Anal. Chem. 1988,60,1285-1290
t TP TS
TYB
X
x x,
percentage point of Student's t distribution test portion (2), or amount of laboratory sample subjected to analysis; in our case either 3 or 6 g of calcium carbonate treated sample (2), or solution from which the analytical signal is obtained; in our case the 200-mL solution where barium sulfate is precipitated total Youden blank, ordinate intercept of the test portion curve net (blank-corrected) analytical signal -
O/n)'EY=+
raw analytical signal (mg of barium sulfate collected)
LITERATURE CITED (1) Taylor, J. K. Anal. Chem. 1983, 55, 600A-602A, 604A,608A. (2) HorwitZ, W. Nomenclahve of Sampllng in AnalyNcal Chem/stty, IUPAC Commlssh, V. 3,Analytical Nomenclature, Provlslonal Proposal, 14th Draft, 1986. 16 May. (3) K W f f , I. M.; Sandell, E. B.; Meehan. E. J.; Bruckenstein. S. Quantitative Chemlcal Analysis, 4th 4.;MacMillan: London, 1969;Chapter 16. (4) Hunter, J. S.J . Assoc. Off. Anal. Chem. 1981, 6 4 , 574-583. (5) Mitchell, D. G.; Garden, J. S. Talanta 1982, 29, 921-929. (6) Cardone, M. J. J . Assoc. Off. Anal. Chem. 1983, 66, 1257-1282. (7) Cardone, M. J. J . Assoc. Off. Anal. Chem. 1983, 66, 1283-1294. (8) Cardone, M. J. Anal. Chem. 1986, 58, 438-445.
1285
(9) Beukelman, T. E.; Lord, S. S., Jr. Appl. Spectrosc. 1960, 14, 12-17. (IO) Ferriis, R. Anal. Chem. 1987, 59, 2816-2818. (11) Delaney, M. F. Liq. Chromatogr. 1984, 2 , 85-86. (12) Tyson, J. F. Analyst (London) 1984, 109. 313-317. (13) Ferrfis, R.; Torrades. F. Analyst (London) 1985, 1 1 0 , 403-406. (14) Grubbs, F. E.; Beck, G. Technometrics 1972, 14, 847-854. (15) Dlxon, W. J.; Massey, F. J., Jr. Introduction to Statistical Analysis, 4th ed.; McGraw-Hlll: New York, 1983;Chapter 15. (16) Draper, N. R.; Smith, H. Applied Regression Analysis, 2nd ed.; Wiley: New York, 1981;Chapter 1. (17) Franke, J. P.; de Zeeuw. R. A,; Hakkert, R. Anal. Chem. 1978, 5 0 , 1374-1380. (16) Spendley. W. I n Statlstlcal Methods in Research and Productlon, 4th ed.; Davies, 0.L., Goldsmith, P. L., Eds.; Longman: London, 1980; Chapter 4. (19) Mostyn, R. A.; Cunningham. A. F. Anal. Chem. 1966,38, 121-123. (20) Skogerboe, R. K.; Koirtyohann, S. R. Accuracy in Trace Analysis: Sampllng , Sample Handling and Analysis NBS Spec. Pub/. ( U S .) 1978, No. 422, 199-210,through ref 6. (21) Uriano, G. A.; Cali, J. P. I n Validation on the Measurement Process; De Voe, J., Ed.; ACS Symposium Series No. 63;American Chemical Society: Washington, DC, 1977;Chapter 4. (22) Annu. Book ASTMStand. 1980. Part 12: Chemical Analysis of Metals and Metal Bearing Ores. (23) Marchandise, H.; Colinet, E. Fresenius' 2. Anal. Chem. 1983, 316,
669-672. (24) Henning, S.; Jackson, T. L. At. Absorpt. News/. 1973, 12, 100-101. (25) Uriano, G. A.; Gravatt, C. C. CRC, Crit. Rev. Anal. Chem. 1977, 7 , 361-4 1 1. (26) Annu. Book ASTM Stand. 1980. Part 42: Analytical MethodsSpectroscopy; Chromatography; Computerized Systems. Standards E
327-68(1978)and E 572-76,among others.
RECEIVED for review September 9,1987. Accepted February 8, 1988.
Characterization of Multicomponent Mixtures of Polynuclear Aromatic Hydrocarbons with a-Cyclodextrin- Induced Solid-Surface Room-Temperature Luminescence J. M.Bello and R. J. Hurtubise* Chemistry Department, University of Wyoming, Laramie, Wyoming 82071
a-Cyclodextrln-induced solid-surface room-temperature luminescence (CISS-RTL) was used to characterize synthetic mlxtures of polynuclear aromatic hydrocarbons (PAH) adsorbed on a-cyclodextrln-sodlum chloride mixtures. With CISS-RTL, quailtatlve analysis of binary mixtures of benzo[alpyrene and benzo[elpyrene was easily accomplished even at nanogram levels. I n addition, a technique was also developed that was used in characterizing a nine-component PAH mixture. Thls method employs extraction, selective exdtatkm, and detectbn by CISSRTL and sdutbn fluorescence spectroscopy. Nine PAH of various slzes ranging from a two-rlng compound (naphthalene) to a ten-ring compound (decacyclene) were selected as model compounds, and several synthetic mixtures contelning different amounts of the nlne PAH were prepared and were analyzed by this approach. The results showed that between seven to nine PAH were identified In the mixtures ai nanogram levels.
Room-temperature phosphorimetry (RTP) has been proven to be a useful technique for trace organic analysis. The use of RTP in pesticide analysis has been shown by several authors to be analytically viable. Su et al. (I) developed a versatile luminescence sampling system used in the study of the RTP
of certain pesticides. They were able to analyze several synthetic mixtures of phosphorescent pesticides without prior separation by using different substrates and heavy atom perturbers. Vannelli and Schulman (2) also reported that pesticides, which are usually analyzed by gas chromatography, liquid chromatography, and spectrophotometric detection techniques, can be analyzed by RTP. In another area RTP has been useful in the analysis of pharmaceutical compounds ( 3 , 4 )and biologically-important compounds (5). Also, Senthilnathan and Hurtubise (6)developed a quantitative method that employs a combined use of solid-surface room-temperature fluorescence (RTF) and RTP. They were able to analyze various binary and ternary mixtures without prior separation a t the nanogram level with this method. Recently, AsafuAdjaye and Su (7)also used a combined RTP and R T F technique for the analysis of mixtures. With changes in the experimental parameters such as pH of the sample environment, the solid substrate, and the heavy atoms, five and eight luminescent compounds were analyzed in synthetic mixtures. Polynuclear aromatic hydrocarbons (PAH) are an important group of compounds because many PAH have been found to be carcinogenic in laboratory animal tests (8). PAH are found in coal liquid samples or samples originating from coal conversion processes. Vo-Dinh and co-workers have successfully applied a combined use of synchronous fluorescence spectroscopy and the R T P technique to the analysis of PAH in
0003-2700/86/0360-1285$01.50/00 1988 American Chemical Society
1286
ANALYTICAL CHEMISTRY, VOL. 60, NO. 13, JULY 1, 1988
?-
coal liquid samples (9, IO) and of PAH in air samples (11). In this work, a-cyclodextrin-induced solid-surface room-temperature luminescence (CISS-RTL) was used to characterize synthetic mixtures of PAH. With CISS-RTL, qualitative analysis of binary mixtures of benzo[a]pyrene and benzo[elpyrene was easily accomplished. Also, a method that uses extraction, CISS-RTL, solution fluorescence, and selective excitation was developed which can be used to characterize multicomponent mixtures of PAH. With this technique, seven to nine PAH can be qualitatively analyzed in a mixture. EXPERIMENTAL S E C T I O N Apparatus. All RTP, RTF, and solution fluorescence spectra were obtained with a FLUOROLOG 2+2 spectrofluorometer (SPEX Industries, Edison, NJ), and data were processed through a SPEX Datamate computer interfaced with the spectrofluorometer. A cooled Hamamatsu R928 photomultiplier tube was employed. For RTF and solution fluorescence spectra, a 450-W Xe lamp (Osram, Germany) was used. RTP spectra were obtained by employing the SPEX 1934C phosphorimeter accessory with the spectrofluorometer. The phosphorimeter system used a programmable pulsed excitation source and selectable gating of the signal from the photomultiplier tube. All the glassware used were washed with soap, rinsed several times with 50% nitric acidlwater, and then rinsed with distilled water. The glassware were further rinsed with distilled ethanol and then dried in an oven at 100 "C. Reagents. Methanol was Photrex grade and was purchased from J. T. Baker. The cyclohexane (Spectro Grade, Eastman Kodak Co.), 1,2-dichloroethane (Reagent Grade, EM Science), and chloroform (HPLC Grade, J. T. Baker) were used as received. The a-cyclodextrin hydrate (Aldrich) and sodium chloride (Reagent Grade) were washed with distilled ethanol prior to use. Benzo[a]pyrene (Gold Label, Aldrich), naphthalene (Gold Label, Aldrich), coronene (99% pure, Aldrich), benzo[e]pyrene (99% pure, Aldrich), and decacyclene (Aldrich) were used as received. Triphenylene, tetracene, pyrene, and phenanthrene were recrystallized from distilled ethanol prior to use. The nitrogen gas was a high-purity grade and was passed through an Oxy-trap (Alltech Associates) to remove oxygen. Procedures: Samples Preparation for the Characterization of Binary Mixtures of Benzo[a]pyrene and Benzo[elpyrene. Several binary mixture solutions of benzo[a]pyrene and benzo[e]pyrene were prepared from methanol. A 0.2-mL methanol aliquot was added into a test tube, and then a 5-pL aliquot of the methanol solution containing the binary mixture of benzo[a]pyrene and benzo[e]pyrene was added to the methanol solvent. Next, 85 mg of an 80% a-cyclodextrin-sodium chloride mixture was added into the test tube, and the resulting slurry was sonicated for a few seconds. The solution was then placed in an oven a t 100 "C and dried for 15 min. After drying, the powdered sample was mixed, and then 60 mg of the powder was packed loosely onto the depression of the triangular brass plate sample holder. The sample holder was placed in the sample compartment of the SPEX instrument, and the RTP and RTF spectra were obtained. For RTP measurements, the samples were degassed with nitrogen for 10 min prior to recording the spectra and also continued during the span of the measurement. The same procedure was followed for the reference spectra of benzo[a]pyrene and benzo[e]pyrene and for the blanks. To record the RTF emission spectra of the binary mixtures and the benzo[a]pyrene reference, the samples were excited a t 369 and 300 nm, respectively. On the other hand, the RTP emission spectra of the binary mixtures and the benzo[e]pyrene reference were recorded by exciting the samples a t 284 nm. Extraction of P A H from the a-Cyclodextrin-Sodium Chloride Mixtures. A 0.5-pg sample of the PAH was adsorbed onto an a-cyclodextrin-sodium chloride mixture by using the procedure described above for the characterization of binary mixtures of benzo[a]pyrene and benzo[e]pyrene. The cu-cyclodextrin-sodium chloride powder containing the adsorbed PAH was then transferred into a 10-mL beaker. The test tube used to prepare the a-cyclodextrin sample was rinsed 5 times with 1-mL portions of cyclohexane, and after each rinsing the cyclohexane was added to the a-cyclodextrin-sodium chloride powder in the
WIXTURE
adsorb
0.5% a-CD
cyclohexane extract 1
solid 1
1. evaporate cyclohexane
2. redissolve in
methanol 3 . readsorb
1
1 . dry 2 . d e t e c t by RTP and RTF
I n a p h t h a l e n e , phenanthrene, pyrene, t e t r a c e n e
aoz a-CD
Cyclohexane extract 2
Solution fluorescence
solid 2 1 . dry 2 . d e t e c t by RTF and RTP
B(a)P, Bte)P. decacyclene
Figure 1. Schematic diagram of the method employed for the separation and characterization of the multicomponent PAH mixtures: cu-CD, a-cyclodextrin; B(e)P, benzo[e]pyrene; B(a)P,benzo[a Ipyrene.
beaker. By use of a magnetic stirrer, the solution was stirred for about 30 min. The solution was then transferred quantitatively into a test tube and was centrifuged. The cyclohexane extract was decanted into a 5-mL volumetric flask and diluted to volume. The corrected solution fluorescenceemission spectrum of the PAH extract was then recorded at the excitation band maximum of the PAH. Extraction was also carried out on an a-cyclodextrin-sodium chloride blank, and the corrected solution fluorescence emission spectrum of the cyclohexane extract of the a-cyclodextrin-sodium chloride blank was recorded. The spectrum of the blank was subtracted from the spectra of the PAH extracts. In addition, a cyclohexane solution containing the amount of the PAH that would be obtained with a complete extraction was also prepared, and the corrected fluorescence emission spectrum of this solution was obtained. The percentage of PAH extracted from the a-cyclodextrin-sodium chloride mixture was then calculated by taking the ratio of the fluorescence intensities a t the emission band maxima of the solution fluorescence spectra of the PAH extract and the reference. Characterization of a Multicomponent PAH Mixture. A schematic diagram of the method employed for the characterization of the multicomponent PAH mixtures is shown in Figure 1. All five nine-component PAH mixture solutions used were prepared in chloroform. A 5-pL aliquot of the chloroform solution containing the nine PAH was added to a test tube. The test tube was then placed in an oven at 100 "C for 5 min to evaporate the chloroform. Then the test tube was cooled to room temperature, and 0.20 mL methanol was added to the test tube. The solution was then sonicated for several minutes to dissolve the PAH. Next, 150 mg of 0.5% a-cyclodextrin-sodium chloride mixture was added to the solution and the mixture was sonicated for a few seconds. The methanol solvent was then evaporated a t 100 "C for 15 min, and after drying, the powdered sample was cooled and then transferred into a 10-mL beaker. Extraction was then carried out on the 0.5 % a-cyclodextrin-sodium chloride sample with cyclohexane employing the extraction procedure described above. The 0.5% a-cyclodextrin-sodium chloride solid and the cyclohexane extract were separated by centrifuging the solution. As shown in Figure 1, the 0.5% a-cyclodextrin-sodium chloride mixture was dried, and the PAH remaining in this substrate were characterized by RTP and RTF. In the 0.5% a-cyclodextrinsodium chloride mixture, naphthalene and phenanthrene were characterized by RTP, while pyrene and tetracene were characterized by RTF. On the other hand, the cyclohexane extract from the 0.5% a-cyclodextrin-sodium chloride mixture was transferred to a small vial and was treated as follows. First, the cyclohexane was evaporated gently in a hot plate, and then after the vial was cooled to room temperature, 0.2 mL of methanol was added and the mixture was sonicated for several minutes to dissolve the PAH. Next, 85 mg of an 80% a-cyclodextrin-sodium chloride mixture
ANALYTICAL CHEMISTRY, VOL. 60, NO. 13, JULY 1, 1988
was added, and the slurry was sonicated and was placed in an oven at 110 "C for 15 min to evaporate the methanol. After drying, the 80% a-cyclodextrin-sodium chloride powder was transferred quantitativelyinto a 10-mL beaker by rinsing the vial several times with cyclohexane. The cyclohexane solution containing the 80% a-cyclodextrin-sodium chloride sample was then transferred to a test tube and was centrifuged. The 80% a-cyclodextrin-sodium chloride solid was dried, and compounds remaining in the 80% mixture were characterized by RTP and RTF. Benzo[a]pyrene was characterized in the 80% a-cyclodextrin-sodium chloride mixture by RTF, while benzo[e]pyrene and triphenylene were characterized by RTP. On the other hand, the cyclohexane extract from the 80% a-cyclodextrin-sodium chloride mixture was analyzed by solution fluorescence. The PAH that were Characterized in the extract were coronene and decacyclene. The total analysis time for each mixture was about 2 h. In addition, a blank 0.5% a-cyclodextrin-sodium chloride mixture was prepared and treated with the same procedure as discussed. The spectra of the 0.5% and 80% a-cyclodextrin-sodium chloride blanks and the cyclohexane extract from the 80% a-cyclodextrin-sodium chloride mixture blank were obtained and were subtracted from the appropriate spectra of the PAH in the mixture.
1287
3.1BE 86
3
-I-
_VI C aJ -IC
Y
278.88
178.88
678.88
Wave) ength (nml
RESULTS AND DISCUSSION Characterization of Binary Mixtures of Benzo[a ]pyrene and Benzo[e Ipyrene. Recently, we reported that an 80% a-cyclodextrin-sodium chloride mixture can be used as a matrix to obtain R T F and R T P analytical data (12,13). Cyclodextrins are macrocyclic carbohydrate molecules composed of a-(l,l)-linked glucose units arranged in a torus. Cyclodextrins have been shown to form inclusion complexes with a wide variety of organic and inorganic compounds that can fit into their hydrophobic cavity (14,15).In this work, we investigated the possibility of using CISS-RTL to characterize binary mixtures of benzo[a]pyrene and benzo[e]pyrene. It was reported previously that benzo[a]pyrene gave only RTF on an 80% a-cyclodextrin-sodium chloride mixture, whereas benzo[e]pyrene gave both R T F and RTP on this matrix (13). In addition, there was no overlap in the RTF excitation spectra of benzo[a]pyrene and benzo[e]pyrene a t wavelengths greater than 345 nm, with only benzo[a]pyrene absorbing radiation in this region of the spectrum. Furthermore, the R T F intensity of benzo[e]pyrene was 16 times less than the R T F intensity of benzo[a]pyrene. Thus, identification of benzo[a]pyrene and benzo[e]pyrene was straightforward with the a-cyclodextrin matrix. For example, benzo[e]pyrene can be characterized by RTP, whereas benzo[a]pyrene can be characterized by R T F using a 369-nm excitation wavelength. Several binary mixtures of benzo[a]pyrene and benzo[e]pyrene were prepared and then qualitatively analyzed by CISS-RTL. In Figure 2a, the RTF emission spectra of benzo[a]pyrene in several binary mixtures adsorbed on the 80% a-cyclodextrin-sodium chloride mixture are shown along with the standard R T F emission spectrum of benzo[a]pyrene adsorbed on the same matrix. In each of the binary mixtures, the amount of benzo[e]pyrene was kept constant a t 500 ng, while the amount of benzo[a]pyrene was varied from 20 to 500 ng. As can be seen in Figure 2a, benzo[a]pyrene can be readily detected in the presence of benzo[e]pyrene even at low nanogram amounts. Figure 2b shows the RTP emission spectra for several binary mixtures of benzo[e]pyrene and benzo[a]pyrene adsorbed on the 80% a-cyclodextrin-sodium chloride mixture and the standard R T P emission spectrum for benzo[e]pyrene adsorbed on the a-cyclodextrin matrix. By comparison of the spectra of benzo[e]pyrene in the binary mixtures with the R T P reference spectrum of benzo[e]pyrene, it can be seen that benzo[e]pyrene can be identified at nanogram levels with a relatively large amount of benzo[a]pyrene present. These results further show the advantage of using both RTF and R T P in characterizing mixtures (6, 7,16). In
0.BBE 88 488.80
688.88
808.88
Wave)ength [nml Figure 2. (a) RTF spectra of benzo[a]pyrene (-) on 80% a-cyclodextrin-sodium chloride in the presence of 500 ng of benzo[e]pyrene and RTF spectrum of 500 ng of benzo[a]pyrene (---) on 80 % a-cyclodextrin-sodium chloride. Amounts of benzo[a ] pyrene in the mixtures were as follows: (A) 500 ng; (B) 100 ng; (C) 50 ng; (D) 20 ng. (b) RTP spectra of benzo[e]pyrene (-) on 80% acyclodextrin-sodium chloride In the presence of 500 ng of benzo[alpyrene and RTP spectrum of 500 ng of benzo[e]pyrene (---) on 80 % a-cyclodextrin-sodium chloride. Amounts of benzo [e ]pyrene in the mixtures were as follows: (A) 500 ng; (6) 100 ng; (C) 50 ng; (D) 20 ng.
addition, no heavy atom was used in characterizing the mixtures.
Conditions for Characterizing a Multicomponent Mixture of PAH with CISS-RTL. The analytical applicability of CISS-RTL to the qualitative analysis of a multicomponent mixture of PAH was also investigated. Previously, we reported that a large number of PAH give R T P and/or RTF on an 80% a-cyclodextrin-sodium chloride mixture (13). In addition, we have shown that the formation of an inclusion complex of the analyte with the a-cyclodextrin is an important mechanism of interaction for the observation of RTP and RTF in an a-cyclodextrin-sodium chloride mixture (17). Thus, by use of the size exclusion feature of a-cyclodextrin, a mixture of PAH of various sizes could be analyzed by CISS-RTL. For example, large PAH molecules would interact less effectively with a mixture of a-cyclodextrin-sodium chloride which contained a relatively small amount of a-cyclodextrin. However, smaller PAH molecules should readily interact with the a-cyclodextrin-sodium chloride mixture. Thus, larger PAH that would not be included or only partially included in the a-cyclodextrin cavity could be extracted from the acyclodextrin-sodium chloride matrix and analyzed by some other means such as solution fluorescence spectroscopy or could be readsorbed on another a-cyclodextrin matrix containing a larger percentage of a-cyclodextrin and characterized by CISS-RTL. In addition, the PAH remaining in the a-cyclodextrin-sodium chloride matrix after extraction could also
1288
ANALYTICAL CHEMISTRY, VOL. 60, NO. 13, JULY 1, 1988
Table I. Percentage of Analytes Extracted with Cyclohexane in the 0.5% a-Cyclodextrin-Sodium Chloride and in the 80% a-Cyclodextrin-Sodium Chloride Mixtures
Table 11. Conditions for Characterizing a Multicomponent Mixture of PAH
interfering
% extracted
0.5% CY-CD decacyclene coronene benzo[elpyrene benzo[a]pyrene triphenylene pyrene tetracene phenanthrene naphthalene
93
90 84 71 80 29 24 0 0
80% a-CD 89 40 28 6 0 0 4 0 0
be analyzed by CISS-RTL. Preliminary studies were done to determine which PAH could be extracted from a-cyclodextrin-sodium chloride mixtures. Nine PAH were chosen as model compounds, and the nine PAH were of various sizes ranging from a two-ring compound (naphthalene) to a ten-ring compound (decacyclene). Also, several organic solvents such as cyclohexane, a 50% cyclohexane-chloroform mixture, and 1,2-dichloroethane were investigated as possible solvents for the extraction. These solvents were selected because a-cyclodextrin is not soluble in these solvents, and therefore these solvents would not dissolve the included complexes. It was found, however, that the amounts of the PAH extracted in an a-cyclodextrin-sodium chloride mixture were relatively the same in all of the solvents investigated. Thus, cyclohexane was chosen as the solvent for the extraction of PAH. Table I lists the nine PAH and the percentage of analyte extracted in 0.5% and 80% a-cyclodextrin-sodium chloride mixtures. I t can be seen in this table that large amounts of triphenylene, benzo[a]pyrene, decacyclene, coronene, and benzo[e]pyrene were extracted from the 0.5% mixture. However, large portions of tetracene, pyrene, naphthalene, and phenanthrene still remained in the 0.5% a-cyclodextrin-sodium chloride mixture after extraction. On the other hand, the two large compounds, decacyclene and coronene, were extracted at a level of 89% and 40%, respectively, in the 80% a-cyclodextrin-sodium chloride mixture. The results in Table I allowed the development of a scheme of characterizing the nine PAH listed in the table. As shown in Table I, only four compounds remained in the 0.5% a-cyclodextrin-sodium chloride mixture in relatively high amounts after extraction. Thus, by extraction of a large percentage of five of the PAH from the 0.5% mixture, the characterization of the compounds remaining in this substrate was less complicated (Figure 1). In addition, tetracene only gave RTF on the 0.5% a-cyclodextrin-sodium chloride mixture, and its excitation and emission wavelength ranges are from 360 to 450 nm and from 460 to 610 nm, respectively. These ranges are far removed from the RTF excitation and emission spectral range for the other three PAH (pyrene, phenanthrene, and naphthalene) remaining in this substrate. The RTF and R T P spectral characteristics of pyrene, tetracene, phenanthrene, and naphthalene were studied in detail to access interferences from spectral overlap of the various RTF and R T P bands. It can also be seen from Table I that by readsorbing the extract from the 0.5% a-cyclodextrin-sodium chloride mixture, which would contain mainly benzo[a]pyrene, benzo[elpyrene, triphenylene, coronene, and decacyclene, onto an 80% a-cyclodextrin-sodium chloride mixture, and then by performing extraction on the 80% a-cyclodextrin sample, only three PAH, triphenylene, benzo[a]pyrene, and benzo[e]pyrene, would be present in relatively high amounts on the 80% a-cyclodextrin-sodium chloride mixture. Benzo[a]pyrene
component
substrate or
phenannaphthathrene lene naphtha- phenanlene threne benzo[a]- none
0.5% a-CD
compound
pyrene
pyrene
benzo [a]-
solvent
method
excitation wavelength, nm
RTP
223
0.5% a-CD RTP
244
80% a-CD
RTF
369
0.5% u-CD
RTF
339
80% a-CD
RTP
335
pyrene
benzo[e]- coronene pyrene coronene none tetracene
none
triphenylene
none
decanone cyclene
cyclohexane solution fluorescence 0.5% a-CD RTF 80% wCD RTP
444 261
cyclohexane solution
378
301
fluorescence
gives only RTF in the 80% a-cyclodextrin mixture, while benzo[e]pyrene and triphenylene give both R T P and RTF. The R T F emission spectrum of benzo[a]pyrene overlaps slightly with the RTF emission spectra of benzo[e]pyrene and triphenylene in this substrate. However, at wavelengths greater than 360 nm, only benzo[a]pyrene absorbs radiation. On the other hand, the RTP emission bands of triphenylene and benzo[e]pyrene are well separated. Furthermore, triphenylene does not absorb radiation at wavelengths greater than 330 nm. It is also evident from Table I that the extract from the 80% a-cyclodextrin-sodium chloride mixture would contain mostly decacyclene and coronene at this point because a large percentage of benzo[a]pyrene and benzo[e]pyrene would remain on the 80% mixture after extraction and no triphenylene would be removed. Thus, characterization of coronene and decacyclene in the 80% a-cyclodextrin extract would be readily accomplished by solution fluorescence spectroscopy. This is due to the fact that the solution fluorescence emission bands of coronene and decacyclene do not overlap. In addition, the fluorescence emission intensity of coronene is much stronger than decacyclene, and also coronene does not absorb radiation at wavelengths greater than 350 nm. Table I1 summarizes the conditions for the characterization of the nine PAH. In Table 11, it is shown that the interfering component for the pyrene adsorbed on a 0.5% a-cyclodextrin-sodium chloride mixture is benzo[a]pyrene. As indicated in Table I, 29% of benzo[a]pyrene remains on the 0.5% a-cyclodextrin-sodium chloride mixture after extraction. There is a slight overlap between the RTF emission spectra of pyrene and benzo[a]pyrene in this a-cyclodextrin mixture; however, all of the relevant RTF emission bands used in identifying pyrene are situated in the region where benzo[alpyrene gives a minimum emission or does not emit RTF at all. In addition, the excitation wavelength used in characterizing pyrene was selected at a region in the excitation spectrum where benzo[a]pyrene does not absorb radiation strongly. Table I1 also indicates that coronene interferes with the identification of benzo[e]pyrene in the 80% a-cyclodextrin-sodium chloride mixture. In Table I, it is shown that only 40% of coronene was extracted in the 80% a-cyclodextrin mixture. Since a significant overlap between the R T P excitation and emission spectra of coronene and benzo[e]pyrene was observed, the unextracted coronene in the 80% a-cyclodextrin mixture would therefore interfere with the identifi-
ANALYTICAL CHEMISTRY, VOL. 60, NO. 13, JULY 1, 1988 6.50E-81
I
I
1
k
Table 111. Amount of Polynuclear Aromatic Hydrocarbons in the Synthetic Mixtures and Compounds Characterized in the Mixtures
I\
?- t
amount: ng A
VI
C
+aC
'
c-(
0.0BE 08 208. 80
588. m
880.00
Wavel ength (nml 2.10E 86 I
1289
1
1
(b)
I
B
C
phenanthrene naphthalene benzo [ a ]pyrene pyrene benzo[elpyrene coronene tetracene triphenylene decacyclene
D
E
500(+) 500(+) 500(+) 500(+) 500(+) 500(+) 500(+) 500(+) 500(+)
loo(+) 500(+) 500(+)
loo(+) loo(+) 500(+)
50(+)
50(+)
loo(+)
"The compounds that were characterized are indicated by (+) and the compounds that were not characterized are indicated by (-).
,
5.58-02
t-
t
U
1 I
0.80E 88 280.00
i
588.88
880.88
Wavel ength [nml Flgure 3. (a) RTP spectra of the solid 2 fraction (see Figure 1) of mixture D (-) and a standard sample of triphenylene (---) adsorbed on 80% a-cyclodextrin-sodium chloride. (b) RTF spectra of the solid 2 fraction of mixture D (-) and a standard sample of benzo[e ] pyrene (- - -) adsorbed on 80% a-cyclodextrin-sodium chloride.
cation of benzo[e]pyrene in a mixture containing the two compounds. However, the R T P emission bands of both compounds in the 80% a-cyclodextrin-sodium chloride mixture were sharp, and the bands were distinguishable from one another. For example, benzo[e]pyrene has bands a t 538, 548, and 588 nm, while the bands for coronene are at 529,564, and 611 nm. In addition, the R T P of coronene can also be significantly diminished by selecting an excitation wavelength where coronene absorbs little radiation. Characterization of Several Multicomponent Mixtures of PAH. Five synthetic mixtures of the nine PAH were prepared and then were characterized according to the scheme shown in Figure 1. T o identify the nine PAH in a mixture, a t least two emission bands were used to identify the PAH. In addition, the guideline used in matching the emission bands was a i 2 nm difference between the reference bands and the mixture bands. Table I11 lists the amount of each PAH in the five synthetic mixtures and summarizes the results of the characterization of the five PAH synthetic mixtures. As shown in Table I1 the emission bands of decacyclene, triphenylene, tetracene, coronene, and benzo[a]pyrene do not have any interfering components. Thus, as shown in Table I11 these compounds were readily identified in all of the five synthetic mixtures. Representative emission spectra are shown in Figure 3 for triphenylene and benzo[a]pyrene in mixture D. As shown in this figure, there is an excellent match between the spectra obtained from the mixtures and the standard spectra. The RTF emission spectrum of pyrene also overlaps slightly with the R T F emission spectrum of the unextracted benzo[alpyrene in the 0.5% a-cyclodextrin-sodium chloride mixture. However, despite the overlap of the two spectra pyrene was characterized in all of the five synthetic mixtures as shown
0 . m BB 288.88
588. 00
888.88
Wavelength [nml
I
0.BBE 00 208.00
588.88
880. 08
Wavel ength [nml Flgure 4. (a) RTP spectra of the solid 2 fraction (see Figure 1) of mixture 6 (-) and a standard sample of benzo[e]pyrene (---) adsorbed on 80 % a-cyclodextrin-sodium chloride. (b) RTP spectra of the solid 2 fraction of mixture D (-) and a standard sample of benzo[e Ipyrene (- - -) adsorbed on 80 % a-cyclodextrin-sodium chloride.
in Table 111. The RTP emission spectrum of benzo[e]pyrene also overlaps considerably with the spectrum of the coronene for these compounds on the 80% a-cyclodextrin-sodium chloride mixture after extraction. The results of the characterization of benzo[e]pyrene in the mixtures showed that in mixtures B and C, where there were 10-fold and &fold excess of coronene, respectively, benzo[e]pyrene was not characterized. As shown in Figure 4a, the bands of the RTP emission spectrum of the solid 2 fraction of mixture B cannot be matched with the R T P emission bands of the benzo[e]pyrene reference. The bands of the RTP emission spectrum of the solid 2 fraction of mixture B, however, match exactly that of coronene which was shown (Table 11)to interfere with
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ANALYTICAL CHEMISTRY, VOL. 60, NO. 13, JULY 1, 1988
benzo[e]pyrene. On the other hand, benzo[e]pyrene was characterized in the other three mixtures where benzo[e]pyrene was present in excess or as the same amounts of coronene. A representative spectrum of these mixtures is shown in Figure 4b for the solid 2 fraction of mixture D, which shows that at least two of the RTP emission bands for this fraction of mixture D can be matched with that of the benzo[ejpyrene reference. One important feature of the above approach for mixture analysis is the large number of compounds that could be characterized. In the examples presented above, at least seven to nine PAH were characterized in a particular mixture. However, the number of PAH identified in the synthetic mixtures are certainly not the maximum number of compounds that could be characterized with this technique. For example, the possibility of using synchronous fluorescence and derivative spectroscopy with this method was not investigated. For instance, by use of synchronous fluorescence and/or derivative spectroscopy in conjunction with the above approach, it is possible to obtain narrower spectral bands and therefore decrease the spectral interferences from other luminescent compounds. Another advantage of the technique developed is the large number of organic compounds giving RTP and/or RTF on an a-cyclodextrin-sodium chloride mixture (13). For example, a large number of nitrogen heterocycles, carbonyl aromatics, hydroxyl aromatics, and other substituted aromatic compounds gave R T P and/or RTF on this matrix. Thus, it can be seen that the application of this approach is not limited to the analysis of PAH. Also, the simplicity of this approach, and the inherent sensitivity and selectivity of luminescence techniques are added advantages. One further advantage of this method is that only a small amount (5 pL) of the sample is needed for the analysis. One disadvantage of this approach, however, can be found in the extraction procedure employed. It can be seen in Table I that for compounds such as coronene and benzo[a]pyrene complete extraction of these compounds in the a-cyclodextrin-sodium chloride mixture was not achieved. And as discussed above, the unextracted compounds in the a-cyclodextrin-odium chloride matrix can interfere with the identification of some of the PAH. In addition, the less than 100% extraction of the compounds in a-cyclodextrin-sodium chloride would also limit the quantitative application of this approach in mixture analysis at its present stage of development.
With highly complex samples such as industrial samples, an initial isolation of a PAH fraction would have to be carried-out to minimize interference from other luminescent components. A further separation of the fraction might be necessary depending on the complexity of the fraction. This could be accomplished by high-performance liquid chromatography (HPLC). The combined use of HPLC and the luminescence approach discussed in this paper should prove to be a very effective approach for the characterization of PAH. For example, Ford and Hurtubise (18) separated benzomquinoline and phenanthridine from shale oil by open-column chromatography and HPLC and then identified the nitrogen heterocycles by room-temperature solid-surface fluorescence and phosphorescence techniques. Also, Dalterio and Hurtubise (16) have discussed second derivative solid-surface luminescence analysis of two-component HPLC fractions. Registry No. a-CD, 10016-20-3;decacyclene, 191-48-0; coronene, 191-07-1;benz[e]pyene, 192-97-2;benz[a]pyene, 50-32-8; triphenylene, 217-59-4; pyrene, 129-00-0; tetracene, 92-24-0; phenanthrene, 85-01-8; naphthalene, 91-20-3.
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RECEIVED for review October 20, 1987. Accepted February 16, 1988. Financial support for this project was provided by the Department of Energy, Division of Basic Energy Sciences, Grant DE-FG02-86ER13547.
