636
Anal. Chem. 1989,6 1 , 636-638
addition to the field of chemical sensor and biosensor development. Registry No. Pt, 7440-06-4; H202, 7722-84-1; ethidium bromide, 1239-45-8;luminol, 521-31-3.
LITERATURE CITED (1) (2) (3) (4)
Seitz, W. R. Anal. Chem. 1984, 56, 16A-34A. Peterson, J. I.; Vurek, G. G. Science 1984, 224, 123-127. VanDyke, D. A'; Cheng, H.-Y. Anal. Chem. '9889 60, 1256-1260. VanDyke, D. A. Ph.D. Thesis, University of Illinois, 1986. (5) VanDyke, D. A.; Nieman, T. A., submitted for publication in Anal. Chem . ( 6 ) Coleman, J. T.; Eastham, J. F.; Sepaniak, M. J. Anal. Chem. 1984, 56,2249-2251. (7) Le Pecq, J. B.; Paoletti, C. Anal. Blochem. 1966. 1 7 , 100-107.
(8) (9) (10) (11) (12) (13) (14) (15) (16) (17) (18)
Karsten. U.; Wollenberger, A. Anal. Biochem. 1972, 4 6 , 135-148. Karsten, U.; Wollenberger, A. Anal. Blochem. 1977, 7 7 , 464-470. Boer, G. J. Anal. Blochem. 1975, 65,225-231. Holl, W. W.; Webb, R. L., Smith Kline and French Labs, personal communication, 1988. Modern Fluorescence Spectroscopy, Wehry, E. L.,Ed.; Plenum Press: New York, 1976; Vol. 2. Angel, S. M. Spectroscopy 1986, 2, 38-47. Freeman. T. M.; Seitz, W. R. Anal. Chem. 1978, 50, 1242-1246. Kuwana, T. J. Nectroanal. Chem. 1963, 6 , 164. Epstein, B.; Kuwana. T. fhotochem. Photobiol. 1965, 4 , 1157. Epstein, B,; Kuwana, T, J , Electrmna/, Chem, 1967, 16, 3899, Hool, K.; Nieman, T. A. Anal. Chem. 1988, 60,834-837.
RECEIVED for review August 25, 1988. Accepted November 22, 1988.
Reproducible Nuclear Magnetic Resonance Surface Coil Fabrication by Combining Computer-Aided Design and a Photoresist Process Teresa W.-M. Fan*%' Nuclear Magnetic Resonance Facility, University of California, Davis, California 95616
Richard M. Higashi University of California Bodega Marine Laboratory, Bodega Bay, California 94923
INTRODUCTION The vigorous development of in vivo NMR spectroscopy and imaging has generated great interest for applications in biology and medicine. While improved electronics and sophisticated pulse sequences are being developed, the success of nuclear spin manipulation in in vivo NMR experimentation will depend, in part, on the efficiency of NMR probes in delivering the desired pulses. Thus, probe design needs to be optimized and constructed accurately for a given application. Winding copper wire has been the method of choice for making surface probes ( l ) while , tailored copper foil has been commonly used for high-resolution NMR probes. Generally, copper foil should be superior to wire as the coil material in probes due to the following: (a) lower inductance, which makes it easier to tune to the higher frequencies t h a t are increasingly common in in vivo applications; (b) larger conductive surface, giving rise to lower radio frequency (rf) impedance and a better Q factor (2);(c) better rf homogeneity, leading to better line shape. Although both wire and foil tape can be used for simple coil designs without much difficulty, they require specialized skills and tools when one is dealing with even slightly more complicated coil designs such as spirals or when they must be tailored to the shape of the sample. In this note, we report an alternative probe coil fabrication method that exploits recent low-cost advances in personal computers, rudimentary computer-aided design (CAD) programs, and high-quality graphics printers and combines them with the proven photoresistive etching process for printed circuit (PC) boards. This method is easy to implement, uses only commonly available materials and equipment, and offers the flexibility and precision needed for the construction of considerably more complex coil designs. Although constructing NMR surface coils from PC boards by etching (3)and machining (2)has been reported, these two methods were limited to available templates or special skills and tools. The combination of CAD and photoetching of PC boards eliminates these technical requirements and provides the following advantages in addition to the inherent advantages of copper foil:
' Current address: Department of Environmental Toxicology, University of California, Davis, CA 95616.
(a) Coils of any shape and complexity can be readily fabricated to scale for a given application. (b) Theoretical designs can be rapidly and accurately made into real probes for testing. (c) Coils can be made with high precision such that even slightly different coil designs can be fabricated with ease for empirical optimization of performance. (d) Reproducible coils can be made by photocopying a design directly from the pages of a journal such as this. (e) The tight binding of copper on plated boards reduces microphonics or other mechanical ringing that can occur with free-standing wire coils (4).
EXPERIMENTAL SECTION The procedure for fabricating photoetched coils was straightforward. Coils were drawn with Cricket Draw program (Cricket Software, Malvern, PA) running on a Macintosh Plus computer (Apple Computer, Inc., Cupertino, CA) and printed onto transparencies by a 300 dots-per-in. (dpi) resolution Laserwriter Plus (Apple). There were no freehand drafting skills involved; in this case, spiral coils were created by using the starburst and radial grate tools of Cricket Draw. If a high-resolution graphics printer is not available, coil designs could be printed oversized with a lower resolution printer and reduction-copied onto transparencies by using a photocopier, the entire process being scaled such that the resulting transparency is 300 dpi or better. Photosensitized single-sided PC boards (35.6 wm thick, 3 in. X 4 in. or 3 in. X 6 in., positive type, GC Electronics, Rockford, IL) were then exposed under the transparencies according to the manufacturer's recommendation. We found that an exposure time of 20 min at a distance of 6-7 in. from a General Electric (GE) Fl5BL ultraviolet lamp was sufficient. After removal of the photoexposed material with the developer (GC Electronics) according to the manufacturer's instruction, the boards were etched with potassium persulfate in deionized H20at 40-50 OC with stirring. Although we avoided the more commonly employed FeC1, etchant because of possible paramagnetic contamination, we experienced no consistent difficulty with using it. After etching was complete, the unexposed photoresist coating was removed with stripping solution (GC Electronics) or ethanol. The coils were then turned to desired frequencies and matched to 50-R impedance to complete the probe construction. We have chosen the balanced matching circuitry approach ( 5 ) for better Q factor and performance on conductive samples. The variable capacitors used were 1-10- and 1-30-pF nonmagnetic air piston
0003-2700/89/0361-0636$01.50/0 1989 American Chemical Society
ANALYTICAL CHEMISTRY, VOL. 61, NO. 6, MARCH 15, 1989
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637
ARG-P
V
b 1H
PPM
85.5MHz
13C 21.5
MHz
'
1
0 -20 -40 -60 I n vivo 31P NMR spectra of a marine mussel (Mytilus Californianus) taken with (a) photoetched foil probe and (b) commercial wire probe. Both spectra were taken with visual localization on the adductor muscle of the mussel, using a pulse width of 15 ks, interpulse delay of 1 s, sweep width of f2500 Hz, 2K data points, and 2048 transients and were processed by using a line broadening factor of 15 Hz before Fourier transformation. Flgure 2.
2
Flgure 1. Photoetched 13C-'H double coils. (a) Coil designs and actual sizes: 'H, 6.3 cm: I3C, 3.5 cm. (b) Circuit diagram: C,, C3, and C, = 11, 24 + 5.6, and 200 10 pF, respectively: C, and C, = 0.6-6 pF: C,, C,, and C8 = 1-30 pF.
1
+
capacitors from Johanson (Boonton, NJ) while ceramic chip capacitors with fixed capacitance were purchased from American Technical Ceramics (Huntington Station, NY). For comparison purposes, we constructed a 2-cm, 5-turn 31P planar spiral probe identical in coil configuration with a commercial wire probe provided with a GE CSI-2T system. The tuning and matching circuitry was photoetched on a separate PC board (2 in. X 4 in.), which was mounted with variable capacitors and attached to the coil board with 22-gauge copper wires. The distance between the coil and capacitor boards was at least 2 cm. We also employed the photoetch fabrication technique to construct a 3.5-cm, 2-turn planar spiral 13Cprobe with a concentric 6.3-cm, 2-loop 'H decoupling coil as shown in Figure la, which displays the actual coil sizes. The 'H coil was of a modified planar concentric loop-gap resonator design (6),which allowed easier tuning of larger coils to the 'H frequency and should exhibit some intrinsic rf isolation from the 13Ccoil (6). The width of the copper foil and the spacing between turns were adjusted to be equal (Figure la), and a h / 4 cable was included in the circuit (Figure l b ) to reduce noise resulting from 'H broad-band decoupling procedures (2). All experiments were performed with the GC CSI-2T system, which has a 200-mm horizontal bore magnet. Reflectance was measured with a Wavetek Model 1061 sweep generator and a Tektronix Model 2465 oscilloscope, from which the Q factor was calculated (7).
RESULTS AND DISCUSSION Table I lists the Q factor and performance of photoetched and commercial wire 31Pprobes on standards. T h e performance of these two probes was also compared on a conductive in vivo sample, a marine mussel (Mytilus californianus) collected from Bodega Bay, CA, as illustrated in Figure 2. It is clear that the commercial wire probe performance was easily
6
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-2000
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FREQUENCY (HZ) Natural abundance in vivo 13C NMR spectrum of avocado. The spectrum was taken by using a pulse width of 15 ps, an interpulse delay of 0.2 s, a sweep width of zt2500 Hz, 4K data points, and 256 transients. Broad-band decoupling was implemented by using the MLEV-64 decoupling scheme with a 90' 'H pulse width of 197 ps. The measured decoupling power during acquisition and interpulse delay was 3.5 W and 400 mW, respectively. The spectrum was processed with Gaussian weighting of 10 Hz before Fourier transformation. The spectral assignments are as follows: 1, terminal methyl carbon of lipid fatty acid chains; 2, methylene carbon of lipid fatty acid chains: 3 and 4, lipid glycerol backbone: 5, polyunsaturated carbon of lipid fatty acid chains; 6, monounsaturated carbon of lipid fatty acid chains: 7, carbonyl carbon of lipid fatty acid chains. Flgure 3.
matched by the CAD/photoetched probe. The photoetched 13C coil had unloaded and loaded Q factors of 307 and 239, respectively, while the concentrically etched 'H coil exhibited unloaded and loaded Q factors of 204 and 68, respectively, with use of the same phantom as in Table I for the loaded Q. T h e line width obtained on the methyl resonance ('H decoupled) of 95% ethanol was typically 2-3 Hz. The in vivo performance of this 13C-'H probe was tested on an avocado (purchased from a local market). A broad-band
638
Anal. Chem. 1989, 6 1 , 638-640
T a b l e I. Comparison of Performance between Wire Probe and Photoetched Probe
coil
Q(un1oadedIa
Q(1oadedIb
photoetched probe wire probe
266 256
231 192
optimal pulse width,‘ SINd LWe 15 16
42 27
3.0
4.2
‘Q = 2n(fo/Af0,71).where fo denotes resonance frequency; b.fo,71, bandwidth at 0.71 peak height. * Q estimated with a phantom containing 100 mM bicarbonate, 10 mM ATP, 20 mM PCr, 1 mM EDTA, 1% NaN,, and 3.7% gelatin, pH 7.3. CMeasuredwith peak heights of a single scan by using a standard containing 100 mM phosphoric acid. dThe ratio of the peak height to rms noise on one scan of 100 mM phosphoric acid. eApproximate line width of 100 mM phosphoric acid in hertz. decoupled natural abundance 13C spectrum of the avocado is shown in Figure 3. The ‘H decoupling probe employed a modified concentric loop-gap resonator design for efficient proton decoupling, although this use of the coil design has not been previously reported. I t is clear that considerable flexibility and precision are inherent in this fabrication technique in terms of coil sizes, shapes, width of copper foil, and interturn spacing. For example, in cases where a coil conforming to the sample shape (e.g. kidney) is desired, a photograph of the sample can be digitized as a bitmap into the computer by using a common optical scanner, and the coil can be drafted accurately on the basis of the sample profile by tracing the bitmap from the
CAD program. As many CAD programs are also capable of producing drawings based on complex calculations, this fabrication scheme can reduce the gap between theoretical designs and actual testing of such probes. Finally, although we demonstrated this fabrication techniuqe on planar coils, it can be readily extended to three-dimensional coils such as Helmholtz or solenoid by turning to three-dimensional CAD programs capable of “unwrapping” a three-dimensional design for two-dimensional transparency printing, and by wrapping the transparency over cylindrical or flexible PC boards (3) to photoetch. Perhaps the most rewarding probe designs involve the testing of coils that integrate all three approaches, a task facilitated by the fabrication method described here. Registry No. ATP, 56-65-5; Cu, 7440-50-8.
LITERATURE CITED (1) Gadian, D. G. NMR and Its Applicatbns to Living Systems; Oxford University Press: Oxford, England, 1982. (2) Tiffon, B.; MispeRer, J.; Lhoste, J-M. J . Magn. Reson. 1988, 68, 544. (3) Barker, P.; Freeman, R. J . Magn. Reson. 1085, 6 4 , 334. (4) Fukushlma, E.: Roeder, S. E. W. €xper/menta/ Puke NMR. A Nuts and Bolts Approach; Addison-Wesley Publishing Co.: London, 1981; pp 379, 380. (5) Murphy-Boesch, J.; Koretsky, A. P. J. M g n . Reson. 1083, 5 4 , 526. (6) Hyde. J. S.; Froncisz, W.; Jesmanowicz. A,; Kneeland, J. B. Med. Phys. 1987, 13, 1. (7) Chang, H.; Chew, W. M.; Weinstein, P. R.; James, T. L. J . Magn. Reson. 1087. 72, 168.
RECEIVED for review October 12,1988. Accepted November 28, 1988. This work was supported in part by NIH Grant P41RR02479.
Coupled Ion-Selective Electrode Measurement of Aqueous Carbonate and Bicarbonate Ion Activities John D. Pigott’ Laboratoire de Geologie d u M u s e u m National d’Histoire Naturelle, 43 rue de B u f f o n , 75005 Paris, France, and School of Geology and Geophysics, University of Oklahoma, 100 East Boyd, Norman, Oklahoma 73019 INTRODUCTION Previous development of carbonate and bicarbonate ion selective electrodes has been hindered either by poor selectivities (e.g. chloride interference: (1-3)) or by restricted pH ranges (C8.5 for 4 X M COS2-for the membrane electrode of Herman and Rechnitz ( 4 ) ) . Nonetheless, it is surprising that the potential uses and demands for these electrodes have not as yet led to their commercial manufacture, especially with the increasing applied interest in the field of ion selective electrodes ( 5 ) . Reported herein are the theoretical development and initial experimental results of carbonate and bicarbonate ion electrodes based upon a novel use of available products.
dissociation constants, etc. (6))works well in open oceans where major ions are fairly conservative with respect to chlorinity. However, the surface and pore waters of estuarine to normal marine to hypersaline natural systems are not amenable to such assumptions of major ion conservation. In these cases, conventional wisdom dictates a total chemical analysis with subsequent calculations for ionic strengths and activity coefficients. However, much of the tedium, calculative procedure, and incumbent error by measuring concentrations and then indirectly assessing carbonate ion activities could be minimized by using the coupled electrode technique. First, we need K1 and K2,the first and second dissociation constants of carbonic acid, respectively (7), a t 25 “C
K, = a H C O s ~ H + / a H 2 ~ 0 ,=p K2 = ~ c o ~ ~ ~ H + /=~ 10-10.329 Hco~-
THEORETICAL DEVELOPMENT Mass Action Fundamentals. Research in field aquatic systems, principally with carbon dioxide dynamics, requires the determination of the activities of the carbonate ion. This ability to quantify C032-is fundamental to an ability to resolve saturation states of lagoons, open oceans, and pore waters with respect to a variety of calcium carbonate phases, e.g. calcite, aragonite, and a spectrum of magnesian calcites. The classic oceanographic approach (titration alkalinity, pH, apparent
(2)
where (7) H2C03* = H 2 C 0 3 + C02(dissolved)
(3)
and a represents activity. Second, by combining eq 1 and 2, one has aco32- =
’Present address: University of Oklahoma.
(1)
K & I ~ H ~ C O ~ * / ~ H + ~ (4)
That is, measuring the activities of the hydrogen ion and
0003-2700/89/0361-0638$01.50/0 0 1989 American Chemical Society
