度が６０
％以上である特許請求の範囲第１項または第２項記載の
微生物粒子分離用多孔膜。 ４、極小平均孔径が分離すべき微生物粒子の最小の粒子
の粒子直径の０．９倍以下、０．５倍以上で＠ｒ＠＿４
／＠ｒ＠＿３≦１．１５を満足する孔径分布を有する特
許請求の範囲第１項から第３項までのいずれかに記載の
微生物粒子分離用多孔膜。 ５、膜厚（ｄ）が１０μｍ〜５００μｍであり、かつ極
小平均孔径を与える層が膜表面から０．９ｄ〜０．１ｄ
の間に存在する特許請求の範囲第１項から第４項までの
いずれかに記載の微生物分離用多孔膜。 ６、分離すべき微生物粒子がリケッチャまたはウィルス
粒子である特許請求範囲第１項から第５項までのいずれ
かに記載の微生物分離用多孔膜。[Claims] 1. A copper ammonium cellulose porous membrane having a minimum average pore diameter of 0.02 to 0.2 μm and a minimum in-plane porosity of 10% or more,
The ratio of the average pore size of the membrane surface to the minimum average pore size is 4 or more, and the minimum average pore size is 1.0 times or less and 0.3 times or more the particle diameter of the smallest microorganism particle to be separated, and Fourth-order average hole radius @r@_4 and third-order average hole radius @r@_3
A cellulose porous membrane for separating microorganism particles, characterized in that it has a pore size distribution in which the ratio @r@_4/@r@_4 is 1.3 or less. 2. The cellulose porous membrane according to claim 1, which is a high-purity cellulose as the cellulose raw material for copper ammonium cellulose. 3. Minimum in-plane porosity is 20% or more and 60% or less,
The degree of in-plane orientation of cellulose molecules constituting the porous membrane is 60
% or more, the porous membrane for separating microorganism particles according to claim 1 or 2. 4. If the minimum average pore diameter is 0.9 times or less and 0.5 times or more the particle diameter of the smallest microorganism particle to be separated, @r@_4
The porous membrane for separating microorganism particles according to any one of claims 1 to 3, which has a pore size distribution satisfying /@r@_3≦1.15. 5. The film thickness (d) is 10 μm to 500 μm, and the layer providing the minimum average pore diameter is 0.9 d to 0.1 d from the film surface.
A porous membrane for separating microorganisms according to any one of claims 1 to 4 existing between the claims 1 to 4. 6. The porous membrane for separating microorganisms according to any one of claims 1 to 5, wherein the microorganism particles to be separated are rickettchers or virus particles.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP11680585A JPS61274707A (en) | 1985-05-31 | 1985-05-31 | Regenerated porous membrane for separating bacteria |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP11680585A JPS61274707A (en) | 1985-05-31 | 1985-05-31 | Regenerated porous membrane for separating bacteria |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| JPS61274707A true JPS61274707A (en) | 1986-12-04 |
Family
ID=14696095
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| JP11680585A Pending JPS61274707A (en) | 1985-05-31 | 1985-05-31 | Regenerated porous membrane for separating bacteria |
Country Status (1)
| Country | Link |
|---|---|
| JP (1) | JPS61274707A (en) |
Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US4808315A (en) * | 1986-04-28 | 1989-02-28 | Asahi Kasei Kogyo Kabushiki Kaisha | Porous hollow fiber membrane and a method for the removal of a virus by using the same |
| US4818258A (en) * | 1987-11-05 | 1989-04-04 | L&H Technologies, Inc. | Filter module |
| US4941897A (en) * | 1987-11-05 | 1990-07-17 | L & H Technologies, Inc. | Microporous filter and method |
| WO2009060836A1 (en) * | 2007-11-05 | 2009-05-14 | Asahi Kasei Fibers Corporation | Cellulose porous membrane |
Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPS5889628A (en) * | 1981-11-25 | 1983-05-28 | Asahi Chem Ind Co Ltd | Regenerated cellulose porous membrane |
| JPS5889626A (en) * | 1981-11-25 | 1983-05-28 | Asahi Chem Ind Co Ltd | Tough regenerated cellulose porous membrane |
| JPS59199728A (en) * | 1983-04-28 | 1984-11-12 | Asahi Chem Ind Co Ltd | Manufacture of regenerated cellulose membrane having large pore size |
| JPS59204912A (en) * | 1983-05-02 | 1984-11-20 | Asahi Chem Ind Co Ltd | Preparation of hollow yarn of regenerated cellulose |
-
1985
- 1985-05-31 JP JP11680585A patent/JPS61274707A/en active Pending
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPS5889628A (en) * | 1981-11-25 | 1983-05-28 | Asahi Chem Ind Co Ltd | Regenerated cellulose porous membrane |
| JPS5889626A (en) * | 1981-11-25 | 1983-05-28 | Asahi Chem Ind Co Ltd | Tough regenerated cellulose porous membrane |
| JPS59199728A (en) * | 1983-04-28 | 1984-11-12 | Asahi Chem Ind Co Ltd | Manufacture of regenerated cellulose membrane having large pore size |
| JPS59204912A (en) * | 1983-05-02 | 1984-11-20 | Asahi Chem Ind Co Ltd | Preparation of hollow yarn of regenerated cellulose |
Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US4808315A (en) * | 1986-04-28 | 1989-02-28 | Asahi Kasei Kogyo Kabushiki Kaisha | Porous hollow fiber membrane and a method for the removal of a virus by using the same |
| US4818258A (en) * | 1987-11-05 | 1989-04-04 | L&H Technologies, Inc. | Filter module |
| US4941897A (en) * | 1987-11-05 | 1990-07-17 | L & H Technologies, Inc. | Microporous filter and method |
| WO2009060836A1 (en) * | 2007-11-05 | 2009-05-14 | Asahi Kasei Fibers Corporation | Cellulose porous membrane |
| JPWO2009060836A1 (en) * | 2007-11-05 | 2011-03-24 | 旭化成せんい株式会社 | Cellulosic porous membrane |
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