Sec. 79.57. Emission generation. This section specifies the equipment and procedures that must be used in generating the emissions which are to be subjected to the characterization procedures and/ or the biological tests specified in Secs. 79.52( b) and 79.53 of these regulations. When applicable, they may also be required in conjunction with testing under Secs. 79.54 and 79.58( c). Additional requirements concerning emission generation, delivery, dilution, quality control, and safety practices are outlined in Sec. 79.61. ( a) Vehicle and engine selection criteria. ( 1) All vehicles and engines used to generate emissions for testing a fuel or additive/ fuel mixture must be new ( i. e., never before titled) and placed into the program with less than 500 miles on the odometer or 12 hours on the engine chronometer. The vehicles and engines shall be unaltered from the specifications of the original equipment manufacturer. ( 2) The vehicle/ engine type, vehicle/ engine class, and vehicle/ engine subclass designated to generate emissions for a given fuel or additive shall be the same type, class, and subclass which, over the previous three years, has consumed the most gallons of fuel in the fuel family applicable to the given fuel or additive. No distinction shall be made between light duty vehicles and light duty trucks for purposes of this classification. ( 3) Within this vehicle/ engine type, class, and subclass, the specific vehicles and engines acceptable for emission generation are those that represent the most common fuel metering system and the most common of the most important emission control system devices or characteristics with respect to emission reduction performance for the model year in which testing begins. These vehicles will be determined through a survey of the previous model year's vehicle/ engine sales within the given subclass. These characteristics shall include, but need not be limited to, aftertreatment device( s), fuel aspiration, air injection, exhaust gas recirculation, and feedback type. ( 4) Within the applicable subclass, the five highest selling vehicle/ engine models that contain the most common such equipment and characteristics shall be determined. Any of these five models of the current model year ( at the time testing begins) may be selected for emission generation. ( i) If one or more of the five models is not available for the current model year, the choice of model for emission generation shall be limited to those remaining among the five. ( ii) If fewer than five models of the given vehicle/ engine type are available for the current model year, all such models shall be eligible. ( 5) When the fuel or fuel additive undergoing testing is not commonly used or intended to be used in the vehicle/ engine types prescribed by this selection procedure, or when rebuilding or alteration is required to obtain a suitable vehicle/ engine for emission generation, the manufacturer may submit a request to EPA for a modification in test procedure requirements. Any such request must include objective test results which support the claim that a more appropriate vehicle/ engine type is needed as well as a suggested substitute vehicle/ engine type. The vehicle/ engine selection in this case shall be approved by EPA prior to the start of testing. ( 6) Once a particular model has been chosen on which to test a fuel or additive product, all mileage accumulation and generation of emissions for characterization and biological testing of such product shall be conducted on that same model. ( i) If the initial test vehicle/ engine fails or must be replaced for any reason, emission generation shall continue with a second vehicle/ engine which is identical to, or resembles to the greatest extent possible, the initial test vehicle/ engine. If more than one replacement vehicle/ engine is necessary, all such vehicles/ engines shall be identical, or resemble to the greatest extent possible, the initial test vehicle/ engine. ( ii) Manufacturers are encouraged to obtain, at the start of a test program, more than one emission generation vehicle/ engine of the identical model, to ensure the availability of back up emission generator( s). All backup vehicles/ engines must be conditioned and must have their emissions fully characterized, as done for the initial test vehicle/ engine, prior to their use as emission generators for biological testing. Alternating between such vehicles/ engines regularly during the course of testing is permissible and advisable, particularly to allow regular maintenance on such vehicles/ engines during prolonged health effects testing. ( b) Vehicle/ engine operation and maintenance. ( 1) For the purpose of generating combustion emissions from a fuel or additive/ base fuel mixture for which the relevant class is light duty, either a light duty vehicle shall be operated on a chassis dynamometer or a light duty engine shall be operated on an engine dynamometer. When the relevant class is heavy duty, the emissions shall be generated on a heavy duty engine operated on an engine dynamometer. In both cases, the vehicle or engine model shall be selected as described in paragraph ( a) of this section and shall have all applicable fuel and emission control systems intact. ( 2) Except as provided in Sec. 79.51( h)( 2)( iii), the fuel or additive/ base fuel mixture being tested shall be used at all times during operation of the test vehicle or engine. No other fuels or additives shall be used in the test vehicle or engine once mileage accumulation has begun until emission generation for emission characterization and biological testing purposes is completed. ( 3) Scheduled and unscheduled vehicle/ engine maintenance. ( i) During emission generation, vehicles and engines must be maintained in good condition by following the recommendations of the original equipment manufacturer ( OEM) for scheduled service and parts replacement, with repairs performed only as necessary. Modifications, adjustments, and maintenance procedures contrary to procedures found in 40 CFR part 86 for the maintenance of test vehicles/ engines or performed solely for the purpose of emissions improvement are not allowed. ( ii) If unscheduled maintenance becomes necessary, the vehicle or engine must be repaired to OEM specifications, using OEM or OEMapproved parts. In addition, the tester is required to measure the basic emissions pursuant to Sec. 79.52( b)( 2)( i) after the unscheduled maintenance and before resuming testing to ensure that the postmaintenance emissions shall be within 20 percent of pre maintenance emissions levels. If the basic emissions cannot be brought within 20 percent of their previous levels, then the manufacturer shall restart the emissions characterization and health testing of its products combustion emissions using a new vehicle/ engine. ( c) Mileage accumulation. ( 1) A vehicle/ engine break in period is required prior to generating emissions for characterization and/ or biological testing under this subpart. The required mileage accumulation may be accomplished on a test track, on the street, on a dynamometer, or using any other conventionally accepted method. ( 2) Vehicles to be used in the evaluation of baseline and nonbaseline fuels and fuel additives shall accumulate 4,000 miles prior to emission testing. Engines to be used in the evaluation of baseline and non baseline fuels and fuel additives shall accumulate 125 hours of operation on an engine dynamometer prior to emission testing. ( 3) When the test formulation is classified as an atypical fuel or fuel additive formulation ( pursuant to definitions in Sec. 79.56( e)( 4)( iii)), the following additional mileage accumulation requirements apply: ( i) The test vehicle/ engine must be operated for a minimum of 4,000 vehicle miles or 125 hours of engine operation. ( ii) Thereafter, at intervals determined by the tester, all emission fractions ( i. e., vapor, semi volatile, and particulate) shall be sampled and analyzed for the presence and amount of the atypical element( s) and/ or other atypical constituents. Pursuant to paragraph ( d) of this section, the sampled emissions must be generated in the absence of an intact aftertreatment device. Immediately before the samples are taken, a brief warmup period ( at least ten miles or the engine equivalent) is required. ( iii) Mileage accumulation shall continue until either 50 percent or more of the mass of each atypical element ( or other atypical constituent) entering the engine can be measured in the exhaust emissions ( all fractions combined), or the vehicle/ engine has accumulated mileage ( or hours) equivalent to 40 percent of the average useful life of the applicable vehicle/ engine class ( pursuant to regulations in 40 CFR part 86). For example, the maximum mileage required for light duty vehicles is 40 percent of 100,000 miles ( i. e., 40,000 miles), while the maximum time of operation for heavy duty engines is the equivalent of 40 percent of 290,000 miles ( i. e., the equivalent in engine hours of 116,000 miles). ( iv) When either condition in paragraph ( c)( 3)( iii) of this section has been reached, additional emission characterization and biological testing of the emissions may begin. ( d) Use of exhaust aftertreatment devices. ( 1) If the selected test vehicle/ engine, as certified by EPA, does not come equipped with an emissions aftertreatment device ( such as a catalyst or particulate trap), such device shall not be used in the context of this program. ( 2) Except as provided in paragraph ( d)( 3) of this section for certain specialized additives, the following provisions apply when the test vehicle/ engine, as certified by EPA, comes equipped with an emissions aftertreatment device. ( i) For mileage accumulation: ( A) When the test formulation does not contain any atypical elements ( pursuant to definitions in Sec. 79.56( e)( 4)( iii)), an intact aftertreatment device must be used during mileage accumulation. ( B) When the test formulation does contain atypical elements, then the manufacturer may choose to accumulate the required mileage using a vehicle/ engine equipped with either an intact aftertreatment device or with a non functional aftertreatment device ( e. g., a blank catalyst without its catalytic wash coat). In either case, sampling and analysis of emissions for measurement of the mass of the atypical element( s) ( as described in Sec. 79.57( c)( 3)) must be done on emissions generated with a non functional ( blank) aftertreatment device. ( 1) If the manufacturer chooses to accumulate mileage without a functional aftertreatment device, and if the manufacturer wishes to do this outside of a laboratory/ test track setting, then a memorandum of exemption for product testing must be obtained by applying to the Director of the Field Operations and Support Division ( see Sec. 79.59( a)( 1)). ( 2) [ reserved] ( ii) For Tier 1 ( Sec. 79.52), the total set of requirements for the characterization of combustion emissions ( Sec. 79.52( b)) must be completed two times, once using emissions generated with the aftertreatment device intact and a second time with the aftertreatment device rendered nonfunctional or replaced with a non functional aftertreatment device as described in paragraph ( d)( 2)( i)( B) of this section. ( iii) For Tier 2 ( Sec. 79.53), the standard requirements for biological testing of combustion emissions shall be conducted using emissions generated with a non functioning aftertreatment device as described in paragraph ( d)( 2)( i)( B) of this section. ( iv) For alternative Tier 2 requirements ( Sec. 79.58( c)) or Tier 3 requirements ( Sec. 79.54) which may be prescribed by EPA, the use of functional or nonfunctional aftertreatment devices shall be specified by EPA as part of the test guidelines. ( v) In the case where an intact aftertreatment device is not in place, all other manufacturer specified combustion characteristics ( e. g., back pressure, residence time, and mixing characteristics) of the altered vehicle/ engine shall be retained to the greatest extent possible. ( 3) Notwithstanding paragraphs ( d)( 1) and ( d)( 2) of this section, when the subject of testing is a fuel additive specifically intended to enhance the effectiveness of exhaust aftertreatment devices, the related aftertreatment device may be used on the emission generation vehicle/ engine during all mileage accumulation and testing. ( e) Generation of combustion emissions ( 1) Generating combustion emissions for emission characterization. ( i) Combustion emissions shall be generated according to the exhaust emission portion of the Federal Test Procedure ( FTP) for the certification of new motor vehicles, found in 40 CFR part 86, subpart B for light duty vehicles/ engines, and subparts D, M and N for heavy duty vehicles/ engines. The Urban Dynamometer Driving Schedule ( UDDS), pursuant to 40 CFR part 86, appendix I( a), shall apply to light duty vehicles/ engines and the Engine Dynamometer Driving Schedule ( EDS), pursuant to 40 CFR part 86, appendix I( f)( 2), shall apply to heavy duty vehicles/ engines. The motoring portion of the heavy duty test cycle may be eliminated, at the manufacturer's option, for the generation of emissions. ( A) For light duty engines operated on an engine dynamometer, the tester shall determine the speed torque equivalencies (   trace'') for its test engine from valid FTP testing performed on a chassis dynamometer, using a test vehicle with an engine identical to that being tested. The test engine must then be operated under these speed and torque specifications to simulate the FTP cycle. ( B) Special procedures not included in the FTP may be necessary in order to characterize emissions from fuels and fuel additives containing atypical elements or to collect some types of emissions ( e. g., particulate emissions from light duty vehicles/ engines, semivolatile emissions from both light duty and heavy duty vehicles/ engines). Such alterations to the FTP are acceptable. ( ii) Pursuant to Sec. 79.52( b)( 1)( i) and Sec. 79.57( d)( 2)( ii), emission generation and characterization must be repeated three times when the selected vehicle/ engine is normally operated without an emissions aftertreatment device and six times when the selected vehicle/ engine is normally operated with an emissions aftertreatment device. In the latter case, the emission generation and characterization process shall be repeated three times with the intact aftertreatment device in place and three times with a non functioning ( blank) aftertreatment device in place. ( iii) From both light duty and heavy duty vehicles/ engines, samples of vapor phase, semi volatile phase, and particulate phase emissions shall be collected, except that semi volatile phase, and particulate emissions need not be sampled for fuels and additives in the methane and propane families ( pursuant to Sec. 79.56( e)( 1)( v) and ( vi)). The number and type of samples to be collected and separately analyzed during one emission generation/ characterization process are as follows: ( A) In the case of combustion emissions generated from light duty vehicles/ engines, the samples consist of three bags of vapor emissions ( one from each segment of the light duty exhaust emission cycle) plus one sample of particulate phase emissions and one sample of semivolatile phase emissions ( collected over all segments of the exhaust emission cycle). If the mass of particulate emissions or semi volatile emissions obtained during one driving cycle is not sufficient for characterization, then the driving cycle may be performed again and the extracted fractions combined prior to chemical analysis. Particulatephase emissions shall not be combined with semi volatile phase emissions. ( B) In the case of combustion emissions generated from heavy duty engines, the samples consist of one sample of each emission phase ( vapor, particulate, and semi volatile) collected over the entire coldstart cycle and a second sample of each such phase collected over the entire hot start cycle ( see 40 CFR 86.334 through 86.342). ( iv) Emission collection and storage. ( A) Vapor phase emissions shall be collected and stored in Tedlar bags for subsequent chemical analysis. Storage conditions are specified in Sec. 79.52( b)( 2). ( B) Particulate phase emissions shall be collected on a particulate filter ( or more than one, if required) using methods described in 40 CFR 86.1301 through 86.1344. These methods, ordinarily applied only to heavy duty emissions, are to be adapted and used for collection of particulates from light duty vehicles/ engines, as well. The particulate matter may be stored on the filter in a sealed container, or the soluble organic fraction may be extracted and stored in a separate sealed container. Both the particulate and the extract shall be shielded from ultraviolet light and stored at  20 deg. C or less. Particulate emissions shall be tested no later than six months from the date they were generated. ( C) Semi volatile emissions shall be collected immediately downstream from the particulate collection filters using porous polymer resin beds, or their equivalent, designed for their capture. The soluble organic fraction of semi volatile emissions shall be extracted immediately and tested within six months of being generated. The extract shall be stored in a sealed container which is shielded from ultraviolet light and stored at  20 deg. C or less. ( D) Particulate and semi volatile phase emission collection, handling and extraction methods shall not alter the composition of the collected material, to the extent possible. ( v) Additional requirements for combustion emission sampling, storage, and characterization are specified in Sec. 79.52( b). ( 2) Generating whole combustion emissions for biological testing. ( i) Biological tests requiring whole combustion emissions shall be conducted using emissions generated from the test vehicle or engine operated in general accordance with the FTP procedures cited in this section. The emissions shall be generated continuously throughout the animal exposure periods, diluted by an amount appropriate for the test being performed as specified in Sec. 79.61( d)( 3), passed through a mixing chamber, and routed to the biological test chamber. ( ii) Light duty test vehicles/ engines shall be operated over the Urban Dynamometer Driving Schedule ( or equivalent engine dynamometer trace, per paragraph ( e)( 1)( i)( A) of this section) and heavy duty test engines shall be operated over the Engine Dynamometer Schedule ( see 40 CFR part 86, appendix I). ( A) The tolerances of the driving cycle shall be two times those of the Federal Test Procedure and must be met 95 percent of the time. ( B) The driving cycle shall be repeated as many times as required for the biological test session. ( C) Light duty dynamometers shall be calibrated prior to the start of a biological test ( 40 CFR 86.118 78), verified weekly ( 40 CFR 86.118 78), and recalibrated as required. Heavy duty dynamometers shall be calibrated and checked prior to the start of a biological test ( 40 CFR 86.1318 84), recalibrated every two weeks ( 40 CFR 86.1318 84( a)) and checked as stated in 40 CFR 86.1318 84( b) and ( c). ( D) The fuel reservoir for the test vehicle/ engine shall be large enough to operate the test vehicle/ engine throughout the daily biological exposure period, avoiding the need for refueling during testing. ( iii) An apparatus to integrate the large concentration swings typical of transient cycle exhaust is to be used between the FTPConstant Volume Sampler ( CVS) source of emissions and the exposure chamber containing the animal test cage( s). The purpose of such apparatus is to decrease the variability of the biological exposure atmosphere. ( A) A large mixing chamber is suggested for this purpose. The mixing chamber would be charged from the CVS at a constant rate determined by the exposure chamber purge rate. Flow to the exposure chamber would begin at the conclusion of the initial transient cycle with the associated mixing chamber charge. ( B) A potential alternative apparatus is a mini diluter ( see, for example, AIGER/ CRADA, February, 1994 in Sec. 79.57( g)). ( C) The mixing chamber ( or any alternative emission moderation apparatus) must function such that the average concentration of total hydrocarbons leaving the apparatus shall be within 10 percent of the average concentration of hydrocarbons entering the chamber. ( iv) Emission dilution. ( A) Dilution air can be pre dried to lower the relative humidity, thus permitting a lower dilution rate and a higher concentration of hydrocarbons to be achieved without condensation of water vapor. ( B) With gasoline fuels, a minimum dilution ratio of about 1: 5 raw exhaust ( dewpoint about 125 deg. F) with dry, clean filtered air is required to reduce the water concentration to a dewpoint of about 68 deg. F. The minimum dilution ratio ( maximum exhaust flow rate) occurs at about 200 seconds into the UDDS transient driving cycle. Larger minimum dilution ratios are required if the dilution air includes water vapor. However, the minimum dilution ratio will vary with fuel composition. Fuels which generate greater engine exhaust water concentrations ( e. g., alcohol and natural gas fuels) will require greater initial dilutions. Heated transfer ducts or tubing can be used to avoid water condensation in much of the system, but the mixing chamber described in paragraph ( e)( 2)( iii) of this section will generally be at or near laboratory temperature, and CVS dilution will have to be adequate to assure that the cumulative dew point in the chamber remains below laboratory temperature at all times ( further guidance on this topic may be found in Black and Snow, 1994 in Sec. 79.57( g)). ( C) After the initial exhaust dilution to preserve the character of the exhaust, the exhaust stream can be further diluted in the mixing chamber ( and/ or after leaving the chamber) to achieve the desired biological exposure concentrations. ( v) Verification procedures. ( A) The entire system used to dilute and transport whole combustion emissions ( i. e., from exhaust pipe to outlet in the biological testing chamber) shall be verified before any animal exposures begin, and verified at least weekly during testing. ( See procedures at 40 CFR 86.119 90 for light duty vehicles and Sec. 86.1319 90 for heavy duty engines.) Verification testing shall be accomplished by introducing a known sample at the end of the vehicle/ engine exhaust pipe into the dilution system and measuring the amount exiting the system. For example, an injected hydrocarbon sample could be detected with a gas chromatograph ( GC) and flame ionization detector ( FID) to determine the recovery factor. ( B) Verification of the integrity of the mixing chamber ( or alternative apparatus) shall be determined before animal exposures begin and at least weekly thereafter. Composite values for weight percent total hydrocarbons shall be determined for the test vehicle/ engine's dilute exhaust stream entering and exiting the mixing chamber apparatus. These values must be within 10 percent of each other. ( vi) Emission exposure quality control. ( A) The tester shall incorporate the additional quality assurance and safety procedures outlined in Sec. 79.61( d) to control variability of emissions during the generation of exposure emissions during health effect testing. ( B) These procedures include requirements that the mean exposure concentration in the inhalation test chamber shall be within 10 percent of the target concentration ( established in the developmental phase of testing) on 90 percent or more of exposure days and that daily monitoring of CO, CO< INF> 2, NO< INF> X, SO< INF> X, and total hydrocarbons in the exposure chamber shall be required. Analysis of the particle size distribution shall also be performed to establish the stability and consistency of particle size distribution in the test exposure. ( C) The testing facility shall allow an audit of its premises, the qualifications, e. g., curriculum vitae, of its staff assigned to testing, and the specimens and records of the testing for registration purposes ( as specified in Sec. 79.60). ( vii) In order to allow for unforeseen problems with the emission generation or dilution equipment, emission generation may be interrupted for up to four hours on a maximum of two occasions in any four week period of testing. The amount of time for which emission generation was interrupted shall subsequently be added after the equipment problem is corrected. If the equipment problem causes more than four consecutive hours of emission generation to be interrupted, or if more than two such occasions occurs in any four week period during testing, the interrupted tests shall be void. Testers shall be aware of concerns for backup vehicles/ engines cited in paragraph ( a)( 7)( ii) of this section. ( 3) Generating particulate and semi volatile emissions for biological testing. ( i) Salmonella mutagenicity testing, pursuant to Sec. 79.68, shall be conducted on extracts of the particulate and semivolatile emission phases separately. These emissions shall be generated by operating the test vehicle/ engine over the appropriate FTP driving cycle ( see paragraph ( e)( 2)( ii) of this section) and collected and analyzed according to methods described in 40 CFR 86.1301 through 1344 ( further information on this subject may be found in Perez, et al. CRC Report No. 551, 1987 listed in Sec. 79.57( g)). ( A) Particulate emissions shall be collected on particulate filters and extracted from the collection equipment for use in biological tests. The particulate emissions from all segments of the FTP or from multiple FTP cycles may be collected on one or more filters, as necessary. The time spent collecting sufficient quantities of the test substances in emissions samples will vary, depending on the emission characteristics of the engine and fuel or additive/ base fuel mixture and on the requirements of the biological test protocol. ( B) Semi volatile emissions shall be collected immediately downstream from the particulate collection filters using porous polymer resin beds, or their equivalent, designed for their capture. Semivolatile phase emissions shall be collected on one apparatus. The time spent collecting sufficient quantities of the test substances in emissions samples will vary, depending on the emission characteristics of the engine and fuel or additive/ base fuel mixture and on the requirements of the biological test protocol. ( ii) The extraction method shall be determined by the specifications of the biological test for which the emissions are used. ( iii) Particulate and semi volatile emission storage requirements are as specified in Sec. 79.57( e)( 1)( iv). ( iv) Particulate and semi volatile phase emission collection, handling and extraction methods shall not alter the composition of the collected material, to the extent possible. ( v) Particulate emissions shall not be combined with semi volatile phase emissions. ( f) Generation of evaporative emissions for characterization and biological testing. ( 1) Except as provided in paragraph ( f)( 5) of this section, an evaporative emissions generator shall be used to volatilize samples of a fuel or additive/ base fuel mixture for evaporative emissions characterization and biological testing. Emissions shall be collected and sampled using equipment and methods appropriate for use with the compounds being characterized and the requirements of the emission characterization analysis. In the case of potentially explosive test substance concentrations, care must be taken to avoid generating explosive atmospheres. The tester is referred to Sec. 79.61( d)( 8) for considerations involving explosivity. ( 2) Evaporative Emissions Generator ( EEG) Description. An EEG is a fuel tank or vessel to which heat is applied causing a portion of the fuel to evaporate at a desired rate. The manufacturer has flexibility in designing an EEG for testing a particular fuel or fuel additive. The sample used to generate emissions in the EEG shall be renewed at least daily. ( i) The evaporation chamber shall be made from materials compatible with the fuels and additives being tested and shall be equipped with a drain. ( ii) The chamber shall be filled to 40 < plus minus> 5 percent of its interior volume with the fuel or additive/ base fuel mixture being tested, with the remainder of the volume containing air. ( iii) The concentration of the evaporated fuel or additive/ base fuel mixture in the vapor space of the evaporation chamber during the time emissions are being withdrawn for testing shall not vary by more than 10 percent from the equilibrium concentration in the vapor space of emissions generated from the fresh fuel or additive/ base fuel mixture in the chamber. ( A) During the course of a day's emission generation period, the level of fuel in the EEG shall be maintained to within 7 percent of its height at the start of the daily exposure period. ( B) The fuel used in the EEG shall be drained at the end of each daily exposure. The EEG shall be refilled with a fresh supply of the test formulation before the start of each daily exposure. ( C) The vapor space of the evaporation chamber shall be well mixed throughout the time emissions are being withdrawn for testing. ( iv) The size of the evaporation chamber shall be determined by the rate at which evaporative emissions shall be needed in the test animal exposure chambers and the rate at which the fuel or the additive/ base fuel mixture evaporates. The rate of evaporative emissions may be adjusted by altering the size of the EEG or by using one or more additional EEG( s). Emission rate modifications shall not be adjusted by temperature control or pressure control. ( v) The temperature of the fuel or additive/ base fuel mixture in the evaporation chamber shall be 130 deg. F< plus minus> 5 deg. F. The vapors shall maintain this temperature up to the point in the system where the vapors are diluted. ( vi) The pressure in the vapor space of the evaporation chamber and the dilution and sampling apparatus shall stay within 10 percent of ambient atmospheric pressure. ( vii) There shall be no controls or equipment on the evaporation chamber system that change the concentration or composition of the vapors generated for testing. ( viii) Manufacturers shall perform verification testing of evaporative emissions in a manner analogous to the verification testing performed for combustion emissions. ( 3) For biological testing, vapor shall be withdrawn from the EEG at a constant rate, diluted with air as required for the particular study, and conducted immediately to the biological testing chamber( s) in a manner similar to the method used in Sec. 79.57( e), excluding the mixing chamber therein. The rate of emission generation shall be high enough to supply the biological exposure chamber with sufficient emissions to allow for a minimum of fifteen air changes per exposure chamber per hour. Interruption of evaporative emissions exposures during biological testing for more than four consecutive hours, or on more than two separate occasions within a four week period for less than four consecutive hours, shall cause the affected test( s) to be void. ( 4) For characterization of evaporative emissions, samples of equilibrated emissions to the vapor space of the EEG shall be withdrawn into Tedlar bags, then stored and analyzed as specified in Sec. 79.52( b). ( 5) A manufacturer ( or group of manufacturers) may submit to EPA a request for approval of an alternative method of generating evaporative emissions for use in emission characterization and biological tests required under this subpart. ( i) To be approved by EPA, the request must fully explain the rationale for the proposed method as well as the technical procedures, quality control, and safety precautions to be used, and must demonstrate that the proposed method will meet the following criteria: ( A) The emission mixture generated by the proposed procedures must be reasonably similar to the equilibrium composition of the vapor which occurs in the vehicle fuel tank head space when the subject fuel or additive/ base fuel mixture is in use and near maximum in use temperatures are encountered. ( B) The emissions mixture generated by the proposed method must be sufficiently concentrated to provide adequate exposure levels in the context of the required toxicologic tests. ( C) The proposed method must include procedures to ensure that the emissions delivered to the biologic exposure chambers will provide a reasonably constant exposure atmosphere over time. ( ii) If EPA approves the request, EPA will place in the public record a copy of the request, together with all supporting procedural descriptions and justifications, and will notify the public of its availability by publishing a notice in the Federal Register. ( g) References. For additional background information on the emission generation procedures outlined in this paragraph ( g), the following references may be consulted. Additional references can be found in Sec. 79.61( f). ( 1) AIGER/ CRADA ( American Industry/ Government Emissions Research Cooperative Research and Development Agreement,    Specifications for Advanced Emissions Test Instrumentation'' AIGER PD 94 1, Revision 5.0, February, 1994 ( 2) Black, F. and R. Snow,    Constant Volume Sampling System Water Condensation'' SAE # 940970 in    Testing and Instrumentation'' SP 1039, Society of Automotive Engineers, Feb. 28 Mar. 3, 1994. ( 3) Perez, J. M., Jass, R. E., Leddy, D. G., eds.    Chemical Methods for the Measurement of Unregulated Diesel Emissions ( CRC APRAC Project No. CAPI 1 64), Coordinating Research Council, CRC Report No. 551, August, 1987. ( 4) Phalen, R. F.,    Inhalation Studies: Foundations and Techniques'', CRC Press, Inc., Boca Raton, Florida, 1984. Sec. 79.58 Special provisions.

( a) Relabeled Additives. Sellers of relabeled additives ( pursuant to Sec. 79.50) are not required to comply with the provisions of Secs. 79.52, 79.53 or 79.59, except that such sellers are required to comply with Sec. 79.59( b). ( b) Low Vapor Pressure Fuels and Additives. Fuels which are not designated as    evaporative fuels'' and fuel additives which are not designated as    evaporative fuel additives'' pursuant to the definitions in Sec. 79.50 need not undergo the emission characterization or health effects testing specified in Secs. 79.52 and 79.53 for evaporative emissions. At EPA's discretion, the evaporative emissions of such fuels and additives may be required to undergo Tier 3 testing, pursuant to Sec. 79.54. ( c) Alternative Tier 2 Provisions. At EPA's discretion, EPA may modify the standard Tier 2 health effects testing requirements for a fuel or fuel additive ( or group). Such modification may encompass substitution, addition, or deletion of Tier 2 studies or study specifications, and/ or changes in underlying engine or equipment requirements, except that a Tier 2 endpoint will not be deleted in the absence of existing information deemed adequate by EPA or alternative testing requirements for such endpoint. If warranted by the particular requirements, EPA will allow additional time for completion of the alternative Tier 2 testing program. ( 1) When EPA intends to require testing in lieu of or in addition to standard Tier 2 health testing, EPA will notify the responsible manufacturer ( or group) by certified letter of the specific tests which EPA is proposing to require in lieu of or in addition to Tier 2, and the proposed schedule for completion and submission of such tests. A copy of the letter will be placed in the public record. EPA intends to send the notification prior to November 27, 1995, or in the case of new fuels and additives ( as defined in Sec. 79.51( c)( 3)), within 18 months of EPA's receipt of an intent to register such product. However, EPA's notification to the manufacturer ( or group) may occur at any time up to EPA's receipt of Tier 2 data for the product( s) in question. EPA will provide the manufacturer with 60 days from the date of receipt of the notice to comment on the tests which EPA is proposing to require and on the proposed schedule. If the manufacturer believes that undue costs or hardships will occur as a result of EPA's delay in providing notification of alternative Tier 2 requirements, then the manufacturer's comments should describe and include evidence of such hardship. In particular, if the standard Tier 2 toxicology testing for the fuel or additive in question has already begun at the time the manufacturer receives EPA's notification of proposed alternative Tier 2 requirements, then EPA shall refrain from requiring alternative Tier 2 tests provided that EPA receives the standard Tier 2 data and report ( pursuant to Sec. 79.59( c)) within one year of the date on which the toxicology testing began. ( 2) EPA will issue a notice in the Federal Register announcing its intent to require special testing in lieu of or in addition to the standard Tier 2 testing for a particular fuel or additive manufacturer or group, and that a copy of the letter to the manufacturer or group describing the proposed alternative Tier 2 testing for that manufacturer or group is available in the public record for review and comment. The public shall have a minimum of 30 days after the publication of this notice to comment on the proposed alternative Tier 2 testing. ( 3) EPA will include in the public record a copy of any timely comments concerning the proposed alternative Tier 2 testing requirements received from the affected manufacturer or group or from the public, and the responses of EPA to such comments. After reviewing all such comments received, EPA may adopt final alternative Tier 2 requirements by sending a certified letter describing such final requirements to the manufacturer or group. In that event, EPA will also issue a notice in the Federal Register announcing that it has adopted final alternative Tier 2 requirements and that a copy of the letter adopting the requirements has been included in the public record. ( 4) After EPA's receipt of a manufacturer's ( or group's) submittals, EPA will notify the responsible manufacturer ( or group) regarding the adequacy of the submittal and potential Tier 3 testing requirements according to the same relative time intervals and by the same procedures as specified in Sec. 79.51 ( c) and ( d) for routine Tier 1 and Tier 2 submittals. ( d) Small Business Provisions. ( 1) For purposes of these provisions, when subsidiary, divisional, or other complex business arrangements exist, manufacturer is defined as the business entity with ultimate ownership of all related parents, subsidiaries, divisions, branches, or other operating units. Total annual sales means the average of the manufacturer's total sales revenue in each of the three years prior to such manufacturer's submittal to EPA of the basic registration information pursuant to Sec. 79.59 ( b)( 2) through ( b)( 5). ( 2) Provisions Applicable to Baseline and Non baseline Products. A manufacturer with total annual sales less than $ 50 million is not required to meet the requirements of Tier 1 and Tier 2 ( specified in Secs. 79.52 and 79.53) with regard to such manufacturer's fuel and/ or additive products which meet the criteria for inclusion in a Baseline or Non baseline group pursuant to Sec. 79.56. Upon such manufacturer's satisfactory completion and submittal to EPA of basic registration data specified in Sec. 79.59( b), the manufacturer may request and EPA shall issue a registration for such product, subject to Sec. 79.51( c) and paragraphs ( d)( 4) and ( d)( 5) of this section. ( 3) Provisions Applicable to Atypical Products. A manufacturer with total annual sales less than $ 10 million is not required to meet the requirements of Tier 2 ( specified in Sec. 79.53) in regard to such manufacturer's fuel and/ or additive products which meet the criteria for inclusion in an Atypical group pursuant to Sec. 79.56. Upon such manufacturer's satisfactory completion and submittal to EPA of basic registration data specified in Sec. 79.59( b) and Tier 1 information specified in Sec. 79.52 for an Atypical fuel or additive, the manufacturer may request and EPA shall issue a registration for such product, subject to Sec. 79.51( c) and paragraphs ( d)( 4) and ( d)( 5) of this section. Compliance with Tier 1 requirements under this paragraph may be accomplished by the individual manufacturer or as a part of a group pursuant to Sec. 79.56. ( 4) Any registration granted by EPA under the provisions of this section are conditional upon satisfactory completion of any Tier 3 requirements which EPA may subsequently impose pursuant to Sec. 79.54. In such circumstances, the Tier 3 requirements might include ( but would not necessarily be limited to) information which would otherwise have been required under the provisions of Tier 1 and/ or Tier 2. ( 5) The provisions in paragraphs ( d)( 2) and ( d)( 3) of this section are voluntary on the part of qualifying small manufacturers. Such manufacturers may choose to fulfill the standard requirements for their fuels and additives, individually or as a part of a group, rather than satisfying only the requirements specified in paragraphs ( d)( 2) and/ or ( d)( 3) of this section. If a qualifying small manufacturer elects these special provisions rather than the standard requirements for a product, then EPA will generally assume that any additional information submitted by other manufacturers, for fuels and additives meeting the same grouping criteria ( under Sec. 79.56) as that of the small manufacturer's product, is pertinent to further testing and/ or regulatory decisions that may affect the small manufacturer's product. ( e) Aftermarket Aerosol Additives. ( 1) To obtain registration for an aftermarket aerosol fuel additive, the manufacturer shall provide existing information in the form of a literature search, a discussion of the potential exposure( s) to such product, and the basic registration data specified in Sec. 79.59( b). ( 2) The literature search shall include existing data on potential health and welfare effects due to exposure to the aerosol product itself and its raw ( uncombusted) components. The analysis for potential exposures shall be based on the actual or anticipated production volume and market distribution of the particular aerosol product, and its estimated frequency of use. Other Tier 1 and Tier 2 requirements are not routinely required for aerosol products. EPA will review the submitted information and, at EPA's discretion, may require from the manufacturer further information and/ or testing under Tier 3 on a caseby case basis.

Sec. 79.59 Reporting requirements.

( a) Timing. ( 1) The manufacturer of each designated fuel or fuel additive shall submit to EPA the basic registration data detailed in paragraph ( b) of this section. Forms for submitting this data may be obtained from EPA at the following address: Director, Field Operations and Support Division, 6406J Fuel/ Additives Registration, U. S. Environmental Protection Agency, 401 M Street, S. W., Washington, DC 20460. ( i) For existing products ( pursuant to Sec. 79.51( c)( 1)), manufacturers shall submit the basic registration data as specified in Sec. 79.59( b) to EPA by November 28, 1994. ( ii) For registrable products ( pursuant to Sec. 79.51( c)( 2)), manufacturers shall submit the basic registration data as specified in Sec. 79.59( b) to apply for registration for such product. ( iii) For new products ( pursuant to Sec. 79.51( c)( 3)), manufacturers are strongly encouraged to notify EPA of an intent to obtain product registration by submitting the basic registration data as specified in Sec. 79.59( b) prior to starting Tiers 1 and 2. ( 2) The information specified in paragraph ( c) of this section shall be submitted to the address in paragraph ( a)( 1) of this section at the conclusion of activities performed in compliance with Tiers 1 and 2 under the provisions of Secs. 79.52 and 79.53, according to the time constraints specified in Sec. 79.51 ( c) through ( d). ( 3) The information specified in paragraph ( d) of this section shall be submitted to EPA at the address in paragraph ( a)( 1) of this section at the conclusion of activities performed in compliance with Tier 3 under the provisions of Sec. 79.54. ( b) Basic Registration Data. Each manufacturer of a designated fuel or fuel additive shall submit the following data in regard to such fuel or fuel additive: ( 1) The information specified in Sec. 79.11 or Sec. 79.21. If such information has already been submitted to EPA in compliance with subpart B or C of this part, and if such previous information is accurate and up to date, the manufacturer need not resubmit this information. ( 2) Annual production volume of the fuel or fuel additive product, in units of gallons per year if most commonly sold in liquid form or kilograms per year if most commonly sold in solid form. For fuels and fuel additives already in production, the most recent annual production volume and the volume projected to be produced in the third subsequent year shall be provided. For products not yet in production, the best estimate of expected annual volume during the third year of production shall be provided. ( 3) Market distribution of the product. For fuels and bulk additives, this information shall be presented as the percent of total annual sales volume marketed in each Petroleum Administration for Defense District ( PADD). The states comprising each PADD are listed in the following section. For aftermarket additives, the distribution data shall be presented as the percent of total annual sales volume marketed in each state. For a product not yet in production, the manufacturer shall present the distribution ( by PADD or state, as applicable) projected to occur during the third year of production. ( i) The following states and jurisdictions are included in PADD I:

Connecticut Delaware District of Columbia Florida Georgia Maine Maryland Massachusetts New Hampshire New Jersey New York North Carolina Pennsylvania Rhode Island South Carolina Vermont Virginia West Virginia ( ii) The following states are included in PADD II:

Illinois Indiana Iowa Kansas Kentucky Michigan Minnesota Missouri Nebraska North Dakota Ohio Oklahoma South Dakota Tennessee Wisconsin ( iii) The following states are included in PADD III:

Alabama Arkansas Louisiana Mississippi New Mexico Texas ( iv) The following states are included in PADD IV:

Colorado Idaho Montana Utah Wyoming ( v) The following states are included in PADD V:

Alaska Arizona California Hawaii Nevada Oregon Washington ( 4) Any applicable information pursuant to the grouping provisions in Sec. 79.56, as follows: ( i) If the manufacturer has enrolled or intends to enroll the product in a fuel/ additive group, the relevant group and the person( s) or entity expected to submit information on behalf of the group must be identified. ( ii) If the manufacturer intends to rely on registration information previously submitted by another manufacturer ( or group) for registration of other product( s) in the same fuel/ additive group, then the original submitter and its product ( or product group) shall be identified. In such cases, the manufacturer shall provide evidence that the original submitter has been notified of the use of its registration data and that the manufacturer has complied or intends to comply with the proportional reimbursement required under Sec. 79.56( c) of this rule. ( 5) Any applicable information pursuant to the special provisions in Sec. 79.58, as follows: ( i) If the manufacturer claims applicability of the special provisions for relabeled additives, pursuant to Sec. 79.58( a), then the manufacturer and brand name of the original product shall be given. ( ii) If the manufacturer claims applicability of any small business provisions pursuant to Sec. 79.58( d), the average of the manufacturer's total annual sales revenue for the previous three years shall be given. ( iii) If the manufacturer claims applicability of the special provisions for aerosol products, pursuant to Sec. 79.58( e), then the purpose and recommended frequency of use shall be given. ( c) Tier 1 and Tier 2 Reports. If the results of Tiers 1 and 2 are reported to EPA at the same time, then the report shall include the following documents in paragraphs ( c)( 1) through ( 7) of this section. If Tier 1 and Tier 2 results are submitted to EPA separately, then the separate Tier 1 report shall include only documents in paragraphs ( c)( 1) through ( 4), ( c)( 6), and associated appendices in paragraphs ( c)( 7) of this section, and the separate Tier 2 report shall include only documents in paragraphs ( c)( 1) through ( 3), ( c)( 5), ( c)( 6), and associated appendices in paragrpah ( c)( 7) of this section. In addition, pursuant to the requirements in Sec. 79.51( c)( 1)( ii)( B), if the Tier 2 report for registered fuels and fuel additives is not submitted prior to May 27, 1997, then evidence of a suitable arrangement for completion of Tier 2 ( e. g., a copy of a signed contract with a qualified laboratory for applicable Tier 2 services) must be submitted to EPA prior to that date. ( 1) Cover page. ( i) Identification of test substance, ( ii) Name and address of the manufacturer of the test substance, ( iii) Name and phone number of a designated contact person, ( iv) Group information, if applicable, including: ( A) Group name or grouping criteria, ( B) Name and address of responsible organization or entity reporting for the group, ( C) Product trade name and manufacturer of each member fuel and additive to which the report pertains. ( 2) Executive Summary. Text overview of the significant results and conclusions obtained as a result of completing the requirements of Tier 1 and/ or Tier 2, including references if used to support such results and conclusions. ( 3) Test Substance Information. Test substance description, including, as applicable, ( i) Base fuel parameter values ( including types and concentrations of base fuel additives) or test fuel composition ( if a fuel other than the base fuel is used in testing). These values must be provided for each of the fuel parameters specified in Sec. 79.55 for the applicable fuel family. ( ii) Test additive composition and concentration ( 4) Summary of Tier 1 ( i) Literature Search. Pursuant to Sec. 79.52( d), the literature search shall include a text summary of the methods and results of the literature search, including the following: ( A) Identification of person( s) performing the literature search, ( B) Description of data sources accessed, search strategy used, search period, and terms included in literature search, ( C) Documentation of all unpublished in house and other privatelyconducted studies, ( D) Tables summarizing the protocols and results of all cited studies, ( E) Summary of significant results and conclusions with respect to the effects of the emissions of the subject fuel or fuel additive on the public health and welfare, including references if used to support such results and conclusions. ( F) Statement of the extent to which the literature search has produced adequate information comparable to that which would otherwise be obtained through the performance of applicable emission characterization requirements under Sec. 79.52( b) and/ or health effects testing requirements under Sec. 79.53, including justifications and specific references. ( ii) Emission Characterization. Pursuant to Sec. 79.52( b), the emission characterization shall include: ( A) Name, address, and telephone number of the laboratory performing the characterization, ( B) Name and description of analytic methods used for characterization. ( iii) Exposure Analysis. Pursuant to Sec. 79.52( c), the exposure analysis shall include: ( A) A qualitative discussion of the potential exposure( s) of the general and any special at risk populations to the emission products, based on annual and projected production volume, and market distribution data. For group submittals, this discussion shall address the characteristics of the cumulative exposure from the potential use of all fuel or additive products in the group. ( B) Identification of person( s) preparing the analysis. ( 5) Summary of Tier 2. For each health effects test performed pursuant to the provisions of Sec. 79.53, the Tier 2 summary shall contain the following information: ( i) Name, address, and telephone number of the testing facility, ( ii) Summary of procedures ( including quality assurance, quality control and compliance with Good Laboratory Practice Standards as specified in Sec. 79.60), findings, and conclusions, including references if used to support such results and conclusions, ( iii) Description of any problems and their resolution. ( 6) Conclusions. The conclusions shall identify the need for further testing, if that need exists, or justify that current testing and/ or available information is adequate for the tier( s) included in the report. ( 7) Appendices. The appendices shall contain detailed documentation related to the summary information described in this section, including, at a minimum, the following five appendices: ( i) Literature search appendices shall contain: ( A) Copies of literature source outputs, including reference lists and associated abstracts from database searches, printed or on 3\ 1/ 2\ inch IBM compatible computer diskettes; ( B) Summary tables organized by health or welfare endpoint and type of emission ( e. g., combustion, evaporation, individual emission product), presenting in tabular form the following information at a minimum: number and species of test subjects, exposure concentrations/ duration, positive ( i. e., abnormal) findings including numbers of test subjects involved, and bibliographic references; ( C) Complete documentation and/ or reprints of articles for any previous study relied upon for satisfying emission characterization and/ or Tier 2 test requirements; and ( D) Full reports for unpublished/ in house studies. ( ii) Emissions characterization appendices shall contain: ( A) Complete laboratory reports, including documentation of calibration and verification procedures; ( B) Documentation of the emissions generation procedures used; and ( C) Lists of speciated emission products and their emission rates reported in units of grams/ mile. ( iii) Exposure analysis appendices may be submitted to report any detailed documentation of data used in the analyses and/ or calculations determining potential exposures to population( s). If modeling data are used, these should be included in an appendix. ( iv) Tier 2 appendices shall contain, for each test performed: ( A) Complete protocol used; ( B) Documentation of emission generation procedures; and ( C) Complete laboratory report in compliance with the reporting standards in Sec. 79.60, including detailed test results and conclusions, and descriptions of any problems encountered and their resolution. ( v) Laboratory certification/ accreditation information, personnel credentials, and statements of compliance with the Good Laboratory Practices Standards specified in Sec. 79.60 and the requirements in Sec. 79.53( c)( 1). ( d) Tier 3 Report. Subject to applicability as specified in Sec. 79.54, each manufacturer of a designated fuel or fuel additive, or each group of such manufacturers pursuant to the provisions of Sec. 79.56, shall submit the following information with respect to each Tier 3 test conducted for such fuels or fuel additives: ( 1) The test objectives, including a summary of the reason( s) why such additional testing, beyond Tiers 1 and 2, was required; ( 2) Name, address, and telephone number of each testing facility; ( 3) Summary of test procedures, results and conclusions; ( 4) Complete documentation of test protocols and emission generation procedures, complete laboratory reports in compliance with the reporting standards of Sec. 79.60, detailed test results and conclusions, including references if used to support such results and conclusions, and descriptions of any problems encountered and their resolution; and ( 5) Laboratory certification information, personnel credentials, and statements of compliance with the Good Laboratory Practices Standards specified in Sec. 79.60. ( e) Availability of Information. ( 1) All health and safety test data and other information concerning health and welfare effects which is submitted by any manufacturer or group pursuant to Secs. 79.52( c), 79.53, or 79.54, shall be considered to be public information and shall be made available to the public by EPA upon request. A reasonable fee may be charged by EPA for copying such materials. Any manufacturer or group who claims that any information concerning the composition of a fuel or fuel additive product, or any other information, submitted under this subpart is confidential business information must state this claim in writing at the time of the submittal. ( 2) To assert a business confidentiality claim concerning any information submitted under this subpart, the submitter must: ( i) Clearly mark the information as confidential at each location it appears in the submission; and ( ii) Submit with the information claimed as confidential a separate document setting forth the claim and listing each location at which the information appears in the submission. ( 3) If any person subsequently requests access to information submitted under this subpart ( other than health and safety test data and other information concerning health and welfare effects), and such information is subject to a claim of business confidentiality, the request and any subsequent disclosure shall be governed by the provisions of 40 CFR part 2. Sec. 79.60 Good laboratory practices ( GLP) standards for inhalation exposure health effects testing.

( a) General Provisions( 1) Scope. ( i) This section prescribes good laboratory practices ( GLPs) for conducting inhalation exposure studies relating to motor vehicle emissions health effects testing under this part. These directions are intended to ensure the quality and integrity of health effects data submitted pursuant to registration regulations issued under sections 211( b) or 211( e) of the Clean Air Act ( CAA) ( 42 U. S. C. 7545). ( ii) This section applies to any study described by paragraph ( a)( 1)( i) of this section which any person conducts, initiates, or supports on or after May 27, 1994. ( iii) It is EPA's policy that all health effects data developed under sections 211( b) and ( e) of CAA be in accordance with provisions of this section. If data are not developed in accordance with the provisions of this section, EPA may consider such data insufficient to evaluate the health effects of a motor vehicle's fuel or fuel additive emissions, unless the submitter provides additional information demonstrating that the data are reliable and adequate and EPA determines that the data are sufficient. ( 2) Definitions. As used in this section, the following terms shall have the meanings specified: Batch means a specific quantity or lot of a test fuel, additive/ base fuel mixture, or reference substance that has been characterized according to Sec. 79.60( f)( 1)( i). CAA means the Clean Air Act. Carrier means any material which is combined with engine/ motor vehicle emissions or a reference substance for administration to a test system.    Carrier'' includes, but is not limited to, clean, filtered air, water, feed, and nutrient media. Control atmosphere means clean, filtered air which is administered to the test system in the course of a study for the purpose of establishing a basis for comparison with the test atmosphere for chemical or biological measurements. Experimental start date means the first date the test atmosphere is applied to the test system. Experimental termination date means the last date on which data are collected directly from the study. Person includes an individual, partnership, corporation, association, scientific or academic establishment, government agency, or organizational unit thereof, and any other legal entity. Quality assurance unit means any person or organizational element, except the study director, designated by testing facility management to perform the duties relating to quality assurance of the studies. Raw data means any laboratory worksheets, records, memoranda, notes, or exact copies thereof, that are the result of original observations and activities of a study and are necessary for the reconstruction and evaluation of the report of that study. In the event that exact transcripts of raw data have been prepared ( e. g., tapes which have been transcribed verbatim, dated, and verified accurate by signature), the exact copy or exact transcript may be substituted for the original source as raw data.    Raw data'' may include photographs, videotape, microfilm or microfiche copies, computer printouts, magnetic media, including dictated observations, and recorded data from automated instruments. Reference substance means any chemical substance or mixture, analytical standard, or material other than engine/ motor vehicle emissions and/ or its carrier, that is administered to or used in analyzing the test system in the course of a study. A    reference substance'' is used to establish a basis for comparison with the test atmosphere for known chemical or biological measurements, i. e., positive or negative control substance. Specimen means any material derived from a test system for examination or analysis. Sponsor means person who initiates and supports, by provision of financial or other resources, a study or a person who submits a study to EPA in response to the CAA Section 211( b) or 211( e) Fuels and Fuel Additives Registration Rule or a testing facility, if it both initiates and actually conducts the study. Study means any experiment, at one or more test sites, in which a test system is exposed to a test atmosphere under laboratory conditions to determine or help predict the health effects of that exposure in humans, other living organisms, or media. Study completion date means the date the final report is signed by the study director. Study director means the individual responsible for the overall conduct of a study. Study initiation date means the date the protocol is signed by the study director. Test substance means a vapor and/ or aerosol mixture composed of engine/ motor vehicle emissions and clean, filtered air which is administered directly, or indirectly, by the inhalation route to a test system in a study which develops data to meet the registration requirements of CAA section 211( b) or ( e). Test system means any animal, microorganism, chemical or physical matrix, to which the test, control, or reference substance is administered or added for study. This definition also includes appropriate groups or components of the system not treated with the test, control, or reference substance. Testing facility means a person who actually conducts a study, i. e., actually uses the test substance in a test system.    Testing facility'' encompasses only those operational units that are being or have been used to conduct studies. TSCA means the Toxic Substances Control Act ( 15 U. S. C. 2601 et seq.). ( 3) Applicability to studies performed under grants and contracts. When a sponsor or other person utilizes the services of a consulting laboratory, contractor, or grantee to perform all or a part of a study to which this section applies, it shall notify the consulting laboratory, contractor, or grantee that the service is, or is part of, a study that must be conducted in compliance with the provisions of this section. ( 4) Statement of compliance or non compliance. Any person who submits to EPA a test in compliance with registration regulations issued under CAA section 211( b) or section 211( e) shall include in the submission a true and correct statement, signed by the sponsor and the study director, of one of the following types: ( i) A statement that the study was conducted in accordance with this section; or ( ii) A statement describing in detail all differences between the practices used in the study and those required by this section; or ( iii) A statement that the person was not a sponsor of the study, did not conduct the study, and does not know whether the study was conducted in accordance with this section. ( 5) Inspection of a testing facility. ( i) A testing facility shall permit an authorized employee or duly designated representative of EPA, at reasonable times and in a reasonable manner, to inspect the facility and to inspect ( and in the case of records also to copy) all records and specimens required to be maintained regarding studies to which this section applies. The records inspection and copying requirements shall not apply to quality assurance unit records of findings and problems, or to actions recommended and taken, except the EPA may seek production of these records in litigation or formal adjudicatory hearings. ( ii) EPA will not consider reliable for purposes of showing that a test substance does or does not present a risk of injury to health or the environment any data developed by a testing facility or sponsor that refuses to permit inspection in accordance with this section. The determination that a study will not be considered reliable does not, however, relieve the sponsor of a required test of any obligation under any applicable statute or regulation to submit the results of the study to EPA. ( 6) Effects of non compliance. ( i) Pursuant to sections 114, 208, and 211( d) of the CAA, it shall be a violation of this section and a violation of this rule ( 40 CFR part 79, subpart F) if: ( A) The test is not being or was not conducted in accordance with any requirement of this part; or ( B) Data or information submitted to EPA under part 79, including the statement required by Sec. 79.60( a)( 4), include information or data that are false or misleading, contain significant omissions, or otherwise do not fulfill the requirements of this part; or ( C) Entry in accordance with Sec. 79.60( a)( 5) for the purpose of auditing test data is denied. ( ii) EPA, at its discretion, may not consider reliable for purposes of showing that a chemical substance or mixture does not present a risk of injury to health any study which was not conducted in accordance with this part. EPA, at its discretion, may rely upon such studies for purposes of showing adverse effects. The determination that a study will not be considered reliable does not, however, relieve the sponsor of a required test of the obligation under any applicable statute or regulation to submit the results of the study to EPA. ( iii) If data submitted in compliance with registration regulations issued under CAA section 211( b) or section 211( e) are not developed in accordance with this section, EPA may determine that the sponsor has not fulfilled its obligations under 40 CFR part 79 and may require the sponsor to develop data in accordance with the requirements of this section in order to satisfy such obligations. ( b) Organization and Personnel. ( 1) Personnel. ( i) Each individual engaged in the conduct of or responsible for the supervision of a study shall have education, training, and experience, or combination thereof, to enable that individual to perform the assigned functions. ( ii) Each testing facility shall maintain a current summary of training and experience and job description for each individual engaged in or supervising the conduct of a study. ( iii) There shall be a sufficient number of personnel for the timely and proper conduct of the study according to the protocol. ( iv) Personnel shall take necessary personal sanitation and health precautions designed to avoid contamination of test fuel and additive/ base fuel mixtures, test and reference substances, and test systems. ( v) Personnel engaged in a study shall wear clothing appropriate for the duties they perform. Such clothing shall be changed as often as necessary to prevent microbiological, radiological, or chemical contamination of test systems and test, control, and reference substances. ( vi) Any individual found at any time to have an illness that may adversely affect the quality and integrity of the study shall be excluded from direct contact with test systems, fuel and fuel/ additive mixtures, test and reference substances and any other operation or function that may adversely affect the study until the condition is corrected. All personnel shall be instructed to report to their immediate supervisors any health or medical conditions that may reasonably be considered to have an adverse effect on a study. ( 2) Testing facility management. For each study, testing facility management shall: ( i) Designate a study director as described in Sec. 79.60( b)( 3) before the study is initiated. ( ii) Replace the study director promptly if it becomes necessary to do so during the conduct of a study. ( iii) Assure that there is a quality assurance unit as described in Sec. 79.60( b)( 4). ( iv) Assure that test fuels and fuel/ additive mixtures and test and reference substances have been identified as to content, strength, purity, stability, and uniformity, as applicable. ( v) Assure that personnel, resources, facilities, equipment, materials and methodologies are available as scheduled. ( vi) Assure that personnel clearly understand the functions they are to perform. ( vii) Assure that any deviations from these regulations reported by the quality assurance unit are communicated to the study director and corrective actions are taken and documented. ( 3) Study director. For each study, a scientist or other professional person with a doctorate degree or equivalent in toxicology or other appropriate discipline shall be identified as the study director. The study director has overall responsibility for the technical conduct of the study, as well as for the interpretation, analysis, documentation, and reporting of results, and represents the single point of study control. The study director shall assure that: ( i) The protocol, including any changes, is approved as provided by Sec. 79.60( g)( 1)( i) and is followed; ( ii) All experimental data, including observations of unanticipated responses of the test system are accurately recorded and verified; ( iii) Unforeseen circumstances that may affect the quality and integrity of the study are noted when they occur, and corrective action is taken and documented; ( iv) Test systems are as specified in the protocol; ( v) All applicable good laboratory practice regulations are followed; and ( vi) All raw data, documentation, protocols, specimens, and final reports are archived properly during or at the close of the study. ( 4) Quality assurance unit. A testing facility shall have a quality assurance unit which shall be responsible for monitoring each study to assure management that the facilities, equipment, personnel, methods, practices, records, and controls are in conformance with the regulations in this section. For any given study, the quality assurance unit shall be entirely separate from and independent of the personnel engaged in the direction and conduct of that study. The quality assurance unit shall conduct inspections and maintain records appropriate to the study. ( i) Quality assurance unit duties. ( A) Maintain a copy of a master schedule sheet of all studies conducted at the testing facility indexed by test substance and containing the test system, nature of study, date study was initiated, current status of each study, identity of the sponsor, and name of the study director. ( B) Maintain copies of all protocols pertaining to all studies for which the unit is responsible. ( C) Inspect each study at intervals adequate to ensure the integrity of the study and maintain written and properly signed records of each periodic inspection showing the date of the inspection, the study inspected, the phase or segment of the study inspected, the person performing the inspection, findings and problems, action recommended and taken to resolve existing problems, and any scheduled date for re inspection. Any problems which are likely to affect study integrity found during the course of an inspection shall be brought to the attention of the study director and management immediately. ( D) Periodically submit to management and the study director written status reports on each study, noting any problems and the corrective actions taken. ( E) Determine that no deviations from approved protocols or standard operating procedures were made without proper authorization and documentation. ( F) Review the final study report to assure that such report accurately describes the methods and standard operating procedures, and that the reported results accurately reflect the raw data of the study. ( G) Prepare and sign a statement to be included with the final study report which shall specify the dates inspections were made and findings reported to management and to the study director. ( ii) The responsibilities and procedures applicable to the quality assurance unit, the records maintained by the quality assurance unit, and the method of indexing such records shall be in writing and shall be maintained. These items including inspection dates, the study inspected, the phase or segment of the study inspected, and the name of the individual performing the inspection shall be made available for inspection to authorized employees or duly designated representatives of EPA. ( iii) An authorized employee or a duly designated representative of EPA shall have access to the written procedures established for the inspection and may request test facility management to certify that inspections are being implemented, performed, documented, and followed up in accordance with this paragraph. ( c) Facilities( 1) General. Each testing facility shall be of suitable size and construction to facilitate the proper conduct of studies. Testing facilities which are not completely located within an indoor controlled environment shall be of suitable location/ proximity to facilitate the proper conduct of studies. Testing facilities shall be designed so that there is a degree of separation that will prevent any function or activity from having an adverse effect on the study. ( 2) Test system care facilities. ( i) A testing facility shall have a sufficient number of animal rooms or other test system areas, as needed, to ensure proper separation of species or test systems, quarantine or isolation of animals or other test systems, and routine or specialized housing of animals or other test systems. ( ii) A testing facility shall have a number of animal rooms or other test system areas separate from those described in paragraph ( a) of this section to ensure isolation of studies being done with test systems or test, control, and reference substances known to be biohazardous, including volatile atmospheres and aerosols, radioactive materials, and infectious agents. The animal handling facility must operate under the supervision of a veterinarian. ( iii) Separate areas shall be provided, as appropriate, for the diagnosis, treatment, and control of laboratory test system diseases. These areas shall provide effective isolation for the housing of test systems either known or suspected of being diseased, or of being carriers of disease, from other test systems. ( iv) Facilities shall have proper provisions for collection and disposal of contaminated air, water, or other spent materials. When animals are housed, facilities shall exist for the collection and disposal of all animal waste and refuse or for safe sanitary storage of waste before removal from the testing facility. Disposal facilities shall be so provided and operated as to minimize vermin infestation, odors, disease hazards, and environmental contamination. ( v) Facilities shall have provisions to regulate environmental conditions ( e. g., temperature, humidity, day length, etc.) as specified in the protocol. ( 3) Test system supply/ operation areas. ( i) There shall be storage areas, as needed, for feed, bedding, supplies, and equipment. Storage areas for feed and bedding shall be separated from areas where the test systems are located and shall be protected against infestation or contamination. Perishable supplies shall be preserved by appropriate means. ( ii) Separate laboratory space and other space shall be provided, as needed, for the performance of the routine and specialized procedures required by studies. ( 4) Facilities for handling test fuels and fuel/ additive mixtures and reference substances. ( i) As necessary to prevent contamination or mixups, there shall be separate areas for: ( A) Receipt and storage of the test fuels and fuel/ additive mixtures and reference substances; ( B) Mixing of the test fuels, fuel/ additive mixtures, and reference substances with a carrier, i. e., liquid hydrocarbon; and ( C) Storage of the test fuels, fuel/ additive mixtures, and reference substance/ carrier mixtures. ( ii) Storage areas for test fuels and fuel/ additive mixtures and reference substances and for reference mixtures shall be separate from areas housing the test systems and shall be adequate to preserve the identity, strength, purity, and stability of the substances and mixtures. ( 5) Specimen and data storage facilities. Space shall be secured for archives for the storage and retrieval of all raw data and specimens from completed studies. ( d) Equipment( 1) Equipment design. Equipment used in the generation, measurement, or assessment of data and equipment used for facility environmental control shall be of appropriate design and adequate capacity to function according to the protocol and shall be suitably located for operation, inspection, cleaning, and maintenance. ( 2) Maintenance and calibration of equipment. ( i) Equipment shall be adequately inspected, cleaned, and maintained. Equipment used for the generation, measurement, or assessment of data shall be adequately tested, calibrated, and/ or standardized. ( ii) The written standard operating procedures required under Sec. 79.60( e)( 1)( ii)( K) shall set forth in sufficient detail the methods, materials, and schedules to be used in the routine inspection, cleaning, maintenance, testing, calibration, and/ or standardization of equipment, and shall specify, when appropriate, remedial action to be taken in the event of failure or malfunction of equipment. The written standard operating procedures shall designate the person responsible for the performance of each operation. ( iii) Written records shall be maintained of all inspection, maintenance, testing, calibrating, and/ or standardizing operations. These records, containing the date of the operation, shall describe whether the maintenance operations were routine and followed the written standard operating procedures. Written records shall be kept of non routine repairs performed on equipment as a result of failure and malfunction. Such records shall document the nature of the defect, how and when the defect was discovered, and any remedial action taken in response to the defect. ( e) Testing Facilities Operation( 1) Standard operating procedures. ( i) A testing facility shall have standard operating procedures in writing, setting forth study methods that management is satisfied are adequate to insure the quality and integrity of the data generated in the course of a study. All deviations in a study from standard operating procedures shall be authorized by the study director and shall be documented in the raw data. Significant changes in established standard operating procedures shall be properly authorized in writing by management. ( ii) Standard operating procedures shall be established for, but not limited to, the following: ( A) Test system room preparation; ( B) Test system care; ( C) Receipt, identification, storage, handling, mixing, and method of sampling of test fuels and fuel/ additive mixtures and reference substances; ( D) Test system observations; ( E) Laboratory or other tests; ( F) Handling of test animals found moribund or dead during study; ( G) Necropsy or postmortem examination of test animals; ( H) Collection and identification of specimens; ( I) Histopathology ( J) Data handling, storage and retrieval. ( K) Maintenance and calibration of equipment. ( L) Transfer, proper placement, and identification of test systems. ( iii) Each laboratory or other study area shall have immediately available manuals and standard operating procedures relative to the laboratory procedures being performed. Published literature may be used as a supplement to standard operating procedures. ( iv) A historical file of standard operating procedures, and all revisions thereof, including the dates of such revisions, shall be maintained. ( 2) Reagents and solutions. All reagents and solutions in the laboratory areas shall be labeled to indicate identity, titer or concentration, storage requirements, and expiration date. Deteriorated or outdated reagents and solutions shall not be used. ( 3) Animal and other test system care. ( i) There shall be standard operating procedures for the housing, feeding, handling, and care of animals and other test systems. ( ii) All newly received test systems from outside sources shall be isolated and their health status or appropriateness for the study shall be evaluated. This evaluation shall be in accordance with acceptable veterinary medical practice or scientific methods. ( iii) At the initiation of a study, test systems shall be free of any disease or condition that might interfere with the purpose or conduct of the study. If during the course of the study, the test systems contract such a disease or condition, the diseased test systems shall be isolated, if necessary. These test systems may be treated for disease or signs of disease provided that such treatment does not interfere with the study. The diagnosis, authorization of treatment, description of treatment, and each date of treatment shall be documented and shall be retained. ( iv) When laboratory procedures require test animals to be manipulated and observed over an extended period of time or when studies require test animals to be removed from and returned to their housing units for any reason ( e. g., cage cleaning, treatment, etc.), these test systems shall receive appropriate identification ( e. g., tattoo, color code, etc.). Test system identification shall conform with current laboratory animal handling practice. All information needed to specifically identify each test system within the test system housing unit shall appear on the outside of that unit. Suckling animals are excluded from the requirement of individual identification unless otherwise specified in the protocol. ( v) Except as specified in paragraph ( e)( 3)( v)( A) of this section, test animals of different species shall be housed in separate rooms when necessary. Test animals of the same species, but used in different studies, shall not ordinarily be housed in the same room when inadvertent exposure to the test or reference substances or test system mixup could affect the outcome of either study. If such mixed housing is necessary, adequate differentiation by space and identification shall be made. ( A) Test systems that may be used in multispecies tests need not be housed in separate rooms, provided that they are adequately segregated to avoid mixup and cross contamination. ( B) [ reserved] ( vi) Cages, racks, pens, enclosures, and other holding, rearing, and breeding areas, and accessory equipment, shall be cleaned and sanitized at appropriate intervals. ( vii) Feed and water used for the test animals shall be analyzed periodically to ensure that contaminants known to be capable of interfering with the study and reasonably expected to be present in such feed or water are not present at greater than trace levels. Documentation of such analyses shall be maintained as raw data. ( viii) Bedding used in animal cages or pens shall not interfere with the purpose or conduct of the study and shall be changed as often as necessary to keep the animals dry and clean. ( ix) If any pest control materials are used, the use shall be documented. Cleaning and pest control materials that interfere with the study shall not be used. ( x) All test systems shall be acclimatized to the environmental conditions of the test, prior to their use in a study. ( f) Test fuels, additive/ base fuel mixtures, and reference substances( 1) Test fuel, fuel/ additive mixture, and reference substance identity ( i) The product brand name/ service mark, strength, purity, content, or other characteristics which appropriately define the test fuel, fuel/ additive mixture, or reference substance shall be reported for each batch and shall be documented before its use in a study. Methods of synthesis, fabrication, or derivation, as appropriate, of the test fuel, fuel/ additive mixture, or reference substance shall be documented by the sponsor or the testing facility, and such location of documentation shall be specified. ( ii) The stability of test fuel, fuel/ additive mixture, and reference substances under storage conditions at the test site shall be known for all studies. ( 2) Test fuel, additive/ base fuel mixture, and reference substance handling. Procedures shall be established for a system for the handling of the test fuel, fuel/ additive mixture, and reference substance( s) to ensure that: ( i) There is proper storage. ( ii) Distribution is made in a manner designed to preclude the possibility of contamination, deterioration, or damage. ( iii) Proper identification is maintained throughout the distribution process. ( iv) The receipt and distribution of each batch is documented. Such documentation shall include the date and quantity of each batch distributed or returned. ( 3) Mixtures of test emissions or reference solutions with carriers. ( i) For test emissions or each reference substance mixed with a carrier, tests by appropriate analytical methods shall be conducted: ( A) To determine the uniformity of the test substance and to determine, periodically, the concentration of the test emissions or reference substance in the mixture; ( B) When relevant to the conduct of the experiment, to determine the solubility of each reference substance in the carrier mixture before the experimental start date; and ( C) To determine the stability of test emissions or a reference solution in the test substance before the experimental start date or concomitantly according to written standard operating procedures, which provide for periodic analysis of each batch. ( ii) Where any of the components of the reference substance/ carrier mixture has an expiration date, that date shall be clearly shown on the container. If more than one component has an expiration date, the earliest date shall be shown. ( iii) If a chemical or physical agent is used to facilitate the mixing of a test substance with a carrier, assurance shall be provided that the agent does not interfere with the integrity of the test. ( g) Protocol for and conduct of a study( 1) Protocol. ( i) Each study shall have a written protocol that clearly indicates the objectives and all methods for the conduct of the study. The protocol shall contain but shall not be limited to the following information: ( A) A descriptive title and statement of the purpose of the study. ( B) Identification of the test fuel, fuel/ additive mixture, and reference substance by name, chemical abstracts service ( CAS) number or code number, as applicable. ( C) The name and address of the sponsor and the name and address of the testing facility at which the study is being conducted. ( D) The proposed experimental start and termination dates. ( E) Justification for selection of the test system, as necessary. ( F) Where applicable, the number, body weight, sex, source of supply, species, strain, substrain, and age of the test system. ( G) The procedure for identification of the test system. ( H) A description of the experimental design, including methods for the control of bias. ( I) Where applicable, a description and/ or identification of the diet used in the study. The description shall include specifications for acceptable levels of contaminants that are reasonably expected to be present in the dietary materials and are known to be capable of interfering with the purpose or conduct of the study if present at levels greater than established by the specifications. ( J) Each concentration level, expressed in milligrams per cubic meter of air or other appropriate units, of the test or reference substance to be administered and the frequency of administration. ( K) The type and frequency of tests, analyses, and measurements to be made. ( L) The records to be maintained. ( M) The date of approval of the protocol by the sponsor and the dated signature of the study director. ( N) A statement of the proposed statistical method. ( ii) All changes in or revisions of an approved protocol and the reasons therefor shall be documented, signed by the study director, dated, and maintained with the protocol. ( 2) Conduct of a study. ( i) The study shall be conducted in accordance with the protocol. ( ii) The test systems shall be monitored in conformity with the protocol. ( iii) Specimens shall be identified by test system, study, nature, and date of collection. This information shall be located on the specimen container or shall accompany the specimen in a manner that precludes error in the recording and storage of data. ( iv) In animal studies where histopathology is required, records of gross findings for a specimen from postmortem observations shall be available to a pathologist when examining that specimen histopathologically. ( v) All data generated during the conduct of a study, except those that are generated by automated data collection systems, shall be recorded directly, promptly, and legibly in ink. All data entries shall be dated on the day of entry and signed or initialed by the person entering the data. Any change in entries shall be made so as not to obscure the original entry, shall indicate the reason for such change, and shall be dated and signed or identified at the time of the change. In automated data collection systems, the individual responsible for direct data input shall be identified at the time of data input. Any change in automated data entries shall be made so as not to obscure the original entry, shall indicate the reason for change, shall be dated, and the responsible individual shall be identified. ( h) Records and Reports( 1) Reporting of study results. ( i) A final report shall be prepared for each study and shall include, but not necessarily be limited to, the following: ( A) Name and address of the facility performing the study and the dates on which the study was initiated and was completed, terminated, or discontinued. ( B) Objectives and procedures stated in the approved protocol, including any changes in the original protocol. ( C) Statistical methods employed for analyzing the data. ( D) The test fuel, additive/ base fuel mixture, and test and reference substances identified by name, chemical abstracts service ( CAS) number or code number, strength, purity, content, or other appropriate characteristics. ( E) Stability, and when relevant to the conduct of the study, the solubility of the test emissions and reference substances under the conditions of administration. ( F) A description of the methods used. ( G) A description of the test system used. Where applicable, the final report shall include the number of animals or other test organisms used, sex, body weight range, source of supply, species, strain and substrain, age, and procedure used for identification. ( H) A description of the concentration regimen as daily exposure period, i. e., number of hours, and exposure duration, i. e., number of days. ( I) A description of all circumstances that may have affected the quality or integrity of the data. ( J) The name of the study director, the names of other scientists or professionals and the names of all supervisory personnel, involved in the study. ( K) A description of the transformations, calculations, or operations performed on the data, a summary and analysis of the data, and a statement of the conclusions drawn from the analysis. ( L) The signed and dated reports of each of the individual scientists or other professionals involved in the study, including each person who, at the request or direction of the testing facility or sponsor, conducted an analysis or evaluation of data or specimens from the study after data generation was completed. ( M) The locations where all specimens, raw data, and the final report are to be kept or stored. ( N) The statement, prepared and signed by the quality assurance unit, as described in Sec. 79.60( b)( 4)( i)( G). ( ii) The final report shall be signed and dated by the study director. ( iii) Corrections or additions to a final report shall be in the form of an amendment by the study director. The amendment shall clearly identify that part of the final report that is being added to or corrected and the reasons for the correction or addition, and shall be signed and dated by the person responsible. Modification of a final report to comply with the submission requirements of EPA does not constitute a correction, addition, or amendment to a final report. ( iv) A copy of the final report and of any amendment to it shall be maintained by the sponsor and the test facility. ( 2) Storage and retrieval of records and data. ( i) All raw data, documentation, records, protocols, specimens, and final reports generated as a result of a study shall be retained. Specimens obtained from mutagenicity tests, wet specimens of blood, urine, feces, and biological fluids, do not need to be retained after quality assurance verification. Correspondence and other documents relating to interpretation and evaluation of data, other than those documents contained in the final report, also shall be retained. ( ii) All raw data, documentation, protocols, specimens, and interim and final reports shall be archived for orderly storage and expedient retrieval. Conditions of storage shall minimize deterioration of the documents or specimens in accordance with the requirements for the time period of their retention and the nature of the documents of specimens. A testing facility may contract with commercial archives to provide a repository for all material to be retained. Raw data and specimens may be retained elsewhere provided that the archives have specific reference to those other locations. ( iii) An individual shall be identified as responsible for the archiving of records. ( iv) Access to archived material shall require authorization and documentation. ( v) Archived material shall be indexed to permit expedient retrieval. ( 3) Retention of records. ( i) Record retention requirements set forth in this section do not supersede the record retention requirements of any other regulations in this subchapter. ( ii) Except as provided in paragraph ( h)( 3)( iii) of this section, documentation records, raw data, and specimens pertaining to a study and required to be retained by this part shall be archived for a period of at least ten years following the completion of the study. ( iii) Wet specimens, samples of test fuel, additive/ base fuel mixtures, or reference substances, and specially prepared material which are relatively fragile and differ markedly in stability and quality during storage, shall be retained only as long as the quality of the preparation affords evaluation. Specimens obtained from mutagenicity tests, wet specimens of blood, urine, feces, biological fluids, do not need to be retained after quality assurance verification. In no case shall retention be required for a longer period than that set forth in paragraph ( h)( 3)( ii) of this section. ( iv) The master schedule sheet, copies of protocols, and records of quality assurance inspections, as required by Sec. 79.60( b)( 4)( iii) shall be maintained by the quality assurance unit as an easily accessible system of records for the period of time specified in paragraph ( h)( 3)( ii) of this section. ( v) Summaries of training and experience and job descriptions required to be maintained by Sec. 79.60( b)( 1)( ii) may be retained along with all other testing facility employment records for the length of time specified in paragraph ( h)( 3)( ii) of this section. ( vi) Records and reports of the maintenance and calibration and inspection of equipment, as required by Sec. 79.60( d)( 2) ( ii) and ( iii), shall be retained for the length of time specified in paragraph ( h)( 3)( ii) of this section. ( vii) If a facility conducting testing or an archive contracting facility goes out of business, all raw data, documentation, and other material specified in this section shall be transferred to the sponsor of the study for archival. ( viii) Records required by this section may be retained either as original records or as true copies such as photocopies, microfilm, microfiche, or other accurate reproductions of the original records.

Sec. 79.61. Vehicle emissions inhalation exposure guideline.

( a) Purpose. This guideline provides additional information on methodologies required to conduct health effects tests involving inhalation exposures to vehicle combustion emissions from fuels or fuel/ additive mixtures. Where this guideline and the other health effects testing guidelines in 40 CFR 79.62 through 79.68 specify differing values for the same test parameter, the specifications in the individual health test guideline shall prevail for that health effect endpoint. ( b) Definitions. For the purposes of this section the following definitions apply. Acute inhalation study means a short term toxicity test characterized by a single exposure by inhalation over a short period of time ( at least 4 hours and less than 24 hours), followed by at least 14 days of observation. Aerodynamic diameter means the diameter of a sphere of unit density that has the same settling velocity as the particle of the test substance. It is used to compare particles of different sizes, densities and shapes, and to predict where in the respiratory tract such particles may be deposited. It applies to the size of aerosol particles. Chronic inhalation study means a prolonged and repeated exposure by inhalation for the life span of the test animal; technically, two years in the rat. Concentration means an exposure level. Exposure is expressed as weight or volume of test aerosol/ substance per volume of air, usually mg/ m< SUP> 3 or as parts per million ( ppm) over a given time period. Micrograms per cubic meter (< greek m> g/ m< SUP> 3) or parts per billion may be appropriate, as well. Cumulative toxicity means the adverse effects of repeated exposures occurring as a result of prolonged action or increased concentration of the administered test substance or its metabolites in the susceptible tissues. Inhalable diameter means that aerodynamic diameter of a particle which is considered to be inhalable for the organism. It is used to refer to particles which are capable of being inhaled and may be deposited anywhere within the respiratory tract from the trachea to the alveoli. Mass median aerodynamic diameter ( MMAD) means the calculated aerodynamic diameter, which divides the particles of an aerosol in half based on the mass of the particles. Fifty percent of the particles in mass will be larger than the median diameter, and fifty percent will be smaller than the median diameter. MMAD describes the particle distribution of any aerosol based on the weight and size of the particles. MMAD and the geometric standard deviation describe the particle size distribution. Material safety data sheet ( MSDS) means documentation or information on the physical, chemical, and hazardous characteristics of a given chemical, usually provided by the product's manufacturer. Reynolds number means a dimensionless number that is proportional to the ratio of inertial forces to frictional forces acting on a fluid. It quantitatively provides a measure of whether flow is laminar or turbulent. A fluid traveling through a pipe is fully developed into a laminar flow for a Reynolds number less than 2000, and fully developed into a turbulent flow for a Reynolds number greater than 4000. Subacute inhalation toxicity means the adverse effects occurring as a result of the repeated daily exposure of experimental animals to a chemical by inhalation for part ( less than 10 percent) of a lifespan; generally, less than 90 days. Subchronic inhalation study means a repeated exposure by inhalation for part ( approximately 10 percent) of a life span of the exposed test animal. Toxic effect means an adverse change in the structure or function of an experimental animal as a result of exposure to a chemical substance. ( c) Principles and design criteria of inhalation exposure systems. Proper conduct of inhalation toxicity studies of the emissions of fuels and additive/ fuel mixtures requires that the exposure system be designed to ensure the controlled generation of the exposure atmosphere, the adequate dilution of the test emissions, delivery of the diluted exposure atmosphere to the test animals, and use of appropriate exposure chamber systems selected to meet criteria for a given exposure study. ( 1) Emissions generation. Emissions shall be generated according to the specifications in 40 CFR 79.57. ( 2) Dilution and delivery systems. ( i) The delivery system is the means used to transport the emissions from the generation system to the exposure system. The dilution system is generally a component of the delivery system. ( ii) Dilution provides control of the emissions concentration delivered to the exposure system, serving the function of diluting the associated combustion gases, such as carbon monoxide, carbon dioxide, nitrogen oxides, sulfur dioxide and other noxious gases and vapors, to levels that will ensure that there are no significant or measurable responses in the test animals as a result of exposure to the combustion gases. The formation of particle species is strongly dependent on the dilution rate, as well. ( iii) The engine exhaust system shall connect to the first stagedilution section at 90 deg. to the axis of the dilution section. This is then connected to a right angle elbow on the center line of the dilution section. Engine emissions are injected through the elbow so that exhaust flow is concurrent to dilution flow. ( iv) Materials. In designing the dilution and delivery systems, the use of plastic, e. g., PVC and similar materials, copper, brass, and aluminum pipe and tubing shall be avoided if there exists a possibility of chemical reaction occurring between emissions and tubing. Stainless steel pipe and tubing is recommended as the best choice for most emission dilution and delivery applications, although glass and teflon may be appropriate, as well. ( v) Flow requirements. ( A) Conduit for dilute raw emissions shall be of such dimensions as to provide residence times for the emissions on the order of less than one second to several seconds before the emissions are further diluted and introduced to the test chambers. With the high flow rates in the dilute raw emissions conduit, it will be necessary to sample various portions of the dilute emissions for delivering differing concentrations to the test chambers. The unused portions of the emissions stream are normally exhausted to the atmosphere outside of the exposure facility. ( B) Dimensions of the dilute raw exhaust conduit shall be such that, at a minimum, the flow Reynolds number is 70,000 or greater ( see Mokler, et al., 1984 in paragraph ( f)( 13) of this section). This will maintain highly turbulent flow conditions so that there is more complete mixing of the exhaust emissions. ( C) Wall losses. The delivery system shall be designed to minimize wall losses. This can be done by sizing the tubing or pipe to maintain laminar flow of the diluted emissions to the exposure chamber. A flow Reynolds number of 1000 3000 will ensure minimal wall losses. Also, the length of and number and degree of bends in the delivery lines to the exposure chamber system shall be minimized. ( D) Whole body exposure vs. nose only exposure delivery systems. Flow rates through whole body chamber systems are of the order of 100 liters per minute to 500 liters per minute. Nose only systems are on the order of less than 50 liters per minute. To maintain laminar flow conditions, the principles described in paragraph ( c)( 2)( v)( C) of this section apply to both systems. ( vi) Dilution requirements. ( A) To maintain the water vapor, and dissolved organic compounds, in the raw exhaust emissions stream, a manufacturer/ tester will initially dilute one part emissions with a minimum of five parts clean, filtered air ( see Hinners, et al., 1979 in paragraph ( f)( 11) of this section). Depending on the water vapor content of a particular fuel/ additive mixture's combustion emissions and the humidity of the dilution air, initial exhaust dilutions as high as 1: 15 or 1: 20 may be necessary to maintain the general character of the exhaust as it cools, e. g., M100. At this point, it is expected that the exhaust stream would be further diluted to more appropriate levels for rodent health effects testing. ( B) A maximum concentration ( minimum dilution) of the raw exhaust going into the test animal cages is anticipated to lie in the range between 1: 5 and 1: 50 exhaust emissions to clean, filtered air. The minimum concentration ( maximum dilution) of raw exhaust for health effects testing is anticipated to be in range between 1: 100 and 1: 150. Individual manufacturers will treat these ranges as approximations only and will determine the optimum range of emission concentrations to elicit effects in Tier 2 health testing for their particular fuel/ fuel additive mixture. ( 3) Exposure chamber systems( i) Referenced Guidelines. ( A) The U. S. Department of Health and Human Services    Guide for the Care and Use of Laboratory Animals'' ( Guide), 1985 cited in paragraph ( c)( 3)( ii)( A)( 4), and in paragraphs ( d)( 2)( i), ( d)( 2)( ii), ( d)( 2)( iii), ( d)( 4)( ii), and ( d)( 4)( iii) of this section, has been incorporated by reference. ( B) This incorporation by reference was approved by the Director of the Federal Register in accordance with 5 U. S. C. 552( a) and 1 CFR part 51. Copies may be purchased from the Superintendent of Documents, U. S. Government Printing Office, Washington, DC 20402. Copies may be inspected at U. S. EPA, OAR, 401 M Street SW., Washington, DC, 20460 or at the Office of the Federal Register, 800 North Capitol Street NW., suite 700, Washington, DC. ( ii) Exposure chambers. There are two basic types of dynamic inhalation exposure chambers, whole body chambers and nose/ head only exposure chambers ( see Cheng and Moss, 1989 in paragraph ( f)( 8) of this section). ( A) Whole body chambers. ( 1) The flow rate through a chamber shall be maintained at 15 air changes per hour. ( 2) The chambers are usually maintained at a slightly negative pressure ( 0.5 to 1.5 inch of water) to prevent leakage of test substance into the exposure room. ( 3) The exposure chamber shall be designed in such a way as to provide uniform distribution of exposure concentrations in all compartments ( see Cheng et al., 1989 in paragraph ( f)( 7) of this section). ( 4) Animals are housed in separate compartments inside the chamber, where the whole surface area of an animal is exposed to the test material. The spaces required for different animal species shall follow the Guide. In general, the volume of animal bodies occupy less than 5 percent of the chamber volume. ( B) Head/ nose only exposure chambers. ( 1) In head/ nose only exposure chambers, only the head ( oronasal) portion of the animal is exposed to the test material. ( 2) The chamber volume and flow rates are much less than in the whole body exposure chambers because the subjects are usually restrained in a tube holder where the animal's breathing can be easily monitored. The head/ nose only exposure chamber is suitable for shortterm exposures or when use of a small amount of test material is required. ( iii) Since whole body exposure appears to be the least stressful mode of exposure, it is the preferred method. In general, head/ nose only exposure, which is sometimes used to avoid concurrent exposure by the dermal or oral routes, i. e., grooming, is not recommended because of the stress accompanying the restraining of the animals. However, there may be specific instances where it may be more appropriate than whole body exposure. The tester shall provide justification for its selection. ( d) Inhalation exposure procedures( 1) Animal selection. ( i) The rat is the preferred species for vehicle emission inhalation health effects testing. Commonly used laboratory strains shall be used. Any rodent species may be used, but the tester shall provide justification for the choice of that species. ( ii) Young adult animals, approximately ten weeks of age for the rat, shall be used. At the commencement of the study, the weight variation of animals used shall not exceed < plus minus> 20 percent of the mean weight for each sex. Animals shall be randomly assigned to treatment and control groups according to their weight. ( iii) An equal number of male and female rodents shall be used at each concentration level. Situations may arise where use of a single sex may be appropriate. Females, in general, shall be nulliparous and nonpregnant. ( iv) The number of animals used at each concentration level and in the control group( s) depends on the type of study, number of biological end points used in the toxicity evaluation, the pre determined sensitivity of detection and power of significance of the study, and the animal species. For an acute study, at least five animals of each sex shall be used in each test group. For both the subacute and subchronic studies, at least 10 rodents of each sex shall be used in each test group. For a chronic study, at least 20 male and 20 female rodents shall be used in each test group. ( A) If interim sacrifices are planned, the number of animals shall be increased by the number of animals scheduled to be sacrificed during the course of the study. ( B) For a chronic study, the number of animals at the termination of the study must be adequate for a meaningful and valid statistical evaluation of chronic effects. ( v) A concurrent control group is required. This group shall be exposed to clean, filtered air under conditions identical to those used for the group exposed to the test atmosphere. ( vi) The same species/ strain shall be used to make comparisons between fuel only and fuel/ additive mixture studies. If another species/ strain is used, the tester shall provide justification for its selection. ( 2) Animal handling and care. ( i) A key element in the conduct of inhalation exposure studies is the proper handling and care of the test animal population. Therefore, the exposure conditions must conform strictly with the conditions for housing and animal care and use set forth in the Guide. ( ii) In whole body exposure chambers, animals shall be housed in individual caging. The minimum cage size per animal will be in accordance with instructions set forth in the Guide. ( iii) Chambers shall be cleaned and maintained in accordance with recommendations and schedules set forth in the Guide. ( A) Observations shall be made daily with appropriate actions taken to minimize loss of animals to the study ( e. g., necropsy or refrigeration of animals found dead and isolation or sacrifice of weak or moribund animals). Exposure systems using head/ nose only exposure chambers require no special daily chamber maintenance. Chambers shall be inspected to ensure that they are clean, and that there are no obstructions in the chamber which would restrict air flow to the animals. Whole body exposure chambers will be inspected on a minimum of twice daily, once before exposures and once after exposures. ( B) Signs of toxicity shall be recorded as they are observed, including the time of onset, degree, and duration. ( C) Cage side observations shall include, but are not limited to: changes in skin, fur, eye and mucous membranes, respiratory, autonomic, and central nervous systems, somatomotor activity, and behavioral patterns. Particular attention shall be directed to observation of tremors, convulsions, salivation, diarrhea, lethargy, sleep, and coma. ( iv) Food and water will be withheld from animals for head/ noseonly exposure systems. For whole body exposure systems, water only may be provided. When the exposure generation system is not operating, food will be available ad libitum. During operation of the generation system, food will be withheld to avoid possible contamination by emissions. ( v) At the end of the study period, all survivors in the main study population shall be sacrificed. Moribund animals shall be removed and sacrificed when observed. ( 3) Concentration levels and selection. ( i) In acute and subacute toxicity tests, at least three exposure concentrations and a control group shall be used and spaced appropriately to produce test groups with a range of toxic effects and mortality rates. The data shall be sufficient to produce a concentration response curve and permit an acceptable estimation of the median lethal concentration. ( ii) In subchronic and chronic toxicity tests, testers shall use at least three different concentration levels, with a control exposure group, to determine a concentration response relationship. Concentrations shall be spaced appropriately to produce test groups with a range of toxic effects. The concentration response data may also be sufficient to determine a NOAEL, unless the result of a limit test precludes such findings. The criteria for selecting concentration levels has been published ( 40 CFR 798.2450 and 798.3260). ( A) The highest concentration shall result in toxic effects but not produce an incidence of fatalities which would prevent a meaningful evaluation of the study. ( B) The lowest concentration shall not produce toxic effects which are directly attributable to the test exposure. Where there is a useful estimation of human exposure, the lowest concentration shall exceed this. ( C) The intermediate concentration level( s) shall produce minimal observable toxic effects. If more than one intermediate concentration level is used, the concentrations shall be spaced to produce a gradation of toxic effects. ( D) In the low, intermediate, and control exposure groups, the incidence of fatalities shall be low to absent, so as not to preclude a meaningful evaluation of the results. ( 4) Exposure chamber environmental conditions. The following environmental conditions in the exposure chamber are critical to the maintenance of the test animals: flow; temperature; relative humidity; lighting; and noise. ( i) Filtered and conditioned air shall be used during exposure, to dilute the exhaust emissions, and during non exposure periods to maintain environmental conditions that are free of trace gases, dusts, and microorganisms on the test animals. Twelve to fifteen air changes per hour will be provided at all times to whole body exposure chambers. The minimum air flow rate for head/ nose only exposure chambers will be a function of the number of animals and the average minute volume of the animals: