Anal. Chem. 1991, 63,2082-2086
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Microenvironments of Fluorescence Probes in Sodium Taurocholate and Sodium Taurodeoxycholate Bile Salt Media Steven M. Meyerhoffer and Linda B. McGown*
P. M. Gross Chemical Laboratory, Department of Chemistry, Duke University, Durham, North Carolina 27706
Measurements of fluorescence vlbronic band Intensity ratios and lifetimes of polycyclic aromatic hydrocarbon (PAH) probes, including pyrene, phenanthrene, trlphenyiene, benLO[ e Ipyrene, and benzo[gh/]perylene, were used to study binding sites In organized bile salt media formed by sodium taurocholate (NaTC) and sodlum tawodeoxycholate (NaTDC). Comparisons with similar measurements of the PAH probes In simple solvents and In micellar solutions of sodium dodecyl sulfate (NaDS) indicate that the binding environments in NaTDC are most apolar. The NaTC mkeRar soiutbns provide the greatest microenvironmental diversity between probes, with molecular structure and size of the probe molecules affecting the observed solublllzation behavior. Metal salts were added to the NaTC solutions to modify and control behavior, thereby influencing the soiubilizatlon mlcroenvlronments of the probes In the media.
INTRODUCTION Bile salt aggregation in aqueous solution has been an active area of research, with primary emphasis on the physiological role of bile salts in lipid solubilization (1,2). More recently, bile salt media has attracted attention as an alternative to conventional micellar reagents for chemical analysis, including applications in separations (3)and luminescence analysis (4). Because the molecular structure of bile salts is very different from that of conventional detergents, the bile salts exhibit unique behavior with respect to self-association and molecular solubilization (5-8). In conventional detergents, solubilization sites include the micellar surface, the palisade layer, and the hydrophobic inner core (9). Analogous binding sites are not present in the smaller, more rigid bile salt micelles. Instead, solubilization of hydrophobic compounds is accomplished through favorable interactions with the hydrophobic surfaces of the bile salt micelles (5,10, II), primarily at the CISand CI9sites ( 1 0 , I I ) . The resulting solubilization microenvironments in bile salt micelles are highly apolar. In this study, five polycyclic aromatic hydrocarbon (PAH) fluorescence probes were used to study solubilization microenvironments in micellar solutions of two bile salts, sodium taurocholate (NaTC) and sodium taurodeoxycholate (NaTDC), and a conventional detergent, sodium dodecyl sulfate (NaDS). The PAH probes included pyrene, phenanthrene, triphenylene, benzo[e]pyrene (BeP), and benzo[ghi]perylene (BgP). Measurements of vibronic band intensity ratios and fluorescence lifetimes of the probes yielded information on binding site differences between the micellar media. Effects of metal salts on probe microenvironments in NaTC were also studied.
EXPERIMENTAL SECTION The fluorescence probes, metal salts, bile salts, and detergent were all purchased from commercial vendors and used as received. Stock solutions of the fluorescence probes, including pyrene and BeP (99%, AccuStandard), phenanthrene and BgP (99%, U1-
* Corresponding author. 0003-2700/91/0383-2082$02.50/0
trascientific), and triphenylene (98%, Aldrich), were prepared in ethanol. Stock solutions of NaTC (ULTROL Grade, >98%, Calbiochem), NaTDC (Sigma),and NaDS (puriss, >99%, Fluka) were prepared fresh daily by dissolution of the compounds in water. Metal ions, including magnesium(II), aluminum(III), and u the nitrate salts (99.999%,Aldrich). terbium(III), were added t Spectrophotometric or HPLC grade solvents were used for all solution preparations. Solutions of pyrene (0.5 pM),phenanthrene (2.0 pM), triphenylene (2.0 pM), BeP (2.0 pM), and BgP (2.0 pM) in micellar media were prepared by evaporating the ethanol from an appropriate volume of a stock solution of the probe compound in ethanol using a gentle stream of nitrogen, followed by dissolution of the remaining solid in aqueous micellar solution in a volumetric flask and sonication for at least 1 h. The solutions were not deoxygenated prior to measurement. Fluorescence measurements were made with an SLM 48000s multifrequency, phase-modulation spectrofluorometer (SLM Instruments, Inc., Urbana, IL) using a 450-W xenon arc lamp source, excitation and emission monochromators for wavelength selection, and photomultiplier tube detectors. All measurements were made at 25.0 & 0.1 "C. Fluorescence emission spectra were collected at 1-nm scanning intervals in the "5-average" mode, where each measurement is the average of 5 samplings over a 1 s interval. Pyrene was excited at 335 nm, phenanthrene at 293 nm, triphenylene at 284 nm, BeP at 331 nm, and BgP at 362 nm. An excitation bandwidth of 2 nm was used for pyrene, BeP, and BgP, while a 4-nm bandwidth was required for phenanthrene and triphenylene in order to achieve adequate fluorescence intensities. An emission bandwidth of 2 nm was used for all measurements. Fluorescence lifetimes (q) were determined from phase and modulation measurements collected at three or more modulation frequencies. Fluorescence excitation (Aex) and emission (A,) wavelengths were as follows: pyrene b, = 335 nm, X,= 382 nm; phenanthrene A,, = 293 nm, A,, = 363 nm; triphenylene A,, = 284 nm, A, = 352 nm; BeP X, = 331 nm, X,= 387 nm; and BgP A,, = 381 nm, A,, = 418 nm. The measurements were made by using a 100-average setting, where each measurement is the internally performed average of 100 samplings over a 30-s period. T = 0)~ was used ~ An intensity-matched scattering solution ( BS the lifetime reference. Measurement of fluorescence lifetimes in the frequency domain has been described elsewhere (12). Each vibronic band ratio and lifetime value reported here is the mean of anywhere from two to seven individual measurements (three t o four replicates were used in most cases). In all cases, replicates were obtained on different days, often spanning a period of weeks or months, using freshly prepared solutions. Dimensions of the PAH fluorescence probe molecules were calculated by using SYBYL Molecular Modeling Software (v. 5.3, Tripos Associates, Inc., St. Louis, MO).
RESULTS AND DISCUSSION The chemical structures of the PAH fluorescence probes and their molecular dimensions are shown in Figure 1 and Table I, respectively. The length (y) of each probe is approximately 9.1 A. The width of each probe was calculated a t three separate points along the y axis; x1 and x 3 were calculated along their respective axes, while x 2 was calculated through the region of greatest dimension occupied by the central ring($ Aqueous solubilities of the probes (13),which span a range of almost 4 orders of magnitude, are also shown in Table I. Although the five PAH probes were selected primarily on the basis of reported microenvironmental sen0 1991 American Chemical Society
~
~
ANALYTICAL CHEMISTRY, VOL. 63, NO. 19, OCTOBER 1, 1991 4
Y
4
4
Y
Therefore, although it is interesting and useful to compare band ratios for probes in binding environments to those ratios obtained in simple solvents, care must be taken not to interpret the results simply in terms of polarity. We are currently engaged in further studies of the factors that determine vibronic band intensity ratios in binding environments.
Y
w f
1....................
I ....*
u* ..I... .........
Pyrene
..........*
..........L
I1
=1
Xl
Phenanthrene
4
Triphenylene
,““1“1 ’
7
@ . * x ,
................................. ..)... ......... *
c
Xl
Benrolelp yrene
.............................
*
It
.................. L Xa
Bensotghllper~lene
Flgure 1. Chemical structures of pyrene, phenanthrene, triphenylene, benro [e ] pyrene, and benzo [ghi]perylene fluorescent probes.
Table I. Molecular Dimensions and Aqueous Solubilities of the PAH Fluorescent Probes molecular dimens, A PAH compd
solubility,’rM
y
xl
x2
x3
pyrene phenanthrene triphenylene benzo[e]pyrene benzo[ghi]perylene
0.64 6.46 0.19 0.029 0.00094
9.1 9.1 9.1 9.1 9.1
4.3 4.3 4.3 4.3 6.7
6.7 5.6 7.9 9.1 7.9
4.3 4.3 4.3 4.3 6.7
a
2083
Solubilities from ref 13.
sitivities of their vibronic band ratios (14-17),it is also important that they cover a wide range of molecular sizes and aqueous solubilities in order to study differences among their respective binding environments. Solvent-Sensitive PAH Fluorescence Emission. The fluorescence emission spectra of the probes in acetonitrile and cyclohexane are shown in Figure 2. These uncorrected spectra are in general agreement with those published by others (14-16). The spectra for each probe were normalized to yield constant intensity at the least microenvironmentally sensitive band: band I1 for phenanthrene, band I11 for pyrene, triphenylene, and BgP, and band IV for BeP. These bands correspond to allowed transitions and show little change in intensity as compared to the vibronically forbidden 0 transition of band I. The intensity of band I is weak in nonpolar solvents and increases in polar solvents due to specific solute-solvent interactions, such as dipole-dipole coupling, which serve to reduce the symmetry of the molecular probe (1516). Intensity ratios are reported in this paper as the intensity of the forbidden 0-0 transition relative to the intensity of an allowed transition. The calculated ratios will therefore increase with increasing solvent polarity, and can be used to investigate the polarity of the probe microenvironment. Fluorescence probe molecules have been used to study microenvironments in organized media (18),including micelles and cyclodextrins, as well as in the study of the structure of reversed-phase chromatographic stationary phases (19). Effective polarities of microheterogeneous systems are typically inferred from macroscopic values of intensity ratios and lifetimes observed in simple solvents. It is important to recognize, however, that it is not polarity of the solvent per se that affecta transition probabilities (and therefore intensity ratios), but rather the specific interactions that occur between the molecular probe and the solvent cage (the enuironment).
Intensity Ratios in Simple Solvents and Aqueous Micellar Solutions. It is difficult to compare vibronic band intensity ratios obtained in different laboratories due to variations in instrumental and experimental conditions. It is therefore important for each group to generate its own relative polarity scale or “ruler”, by using solutions of probes in simple solvents. The ratios obtained in this work for the five PAH compounds in simple solvents are shown in Table 11. The values obtained in aqueous micellar solutions of NaTC, NaTDC, and NaDS are inserted at the appropriate positions along the ruler for each probe. Values for the probes in NaTC with added metal ion (5 mM Mg2+,A13+,or Tb3+) are also inserted along the rulers. The concentration of bile salt or detergent in all micellar solutions was 20 mM (calculated as monomer), which is sufficiently above the cmc values to provide complete solubilization of the probes. The rulers shown in Table I1 are not linear, and the positioning of the inserted data for the micellar solutions is only a relative indication of polarity with respect to each other and to the simple solvents. The absolute standard deviations in the intensity ratios were determined to be hO.01 for pyrene, f0.08for phenanthrene, f0.02 for triphenylene, f0.03 for BeP, and hO.01 for BgP. Thus, the small differences among values for micellar solutions that are clustered in similar positions on a ruler are generally not significant for phenanthrene, triphenylene, and BeP, which had the largest relative standard deviations. Phenanthrene and triphenylene also show the smallest sensitivity to solvent polarity. Comparison of the probe intensity ratios indicates that the bile salts and the detergent provide different microenvironments for the solubilized probe molecules with the exception of triphenylene, which showed no significant differences in polarity, all values falling within the methanol-propanol range. Of the three micellar systems, NaTDC yields the lowest ratios for the solubilized probes. The ratios obtained for pyrene, phenanthrene, BeP, and BgP fall between the values observed in 1-decanol and cyclohexane. The intensity ratios are significantly greater in NaDS, matching the values found in more polar solvents: pyrene and phenanthrene ratios are similar to those in ethanol, and the BeP ratio is similar to that in methanol, while the BgP ratio is similar to that in 2-propanol. In contrast to NaTDC and NaDS, the binding environments in NaTC show much greater diversity between probes, ranging from polar for BeP and BgP to relatively apolar for pyrene and phenanthrene. Differences among the solubilization microenvironments in NaTC, NaTDC, and NaDS arise from fundamental structural differences between the micellar aggregates of the three surfactants. Dynamic light-scattering measurements have indicated that micelles of NaTC are much smaller than those of NaTDC (20,21).Also, different models of aggregation have been suggested for the two bile salt micelles; back-toback, hydrophobic interactions have been proposed in the formation of micelles of NaTC (22),while helical (23,24)and rodlike (21,25) structures have recently been proposed for micelles of NaTDC. The additional hydroxy group of trihydroxy NaTC compared to dihydroxy NaTDC is the source of these differences, increasing the solubility of the NaTC monomer and influencing the aggregate structure (1). The NaDS monomers, in contrast to the bile salts, self-associate to form spherical micelles of large aggregation number (approximately 62) in which the hydrocarbon tails point inward
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ANALYTICAL CHEMISTRY, VOL.
03, NO. 19, OCTOBER 1, 1991
(A) pyrene
I
350
370
390
4iO
330
450
430
350
370
390
410
430
450
475
500
Emission Wovelength (nm)
Emission Wovelength (nm)
(C) triphenylene
111
350
375
A00
425
450
Emission Wovelength (nm)
Emission Wovelength (nm)
380
400
420
440
460
480
500
520
Emission Wovelength (nm)
Figure 2. Fluorescence emission spectra of the five PAH probes in acetonitrile (-) and cyclohexane (---): (A) pyrene; (B) phenanthrene; (C) trlphenylene; (D) benzo[e]pyrene (BeP); (E) benzo[ghi]perylene (BgP). range of the probes used in this study. In contrast to NaTC to form a hydrophobic core and the ionic head groups from and NaTDC, solubilization of PAH compounds in NaDS often a hydrophilic outer surface in contact with the aqueous sooccurs in the micelle palisade layer (9),allowing interaction lution (26). with ionic groups of the NaDS micelle and resulting in high In addition to the observed differences in the solubilization intensity ratios (18). The binding environment polarities microenvironments due to structural differences between the reported here for the probes in NaDS and in simple solvents micellar hosts, the microenvironments experienced by the agree well with previous studies (5, 14-17). probe molecules are also influenced by the physical properties Metal Salt Effects on Probe Intensity Ratios in NaTC. of the probes themselves. For example, in NaTC the smaller The addition of 5 mM nitrate salts of Mg2+,A13+,and Tb3+ probes (pyrene and phenanthrene) experience relatively apolar to the 20 mM NaTC solutions was observed to decrease the microenvironments, whereas the larger probes (BeP and BgP) vibronic band intensity ratios in the order (least decrease) experience relatively polar microenvironments. While both Mg2+< A13+< Tb3+(most decrease) (Table II). The increased NaTC and NaTDC are capable of providing apolar binding hydrophobic character of the binding environments in the sites to hydrophobic molecules by solubilizing the molecules presence of the metals is consistent with our previous studies between the hydrophobic surfaces of the bile salts, larger molecules solubilized in NaTC micelles may protrude into the of metal salt enhancement of NaTC aggregation (27). Measurements of fluorescence anisotropy of the benzo[k]aqueous solution or disrupt the structure of the NaTC micelle. fluoranthene probe in micellar solutions of NaTC have inSuch effects would account for the high intensity ratios obdicated that metal ions modify the NaTC aggregates, inserved for BeP and BgP in NaTC. Aqueous solubilities of the creasing the rigidity of the probe binding environment in the probes do not appear to significantly affect solubilization order (least increase) Na+ < Mg2+< A13+ < Tb3+ (most inbehavior in the bile salt media, at least over the solubility
ANALYTICAL CHEMISTRY, VOL. 63, NO. 19, OCTOBER 1, 1991
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Table 11. Vibronic Band Intensity Ratios of the PAH Fluorescent Probes
solvent
pyrene (I/III)
phenanthrene (I/II)
intensity ratio triphenylene ((I + II)/III)a
BeP (I/IV)
BgP (I/III)
water acetonitrile
1.88
1.70 1.70
b
b
b
1.84
0.46
1.41 NaTC 1.19
methanol
1.40
1.69
0.41 NaDS 0.41
1.25 NaDS 1.21
1.66 NaDS 1.66
0.40
1.64 NaTC 1.57 Mg*+-NaTC 1.48 1.40 A13+-NaTC 1.39 Tb3+-NaTC 1.38 1.34 NaDS 1.27
1.12
1.60
1.03 NaTC 1.03 Mg2+-NaTC 0.97 AI3+-NaTC 0.93 Tb3+-NaTC 0.92 0.89 NaTDC 0.77
1.56 NaTC 1.55
0.40 NaTDC 0.40 A13+-NaTC 0.40 NaTC 0.39 Mg2+-NaTC 0.39 Tb3+-NaTC 0.38 0.37
ethanol 2-propanol
tert-butyl alcohol
1-decanol
0.59 0.58
cyclohexane n-heptane
1.53 Mg2+-NaTC 1.53 A13+-NaTC 1.53 Tb3+-NaTC 1.52 NaTDC 1.44 1.43 1.34
1.11
NaDS 1.10 Mg2+-NaTC 1.09 1.06 AP+-NaTC 1.02 Tb3+-NaTC 1.00 0.90
1.26
0.76
1.17
0.36
0.72 NaTDC 0.60
1.00 NaTDC 0.76
0.36 0.34
0.47 0.36
0.49 0.48
"Bands I and I1 overlap (see ref 15). bRatios could not be determined due to limited solubility of the probes in water. Table 111. Fluorescence Lifetimes of the PAH Probes solution"
pyrene (f2 ns)b
ethanol water NaDS NaTC Mg2+-NaTC A13+-NaTC Tb3+-NaTC NaTDC
17.3 103 150 263 280 299 306 329
TF, ns phenanthrene (f4 ns)b triphenylene (f2 ns)b
BeP (f2 ns)b
BgP (&l ns)b 18.9
10.8
12.5
14.9
39 42 46 46 42 47
30 31 32 30 33 34
45 51 53 54 54 55
80 95 95 98 98 101
" Micellar solutions 20 mM, metal ions 5 mM. Absolute standard deviations for aqueous micellar solutions. crease) (27). Because NaTC provides a diverse range of microenvironments for the solubilized probe molecules, it is not unreasonable to expect the metal ions to affect the probe microenvironments to different extents. It is difficult, however, to compare relative changes in hydrophobicities of binding environments between probes from measurements of intensity ratios alone since the intensity ratio scales of the five probes are not linearly correlated to one another. Fluorescence Lifetimes. The fluorescence lifetimes of the probe molecules in micellar solutions of NaDS, NaTC (with and without added metal salts),and NaTDC are shown in Table 111. For comparison, the lifetimes for the probes in ethanol and for pyrene in water are shown as well. The absolute standard deviations reported in Table I11 are for the probes in aqueous micellar solution; in ethanol, where the lifetimes are shorter, the absolute standard deviations are expected to be much smaller. The lifetimes of the probes in ethanol (and, for pyrene, in water) are relatively short because the solutions were not deoxygenated. The fluorescence lifetimes increase in micellar solutions, attributed to increased structural rigidity in the micelle and reduction of collisional interactions with dissolved
oxygen and other components in the bulk solution, among other factors (28). Therefore, the consistent observation of longest lifetimes in NaTDC provide further evidence for a strongly apolar, well-protected binding environment in that media. For all of the probes, the lifetime in NaTC falls between those in NaDS and NaTDC. Pyrene has the longest lifetime and shows the greatest range of values. Phenanthrene and triphenylene show little difference in lifetime among the bile salts, reflecting their relatively short lifetimes and low microenvironmental sensitivity. BeP and BgP show trends in lifetime similar to that of pyrene; for these three probes, the lifetime in NaTC is 60-70% of the difference from the lifetime in NaDS to that in NaTDC. The fluorescence lifetime of pyrene in NaTC is significantly longer in the presence of Mg2+,A13+,and Tb3+,supporting the idea of metal cation enhancement of NaTC aggregation. The lifetimes of BeP and BgP may also reflect the same trend but the differences are much smaller. No significant trends were observed for phenanthrene and triphenylene lifetimes in the NaTC media, which may simply reflect the lower sensitivity of these probes to their microenvironments.
Anal. Chem. 1891, 63, 2086-2090
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CONCLUSIONS These studies show that the microenvironments of solubilized probes in bile salts are very different from those in conventional detergents. The bile salts are capable of providing rigid, apolar binding environments to solubilized probes, whereas the binding environments in NaDS are more fluid and polar. Moreover, the microenvironments of the solubilized probes show a much greater probe-to-probe diversity in NaTC than in either NaTDC or NaDS and are much more influenced by the structure and size of the probe itself. This is consistent with the smaller size and aggregation number of the NaTC micelles, where the smaller probes are more easily isolated in well-protected, apolar binding environments than are the less readily accommodated larger probes. Metal salts enhance the aggregation behavior of NaTC and could provide a means for selectively influencing the microenvironments of hydrophobic molecules in the media.
LITERATURE CITED (1) Small, D. M. In The BUe AcMs, Nair, P. P., Krltchevsky, D., Eds.; P i s num Press: New York, 1971;Vol. l., p 249-356. (2) O'Connor, C. J.; Wallace, R. G. A&. CokM Interface Sci. 1985, 22, 1-111. (3)Buckiey, J. J.; Wetleufer, D. B. J . Chrometcgr. 1989, 464, 61-71. (4) McGown, L. 6.; Krelss, D. S. Roc. SPIE-Int. Soc.Opt. Eng. 1988, 910. 73-78. (5) Zana, R.; Guvell, G. J . Phys. Chem. 1985, 8 9 , 1687-1690. (6) Fisher, L.; Oakenfull, D. Aust. J . Chem. 1979, 32, 31-39. (7) Chen, M.; Gratzel, M.; Thomas, J. K. J . Am. Chem. SOC. 1975, 97, 2052-2057. (8) Sugihara, G.; Yamakawa, K.; Murata, Y.; Tanaka, M. J . Phys. Chem. 1982, 86, 2784-2788. (9) Rosen, M. J. Surfactants and Interfaclel phenomena, 2nd ed.;John Wlley 8 Sons: New York, 1988.
(10) Conte, G.; DI Blasl. R.; Giglb, E.; Panetta. A. Pavel, N. V. J. Phys. Chem. 1984, 88, 5720-5724. (11) Kolehmainen, E. J . CdlOM Interface Sci. 1985. 705, 273-277. (12) Lakowicz, J. R. prlnc@les of Fluwescence Spectroscopy; Plenum Press: New York, 1983. (13) Pearlman, R. S.;Yalkowsky, S. H.;Banerjee, S. J . Phys . Chem. Ref. Data 1984, 73,555-562. (14) Nakajima, A. Photochem. Photoblol. 1977, 2 5 , 593-598. (15) Nakajima, A. J . Lumin. 1974. 8 , 266-269. 116) Kalvanasundaram. K.: Thomas. J. K. J . Am. Chem. Soc.1977, 99, 2039-2044. (17) Mukhopadhyay, An'l-AlI A. K.; Georghiou, S. Photochem. Photobld. 1980, 31, ,
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(18)Kalyanasundaram, K. Photochemistry in Microheterogeneous Sys tems; Academic Press: New York, 1987. (19)Carr, J. W.; Harrls, J. M. And. Chem. 1987, 59, 2546-2550. (20)Kratohvil, J. P.; Hsu, W. P.; Jacobs, M. A.; Amlnabhavi, T. M.; Mukunoki, Y. Co/lohl pdym. Sci. 1983, 267, 781-785. (21) Mazer, N. A.; Carey, M. C.; Kwasnick, R. F.; Benedek, G. 6. B h h e m rsby 1979, 78, 3064-3075. (22) Small, D. M.; Penkett, S. A.; Chapman, D. Blochim. Blophys. Acta 1989, 776, 178-189. (23) D'Alagni, 517-521. M.; Gigllo, E.; Petriconi, S. Colloid Polym. Sci. 1987, 265,
- .. - - ..
(24) Campanelll, A. R.; Candeloro De Sanctls, S.; Chlessi, E.; D'Alagnl, M.; Gigllo, E.; Scaramuzza, L. J . Phys. Chem. 1989, 93, 1536-1542. (25) Schurtenberger, P.; Mazer, N.; Kanzig, W. J . Phys. Chem. 1983. 87, 308-315. (26) Fendler, J. H.; Fendler, E. J. Catalysis in Micellar and Mecromoleculer Systems; Academic Press: New York, 1975. 2146-2149. S.M.; McGown, L. 6. J . Am. Chem. SOC. 1091, 773, (27)Meyerhoffer, (28) Gratzel, M.; Thomas, J. K. In Modern Fluorescence Spectroscopy; Wehry, E. L., Ed.; Plenum Press: New York, 1976; Vol. 2, pp 169-216.
RECEIVED for review May 3, 1991. Accepted July 16, 1991. This work was supported by the United States Department of Energy (Grant No. DE-FG05-88ER13931).
Photopyroelectric Spectrometry and Its Application to the Determination of Trace Constituents in Natural Water Teruo Hinoue,* Junya Kaji, and Yu Yokoyama Department of Chemistry, Faculty of Science, Osaka University, 1-1 Machikaneyama, Toyonaka, Osaka 560, Japan
Michihiro Murata Research & Development Department, Murata Manufacturing Company, Ltd., Hakusan, Midoriku, Yokohama 226, Japan
Photopyrodectrlc spectrometry wlth a compact sensor probe containing a PZT pyroelectric ceramic piece Is described. A He-Ne laser is used as the excitation source. The signal induced by the laser irradiation in a step functkn mode or in a modulated mode could be explained by the classic theory on heat conduction. The concentration detection limit of a dye (Brilliant blue FCF), obtained with the iock-ln amplifier was 5.4 X lo-'' mol dm-', and this concentration corresponded to a Napierian absorption coefficient of 1.6 X l o 4 cm-'. The proposed spectrometry was applied to the determination of phosphorus and Bmmonla nHrogen in natural water on the basts of the heteropdy bhm method and the lndopkd method, respectively. The detection limit was 2.7 ppb for phosphorus and 2.1 ppb for ammonia nitrogen.
INTRODUCTION Up to now, several laser-induced photothermal effects, such as the photoacoustic effect, the thermal lens effect, and so on, 0003-2700/91/0363-2086$02.50/0
have been tested for their ability to detect very weak absorption of a liquid sample and successfully employed for highly sensitive spectrometric detection in batchwise colorimetry, flow injection analysis (FIA), and high-performance liquid chromatography (HPLC) (1,2).In addition to these photothermal spectrometries, photpyroelectric (PPE) spectrometry, in which a temperature change is directly detected by heat rising from the optical absorption, has been developed with thin poly(viny1idene difluoride) (PVDF) foils (3,4)and a lead zirconate titanate (PZT)ceramic disk (5). It has been shown that PPE spectrometry is simple and versatile compared with the other photothermal spectroscopies and can be readily treated theoretically (6-9). However, except for the work by Chirtoc et al. (5), most PPE spectrometry uses have been limited to investigations of solids and thin films (3,4, 10-12). We previously reported (13)the use of the PPE technique for a spectrometric analysis of liquid samples with a sensor probe containing a PZT pyroelectric ceramic piece for the determination of a dye in the liquid samples. The present report describes a fundamental study of signal generation in PPE spectrometry and its application to the de@ 1991 American Chemical Society
