P.E.G. Leonards

High-resolution separation of polychlorinated biphenyls by comprehensive two-dimensional gas chromatography

Comprehensive two-dimensional gas chromatography (GC x GC) with micro electron-capture detection (microECD) has been optimised for the separation of polychlorinated biphenyl congeners with emphasis on the separation of 12 toxic non- and mono-ortho chlorinated biphenyls (CBs), viz. CBs 77, 81, 105, 114, 118, 123, 126, 156, 157, 167, 169 and 189. The selection of the first- and second-dimension columns and the temperature programme optimisation were carried out with a mixture of 90 CBs and the results are compared with those of one-dimensional GC. A complete separation of all 12 priority CBs was obtained with two column combinations, HP-1-HT-8 and HP-1-SupelcoWax-10. With the HP-1-HT-8 column set, ordered structures show up in the two-dimensional plane, with the number of chlorine substituents and their position (ortho vs. non-ortho) being the main parameters of interest. This can help with congener identification. Estimated detection limits are excellent, i.e. about 10 fg. To illustrate the potential and the versatility of GC x GC-microECD, a cod liver extract and a standard mixture of the 17 most toxic polychlorinated dibenzo-p-dioxins and polychlorinated dibenzofurans together with 90 CBs were analysed as an application.

Determination of organotin compounds in the foodweb of a shallow freshwater lake in The Netherlands

An extensive study on the presence of nine organotin compounds (OTs) in a freshwater foodweb was made, using newly developed analytical procedures in order to obtain insight in accumulation and degradation processes. Tributyltin (TBT), Triphenyltin (TPT) and their degradation products were detected. Zebra mussels, eel, roach, bream, pike, perch, and pike perch and cormorant showed high OT body concentrations.At the lower trophic levels, phenyltin concentrations were higher in benthic species while butyltin concentrations were higher in pelagic species. This indicates that TBT is passed on primarily via the water, while TPT is passed on to a larger extent via the sediment.At the higher trophic levels, net bioaccumulation of TPT was greater than that of TBT, resulting in relatively higher TPT concentrations. High concentrations of biodegradation products of TBT, but not of TPT, were found in the livers of fish and birds, which indicates that TBT is more easily metabolized than TPT.A comparison with literature data of fish lethal body concentrations revealed that fish in the field may be endangered. With birds, the highest concentrations of OTs were present in liver and kidney and not in subcutaneous fat, which confirms that OTs accumulate via different mechanisms than traditional lipophilic compounds. As a whole the OT concentrations found in the foodweb may be considered to be quite alarming.

Critical review of the analysis of non- and mono-ortho-chlorobiphenyls

The various methods for the determination of non-ortho and mono-ortho-chlorobiphenyls are critically reviewed. Matrix, sample preparation, extraction, clean-up, fractionation and group separation methods, chromatographic separation (gas, liquid and supercritical fluid chromatography), as well as the various detection methods, multi-residue methods, quality control and method validation are discussed. For each topic, an overview is given of the current status of the field and recommendations for the most appropriate analytical approach are presented.

Brominated flame retardants and endocrine disruption

From an environmental point of view, an increasing important group of organohalogen compounds are the brominated flame retardants (BFRs), which are widely used in polymers and textiles and applied in construction materials, furniture, and electronic equipment. BFRs with the highest production volume are the polybrominated diphenyl ethers (PBDEs), tetrabromobisphenol A (TBBP-A), and hexabromocyclododecane (HBCD). Because of their persistence and low biodegradation profile, several of the PBDE congeners accumulate in biota and are widely found in the aquatic food chain. Their levels in the environment and in humans have increased during the last decades, in contrast to compounds such as polychlorinated biphenyls (PCBs) and dichlorodiphenyltrichloroethane (DDT), for example. Humans may be exposed to PBDEs mainly through consumption of fatty food of animal origin (e.g., fish), but exposure through skin contact with textiles protected with flame retardants or through inhalation of BFRs volatilized from electronic and electric equipment may also occur. The levels of PBDEs in Swedish human milk showed a doubling in concentration every five years over the period 1972 to 1997 (2,2,4,4-tetraBDE being the predominant congener). The levels of penta- and hexa-BDEs increased at the same rate in ringed seals collected in the Canadian Arctic from 1981 to 2000. PBDEs exhibit a great variety of biological effects, depending on the bromine substitution pattern. PBDEs are potential endocrine disrupters, based on shared toxicity with the structurally related PCBs, polychlorinated dibenzofurans (PCDFs), and polychlorinated dibenzo-p-dioxins (PCDDs) (partial aryl hydrocarbon- [Ah-] receptor agonist and antagonist activity in vitro, thyroid toxicity, and immune effects), including developmental toxicity. The potency of TBBP-A to interact with thyroid hormone homeostasis is indicated from in vitro studies in which the compound competes with thyroxin (T4) for binding to transthyretin (TTR). So far, the toxicological profile of many BFRs is too incomplete and insufficient to perform an adequate risk assessment, and further information is required regarding the potential for endocrine disruption of these compounds that are of increasing environmental concern.

Attempt to unravel the composition of toxaphene by comprehensive two-dimensional gas chromatography with selective detection

Comprehensive two-dimensional gas chromatography (GC x GC) coupled with micro electron-capture and time-of-flight mass spectrometric (TOF-MS) detection has been used to analyse technical toxaphene. An HP-1 x HT-8 column combination yielded highly structured chromatograms and revealed a complex mixture of over 1000 compounds what is significantly higher number than in any study before. The analysis of a mixture of 23 individual congeners and TOF-MS evaluation of technical toxaphene showed that the chromatogram is structured according to the number of chlorine substituents in a molecule. The nature of the compounds (bornane and camphene) does not appear to have any influence. The sum of the peak areas of all congeners in each group was calculated using laboratory-written software; based on these results, the composition of technical toxaphene as a function of the number of chlorine substituents was provisionally calculated and was found that hepta- and octachlorinated compounds represents 75% of the total toxaphene area.

Bioconcentration of polycyclic heteroaromatic hydrocarbons in the guppy (Poecilia reticula)

Abstract Bioconcentration factors (BCFs) of 19 parent and hetero polycyclic aromatic hydrocarbons (PAH) were obtained from static bioconcentration experiments with Poecilia reticulata by monitoring the decrease in the initial aqueous concentration. The BCFs were obtained by fitting the observed concentrations to a one-compartment first-order kinetic model. The BCFs thus obtained showed a satisfactory linear relationship with the experimental n-octanol/water partition coefficient, log Kow (r = 0.91, s = 0.30). By comparison with a limited number of semi-static tests, it was concluded that the static bioconcentration experiment is a satisfactory alternative for more laborious bioconcentration tests to obtain BCFs of (hetero) PAH. However, the static tests are less valuable for the acquisition of kinetic bioconcentration parameters such as, e.g., uptake or elimination rate constants. For several of the compounds in this study, the estimates resulting from the static tests did not match with those from a limited number of depuration experiments carried out in this study or with values expected from the relationship between the depuration rate constant and the log Kow, reported in the literature. The distribution of three parent PAH within the fish after a 7-day semi-static exposure was also investigated. Relatively high concentrations were found in liver and kidneys. The major part of the body burden was found in muscle and the gastro-intestinal tract.

Novel Analytical Methods for Flame Retardants and Plasticizers Based on Gas Chromatography, Comprehensive Two-Dimensional Gas Chromatography, and Direct Probe Coupled to Atmospheric Pressure Chemical Ionization-High Resolution Time-of-Flight-Mass Spectrometry

In this study, we assess the applicability of different analytical techniques, namely, direct probe (DP), gas chromatography (GC), and comprehensive two-dimensional gas chromatography (GC × GC) coupled to atmospheric pressure chemical ionization (APCI) with a high resolution (HR)-time-of-flight (TOF)-mass spectrometry (MS) for the analysis of flame retardants and plasticizers in electronic waste and car interiors. APCI-HRTOFMS is a combination scarcely exploited yet with GC or with a direct probe for screening purposes and to the best of our knowledge, never with GC × GC to provide comprehensive information. Because of the increasing number of flame retardants and questions about their environmental fate, there is a need for the development of wider target and untargeted screening techniques to assess human exposure to these compounds. With the use of the APCI source, we took the advantage of using a soft ionization technique that provides mainly molecular ions, in addition to the accuracy of HRMS for identification. The direct probe provided a very easy and inexpensive method for the identification of flame retardants without any sample preparation. This technique seems extremely useful for the screening of solid materials such as electrical devices, electronics and other waste. GC-APCI-HRTOF-MS appeared to be more sensitive compared to liquid chromatography (LC)-APCI/atmospheric pressure photoionization (APPI)-HRTOF-MS for a wider range of flame retardants with absolute detection limits in the range of 0.5-25 pg. A variety of tri- to decabromodiphenyl ethers, phosphorus flame retardants and new flame retardants were found in the samples at levels from microgram per gram to milligram per gram levels.

Characterizing field sediments from three European river basins with special emphasis on endocrine effects - A recommendation for Potamopyrgus antipodarum as test organism

The assessment of endocrine disrupting potentials of field sediments has until now been mostly limited to classical chemical analysis, in vitro assays and in vivo bioassays performed with vertebrates. There is an urgent need for easy, cheap and reproducible invertebrate tests which may be applied in certain monitoring activities. Since the mudsnail Potamopyrgus antipodarum is known to be tolerant to natural stressors, but also sensitive to endocrine disrupting chemicals, it is very likely that this organism could be suitable for the assessment of endocrine effects of e.g. field sediments. Within this study the endocrine potential of sediments in three European river basins was assessed. The yeast estrogen screen (YES) and a sediment contact test with P. antipodarum were performed. Furthermore, analyses of physico-chemical properties and concentrations of heavy metals, PAHs, organotins, natural steroids and alkylphenols were done. In the sediment contact test, the reproduction of the snail was promoted by a part of the sediments. This phenomenon could not be explained by their physico-chemical properties. However, at some of those sites a high estrogenic activity was detected in the YES, leading to the assumption that endocrine disrupting compounds could be responsible for those effects. This assumption could be confirmed to some extent with partially high concentrations of xeno-estrogens (e.g. nonylphenol) at the certain sites. Our study demonstrates the applicability of the test with P. antipodarum for a variety of sediments and once again points out the need of suitable in vivo biotests for the risk assessment of field sediments.

Identification of Photosynthesis Inhibitors of Pelagic Marine Algae Using 96-Well Plate Microfractionation for Enhanced Throughput in Effect-Directed Analysis

Because of large-scale production and use of an increasing diversity of chemicals in modern society, estuarine and coastal waters may be contaminated with numerous substances. Some of these compounds have the potential to affect microalgae at the base of the pelagic food chain. Therefore, we identified the main chemical stressors that negatively affect the effective photosystem II efficiency (ϕPSII) in marine microalgae of the Dutch estuarine and coastal waters. An enhanced effect-directed analysis (EDA) was carried out by combining reversed-phase ultra performance liquid chromatography fractionation of extracts from passive samplers, followed by effect assessment using the pulse amplitude modulation fluorometry assay and chemical analysis of biologically active fractions using high-resolution mass spectrometry. This study focuses on a novel microfractionation technique using 96-well plates to enhance throughput in EDA, structure elucidation, and the analytical and effect confirmation of the compounds that are identified. Although there are numerous unknown compounds present in estuarine and coastal waters, our EDA study shows that atrazine, diuron, irgarol, isoproturon, terbutryn, and terbutylazine are the main contributors to the observed effect on the ϕPSII of marine microalgae.

Testing Endocrine Disruption in Biota Samples: A method to Remove Interfering Lipids and Natural Hormones

A cleanup method was developed to remove coextracted lipids and natural hormones from biota samples in order to test the endocrine-disrupting (ED) capacity of their extracts in in vitro bioassays. Unspiked and spiked fish tissues were cleaned with a combination of dialysis, gel permeation chromatography (GPC), and normal-phase liquid chromatography (NP-HPLC). The spiking mixture consisted of a broad range of environmental pollutants (endocrine disruptors and genotoxic compounds). Chemical recoveries of each test compound, and thyroid-hormone-like and (anti)androgenic activities of the cleaned extracts were investigated. Despite the chemical and toxicological complexity of the spiking mixture and the sequential sample treatment, chemical analysis revealed acceptable recoveries on average: 89 ± 8% after each cleanup step separately and 75 ± 3% after the whole extraction and cleanup procedure in the extracts. In addition, recovered activities in the bioassays were in good agreement with the spiking levels. The developed cleanup method proved to be capable of lipid and natural hormone removal from fish extracts, enabling the measurement of selected endocrine-hormone-like activities in T(4)*-TTR and AR-CALUX bioassays. The method can be used as a sample preparation method of biota samples for toxicity profiling and effect-directed analysis (EDA).