1、Designation: D7464 08 (Reapproved 2013)D7464 14Standard Practice forManual Sampling of Liquid Fuels, Associated Materials andFuel System Components for Microbiological Testing1This standard is issued under the fixed designation D7464; the number immediately following the designation indicates the ye
2、ar oforiginal adoption or, in the case of revision, the year of last revision. A number in parentheses indicates the year of last reapproval. Asuperscript epsilon () indicates an editorial change since the last revision or reapproval.INTRODUCTIONThere are several important characteristics that disti
3、nguish microbiological parameters from otherparameters for which manually collected fuel samples are tested.Microbes, when present in fuels or fuel systems are invariably present as contaminants. Similarlyto particulates, microbes are discrete entities rather than dissolved solutes in fuel, however,
4、 unlikeinanimate particles; microbes can proliferate or die during the interval between sampling and testing.An important consequence of this is that microbes introduced into the sample from sources otherthan the sample itself, can proliferate and potentially eclipse the population indigenous to the
5、 sample.Although microbes can be transported in fuel, they require free-water in order to grow andproliferate. Consequently, microbes tend to form colonies that are embedded in hydrophilic matrices.These matrices are most likely to form at system interfaces, including: fuel-water, fuel-structure,bot
6、tom-water-structure and air and fuel-vapor to structure. Microbes growing within these coloniesproduce chemicals (metabolites and biomolecular detritus) that are deteriogenic (can degrade fuel andfuel system components) and diffuse into fuel.These factors combine to require unique practices specific
7、 to the collection of samples that areintended for microbiological testing.1. Scope Scope*1.1 This practice covers aspects of sample device preparation and sample handling that prevent samples from becomingcontaminated with microorganisms not originally contained within the sample.1.2 This practice
8、also covers sample handling considerations that reflect the perishability of samples collected formicrobiological testing.1.3 This practice supplements Practice D4057 by providing guidance specific to the manual sampling of fuels when samplesare to be tested for microbial contamination.1.4 The value
9、s stated in SI units are to be regarded as standard. No other units of measurement are included in this standard.1.5 This standard does not purport to address all of the safety concerns, if any, associated with its use. It is the responsibilityof the user of this standard to establish appropriate sa
10、fety and health practices and determine the applicability of regulatorylimitations prior to use.2. Referenced Documents2.1 ASTM Standards:2D396 Specification for Fuel OilsD910 Specification for Aviation GasolinesD975 Specification for Diesel Fuel OilsD1129 Terminology Relating to Water1 This practic
11、e is under the jurisdiction of ASTM Committee D02 on Petroleum Products Products, Liquid Fuels, and Lubricants and is the direct responsibility ofSubcommittee D02.14 on Stability and Cleanliness of Liquid Fuels.Current edition approved May 1, 2013Oct. 1, 2014. Published August 2013October 2014. Orig
12、inally approved in 2008. Last previous edition approved in 20082013 asD7464 08.D7464 08 (2013). DOI: 10.1520/D7464-08R13.10.1520/D7464-14.2 For referencedASTM standards, visit theASTM website, www.astm.org, or contactASTM Customer Service at serviceastm.org. For Annual Book of ASTM Standardsvolume i
13、nformation, refer to the standards Document Summary page on the ASTM website.This document is not an ASTM standard and is intended only to provide the user of an ASTM standard an indication of what changes have been made to the previous version. Becauseit may not be technically possible to adequatel
14、y depict all changes accurately, ASTM recommends that users consult prior editions as appropriate. In all cases only the current versionof the standard as published by ASTM is to be considered the official document.*A Summary of Changes section appears at the end of this standardCopyright ASTM Inter
15、national, 100 Barr Harbor Drive, PO Box C700, West Conshohocken, PA 19428-2959. United States1D1193 Specification for Reagent WaterD1655 Specification for Aviation Turbine FuelsD2069 Specification for Marine Fuels (Withdrawn 2003)3D2880 Specification for Gas Turbine Fuel OilsD3508 Method for Evaluat
16、ing Water Testing Membrane Filters for Fecal Coliform Recovery (Withdrawn 1995)3D3699 Specification for KerosineD4057 Practice for Manual Sampling of Petroleum and Petroleum ProductsD4814 Specification for Automotive Spark-Ignition Engine FuelD5245 Practice for Cleaning Laboratory Glassware, Plastic
17、ware, and Equipment Used in Microbiological AnalysesD6227 Specification for Unleaded Aviation Gasoline Containing a Non-hydrocarbon ComponentD6469 Guide for Microbial Contamination in Fuels and Fuel SystemsD6751 Specification for Biodiesel Fuel Blend Stock (B100) for Middle Distillate FuelsD6974 Pra
18、ctice for Enumeration of Viable Bacteria and Fungi in Liquid FuelsFiltration and Culture Procedures2.2 American Petroleum Institute (API) Standard:4Manual of Petroleum Measurement Standards Chapter 3Tank Gauging, section 1AStandard Practice for the ManualGauging of Petroleum and Petroleum Products2.
19、3 Petroleum Equipment Institute (PEI) Standard:5900-08 Recommended Practices for the Inspection and Maintenance of UST Systems3. Terminology3.1 For definition of terms used in this method refer to Terminologies D1129 and D4175, Practice D4057 and Guide D6469.3.2 Definitions:3.2.1 aseptic, adjsterile
20、, free from viable microbiological contamination.3.2.2 scrape sample, na portion of residue removed from a surface by forceful strokes of an instrument such as a spatula.4. Summary of Practices4.1 Liquid Sampling:4.1.1 Fuel and fuel-associated bottom-water samples intended for microbiological testin
21、g are collected similarly to conventionalsamples as described in Practice D4057, however specific measures are added to reduce the risk of sample contamination.4.1.2 Sampling devices are disinfected before collecting microbiological samples.4.1.3 Sterile sample containers are used.4.1.4 Unique chain
22、 of custody procedures are used to minimize the potential qualitative, quantitative or both types of changesin the sample between sampling and testing.4.2 Surface Sampling:4.2.1 Sterile swabs are used to collect surface samples for microbiological testing.4.2.2 Swabbed areas are measured to facilita
23、te test result normalization into parameter units per unit surface area (for exampleCFU/cm2).4.2.3 The post-sampling chain of custody procedures for liquid samples apply.4.3 Filter Media:4.3.1 Canister Elements:4.3.1.1 Filter elements are transferred aseptically to sterile plastic bags.4.3.1.2 The p
24、ost-sampling chain of custody procedures for liquid samples apply.4.3.2 Depth Media:4.3.2.1 Media core-samples are collected aseptically and transferred to tared, sterile containers.4.3.2.2 The post-sampling chain of custody procedures for liquid samples apply.5. Significance and Use5.1 Representati
25、ve samples of fuel products and associated substances are required for the determination of microbialcontamination in fuels and fuel systems in order to accurately assess the biodeterioration risk posed to the fuel, fuel-systemcomponents or both. Uncontrolled microbial contamination can affect fuel
26、specification properties adversely.6 As discussed in3 The last approved version of this historical standard is referenced on www.astm.org.4 Available from American Petroleum Institute (API), 1220 L. St., NW, Washington, DC 20005-4070, http:/www.api.org.5 Available from Petroleum Equipment Institute
27、website, www.pei.org.6 Passman, F. J., McFarland, B. L., and Hillyer, M. J., “Oxygenated Gasoline Biodeterioration and its Control in Laboratory Microcosms,” International Biodeteriorationand Biodegradation, Vol 47, No. 2, 2001, pp. 95-106.D7464 142Guide D6469, microbes can cause a variety of operat
28、ional problems, including filter plugging and microbially influenced corrosion(MIC), the latter of which causes valve failure, tank and pipeline failure.5.2 These practices for microbiological sampling decrease the risk of contaminating samples with extraneous microbes, therebyincreasing the probabi
29、lity that the original microbial population in the sample does not change significantly between the time ofsampling and the time of testing.5.3 The objective of sampling for microbiological testing is to obtain a representative samplesamples that isare likely to reflectthe degree and nature of micro
30、bial contamination in the system from which the sample issamples are collected. Manual 477addresses the rational for and design of microbial contamination programs. Recognizing that microbiological contamination is notdistributed uniformly throughout fuel systems, both the number and types of sample
31、s collected will normally be different from thesamples collected per Practice D4057 in order to determine whether product meets specifications.5.4 The physical, chemical and microbiological property tests to be performed on a sample will dictate the sampling procedures,the sample quantity required,
32、and many of the sample handling requirements.5.5 Fuel systems are not normally designed to facilitate optimal microbiological sampling. Consequently, the selection ofsampling device and sample source reflect compromises between accessibility and suitability for meeting the sample collectionobjective
33、.5.6 The guidance provided in Practice D4057 generally applies to this practice as well. Consequently, this practice will addressonly those procedures that apply uniquely to microbiological sampling.6. Apparatus6.1 The general considerations provided in Practice D4057 apply here. Sample containers c
34、ome in a variety of shapes, sizes andmaterials. To paraphrase D4057, Paragraph 6.1, in order to be able to select the right container for a given application one mustensure that there will be no interaction between the sampled material and the container which would affect the integrity of the other.
35、For general microbiological testing, either glass or plastic containers are appropriate. However, containers should be appropriatefor the specific method of analysis intended.6.1.1 Sample Container Cleanliness:6.1.1.1 Sample containers must be clean and should be sterile.6.1.1.2 For the purposes of
36、most microbiological testing, previously unused containers that are received in originalmanufacturers packaging are sufficiently clean to substitute for sterile containers.6.1.1.3 Practice D5245 provides details on cleaning previously used glassware, plasticware and equipment.6.1.1.4 Method D3508 sp
37、ecifies the protocol for sterilizing containers and labware.6.2 Sampling DevicesSampling devices are described in detail under each of the specific sampling procedures.6.2.1 Sampling Device Cleanliness:6.2.1.1 Sampling devices shall be cleaned between use in accordance with 8.2.1, except cleaning is
38、 not necessary betweenrepeated spot samples obtained either for the purpose of filling a single sample container or filling multiple sample containersintended to be used as replicate spot samples. Such replicates may be used to test the sample for different parameters, when thecontents of a single s
39、ample container are used for a single analysis (for example Practice D6974), for obtaining replicate data inorder to determine parameter variability, or both.6.2.1.2 It can be impractical to sterilize some types of sampling devices used to obtain liquid petroleum, petroleum product orfuel-associated
40、, free-water samples (see 8.2).6.3 Funnel20 to 25 cm diameter mouth; 1.9 cm diameter outlet (diameter small enough to fit into mouth of samplecontainer).6.4 Absorbent Spill Pads.6.5 Gloves; SurgicalUsed to prevent the contamination of samples with microorganisms indigenous to human skin.NOTE 1The us
41、e of surgical gloves may create a static electricity discharge risk that presents an explosion hazard when handling certain fuels.Additionally, polymers from which some surgical gloves are manufactured are incompatible with certain fuels, and can disintegrate on contact with suchfuel, thereby creati
42、ng a skin contact hazard. Where either spark, product incompatibility or both types of risk exist, use an alternative, clean, non-porousglove that has been disinfected in accordance with 8.2 in order to address the explosion hazard risk and still minimize the risk of contaminating sampleswith microb
43、es associated with human skin.6.6 SpatulaStainless steel; 1.5 by 10 cm for collecting surface residue samples.6.7 SwabsSterile, ATP-free.7 Hill, G., “Sampling Methods for Detecting Microbial Contamination in Fuels and Fuel Systems,” in Passman, F. J., Ed., ASTM Manual 47Fuel and Fuel SystemMicrobiol
44、ogy: Fundamentals, Diagnosis and Contamination Control, ASTM International, West Conshohocken, PA, 2003.Hill, G., “Sampling Methods for DetectingMicrobial Contamination in Fuels and Fuel Systems,” in Passman, F. J., Ed., ASTM Manual 47Fuel and Fuel System Microbiology: Fundamentals, Diagnosis andCon
45、tamination Control , ASTM International, West Conshohocken, PA, 2003.D7464 1437. Reagents7.1 Alcohol, 70 % methanol, ethanol or isopropanol, technical grade.7.2 WaterType I Reagent Grade or better (Specification D1193; Terminology D1129).8. Manual Sampling Considerations8.1 The considerations detail
46、ed in Practice D4057 Section 7 apply.8.2 Sampling Device Disinfection:8.2.1 Before collecting a sample, the sampling device shall be cleaned and disinfected. Due to the risk of fire and explosionwhen handling liquid fuels with boiling points below 90C, procedures generally used to disinfect apparatu
47、s used formicrobiological sampling cannot be used in the liquid fuel environment. The following procedure shall be used instead:8.2.1.1 Clean the device, taking particular care to remove any liquid and particulate residue remaining from previous samples.8.2.1.2 Rinse device with alcohol (7.1) by fil
48、ling the device approximately 14 to 13 with alcohol and shaking the closed devicefor 30 s.8.2.1.3 Drain the alcohol thoroughly from the device into a suitable disposal container.8.2.1.4 Allow all residual alcohol to evaporate from device surfaces.9. Special Precautions9.1 The precautions enumerated
49、in Practice D4057 Section 8 apply to sampling for microbiological testing.9.2 Contamination ControlAdditional caution is required to prevent the contamination of samples with non-indigenousmicrobes.9.2.1 The normal microflora of healthy skin is 1 103 bacteria/cm2. Precautions shall be taken to minimize the risk ofcontaminating samples with skin microflora. Wearing surgical gloves provides an adequate barrier between the skin, samplingdevices and sample containers. Gloves should either be replaced or rinsed with 70 % alcohol (7.1) between samples. (See N
