2070
Anal. Chem. 1492, 64, 2070-2071
Direct Detection of Laser-Induced Capillary Vibration by a Piezoelectric Transducer Tamao Odake, Takehiko Kitamori, and Tsuguo Sawadat Department of Industrial Chemistry, Faculty of Engineering, The University of Tokyo, 7-3-1 Hongo, Bunkyo, Tokyo 113, Japan For its extremely high separation efficiency, with the theoretical plate number presently reaching nearly 106, capillary zone electrophoresis (CZE) is a powerful separation method, especially for biological materials.l-6 The method has another merit that sampling volume can be reduced to less than a nanoliter. This merit can be taken an advantage especially in ultramicroanalysis. However, conventional absorbance detectors are not sensitive enough for such small samplingvolumes due to the short optical path. Several highly sensitivespectrometric detection methods have been proposed as on-column methods, including direct7-l1 and indirect laser induced fluorometrylz-14 and a laser-induced photothermal beam deflection method.15-17 The authors have already proposed another highly sensitive detection method, i.e. capillary vibration induced by laser (CVL), and details of the signal generation and detection mechanisms have been given in the literature.l*Jg The lower limit of detection for this method was an absorbance of about 10-5. While, the detection limit of typical UV-vis absorbance detectors equipped with commercializedCZE instruments is about 10-3in absorbance. Therefore, the CVL method was about 2 orders of magnitude more sensitive than conventional absorbance detectors. The CVL method was applied to a detector for CZE, and underivatized amino acids of femtomole levelwere detected.19 In the previous reports, the capillary vibration was caused by local tension fluctuation of the capillary irradiated by an intensity-modulated excitation laser beam. Analytes in the capillary tube absorbed this laser beam and generated heat periodicallyaccordingto the photothermal effect, which made the capillary tension fluctuate locally. This vibration emitted an acoustic wave, which was detected by an optical beam deflection (OBD) method.20 In this case, the probe laser beam
* To whom all correspondence should be addressed.
(1) Cobb, K. A.; Novotny, M. Anal. Chem. 1989,61, 2226-2231. (2) Kennedy, R. T.; Oates, M. D.; Copper, B. R.; Nickerson, B.; Jorgenson, J. W. Science 1989,246, 57-63. (3) Drossman, H.; Luckey, J. A.; Kostichka, A. J.; D’Cunha, J.; Smith, L. M. Anal. Chem. 1990.62.900-903. (4) Cohen, A. S.; Terabe, 8.;Smith, J. A.; Karger, B. L. Anal. Chem. 1987,59, 1021-1027. (5) Ewing, A. G.; Wallingford, R. A.; Olefirowicz, T. M. Anal. Chem. 1989,61, 292A-303A. (6) McCormick, R. M. Anal. Chem. 1988,60, 2322-2328. (7) Cheng, Y.-F.; Dovichi, N. J. Science 1988,242, 562-564. (8) Amankwa. L. N.: Scholl.. J.:, Kuhr. W. G. Anal. Chem. 1990. 62. 2184-2193. (9) Sweedler, J. V.; Shear, J. B.; Fishman, H. A.; Zare, R. N.; Scheller, R. H. Anal. Chem. 1991, 63,496-502. (10) Swerdlow, H.; Zhang, J. Z.; Chen, D. Y.; Harke, H. R.; Grey, R.; Wu, S.; Dovichi, N. J.; Fuller, C. Anal. Chem. 1991, 63, 2835-2841. (11) Jorgenson, J. W.; Lukacs, K. D. Anal. Chem. 1981,53,1298-1302. (12) Kuhr, W. G.; Yeung, E. S. Anal. Chem. 1988,60, 1832-1834. (13) Kuhr, W. G.; Yeung, E. S. Anal. Chem. 1988, 60, 2642-2646. (14) Garner, T. W.; Yeung, E. S. J. Chromatogr. 1990,515, 639-644. (15) Yu, M.; Dovichi, N. J. Anal. Chem. 1989, 61, 37-40. (16) Bruno, A. E.; Paulus, A,; Bornhop, D. J. Appl. Spectrosc. 1991, 45,462-467. (17) Bornhop, D. J.; Dovichi, N. J. Anal. Chem. 1987,59,1632-1636. (18) Wu, J.; Kitamori, T.; Sawada, T. Anal. Chem. 1990, 62, 16761678. (19) Wu, J.; Odake, T.; Kitamori, T.; Sawada, T. Anal. Chem. 1991, 63,2216-2218. (20) Murphy, J. C.; Aamodt, L. C. J.Appl. Phys. 1980,51,4580-4588. .
I
passed just above the vibrating capillary and was deflected by the acoustic wave. As described in this report, the capillary vibration is directly detected by a piezoelectric transducer (PZT). The energy conversion process in this direct detection method does not involve energy transfer from mechanical vibration energy to acoustic wave energy, so this method is expected to be more sensitive than the OBD detection. Furthermore, for PZT detection, alignment of the probe beam is not required. Therefore, the instrumentation of PZT detector is much simpler than that of OBD detector.
EXPERIMENTAL SECTION The experimental arrangement was almost the same as in the previous r e p ~ r t . ~ *One J ~ of the holders which supported the capillary tube (50-pm i.d., 150-pm 0.d.) was replaced by a PZT disc (20 mm in diameter, 2 mm thick) as shown in Figure 1.The excitationbeam was an argon ion laser beam of 488-nmwavelength and 100-mWoutput power. The beam intensity was modulated by a light chopper. The beam was focused with a SO-mm focal length lens on the capillarytube, which was tensioned by a hanging weight. The generation mechanism of the capillary vibration was the same as before, and the capillary vibrated like a string between the two supports, one of which was the PZT detector. Mechanical vibration of the capillary was directly detected by the PZT detector. The PZT output was amplified by a 40-dB preamplifier and fed into an autophase lock-in amplifier. The liquid samples were aqueous solutions of sunset yellow dye (absorption peak at 482 nm). The sample concentrations were adjusted by stepwise dilution with distilled water to have to 1.7 X 10-2 absorption coefficients in the range of 1.7 X cm-l.
RESULTS AND DISCUSSION Typical examples of vibration signals obtained from the PZT detector are shown in Figure 2. The samples were distilled water and sunset yellow dye solution of 5.0 X 10-3 cm-1 absorption coefficient, which corresponded to absorbance of 2.5 X 10-5 for 50-fim4.d. capillary tubes. Stable signals were acquired for both samples. These results confirmed the principle of this photothermal detection method of optical absorption, as described in the previous experiments.18JgThe mechanical energy of vibration of the capillary was directly transferred to the PZT disc and converted into an electrical signal by the piezoelectric effect. Figure 3 shows frequency characteristics of the signal magnitude. The resonant peak appeared a t 604 Hz. The resonance corresponded well with the natural frequency of the capillary calculated18from the length of the vibrating part and the applied tension. This resonant peak also verified the string-like vibration of the capillary. Use of this resonance for analytical applications could not be recommended for the followingreasons. First, as pointed out previously, the signalto-noise ratio was not satisfactory at this resonant frequency, because noise induced by mechanical factors occurred at the natural frequencies of the vibration system. Second, the peak was too sharp (the full width of the half maximum was 7.5 Hz), and it was difficult to keep the resonating conditions.
0003-2700/92/0364-2870$03.00/0 0 1992 American Chemical Soclety
ANALYTICAL CHEMISTRY, VOL. 64, NO. 22, NOVEMBER 15, 1992
2871
Detection point
1-
Excitation beam I
I
\I
"
Capillary
I
P Z T disk
P Z T disk -
Vibration Brass cover
t Weight
From pump
-
Figure 1. Experimental arrangement of the direct detection system for CVL with a PZT detector. Insert: enlargement of the detector.
( b )
( a )
1 min
t I + t Laser O N OFF ON OFF Figure 2. Typical examples of signals. (a) Distilled water, and (b) sunset yellow dye solution of 2.5 X absorbance.
I
400
. .
-= I = 200 .-G v)
I. .* *
0
*
e
.
I
200
.
I
600 800 M o d u l a t i o n Frequency ( H z 1 Figure 3. Frequency characteristics of the signal magnitudes. 400
Therefore, a nonresonant frequency at 710 Hz was used in the following experiments. The dependence of signal magnitudes on dye concentration was measured. For 50-pm4.d. capillary, absorbances of the prepared samples corresponded to 8.4 X 10-6 to 8.4 X absorbance. The signal magnitudes showed linearities for absorbances of the sample as often occurs with the photothermal effect (correlation coefficient was 0.99), because
0
0
1
2
3
Time (min) Flgure 4. Electropherogram of riboflavlnobtained with the CVL method. Separation voltage was 20 kV.
absorbances of the samples were sufficiently low. The detection limit (S/N = 2) of absorbance was calculated as 8.5 X 10-7 for the 50-pm capillary. For sunset yellow dye solution, the lower limit of detection for concentration was calculated to be 6.8 X 10-9M. Considering the optical sampling volume estimated to be 1.0 X 10 L, the detection limit of absolute amount was 6.8 X 10-20 mol. Considering that the detection limit reported previously in the OBD method was 1.5 X 10-5 for the same capillary, the PZT detection was proved to be at least 1 order of magnitude more sensitive than the OBD method. Comparing the energy conversion process of the two detection methods of CVL, energy loss in the PZT detection process is smaller. Energy conversionsfrom opticalto thermal and to mechanic energy of vibration by the photothermal effect are common for both methods. However, the mechanical energy is directly converted to electric energy in the PZT detection, while there are two processes before the final conversion for the OBD detector, i.e. acoustic emission of vibrations and interaction between the acoustic wave and the probe beam. Therefore, the total energy loss in the PZT detecting process is considered to be smaller than for OBD detection, and this results in superior sensitivity. As the PZT detector is very sensitive to any electric field because it is electrically equivalent to the combination of a capacitor and a resistor, poor base-line stability and noise due to the separation current and electrophoresis potential are possible. However, for the experimental results (Figure 4)from applying this method as a CZE detector, no serious effect from the separation current was observed. In this experiment the sample was riboflavin (200-300 fmol) and the separation conditions were the same as in the previous report.19 In addition to the superior sensitivity, the direct detection method does not need optical alignment for probing. This results in favorable reproducibility and simpler instrumentation.
RECEIVED for review May 15, 1992. Accepted August 24, 1992. Registry No. Sunset yellow, 2783-94-0; riboflavin, 83-88-5.
