Anal. Chem. 1991, 63, 886-890
886
Radial Mass Sensitivity of the Quartz Crystal Microbalance in Liquid Media Michael D. Ward*
Department of Chemical Engineering and Materials Science, University of Minnesota, Amundson Hall, 421 Washington Avenue SE,Minneapolis, Minnesota 55455 E d w a r d J. Delawski
Central Research and Development Department, Experimental Station, E. I . d u Pont de Nemours & Co., Inc., P.O. Box 80328, Wilmington, Delaware 19880-0328
The radial sensltivtty function of the quartz crystal microbalance (OCM), employing an AT-cut quartz resonator In aqueous media, has been determined by simultaneous in situ measurement of frequency and charge during copper electrodeposltlon in holes etched in a photoresist polymer on the OCM. Frequency-charge correlations are determlned by copper depositlon either in small (0.025 in.) holes with dlfferent angular and radial locations or in concentric clrcular holes with dtfferent radii. The data lndlcate a Gausslan mass sensitlvlty distribution, with the greatest dtfferentiai sensttlvity in the center of the resonator, decreasing monotonically with increasing radius. Narrower sensitlvity distributions and less mass sensitivity beyond the electrode edges were observed with plano-convex crystals, conslstent with a greater degree of energy trapping that results from the larger mass in the center of contoured crystals. The results demonstrate, as with applications of the QCM In the gas phase, that coatlng uniformity Is Important for accurate measurements of mass with the QCM when used in ilquid media. The signlflcant sensitivity observed beyond the electrode boundary in piano-piano resonators indicates that sensors for iiquld-phase applications must be properly callbrated to account for this property.
INTRODUCTION The quartz crystal microbalance (QCM), comprising a thin quartz wafer sandwiched between two metal excitation electrodes, has been employed extensively for measurement of interfacial mass changes that occur at one of the excitation electrodes (I).For thin films exhibiting elastic behavior (21, mass changes a t the QCM surface generally are determined from the Sauerbrey relationship (3)
A f = --
2f02Am (1)
where Af is the measured frequency shift, fo is the parent frequency of the quartz crystal, Am is the mass change, A is the piezoelectrically active area, pq is the density of quartz (2.648g ~ m - ~and ) , M, is the shear modulus (2.947 x 10" dynes cm-* for AT-cut quartz). The frequency-mass correspondence of the QCM has led to is widespread use as a thickness monitor in metal evaporations. Numerous approaches to chemical and biological sensors using chemically and immunologically active films immobilized on the QCM also have been reported for both gas- and liquid-phase applications ( 4 , 5 ) . Indeed, much of the recent interest in quartz resonators can be attributed to their ability to measure mass changes in solution. Notably, the QCM excitation electrode that faces solution in liquid-
phase applications also can be employed in a conventional electrochemical cell to measure electrochemically induced interfacial mass changes (6-11). The Sauerbrey relationship assumes that the frequency shift resulting from a mass deposited in a small region of the QCM will be identical with the contribution of that m m to the total frequency change when it is a portion of a film that covers the entire piezoelectrically active area of the quartz crystal. The frequency response to that mass is dictated by the differential sensitivity constant, cf, which represents the differential frequency shift for a mass change on a region of the QCM (eq 2). Integration of cf over the total surface area of
cf = d f / d m
(2)
(3) A f = -CfAm/A
(4)
the QCM affords the integral sensitivity constant, Cf (eq 3). The frequency shift expected for the deposition of a uniform film can be expressed in terms of C f according to eq 4, which is equivalent to eq 1. Application of eq 1 or 4 requires, however, that the deposited film have uniform thickness across the entire active region of the resonator. This condition is necessary because cf is a function of distance from the center of the resonator. Indeed, previous studies in which metal films were evaporated or sputtered onto localized areas of a QCM demonstrated that cf is highest a t the center of the QCM and decreases monotonically in a Gaussian manner, becoming negligible a t and beyond the electrode boundary (3,12).This behavior is attributed to the decrease in shear amplitude of the crystal oscillations with increasing r [cf (shear amplit~de)~]. Charge polarization (13)and admittance (14)experiments using small probe electrodes in air have corroborated these studies, demonstrating that the shear amplitude also exhibits a Gaussian distribution. A recent study using a tungsten probe to measure the radial dependence of shear amplitude of AT-cut quartz resonators immersed in water has indicated similar behavior (15). Notably, the shear amplitude in liquid was less than that measured in air, and shear motion extended beyond the electroded region of the resonator. This latter property, referred to as field fringing, was attributed to the dielectric contribution from water. In addition, larger frequency shifts were observed upon the probe contacting the resonator in water, which was attributed to viscous coupling (16). These experiments have revealed that the physical behavior of quartz resonators is significantly different in gas and liquid media. Most QCM experiments performed in liquid, however, are interpreted on the well-characterized behavior of quartz resonators in the gas phase. While the integral mass sensitivity
0003-2700/9 1/0363-0886$02.50/0 0 199 1 American Chemical Society
ANALYTICAL CHEMISTRY, VOL. 63, NO. 9, MAY 1, 1991
Un8torm hale w e
Concentric holes
Flguro 1. Experimental procedure for the electroplating of copper in circular features on the QCM.
in liquids has been implicitly determined in numerous studies, the radius-dependent differential sensitivity has not been examined. The observations of field fringing and diminished shear amplitudes in liquid prompted us to examine the radial dependence of cy, since mass sensitivity distribution will play a significant role in the QCM response when nonuniform films are present or mass changes occur beyond the electrode boundary. For example, corrosion monitors may involve nonuniform films due to nonuniform current densities, and QCM sensors commonly have active films that extend beyond the excitation electrode boundary. There is also a surprising absence of mass sensitivity distribution measurements for contoured resonators, which are expected to exhibit less field fringing because of their more efficient energy trapping. If this is indeed the case, contoured crystals may prove to be more reliable for sensor applications. We describe herein investigations of the mass sensitivity distribution on planar and contoured AT-cut quartz resonators, as determined by in situ measurements of copper electrodeposition.
EXPERIMENTAL SECTION Apparatus. The experimental apparatus comprised a 5-MHz AT-cut quartz crystal (Valpey-Fisher,Hopkinton, MA, or McCoy Electronics, Mt. Holly, PA) and a homemade oscillator designed to drive the crystal at its resonant frequency. Gold electrodes (2000 A thick) were deposited on chromium underlayers (200 A) on both sides of the crystal by using evaporative techniques. The patterns were arranged so that the gold leads from the outer edges of thecrystal to the center circular pad on opposite sides did not overlap. The quartz crystal was mounted between two O-rings confined by standard glass fittings and a metal clamp. The areas of the top (facing solution) and bottom (facing air) electrodeswere identical for experiments in which A = 0.18 cm2 (re = 0.24 cm) and A = 0.32 cm2 (re = 0.32 cm). Experiments with 0.08-cm2 electrodes (re = 0.16 cm) were performed, however, with a smaller bottom electrode (re = 0.16 cm) and a larger top electrode (re = 0.32 cm), which facilitated collection of a sufficient number of data points during electrodeposition. According to the Sauerbrey equation, the sensitivity of the 5-MHz QCM is 0.057 Hz cm2ng-l. The frequency of the QCM was monitored with a Hewlett Packard 5384A frequency counter and recorded with a Digital Equipment Corp. PDP-11/73 computer. Impedance analyses were performed with a Hewlett Packard 4194A impedance/gain-phase analyzer. A commercially available potentiostat (either a Princeton Applied Research 173 or 273) was used with the working electrode at hard ground and the current determined by measurement of the voltage drop across a 1000-R resistor between the potentiostat and the counter electrode, as described previously (9). Polymer and copper film thicknesses were measured with a Sloan Dektak profilometer. Preparations of Electrodes. All spin coating and developing procedures were performed in a class 100 clean room in order to avoid contamination by dust particles that compromise the integrity of the thin polymer films. The crystals with electrodes were cleaned on a spin coater by adding acetone to the surface while spinning at 3000 rpm (Figure 1). Then the crystal was spin-coated with positive photoresist (Shipley Microposit 1813, Shipley Co., Newton, MA) at 3000 rpm for 20 s, followed by curing at 100 'C for 120 s. This affords an approximately 1.5 pm thick polymer film. The films were then exposed through 0.010-in.
887
Kapton films with the appropriate hole size to 17.5 W cm-2 of 366-nm light for 8 s. Development was performed by immersion of the coated crystal in Shipley 321 developer solution for 35 s followed by immersion for 5 s in a second bath containing the same solution. The crystals were then washed with water and dried with clean air, and a circular feature identical with the size and location of the hole in the Kapton mask was evident in the polymer, exposing the gold electrode in this region. Electroplating of copper in these circular features was then performed by immersing the photoresist side of the crystal in 0.1 M HzSOl conM CuS04, dynamically measuring the charge taining 20 X and frequency shift associated with copper deposition. Electroplating was performed at minimum overpotentials in order to minimize high current densities at electrode edges. We found that electroplating could be accomplished conveniently at -0.07 V vs Ag/AgCl (Bioanalytical Systems, Inc.). The radial sensitivity was probed by two methods. The first method involved etching a small hole (0.025 in.) in the photoresist film on an AT-cut quartz crystal so that a small circular area of the gold excitation/working electrode was exposed. This was repeated for several crystals at different values of r and B (in the following discussions we only illustrate data collected for B = 0' (parallel to the 3c axis, the direction of shear) and 90' (perpendicular to the x axis); the angular dependences generally were not significant enough to warrant extensive discussion). The second method involved electroplating in concentric circular areas with different radii. In both cases, regression analysis of frequency change vs electrochemical charge plots generally afforded correlation coefficients >0.99, and the slopes of these plots ( A f / Q ) were used to determine the frequency-charge ratios, which is equivalent to the sensitivity normalized to the mass deposited. This value can be further normalized to thickness by conversion to the sensitivity constant, which has units of Hz cm2g-l. The electroplating was performed so that the copper film thickness never exceeded 0.5 pm, which was one-third the height of the film. Network analysis did not indicate any viscoelastic effects or solvent drag from trapping in the etched features; conductance peaks and quality factors measured with the etched polymer films before or after electroplating were identical with those of bare gold surfaces when compared under the same conditions.
RESULTS AND DISCUSSION Concentric Circular Patterns. The radial dependence of the differential mass sensitivity of an AT-cut quartz resonator in aqueous medium can be demonstrated readily by electroplating copper on the exposed gold electrode in lithographically developed circular regions of different areas centered about r = 0, while dynamically, and simultaneously, measuring electrochemical charge and frequency. These experiments are conceptually equivalent to measuring the stepwise change in integral sensitivity for incremental additions of annular rings with increasing r values. The change in integral sensitivity during each step is equivalent to the differential sensitivity averaged over all angles of 8, that is, cf integrated over 27rr dr (eq 3). These experiments therefore absorb any angular dependence of cp The mass sensitivity at different r values can be expressed conveniently in terms of an integral sensitivity, C; (eq 5), which for r < lrel (reis the actual radius of the excitation electrode) will be less than Cf (ideally, a t r = &re, C; = Cf). The results from these experiments can be plotted as Af/Q, which can be related to the sensitivity constant by eq 6, where MW is the molar mass of C i = Jrcf27rr dr
A f / Q = C/MW/BFA' copper and A' is the area of electrodeposition. The quotient A f / Q is equivalent to the frequency change normalized to the mass of copper deposited and can be obtained directly from the slope of a plot of Af vs Q . Plots of A f / Q revealed a Gaussian mass sensitivity distribution (Figure 2a) with the largest A f / Q values at the center.
ANALYTICAL CHEMISTRY, VOL. 63, NO. 9, MAY 1, 1991
888
140000
100000
i
. a
80000 90000
120000
70000
-
5 a 80000 I
60000
-
9a
50000
-
100000
h
% 2 9
60000
40000
4
20000
40000 -
'
'
30000
'
"
'
'
'
'
'
"
'
0' -0.6
I
0.2
0.4
0.6
Flgwe 3. Sensitivity dependence of plano-plano crystals on the radial position of 0.025-in. holes aligned parallel (0)and perpendicular (m) to the x axis (0 = 0 and go', respectively),expressed as A f / Qratios measured during copper electroplating: Ir,l = 0.32 cm. The points depicted at -r were obtained by reflection through zero of the points at + r .
0.05
b
0.04
r > f r , (Figure 2b) due to the decrease in mass sensitivity beyond the electrode edge. In this region the A f / Q values, in the absence of field fringing, ideally should follow eq 7,
0.03
A f / Q = CfMW/(2F[are2+ 2(r - re)wll
0.02 0.01
(7)
where w is the width of the electrode tabs which are exposed when r > re. Therefore, A f / Q in this region should decrease with l / [ r r ; 2(r - r,)w]. If field fringing is operative and maw sensitivity extends beyond re, 4 / Q will not fall as rapidly as predicted by eq 7 . Indeed, comparison of the data from Figure 2a a t values of r > Ir,l with those expected from eq 7 indicates this behavior (Figure 2c). The mass sensitivity a t r > Jrelis elucidated more readily by electrodeposition in small holes in the polymer film at different values of r, as described in the following section. Electroplating in Small Holes. In order to determine directly the differential sensitivity along specified values of 0, electrodeposition of copper was performed in 0.025-in. holes on AT-cut quartz crystals at different values of r. Since this method compares the mass sensitivity in regions of identical areas, the ratio A f / Q is a direct measurement of the differential sensitivity, as given by eq 8, where A" is the area of the 0.025-in. hole.
+
0.00 0.0
0.1
0.2
0.3
0.5
0.4
' (cm)
32000
0.0
r(cm)
hole radius (cm)
..
-0.4 -0.2
IC
r
300001
0.32
A f / Q = -cfMW/2FA"
re i
0.34
'
i
0.36
.
'
0.38
L
0.40
'
I
0.42
Flgure 2. (a) Sensitivity distribution of phno-plano crystals for circular patterns with radius r whose centers are located at r = 0, expressed as A f l Qratios measured during copper electroplating (Ir,l = 0.32 cm). The points depicted at -rare obtained by reflection through zero of the points at + r . (b) Sensitivity constants for circular patterns with radius r whose centers are located at r = 0. (c) Expanded view of the sensitivity distribution In (a)for values or r > Ir,l. The open squares (0)depict the measured value of A f l Q , and the filled squares p), the expected trend if mass sensitivity did not extend beyond the electrode
boundary. Conceptually, this indicates that deposition in a small area about the center affords a greater frequency shift than when a deposit of equivalent mass is distributed over a larger area of the actively vibrating area. This is consistent with larger vibrational amplitude and more sensitivity a t the center of the resonator. The integral sensitivity constant determined in these experiments clearly increases as r becomes larger, approaching Cf near r = re, and then decreases at values of
(8)
Data collected along the x axis (0 = Oo) indicated that A f / Q was clearly larger a t the center of the electrode ( r = 0) than near the electrode edge at r = *re (Figure 3). Our results with plano-plano crystals with 0.32 diameter electrodes indicate a 3-4-fold greater sensitivity near the center than just inside the edge of the electrode. Similar sensitivity profiles were observed when r was varied in a direction perpendicular to the x axis (0 = 90'). In both cases, the electrodes were configured so that electrodeposition could be performed on the electrode tabs at r > lrel (the tabs of the top and bottom electrodes did not overlap). The mass sensitivity distributions qualitatively resemble the Gaussian profiles reported previously in gas-phase measurements involving metal evaporation and sputtering (3,12);however, they are broader and exhibit significantly greater mass sensitivity at r > lrel (in the gas phase the resonators exhibit negligible mass sensitivity at r > Ir,l). This is in agreement with the extensive field fringing noted in probe experiments with these resonators in water (15). The origin of the differential mass sensitivity distribution is attributed to confinement of the crystal oscillations to the electroded region by "energy trapping". This property is a direct result of mass loading by the electrodes, which increases
ANALYTICAL CHEMISTRY, VOL. 63, NO. 9, MAY 1, 1991
300000
L El
-Q
200000
-
4-k I
I
01
I ““/I I
I
0 a 100000
I
I
\
.. \
\El
\
\”
10
-
t
00.6
40000
30000
o^ E 20000
P d 10000
Figure 4. (a) Sensitivity dependence on the radial position of 0.025-in. holes aligned parallel to the x axis (0 = 0’) for lrsl = 0.32 cm and quartz crystals, expressed as plano-plano (W)and plano-convex (0) Af 10 ratios measured during copper electroplating. (b) Sensitivity dependence on the radial position of 0.025-in. holes aligned parallel to the x axis (0 = 0’) for 1r.l = 0.16 cm and piano-plano (W)and quartz crystals, expressed as Af10 ratios measured plano-convex (0) during copper electroplating. In both cases, the points depicted at -r are obtained by reflection through zero of the points at + r . The plano-convex crystals were coated with the photoresist polymer on the plan0 side, with this side immersed in solution.
the effective density in this region compared to the unplated portion of the crystal. As a result, the electroded and unplated regions have cutoff frequencies below which acoustic waves cannot propagate without attenuation, designated as oeand wq for the electroded and unplated quartz regions, respectively. These cutoff frequencies are equivalent to the frequencies of the fundamental resonant thickness-shear modes in these regions, f r ( e ) and fr(q). Accordingly, the amplitudes of acoustic waves with frequencies between we and wq decrease exponentially, with the most severely attenuated. Frequencies greater than wq propagate freely through the unplated region until dampened by clamps or contacts. The result is that the energy of the fundamental mode of interest is trapped in the electroded region, with the amplitude greatest in the center and approaching negligible values near the edges. We attribute the significant mass sensitivity a t r > lrel to field fringing that is enhanced by the presence of the liquid or the polymer in the unplated region as well as the presence of the electrode tabs upon which deposition occurs at r > pel. These tabs effectively lower the cutoff frequency in this region compared to the ideal case in which the tabs have zero mass, resulting is less attenuation of the shear amplitude. As a result, significant mass sensitivity in this region can be observed. The role of energy trapping can be illustrated by measurements of the differential sensitivity on plano-convex crystals. The mass sensitivity distributions of planc-convex crystals are narrower and exhibit greater amplitudes a t r =
plano-plan0
888
u I
I
plano-convex
0 compared to plano-plano resonators (Figure 4). The data depicted in Figure 4 were obtained with the plano side of the crystal coated with the polymer and facing solution. The mass sensitivity at values of r > Irel is significantly reduced for the plano-convex resonators, indicating less fringing. The sensitivity at r = 0 is approximately 2 times higher than that observed for planc-plan0 crystals. These results are consistent with the larger effective mass in the center due to the contouring of the plano-convex crystals. The large effective mass traps more of the energy of the acoustic wave in the center, and thus a greater mass sensitivity is observed in this region. This clearly is evident for resonators possessing either pel = 0.32- and Irel = 0.16-cm electrodes. The latter also exhibits greater amplitude a t r = 0, as expected for smaller diameter electrode. For the resonator with lrel = 0.32 cm, the integral sensitivities of the plano-plano and plano-convex resonators are similar. The plano-convex resonator with the smaller Irel = 0.16 cm electrode, however, appears to have a greater integral sensitivity compared to the plano-plan0 resonator with the same electrode size. The energy trapping effect should also be evident for quartz crystals with different electrode thicknesses. We did not observe any differences, however, between data collected on crystals possessing 2000 and 4000 8, thick gold electrodes. In principle, the mass sensitivity of the QCM is not dependent upon electrode area. That is, the integral sensitivity is identical for all electrode sizes. This therefore requires a narrower differential mass sensitivity distribution for smaller electrodes with a greater maximum. Comparison of data collected with 0.32- and 0.28-cm2electrodes (re= 0.32 and 0.24 cm, respectively) suggest this trend, although we have found that the differences between these two sizes are not particularly dramatic. One data set with the smaller electrode along the x axis (0 = Oo) exhibited a greater mass sensitivity in the center of the resonator and a narrower distribution throughout all r values, indicating similar integral sensitivities for both sizes. In another data set collected at = 90°, the maxima were identical, but the profile was narrower for the smaller electrode, suggesting a somewhat different integral sensitivity along this direction. The apparent difference in behavior along the two directions is not entirely unexpected, since the amplitude distributions are anisotropic, although Figure 3 suggests similar mass sensitivity distributions along these two directions. Electrodeposition on bare gold electrodes (without photoresist) of different sizes, however, did not reveal significant differences in overall sensitivity for electrodes ranging from 0.16 < Irel < 0.32 cm. The experiments with the small holes, only collected along 0 = 0 and 90°, may not account adequately for the differential sensitivity throughout all values of 0. The apparent discrepancies in the differential sensitivity also may be due to irreproducibilities in different quartz resonators. In general, we have found that smaller diameter electrodes afford narrower mass sensitivity distributions, but the integral sensitivities are similar within the range of electrode sizes investigated. In conclusion, the differential sensitivity of the QCM in liquid media clearly is not uniform across the active region. Indeed, the radial dependence of cf closely resembles that previously obtained in vacuum by metal evaporation and sputtering and is consistent with the radial dependence of the shear vibrational amplitude of the quartz oscillator. Unlike previously reported gas-phase mass sensitivity measurements, significant mass sensitivity is evident beyond the edge of the circular electrode in liquid media, particularly with planoplano resonators. We estimate that for immersed plano-plan0
Anal. Chem. 1991. 63,890-893
890
resonators coated with the photoresist polymer as much as 10-15% of the integral sensitivity stems from the region beyond the electrode boundary. This behavior is attributed to the dielectric contribution and mass loading of the liquid medium and polymer film, as well as the mass loading of the electrode tabs in this region. Our results clearly indicate that calibration of the QCM based on geometric measurements alone can lead to erroneous results. It is therefore important to calibrate the QCM, preferably by electroplating and stripping, to guarantee accurate measurements of mass with the QCM in liquids. In addition, the observation of appreciable mass sensitivity at r > Ir,l for plano-plano crystals is especially significant for calibration of QCM sensors, which commonly employ designs in which active films are present, and therefore mass changes occur, in nonelectroded regions of the resonator. Our results suggest that plano-convex resonators, which exhibit negligible sensitivity at r > IrJ, may be more suitable for mass-sensing applications.
ACKNOWLEDGMENT We gratefully acknowledge the assistance of J. Howe (Du Pont). Registry NO.CU, 7440-50-8;Ag, 7440-57-5; quartz, 14808-60-7. LITERATURE CITED (1) (a) Bruckensteln, S.; Shay, M. Electrochim. Acta 1985, 30, 1295. (b) Buttry. D. A. I n Electroanalytical Chemistry; Bard, A. J., Ed.; Marcel Dekker: New York, 1990; Vol. 17, p 1. (c) Ward, M. D.: Buttry, D. A. Science 1990, 249, 1000. (2) Lu, C.; Lewis, 0. J . Appl. fhys. 1972, 43, 4385. (3) Sauerbrey, G. 2.Phys. 1959, 155, 206. (4) (a) Gullbault. G. 0. Ion-Sel. Electrode Rev. 1880, 2 , 3. (b) Guilbault, G. G.; Jordan, J. M. CRC Crn. Rev. Anal. Chem. 1988, 79, 1 and references therein.
(5) (a) Ngeh-Ngwainbi, J.; Foley, P. H.; Kuan, S. S.; Gullbault, 0. 0. J . Am. Chem. Soc. 1988, 108, 5444. (b) Muramatsu. H.; Dicks, J. M.; Tamiya, E.; Karube, 1. Anal. Chem. 1987, 59. 2760. (c) Shons, A.; Dorman, F.; Najarian, J. J . Biomed. Mater. Res. 1972, 6 , 565. (d) U. S. Patent 4,236,893. (e) Roederer, J. E.; Bastiaans. 0. J. Anal. Chem. 1983, 55, 2333. (f) U. S. Patent 4,242,096. (9) Ebersole, R. C.; Ward, M. D. J . Am. Chem. SOC. 1988, 710, 8623. (h) Ebersole, R. C . ; Miller, J. A.; Moran. J. R.; Ward, M. D. J. Am. Chem. Soc. 1990, 772, 3239. (I) Lasky, S. J.; Buttry, D. A. ACS Symp. Ser. 1989, 403, 237. (6) (a) Melroy, 0.: Kanazawa, K.; Gordon, J. G., 11.; Buttry, D. Langmulr 1988, 2, 697. (b) Deakln. M. R.; Melroy, 0. J . Electroanal. Chem. Interfacial Electrochem. 1988. 239, 321. (7) (a) Kaufman, J. H.; Kanazawa, K. K.; Street, G. B. fhys. Rev. Lett. 1984, 53, 2461. (b) Varineau. P. T.; Buttry, D. A. J . Phys. Chem. 1987, 9 7 , 1292. (c) Ward, M. D. J. Electrochem. SOC. 1988, 735, 2747. (d) Orata. D.;Buttry, D. A. J . Am. Chem. Soc. 1987, 709, 3574. (8) Masuda, H.; Baba, N. Chem. Lett. 1987, 1877. (9) Ward, M. D. J . fhys. Chem. 1988, 9 2 , 2049. (10) (a) Schumacher, R.; Mueller, A.; Stoeckel. W. J . Electroanal. Chem. InterfacialElectrochem. 1987, 279, 311. (b) Schumacher, R.; Gordon, J. G.; Melroy. 0. J . Electrmnal. Chem. Interfacial Electrochem. 1987, 276, 127. (c) Schumacher, R.; Borges, G.; Kanazawa, K. K. Surf. Sci. 1985, 163, L261. (11) Baker, C. K.; Reynolds, J. R. J . Electroanal. Chem. InterfaclalElectrochem. 1988, 257, 307. (12) Ullevig, D. M.; Evans, J. F.; Albrecht, M. G. Anal. Chem. 1982, 5 4 , 2341. (13) (a) Koga, I.; Fukuyo. H. J . Instrum. Electr. Commun. Eng. Jpn. 1953, 36. 59. (b) Fukuyo, H.; Yokoyama, A.; Ooura, N.; Nonaka, S. Bull. Tokyo Inst. Tschnol. 1965, 72, 1. (c) Koga, I; Tsuzuki, Y.; Wln, S. N., Jr.; Bennett, A. L. Roc. Annu. Freq. Control Symp, 1960, 74, 53. (14) (a) van Dyke, K. S. Roc. Annu. Freq. Control Symp. 1957, 7 7, 000. (b) van Dyke, K. S. Roc. Annu. Freq. Control Symp. 1958, 10, 1. (15) Martin, B. A.; Hager, H. E. J . Appl. Fhys. 1989, 65, 2630. (16) Kanazawa, K. K.: Gordon, J. G., 11. Anal. Chem. 1985, 5 7 , 1770.
RECEIVED for review November 26, 1990. Accepted January 29,1991. This paper is contribution no. 5622 from the Central Research and Development Department (Du Pont).
Determination of Boron in Tissues and Cells Using Direct-Current Plasma Atomic Emission Spectroscopy Rolf F. Barth,* Dianne M. Adams, Albert H. Soloway, Eugene B. Mechetner,' Fazlul Alam? and Abul K. M. Anisuzzaman Department of Pathology and College of Pharmacy, The Ohio State University, Columbus, Ohio 43210 We have developed a safe, slmpie, and efflclent method for boron determlnatlon by means of dlreckurrent plasma atomic emlsslon spectroscopy. Tissues were soiubllized by using concentrated sulfuric acld and 70 % hydrogen peroxlde to dlgest the samples wlthout the need of hlgh temperatures and pressures. Boron duster compounds coukl be measured wlth sensitlvlty, preclslon, and accuracy slmllar to those of boric acld standards. Results obtalned with [(C,H5)3NH]2B12H12, Cs2Bl,H11SH*H,0, and ClSH,,Bl,O, show that thls analytical method ls applicable to a variety of compounds with different chemlcal structures. A sensltlvlty of 0.1 ppm has been obtained with known standards alone and In a varlety of tissue matrices includlng tumor, blood, liver, skin, and cell suspensions. The measurement of total boron by dlrect-current plasma atomlc emlssion spectroscopy (DCP-AES) has been achleved wlth as little as 50 mg of tissue or as few as 5 X lo7 cells. The procedure ls applkabk to the analysis of boron In the ppm range with a hlgh degree of precision and accuracy. Present address: Department of Genetics, University of Illinois, College of Medicine, 808 S. W o o d St., Chicago, IL 60612. Present address: US. Borax, 412 Crescent Way, Anaheim, CA
92801.
INTRODUCTION The accurate measurement of total boron content in biological samples with a sensitivity in the ppm range is essential for evaluating the potential usefulness of various tumorlocalizing boron-containing compounds for boron neutron capture theory (BNCT) ( I ) . Among the procedures that have been used is spectrophotometric analysis involving various complexing agents such as 1,l'-dianthrimide (2-4), methylene blue (5), and curcumin (6, 7). These methods are time consuming and require that the boron compounds be oxidized to boric acid. Relatively low sensitivity and interference from various contaminants are further limitations in the use of colorimetric assays. A second group of analytical procedures for boron are those involving nuclear methods. These include the detection of a particles resulting from the ' O B ( ~ , C Y , ~reaction ) ~ L ~ by means of a track autoradiography (8-11) and the measurement of y photons, by means of prompt y analysis (12,13). A major advantage in the use of nuclear methods is the fact that sample decomposition is not required. However, sample geometry is important and the a particles and y photons resulting from the 'OB(n,cu,~)~Li reaction are produced only from boron-10, which comprises 19.8% of natural elemental boron. The remaining 8O.2%, consisting of boron-11, is not detected by
0003-2700/91/0363-0890$02.50/00 1991 American Chemical Society
