1、Designation: E 981 04Standard Test Method forEstimating Sensory Irritancy of Airborne Chemicals1This standard is issued under the fixed designation E 981; the number immediately following the designation indicates the year oforiginal adoption or, in the case of revision, the year of last revision. A
2、 number in parentheses indicates the year of last reapproval. Asuperscript epsilon (e) indicates an editorial change since the last revision or reapproval.1. Scope1.1 This laboratory test method provides a rapid means ofdetermining sensory irritant potential of airborne chemicals ormixtures. It may
3、also be used to estimate threshold limit values(TLV) for man. However, it cannot be used to evaluate therelative obnoxiousness of odors.1.2 This test method is intended as a supplement to, not areplacement for, chronic inhalation studies used to establishallowable human tolerance levels.1.3 This sta
4、ndard does not purport to address all of thesafety concerns, if any, associated with its use. It is theresponsibility of the user of this standard to establish appro-priate safety and health practices and determine the applica-bility of regulatory limitations prior to use. Specific hazardinformation
5、 is given in Section 6.2. Summary of Test Method2.1 This test method quantitatively measures irritancy asindicated by the reflex inhibition of respiration in mice exposedto sensory irritants.2.2 Four mice are simultaneously exposed to the airbornechemical. Usually a sufficient number of groups of an
6、imals areexposed to a geometric series of concentrations so that aconcentration-response curve can be constructed. For simplepreliminary comparisons, however, a single group of fouranimals at one concentration will suffice.2.3 The mice are placed in a body plethysmograph attachedto an exposure chamb
7、er so that only the head is exposed to thetest material. The plethysmographs are connected to pressuretransducers, which sense changes created by inspiration andexpiration. The amplified signals are transmitted to a polygraphrecorder.2.4 The concentration of airborne irritant that produces a50 % dec
8、rease in respiratory rate (RD50) is determined fromthe concentration-response curve constructed from the variousdata points obtained with a series of concentrations.3. Significance and Use3.1 This test method was developed to meet the followingcriteria:3.1.1 It provides positive recognition of senso
9、ry irritants ofwidely varying potencies.3.1.2 It is sufficiently simple to permit the testing of largenumbers of materials.3.1.3 This test method is capable of generatingconcentration-response curves for purposes of compound com-parison.3.1.4 This test method has good reproducibility.3.2 This test m
10、ethod can be used for a variety of divergentpurposes, including the assessment of comparative irritancy ofcompounds or formulations and setting interim exposure levelsfor the workplace (1, 2).23.3 It has been shown that for a wide variety of chemicalsand mixtures, a perfect rank order correlation ex
11、ists betweenthe decreases in respiratory rate in mice and subjective reportsof sensory irritation in man (1, 3, 4, 5).3.4 A quantitative estimate of the sensory irritancy of a widevariety of materials can be obtained from concentration-response curves developed using this method (1, 3, 4, 6, 7, 8,9)
12、.3.5 Although this test method is intended to measure sen-sory irritation of the nasal mucosa, the cornea is innervated bythe same nerve. This animal model will, therefore, allow anestimate of the irritant potential of cosmetic ingredients orother household products to the eye, assuming that they ca
13、n beaerosolized (10).3.6 This test method is recommended for setting interimguidelines for exposure of humans to chemicals in the work-place, to assess acute sensory irritation resulting from inadvert-ent spills of household products, and to assess the comparativeirritancy of formulations or materia
14、ls intended for a variety ofuses (see Appendix X2).3.7 This test method will detect irritating effects at concen-trations far below those at which pathological changes areobserved (9).NOTE 1A good overview of the toxicological evaluation of irritantcompounds is given in Ref (8).1This test method is
15、under the jurisdiction of ASTM Committee E35 onPesticides and is the direct responsibility of Subcommittee E35.26 on Safety to Man.Current edition approved April 1, 2004. Published May 2004. Originallyapproved 1984. Last prvious edition approved in 2000 as E 981 84 (2000).2The boldface numbers in pa
16、rentheses refer to the list of references at the end ofthis standard.1Copyright ASTM International, 100 Barr Harbor Drive, PO Box C700, West Conshohocken, PA 19428-2959, United States.4. Apparatus4.1 The apparatus required to perform this test is listedbelow. The basic components for testing any typ
17、e of materialare the same. A list of suitable apparatus and suppliers is foundin Appendix X1.4.2 Plethysmograph Tubes.4.3 Exposure Chamber, constructed entirely of glass, with avolume of 2.3 L.4.4 S.T.103/60 Ground Glass Joint, that allows access to theinside of the exposure chamber.4.5 Perforated R
18、ubber Dental Dam, reinforced with electri-cal tape.4.6 Rubber Stoppers.4.7 “T” Tube, with a tube 6 cm long and the “T” 12 cm long.4.8 Vacuum Pump.4.9 Flowmeter.4.10 Absolute Filter.4.11 Sodium Carbonate-Activated Charcoal Filter.4.12 Pressure Transducer.4.13 Polygraph Recorders.4.14 Frequency-to-Vol
19、tage Converter, operating in the av-eraging mode instead of the pulse mode. See Appendix X1.7.4.15 Voltage Addition and Division Equipment, to obtain thesignal average for four mice.4.16 Signal Averages.4.17 Oscillograph.4.18 Aerosol Generator.4.19 Timer.4.20 Control Valve.5. Reagents5.1 Technical r
20、eagents may be used in all tests wheresolvents other than water are required.5.2 Solutions containing 1 to 3 % of the test material areused for comparative studies.6. Hazards6.1 Not all compounds that cause a decrease in respiratoryrate are sensory irritants. To be characterized as a sensoryirritant
21、, a compound must produce a net decrease in respiratoryrate as a result of the characteristic pause during expiration asshown in Fig. 1. This pause differentiates sensory irritants frompulmonary irritants, general anesthetics, and asphyxiants,which also reduce respiratory rate, but as a result of a
22、pausebetween breaths as shown in Fig. 2.6.2 It is possible for one component to alter the effect ofanother in a mixture, depending on their respective concentra-tions (11). Additive and antagonistic responses are possible.NOTE 1Taken from Ref. (3).FIG. 1 Typical Tracing of Normal Mouse Respiration (
23、Top), and of a“ Moderate” Sensory Irritant Response (Bottom)NOTE 1Taken from Ref. (8).FIG. 2 Typical Tracing of Normal Mouse Respiration (Top), aModerate Pulmonary Irritant Response (Center), and an ExtremePulmonary Irritant Response (Bottom)E981042For this reason the effects of each compound in a f
24、ormulationshould be assessed before any test is made for interactions.6.3 Although the test procedure has been found to show ahigh correlation for sensory irritants with established TLVvalues for man, it may well predict values that are too high forcompounds of low reactivity that are metabolically
25、activated,and also for pulmonary irritants (10).7. Test Animals7.1 Mice are the subjects to be used for this test. It isimperative that they meet the specifications outlined here.Although any mouse of the proper size could be used, markeddifferences have been observed between different strains andse
26、xes (2).7.1.1 Male Swiss-Webster mice shall be used as the testsubjects.7.1.2 Only animals weighing between 22 and 28 g may beused. Smaller mice might be able to crawl into the exposurechamber, while larger ones may not be able to breathe normallyin the apparatus.7.1.3 The same system can be used wi
27、th guinea pigs or ratswith an airflow of 2 L/min when using head dome (9).8. Preparation of Apparatus8.1 Exposure Chamber:8.1.1 The heads of each of four mice extend into theexposure chamber, and the bodies are contained in plethysmo-graph tubes. Perforated rubber dental dam reinforced withelectrica
28、l tape provides tight but comfortable seals around theanimals necks, and rubber stoppers prevent them from back-ing out of the tubes, and provides an airtight body plethysmo-graph (see Fig. 3).8.1.1.1 The “T” tube is of the same diameter as the inlet tothe chamber. The gas or aerosol from the genera
29、tor enters oneside of the “T” and the makeup air enters on the other. Thus thetube acts as a miniature mixing chamber, eliminating the needfor a baffle plate. The “T” tube is not shown in Fig. 3.NOTE 1Dimensions are in centimetres.NOTE 2Taken from Ref. (19).FIG. 3 Glass Exposure Chamber with Attache
30、d Body PlethysmographsNOTE 1Taken from Ref. (19).FIG. 4 Diagram of Test ApparatusE9810438.1.2 Chamber Equilibration:8.1.2.1 It is desirable to reach equilibrium of the testmaterial in the exposure chamber in as short a time as possible.In no case should this time exceed one-tenth of the totalexposur
31、e time. The validity of the data for extrapolation to manrequires rapid attainment of maximum concentration.8.1.2.2 Equilibration time in minutes is 5.0 times the cham-ber volume in litres divided by airflow through the chamber inlitres per minute (12).8.2 A vacuum pump with a control valve monitore
32、d by aflowmeter provides a constant airflow through the exposurechamber. Chamber effluent is passed through an absolute filterand then a sodium carbonate-activated charcoal filter beforeexhausting, preferably into a fume hood. (See Fig. 4.)8.3 Each of the four plethysmograph tubes is connected to ap
33、ressure transducer. As the mouse inhales, a positive pressureis created and exhalation results in a negative pressure. Theamplified signals are recorded on a polygraph, which has thepolarity set so that an upward deflection is obtained duringinspiration and a downward deflection is obtained duringex
34、piration. The signal from each transducer is also fed into afrequency-to-voltage converter, and then fed into a signalaverager. The output of the averager is displayed on a secondrecorder, thus permitting continuous monitoring of the averagerespiratory rate of the four mice. (See Fig. 4.)8.4 A suita
35、ble generator for this test is a glass Dautrebande-type generator modified to allow continuous feed of testmaterial.3This generator can be used for volatile or nonvolatileliquids, solutions, or suspensions of solids. It is depictedschematically in Fig. 5.8.4.1 For aqueous solutions, liquid is delive
36、red via a pumpregulated at 1.0 mL/min to the right-hand tube. This deliveryrate can be varied by a factor of 3 to 4. Air is delivered at 10to 12 psig when a water solution is used, and 8 to 10 psig whenacetone solution is used. With acetone the amount of solutiondelivered is restricted so that no mo
37、re than 3000 ppm acetonevapor is produced in the exposure chamber. The calculation ismade from the total airflow used in the chamber. At thestandard flow rate of 20 L/min through the chamber, deliveryto the generator of 0.22 mL of acetone per minute will result ina concentration of 2800 to 3000 ppm.
38、 With acetone there willbe no liquid overflow, but with aqueous solutions, 1.0 mL/minis high enough so that liquid will fall to the bottom of thegenerator. This is collected in a reservoir via the overflow tube.8.4.2 Arrows in Fig. 5 indicate the path that the aerosol willfollow. Polyethylene Glycol
39、 200 (PEG 200) can be used as asolvent instead of water. The air pressure should be about 20 to25 psig with this solvent. Dry air must be used with PEG 200,which is hygroscopic. Using this generator with a 1 % solutionof test material in water and 20 L/min flow rate through theexposure chamber, the
40、concentration in the chamber will bebetween 10 to 20 mg/m3and most particles will be submi-cronic.8.4.3 The Dautrebande-type generator can also be used tovaporize liquids for exposure of animals to vapors. For thispurpose, the liquid is delivered at a known rate by a regulatedpump and airflow is set
41、 at 10 to 20 psig. For liquids of lowervapor pressure, heating tape can be used around the generatorto increase vaporization efficiency. For aerosols or vaporslikely to oxidize rapidly in air, dry nitrogen should be usedinstead of air. When this is done, pure oxygen is added to thechamber airflow to
42、 maintain 18 to 20 % O2in the exposurechamber. When suspensions are to be tested, the suspendedmaterial must be very fine to prevent clogging of the tip on thegenerator. Although larger tips can be used if required, adegradation of aerosolizing performance will result from theiruse.8.5 To start and
43、stop test material generation, a timer and anassociated control valve are needed in conjunction with theaerosol generator.8.6 When using water or acetone a “dry” particle will beproduced, since both solvents will evaporate. However, PEG200 will not evaporate and a liquid droplet is obtained. Masscon
44、centration in the chamber should be obtained by samplingon filters and weighing on an appropriate balance. A bettermethod, but one not required in a screening experiment, isappropriate chemical analysis. When acetone is used, itsconcentration in the chamber should be verified. Indicator tube3Pitt No
45、. 1 aerosol generator available from Scientific Glassblowing Laboratory,McKees Rocks, PA 15136, has been found suitable.NOTE 1Taken from Ref. (12).FIG. 5 Schematic Representation of the Pitt No. 1 AerosolGeneratorE981044analysis is adequate, or an infrared analyzer or gas chromato-graphic analysis c
46、an be used.8.7 Gases are delivered directly into the exposure chambervia an appropriate flowmeter.8.8 With the exception of the exposure chamber which isessentially a unique piece of apparatus, other parts can besubstituted by similar equipment. Also, minicomputers can beused to replace the frequenc
47、y-to-voltage converter and signal-averaging device. The magnetic tape is not required, and afour-trace oscilloscope with storage capability can replaceoscillograph No. 1.9. Sample Preparation9.1 Because of the large variety of chemicals and formula-tions that can be tested by this procedure, and the
48、 tremendousdifferences in irritant potential between them, no specificstipulation for sample preparation can be made. The onlyrequirement for concentration is that the levels to be tested arespaced at even logarithmic intervals to allow goodconcentration-response curves to be generated from the data
49、obtained. The information provided in the succeeding para-graphs of this section is therefore intended for general guidanceonly.9.2 For solids and nonvolatile liquids, solutions are pre-pared in an appropriate solvent. Water and polyethylene glycol200 (PEG 200) are the most commonly used for this purpose,although 0.1 N HCl, 0.1 N NaOH, and acetone can also beused. In the case of acetone, which is a mild irritant, theconcentration in the chamber should be kept below 3000 ppmto avoid irritation from the solvent.9.3 As an indication of concentrations to be expected, 1 %aqueous ba
