JPH09121889A - Nondestructive viability evaluation of tightly sealed and freeze-dried microorganism - Google Patents
Nondestructive viability evaluation of tightly sealed and freeze-dried microorganismInfo
- Publication number
- JPH09121889A JPH09121889A JP28358995A JP28358995A JPH09121889A JP H09121889 A JPH09121889 A JP H09121889A JP 28358995 A JP28358995 A JP 28358995A JP 28358995 A JP28358995 A JP 28358995A JP H09121889 A JPH09121889 A JP H09121889A
- Authority
- JP
- Japan
- Prior art keywords
- freeze
- microorganism
- sealed
- container
- light
- 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.)
- Pending
Links
- 244000005700 microbiome Species 0.000 title claims abstract description 39
- 230000035899 viability Effects 0.000 title claims abstract description 8
- 238000011156 evaluation Methods 0.000 title abstract 2
- CURLTUGMZLYLDI-UHFFFAOYSA-N Carbon dioxide Chemical compound O=C=O CURLTUGMZLYLDI-UHFFFAOYSA-N 0.000 claims abstract description 34
- 239000007789 gas Substances 0.000 claims abstract description 20
- QVGXLLKOCUKJST-UHFFFAOYSA-N atomic oxygen Chemical compound [O] QVGXLLKOCUKJST-UHFFFAOYSA-N 0.000 claims abstract description 17
- 229910002092 carbon dioxide Inorganic materials 0.000 claims abstract description 17
- 239000001569 carbon dioxide Substances 0.000 claims abstract description 17
- 239000001301 oxygen Substances 0.000 claims abstract description 17
- 229910052760 oxygen Inorganic materials 0.000 claims abstract description 17
- 238000001069 Raman spectroscopy Methods 0.000 claims abstract description 6
- 230000002452 interceptive effect Effects 0.000 claims abstract description 4
- 238000001514 detection method Methods 0.000 claims abstract 2
- 239000000463 material Substances 0.000 claims abstract 2
- 238000001237 Raman spectrum Methods 0.000 claims description 21
- 238000004458 analytical method Methods 0.000 claims description 20
- 230000004083 survival effect Effects 0.000 claims description 13
- 238000000034 method Methods 0.000 claims description 9
- 230000003287 optical effect Effects 0.000 claims description 3
- 238000000605 extraction Methods 0.000 claims 1
- 230000001788 irregular Effects 0.000 claims 1
- 229940026085 carbon dioxide / oxygen Drugs 0.000 abstract 1
- IJGRMHOSHXDMSA-UHFFFAOYSA-N nitrogen Substances N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 description 11
- 229910052757 nitrogen Inorganic materials 0.000 description 5
- 239000003708 ampul Substances 0.000 description 4
- 239000000203 mixture Substances 0.000 description 4
- 238000001228 spectrum Methods 0.000 description 4
- 241000894006 Bacteria Species 0.000 description 3
- 230000034994 death Effects 0.000 description 3
- 231100000517 death Toxicity 0.000 description 3
- 230000000694 effects Effects 0.000 description 3
- 240000004808 Saccharomyces cerevisiae Species 0.000 description 2
- 230000002411 adverse Effects 0.000 description 2
- 238000007796 conventional method Methods 0.000 description 2
- 230000001066 destructive effect Effects 0.000 description 2
- PXEDJBXQKAGXNJ-QTNFYWBSSA-L disodium L-glutamate Chemical compound [Na+].[Na+].[O-]C(=O)[C@@H](N)CCC([O-])=O PXEDJBXQKAGXNJ-QTNFYWBSSA-L 0.000 description 2
- 239000003814 drug Substances 0.000 description 2
- 229940079593 drug Drugs 0.000 description 2
- 230000000813 microbial effect Effects 0.000 description 2
- 235000013923 monosodium glutamate Nutrition 0.000 description 2
- 239000003223 protective agent Substances 0.000 description 2
- 235000020183 skimmed milk Nutrition 0.000 description 2
- 229940073490 sodium glutamate Drugs 0.000 description 2
- 238000001845 vibrational spectrum Methods 0.000 description 2
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 2
- RNLHGQLZWXBQNY-UHFFFAOYSA-N 3-(aminomethyl)-3,5,5-trimethylcyclohexan-1-amine Chemical compound CC1(C)CC(N)CC(C)(CN)C1 RNLHGQLZWXBQNY-UHFFFAOYSA-N 0.000 description 1
- 241000186361 Actinobacteria <class> Species 0.000 description 1
- MYMOFIZGZYHOMD-UHFFFAOYSA-N Dioxygen Chemical compound O=O MYMOFIZGZYHOMD-UHFFFAOYSA-N 0.000 description 1
- 241000233866 Fungi Species 0.000 description 1
- 229910001873 dinitrogen Inorganic materials 0.000 description 1
- 229910001882 dioxygen Inorganic materials 0.000 description 1
- 238000004090 dissolution Methods 0.000 description 1
- 230000005284 excitation Effects 0.000 description 1
- 238000004108 freeze drying Methods 0.000 description 1
- 239000011521 glass Substances 0.000 description 1
- 230000005764 inhibitory process Effects 0.000 description 1
- 238000007689 inspection Methods 0.000 description 1
- 239000012528 membrane Substances 0.000 description 1
- 230000004060 metabolic process Effects 0.000 description 1
- 229910052751 metal Inorganic materials 0.000 description 1
- 239000002184 metal Substances 0.000 description 1
- VUZPPFZMUPKLLV-UHFFFAOYSA-N methane;hydrate Chemical compound C.O VUZPPFZMUPKLLV-UHFFFAOYSA-N 0.000 description 1
- 230000035772 mutation Effects 0.000 description 1
- QJGQUHMNIGDVPM-UHFFFAOYSA-N nitrogen(.) Chemical compound [N] QJGQUHMNIGDVPM-UHFFFAOYSA-N 0.000 description 1
- 239000012466 permeate Substances 0.000 description 1
- 239000000047 product Substances 0.000 description 1
- 230000029058 respiratory gaseous exchange Effects 0.000 description 1
- 238000002054 transplantation Methods 0.000 description 1
Landscapes
- Investigating, Analyzing Materials By Fluorescence Or Luminescence (AREA)
- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
Abstract
Description
【０００１】[0001]
【発明の属する技術分野】本発明は、微生物の生存に関
する非破壊検査法、更に詳しくは、凍結乾燥して長期間
密封保存した微生物（ライオファイル微生物）を当該密
封保存容器を破壊することなく、そのまま、即ち容器ご
とラマンスペクトル分析に付し、該容器内気体の炭酸ガ
ス対酸素の比に基いて該微生物が生存しているか死滅し
ているかを判定する方法に関する。TECHNICAL FIELD The present invention relates to a non-destructive inspection method for survival of microorganisms, more specifically, a microorganism (lyofile microorganism) freeze-dried and stored for a long period of time without destroying the sealed storage container. As it is, that is, the container is subjected to Raman spectrum analysis to determine whether the microorganism is alive or dead based on the ratio of carbon dioxide gas to oxygen in the gas in the container.
【０００２】[0002]
【従来の技術】無菌状態にある医薬品アンプル中の窒素
及び酸素ガスを回転ラマンスペクトル分析法により測定
し、アンプル中のガス充填が所定通りになされているか
否かを検査する方法が知られている（GLEN F. BAILEY a
nd HERBERT A. MOORE. JR.：Journal of the Parentera
l Drug Association, 34, (2)127-133(1980)）。しか
し、密封保存した凍結乾燥微生物を開封することなく、
振動ラマンスペクトル分析に付し、該微生物の生存性を
容器内気体における炭酸ガス対酸素の比に基いて判断す
る方法は知られていない。2. Description of the Related Art A method is known in which nitrogen and oxygen gas in an aseptic drug ampoule are measured by a rotating Raman spectrum analysis method to inspect whether or not the ampoule is filled with gas in a predetermined manner. (GLEN F. BAILEY a
nd HERBERT A. MOORE. JR .: Journal of the Parentera
l Drug Association, 34 , (2) 127-133 (1980)). However, without opening the freeze-dried microorganisms that have been sealed and stored,
There is no known method for determining the viability of the microorganism based on the ratio of carbon dioxide gas to oxygen in the gas in the container, which is subjected to vibrational Raman spectrum analysis.
【０００３】従来、微生物は、例えば当該微生物を凍結
乾燥し、アンプルに封入して保存している。そして、保
存中の生存を確認するためには、作製したアンプルの内
の何本かを実際に破壊していわゆる生存試験を行ない、
その結果から他の保存アンプルの生存又は死滅を推定し
ているものであって、目的のアンプルの生存又は死滅そ
のものを直接に判定しているわけではない。Conventionally, microorganisms are stored by, for example, freeze-drying the microorganisms and enclosing them in ampoules. Then, in order to confirm the survival during storage, some of the prepared ampoules were actually destroyed to perform a so-called survival test,
The result estimates the survival or death of other stored ampoules, and does not directly determine the survival or death itself of the target ampoule.
【０００４】[0004]
【発明が解決しようとする課題】前項記載の従来技術の
背景下に、本発明は、密封保存した凍結乾燥微生物を、
その容器を破壊することなしに容易に生存しているか否
かを判定することのできる方法を開発することを目的と
する。DISCLOSURE OF THE INVENTION Under the background of the prior art described in the preceding paragraph, the present invention provides a freeze-dried microorganism that has been sealed and stored,
It is an object to develop a method capable of easily determining whether or not a container is alive without destroying the container.
【０００５】[0005]
【課題を解決するための手段】本発明者は、前項記載の
目的を達成すべく鋭意研究の結果、凍結乾燥微生物を収
容した密封保存容器内の気体の組成、より具体的には該
気体における炭酸ガス対酸素の比と該微生物の生存性と
の間に相関関係があること、及び該密封保存容器内の気
体の組成は、ラマンスペクトル分析を利用すれば、容器
を破壊せずとも極めて容易に測定しうることを見出し、
このような知見に基いて本発明を完成した。Means for Solving the Problems As a result of intensive research to achieve the object described in the preceding paragraph, the present inventor has found that the composition of gas in a hermetically-sealed storage container containing freeze-dried microorganisms, more specifically, in the gas There is a correlation between the carbon dioxide to oxygen ratio and the viability of the microorganism, and the composition of the gas in the hermetically-sealed storage container is extremely easy to use using Raman spectrum analysis without destroying the container. To find that
The present invention has been completed based on such knowledge.
【０００６】すなわち、本発明は密封保存凍結乾燥微生
物をその容器を開封破壊することなくそのままラマンス
ペクトル分析に付し、容器内気体における炭酸ガス対酸
素の比に基いて該微生物の生存性を判定することを特徴
とする密封保存凍結乾燥微生物の非破壊生存判定法、及
びこのような方法の実施に適するラマンスペクトル分析
法に関する。That is, according to the present invention, the freeze-dried microorganisms stored in a sealed container are directly subjected to Raman spectrum analysis without breaking the container, and the viability of the microorganism is judged based on the ratio of carbon dioxide gas to oxygen in the gas in the container. And a Raman spectrum analysis method suitable for carrying out such a method.
【０００７】[0007]
【発明の実施の形態】以下、本発明を詳細に説明する。BEST MODE FOR CARRYING OUT THE INVENTION Hereinafter, the present invention will be described in detail.
【０００８】本発明にいう密封保存凍結乾燥微生物の微
生物としては、酵母、細菌、放線菌、糸状菌などの微生
物を挙げることができる。Examples of the microorganisms of the freeze-dried microorganisms that have been sealed and stored according to the present invention include microorganisms such as yeast, bacteria, actinomycetes and filamentous fungi.
【０００９】密封保存凍結乾燥微生物とは、上記微生物
をライオファイルチューブにスキムミルク、グルタミン
酸ナトリウム等の保護剤と共に封入し、凍結乾燥して減
圧下又は常圧下で封管したチューブを言う。The sealed-preserved lyophilized microorganism means a tube obtained by encapsulating the above microorganism in a lyofile tube together with a protective agent such as skim milk and sodium glutamate, lyophilized and sealed under reduced pressure or normal pressure.
【００１０】本発明によれば、このよう密封保存容器
は、微生物の凍結乾燥物（ライオファイル微生物）を収
容したままラマンスペクトル分析に付されるので、その
器壁は少なくとも一部が光線が容器内気体を貫通できる
ものではなくてはならない。According to the present invention, such a sealed storage container is subjected to Raman spectrum analysis while containing a lyophilized product of a microorganism (Liofile microorganism). It must be able to penetrate the internal gas.
【００１１】また、例えば、ライオファイルチューブ入
りの凍結乾燥微生物をそのまま直接測定した場合、ガラ
スチューブやアンプルによる乱反射の影響が大き過ぎる
ときは、乱反射に由来する妨害光を遮断するために、光
源波長を透過しない金属製等の特別の形状の容器を使用
するか又はフィルターを使用するか或いは両者を併用す
ることで、このような影響を除くことができる。そのよ
うな特別な形状の容器として、例えば、図１に示す筒状
容器を挙げることができる。この容器の径は、密封保存
容器（封管）の径に応じてある範囲で可変のものとする
こともできる。Further, for example, when the freeze-dried microorganisms contained in the lyofile tube are directly measured as they are, and the influence of the diffused reflection due to the glass tube or the ampoule is too great, the light source wavelength is used to block the interfering light derived from the diffused reflection. Such an influence can be eliminated by using a container having a special shape such as a metal that does not permeate through, a filter, or a combination of both. As a container having such a special shape, for example, a cylindrical container shown in FIG. 1 can be cited. The diameter of this container may be variable within a certain range depending on the diameter of the hermetically-sealed storage container (sealed tube).
【００１２】凍結乾燥微生物を密封保存容器に収容した
ままラマンスペクトル分析に付する方法は、例えば、次
のようにして行なうことができる。The method of subjecting the freeze-dried microorganisms to Raman spectrum analysis while keeping them in a sealed storage container can be carried out, for example, as follows.
【００１３】特別の形状の容器を使用する場合は、図１
に示すように、これに凍結乾燥微生物入りの密封保存容
器を収納してラマンスペクトル分析に付する。フィルタ
ーを使用する場合は、図２に示すように、光路の適当な
位置にフィルターをおいてラマンスペクトル分析に付す
る。図２においてフィルターとしては、励起レーザー光
波長のみを除去できるフィルターを使用することができ
る。なお、Ｓはサンプル（試料）を、そしてＩＰＤＡは
ダイオードアレイ（ｉｎｔｅｎｓｉｆｉｅｄｐｈｏｔｏ
ｄｉｏｄｅ ａｒｒａｙ）を意味する。他に、ＣＣＤ
（ｃｈａｒｇｅｃｏｕｐｌｅｄ ｄｅｖｉｃｅ）やＩＣ
ＣＤ（ｉｎｔｅｎｓｉｆｉｅｄ ＣＣＤ）なども用いる
ことができる。If a specially shaped container is used, FIG.
As shown in, a sealed storage container containing freeze-dried microorganisms is housed in this and subjected to Raman spectrum analysis. When a filter is used, the filter is placed at an appropriate position in the optical path as shown in FIG. 2 and subjected to Raman spectrum analysis. As the filter in FIG. 2, a filter that can remove only the wavelength of the excitation laser light can be used. In addition, S is a sample (sample), and IPDA is a diode array (intensified photo).
It means a diode array). Besides, CCD
(Charge coupled device) and IC
A CD (intended CCD) or the like can also be used.
【００１４】スペクトルとしては、振動スペクトル及び
回転スペクトルがあるが、本発明方法の実施において
は、そのいずれに拠ることもできる。ただし、前者の方
が、レーリー散乱光から波長的に遠いために乱反射によ
り妨害され難く、かつ低分解能の条件でも成分のスペク
トルが十分に分離して検査できる点で後者よりも優れて
いる。The spectrum includes a vibration spectrum and a rotation spectrum, which may be used in the practice of the method of the present invention. However, the former is superior to the latter in that it is less likely to be disturbed by diffuse reflection because it is far from the Rayleigh scattered light in terms of wavelength, and that the spectrum of the component can be sufficiently separated and inspected even under the condition of low resolution.
【００１５】ラマンスペクトル分析用光源としては、試
料に損傷を与えず、密封保存容器の器壁を透過するもの
であれば、その波長を問わず、例えば、Ａｒ+ レーザー
のレーザー光で 514.5ｎｍ、 488.0ｎｍなどの波長のも
のを使用することができる。振動スペククトルの場合、
Ｎ2 、Ｏ2 及びＣＯ2 は、それぞれ、ラマンシフトが23
31、1555及び1388ｃｍ-1（他にフェルミ共鳴により1286
ｃｍ-1にも出現）である。As a light source for Raman spectrum analysis, as long as it does not damage the sample and transmits through the vessel wall of the hermetically-sealed storage container, regardless of its wavelength, for example, a laser beam of Ar + laser 514.5 nm, A wavelength of 488.0 nm or the like can be used. For vibration spectrum,
N 2 , O 2 and CO 2 each have a Raman shift of 23.
31, 1555 and 1388 cm -1 (1286 by Fermi resonance
It also appears in cm -1 ).
【００１６】このような機器を使用してデータを採取
し、そして採取したデータの処理は、例えば、次のよう
にして行なうことができる。サンプルの凍結乾燥微生物
入り密封保存容器と同様の空の容器に空気を封入したも
のをラマンスペクトル分析に付してブランクとして測定
し、機器の補正を行なう。次に、サンプルの密封保存容
器中の窒素、酸素及び二酸化炭素をラマンスペクトル分
析に付して測定し、窒素に対する酸素の比（Ｏ2 ／
Ｎ2 ）及び窒素に対する二酸化炭素の比（ＣＯ2 ／
Ｎ2 ）を算出し、前者の比で後者の比を除して酸素に対
する二酸化炭素の比（ＣＯ2 ／Ｏ2 ）を算出する。ここ
で、それぞれの比の値は、本来、スペクトルのピークの
面積の比を求めるべきであるが、半値幅ほぼ等しい場合
においてはそのピークの高さの比を用いてもかまわない
（後掲実施例１はこれによっている）。酸素に対する二
酸化炭素の比は、比較の便宜上から空気中の酸素に対す
る二酸化炭素の比を１として、これとの比較の値（相対
値）で表わすこともできる。また、もちろん、ラマンス
ペクトル上での、ブランク値との絶対強度の比較から二
酸化炭素及び酸素の存在量そのものを定量又は半定量
し、これらの値を使用することもできる。Data can be collected using such a device, and the collected data can be processed, for example, as follows. An empty container similar to the sealed storage container containing freeze-dried microorganisms of the sample filled with air is subjected to Raman spectrum analysis and measured as a blank, and the instrument is corrected. Next, the nitrogen, oxygen and carbon dioxide in the sealed storage container of the sample were measured by Raman spectrum analysis, and the ratio of oxygen to nitrogen (O 2 /
N 2 ) and the ratio of carbon dioxide to nitrogen (CO 2 /
N 2) was calculated, by dividing the latter ratio in the former ratio is calculated the ratio of carbon dioxide to oxygen (CO 2 / O 2). Here, for each ratio value, the ratio of the peak areas of the spectrum should be obtained originally, but if the half widths are almost equal, the ratio of the peak heights may be used (see below). Example 1 depends on this). For convenience of comparison, the ratio of carbon dioxide to oxygen can be expressed as a value (relative value) for comparison with the ratio of carbon dioxide to oxygen in the air being 1. Further, of course, it is also possible to quantitatively or semiquantitatively determine the existing amounts of carbon dioxide and oxygen based on the comparison of the absolute intensity with the blank value on the Raman spectrum and use these values.
【００１７】本発明者の知見によれば、このようにラマ
ンスペクトル分析によって得られた密封容器の気体にお
ける炭酸ガス対酸素の体積比（ＣＯ2 ／Ｏ2 ）が大きい
ものは当該微生物が死滅しており、小さいものは生存し
ている。後掲実施例参照。因みに、これは、炭酸ガス対
酸素の比が大きいものは、保存中も微生物菌体が呼吸し
て酸素が消費され、炭酸ガス及び水が生成し、これらが
菌体の生存にとって、膜構造の変化、代謝阻害、水への
ＣＯ2 の溶解によるｐＨ低下などの悪影響を及ぼしたも
のと推定され、一方、小さいものは、保存中の菌体によ
る呼吸がほとんどなく、そのような悪影響がなかったも
のであることによるものと推定される。According to the knowledge of the inventor of the present invention, the microorganism is killed in the case where the volume ratio of carbon dioxide gas to oxygen (CO 2 / O 2 ) in the gas in the sealed container obtained by the Raman spectrum analysis is large. And small ones are alive. See the examples below. By the way, when the ratio of carbon dioxide gas to oxygen is large, microbial cells breathe and oxygen is consumed even during storage, carbon dioxide and water are produced, and these are factors of the membrane structure for the survival of the cells. It is presumed that adverse effects such as change, inhibition of metabolism, and pH decrease due to dissolution of CO 2 in water were exerted. On the other hand, small ones had almost no respiration by stored cells and did not have such adverse effects. It is presumed that this is due to the fact that it is a thing.
【００１８】そこで、ある密封保存凍結乾燥微生物につ
いて、ラマンスペクトル分析に付して得られた炭酸ガス
対酸素の比と、これを従来法により生存試験を行なって
得られた生存状況との関係は、同様の条件で保存されて
きた他の密封保存凍結乾燥微生物にも当てはまるので、
後者の微生物については、従来法による生存試験を行な
わなくても、前者の微生物についての生存状況から推定
してその生存状況が容易に判定できるのである。Therefore, the relationship between the ratio of carbon dioxide gas to oxygen obtained by subjecting a certain lyophilized microorganism to be preserved in a sealed state to Raman spectrum analysis and the survival condition obtained by conducting a survival test by the conventional method is as follows. , As it applies to other sealed and preserved freeze-dried microorganisms that have been preserved under similar conditions,
For the latter microorganisms, the survival status can be easily estimated by estimating from the survival status of the former microorganisms without conducting a survival test by the conventional method.
【００１９】[0019]
【実施例】以下、実施例により本発明を更に説明する。The present invention will be further described with reference to the following examples.
【００２０】実施例１ ライオファイルチューブ（保護剤：スキムミルク10％、
グルタミン酸ナトリウム２％）で31年間10℃で保存した
酵母12株を、それぞれ、ライオファイルチューブ入りの
まま乱反射防止用アダプター（図１の特殊形状の容器）
に収納しＡｒ+ レーザー光を光源とするラマンスペクト
ル分析に付し、直接チューブ中のガス組成を測定した。
窒素、酸素及び二酸化炭素を測定し、空気中の窒素を基
準としてＯ2 ／Ｎ2 及びＣＯ2 ／Ｎ2 の値からＣＯ2 ／
Ｏ2 の値を算出した。Example 1 Lyofile tube (protective agent: skim milk 10%,
12 yeast strains stored at 10 ° C for 31 years in sodium glutamate (2%), respectively, with an adapter for anti-diffuse reflection in a lyofile tube (container of special shape in Fig. 1)
The sample was housed in a tube and subjected to Raman spectrum analysis using Ar + laser light as a light source to directly measure the gas composition in the tube.
Nitrogen, oxygen and carbon dioxide were measured, O and nitrogen in the air as the reference 2 / N 2 and CO 2 / N 2 of the CO from the value 2 /
The value of O 2 was calculated.
【００２１】その後、ガス組成を測定したライオファイ
ルチューブを無菌的に開管し、いわゆる生存試験に付し
てチューブ中の生菌数の計数を行なった。Then, the Lyophile tube whose gas composition was measured was opened aseptically and subjected to a so-called survival test to count the number of viable bacteria in the tube.
【００２２】結果を下記第１表に示す。The results are shown in Table 1 below.
【００２３】[0023]
【表１】 [Table 1]
【００２４】第１表から理解されるように、空気中のＣ
Ｏ2 ／Ｏ2 ＝0.0016との相対値が 340以上のチューブで
は、いずれも生菌が検出されず、死滅していたが、一
方、 230以下のものは生存が確認されたので、同様の条
件における保存株については、300を境界として、それ
以下を生存、そしてそれ以上を死滅とする判定を非破壊
的に行なうことが可能である。As can be seen from Table 1, C in air
In the tubes with a relative value of O 2 / O 2 = 0.0016 of 340 or more, no viable bacteria were detected and the cells died. On the other hand, those of 230 or less were confirmed to be alive. With respect to the preserved strains in No. 3, it is possible to make a non-destructive determination that the number of surviving points is less than 300 and the number of deaths is more than 300, with 300 as the boundary.
【００２５】[0025]
【発明の効果】密封保存冷凍乾燥微生物が生存している
か死滅しているかを密封保存容器を開封破壊することな
く短時間で直接判定することができる。このことから、
密封保存凍結乾燥微生物の作成本数を大幅に削減するこ
とができ、また、無駄な植え継ぎによる微生物の変異の
危険を防止できる。さらに、ＣＯ2 ／Ｏ2 の比を経時的
に測定することにより、当該保存微生物の死滅時期を予
想することが容易となる。EFFECTS OF THE INVENTION Whether the freeze-dried microorganisms stored in a sealed state are alive or dead can be directly determined in a short time without opening and breaking the sealed storage container. From this,
The number of freeze-dried microorganisms stored in a sealed manner can be significantly reduced, and the risk of microbial mutation due to unnecessary transplantation can be prevented. Further, by measuring the CO 2 / O 2 ratio with time, it becomes easy to predict the dead time of the stored microorganism.
【図１】ラマンスペクトル分析を行なうときの、密封保
存容器を収納すべき容器を例示する。FIG. 1 illustrates a container in which a hermetically-sealed storage container is to be housed when performing Raman spectrum analysis.
【図２】ラマンスペクトル分析を行なうときの、光路に
おくべきフィルターの位置を例示する。FIG. 2 illustrates the position of a filter to be placed in the optical path when performing Raman spectrum analysis.
───────────────────────────────────────────────────── フロントページの続き (72)発明者 石原 勝 神奈川県川崎市川崎区鈴木町１−１ 味の 素株式会社中央研究所内 (72)発明者 山中 茂 神奈川県川崎市川崎区鈴木町１−１ 味の 素株式会社中央研究所内 ─────────────────────────────────────────────────── ─── Continuation of the front page (72) Inventor Katsu Ishihara 1-1, Suzuki-cho, Kawasaki-ku, Kawasaki-shi, Kanagawa Ajinomoto Co., Inc. Central Research Laboratory (72) Inventor Shigeru Yamanaka 1-Suzuki-cho, Kawasaki-ku, Kawasaki-shi, Kanagawa 1 Central Research Institute of Ajinomoto Co., Inc.
Claims (2)
封することなくそのままラマンスペクトル分析に付し、
容器内気体における炭酸ガス対酸素の比に基いて該微生
物の生存性を判定することを特徴とする密封保存凍結乾
燥微生物の非破壊生存判定法。1. A sealed and freeze-dried microorganism is directly subjected to Raman spectrum analysis without opening the container,
A nondestructive method for determining the survival of a freeze-dried microorganism in a hermetically sealed state, which comprises determining the viability of the microorganism based on the ratio of carbon dioxide gas to oxygen in the gas in the container.
害光を遮断すべく、光源波長を透過しない材質からな
る、入射光入り口、透過光出口及びラマン散乱光取り出
し口を有する容器に該密封保存容器を収納して、並びに
／或いは、該妨害光のみを遮断する又は検出光波長近辺
のみを透過するフィルターを光路に使用してラマンスペ
クトル分析を行うことを特徴とする請求項１記載の方
法。2. A container having an incident light inlet, a transmitted light outlet, and a Raman scattered light extraction port, which is made of a material that does not transmit a light source wavelength, in order to block disturbing light caused by irregular reflection by the hermetically-sealed storage container. The method according to claim 1, characterized in that the Raman spectrum analysis is carried out by using a filter for accommodating and / or storing only the interfering light or transmitting only the vicinity of the detection light wavelength in the optical path.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP28358995A JPH09121889A (en) | 1995-10-31 | 1995-10-31 | Nondestructive viability evaluation of tightly sealed and freeze-dried microorganism |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP28358995A JPH09121889A (en) | 1995-10-31 | 1995-10-31 | Nondestructive viability evaluation of tightly sealed and freeze-dried microorganism |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| JPH09121889A true JPH09121889A (en) | 1997-05-13 |
Family
ID=17667471
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| JP28358995A Pending JPH09121889A (en) | 1995-10-31 | 1995-10-31 | Nondestructive viability evaluation of tightly sealed and freeze-dried microorganism |
Country Status (1)
| Country | Link |
|---|---|
| JP (1) | JPH09121889A (en) |
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