Anal. Chem. 1994,66,874-881
Cloud Point Preconcentration and Hlgh-Performance Liquid Chromatographic Determination of Polycyclic Aromatic Hydrocarbons with Fluorescence Detection Carmelo Garcia Pinto, J o d LUIS P6rez Pavh, and Bernard0 Moreno Cordero'
Departamento de Qujmica Anahica, Nutricfin y Bromatolog?a, Facultad de Qujmica, Universldad de Salamanca, 37008 Salamanca, Spain
In chromatographic analysis of polycyclic aromatic hydrocarbons (PAH), three important aspects must be taken into account: losses by sorption onto the containers, association with dissolved organic material, and the need for high preconcentration factors. In this work, fluorescence detection was used for liquid chromatographic analysis of polycyclic aromatic hydrocarbons after cloud point preconcentrationwith the nonionic surfactant Triton X-114. Negligible sorption of PAH onto the containers and high recoveries, even in the presence of considerable levels of humic acids, are additional advantages of the proposed micelle-mediated methodology. Bottled and river water samples afforded almost complete recoveries when the procedure described was applied. Furthermore, micellar solutions of Triton X-114 effectively extracted PAH from solid samples. Polycyclic aromatic hydrocarbons (PAH) currently form one of the groups of organic compounds of greatest environmental impact owing to their widespread distribution in the milieu; this is because they are generated both by natural causes (forest fires, volcanic activity, etc.) and by incomplete combustion of fossil fuels and other organic materials.'v2 The mutagenic and/or carcinogenic nature of these compounds3 means that their presence in different matrices should be rigorously monitored; among such matrices, of exceptional importance is drinking water, both from the supply network and in bottles. In view of the maximum levels permitted, around a few tenths ppb, their determination is difficult and therefore a preconcentration step is essential. Previously, the preconcentration step has involved the use of either solid sorbent^^.^ or liquid-liquid e x t r a c t i ~ n . ~ ? ~ It is known that aqueous solutions of certain surfactants, both nonionic and zwiterionic when heated above a certain temperature (cloud point temperature) exhibit the property of separating into two phases.*-10 Once formed, and after a (1) Bjorscth, A. Anal. Chim. Acra 1977, 94, 21-27. (2) Auer, W.; Malissa, H., Jr. Anal. Chim. Acta 1990, 237, 451457. (3) Harvey, R. G., Ed. Polycyclic Hydrocarbons and Carcinogenesis; American Chemical Society: New York, 1985. (4) Symons, R. K.; Crick, I. Anal. Chim. Acta 1983, 151, 237-243. (5) Van Rmsum, P.; Webb, R. G. J. Chromatogr. 1978, 150, 381-392. (6) Jungclans, G.A.; Games, L. M.; Hites, R. A. Anal. G e m . 1976,48, 18941896. (7) Achcson, M. A.; Harrison, R. M.; Perry, R.; Wellings, R. A. WaterRes. 1976, 10, 207-212. (8) Corti, M.; Minero, C.; Degiorgio,V. J. Phys. Chem. 1984, 88, 309-317. (9) Degiorgio,W.; Piazza, R.; Corti, M.;Minero, C. J. Chem. Phys. 1985, 82.
1025-1031. (10) Goldstein, R. E. J. Phys. Chem. 1986, 84, 3367-3378.
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Analytical Chemlsw, Vol. 66,No. 6,M r c h 15, 1994
given time, which can be accelerated by centrifugation, two transparent liquid phases are obtained; one contains most of the surfactant (the surfactant-rich phase) and the other is an aqueous phase in equilibrium with the former, with a surfactant concentration close to the critical micelle concentration (cmc). The surfactant-rich phase can be used for the preconcentration of certain analytes before injection in HPLC. The analytical potential of the cloud point phenomenon has been discussed by several authors11-15 and can be summarized as follows: the small volume of the surfactantrich phase obtained, as a function of the amount of surfactant in solution, can be used for the preconcentration of analytes of both hydrophilic (Le., metal ions after reaction with a suitable ligand) and hydrophobic nature as a step prior to their determination by HPLC16J7 or FIA,18 obtaining preconcentration factors similar to those of other conventional extraction techniques. Recently, the cloud point separation of some PAH with Genapol X80 and synchronous fluorescence detection have been described.19 The experimental procedure is very versatile since there are many commercially available surfactants with cloud point temperatures suitable for analytical purposes (e.g., separation of heat-sensitive molecules), and others of more specific functional nature can be synthesized that will allow improvements in selectivity. The methodology is simple since one only needs to heat the solutions above the cloud point temperature; it is inexpensive with respect to the reagent used and simple apparatus required and is less toxic than extraction with organic solvents. The surfactant-rich phase obtained in the separation process is compatible with micellar mobile phases, hydroorganic mobile phases, and the carriers usually employed in FIA methodology; this facilitates latter determination of the analyte by any hydrodynamic analytical method. One of the most important drawbacks when this methodology is used prior to HPLC is (11) Bordier, C. J. Biol. Chem. 1981, 25, 1604-1607. (12) Watanabc, H. In Solution Behmiour of Surfacrants; Mittal, K. L., Fendler, E. F., Eds.; Plenum Press: New York, 1982; Vol. 2, pp 1305-1313. (13) Hinze,W. L. Ann. Chim. 1987, 7 7 , 167-207.
L. In Ordered Media in Chemical Separations; Hinze, W. L., Amstrong, D. W., Eds.; ACS Symposium Serics 342; American Chemical Society: Washington, DC,1987. (15) Pramauro, E. Ann. Chim. 1990.80, 101-109. (16) Saitoh, T.;Hinzc, W. L.Anal. Chem. 1991, 63, 2520-2525. (17) Garcla Pinto, C.; Perez Pav6n, J. L.; Moreno Cordero, B. Anal. Chem. 1992, (14) Hinze,W.
64, 2334-2338. (1 8) Fernindez Lacspada, M. E.; Perez Pav611, J. L.; Moreno Cordero, E.Analyst 1993,118, 209-212. (19) Bockelen, A.; Nicssner, R. Fresenius J. Anal. Chem. 1993, 346, 435440.
0003-2700/94/036&0874$04,50/0
0 1994 Amerlcan Chemlcal Soclety
the high background absorbance in the UV region. Electrochemical detection has recently been proposedI7as a way to overcome this problem. In the present work, we propose for the first time the use of the nonionic surfactant Triton X- 114 as a real and efficient alternative for the preconcentration of PAH using the cloud point methodology as a step prior to their determination by HPLC with fluorescencedetection. The effect ofTriton X-114 on two of the causes that most often lead to low recoveries of these compounds are studied: i.e., adsorption of the PAH on the container surface used for collecting aqueous samples and their interaction with the organic matter present in the water (Le., humic acids). Also studied is the possible use of this surfactant for the extraction of PAH from solid samples as a step prior to their preconcentration. Triton X-114 was selected owing to its favorablecharacteristics regarding price, toxicity, and commercial availability.
EXPERIMENTAL SECTION Reagents. The nonionic surfactant Triton X-114 was obtained from Fluka and used without further purification. HPLC-grade acetonitrile was obtained from Carlo Erba. Individual PAH standards were purchased as 100 pg/mL solutions in methanol from Sugelabor Chemical Service. All other reagents were of analytical grade. All solvents and analytes were filtered through 0.45-pm nylon membrane filters (Millipore), and ultra-high-quality water obtained from a Elgastat UHQ water purificationsystem was used throughout. Apparatus. A modular component liquid chromatographic system was used consisting of a Spectra Physics SP 8800 ternary pump, a Spectra FL 2000 fluorescence detector, and an S P 4290 integrator. In all experiments, a Rheodyne 7125 injection valve with a IO-pL sample loop was used. The stationary-phasecolumns were a 224 X 4.6 mm Spheri 5 ODS224 column from Brownlee and a 15 X 4.6 mm Vydac 201TP5415 5-pm particles from Phenomenex. Easyspin Sorval Instruments Du Pont and Kokusan H-103 N centrifuges were also used. Cloud Point Preconcentration. Appropriate aliquots (1050 mL) of the cold solutions containing the analytes in the presence of Triton X- 114 were kept for 5 min in a thermostated bath at 40 OC. Separation of the two phases was achieved by centrifugation for 5 min at 3500 rpm. The determination of cloud point temperature and the phase ratio has been described elsewhere.17.18 Liquid Chromatographic Analysis. Most studies were carried out with the Spheri 5 ODS-224 column. From the surfactant-rich phase obtained after cloud point preconcentration, 60 pL was collected using a Hamilton syringe of which 10 pL was injected into the chromatographic system. The separation and detection of the PAH was carried out using a mobile phase consisting of 75:25 (v/v) acetonitribwater accompanied by excitation-mission wavelength programming. Detection parameters were optimized for minimum background signal and maximum sensitivity for 10 PAH. The excitation-emissionwavelengths (in nanometers) used for each compound wereas follows: 334-348 for anthracene (l), pyrene (3), and benz[a]anthracene (4); 348-450 for fluoranthene (2); 300-430 for benzo[b]fluoranthene ( 5 ) , benzo[k]fluo-
ranthene (a),benzo [a]pyrene (7), dibenzo[a,h]anthracene (8), and benzo[ghi]perylene (9); 300-500 for indeno[ 1,2,3-c4pyrene (10). Numbers in parentheses indicate the elution order and will be used to identify peaks in the figures. Benzo[ghi]perylene and indeno[ 1,2,3-cdpyrene coelute when the Spheri 5 ODS-224 column is used. Therefore, two injections with different detector settings were made for quantification of these compounds in synthetic mixtures containing both of them. Chromatographic analysis of water samples was performed with the Vydac 201TP5415 column, with which the two above compounds elute separately. Due to a higher background signal caused by the surfactant with this column, 100pL of acetonitrilewas added to the surfactantrich phase before its injection into the system. Sorption of PAH onto Containers. The sorption of PAH onto the containers was measured by taking aliquots of the preparedsolutionat selectedtimeintervals; 0.596TritonX-114 was added and cloud point preconcentration was carried out, quantifying the recovery after chromatographic analysis. A similar procedurewas used to study the effect of the surfactant on sorption; in this case, 0.5% Triton X-114 was added to the solution at the beginning, and aliquots were preconcentrated and analyzed chromatographically. Recovery of PAH from Humic Acid Solutions. An optically standardized humic acid solution was prepared according to the procedure described by Johnson at a1.,2O spiked with appropriate amounts of PAH, and left overnight. Following this, 0.5% Triton X-114 was added and aliquots taken at 15 and 45 min were cloud point preconcentrated and analyzed chromatographically. Recovery of PAH from Spiked Water Samples. Triton X-ll4,0.1%, was added to 50-mL water samples immediately after collection. Aliquots, 15 mL, were cloud point preconcentrated, and after phase separation, 100 pL of acetonitrile was added to the surfactant-rich phase, 10 pL of this being injected into the chromatographic system. River water samples were passed through glass wool filters prior to preconcentration. Micelle-MediatedExtraction and Cloud Point Preconcentration of PAH from Solid Samples. Solid samples, wood ashes, and smoke particulates were collected from a house fireplace. Appropriate amounts of the solids (6-40 mg of smoke particulates or 0.1-2 g of wood ash) were extracted with an aqueous 0.5% Triton X-114 solution. The extracts were filtered and injected into the chromatographic system either directly (smoke particulate samples) or after cloud point preconcentration (wood ash samples). Cloud point preconcentration of wood ash extracts was always performed at 40 OC, independent of the temperature at which the solid was extracted with the surfactant solution.
RESULTS AND DISCUSSION Adsorptiononto Containers. The Occurrence of losses due to the sorption of PAH onto containers is a well-known phenomenon that poses problems, above all when extraction with solid sorbents is used in the preconcentration method. To avoid this, it is recommended that solvents such as methanol be used at quite high concentrations ( ~ 2 0 % )The ~ use of (20) Johnson,W. E.; Fcndingcr. N. J.: Plimmcr,J. R. AMI. Chcm. 1991.63.1 5 10-
1513.
Analytical Chemistry, Vol. 66, No. 6, M r c h 15, 1004
875
41
Tabk 1. Sorptlon of PAH onto a a r r Contalwr
compound'
l h
% recovery 8h 12h
fluoranthene benzo[ai pyrene benzo[ghi]perilene
100 92 95
72 21 17
0
12hb
56 16 11
99 100 100
0
3 -
Concentration, 2 ppb. Sample stored in the presence of 0.5% Triton X-114.
2 -
TaMo 2. Sorptlon of PAH onto Polyothylono Contalnon % recovery
1 -
compound(ppb)
2h
6h
2 ha
2hb
fluoranthene (4) benzo[a]pyrene(2) benzo[ghi]perilene(2)
ND 3 12
ND ND
91 85 99
43 40 53
ND
78 65 87
90 77 95
'Sample stored in the presence of 0.6% Triton X-114 from the beginning. b After 6 h in the container, 0.5 % Triton X-114 waa added (for details, see text). Tablo 3. Rlcovorlor of PAH from Humlc Acld 8dutlon
% recovery 15 min' 45 min'
fluoranthene (4.8) benzo[a]pyrene (2.4) benzo[ghi]perylene (2.4)
\
Fluorsntbe
' 0
10hb 20hb
n " l 0,O
compound (ppb)
\ \
F
94 94 98
99 97 98
I
I
I
I
0,1
0,2
0,3
0,4
I
0,5
0,6
Trlton X-114/% Flguro 1. ReiaUve fluorescence intensity versw) Trlton X-114 conCentratkn.Samples: 0.70ppbfkroram, 0.12ppbbw0[8]pyr0n0, and 0.10 ppb benro[ghllpc#ylene. ChrorrmtograpMc condltkns as described In the text. 3
a
Time elapsed between the addition of Triton X-114 (0.5%)and the cloud point preconcentration. Tablo 4. Sorptlon of PAH onto Qlarr Contalnon: Effsot of Humlc Aclda % recovery after 48 h
compound'
water
humic acid solution
fluoranthene benzo[al pyrene benzokhilperylene
18 5 4
55 71 59
3
Concentration, 2 ppb.
b 0.30 FU
surfactant species has also been described as an alternative to hydroorganic media.21*22The use of Triton X-I14 as a surfactant for the preconcentrationof PAH has the advantage that this medium effectively avoids the sorption of the species of interest onto thecontainers. Table 1 summarizesthe results obtained for three PAH of different hydrophobicities. It may be seen that the presence of Triton X-114,used in this work for the preconcentrationof the PAH, also inhibitstheir sorption onto the glass containers with no modifications being observed in the signal for at least 3 days. Adsorption onto polyethylene containers is even more pronounced, as can be seen in Table 2; after 6 h of contact, none of the three species tested could be detected. However, in the presence of Triton X-1 14,recoverieswere almost 100%. The surfactant is also able to desorb the PAH retained, although the process is slow. When a solution of PAH was (21) Hinzc, W. L.; Singh, H. N.; Fu, Z. W.; William, R. W.; Kippcnberger, D. J.; Morris, M. D.; Sad& F. S.In Chemical Analysis of Polycyclic Aromatic Compounds; Vo-Dinh, T., Ed.; Wilcy: New York, 1989; Chapter 5, pp 151-
169. (22) Mpcz Garcla, A.; Blanco Gondlez, E.; Garcia Alonso, J. I.; Sanz Mcdel, A. Anal. Chim. Acta 1992, 264, 241-248.
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Analytical Chemistry, Vol. 66,No. 6,March 15, 1994
II
I
I
~~
0
4
8 12 Timdmin
16
Flgurr 2. Chromatograms obtalnrd for the injection of a Surfactantrich phase of 0.12% (a) and 0.27% (b) Trlton X-114 solution8 after cloud point preconcentratlon. Chromatographk condltbn8 and peak assignment as cksorlbed in the Experimental Seotkn.
left for 6 h in the container and Triton X- 1 14 was added, -20 h was required for high recoveries to be attained (see Table 2). This ability of the surfactant to prevent sorption onto containers facilitates the procedure, and no added chemicals other than Triton X-114are required to prevent such losses. This same surfactant is used in the preconcentrationstep. The recoveries obtained are similar to those reported for liquid-
0.01 A U
0 0
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30
45
60
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15
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Sample v o l u m e h l Flgurr 3. Relative fluorescence intensity versus sample volume preconcentrated. For details, see text.
liquid extraction or for solid-liquid extraction in the presence of about 20% methanol! Recovery of PAH from Humic Acid Solutions. The interference caused by humic acids in liquid-solid extraction of organicshas been describedpreviously. The results obtained by Johnson et alazoindicate that species such as pesticides associated with humic acid material, either as a complex or by simple adsorption, are not retained by the solid sorbent, leading to lower recoveries than those obtained by liquidliquid extraction. In order to investigate the effect of humic acids on the cloud point preconcentration of PAH, a solution of humic acids (- 10 ppm organic carbon content) was spiked with 4.8 ppb fluoranthene, 2.4ppb benzo[a]pyrene, and 2.4ppb benzo[ghilperylene. As the surfactant was added to the whole solution, no attention was paid to the possible sorption onto the container, which, as shown above, is negligible in the presence of Triton X-114. The recoveries obtained in the cloud point preconcentration are shown in Table 3; 45 min after the addition of Triton X-114,the removal of PAH into the surfactant-rich phase was almost complete despite the possible association with the organic material. Some evidence of association is provided by the fact that adsorption of PAH on the walls of a glass container was lower when humic acids were present in the solution (Table 4). Cloud Point Preconcentrationand Liquid Chromatographic Analysis. The surfactant-rich phase volume obtained after cloud point preconcentration depends on the Triton X-114 concentrationand determines the maximum preconcentration factor attainable. This allows one to design analytical schemes with a preselected preconcentration factor as a function of the amount of analyte to be determined, of the volume of sample available, and of the analytical technique to be ~sed.~'J*
I
0
I+
1
+k+
1
4
8
12
16
+, 20
I
24
Timdmin
Figu. 4. Blankchromatog" obtahedfor the lnjecfbnof a sufactantrich phase of Triton X-114 alone after cloud point preconcentratbn wtth UV detection at 254 nm. Arrows Indicate the retention times of six PAH. Chrometographicc o n d k b and ~ peakas8lanmentas deacrbd in the Experlmentai Section.
Figure 1 shows the relationship between the relative fluorescence intensity and the Triton X-114 concentration when 15-mLsamples are preconcentrated and the surfactantrich phases obtained are analyzed chromatographically. Three model PAH were used for this experience: 0.7 ppb fluoranthene, 0.12 ppb benzo[a]pyrene, and 0.1 ppb benzo[ghi]perylene. The chromatogramsin Figure 2 correspond to 0.12% and 0.27% Triton X-114,respectively. For a fixed amount of surfactant, the preconcentration factor, and hence the intensity of the analytical signal, depend on the sample volume preconcentrated. Figure 3 shows the relationship between the relative fluorescence intensity and the preconcentrated sample volume of solutions containing 0.1 ppb of three PAH. The data in Table 5 show the enhancement factor (the ratio of peak intensities with and without preconcentration) for six PAH. A very high concentration of Triton X-114(3%) was used in this experience in order to accurately measure the
T a b 5. Enhammonl Factor
compound
enhancements factor
compound
enhancement? factor
fluoranthene benzo[b] fluoranthene bozo [kl fluoranthene
14.1 30.3 20.3
benu, [a]pyrene benzoCghilperylene indeno[ 1,2,3-cd]pyrene
30.2 20.7 21.1
a Ratio
of fluorescence peak intensities of preconcentrated sample to that obtained without any preconcentration step. AnaWcal Chemlstry, Vd. 66, No. 6, M r c h 15, 1994
811
Tabh 6. Analytlcri Charaotwbtb d th.Mothod'
compound
range (ppb)
anthracene fluoranthene Pyrene benz [a]anthracene benzo[b]fluoranthene benzo[k] fluoranthene benzo[a]pyrene dibenzo[a,hlanthracene benzo[ghi]perylene indeno[ 1,2,3-cd]pyrene
0.15-2.0 0.10-2.0 0.03-0.3 0.15-1.5 0.08-1.0 0.01-0.1 0.04-0.3 0.08-1.0 0.03-0.3 0.3-3.0
slope (4.2 (4.6 (7.6 (5.0 (1.79 (1.11 (3.8 (1.36 (2.00 (2.21
intercept
1V ** 0.1) 0.2) X 1V X l@ * 0.3) 0.2) l V 0.06) 1@ * 0.03) 109 * 0.1) x l@ 0.05) X 1@ ** 0.05) X l@ 0.06) x 104 X
X
X X
* 1) x 109 ** 3)1) xx 109 ** 1)2) xx 109 109 ** 1)1) xx 109 109 109 **1)6)x 109102 * 7) x 102
(2 (1 (4 (3 (1 (1 (1 (1 (-5 (5
X
P
% rsdb
L O P (Ppb)
0.9988 0.9980 0.9979 0.9984 0.9986 0.9989 0.9990 0.9983 0.9989 0.9989
4.3 (0.4) 3.3 (0.8) 3.5 (0.08) 4.5 (0.6) 3.3 (0.25) 3.7 (0.04) 4.8 (0.12) 4.4 (0.25) 5.2 (0.1) 4.1 (1.0)
0.07 0.06
0.004 0.006 0.02 0.002 0.007 0.02 0.01 0.12
0 Samples, 15 mL, with 0.5% Triton X-114. Du licate injection. * Values in parentheses are the compound concentrations for which red was obtained. e LOD, hmit of detection (calculate8aa twice the noise).
6
volume of the surfactant-rich phase obtained. Since the ratio of phases under the described experimental conditions was 15 and the enhancement factor found for most of the compounds studied was greater than this value, the additional increase in sensitivity can be attributed to the modifications in the microenvironment of the analytes when they reach the detector in the presence of the surfactant. Figure 4 shows the chromatogram obtained with UV detection at 254 nm when a solution of Triton X-114is preconcentrated and injected into the chromatographic system. The arrows indicate the retention times of the compounds in Table 5. Calibration graphs were constructed for 15-mLsamples with 0.5% Triton X-114. This concentration of surfactant ensures a sufficient surfactant-rich phase volume to make two injections per sample, although lower Triton X-114 percentages would lead to greater sensitivity. The ratio of phases under these experimental conditions was 70. Linear relationships between the relative fluorescence intensity and concentration were found for all the compounds studied. The parameters of the least squares fittings are shown in Table 6 together with the calculated detection limits (twice the noise) and the relative standard deviation for 10 samples to which the complete procedure (cloud point preconcentration and chromatographicseparation) was applied. When 0.2% Triton X-114is used, the slopes of the calibration graphs and the detection limits for the different compounds are improved by a mean factor of 2.2,but only one injection per sample can be made. Under these experimental conditions the ratio of phases was 140. Preconcentration of PAH in Water Samples. In order to test the reliability of the proposed methodology for the separation and preconcentration of PAH, it was applied to samples of bottled water and of water from the River Tormes (Salamanca, Spain). No peaks correspondingto the species studied were detected in the blank chromatogram of the bottled water (Figure 5a). The recoveries obtained for samples spiked at four different levels are in Table 7, and the chromatogram in Figure 5b corresponds to spike 2 in Table 7. The blank chromatogram obtained after preconcentration of 15 mL of River Tormes water shows one peak suspected to correspond to pyrene. The chromatogram in Figure 6a corresponds to an unspiked river water sample. In order to better identify the compound detected, chromatograms were recorded at two different sets of excitation-emission wavelengths (334-384 and 340-400 nm); the ratios of peak 878 Am!~tlcaIChem&try,Vol. 66, No. 6, March 15, 1994
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8 12 16 Time /min. Flguro 8. Chromatograms obtalned for the Injection of a surfactantrich phase of a bottled water sample (a) and of a @ked bottled water sample (b) after cloud point preconcentratkn. Chromatographk conditions and peak asslgnment as described in the Experlmentai Section. 0
4
intensities in the unspiked and spiked sample were 2.36 and 2.34,respectively (less than 1% difference). A concentration of 0.14 ppb was found for pyrene by direct interpolation in the calibration graph and by standard addition. Even though the absolute identification of individual PAH is difficult due to the diversity of isomers, the results obtained indicate, with a high level of confidence, that the detected compound was indeed pyrene. Extraction and Preconcentration of PAH from Solid Samples. The use of aqueous solutions of surfactants as alternative extractants to organic solvents was reported previously,I6although their use in analytical methodology is not widespread. In this work, Triton X-1 14 is proposed as an extractant for PAHs in different solid samples. Figure 7 shows the chromatogram obtained when a sample of 0.04 g of smoke particulates was extracted with 100 mL of a 0.5% aqueous solution of Triton X-114over 5 h and 10 p L of the solution filtered was injected into the chromatographic system. The retention times of seven of the peaks appearing in the figure correspond to the retention times of seven PAH of those studied. As for pyrene in the river water sample, these seven peaks were identified by their retention times and the peak intensity ratios using different pairs of excitation and emission wavelengths for the sample and for solutions of standards. These ratios differ by less than 5%.
~~
Table 7. R e c o v of ~ PAH trom Splked Water Sampkr
% recovery
spike
compound
added (ppb)
bottled water
river water
1
fluoranthene benzo[b]fluoranthene benzo[k]fluoranthene benzo[a]pyrene benzolqhi]perylene indeno[l,2,3-cd]pyrene fluoranthene benzo[ b]fluoranthene benzo[k]fluoranthene benzo[alpyrene benzolqhi]perylene indeno[l,2,3-cdlpyrene fluoranthene benzo[b]fluoranthene benzo [klfluoranthene benzo[a]pyrene benzolqhil perylene indeno[l,2,3-cdlpyrene fluoranthene benzo[b]fluoranthene benzo[kl fluoranthene benzo[a]pyrene benzolqhi]perylene indeno[1,2,3-cd]pyrene
0.10 0.05 0.01 0.05 0.05 0.30 0.20 0.10 0.02 0.10 0.10
76 88 100 100 92 110 96 62 110 102 90 99 90 106 103 106 97 105 95 97 100 105 97 98
81 92 92 104 99 92 98 96 98 96
2
0.60
0.30 0.15 0.03 0.15 0.15 0.90 0.40
0.20 0.04 0.20 0.20 1.20
94
92 101 103 101 101 94
91 98 96 96 97 95 90
3
3
I
6
I
Y
a
1.lOFu
3
-F
h
m
1
I
I
4
I
8
I
12
I
16
Time/min. Figure 6. Chromatogramsobtalnedfor the injectionof the surfactantrlch phases of a river water sample (a) and of a splked river water sample (b) after cloud polnt preconcentratlon. Chromatographk condltlons and peak assignment as described in the Experlmental Section.
The PAH identified in the sample of smoke particulates were the following: fluoranthene (1 12 ppm), pyrene (120 ppm), benz[a]anthracene (62 ppm), benzo[b]fluoranthene (1 9 ppm),
h
I
7
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I
I
12 16 20 TImC/mih Figure 7. Chromatogram obtalned for the Injection of a 0.6% Trlton X-114 soldon after 5-h contact with 0.04 g of smoke pertlcuhtea. Chromatographk conditions and peak asdgnmemtas &scribed kr the Experimental S e n . 0
0
1
e
4
8
benzo[k]fluoranthene (9 ppm), benzo[a]pyrene (20 ppm), and benzo[ghi]perylene (1 1 ppm). In order to compare extraction yield when a micellar solution is used with that obtained with the use of organic solvents, an experiment was conducted in which different portions of sample were extracted with 30 mL of methanol Analytical Chemktty, Vol. 66. No. 6, March 15, l9Q4
879
Table 8. Relrtlonahlp botween the Slgnal Obtalned by Extractlng wtth 0.5% Trtton X-114 and wHh Methanol
R" compound fluoranthene Pyrene benz[a]anthracene benzo[b] fluoranthene benzo[k]fluoranthene benzo[a1pyrene benzo[ghil perylene
2h
4h
6h
0.63 0.64 0.61 0.63 0.56 0.47 0.38
1.07 0.98 1.10 0.98 1.01 0.95 0.75
1.19 1.02 1.24 1.10 1.24 1.17 1.02
11
a Ratio of fluorescence peak intensities of samples extracted with 0.5% Triton X-114to that obtained when extracting over 5 hours with methanol.
~~
~~
~~
~~
~~
~~
Table 0. Relatlve Standard Deviation of the PAH Micellar Extractlon Process from Smoke Partlculater compound % rad
fluoranthene Pyrene benz[a]anthracene benzo[b] fluoranthene benzo [k] fluoranthene benzo[aI pyrene benzolqhilperylene
9.8 5.0 4.6 4.8 6.9 5.5 7.5
and acetonitrile and an aqueous solution of 0.5% Triton X-1 14 at room temperature and different contact times. Table 8 shows the relationship between the signal obtained following extraction with Triton X-114and the signal afforded by the sample following extraction over 5 h with methanol. The results show that for this type of sample extraction with the surfactant solution is as effective as when it is performed with organic solvents. The precision of the method applied to smoke samples was determined by preparing eight different samples (-6 mg) that were extracted with 30 mL of an aqueous solution of 0.5% Triton X-114. To do so, after keeping the solution in contact with the solid sample for 1 h, it was filtered through glass wool and 10pL of the solution was injected directly into the chromatographic system. The values of the relative standard deviation obtained for the different compounds identified in the sample are shown in Table 9. It may be seen that these values lie within the normal limits of extraction processes that employ organic solvents. Similar experiments were carried out on samples of wood ash in which the concentration of PAH was much lower than in the samples of smoke particulates. Figure 8 shows the chromatograms corresponding to the extraction and later preconcentration of a 2-gsample of ash (a) and a sample of 0.1 g of ash (b) with 100 mE of an aqueous solution of Triton X-114 at 0.5%. In both cases, the time of contact of the solution with the solid was 5 h. When the amount of sample is large, eight peaks corresponding to eight PAHs of those studied can be identified, while when the amount is small, only the pyrene peak can be identified. The PAH identified in the wood ash samples are as follows: anthracene (30 ppb), fluoranthene (21 ppb), pyrene (20ppb), benz[a]anthracene (9 ppb), benzo[b]fluoranthene (6ppb), benzo[k]fluoranthene (2ppb), benzo[a]pyrene (5 ppb) and benzo[ghi]perylene (3 PPb). In order to check the effect of temperature on the extraction process, a series of experiments was performed in which 000
Analytical Chemistry. Vol. 66, No. 6, March 15, 1994
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0
4
S Timehnin.
1
2
1
6
0
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8
rim.
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Flgure 8. Chromatogram obtained for the injection of the surfactantrich phase obtained after cloud point preconcentration of a 0.5% Triton X-114 solution after 5-h contact with 2 g (a)and 0.1 g (b) of wood ash. Chromatographic conditions and peak assignment as described in the Experimental Section. Table 10. Recoverler of PAH from Splked Wood Aah Samples
compound
concn (ppb) added found
% recovery
fluoranthene
200 400 pyrene
25 50 benzo[k]fluoranthene
8 16
196 376 12.2 37.2 61.6 0.8 8.4 16.8
98 94
100 99 95 100
different portions of sample were extracted (at different times) with the 0.5% solution of Triton X-114 at 25 and 60 OC. Solutions were always reequilibrated at 40 OC for 5 min before cloud point preconcentration was performed. The results obtained indicate that extraction eficiency at 60 O C is -3 times higher than that achieved at 25 OC,attributable to faster liquid-solid extraction kinetics at higher temperature. This ratio remains almost constant for all the PAH identified. In order to check the recoveries of these compounds in the samples of wood ash, three samples of 0.1 g of solid were prepared and extracted with 20 mL of a 0.5% solution of Triton X-114at 60 "C. Two of these samples were spiked with a solution of PAH at two concentrations. The three samples were kept in contact with the solution of surfactant for 30 min and then filtered and subjected to the preconcentration process. The same experiment was carried out at 60,120,180,and 240 min. The recovery values for the spiked samples are good and almost constant after 120 min. As an example, Table 10 shows the recovery for three polycyclic aromatic hydrocarbons for a fixed contact time of 120 min. For the rest of the compounds studied, the values are similar.
CONCLUSIONS The results obtained prove that the use of Triton X-114 provides several advantages for the preconcentration of PAH from water samples. The micelle-mediated methodology avoids sorption of the compounds onto the containers and makes it possible to attain high preconcentration factors through the cloud point phenomenon. The presence of high contents of dissolved organic matter in water samples has no detrimental effect on the recoveries. The high efficacy in the preconcentration step is probably due to the close contact attained between the extractant and the analytes in the sample owing to the fact that, at temperatures lower than the cloud point temperature, the extractant and sample form a homogeneous solution. This latter degree of interaction can never be obtained with waterinmiscible organic solvents in conventional extractions. Furthermore, injection of the surfactant-rich phase containing preconcentrated PAH in the chromatographic system provides additional sensitivity enhancement for fluorescence
detection, probably due to the modification of the microenvironment of the analytes when they reach the detector (viscosity changes and protection of the excited states due to the coeluted surfactant). It has also been demonstrated that micellar solutions of Triton X-114 are as effective as traditional organic solvents for the extraction of PAH from solid matrices, leading to procedures in which organic solvents are eliminated in the sample pretreatment and preconcentration step. Although PAH were used in this work, similar results should be expected for many other neutral compounds.
ACKNOWLEDGMENT This work was supported by DGICYT (Project PB910185). C.G.P. acknowledges the financial support by the Spanish Government (PFPI). Received for review June 17, 1993. Accepted December 3, 1993."
* Abstract
published in Aduunce ACS Abstructs, January 15, 1994.
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