Metabolites of 2,4,4′-Tribrominated Diphenyl Ether (BDE-28) in Pumpkin after In Vivo and In Vitro Exposure Miao Yu, Jiyan Liu,* Thanh Wang, Jianteng Sun, Runzeng Liu, and Guibin Jiang State Key Laboratory of Environmental Chemistry and Ecotoxicology, Research Center for Eco-Environmental Sciences, Chinese Academy of Sciences, Beijing, 100085, China * S Supporting Information ABSTRACT: There is currently limited knowledge on PBDE metabolism in plants although they could play an important role in the environmental transformation of these persistent organic pollutants. In this study, pumpkin (Cucurbita maxima × C. moschata) was chosen as the model to understand the fate of BDE-28 in plants. MeO-tri-BDEs, OH-tri-BDEs, and OH-tri-BDEs were found as metabolites in plant samples of both in vivo hydroponic and in vitro tissue culture exposure. Three MeO-tri-BDEs were further identified as para-substituted metabolites. MeO-BDEs and OH-BDEs, respectively, accounted for about 1.6% and 1.5% (recovery corrected) of initial amount of BDE-28 according to the semiquantitative results. Other PBDEs, especially less brominated PBDEs as impurities in the standard of BDE-28, were also detected. The impurities and evaporation of the standard must be considered when trace metabolites are studied in exposure experiments. ■ INTRODUCTION As a class of additive brominated flame retardants, polybromi- nated diphenyl ethers (PBDEs) have been widely used in many commercial products. 1 The penta- and octa-BDE technical mixtures have recently been listed in the Stockholm Convention on Persistent Organic Pollutants (SC-POPs) in May 2009 2,3 due to their persistence, bioaccumulation and toxicity. As analogues of PBDEs, hydroxylated polybrominated diphenyl ethers (OH-PBDEs) and methoxylated polybromi- nated diphenyl ethers (MeO-PBDEs), have also been detected in various environmental matrices. 4-10 However, the relation- ships between OH-PBDEs, MeO-PBDEs, and corresponding PBDEs are still a matter of discussion, 11,12 which is important to elucidate the fate of PBDEs in the environment. Metabolic transformation of organic contaminant in plants is an important biotransformation process in ecosystems. 13 However, only limited studies have focused on the metabolism of PBDEs in plants. Wang et al. found six debromination products, seven hydroxylated metabolites and four methoxy- lated products in maize after exposure to three BDE congeners. 14 Our previous work 12 investigated the metabolism of 2,2′,4,4′-tetrabromodiphenyl ether (BDE-47) in young whole pumpkin plant and found 2,4,4′-tribrominated diphenyl ether (BDE-28) as a debromination product, four hydroxylated metabolites and a methoxylated product. The fate of PBDEs in the plants affects the behavior of PBDEs in the food web because the plants are the primary trophic level. Thus studies on the metabolism of PBDEs in the plants not only provide the information on health and environmental risks of PBDEs but also supply an important clue for the remediation of those contaminants. In vivo exposure of intact plants is an effective method to study the fate of certain compounds in plants, and can provide information on the distribution of parent compounds and metabolites. However, in vivo studies 12,14-17 cannot completely eliminate the effects of microorganisms even with rigorous protocols. In vitro studies using tissue culture under axenic conditions can usually solve the problems caused by microbes. However, it is usually difficult to extrapolate the results from in vitro to in vivo conditions due to the complexity of the study organisms. Therefore, integrating in vivo and in vitro studies is an effective way to explain the metabolism of contaminants in plants. Some lower brominated diphenyl ethers such as 2,4,4′- tribrominated diphenyl ether (BDE-28) are frequently detected in the environment at relatively high concentrations 18,19 and tend to be bioaccumulative. 20 BDE-28 was also found as a debromination product of BDE-47 in the pumpkin plant. 12 Pumpkin has also been found to have a high bioaccumulation capacity for POPs such as polychlorinated biphenyls (PCBs) 21 and DDT. 22 Therefore, intact pumpkin (Cucurbita maxima × C. moschata) seedlings and tissue cultures were hydroponically exposed to BDE-28 to study the fate of BDE-28 in plants in this work. ■ EXPERIMENTAL SECTION Chemicals. The standard of BDE-28 (solid form, 5 mg, 99.3%) and standard solutions of potential BDE metabolites Received: September 17, 2013 Revised: October 31, 2013 Accepted: November 5, 2013 Published: November 5, 2013 Article pubs.acs.org/est © 2013 American Chemical Society 13494 dx.doi.org/10.1021/es404144p | Environ. Sci. Technol. 2013, 47, 13494-13501