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Anal. Chem. 1991, 63, 1506-1509
Determination of Butyltins in Mussel by Gas Chromatography with Flame Photometric Detection Using Quartz Surface- Induced Luminescence G . B. Jiang,' P.S.Maxwell, K. W.M. Siu,* V. T. Luong, and S. S. Berman I n s t i t u t e for Environmental Chemistry, National Research Council of Canada, Montreal Road, Ottawa, Ontario, Canada K I A OR9
A flame photometric detector optlmlzed for the generatlon of a quartz surface-Induced tln lumlnescence was constructed and evaluated for quantlficatlon of butyltlns In a mussel tissue. The butyltlns were flrst extracted with toluene, pentylated, preseparated from Ilplds, and then separated by uslng gas chromatography. Thls study conflrmed that this quartz surface-Induced lumlnescence for tin, which has an emlsslon maxlmum at ca. 390 nm, Is slgnlflcantly more sensitive than the much more commonty used SnH gasphase lumlnescence at 810 nm. It demonstrated, for the flrst time, that this blue lumlnescence Is sufflclently stable, reproducible, and rugged to be used for environmental sample analysls provlded adequate cleanup Is performed. The amount of cleanup practiced In thls study was no more rlgorous than customarily employed In an analytlcal laboratory followlng establlshed protocols. The mlnlmum detectable amounts (MDAs) (defined as the slgnals that equal 3 tlmes the devlatlons of the nolse) were found to be about 0.3 pg of Sn for tetrapropyltln and about 2-3 pg of Sn for the pentylated derlvatlves of trl-, dl-, and monobutytlln. These MDAs are approximately 30 thnes better than those reported for using SnH emlsslon. For the mussel samples, the relatlve MDAs for the butyltlns were measured to be about 150 pg of Sn/g of tlssue, some 100 tlmes better than those found for using gas-phase lumlnescence under Identical sample workup and slmllar gas chromatographlc condltlons.
INTRODUCTION Although originally developed for sulfur and phosphorus detection ( I ) , flame photometric detection (FPD) has become one of the two major detection techniques for organotin compounds. The high sensitivity of FPD for tin in gas chromatography (GC)was first recognized in 1972 ( 2 ) ,and in a series of studies, Aue and co-workers (3-5) established FPD as one of the best detection techniques for tin. In the last decade, numerous monitoring studies of butyltins in the environment (e.g., refs 6-8) were made by employing gas chromatography with flame photometric detection, thus placing the technique in the forefront of butyltin analysis. Under the appropriate conditions in a hydrogen/air flame, tin compounds are converted to SnH, which yields a red emission in the gas phase (centers a t ca. 610 nm) (9). This red emission forms the basis of tin flame photometric detection in all butyltin analytical work to date. Tributyltin (TBT) is one of the most effective marine antifouling agents and has been extensively used in marine paints until recently. The first large-scale environmental effect of TBT was reported in an oyster-growing area in France in 1980
* Author to whom correspondence should be addressed.
On leave from the Research Centre for Eco-EnvironmentalSciences, Academia Sinica,Beijing, China.
(10). Similar effects were observed in other parts of the world. These observations led eventually to many countries drafting legislations severely limiting the use of TBT. As a result, TBT levels in some highly affected areas have been reported to exhibit a slow but steady decline (11). To be able to monitor decreasing T B T concentrations in the environment, increasingly sensitive analytical techniques are desired. This paper describes our experience in utilizing a highly sensitive but supposedly somewhat unstable and/or irreproducible tin emission mode for real sample analysis. That emission mode was first reported in 1977 by Aue and Flinn (3)and later identified as a quartz surface-induced tin luminescence, which has an emission maximum at ca. 390 nm ( 4 ) . The identity of the emitting species is unknown. Despite its subpicogram limit of detection, this luminescence has not been employed for tin determination, while the less sensitive gas-phase emission is extensively used. One possible reason is that this surface luminescence requires a clean quartz surface in the vicinity of the flame and that only one commercial detector (the Shimadzu) has this feature. A second reason is that it has acquired a reputation for being unstable and easily quenched, which may have discouraged attempts to apply it for quantification. In this paper, we are reporting the construction of a flame photometric detector to exploit this quartz surface-mediated tin emission, the optimization of parameters, the application of this emission mode for the determination of butyltins in a mussel sample, and the comparison of results with those obtained by using the established gas-phase emission mode.
EXPERIMENTAL SECTION Instrumentation. A schematic diagram of the flame photometric detector is shown in Figure 1. The detector body was machined from stainless steel. The burner was assembled from three stainless steel tubes: 1/16-in.o.d., 0.5-mm i.d.; '/*-in. o.d., 2-mm id.; and 1/4-in.o.d., 4.5-mm i.d. arranged coaxially and silver-soldered in place. Air and hydrogen were fed into the outer two tubes; the preferred configuration was hydrogen in the 1/4-in. and air in the l/s-in. 0.d. tubes with the flame being hydrogen rich. The inner most tube (1/16-in.0.d.) carried the column effluent. A quartz tube, 7-mm i.d. X 6.5 cm high, was slipped over the burner assembly to provide the necessary surface for the blue tin emission. Heating of the detector was effected by two cartridges (Watlow) controlled by a variable transformer. To minimize electronic noise, the light pipe and the photomultiplier tube (PMT; RCA, Model IP28) housing were water cooled and purged with nitrogen. A high-voltage power supply (Fluke,Model 412B) was used for PMT biasing (typidy -700 V) while the signal was processed by means of a picoammeter (Keithley, Model 417) and recorded by means of a strip chart recorder (Fisher, Series 5000). Purging of the PMT housing was necessary to keep the humidity and consequently the dark current low. The detector was operated mostly under lopen" (no filter) mode, although some runs were made with a 394-nm band filter or a 610-nm interference filter in place. The detector was housed in the detector compartment of a gas chromatograph (Varian Aerograph, Model 2800). For the separation of butyltin compounds, a methyl/phenylsilicone fused silica
0003-2700/91/0363-1506$02.50/0 Published 1991 by the American Chemical Society
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Flgure 1. Schematic diagram of the flame photometrlc detector.
YmegabOrencolumn (DB-608,J&W Scientific,0.53" i.d. X 15 m, 0.83 pm thickness) was employed. The typical temperature program was initial, 120 OC for 2 min; ramp, 2 OC/min; and final, 150 OC. The temperatures for other heated zones were injector, 200 OC; transfer, 170 O C ; and detector body, 100 OC. This detector body temperature used was much lower than what would have normally been employed but was actually an optimized temperature for maximum analyte signal intensity (see Results and Discussion). Also, as the detector thermal couple was located rather close to a surface that was poorly insulated, the actual temperature of the burner assembly was probably higher than 100 OC. The carrier gas was nitrogen (Linde, Ultra High Purity grade), which had been further purified by sequential passage through a molecular sieve 5A trap and a heated oxygen scavenger unit (both from Supelco). The typical flow rate was 8 mL/min. For the detector gases, typical flow rates were hydrogen, 300 mL/min, and air (both from Linde, High Purity grade), 160 mL/min. Reagents. Organotin compounds were purchased from commercial sources (Alfa and Aldrich). Their purity had previously been established by GC (12) and liquid chromatography-inductively coupled plasma mass spectrometry (13). Stock solutions of tributyltin chloride, dibutyltin (DBT) dichloride, monobutyltin (MBT) trichloride, and tzipropyltin (TPT) chloridewere prepared in hexane and refrigerated when not in use. Tetrapropyltin (TePT) solutions in toluene were used for the optimizationof FPD conditions. All solvents were 'Distilled-in-glass" grade (Caledon). Butyltin chloride solutions were pentylated with pentylmagnesium bromide (2 M solution in anhydrous ether, Aldrich) prior to gas chromatography. Mussel Sample. A batch of mussels were made available to us by courtesy of S. Freitas (Battelle Ocean Sciences, Duxeury, WA). It was harvested in Boston Harbor and delivered frozen. The mussels were shelled, pooled, homogenized, freeze dried, and blended. The dry powder was stored in the dark in a cold room (ca. 5 "C) until analysis. Sample Extraction, Pentylation, and Cleanup. A 0.50-g portion of the dry muasel sample was placed in a WmL centrifuge tube. The appropriate amount of TPT was added as an internal standard; for standard additions runs, the appropriate amounts of TBT, DBT, and MBT were also added. The mixture was allowed to equilibrate ovemight. A 30-mL portion of toluene was then added followed by 1mL of 1.5% tropolone (Aldrich)solution in toluene. The mixture was sonicated in an ultrasonic bath (Branson, Model 2200) for 30 min and then centrifuged at 2000 rpm for 20 min (IEC HN-SI1 centrifuge). The toluene phase was removed by means of a Pasteur pipet. The extraction was repeated with another 30-mL portion of toluene plus 1mL of 1.5% tropolone solution. The toluene phases were combined, fitered over 2 g of anhydrous sodium sulfate, and reduced to almost dryness in a rotary evaporator. Two milliliters of hexane was
FIgun 2. Gas chromatographic separation of organotin compounds: peak 1, solvent; peak 2,20 pg of Sn as TepT; peak 3, 100 pg of Sn as TPT; peak 4, 100 pg of Sn as TBT; peak 5, 100 pg of Sn ea DBT; peak 6, 100 pb of Sn as MBT. Peaks 3-6 were the pentylated derivatlves of the organotin chlorides. See text for experimental con-
ditions.
added to dissolve the brown syrup, and the solution was transferred into a 10-mL Reacti-vial (Pierce) followed by 2 mL of hexane wash. Two milliliters of 2 M pentylmagnesium bromide solution in ether was added, and the pentylation reaction was allowed to proceed at 80 O C for 3 h. The mixture was then cooled and chilled on ice. One milliliter of deionized, distiUed water was added dropwise followed by 1mL of 6 M hydrochloric acid. The mixture was centrifuged at 2000 rpm for 15 min, after which the organic phase was removed for column cleanup. The column (a modified 50-mL buret) was dry packed, in successive order, with 10 g of Florisil, 60-100 mesh, which had been activated at 170 OC for 2 h; 10 g of alumina, 40-60 mesh, previously activated at 400 OC for 2 h; and 10 g of anhydrous sodium sulfate. The packing was k t conditioned by the passage of 100 mL of hexane. The derivatized mussel extract was then placed on the column head, and the penytylated butyltins were eluted with 140 mL of hexane. The colorless eluate was reduced to approximately 2 mL in a rotary evaporator and further evaporated to 0.3 mL by means of a stream of nitrogen. This solution was then ready for GC analysis,which typically required 1pL of sample.
RESULTS AND DISCUSSION Numerous extraction and cleanup procedures for butyltin determination have been published. The one used in this study was a modified version of the procedure adopted by Wade et al. (14),the only significant change being the use of toluene instead or dichloromethane as the extraction solvent to ensure quantitative butyltin recoveries. The sample treatment was arguably complicated and labor-intensive, but this was deemed necessary for the removal of interferences that would have degraded analytical performance. This scale of complexity was almost universally employed in all sample work-up procedures for biological tissues. The gas chromatographic performance of the DB-608 column for butyltins has previously been reported (12). The separation of TePT, TFT, TBT, DBT, and MBT was straightforward and could be accomplished easily in a linear temperature program (Figure 2). Similar to the emission of other species in FPD, the blue luminescence of tin is heavily dependent on the flame environment. During the initial stages of this study, much time was spent on the optimization of several interrelated key parameters, e.g., detector gas flow rates, burner assembly height, gas flow configuration, and quartz tube diameter; the
ANALYTICAL CHEMISTRY, VOL. 63,NO. 15, AUQUST 1, 1991
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FIguro 4. Response of 50 pg of tetrapropyttln under various FPO conditkns: (a)opem (no fllter) mode, (b) with 394-nm filter, (c) with 6 1 h m filter, (d) open rode, quartz encloswe replaced with Pyrex enclosure. See text for experimental c c " s .
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Flgur. 9. (a, top) Detector r o q " versus hydrogen flow rate: air now, 160 M m h . @. battom) Detector response v(~glg a t flow raw hydrogen flow, 300 mL/min. (0)TePT, 20 pg of Sn Injected; pentyl dotivathe8 of (X) TPT, (+) TBT, (A)DBT, and (B) MBT. 100 pg of Sn injected.
conditions or numbers reported here were optimal for the present detector. For the gases, the optimum condition was achieved when hydrogen was fed to the l/&.-o.d. tube and air was fed to the 1/8-in.-o.d. tube a t rates of 300 and 160 mL/min, respectively, and when the quartz tube dimensions and the viewing height were those shown in Figure 1. Figure 3 illustrates the degree of response variation versus hydrogen as well as air flow rate. That the emission observed in this study was the blue quartz surface-induced luminescence and not the more common red gas-phase emission was evident from the following observations the tin signal vanished when the quartz tube was replaced with a Pyrex tube of identical dimensions; the tin signal/noise decreased by a factor of about 2 (although both the absolute signal and the absolute noise decreased by a much larger factor) when a 394-nm band-pass filter was installed but dropped to zero when a 610-nminterference filter was in place (Figure 4). These observations established that the l u m i n m required the preaence of quartz and centered in the blue region rather than the red region. The simple cylindrical quartz tube performed best when it came to sensitivity, peak symmetry, and reproducibility. The performance of other designs, such as bisected, constricted, and quartz wool (3-51,was either comparable or inferior. For the cylindrical flame enclosure, the critical dimension was the tube inner diameter, which presumably defined the degree and the area of contact between the quartz and the flame. The optimum diameter found for the present
20
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Flguro 5. FPD response of 10 pg of tetrapropyltin versus detector temperature.
detector when it was operated under typical conditions was about 7-mm i.d. As well, freons were found ineffective as cleaners to counteract detector poisoning as previously reported (3, 4). Methanol worked to a certain extent, but the best solution we found was to soak the quartz tube in aqua regia overnight. This, however, was needed only infrequently; our last cleaning was done some 3 months ago, and we have been using the system for mussel extract analysis. Detedor temperature appeared to have a strong effect on the intensity of the surface luminescence. Figure 5 shows a plot of FPD response versus detector temperature for tetrapropyltin, which exhibits an optimum range from 80 to 120 OC. This range was lower than what would normally be considered as a prudent detector temperature in view of the final column program temperature of 150 O C . As it turned out, no evidence of butyltin condensation was apparent even after months of operating our flame photometric detector at 100 "C. It should be emphasized that the detector temperature was measured at a point adjacent to a rather poorly insulated surface and that the burner assembly temperature
ANALYTICAL CHEMISTRY, VOL. 63, NO. 15, A W S T 1, 1991 sample
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Table I. Analysis of Mussel Sampleso
ua of Snla of tissue surface luminescence gas-phase emission
1
TBT DBT MBT
0.0954 f 0.0049* 0.0481 h 0.0032 0.0439 f 0.0076
0.0946 f 0.0029 0.0501 f 0.0067 0.0493 f 0.0083
Six redicate analvses. Standard deviation.
Flguro 6. Mussel analysis (a) quartz surfacelnduced luminescence mode and (b) gas-phase emission mode (Varian detector). Peaks 1, 2, 3, and 4 are pentyl derivatives of TPT, TBT, DBT, and MBT, respectively. The standard solutions contained the equivalent amounts of TPT, TBT, DBT, and MBT added to the spiked samples. The solutions analyzed by the gas-phase emissbn mode were 8 times more concentrated then those for the surfaceinduced luminescence mode.
was probably higher than that. The figures of merit of this surface-inducedluminescence for tin were noteworthy. The overall reproducibility of 10 sequential injections of the same standard solution was always better than 8% (standard deviation). The detector linear range spanned over about 4 decadea; for tetrapropyltin, it was from 0.3 pg to about 1 ng of Sn. The minimum detectable amounts (MDAs), defined as the signals that equal 3 times the deviations of the noise, were about 0.3 pg for tetxapropyltin and 2-3 pg for the pentylated derivatives of tri-, di-, and monobutyltin. These MDAs are approximately 30 times better than those reported for using SnH emission in a commercial detector (12). This more sensitive surfaceinduced luminescence mode was applied to the determination of butyltim in a mussel sample, which we deemed to be one of the best ways to assess the sensitivity,ruggedness, and reproducibility of this method for real sample analysis. As well, to gauge its accuracy and performance versus its closest rival, the gas-phase SnH emission, parallel analyses were performed on a commercial flame photometric detector (Varian) (12). To make up for the lack of sensitivity in the gas-phase emission, 8 times more (4g) mussel tissue was extracted under otherwise identical conditions to bring the butyltin concentrations in the cleaned up and concentrated extract to within usable range of the commercial detector (Figure 6). Occasionally, some of these extracts were diluted 8 times for analysis by means of surface-induced luminescence for a direct comparison. Table I summarizes the results. Good agreement is evident between
the two sets of results for TBT, DBT, and MBT. Measurementa were performed by means of both standard calibrations and standard additions; their results were comparable. The surface-mediated luminescence mode performed impeccably under heavy use. The emission was stable and reproducible, and no signs of poisoning or quenching were apparent even after months of use in mussel analysis. The relative MDAs for the butyltins were estimated to be about 150 pg of Sn/g of tissue; this was about 100 times better than those found for the gas-phase emission mode using the commercial detector. In conclusion, the quartz surfaceinduced luminescence for tin was found to be significantly more sensitive than, and equally as rugged and reproducible as, the gas-phase emission mode. These aspects were fully demonstrated in the application of this surface emission mode to the analysis of cleaned up and concentrated mussel extract.
ACKNOWLEDGMENT We thank S. Freitas for making the mussel sample available to us, and W. A. Aue for discussion. Registry No. DBT, 683-18-1; MBT, 1118-46-3; tributyltin chloride, 1461-22-9.
LITERATURE CITED (1) (2) (3) (4) (5)
(8) (7) (8) (9)
IO) 11) 12) (13) (14)
Brody. S. S.; Chaney, J. E. J . Gas. chrometugr. 1988, 4 , 42-46. Aue, W. A.; Hill, H. H.; Jr. J . chrometogr. 1972, 70, 158-161. Am, W. A.; Fllnn, C. 0. J . chnrmetog.1977, 142, 145-154. Fllnn, C. 0.;Aue. W. A. Can. J . Spectrarc. 1980, 25, 141-148. Aue, W. A.; Fllnn, C. a. Anal. &em. 1980, 52, 1537-1538. Maguke, R. J.; Huneautt, H. J . chrometogr. 1981, 209, 456-462. Magulre, R. J. En&. Scl. T M . 1984, 18, 291-294. Mueller, M. D. Fresenlus' Z . Anal. Chem. 1984, 317, 32-36. Degnall, R. M.; Thompson, K. C.; West, T. S. AM&t 1988, 09, 518-521. AWeu, C.; Thibauk, Y.; Herel, M.; Boutler, B. Rev. Tmv. Inst. F " s M .1980, 44,301-349. Vatklrs, A. 0.; Davidson, B.; Fransham, R. L.; Grovhwg, J. G.; Sei& man, P. F. Prtxsdhp of Ilw, 3rd Infemelknel w n o t h Synpodm, Monaco, April 17-20, 1990 pp 151-158. Slu, K. W. M.; Maxwell, P. S.; Berman, S. S. J . chrometog. 1989, 475, 373-379. W r e n , J. W.; Slu, K. W. M.; Lam, J. W.; W W , S. N.; MaxweU, P. S.; Palepu, A.; Koether, M.; Berman, S. S. Freseni" Z . Anal. Chem. 1990, 337, 721-728. Wade, T. L.; Garcle-Romero, B.; Brooks, J. M. En&. Sd. T M . 1988, 22, 1488-1493.
RECEIVED for review January 22, 1991. Accepted April 24, 1991.
