CA1247055A - Generation of anaerobic or microaerophilic atmosphere - Google Patents
Generation of anaerobic or microaerophilic atmosphereInfo
- Publication number
- CA1247055A CA1247055A CA000478712A CA478712A CA1247055A CA 1247055 A CA1247055 A CA 1247055A CA 000478712 A CA000478712 A CA 000478712A CA 478712 A CA478712 A CA 478712A CA 1247055 A CA1247055 A CA 1247055A
- Authority
- CA
- Canada
- Prior art keywords
- level
- acid
- package
- water soluble
- accordance
- 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.)
- Expired
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Classifications
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
- C12M29/00—Means for introduction, extraction or recirculation of materials, e.g. pumps
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
- C12M41/00—Means for regulation, monitoring, measurement or control, e.g. flow regulation
- C12M41/30—Means for regulation, monitoring, measurement or control, e.g. flow regulation of concentration
- C12M41/34—Means for regulation, monitoring, measurement or control, e.g. flow regulation of concentration of gas
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- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y10—TECHNICAL SUBJECTS COVERED BY FORMER USPC
- Y10S—TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y10S435/00—Chemistry: molecular biology and microbiology
- Y10S435/801—Anerobic cultivation
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- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y10—TECHNICAL SUBJECTS COVERED BY FORMER USPC
- Y10S—TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y10S435/00—Chemistry: molecular biology and microbiology
- Y10S435/81—Packaged device or kit
Abstract
GENERATION OF ANAEROBIC OR
MICROAEROPHILIC ATMOSPHERE
ABSTRACT OF THE DISCLOSURE
A package is provided for all or part of the oxygen in a gaseous atmosphere within the package.
The package includes a gas impermeable, sealable container having a sachet disposed therein. The sachet contains an oxygen reactive material, pref-erably powdered iron. A compartment is provided within the container which is in fluid communication with the sachet location. The compartment is adapted to receive a predetermined level of water.
MICROAEROPHILIC ATMOSPHERE
ABSTRACT OF THE DISCLOSURE
A package is provided for all or part of the oxygen in a gaseous atmosphere within the package.
The package includes a gas impermeable, sealable container having a sachet disposed therein. The sachet contains an oxygen reactive material, pref-erably powdered iron. A compartment is provided within the container which is in fluid communication with the sachet location. The compartment is adapted to receive a predetermined level of water.
Description
GENERATION OF ANAEROsI~ OR
MIC~OAEROP~ILIC ATMOSPHERE
Field of the Invention The present invention relates generally to an apparatus and method for the generation of an anaero=
bic or microaerophilic atmosphere which is conducive to the growth of certain microorganisms.
Description of the Prior Art It is well known that some microorganisms require an aerobic atmosphere for growth, others require an anaerobic atmosphere, and still others require a microaerophilic atmosphere in which the oxygen level is between aerobic and anaerobic. In addition, some microorganisms require elevated levels of carbon dioxide for enhanced growth.
The attainment of an aerobic atmosphere is relatively simple, in most cases merely requiring aeration of the culture media~ Anaerobic conditions are more difficult to attain and the prior art contains many devices and processes for producing anaerobic atmospheres. U.S. Pat. No. 3,246,959, to Brewer discloses a device for generating such anaerobic atrnospheres by generation of hydrogen for reaction with oxygen in the atmosphere of a gas-tight apparatus such as that disclosed in United States Patent No. 3,483,089 to Brewer. The reaction between the hydrogen and oxygen is catalyzed by a platinum ' catalyst in the anaerobic apparatus.
United States Patent No. 4,347~222 to Beall discloses disposition of the catalyst in one recep-tacle of a unitary apparatus. This apparatus re-quires means for puncturing seals between several of ,,,. ~ ;~
~ 5~ P-816
MIC~OAEROP~ILIC ATMOSPHERE
Field of the Invention The present invention relates generally to an apparatus and method for the generation of an anaero=
bic or microaerophilic atmosphere which is conducive to the growth of certain microorganisms.
Description of the Prior Art It is well known that some microorganisms require an aerobic atmosphere for growth, others require an anaerobic atmosphere, and still others require a microaerophilic atmosphere in which the oxygen level is between aerobic and anaerobic. In addition, some microorganisms require elevated levels of carbon dioxide for enhanced growth.
The attainment of an aerobic atmosphere is relatively simple, in most cases merely requiring aeration of the culture media~ Anaerobic conditions are more difficult to attain and the prior art contains many devices and processes for producing anaerobic atmospheres. U.S. Pat. No. 3,246,959, to Brewer discloses a device for generating such anaerobic atrnospheres by generation of hydrogen for reaction with oxygen in the atmosphere of a gas-tight apparatus such as that disclosed in United States Patent No. 3,483,089 to Brewer. The reaction between the hydrogen and oxygen is catalyzed by a platinum ' catalyst in the anaerobic apparatus.
United States Patent No. 4,347~222 to Beall discloses disposition of the catalyst in one recep-tacle of a unitary apparatus. This apparatus re-quires means for puncturing seals between several of ,,,. ~ ;~
~ 5~ P-816
-2-the receptacles. The puncture means is supplied by a device which is separate from the gas generating apparatus or as part of a mated container.
The reaction between hydrogen and oxygen in the presence of a catalyst is strongly exothermic.
Flashing, and even explosion, can occur at the catalyst surface, particularly if the catalyst is finely divided and no means are provided to dissipate the heat generated.
United States Patent No. 4,013,422 to Spinner discloses a container having a material for gener-ating a reducing gas, such as hydrogen, for reaction, in the presence of a catalyst, with oxygen. The Spinner apparatus, however, uses an exposed catalyst pellet with no pro~ision for heat removal. In addition, it relies on the breaking of an ampoule containing a liquid and thereby instantaneous release of the liquid into contact with the material for generating the reducing gas. Thus no means are provided for control]ing the rate at which the contact is made. Experience has shown that contact at a slow and controlled rate is essential for accurate attainment of pre-determined final oxygen levels, particularly in those cases where it is desired to reduce, but not eliminate, the oxygen in the atmosphere.
U.S. Pat. No. 4,287,306 to Brewer describes a further apparatus for generating anaerobic atmo-spheres. In accordance with this patent, a flexible sealed package is provided with a catalyst coated onto the exterior surface of the package for use in 7~5~i P- 816 catalyzing the reaction between oxygen outside the package and hydrogen generated within the package.
This apparatus, like the Spinner apparatus, has the disadvantage of exposed active catalyst.
United States Patent No. 4,289,855 to Whitley discloses a safety catalyst package designed to reduce the danger of flashing or explosion. The Whitley package encloses a catalyst within holes and folds in a metal foil net. The net, which is inside of a container having holes for gas exchange, is composed of a heat conducting material, and thereby functions to remove heat from the catalyst vicinity. The Whitley disclosure is of a safety catalyst package only, and makes no provision for supply of hydrogen.
A package, for use in an anaerobic jar and specifically designed for attainment of a microaero-philic atmosphere, is disclosed in United States Patent 4,377,554 to Johnson. The Johnson invention relies on control of "wetover" and "condensation"
times for successful generation of microaerophilic atmospheres, and uses conventional exposed catalysts attached either to the outside of the package or to the lid of the jar.
A cardboard package for use in generating an anaerobic atmosphere is commercially available from the BBL Microbiology Systems Division of Becton DicXinson Company. This package includes a catalyst chamber mounted in the side of the box. The catalyst chamber is porous on both sides to permit generated hydrogen to flow through the chamber and thus react with the catalyst.
The prior art discussed above generally de-scribes apparatus for attaining an anaerobic atmo-sphere wherein hydrogen is generated and caused ~o react with the oxygen present in a vessel by means of a catalyst. Great Britain Patent No. 2109406 to Kasugai describes a process for culturing anaerobic bacteria wherein an agent which is reactive with oxygen is enclosed in an air tight closed vessel.
The agent contains a composition which is capable of removing oxygen and generating carbon dioxide in a volumetric ratio of about approximately twice as much oxygen removed as carbon dioxide generated. The preferred oxygen reactive material disclosed in the Kasugai patent is powdered iron.
Thus the invention provides in a broad aspect a package for removing oxygen from a gaseous mixture within a confined space comprising: (a) a sealable envelope, (b) a gas permeable sachet located within said envelope containing (l) an oxygen reactive material selected from the group consisting of iron powder, copper powder and aluminum powder, (2) a carbon dioxide generating composition including a water soluble, solid acid and a water soluble carbonate, (3) a filler selected from the group consisting of diato-maceous earth, charcoal, cellulose fiber, silica gel and barium sulfate and (4) a water dispersive and wetting surfactant, (c) a compartment within said envelope in fluid communication with said sachet location for receiving a predetermined level of water.
In another aspect the invention provides an article for use in removing oxygen from a gaseous mixture in a confined space comprising: a gas permeable sachet having the following components ~;~47~55 -4a-disposed therein: tl) an oxygen reactive material selected from the group consisting of iron powder, copper powder and aluminum powder, (2) a carbon dioxide generating composition including a water soluble, solid acid and a water soluble carbonate, (3) a filler selected from the group consisting of diatomaceous earth, charcoal, cellulose fiber, silica gel and barium sulfate;and (4) a waterdispersive andwetting surfactant.
Summary of the Invention In accordance with one embodiment of the present invention, there is provided a package for removing all or part of the oxygen in a gaseous atmosphere within the package. The package includes a gas impermeable, sealable container having a sachet disposed therein. The sachet contains an oxygen reactive material. A compartment is provided within the container which is in fluid communication with the sachet location. The compartment is adapted to receive a predetermined level of water.
In use, one or more petri dishes having a prepared media and havinq been inoculated with a sample suspected of containing an anaerobic micro-organism are inserted into the container adjacent to the sachet. A pipette or a vial is then used to ~Z~7~S5 P-81~
dispense a predetermined level of water into the compartment. The water hastens reaction between any oxygen contained within the envelope and the oxygen reactive material. The container is then sealed by suitable means. In a preferred embodiment of the invention the container is a flexible envelope and oxygen reactive material is powdered iron. The oxygen reacts with the powde~ed iron to form iron oxide. After a suitable period of time has elapsed the oxygen level contained within the envelope is, reduced to a level appropriate for the growth for anaerobic organisms, e.g. less than about 2 percent by volume.
The present invention provides an extremely simple, easily used, self contained device for generation of an anaerobic atmosphere without the hazards associated with the use of hydrogen and a catalyst.
Brief Description of the Drawings Figure 1 is an exploded view of the anaerobic generating apparatus of the invention showing the sequence of assembling the package;
Fiyure 2 is a front view of the assembled package;
Figure 3 is a side view of the assembled pack-age; and Figure 4 is a view of the water containing vial of the invention showing the arrangement of the tear strip used to seal the end of the vial.
Detailed Description of the Invention Referring now to Figure 1, there is shown a package for removing oxygen from a gaseous atmo-sphere. The package is in the form of an envelope 11. Envelope 11 is made of a suitable material which is impervious to the atmosphere and moisture. Thus, for example, the envelope 11 may be formed of a laminated plastic material, such as polyvinylidene chloride/polyester/polyethylene. The envelope 11 may be formed from two panels suitably secured together around the edges by heat sealing.
The interior of the envelope contains a sachet 13 which includes an oxygen reactive material. The sachet 13 is secured in place within the envelope 11 by any suitable means. One suitable means is to place one edge of the sachet between the two panels of plastic which are used to form the envelope prior to heat sealing and to heat seal the plastic over a portion of the sachet. Another suitable means would be to heat seal an area of the envelope adja-cent to the sachet so as to provide an open pocket for restraining the sachet in a suitable position~
The sachet is formed of a porous material, such as water resistant paper, to permit the water to pene-trate to the inside of the sachet and to permit reaction of the gaseous atmosphere within the en-velope with the contents of the sachet.
,~ As indicated, the sachet contains a material which is reactive with oxygen so as to remove the oxygen from the gaseous atmosphere within the en-velope. A suitable o~ygen reactive material is a ~2~55 P-816 metal powder provided in a red1ced state of oxygena-tion. Suitable metal powders include iron, copper, aluminum. A preferred metal powder is powdered iron. It has been determined tha~ the use of iron powder having a particle size 95 percent less than 325 mesh U.S. Standard Sieve Size is a most preferred oxygen reactive material for use in the invention.
For an embodiment of the invention wherein it is desired to provide an atmosphere having a predeter-mined level of carbon dioxide, a carbon dioxide generating composition is provided. The carbon dioxide generating composition includes a water soluble solid acid and a water soluble carbonate in amounts suitable for generating carbon dioxide~ As representative examples of suitable acids there may be mentioned: citric, tartaric, ascorbic, succinic, malic, fumaric, lactic acids and the like. AS
representative examples of suitable carbonates, there may be mentioned sodium bicarbonate, sodium car-bonate, potassium carbonate, sodium sesquicarbonate, etc. A preferred composition includes citric acid and sodium bicarbonate~ The carbon dioxide gener-ating composition is preferably employed in the form of a powder which is 100 percent less than 200 mesh U.S. Standard seive size and which is dispersed with the powdered iron.
It has also been determined that to provide a suitable low level of oxygen required for anaerobic conditions within an appropriate time, the presence of a water absorbing filler is important. Suitable fillers include diatomaceous earth, charcoal, cellu-lose fiber, silica gel and barium sulfate. In ~L24~ 5~
particular, diatomaceous earth has been found to be suitable to provide water absorptive properties and iron powder spacing properties. The filler is preferably 100 percent less than 200 mesh U.S.
Standard Sieve Size. A surfactant is also advanta-geous to provide water dispersive properties and wetting properties to the oxygen reactive composition of the invention. Suitable surfactants include sodium laurel sulfate, polysorbate and sodium dodecyl sulfate.
Preferably, the powdered iron is present in the composition at a level of from about 25 to about 35 percent, the filler is present at a level of from about 40 to about 60 percent, the acid is present at a level of from about 12 to about 21 percent, the carbonate is present at a level of from about 1 to about 4 Percent and the surfactant is present at a level of from about .05 to about .4 percent. In a most preferred composition, the powdered iron is present at a level of from about 28 to about 30 percent, the diatomaceous earth is present at a level of from about 50 to about 54 percent, the acid is present at a level of from about 15.5 to about 17.5 percent, the carbonate is present at a level of from about 1.7 to about 2.7 percent and the surfac-tant is present at a level of from about .15 to about .25 percent. All percentages used herein are by weight, unless otherwise indicated.
In general, the oxygen reactive composition of the present invention should be present in the container at a level of from about 4.5 to 6.0 grams, ~2~7~ p-816 _g_ preferably from about 4.7 to about 5.6 grams, for providing suitable oxygen reactive properties with a volume of air of from about 250 cc to 350 cc. This size of container is sufficiPnt to process one or two petri dishes. PreferabIy~ the container is a flexible, plastic, gas impermeable envelope having a width of from about 14 to about 18 cm and a length of from about 23 to about 30 cm. Of course, the en-velope can have any length longer than the preferred length since the excess length can be compensated for during the process of sealing the bag. The envelope has a compartment 29 formed therein by heat sealing a section 31 as shown in Figure 1.
The goal of the present invention is to attain less than 2 percent oxygen within the container and from about 4 to about 12 percent carbon dioxide within the container in a period of less than about 2 hours. To achieve this goal, it has been determined that the level o~ water added to the oxygen reactive composition of the invention is critical. To achieve the oxygen reducing goal of the invention, the water should be added to the composition at a level of from about 60 to about 80 percent of the weight of the oxygen reactive composition. Most preferably, the water should be added to the composi-tion at a level of from about 65 to abot 75 percent of the weight of the composition. In one embodiment ,~ of the invention, the oxygen reactive material in the sachet can be premoistened with a suitable level of water prior to sealing of the pacXage without inser-tion of the petri dish or dishes. The oxygen within ;;5 --1 o--the package is removed by reaction with the oxygen reactive material, which retains a residual ability to react with additional oxygen. The package can then subsequently be reopened for insertion of the petri dish and reaction with the oxygen introduced by the reopening of the package.
In a preferred embodiment of the invention, a suitable predetermined level of water is provided for use with the oxygen reactive composition of the invention as a separate vial, 15, as shown in Figure 4. The water containing vial 15 is made of heat sealed plastic having a tube 17 shaped therein. A
spout 19 is provided at the end of the tube. The spout can be opened by use of the tear strip 21 provided in the vial.
In use, as shown in Figure 1, the end of the vial is torn from the vial to expose the tip of the spout and the vial is inserted within the envelope 11 into compartment 29 in fluid communication with the sachet containing the oxygen reactive material. One petri dish miniteter panels or microtiter-type panels 23 are inserted in the container and the container is sealed.
A suitable means for sealing a flexible envelope is shown in Figures 1 and 2. As shown in Figure 2, the open end of the envelope 11 is wrapped around tubular rod 25. ~ semi-circular clamping rod 27 is clamped over the end of the envelope and the rod 25, as shown in Figure 2. The water is then dispensed from the vial by squeezing the vial while it is in place in the envelope. Thereafter, the package is ~2~S~
placed in a suitable incubation temperature for generation of an oxygen free atmosphere and growth of the anaerobic organism.
In a commercial embodiment of the present inven-tion, a plurality o~ envelopes having the sachet suitably positioned in the envelope would be packaged with a matching plurality of vials containing a predetermined level of water. The package would include at least one device for sealing the end of the envelope.
The following examples further illustrate various features of the present invention but are intended to in no way limit the scope of the inven-tion which is defined in the appended claims.
Example An oxygen reactive and carbon dioxide generating composition is prepared by blending together the Eollowing materials:
Oxygen Reactive Material Material % by Weight Keselguhr (diatomaceous éarth 51.6 from J. T. Baker, Catalog No. 2244) Powdered Iron (95~ less than 325 mesh 29.2 ., U.S.S. obtained from Glidden Co.
(A-131 SCM) Citric Acid (anhydrous, free 16.7 flowing powder) Sodium Carbonate (amorphous powder) 2.3 Sodium Lauryl Sulfate (crystals)0.2 ~7~ P-81 6 The above blend of materials was placed in a ball mill and milled for one hour to uniformly disperse the materials.
Sachets were prepared having 5.14 ~ 0.4 grams of the above oxygen Reactive Material. The sachets were prepared using the same porous water-resistant paper as are used for the preparation of tea bags.
The sachets were placed in position between two 16 cm x 27 cm sheets of heat sealable gas impermeable plastic, comprising a lamination of polyester, polyvinylidene chloride and polyethylene, with the polyester layer being the inside layer. The sachet was located near a corner of the sheets. The sheets were then heat sealed around three sides to form an envelope. The sachet was located in a position such that the bottom heat seal line passed over an edge of the sachet to secure the sachet in place. An addi-tional heat seal line was provided to form a compart-ment extending fro,m the top of the envelope and above the sachet.
The reaction between hydrogen and oxygen in the presence of a catalyst is strongly exothermic.
Flashing, and even explosion, can occur at the catalyst surface, particularly if the catalyst is finely divided and no means are provided to dissipate the heat generated.
United States Patent No. 4,013,422 to Spinner discloses a container having a material for gener-ating a reducing gas, such as hydrogen, for reaction, in the presence of a catalyst, with oxygen. The Spinner apparatus, however, uses an exposed catalyst pellet with no pro~ision for heat removal. In addition, it relies on the breaking of an ampoule containing a liquid and thereby instantaneous release of the liquid into contact with the material for generating the reducing gas. Thus no means are provided for control]ing the rate at which the contact is made. Experience has shown that contact at a slow and controlled rate is essential for accurate attainment of pre-determined final oxygen levels, particularly in those cases where it is desired to reduce, but not eliminate, the oxygen in the atmosphere.
U.S. Pat. No. 4,287,306 to Brewer describes a further apparatus for generating anaerobic atmo-spheres. In accordance with this patent, a flexible sealed package is provided with a catalyst coated onto the exterior surface of the package for use in 7~5~i P- 816 catalyzing the reaction between oxygen outside the package and hydrogen generated within the package.
This apparatus, like the Spinner apparatus, has the disadvantage of exposed active catalyst.
United States Patent No. 4,289,855 to Whitley discloses a safety catalyst package designed to reduce the danger of flashing or explosion. The Whitley package encloses a catalyst within holes and folds in a metal foil net. The net, which is inside of a container having holes for gas exchange, is composed of a heat conducting material, and thereby functions to remove heat from the catalyst vicinity. The Whitley disclosure is of a safety catalyst package only, and makes no provision for supply of hydrogen.
A package, for use in an anaerobic jar and specifically designed for attainment of a microaero-philic atmosphere, is disclosed in United States Patent 4,377,554 to Johnson. The Johnson invention relies on control of "wetover" and "condensation"
times for successful generation of microaerophilic atmospheres, and uses conventional exposed catalysts attached either to the outside of the package or to the lid of the jar.
A cardboard package for use in generating an anaerobic atmosphere is commercially available from the BBL Microbiology Systems Division of Becton DicXinson Company. This package includes a catalyst chamber mounted in the side of the box. The catalyst chamber is porous on both sides to permit generated hydrogen to flow through the chamber and thus react with the catalyst.
The prior art discussed above generally de-scribes apparatus for attaining an anaerobic atmo-sphere wherein hydrogen is generated and caused ~o react with the oxygen present in a vessel by means of a catalyst. Great Britain Patent No. 2109406 to Kasugai describes a process for culturing anaerobic bacteria wherein an agent which is reactive with oxygen is enclosed in an air tight closed vessel.
The agent contains a composition which is capable of removing oxygen and generating carbon dioxide in a volumetric ratio of about approximately twice as much oxygen removed as carbon dioxide generated. The preferred oxygen reactive material disclosed in the Kasugai patent is powdered iron.
Thus the invention provides in a broad aspect a package for removing oxygen from a gaseous mixture within a confined space comprising: (a) a sealable envelope, (b) a gas permeable sachet located within said envelope containing (l) an oxygen reactive material selected from the group consisting of iron powder, copper powder and aluminum powder, (2) a carbon dioxide generating composition including a water soluble, solid acid and a water soluble carbonate, (3) a filler selected from the group consisting of diato-maceous earth, charcoal, cellulose fiber, silica gel and barium sulfate and (4) a water dispersive and wetting surfactant, (c) a compartment within said envelope in fluid communication with said sachet location for receiving a predetermined level of water.
In another aspect the invention provides an article for use in removing oxygen from a gaseous mixture in a confined space comprising: a gas permeable sachet having the following components ~;~47~55 -4a-disposed therein: tl) an oxygen reactive material selected from the group consisting of iron powder, copper powder and aluminum powder, (2) a carbon dioxide generating composition including a water soluble, solid acid and a water soluble carbonate, (3) a filler selected from the group consisting of diatomaceous earth, charcoal, cellulose fiber, silica gel and barium sulfate;and (4) a waterdispersive andwetting surfactant.
Summary of the Invention In accordance with one embodiment of the present invention, there is provided a package for removing all or part of the oxygen in a gaseous atmosphere within the package. The package includes a gas impermeable, sealable container having a sachet disposed therein. The sachet contains an oxygen reactive material. A compartment is provided within the container which is in fluid communication with the sachet location. The compartment is adapted to receive a predetermined level of water.
In use, one or more petri dishes having a prepared media and havinq been inoculated with a sample suspected of containing an anaerobic micro-organism are inserted into the container adjacent to the sachet. A pipette or a vial is then used to ~Z~7~S5 P-81~
dispense a predetermined level of water into the compartment. The water hastens reaction between any oxygen contained within the envelope and the oxygen reactive material. The container is then sealed by suitable means. In a preferred embodiment of the invention the container is a flexible envelope and oxygen reactive material is powdered iron. The oxygen reacts with the powde~ed iron to form iron oxide. After a suitable period of time has elapsed the oxygen level contained within the envelope is, reduced to a level appropriate for the growth for anaerobic organisms, e.g. less than about 2 percent by volume.
The present invention provides an extremely simple, easily used, self contained device for generation of an anaerobic atmosphere without the hazards associated with the use of hydrogen and a catalyst.
Brief Description of the Drawings Figure 1 is an exploded view of the anaerobic generating apparatus of the invention showing the sequence of assembling the package;
Fiyure 2 is a front view of the assembled package;
Figure 3 is a side view of the assembled pack-age; and Figure 4 is a view of the water containing vial of the invention showing the arrangement of the tear strip used to seal the end of the vial.
Detailed Description of the Invention Referring now to Figure 1, there is shown a package for removing oxygen from a gaseous atmo-sphere. The package is in the form of an envelope 11. Envelope 11 is made of a suitable material which is impervious to the atmosphere and moisture. Thus, for example, the envelope 11 may be formed of a laminated plastic material, such as polyvinylidene chloride/polyester/polyethylene. The envelope 11 may be formed from two panels suitably secured together around the edges by heat sealing.
The interior of the envelope contains a sachet 13 which includes an oxygen reactive material. The sachet 13 is secured in place within the envelope 11 by any suitable means. One suitable means is to place one edge of the sachet between the two panels of plastic which are used to form the envelope prior to heat sealing and to heat seal the plastic over a portion of the sachet. Another suitable means would be to heat seal an area of the envelope adja-cent to the sachet so as to provide an open pocket for restraining the sachet in a suitable position~
The sachet is formed of a porous material, such as water resistant paper, to permit the water to pene-trate to the inside of the sachet and to permit reaction of the gaseous atmosphere within the en-velope with the contents of the sachet.
,~ As indicated, the sachet contains a material which is reactive with oxygen so as to remove the oxygen from the gaseous atmosphere within the en-velope. A suitable o~ygen reactive material is a ~2~55 P-816 metal powder provided in a red1ced state of oxygena-tion. Suitable metal powders include iron, copper, aluminum. A preferred metal powder is powdered iron. It has been determined tha~ the use of iron powder having a particle size 95 percent less than 325 mesh U.S. Standard Sieve Size is a most preferred oxygen reactive material for use in the invention.
For an embodiment of the invention wherein it is desired to provide an atmosphere having a predeter-mined level of carbon dioxide, a carbon dioxide generating composition is provided. The carbon dioxide generating composition includes a water soluble solid acid and a water soluble carbonate in amounts suitable for generating carbon dioxide~ As representative examples of suitable acids there may be mentioned: citric, tartaric, ascorbic, succinic, malic, fumaric, lactic acids and the like. AS
representative examples of suitable carbonates, there may be mentioned sodium bicarbonate, sodium car-bonate, potassium carbonate, sodium sesquicarbonate, etc. A preferred composition includes citric acid and sodium bicarbonate~ The carbon dioxide gener-ating composition is preferably employed in the form of a powder which is 100 percent less than 200 mesh U.S. Standard seive size and which is dispersed with the powdered iron.
It has also been determined that to provide a suitable low level of oxygen required for anaerobic conditions within an appropriate time, the presence of a water absorbing filler is important. Suitable fillers include diatomaceous earth, charcoal, cellu-lose fiber, silica gel and barium sulfate. In ~L24~ 5~
particular, diatomaceous earth has been found to be suitable to provide water absorptive properties and iron powder spacing properties. The filler is preferably 100 percent less than 200 mesh U.S.
Standard Sieve Size. A surfactant is also advanta-geous to provide water dispersive properties and wetting properties to the oxygen reactive composition of the invention. Suitable surfactants include sodium laurel sulfate, polysorbate and sodium dodecyl sulfate.
Preferably, the powdered iron is present in the composition at a level of from about 25 to about 35 percent, the filler is present at a level of from about 40 to about 60 percent, the acid is present at a level of from about 12 to about 21 percent, the carbonate is present at a level of from about 1 to about 4 Percent and the surfactant is present at a level of from about .05 to about .4 percent. In a most preferred composition, the powdered iron is present at a level of from about 28 to about 30 percent, the diatomaceous earth is present at a level of from about 50 to about 54 percent, the acid is present at a level of from about 15.5 to about 17.5 percent, the carbonate is present at a level of from about 1.7 to about 2.7 percent and the surfac-tant is present at a level of from about .15 to about .25 percent. All percentages used herein are by weight, unless otherwise indicated.
In general, the oxygen reactive composition of the present invention should be present in the container at a level of from about 4.5 to 6.0 grams, ~2~7~ p-816 _g_ preferably from about 4.7 to about 5.6 grams, for providing suitable oxygen reactive properties with a volume of air of from about 250 cc to 350 cc. This size of container is sufficiPnt to process one or two petri dishes. PreferabIy~ the container is a flexible, plastic, gas impermeable envelope having a width of from about 14 to about 18 cm and a length of from about 23 to about 30 cm. Of course, the en-velope can have any length longer than the preferred length since the excess length can be compensated for during the process of sealing the bag. The envelope has a compartment 29 formed therein by heat sealing a section 31 as shown in Figure 1.
The goal of the present invention is to attain less than 2 percent oxygen within the container and from about 4 to about 12 percent carbon dioxide within the container in a period of less than about 2 hours. To achieve this goal, it has been determined that the level o~ water added to the oxygen reactive composition of the invention is critical. To achieve the oxygen reducing goal of the invention, the water should be added to the composition at a level of from about 60 to about 80 percent of the weight of the oxygen reactive composition. Most preferably, the water should be added to the composi-tion at a level of from about 65 to abot 75 percent of the weight of the composition. In one embodiment ,~ of the invention, the oxygen reactive material in the sachet can be premoistened with a suitable level of water prior to sealing of the pacXage without inser-tion of the petri dish or dishes. The oxygen within ;;5 --1 o--the package is removed by reaction with the oxygen reactive material, which retains a residual ability to react with additional oxygen. The package can then subsequently be reopened for insertion of the petri dish and reaction with the oxygen introduced by the reopening of the package.
In a preferred embodiment of the invention, a suitable predetermined level of water is provided for use with the oxygen reactive composition of the invention as a separate vial, 15, as shown in Figure 4. The water containing vial 15 is made of heat sealed plastic having a tube 17 shaped therein. A
spout 19 is provided at the end of the tube. The spout can be opened by use of the tear strip 21 provided in the vial.
In use, as shown in Figure 1, the end of the vial is torn from the vial to expose the tip of the spout and the vial is inserted within the envelope 11 into compartment 29 in fluid communication with the sachet containing the oxygen reactive material. One petri dish miniteter panels or microtiter-type panels 23 are inserted in the container and the container is sealed.
A suitable means for sealing a flexible envelope is shown in Figures 1 and 2. As shown in Figure 2, the open end of the envelope 11 is wrapped around tubular rod 25. ~ semi-circular clamping rod 27 is clamped over the end of the envelope and the rod 25, as shown in Figure 2. The water is then dispensed from the vial by squeezing the vial while it is in place in the envelope. Thereafter, the package is ~2~S~
placed in a suitable incubation temperature for generation of an oxygen free atmosphere and growth of the anaerobic organism.
In a commercial embodiment of the present inven-tion, a plurality o~ envelopes having the sachet suitably positioned in the envelope would be packaged with a matching plurality of vials containing a predetermined level of water. The package would include at least one device for sealing the end of the envelope.
The following examples further illustrate various features of the present invention but are intended to in no way limit the scope of the inven-tion which is defined in the appended claims.
Example An oxygen reactive and carbon dioxide generating composition is prepared by blending together the Eollowing materials:
Oxygen Reactive Material Material % by Weight Keselguhr (diatomaceous éarth 51.6 from J. T. Baker, Catalog No. 2244) Powdered Iron (95~ less than 325 mesh 29.2 ., U.S.S. obtained from Glidden Co.
(A-131 SCM) Citric Acid (anhydrous, free 16.7 flowing powder) Sodium Carbonate (amorphous powder) 2.3 Sodium Lauryl Sulfate (crystals)0.2 ~7~ P-81 6 The above blend of materials was placed in a ball mill and milled for one hour to uniformly disperse the materials.
Sachets were prepared having 5.14 ~ 0.4 grams of the above oxygen Reactive Material. The sachets were prepared using the same porous water-resistant paper as are used for the preparation of tea bags.
The sachets were placed in position between two 16 cm x 27 cm sheets of heat sealable gas impermeable plastic, comprising a lamination of polyester, polyvinylidene chloride and polyethylene, with the polyester layer being the inside layer. The sachet was located near a corner of the sheets. The sheets were then heat sealed around three sides to form an envelope. The sachet was located in a position such that the bottom heat seal line passed over an edge of the sachet to secure the sachet in place. An addi-tional heat seal line was provided to form a compart-ment extending fro,m the top of the envelope and above the sachet.
3.5 grams of water were pipetted into the compartment above the sachet and the envelope was sealed. The oxygen level in the envelope after a period of two hours was found to be 1.18 percent by volume.
Claims (13)
1. A package for removing oxygen from a gaseous mixture within a confined space comprising:
(a) a sealable envelope (b) a gas permeable sachet located within said envelope containing (1) an oxygen reactive material selected from the group consisting of iron powder, copper powder and aluminum powder, (2) a carbon dioxide generating composition including a water soluble, solid acid and a water soluble carbonate, (3) a filler selected from the group consisting of diatomaceous earth, charcoal, cellulose fiber, silica gel and barium sulfate and (4) a water dispersive and wetting surfactant, (c) a compartment within said envelope in fluid communication with said sachet location for receiving a predetermined level of water.
(a) a sealable envelope (b) a gas permeable sachet located within said envelope containing (1) an oxygen reactive material selected from the group consisting of iron powder, copper powder and aluminum powder, (2) a carbon dioxide generating composition including a water soluble, solid acid and a water soluble carbonate, (3) a filler selected from the group consisting of diatomaceous earth, charcoal, cellulose fiber, silica gel and barium sulfate and (4) a water dispersive and wetting surfactant, (c) a compartment within said envelope in fluid communication with said sachet location for receiving a predetermined level of water.
2. A package in accordance with claim 1 wherein said oxygen reactive material is iron powder having a particle size 95% less than 325 mesh U.S. Standard Sieve Size.
3. A package in accordance with claim 1 wherein said water soluble carbonate is selected from the group consisting of sodium bicarbonate, sodium carbonate, potassium carbonate, sodium sesquicarbonate and said water soluble acid is selected from the group consisting of citric acid, tartaric acid, ascorbic acid, succinic acid, malic acid, fumaric acid and lactic acid.
4. A package in accordance with claim 1 wherein said filler is diatomaceous earth having a particle size of 100% less than 200 mesh U.S. Standard Sieve Size.
5. A package in accordance with claim 1 wherein said sachet is fixed to said envelope.
6. A package in accordance with claim 1 which further comprises a separate vial containing a pre-determined level of water.
7. A package in accordance with claim 1 wherein said oxygen reactive material comprises said powdered iron at a level of from about 25 to about 35%, said filler at a level of from about 40 to about 60%, said water soluble acid at a level of from about 12 to about 20%, said water soluble carbonate at a level of from about 1 to about 4% and said surfactant at a level of from about 0.05 to about 0.4%.
8. A package in accordance with claim 7 wherein said package further includes a vial containing water at a level of from about 60 to about 80% by weight of water based on the weight of said oxygen reactive material.
9. An article for use in removing oxygen from a gaseous mixture in a confined space comprising:
a gas permeable sachet having the following compo-nents disposed therein:
(1) an oxygen reactive material selected from the group consisting of iron powder, copper powder and aluminum powder, (2) a carbon dioxide generating composition including a water soluble, solid acid and a water soluble carbonate, (3) a filler selected from the group consisting of diatomaceous earth, charcoal, cellulose fiber, silica gel and barium sulfate; and (4) a water dispersive and wetting surfactant.
a gas permeable sachet having the following compo-nents disposed therein:
(1) an oxygen reactive material selected from the group consisting of iron powder, copper powder and aluminum powder, (2) a carbon dioxide generating composition including a water soluble, solid acid and a water soluble carbonate, (3) a filler selected from the group consisting of diatomaceous earth, charcoal, cellulose fiber, silica gel and barium sulfate; and (4) a water dispersive and wetting surfactant.
10. An article in accordance with claim 9 wherein said oxygen reactive material is iron powder having a particle size 95% less than 325 mesh U.S. Standard Sieve Size.
11. An article in accordance with claim 9 wherein said water soluble carbonate is selected from the group consisting of sodium bicarbonate, sodium carbonate, potassium carbonate, sodium sesquicarbonate and said water soluble acid is selected from the group con-sisting of citric acid, tartaric acid, ascorbic acid, succinic acid, malic acid, fumaric acid and lactic acid.
12. An article in accordance with claim 9 wherein said filler is diatomaceous earth having a particle size of 100% less than 200 mesh U.S. Standard Sieve Size.
13. An article in accordance with claim 9 wherein said oxygen reactive material comprises said powdered iron at a level of from about 25 to about 35%, a filler at said level of from about 40 to about 60%, said water soluble acid at a level of from about 12 to about 20%, said water soluble carbonate at a level of from about 1 to about 4% and said surfactant at a level of from about 0.05 to about 0.4%.
Applications Claiming Priority (2)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| US628,670 | 1984-07-06 | ||
| US06/628,670 US4588561A (en) | 1984-07-06 | 1984-07-06 | Package for removing oxygen from a gaseous mixture |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CA1247055A true CA1247055A (en) | 1988-12-20 |
Family
ID=24519838
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CA000478712A Expired CA1247055A (en) | 1984-07-06 | 1985-04-10 | Generation of anaerobic or microaerophilic atmosphere |
Country Status (9)
| Country | Link |
|---|---|
| US (1) | US4588561A (en) |
| EP (1) | EP0167229B1 (en) |
| JP (1) | JPS6121083A (en) |
| AU (1) | AU572287B2 (en) |
| CA (1) | CA1247055A (en) |
| DE (1) | DE3579760D1 (en) |
| DK (1) | DK170407B1 (en) |
| FI (1) | FI85875C (en) |
| MY (1) | MY102069A (en) |
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| DE3804488A1 (en) * | 1987-02-14 | 1988-08-25 | Mitsubishi Gas Chemical Co | OXYGEN ABSORPTION AGENT |
| EP0290781A3 (en) * | 1987-05-01 | 1989-02-08 | Becton, Dickinson and Company | Method for modifying a contained gaseous mixture |
| US5284871A (en) * | 1987-09-25 | 1994-02-08 | The Pillsbury Company | Oxygen removal |
| US5270337A (en) * | 1987-09-25 | 1993-12-14 | The Pillsbury Company | Oxygen removal |
| JP2503560B2 (en) * | 1987-12-29 | 1996-06-05 | 凸版印刷株式会社 | Oxygen absorber |
| JP2701999B2 (en) * | 1990-05-16 | 1998-01-21 | 株式会社上野製薬応用研究所 | Oxygen absorber |
| US5236617A (en) * | 1990-05-16 | 1993-08-17 | K.K. Ueno Seiyaku Oyo Kenkyujc | Oxygen absorbent |
| US5667863A (en) * | 1991-01-07 | 1997-09-16 | Multisorb Technologies, Inc. | Oxygen-absorbing label |
| ATE157932T1 (en) * | 1991-01-07 | 1997-09-15 | Multisorb Tech Inc | OXYGEN ABSORBING LABEL |
| US5686161A (en) * | 1991-01-07 | 1997-11-11 | Multisorb Technologies, Inc. | Moisture-sensitive label |
| JPH056867U (en) * | 1991-07-10 | 1993-01-29 | 株式会社三協精機製作所 | Printed circuit board |
| US6209289B1 (en) | 1992-01-30 | 2001-04-03 | Multisorb Technologies, Inc. | Composition for and method of absorbing oxygen in an oxygen/carbon dioxide environment |
| US6369148B2 (en) * | 1993-07-16 | 2002-04-09 | Ciba Specialty Chemicals Corporation | Oxygen-scavenging compositions and articles |
| US5391350A (en) * | 1994-03-09 | 1995-02-21 | Nalge Company | Apparatus and method for sterilizing containers in an autoclave |
| US5698250A (en) | 1996-04-03 | 1997-12-16 | Tenneco Packaging Inc. | Modifield atmosphere package for cut of raw meat |
| US5830545A (en) * | 1996-04-29 | 1998-11-03 | Tetra Laval Holdings & Finance, S.A. | Multilayer, high barrier laminate |
| US5928560A (en) | 1996-08-08 | 1999-07-27 | Tenneco Packaging Inc. | Oxygen scavenger accelerator |
| US6395195B1 (en) | 1996-08-08 | 2002-05-28 | Pactiv Corporation | Oxygen scavenger accelerator |
| EP0869172B1 (en) * | 1997-04-01 | 2003-10-15 | Mitsubishi Gas Chemical Company, Inc. | Atmosphere regulator and method for culturing anaerobic bacteria |
| US6054153A (en) * | 1998-04-03 | 2000-04-25 | Tenneco Packaging Inc. | Modified atmosphere package with accelerated reduction of oxygen level in meat compartment |
| US6231905B1 (en) | 1998-10-08 | 2001-05-15 | Delduca Gary R. | System and method of making a modified atmosphere package comprising an activated oxygen scavenger for packaging meat |
| WO2000028839A1 (en) * | 1998-11-13 | 2000-05-25 | Pactiv Corporation | Oxygen scavenger accelerator |
| US6321509B1 (en) | 1999-06-11 | 2001-11-27 | Pactiv Corporation | Method and apparatus for inserting an oxygen scavenger into a modified atmosphere package |
| WO2001025369A1 (en) * | 1999-10-01 | 2001-04-12 | Pactiv Corporation | Rapid oxygen absorption by using activators |
| CN1545386A (en) * | 2001-03-02 | 2004-11-10 | 宝洁公司 | Devices and methods for prolonging the storage life of produce |
| US20030054074A1 (en) * | 2001-07-25 | 2003-03-20 | Delduca Gary R. | Modified atomsphere packages and methods for making the same |
| US20030054073A1 (en) * | 2001-07-25 | 2003-03-20 | Delduca Gary R. | Modified atmosphere packages and methods for making the same |
| US7125498B2 (en) * | 2004-02-04 | 2006-10-24 | Multisorb Technologies, Inc. | Oxygen-absorbing compositions and method |
| GB0619060D0 (en) * | 2006-09-27 | 2006-11-08 | Glaxosmithkline Biolog Sa | Novel process |
| US8048201B2 (en) * | 2009-03-26 | 2011-11-01 | Desiccare, Inc | Oxygen scavenger and a method for scavenging oxygen |
| CA2832705A1 (en) * | 2011-04-18 | 2012-10-26 | Bret T. BARNHIZER | Improved flat packaging of petri dishes for prolonged preservation and method of producing the same |
| US9078939B2 (en) * | 2012-01-23 | 2015-07-14 | Star-Brite Distributing, Inc. | Disinfectant system |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| US2908555A (en) * | 1955-07-15 | 1959-10-13 | Drager Otto H | Gas detecting device |
| US2825651A (en) * | 1957-07-01 | 1958-03-04 | Carnation Co | In-package oxygen remover |
| US3246959A (en) * | 1963-03-21 | 1966-04-19 | John H Brewer | Apparatus for generating gas |
| US4013422A (en) * | 1975-12-22 | 1977-03-22 | Marion Laboratories, Inc. | Gas generating apparatus |
| US4038148A (en) * | 1975-12-22 | 1977-07-26 | Marion Laboratories, Inc. | Anaerobic environmental system for bacteria culture testing |
| GB1544995A (en) * | 1977-01-11 | 1979-04-25 | Oxoid Ltd | Gas-producing device |
| DE2742874A1 (en) * | 1977-09-23 | 1979-04-05 | Merck Patent Gmbh | OXYGEN ABSORBING AGENTS AND METHODS FOR CREATING A LOW-OXYGEN ATMOSPHERE |
| US4230595A (en) * | 1978-03-13 | 1980-10-28 | Teijin Limited | Oxygen scavenging and heat-generating compositions, and deoxygenating and heat-generating structures |
| US4299719A (en) * | 1978-10-23 | 1981-11-10 | Mitsubishi Chemical Ind., Ltd. | Deoxidizer |
| US4287306A (en) * | 1979-04-02 | 1981-09-01 | Becton, Dickinson And Company | Apparatus for generation of anaerobic atmosphere |
| JPS5642903A (en) * | 1979-09-14 | 1981-04-21 | Matsushita Electric Works Ltd | Lighting device for low temperature |
| JPS6038330B2 (en) * | 1980-05-20 | 1985-08-31 | 松下電器産業株式会社 | Carbon dioxide supply device |
| JPS5739776A (en) * | 1980-08-20 | 1982-03-05 | Mitsubishi Gas Chem Co Inc | Cultivation of anaerobic bacteria |
| DE3071273D1 (en) * | 1980-08-21 | 1986-01-16 | Daishiro Fujishima | Disoxidant and process for preparing same |
| JPS5816673A (en) * | 1981-07-21 | 1983-01-31 | Showa Yakuhin Kako Kk | Device for ready cultivation of anaerobic bacteria |
| US4377554A (en) * | 1981-08-26 | 1983-03-22 | Becton, Dickinson And Company | Generation of microaerophilic atmosphere |
| JPS5856679A (en) * | 1981-09-30 | 1983-04-04 | Meito Kk | Method for culturing anaerobic bacteria and agent for controlling cultivation atmosphere |
| IE54140B1 (en) * | 1982-03-19 | 1989-06-21 | Johnson Matthey Plc | Deoxygenation process |
| US4562051A (en) * | 1984-02-27 | 1985-12-31 | Becton, Dickinson And Company | Generation of anaerobic or microaerophilic atmosphere |
-
1984
- 1984-07-06 US US06/628,670 patent/US4588561A/en not_active Expired - Lifetime
-
1985
- 1985-04-10 CA CA000478712A patent/CA1247055A/en not_active Expired
- 1985-04-17 AU AU41372/85A patent/AU572287B2/en not_active Ceased
- 1985-04-19 EP EP85302775A patent/EP0167229B1/en not_active Expired
- 1985-04-19 DE DE8585302775T patent/DE3579760D1/en not_active Expired - Fee Related
- 1985-05-10 FI FI851862A patent/FI85875C/en not_active IP Right Cessation
- 1985-06-04 DK DK252785A patent/DK170407B1/en not_active IP Right Cessation
- 1985-06-07 JP JP60124066A patent/JPS6121083A/en active Granted
-
1987
- 1987-09-29 MY MYPI87002175A patent/MY102069A/en unknown
Also Published As
| Publication number | Publication date |
|---|---|
| JPS6121083A (en) | 1986-01-29 |
| FI851862L (en) | 1986-01-07 |
| FI851862A0 (en) | 1985-05-10 |
| EP0167229A3 (en) | 1987-09-16 |
| DK170407B1 (en) | 1995-08-21 |
| FI85875B (en) | 1992-02-28 |
| EP0167229A2 (en) | 1986-01-08 |
| DK252785D0 (en) | 1985-06-04 |
| EP0167229B1 (en) | 1990-09-19 |
| AU572287B2 (en) | 1988-05-05 |
| FI85875C (en) | 1992-06-10 |
| MY102069A (en) | 1992-03-31 |
| AU4137285A (en) | 1986-01-09 |
| US4588561A (en) | 1986-05-13 |
| JPS6260063B2 (en) | 1987-12-14 |
| DE3579760D1 (en) | 1990-10-25 |
| DK252785A (en) | 1986-01-07 |
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