Iowa Superfund Basic Research Program: Program Support Core: Synthesis Core- Past Highlights

Synthesis Core- Past Highlights

Studies of environmental and toxic effects of polychlorinated biphenyls (PCBs) are ideally performed with PCB mixtures reflecting the composition of environmental PCB profiles to mimic actual effects and to account for complex interactions among individual PCB congeners. Unfortunately, only a few laboratory studies employing synthetic PCB mixtures have been reported, in part because of the challenges associated with the preparation of complex PCB mixtures containing many individual PCB congeners.

The objective of this study was to develop a PCB mixture that resembles the average PCB profile recorded from 1996 to 2002 at a satellite station of the Integrated Atmospheric Deposition Network located at the Illinois Institute of Technology (IIT) in Chicago, Illinois, using commercial PCB mixtures. Initial simulations, using published Aroclor profiles, showed that a mixture containing 65% Aroclor 1242 and 35% Aroclor 1254 was a good approximation of the target profile. A synthetic Chicago air mixture (CAM) was prepared by mixing the respective Aroclor’s in this ratio, followed by GC/MS/MS analysis. Comparison of the PCB profile of the synthetic mixture with the target profile suggests that the synthetic PCB mixture is a good approximation of the average IIT Chicago air profiles (similarity coefficient cos θ = 0.82; average relative percent difference = 84%). The synthetic CAM was also a reasonable approximation of the average of 184 PCB profiles analyzed in 2007 at 37 sites throughout Chicago as part of the University of Iowa Superfund Research Program (isrp), with a cos θ of 0.70 and an average relative percent difference of 118%.

While the CAM and the two Chicago air profiles contained primarily di- to pentachlorobiphenyls, higher chlorinated congeners, including congeners with seven or eight chlorine atoms, were underrepresented in the synthetic CAM. The calculated TCDD toxic equivalency quotients of the synthetic CAM (2.7 ng/mg PCB) and the IIT Chicago air profile (1.6 ng/mg PCB) were comparable, but lower by two orders of magnitude than the isrp Chicago air profile (865 ng/mg PCB) due to surprisingly high PCB 126 levels in Chicago air. In contrast, the calculated neurotoxic equivalency quotients of the CAM (0.33 mg/mg PCB) and the two Chicago air profiles (0.44 and 0.30 mg/mg PCB, respectively) were similar. This study demonstrates the challenges and methods of creating and characterizing synthetic, environmental mixtures of PCBs.

Figure 1: Comparison to the synthetic CAM with the average PCB profile in Chicago air. (A) Average PCB IIT Chicago air profile; (B) PCB profile of the synthetic CAM consisting of 65% Aroclor 1242 and 35% Aroclor 1254; (C) difference in percentage of the PCB congener profiles of both mixtures.

Figure 2. Homologue composition of (A) Aroclor 1242, (B) Aroclor 1254, (C) the synthetic CAM, (D) the average IIT Chicago air profile and (E) the average isrp Chicago air profile.

Several polychlorinated biphenyls (PCBs) and their hydroxylated metabolites display axial chirality. The Synthesis Core provided a series of chiral methoxylated PCB (MeO-PCB) standards to develop an enantioselective, gas chromatographic separation of MeO-PCBs using a chemically bonded -cyclodextrin column (Chirasil-Dex). The atropisomers of several MeO-PCBs could be separated on this column with resolutions ranging from 0.42-0.87 under isothermal or temperature programmed conditions. In addition, the enantiomeric fraction of OH-PCB 136 metabolites was determined in male and female rats treated with racemic PCB 136. As shown in the Figure, the second eluting enantiomer (E2) of 3’-MeO-PCB 150 and 5-MeO-PCB 136 was enriched in the liver of male (A) and female (B) Sprague-Dawley rats. Rats received two intraperitoneal injection of racemic PCB 136 (2 × 100 mol/kg body weight, 2 × 36.1 mg/kg body weight) dissolved in corn oil (5 ml/kg body weight) on day 1 and 4, and were euthanized on day 7 (IS = PCB 204 as internal standard) (I. Kania-Korwel et al.; J. Chromatogr. A 2008: 1207, 146-154 ).

We recently reported improved syntheses of PCB congeners (and their metabolites) containing two or more ortho-chlorine substituents. (A) The Suzuki coupling reaction at 110 °C yielded PCB congeners with a 2,2’-substitution pattern in good yields (78-99%), but failed to give PCB congeners with 3 or 4 ortho chlorine substituents. (B) Symmetrically substituted PCB congeners with multiple ortho chlorine substituents were obtained in 20-52% yields using a modified Ullmann coupling reaction. The yield of the coupling increased with increasing degree of chlorination of the starting material. The modified Ullmann coupling reaction employed much milder reaction conditions (copper-bronze/CuCl in N-methylpyrrolidinone, 110°C) and, therefore, appears to be advantageous compared to the classical Ullmann coupling reaction (copper-bronze, no solvent, 230°C). These modified reaction conditions allow the synthesis of large quantities of pure, non-dioxin-like PCB congeners for environmental and toxicological studies by overcoming problems associated with classical PCB synthesis strategies (Telu et al., Environ. Int., 2009).

We developed a a straightforward synthesis of a series of ten PCB sulfate monoesters from the corresponding hydroxylated PCBs for Project #3. The hydroxylated PCBs were synthesized by coupling chlorinated benzene boronic acids with appropriate brominated (chloro-)anisoles, followed by demethylation with boron tribromide. The hydroxylated PCBs were sulfated with 2,2,2-trichloroethyl chlorosulfate using DMAP as base. Deprotection with zinc powder/ammonium formate yielded the ammonium salts of the desired PCB sulfate monoesters in good yields. These compounds are now availalbe to study their chemical stability, physicochemical properties and interaction with biological targets, such as albumin and other proteins (Li et al., Environ. Int., 2009).

PCB sulfate monoesters are currently investigated by Project #3. Their sensitive and selective analysis by gas chromatography-mass spectrometry (GC-MS) requires their derivatization, for example, as PCB 2,2,2-trichloroethyl (TCE) sulfate monoesters. To aid in the identification of unknown PCB sulfate metabolites isolated from biological samples, the electron impact MS fragmentation pathways of selected PCB TCE sulfate diesters were analyzed and compared to the fragmentation pathways of the corresponding methoxylated PCBs. As shown for sulfuric acid 2´,5´-dichlorobiphenyl-4yl ester 2,2,2-trichloroethyl ester, the most abundant and characteristic fragment ions of PCB TCE sulfate diesters were formed by releasing CHCCl3, SO3, HCl2 and/or CCl3 from the TCE sulfate moiety and Cl2, HCl, ethyne and chloroethyne from an intermediate phenylcyclopentadienyl cation. Knowledge of the fragmentation patterns of PCB TCE sulfate diesters will greatly aid in determining the position of sulfate moiety (ortho vs. meta/para) of unknown PCB sulfate metabolites isolated from environmental or laboratory samples (Li et al., Chem. Cent. J., 2009).

Polychlorinated biphenyls (PCBs) can be metabolized via hydroxylated and dihydroxylated metabolites to PCB quinone intermediates. We have recently demonstrated that both dihydroxy PCBs and PCB quinones can form semiquinone radicals (SQ·-) in vitro. These semiquinone radicals are reactive intermediates that have been implicated in the toxicity of lower chlorinated, airborne PCB congeners. To further investigate this observation, we synthesized a series of PCB hydroquinones and quinones with different degrees of chlorination of the (hydro-)quinone ring system. These PCB (hydro-)quinones readily react with oxygen or via comproportionation to yield the corresponding semiquinone free radicals, as detected by electron paramagnetic resonance spectroscopy. Surprisingly, the greater the number of chlorines on the (hydro-)quinone (oxygenated) ring, the higher the steady-state level of the resulting semiquinone radical at near neutral pH (Y. Song et al., J. Org. Chem. 2008:73, 8296-8304. DOI: 10.1021/jo801397g)

Crystal structure of PCB 77 (3, 3’, 4, 4’ -tetrachlorobiphenyl), a dioxin-like PCB congener. (a) Molecular structure of PCB 77 showing the atom-labeling scheme and (b) view of PCB 77 along the C1–C1′ axis illustrating the non-planar conformation of the molecule. Displacement ellipsoids are drawn at the 50% probability level. Unlabeled atoms are at the symmetry position (1 − x;1 − y;z). (Shaikh et al., Chemosphere 2008)

doi:10.1016/j.chemosphere.2007.07.017
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