优择Designation:F1608−16
Standard Test Method for
Microbial Ranking of Porous Packaging Materials (Exposure Chamber Method)1
This standard is issued under the fixed designation F1608;the number immediately following the designation indicates the year of original adoption or,in the ca of revision,the year of last revision.A number in parenthes indicates the year of last reapproval.A superscript epsilon (´)indicates an editorial change since the last revision or reapproval.
1.Scope
1.1This test method is ud to determine the passage of airborne bacteria through porous materials intended for u in packaging sterile medical devices.This test method is designed to test materials under conditions that result in the detectable passage of bacterial spores through the test material.
1.1.1A round-robin study was conducted with eleven labo-ratories participating.Each laboratory tested duplicate samples of six commercially available porous materials to determine the Log Reduction Value (LRV)(e calculation in Section 12).Materials tested under the standard conditions described in this test method returned average values that range from LRV 1.7to 4.3.
1.1.2Results of this round-robin study indicate that caution should be ud when comparing test data and ranking materials,especially when a small number of sample replicates are ud.In addition,further collaborative work (such as described in Practice E691)should be conducted before this test method would be considered adequate for purpos of tting performance standards.
1.2This test method requires manipulation of microorgan-isms and should be performed only by trained personnel.The U.S.Department of Health and Human Services publication Biosafety in Microbiological and Biomedical Laboratories (CDC/NIH-HHS Publication No.84-8395)should be con-sulted for guidance.
1.3The values stated in SI units are to be regarded as standard.No other units of measurement are included in this standard.
1.4This standard does not purport to address all of the safety concerns,if any,associated with its u.It is the responsibility of the ur of this standard to establish appro-priate safety and health practices and determine the applica-bility of regulatory limitations prior to u.
yah2.Referenced Documents
2.1ASTM Standards:2
E691Practice for Conducting an Interlaboratory Study to Determine the Precision of a Test Method 3.Terminology
3.1Definitions:
3.1.1porous packaging material,n—a material ud in medical packaging which is intended to provide an environ-mental and biological barrier,while allowing sufficient air flow to be ud in gaous sterilization methods (for example,ethylene oxide,steam,gas plasma).
4.Summary of Test Method
4.1Samples of porous materials are subjected to an aerosol of Bacillus atrophaeus spores within an exposure chamber.Spores which pass through the porous sample are collected on membrane filters and enumerated.The LRV is calculated by comparing the logarithm of the number of spores passing through the porous material with the logarithm of the microbial challenge.
4.2Standard Set of Conditions—This test method specifies a standard t of conditions for conducting the exposure chamber test method.A standard t of conditions is required to enable evaluation of materials between laboratories.The con-ditions stated in this test method were chon for veral reasons.First,it is difficult to maintain an aerosol of spores over long periods of time.(Also,if the spore challenge time is long,the cost of the test increas).Second,to determine the differences between materials,it is necessary to test the materials under conditions which allow passage of bacterial spores.If a material does not allow any passage of spores,all that can be stated is that it has better resistance to penetration than the verity of the challenge conditions.Third,it is necessary to have a large spore challenge level to be able to detect the passage of spores through the entire range of
1
This test method is under the jurisdiction of ASTM Committee F02on Flexible Barrier Packaging and is the direct responsibility of Subcommittee F02.15on Chemical/Safety Properties.
Current edition approved May 1,2016.Published June 2016.Originally approved in 1995.Last previous edition approved in 2009as F1608–00(2009).DOI:10.1520/F1608-16.
2
For referenced ASTM standards,visit the ASTM website,www.astm,or contact ASTM Customer Service at rvice@astm.For Annual Book of ASTM Standards volume information,refer to the standard’s Document Summary page on the ASTM website.
softskillsCopyright ©ASTM International,100Barr Harbor Drive,PO Box C700,West Conshohocken,PA 19428-2959.United States
This international standard was developed in accordance with internationally recognized principles on standardization established in the Decision on Principles for the
Development of International Standards,Guides and Recommendations issued by the World Trade Organization Technical Barriers to Trade (TBT)Committee.
commercially available porous packaging materials.The stan-dard conditions stated in this test method are bad upon the factors.(Additional information may be found in the Refer-ences ction).However,since many factors influence the determination of an appropriate porous material(outlined in 5.1.1–5.1.4),each ur may modify the conditions(that is, bacterial challenge,time,flow rate)afterfirst conducting studies at the specified standard conditions.The standard t of target parameters for conducting the test method are as follows: 4.2.1Flow Rate Through Sample—2.8L/min.
4.2.2Exposure Time—15min.
4.2.3Target Microbial Challenge—1×106colony forming units(CFU)/sample port.
5.Significance and U
5.1The exposure-chamber method is a quantitative proce-dure for determining the microbial-barrier properties of porous materials under the conditions specified by the test.Data obtained from this test is uful in asssing the relative potential of a particular porous material in contributing to the loss of sterility to the contents of the package versus another porous material.This test method is not intended to predict the performance of a given material in a specific sterile-packaging application.Th
e maintenance of sterility in a particular pack-aging application will depend on a number of factors, including,but not limited to the following:
5.1.1The bacterial challenge(number and kinds of micro-organisms)that the package will encounter in its distribution and u.This may be influenced by factors such as shipping methods,expected shelf life,geographic location,and storage conditions.
5.1.2The package design,including factors such as adhe-sion between materials,the prence or abnce of condary and tertiary packaging,and the nature of the device within the package.
5.1.3The rate and volume exchange of air that the porous package encounters during its distribution and shelf life.This can be influenced by factors including the free-air volume within the package and pressure changes occurring as a result of transportation,manipulation,weather,or mechanical influ-ences(such as room door closures and HV AC systems).
5.1.4The microstructure of a porous material which influ-ences the relative ability to adsorb or entrap microorganisms, or both,under different air-flow conditions.
6.Apparatus
6.1This procedure should be conducted in a microbiologi-cal laboratory by trained personnel.As a result,it is assumed that basic microbiological equipment and supplies for conduct-ing routine microbiological manipulations(that is,standard plate counts,sterilization with an autoclave,and so forth)will be available.
6.2Exposure Chamber,constructed primarily from acrylic sheeting and consists of two major ctions,as illustrated in Fig.1.The bottom ction contains a six-place
manifold FIG.1Example of an Exposure
Chamber
connected to sixflowmeters,one per port,containing hos attached to sixfiltering units.The port to the manifold is attached to a vacuum source.A vacuum gauge is mounted between the manifold and the vacuum source.The upper chamber contains a fan for dispersion of the bacterial aerosol, a port for attachment of the nebulizer,a port for exhausting the chamber,and a plate for attachment of disposable or steriliz-ablefilter units.The chamber may u disposablefilter units or reusablefilter units,or both.
7.Materials
7.1Bacillus atrophaeus(ATCC9372),aqueous spore sus-pension in water.
7.2Soybean Cain Digest Agar/Tryptic Soy Agar—Bottles for pour plates and pre-poured plates(;25mL in100by 15-mm plates)prepared commercially or in accordance with standard techniques.
7.3Sterile Cellulo Nitrate Filters,47or50-mm diameter, depending uponfilter unit specification,0.45-µm pore size.
7.4Sterile Bottle-Top Filter Units,(Falcon-type7104or filter holders with funnel310-4000or equivalent).
7.5Glass Nebulizer.
7.6Sterile Forceps.
7.7Incubator,30to35°C.
7.8Disk Cutter,47or50-mm diameter,depending upon filter unit specification.
7.9Sterile Gloves.
7.10Sterile Syringe,3-cm3with needle or micropipette. 7.11Sterile Pipettes,to deliver0.1,1,10,and25mL. 7.12Blender,with sterile1⁄2-pt jar(s).
造化弄人英文7.13Vortex Mixer.
考个会计证要多少钱7.14Vacuum Pump,with airfilter.
7.15NIST Traceable Calibrated Timer.trashbin
7.16NIST Traceable Calibrated Flowmeters—One pressure flowmeter with a range from5to30L/min;six vacuum flowmeters each with a range from1.0to5.0L/min.
7.17Sterile Petri Plates.
7.18Sterile Water,100and9.9-mL aliquots,or other appro-priate volumes for membrane grinding and dilutions.观鸟大年
7.19Hos and Piping—See Section9for lengths and diameters.
7.20Rubber Stoppers with Holes—See Section9for sizes.
7.21Trap Jar.
7.22NIST Traceable Calibrated Vacuum Gauge.
7.23Compresd Air Source,with airfilter.
7.24Biocontainment Hood.
7.25Chlorine Bleach,or suitable sporocide.8.Sample Preparation
8.1Cut random samples of material into disks in accordance with the size required for thefilter holder being ud(47or50 mm)using a disk cutter.It is suggested that additional samples be cut to allow for errors during the procedure.Typically,the sample disks are sterilized prior to testing using a test method appropriate for the specific material.Materials may also be tested before or after they are subjected to other conditions such as heat or cold,relative humidity,different sterilization process,real time,or accelerated aging.The samples may be stored in sterile petri plates or other suitable sterile containers before testing.
8.2The minimum sample size for a given material is two, which was ud in the round-robin study of this test method. However,it is strongly suggested that more samples be ud to improve precision and bias(Section14).
9.Apparatus Preparation
9.1Since aerosols containing bacterial spores are formed during the u of this apparatus,the exposure chamber(e Fig.
1)should be asmbled and ud within a biological safety cabinet.
9.1.1Place the top of the chamber on the bottom ba. 9.1.2Connect the top of each of the sixflowmeters to the manifold using0.65-cm inside diameter hos.Connect the manifold to afiltered vacuum source.
9.1.3Connect the bottom of each sampleflowmeter to a filter unit with0.65-cm inside diameter ho using an end connector.
9.1.4Using a rubber ho,attach the nebulizer to a tee connector made of0.65-cm PVC and three pieces of0.6-cm inside diameter PVC piping approximately7.5cm long.
9.1.5Attach the vertical leg of the tee to a trap jar using a rubber stopper with a0.65-cm diameter hole.The trap jar is intended to retain any unsuspended droplets produced by the nebulizer.
9.1.6Attach the cond end of the tee to a1.3-cm inside diameter rubber tubing approximately3.8cm long and connect to the front port of the chamber.
9.1.7Attach a1.3-cm inside diameter rubber tubing ap-proximately16cm long to the mouth of the nebulizer.Connect the loo end of the tubing to the third end of the tee.
9.1.8Connect the nebulizer inlet port with a0.5-cm inside diameter rubber tubing to the top port of a calibratedflowmeter (from5to30-L/min range).
9.1.9Connect the bottom port of theflowmeter to afiltered air source.
9.1.10Attach the exhaust port of the chamber that is ud for evacuation to a1.3-cm inside diameter tubing which,in turn,leads to an airfilter and to a vacuum source.
9.2Filter Unit-Holder Preparation:
9.2.1Wrap the non-sterile sterilizablefilter unit in a steril-izable wrap.
9.2.2Sterilize thefilter units as specified by the manufac-turer.Presterilizedfilter units do not need to be
resterilized.
中俄翻译
10.Apparatus Validation
10.1The test apparatus (e Fig.1)must be validated for bacterial challenge to each port.This step should be performed upon first u of the chamber and a minimum of three runs should be conducted.The following description outlines the validation of the test procedure for a challenge of 1×106colony forming units (CFU)per port in 15min at a flow rate of 2.8L/min.If testing is to be conducted using other parameters,a validation should be conducted using tho parameters.10.1.1Place the sterile filtering apparatus in a biological safety cabinet.
10.1.2Aptically prepare six filter units by placing a sterile 0.45-µm membrane filter on the ba of each filter unit using sterile forceps and gloves (Fig.2B).
10.1.3Attach the top of each filter unit to the bottom of the exposure chamber.Then attach each filter
unit to its respective flowmeter.
10.1.4Dispen 3.0mL of the spore suspension into the nebulizer.When using the DeVilbiss #40nebulizer,a volume of 3.0mL at a concentration of 5×107spores/mL is necessary to achieve a challenge of 1×106CFU (60.5log)per port in 15min.
10.1.5Turn on the chamber fan.
10.1.6Adjust port flowmeters to 2.8L/min.It is important that all ports be t to the same flow and monitored during the exposure period.Before adjusting each flowmeter,open each
valve completely,then slowly open the vacuum and fine adjust until the desired flow is achieved.
continue的意思
10.1.7Adjust the nebulizer flow rate as recommended by the nebulizer manufacturer to produce droplets that are within the appropriate particle size range.When using the DeVilbiss #40nebulizer,a flow rate of 8.5L/min is ud.
10.1.8Immediately start the 15-min timer.At regular intervals,obrve and adjust (if necessary)all flowmeters to maintain the appropriate flow rate ttings during the 15-min test period.
10.1.9After exposure,turn off the vacuum,the fan,and the air flow to the nebulizer.Open the filtered exhaust port at the back of the chamber.
10.1.10Evacuate the chamber for 15min by connecting the vacuum source to the front of the chamber through a microbial filter asmbly.
10.1.11Disconnect the hos from each of the filter units and remove the units from the bottom plate of the exposure chamber.
10.1.12Disinfect the outside of each filtering unit with a suitable sporicide being careful not to compromi the test material.
10.1.13Remove the filter membranes aptically,one at a time,and enumerate the organisms on each membrane (Fig.3).Since more than 100CFU are anticipated,the spores must be eluted from the membrane by grinding the membrane for 1min in a suitable blender that has been validated containing 100.0mL of sterile water.Samples are then rially diluted prior to performing standard plate counts to accurately determine the number of spores.A dilution and plating scheme,which
was
FIG.2Sample and Control Material
Setup
FIG.3Example of Possible Dilution
Scheme
ud in the round-robin study,includes plating10.0,1.0,and 0.1-mL aliquots of the blended membrane in duplicate.An additional1to100dilution is prepared by placing0.1mL in 9.9mL of sterile water and plating1.0and0.1-mL aliquots of this dilution in duplicate.This scheme produces dilution factors of10−1,10−2,10−3,10−4,and10−5.Other validated extraction and enumeration protocols may be ud.Plates having between25and250CFU should be ud for enumera-tion.If alternative test conditions are ud,then the previously described dilution scheme may not be appropriate.In instances where colony counts are less than30CFU,the limit of detection is dependent upon the volume of the undiluted aliquot plated from the blender jar.Duplicate10-mL samples will result in a limit of detection of5CFU/membrane.If a lower limit is desired,plate an appropriately larger volume. However,there is incread statistical variation with the low numbers.If the membrane grinding and plating procedure consistently results in counts less than25CFU from all dilutions,enumeration c
an be accomplished by placing the membrane directly onto the surface of a SCDA plate with the challenge side up.
10.1.14Enumeration cultures are incubated for a minimum of24h at30to35°C.If incubated longer than24h,care should be taken to ensure that individual colonies remain discrete and overgrowth does not occur and that the growth media does not dry out.
10.1.15After incubation,count and record the number of CFUs and dilution factor for eachfilter.
10.1.16A minimum distribution of1×106(60.5log) spores is recommended.To increa the challenge per port, increa the concentration of the aqueous spore suspension rather than the volume.
10.1.17All ports must receive the same bacterial challenge (60.5log)for successful validation.
10.2Revalidation and Validation of Alternative Test Parameters—After validation has been performed using the standard test parameters as described in10.1–10.2,an appropriate number of validation runs should be performed when changes are made which have potential effects on bacterial distribution.Environmental conditions,equipment modifications,and changes in test parameters may be neces-sary considerations.If it is desired to perform testing using different test parameters,the test apparatus should be revali-dated using tho conditions.
11.Microbial Procedures for Testing Samples
11.1Place the sterilefiltering apparatus in a biological safety cabinet.
11.2Aptically place a sterile0.45-µm membranefilter on the ba of each unit,using sterile forceps and gloves(Fig. 2A).
11.3Place an appropriate diameter disk of test material on top of the membrane(Fig.2A).
11.3.1If the materials to be tested are known to produce high LRV values,it may be advisable to include a sample of a known material which allows substantial passage of spores in each run.A known material will also provide a frame of reference.In this ca,refer to the sample as the positive control.
11.4Prepare one of the six units with the challenge control membrane(N0)on top of the sterile0.45-µm membranefilter (Fig.2B).
11.4.1This bottom0.45-µm membranefilter rves as the negative control.
11.5Attach the top of eachfilter unit to the bottom of the exposure chamber.Then attach eachfilter unit to its respective flow meter.
11.6Dispen3.0mL of spore suspension into the nebu-lizer.U the concentration of spore suspension required to achieve the desired challenge level as determined during the apparatus validation.The suspension should be thoroughly mixed prior to u.
11.7Turn on the chamber fan.
11.8Adjust portflowmeters to2.8L/min.It is important that all ports be t to the sameflow and monitored during the exposure period.Before adjusting eachflowmeter,open each valve completely,then slowly open the vacuum andfine adjust until the desiredflow is achieved.
N OTE1—For comparing different materials,the standard test param-eters are aflow rate of2.8L/min to provide a minimum challenge of 1.0×106CFU60.5log per sample within15min.The comparison achieved with the parameters does not necessarily correlate with or predict performance of the material when different parameters are ud.
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11.9Adjust the nebulizerflowmeter to aflow rate that will produce droplets that are within the appropriate particle size range.
11.10If necessary,adjust the system vacuum to achieve the desiredflow rate.
N OTE2—When testing samples of materials,take care to apply enough vacuum to achieve the desiredflow rate,but not enough to cau damage to the0.45-µm membranefilter or the test material.If the porosity of a material sample is too low to apply the statedflow rate without damaging the membrane or material,a lowerflow rate should be ud and documented.If a sufficientflow cannot be attained,this test method should not be ud.Generally,an applied vacuum of less than12in.Hg will not result in damage to the membrane or materials.
11.11Immediately start the15-min timer.At regular intervals,obrve and adjust(if necessary)allflowmeters to maintain the appropriateflow rate ttings during the15-min test period or other validated time interval.Document any adjustments.
11.12After exposure,turn off the vacuum,and the fan and the airflow to the nebulizer;Open thefiltered exhaust port at the back of the chamber.
11.13Evacuate the chamber for15min by connecting the vacuum source to the chamber through a microbialfilter asmbly.
11.14Disconnect the hos from thefilter units and remove the units from the bottom plate of the exposure chamber. 11.15Disinfect the outside of eachfiltering unit with a suitable sporicide being car
eful not to compromi the test
material.
