in Pacifiers:
Jacksan 0. Lay, Jr. Barbara J. Miller National Center for Toxicological Research Jefferson, Ark. 72079
Phthalate esters are widely used plasticizers in flexible polyvinyl chloride (PVC). Of the phthalates, dioctyl phthalate bas been the most commonly used; in 1979 it represented about 26% of the total phthalate plasticizer consumption in the United States. The dioctyl phthalate isomer used for plasticization, 12-benzenedicarboxylic acid bis(2-ethylhexyl)ester, also frequently named di(2-ethylbexyl) phthalate or DEHP, is reported to be an animal carcinogen. DEHP is found in such diverse materials a5 children’s products, clothing, footwear, flooring, home furnishings, housewares, hardware, sporting goods, and personal products. Furthermore, DEHP is typically used in the PVC contained in these products at levels as high as 50%to achieve complete plasticization. Because DEHP is not chemically bound to PVC, significant migration of the plasticizer into the environment i6 possible. Indeed, DEHP has been detected in air, food, and water samples. These findings, as well as other studies, which have shown that DEHP could be detected in blood that had been stored in plastic bags, have resultlhis attide not subject Io US. Copyright Published 1987 American Chemical Socier,
ANALYTICAL APPROACH
ed in an increased awareness of the POtential for human exposure to DEHP resulting from the use of PVC products. In addition, migration of DEHP from PVC might also lead to human exposure via direct dermal or oral contact. For example, the presence of DEHP in some baby pacifiers may have exposed infants via direct oral uptake of DEHP from PVC. This specific concern has led to a voluntary ban on the use of dioctyl phthalate in baby pacifiers. Attempts to veri.
fy that this or other uses of DEHP have been discontinued require the development of very rapid analytical methods so that large numbers of samples can be examined in a timely manner. We have recently investigated the analysis of PVC pacifiers for DEHP (dioctyl phthalates) at levels of about 30% by weight. Although alternative methods had been developed for the analysis of DEHP in PVC based on lengthy extraction followed by gas chromatogra-
mg DEHPllO mg DDP
Figure 1. Relative response for MHP and W P [MH]+ ions. ANALYTICAL CEMISTRY. VOL. 59, NO. 22. NOVEMBER 15, 1987
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In addition, we found that FAB-MS could be utilized for qualitative screening of PVC samples for phthalates by direct introduction of the solid PVC material into the mass spectrometer ion source. R e p a r a h d sa-
phy or gas chromatographylmass spectrometry (GC or GCIMS), we believed a simpler and faster method for the determination of dioctyl phthalates in PVC could he developed that would require neither extraction nor chromatographic separation. As described below, we were able to develop a method for the determination of DEHP (dioctyl phthalate) in PVC at levels typically found in consumer products (-30%). This method is based on the addition of a reference material, didecyl phthalate (DDP), to a solution of the pacifier in tetrahydrofuran (THF) with qnantitation hy fast atom bombardment mass spectrometry (FAB-MS). Previous reports of quantitative measurements employing FAB-MS suggested to us that this ionization technique might be amenable to our analytical problem, especially if we employed an internal standard. We were interested in FAB not for the usual reasons associated with the analyte being too polar or unstable for the application of an alternative ionization method, but rather because of our desire to minimize or eliminate all chromatographic and extraction steps. We reasoned that it ought to be possible to quantitate the amount of DEHP based on the protonated molecules of DEHP and DDP. Also,we expected that some selective ionization of the analyk and reference standard rather than bulk PVC might occur because of the high molecular weight of the PVC polymer. 1324,.
For the quantitative studies standard solutions were prepared that contained a fixed amount of DDP (50 mg) and a variable amount of DEHP (0, 10,25, or 50 mg) in 5 mL of THF. The samples were prepared hy dissolving a weighed pacifier sample of ahout 10 mg in 1.0mL of THF containing 10mglmL of the internal standard, DDP. Then FAB mass spectra were acquired using a Kratos (Manchester, England) MS-50 equipped with an M-Scan (Ascot on Berkshire, England) fast atom source and gun. Full-scan spectra were acquired by scanning down from about
1000 to 100 daltons in 30 8. Samples of either the standard solution or the pacifier solution (1 rL) were applied to a stainless steel target along with 2-3 p L of thioglycerol. To survey or screen the pacifiers qualitatively for phthalates, a small sliver of the pacifier material was removed from the bulk sample with a razor. This sliver (about 1mm X 6 mm and about 0.5 mm thick) was then affixed to the target when the stainless steel surface was wetted with thioglycerol, which was also then added to the upper surface of the sample (upon which the fast atom beam impinged) before being admitted through the vacuum lock into the MS ion source.
Quantitation of DEHP
Quantitation of DEHP in the PVC pacifiers was based on the relative signal levels of the [MH]+ions of DEHP and
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I
Figure 2. The partial FABMS specbum of a sampie of PVC pacifier (sample A)
Ions dissolved in 1 mL of THF containing 10 m g l d DDP. showing the intense [MI+ for DEHP and DDP.
ANALYTICAL CHEMISTRY, VOL. 59. NO. 22, NOVEMBER 15. 1987
DDP (internal standard) obtained from full-scan FAB spectra. Full-scan spectra were used so that the presence of other plasticizers (including any phthalates in addition to the dioctyl phthalates) might be detected. Knowledge of the relative response factors for DEHP and DDP from the calibration curve allowed the percent DEHP to be rapidly determined simply hy OhseNing the ratio of the intensities of the two protonated molecules (DEHP and DDP) in the sample solutions. Data for the calibration curve were obtained hy the FAB-MS analysis of the standard solutions, which contained 10 mg/mL DDP and 0, 1.0, 5.0, or 10 mg/mL DEHP. The calibration curve was prepared by applying a linear least-squares fit of the data (see box), plotted with milligrams of DEHP per 10 mg DDP internal standard as a function of the ratio of the intensity for the [MH]+ ions from DEHP and DDP (Figure 1). The least-squares line has an intercept of -0.056 ( u = 0.042) and a slope of 0.238 ( u = 0.0069).The value of 0.238 for the slope indicates that the DEHP gives more than two times the signal per unit mass for its [MH]+ ion than does DDP. The region of the mass spectra encompassing the protonated molecules of the pacifier solutions showed large signals for the [MH]+ ions of DEHP and DDP (Figure 2). In addition, a single large signal corresponding to a wellknown phthalate fragment ion was also observed at mlz 149. No other ionsincluding any associated with the bulk of the PVC polymer-were observed, even though no chromatographic separation or sample cleanup was employed. In this instance, selective ionization (or perhaps an inability to detect ions from the high-molecularweight polymer) led to significant enhancement of the analytically useful ions in the FAB m a s spectra. Analysis of four commercial pacifiers (A-D) showed DEHP concentrations of 3142% by weight. These values are
the same, within the experimental error, as the results determined for the same lots of PVC pacifiers obtained by extraction with analysis hy GC. Replicate analyses on the same solutions showed relative standard deviations (due to variability in intensity ratio measurements) as high as lo%,but they were generally lower with an average of about 5% (Table I). We have also investigated the use of direct FAB-MS on a small section of untreated pacifier sample introduced directly into the mass spectrometer ion source for qualitative screening of samples containing high levels of DEHP. A small sliver of material was affixed to the target using thioglycerol,which was also applied to the surface of the sample in the path of the fast atom beam. All four PVC pacifiers showed intense signals for DEHP and no other signals except those from the FAB matrix (Figure 3). A sample of a latex pacifier known not to contain DEHP, on the other hand, showed no phthalate signals (spectrum not shown). The use of this direct introduction technique should allow several pacifier samples to he screened for DEHP in as little kw an hour with no s m p l e preparation. We have developed an analytical method employing FAB-MS for the analysis of DEHP (including any isomeric dioctyl phthalates) in baby pacifiers that does not require sample extraction, cleanup, or chromatographic separation. Using this method we have found levels of DEHP in four haby pacifiers that were, within the experimental error, equivalent to the values determined using the more traditional method-extraction followed hy GC. We believe that this FAB-MS method is potentially more efficient than alternative methods for the analysis of DEHP in baby pacifiers and other PVC products containing high levels of DEHP because it eliminates all of the extraction and cleanup steps. The FAB-MS method is considerably faster than alternative methods, and any er-
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P*
Figure 3. The FAB-MS spectrum of a portion of intact PVC pacifier (sample A) ahered directly to the FAB probe tip.
ror or loss of precision associated with
sample handling should he reduced when it is used. In principle, potential prohlems associated with sample homogeneity could also he reduced or eliminated by the analysis of solutions containing whole (or even several) pacifiers. Direct FAB-MS of untreated sections of pacifiers has also been shown to he a very rapid method for the screening of PVC samples for high levels of DEHP.
Jackson 0. Lay, Jr., receiued a Ph.D. from the University of Nebraska in 1982. I n 1985 he joined the staff of the National Center for Toxicological Research, where he is currently a research chemist in the Division of Biochemical Toxicology’s mass spectrometry laboratory. He also is an adjunct assistant professor of chemistry with the University of Arkansas at Little Rock. His research interests include the mass spectral studies of conjugated metabolites, DNA-adducts, and other biomolecules, as well (IS the deuelopment and application of mass spectral methods designed to inuestigate the oxidative modification of DNA.
Barbara J. Miller receiued B.S. and M.S.degrees from Central State Uniuersity (Edmond, Okla.) in 1969 and 1979, respectively. She joined the National Center for Toxicological Researchin 1979, whereshe is currently a chemist with the Diuision of Chemistry. Her research interests include the identification of N-nitrosamines in foodstuffs, baby pacifiers, and other consumer products, as well as the deuelopment of new methods for the analysis of uarious components in animal feeds.
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