I N I T I A L CONDITIONS
0 0
0
150 ppbc, NMHC XOppbC, NMHC 250 ppbc. NMHC
0 0
ozoc
M o p p b c , NMHC E O p p b C . NMHC Mo ppbC. NMHC
C
ppb
loo0
12W 14w LOCAL DAYLIGHT TIME
1#l
1m
Figure 4. Ozone concentration as a function of time for an air parcel traveling at 8.6 km/h
om
1wO
lm,
1400
l#l
1800
LOCAL DAYLIGHT TIME
Figure 5. Ozone concentration as a function of time for an air parcel traveling at 13.9km/h
1979.
Concluding Remarks Initial input to the photochemid model of levels of NMHC from 250 to 350 ppbC is required in order to match computed with measured and inferred 03. This is well within the expected NMHC range (7, 8, 16). Since traveling air parcel photochemical model calculations with a contemporary mechanism (4)and reasonable assumptions yield results comparable to field measurements, consistency of data, mechanism, and procedure is suggested. The minimal description of NMHC as 25% olefin and 75% paraffin appears adequate for the present conditions. Literature Cited (1) Wagner, H. S., Gregory, G. L., Buglia, J. J., paper presented at
the 71st Annual Meeting of the Air Pollution Control Association, Houston, 1978. (2) J. Air Pollut. Control Assoc., 28(4), 378-80 (1978). (3) DeMarrais, G. A., “The Ozone Problem in the Norfolk, Virginia Area”, EPA-600/4-78-006,1978. (4) Falls, A. H., Seinfeld,J. H., Enuiron. Sci. Technol., 12,1398-406 (1978). ( 5 ) Hecht, T. A., Seinfeld, J. H., Dodge, M. C., Enuiron. Sci. Technol., 8,327-39 (1974). (6) Dodge, M. C., Hecht, T. A., Int. J. Chem. Kinet., Symp. No. 1 (1975). (7) Cofer, W. R. 111,paper 79-57.3 presented at the 72nd Air Pollution Control Association Annual Meeting and Exhibition, Cincinnati,
(8) Copeland, G. E., Davis, R., Maroulis, P., Bandy, A. R., Denyszyn,
R., “Ambient Atmospheric Hydrocarbon Content as Determined by Gas Chromatographic Techniques for Rural Tidewater Virginia in Late Spring 1974”, NASA CR-147922,1975. (9) Hampson, R. F., Jr., Garvin, D., Natl. Bur. Stand. (U.S.), Spec. Publ., No. 513 (1978). (10) Meyers, E. L., Jr., Summerhays, J. E., Feas, W. P., “Uses, Limitations and Technical Basis of Procedure for Quantifying Relationships between Photochemical Oxidants and Precursors”, EPA-450/2-77-021a, 1977. (11) Carter, W. P. L., Lloyd, A. C., Sprung, J. L., Pitts, J. N., Jr., Znt. J . Chem. Kinet., 11,45-101 (1979). (12) Dodge, M. C., Arnts, R. R., Int. J. Chem. Kinet., 11,399-410 (1979). (13) Schere, K. L., Demerjian, K. L., “Calculation of Selected Photolytic Rate Constants over a Diurnal Range: A Computer Algorithm”, EPA-600/4-77-015,1977. (14) Brewer, D. A., Remsberg, E. E., Woodbury, G. E.,Quinn,L. C., “1977 Emissions Inventory for Southeastern Virginia”, NASA TM-80119.1979. (15) Schere, K. L., Demerjian, K. L., in “Proceedings of the 4th Joint Conferenceon Sensing of Environmental Pollutants”,New Orleans, La., 1977, pp 427-33. (16) Freas, W. P., Summerhays, J. E., Martinez, E. L., Meyer, E. C., Jr., Possiel, N. P., Sennett, D. H., “Procedures for Quantifying Relationships between Photochemical Oxidants and Precursors: Supporting Documentation”, EPA-450/2-77-021b, 1978. Received for review October 31, 1979. Accepted April 18, 1980.
A New Technique for the Scanning Electron Microscopy of Particles Collected on Membrane Filters Jean M. M. Le Guen, Stephen J. Rooker’, and Nicholas P. Vaughan Health and Safety Executive, Occupational Medicine and Hygiene Laboratories, 403, Edgware Road, London NW2 6LN, U.K.
A method of treating membrane filters that makes the surface flat and featureless when viewed by a scanning electron microscope (SEMI is described. The good collection characteristics of membrane filters can thus be combined with the advantages of SEM examination. The method involves a minimum of sample preparation, and the filters show no sign of radiation damage even at high accelerating voltages and 1008
Environmental Science & Technology
beam currents. Loss of collected particles during the treatment is negligible. With the new technique, it has been possible to compare directly the same field of view under the SEM and under an optical microscope with phase contrast as used for fiber counting. These studies show that fibers of 0.3 pm diameter are readily visible with the optical system. Work is in progress on other applications.
0013-936X/80/0914-1008$01.OO/O Published 1980 American Chemical Society
Membrane filters have a high collection efficiency, even for particles much smaller than their nominal pore size ( I , 2), which is a great advantage in sampling airborne particulates of small aerodynamic diameter. They have also a low resistance to airflow, so that they can be used to sample particles isokinetically in fast moving airstreams or when the sample is collected with the aid of a small pump. Unfortunately, ordinary membranes cannot generally be used when particles are to be examined by a scanning electron microscope (SEM). The SEM can magnify particles from 20 up to lOOOOOX and can yield information about their morphology and surface characteristics. When it is fitted with an energy-dispersive X-ray analyser, the SEM can be used to determine the elemental composition of particles. However, direct analysiswhich requires minimal sample preparation-by the SEM of collected particles on the surface of membrane filters is severely limited by the fibrous appearance of these filters; small compact and fibrous particles are frequently indistinguishable from the filter matrix. The advantages of using membrane filters are so great that attempts have been made to overcome this problem. Thus, De Nee ( 3 ) has reported some limited improvement when the SEM is used in the back-scattered mode. Stewart ( 4 ) first coated the dust-laden filters with a thin layer of carbon (10 to 20 nm) and then dissolved the filter away either by condensation washing or by the use of a Jaffi. wick, to leave a carbon replica with particles attached. This technique is more generally used for particle examination with the transmission electron microscope (TEM);however, it does give improved contrast between the particles and the filter replica. There are problems associated with both methods used to dissolve the filters. Particle losses of up to 60% have been reported for the condensation washing technique, and the carbon replica is liable to rupture when the Jaff6 wick is used ( 5 ) .Both these techniques are also severely limited by the long sample preparation time required (4-21 h). Zumwalde and Dement (6) and Ortiz and Isom ( 7 ) have described a modified technique for producing a featureless filter background. The dust-laden filter is exposed to acetone vapor which partially dissolves the filter, leaving a microscopically smooth surface with the particles embedded in the top layer. This is then coated with either carbon or carbon/ chromium, and the rest of the filter is dissolved away so that a replica with particles attached remains. Ortiz and Isom claim that losses are less than 10% for particles generally and less than 3% for fibers. However, particles will inevitably be completely coated with partially dissolved filter and will thus be lost in the dissolution process. Indeed, the electron photomicrograph presented in their paper shows a cast of an embedded fiber which was not attached to the carbodchromium film. These difficulties have prompted the use of Nuclepore filters, which offer, apart from the pores, an essentially flat and featureless background. There are a number of disadvantages, however: the particles change position on the filter; some are lost during handling; particle counts of samples, taken from the same site, using both membrane and Nucleopore filters give consistently lower results for Nucleopore filters ( 4 ) .The lower particle counts may be due not to losses but to the less efficient filtration of Nuclepore filters. Liu and Lee (2) and Parker e t al. (8)showed that collection efficiency was lower for particles with a diameter smaller than the pore size of the filter. Spurny et al. ( 9 )found that the efficiency of the filter for collecting airborne fibers was dependent mainly on the fiber diameter. Low efficiencies were obtained for fibers of diameter smaller than the pore size of the filter. They also showed that the pressure drop across Nuclepore filters is considerably greater than that across membrane filters with the same nominal size. In addition, the Nuclepore filter has
practical drawbacks; it has a lower load capacity than the membrane filter, and it is difficult to use in the field as it is very thin and fragile and tends to curl up because of its electrostatic properties. These disadvantages have seriously limited the usefulness of Nuclepore filters in many situations and, consequently, the usefulness of the SEM in this application. For example, their high-pressure drop has prevented their use in personal samplers for particles of aerodynamic diameters less than 1 pm (such particles are prevalent in clouds of fibrous dust); they cannot be used for sampling isokinetically fast-moving air streams usually present in stacks and ducts because of their high resistance to airflow and their fragility. The work described below demonstrates a new, rapid, and reliable method of preparing membrane filters for SEM examination with negligible particle loss. The method also clears the filters so that it is possible to examine them by optical microscopy. The paper reports, as an example of the potential of the method, the preliminary results obtained when a direct comparison was made of the same field of view as seen by the SEM and the optical microscope, using the phase contrast technique for fiber counting. P r e p a r a t i o n of Filters
Two types of membrane, Gelman DM450 and DM800, made of a copolymer of poly(viny1 chloride) and acrylonitrile, with nominal pore sizes of 0.45 and 0.8 pm, respectively, were used to sample a cloud of mixed Union Internationale Contre le Cancer (UICC) chrysotile and amosite asbestos fibers generated in a dust chamber. The membranes were then prepared for microscopic examination by the following method: 60 to 80 pL of clearing solution made of 33% dioxane and 67%cyclohexanone was placed on a clean microscope slide with a push-button micropipet and smeared out over an area about the size of the filter. The filter, sample face upward, was quickly and carefully laid on top of the solution. Great care was taken not to trap any air bubbles. The solution evaporated fairly quickly, and best results were obtained when the microscope slide was placed on a flat bench and the filter was held ready in a pair of tweezers before the clearing solution was smeared. Any solution not absorbed by the filter was removed with a tissue, and the filter and microscope slide were left in a dust-free environment for about 3 min and then dried in an oven a t 60-75 "C for 10 min. During this time the filter collapsed to about 15% (20 pm) of its original thickness, and the clearing solution evaporated to leave a permanent thin transparent plastic film in which the fibers were embedded in the uppermost layer and all lay close to one plane. The distortion of the filter was minimal. The cleared filter on the slide was etched in a plasma oven (Nanotech, Plasmaprep P100) for 7 min with an oxygen flow rate of 8 cm3/min and forward and reflected radiofrequency power of 100 and 2 W, respectively, as determined by the meters on the instrument. The filter was freed from the microscope slide with the aid of a scalpel transferred onto an SEM stub and secured with conducting Silver Dag. The filter was cleared on a microscope slide so that it could be examined by optical transmission microscopy before it was mounted for SEM examination. However, when this facility was not required, the preparation could be simplified by clearing the filter directly onto an SEM stub. The filter and stub may then be coated with gold or carbon. Carbon coating is preferable when energy-dispersive X-ray analysis is intended. The etching rate and time have been established by trial and error. An etching time of 5 min, although sufficient to expose most fibers, leaves some partially embedded in the collapsed filter. Longer periods, around 10 min, leave the fibers mounted on ridges of unetched filter. Experience has shown that an etching time of 7 min is sufficient to leave the Volume 14, Number 8, August 1980
1009
fibers clear of the filter matrix and also to expose small par. tirulates trapped a,ut in the upper layers of the original iilcer. Blank filters prepared under these conditions have tlat and teat ureless surfaces. The Gelman DM filters were rwnplrtely stahle under the electron beam and showed no sign of radiation damage even afier prchmged e x l ~ s u r eto thr highest arcelrrating vdtage (50kV1 and beam current (500 AI ohtainable on the Camhridgr Stcreoxm 180 mirruscope. Collapsed cellulosic filters t 1 0 1 [mixed cellulose aretate and nitrate) showed very rapid and severe radiation damage and were nut suitahle for SFM work by this technique. C'ornpnrison o/SE.\I and P h a Conlru.,r ~ Microscopy for Fi her Sizinfi
The resolving power of an optical micruscope depends on the numerical aprrture of the oh,iective used and is defined as the smallest distance hetwren twli or more strurtural details which may be seen as separaw entities. Where suftirient image cuntrast and magnifirntion exist, thr human eye is able to distinguish an isolated partirle from itssurroundings even if it is n d fully r r w l w l . This means that for objects with strong image contrasts such as those aswriated with phase riintrast microscopy, where contrast is enhanced by interferenre techniques, particles smaller than the resolving power of the
Figure 1. Photomicrograph of gmosite and chrysotile on a collapsed and cleared DM 800 filter. Optical magnification SOOX, phase contrast
Figure 2. Electron micrograph of identical field as seen by Cambridge 180 stereoscan SEM
1010 Environmental Science & Technology
objective become visible (11, 12). Clearly, it is of interest t o know the lower limit of detectability of fibrous particles under phase contrast with a typical magnification (about 500X) used for counting and sizing (13). Gelman DM membrane filters, laden with asbestos, were treated by the etching process given above, but omitting the coating procedure. An identification grid was scratched on the filter t o aid in the location of the fields. A drop of trichloroethene was placed on the filter and a cover slip laid on top. Trichloroethene was used as a contact liquid between the cleared filter and the cover slip for two reasons. Firstly, it has a refractive index of 1.48 and therefore mimics the conditions given by Galvin and LeGuen ( 1 4 ) for clearing membrane Millipore RA filters so that particles collected on them can be counted and sized. Secondly, trichloroethene is very volatile and can be easily removed by evaporation without disturbing the particles on the surface of the cleared filter. Photomicrographs of various fields were taken using a Reichert Zetopan microscope with an optical magnification of 500X under phase contrast. The microscope slide was placed in an oven at 60 "C to evaporate the trichloroethene. When the slide was dry, the cover slip was removed and the filter was mounted and titanium coated on an SEM stub. Electron micrographs were made of the same fields as had been photographed on the optical microscope, and selected areas of those fields were
Figure 3. Electron micrograph of region 1 in Figures 1 and 2
Figure 4. Electron micrograph of region 2 in Figures 1 and 2
photographed a t magnifications up to l O O O O x , with point to point resolution of better than 80 nm. Figures 1 and 2 show identical fields as seen by the optical microscope and the SEM. A comparison of the two micrographs shows that no particles are lost during the etching and coating process. Since it is unlikely that particles are lost during the clearing stage, it has been concluded that particle losses for the overall process are negligible. The regions marked 1 and 2 on Figures 1 and 2 are shown a t higher magnification in Figures 3 and 4, respectively. Measurement of fiber A (Figures 3 and 4) gives values of 0.3 ym for the diameter and 13 ym for the length; the fiber labeled B has a length of 3.5 ym and a diameter that varies along its length between 0.35 and 0.18 ym. Fiber A is clearly visible in Figure 1, while fiber B is hardly discernible from the background. The visibility of particles under phase contrast is dependent on several factors, including the absorption of the phase rings, the difference in refractive indices between the mounting medium and the particle, and the angle subtended by the particle and the eye ( I 5 ) .Under the conditions described (65% absorption phase ring, refractive index of the mounting medium 1.48, and optical magnification 500X), fibers of diameter greater than 0.3 pm are visible.
Conclusion A method is described of preparing asbestos fibers collected on Gelman DM filters for examination with the SEM. The filters show no sign of radiation damage, even a t the highest accelerating voltage and beam current obtainable on most SEMs. The available evidence indicates that particle losses from the filter are negligible. I t has been possible to compare directly the same field of view as seen by the optical microscope with phase contrast and by the SEM. The studies show that fibers of diameter greater than 0.3 ym are readily visible with the optical microscope under the conditions described.
Work is in progress on other applications of the technique, including the isokinetic sampling of particles of small aerodynamic diameter (such as fibers) in fast-moving airstreams and the identification of small particles. The latter is possible only with particles that are unaffected by the etching process. However, since these particles are no longer embedded in a matrix of fixed refractive index, they can be readily identified by various optical microscopic techniques ( I O ) or X-ray energy dispersive analyzers.
Literature Cited (1) Spurny, K., Pich, J., Int. J . Air Water Pollut., 8, 193-6 (1964). ( 2 ) Liu, B. Y. H., Lee, K. W.,Enuiron. Sci. Technol., 10, 345-50 (1976). (3) De Nee, P. B., Proc. S y m p . Electron Microsc. Microfibers, 68 (1976). (4) Stewart, I. M., Proc. Symp. Electron Microsc. Microfibers, 93 (1976). (5) Beaman, D. R.,Walker, H. J., Proc. S j m p . Electron Microsc. Microfibers, 98 (1976). (6) Zumwalde, R. D., Dement, J. M., Proc. S j m p . Electron Microsc. Microfibers, 139 (1976). ( 7 ) Ortiz, L. IT., Isom, B. L., Proc. Electron Microsc. Soc. Am., 32nd, 554 (1974). ( 8 ) Parker, R. D., Buzzard, G . H., Dzubay, T. G., Bel1,J. P., Atmos. Enuiron., 11,617-21 (1977). (9) Spurny,K. R., Stober, W., Ackerman, E. R., Lodge, J. P., Spurny, K., J . Air Pollut. Control Assoc., 26,496-8 (1976). (10) LeGuen, J. M. M., Rooker, S. J., Vaughan, N. P., Health & Safety Executive, Internal Report No. IR/L/FD/80/23 (available from authors). (11) Van Duijn, C., Microscope, 11,301-9 (1957-8). (12) \’on Gies Heidermanns, G., Staub-Reinhalt. Luft, 38(10),423-5 (1978). (13) Asbestosis Research Council, “The Measurement of Airborne Asbestos by the Membrane Filter Method”, Technical Note No. 1. Rochdale, Lancashire, U.K., 1971. (14) Galvin, S., LeGuen, J. M. M.:Ann. Occup. Hyg., in press. (15) LeGuen, J. M. M., Proc. Workshop Warmensteinach, 68 (1977). Received f o r reuiew October 19, 1979. Accepted April 21, 1980
CORRESPONDENCE
SIR: An article on the budgeting of polychlorinated biphenyl (PCB)fluxes in and out of Lake Superior by Eisenreich e t al. ( I ) has a computational error concerning the direction and magnitude of an estimated PCB flux. The error is in the equation chosen to describe the transfer of PCB in the vapor phase from the atmosphere to the lake water. This equation is of the form: flux = K,C,
(1)
where K , and C,, respectively, are the mass transfer coefficient and concentration of atmospheric PCB in the vapor phase. The formulation of this equation is credited to Bidleman e t al. (21,who derived it as a special case of the general equation that describes the rate of mass transfer in the two-film model (3). Two assumptions were made, namely, that the exchange of chlorinated hydrocarbons between air and water is gas-phase controlled and that the water surface acts as a “perfect absorber” of these substances. Neither of these as0013-936X/80/0914-1011$01.00/0
@
1980 American Chemical Society
sumptions appears to be valid, as laboratory and field studies have demonstrated the volatilization of PCB from aqueous solutions and that this process is liquid-phase controlled ( 4 , 5). Using the values provided by Eisenreich et al. for C, ( I ) and the values given by Mackay and Leinonen for the Henry’s law constants of the different PCB groups (61, it can be shown that in the case of Lake Superior the atmospheric concentration of PCB can be assumed to be negligible with respect to the aqueous PCB.Furthermore, it can be assumed that a negligible portion of the aqueous PCB is adsorbed to suspended sediments ( 7 ) .With these approximations in mind the general mass transfer equation of the two-film model can be simplified to: flux = -KIC1
(2)
where K1 is the mass transfer coefficient of PCB in the lake water and C1 is the aqueous PCB concentration. Note that the direction of the flux is from the water to the atmosphere. Volume 14, Number 8, August 1980
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