A UPLC-MS/MS method for quantification of metabolites in the ethylene biosynthesis pathway and its biological validation in Arabidopsis.

The plant hormone ethylene is of vital importance in the regulation of plant development and stress responses. Recent studies revealed that 1-aminocyclopropane-1-carboxylic acid (ACC) plays a role beyond its function as an ethylene precursor. However, the absence of reliable methods to quantify ACC and its conjugates malonyl-ACC (MACC), glutamyl-ACC (GACC), and jasmonyl-ACC (JA-ACC) hinders related research. Combining synthetic and analytical chemistry, we present the first, validated methodology to rapidly extract and quantify ACC and its conjugates using ultra-high-performance liquid chromatography coupled to tandem mass spectrometry (UPLC-MS/MS). Its relevance was confirmed by application to Arabidopsis mutants with altered ACC metabolism and wild-type plants under stress. Pharmacological and genetic suppression of ACC synthesis resulted in decreased ACC and MACC content, whereas induction led to elevated levels. Salt, wounding, and submergence stress enhanced ACC and MACC production. GACC and JA-ACC were undetectable in vivo; however, GACC was identified in vitro, underscoring the broad applicability of the method. This method provides an efficient tool to study individual functions of ACC and its conjugates, paving the road toward exploration of novel avenues in ACC and ethylene metabolism, and revisiting ethylene literature in view of the recent discovery of an ethylene-independent role of ACC.

Growth Hormones in Broad Bean (Vicia faba L.) and Radish (Raphanus raphanistrum subsp. sativus L.) Are Associated with Accumulated Concentrations of Perfluoroalkyl Substances.

In this study, we grew radish (Raphanus raphanistrum subsp. sativus L.) and broad beans (Vicia faba L.) in a greenhouse on soils spiked with a mixture of 15 per- and polyfluoroalkyl substances (PFASs) and investigated the association between accumulated ∑PFAS concentrations, growth, and hormone levels. Short-chained PFASs dominated aboveground tissues, whereas long-chained PFASs were most abundant in the plant roots. Our results showed that the presence or absence of exodermal Casparian strips, as well as the hydrophobicity and anion exchange capacities of PFASs, could explain the translocation of PFASs within plants. Significant associations found between accumulated PFAS concentrations and levels of gibberellins (GA1 and GA15), methionine, and indole-3-acetic acid (IAA) imply potential effects of PFASs on plant development and growth. This study provides the first evidence of associations between PFAS accumulation in plants and growth hormone levels, possibly leading to growth reduction of the apical dome and effects on the cell cycle in pericycle cells and methionine metabolism in plants.

Assessment of poly- and perfluoroalkyl substances (PFAS) in commercially available drinking straws using targeted and suspect screening approaches.

Many food contact materials (FCMs) and reusable plastics in the food industry contain poly- and perfluoroalkyl substances (PFAS), a group of synthetic pollutants that are known to be potentially harmful for wildlife, humans, and the environment. PFAS may migrate from FCMs to food consumed by humans. As a replacement for plastics, often paper and other plant-based materials are used in commercial settings. This also applies to drinking straws, where plant-based and other presumably eco-friendly straws are increasingly used to reduce plastic pollution. In order to make these materials water-repellent, PFAS are added during manufacturing but can also already be present early in the supply chain due to the use of contaminated raw materials. In the present study, we examined the PFAS concentrations in 39 different brands of straws, made from five materials (i.e. paper, bamboo, glass, stainless steel, and plastic) commercially available on the Belgian market. We combined both targeted and suspect-screening approaches to evaluate a wide range of PFAS. PFAS were found to be present in almost all types of straws, except for those made of stainless steel. PFAS were more frequently detected in plant-based materials, such as paper and bamboo. We did not observe many differences between the types of materials, or the continents of origin. The presence of PFAS in plant-based straws shows that they are not necessarily biodegradable and that the use of such straws potentially contributes to human and environmental exposure of PFAS.

Accumulation and effects of perfluoroalkyl substances in Arabidopsis thaliana in a temperature-dependent manner: an in vitro study.

The replacement of long-chained per- and polyfluoroalkyl substances (PFAS) with their short-chained homologues may have an impact on the accumulation in plants. The extent to which PFAS are absorbed by plants may differ among species and may depend on environmental factors, including temperature. The effect of an increased temperature on root uptake and translocation of PFAS in plants has been poorly studied. In addition, very few studies have examined toxicity of environmentally realistic PFAS concentrations to plants. Here, we investigated the bioaccumulation and tissue-distribution of fifteen PFAS in Arabidopsis thaliana L. grown in vitro at two different temperatures. Additionally, we examined the combined effects of temperature and PFAS accumulation on plant growth. Short-chained PFAS mainly accumulated in the leaves. The perfluorocarboxylic acid (PFCA) concentrations in roots and leaves, and the relative contribution of PFCAs to the ΣPFAS concentrations increased with carbon chain length regardless of temperature, with the exception of perfluorobutanoic acid (PFBA). An increased uptake of PFAS in leaves and roots at higher temperatures was observed for PFAS containing either eight or nine carbon atoms and could hence potentially result in higher risks for human intake. Leaf:root ratios of PFCAs followed a U-shaped pattern with carbon chain length, which is attributed to both hydrophobicity and anion exchange. Overall, no combined effects of realistic PFAS concentrations and temperature on the growth of A. thaliana were observed. PFAS exposure positively affected early root growth rates and root hair lengths, indicating a potential effect on factors involved in root hair morphogenesis. However, this effect on root growth rate became negligible later on in the exposure, and solely a temperature effect was observed after 6 days. Temperature also affected the leaf surface area. The underlying mechanisms on how PFAS stimulates root hair growth require further examination.

Perfluoroalkylated compounds in the eggs and feathers of resident and migratory seabirds from the Antarctic Peninsula.

In this study, we investigated factors that influence the differences in exposure of perfluoroalkyl acids (PFAAs) from eight species of Antarctic seabirds, including Pygoscelis penguins, Stercorarius maccormicki, and Macronectes giganteus. We analyzed the relationship between foraging ecology (based on δ13C, δ15N, and δ34S values) and PFAAs accumulated in eggs and breast feathers. Ten out of 15 targeted PFAAs were detected in eggs compared to eight in feathers. Mean ∑PFAA concentrations in feathers ranged from 0.47 in P. antarcticus to 17.4 ng/g dry weight (dw) in S. maccormicki. In eggs, ∑PFAA concentrations ranged from 3.51 in P. adeliae to 117 ng/g dw in S. maccormicki. The highest concentrations of most PFAAs were found in trans-equatorial migrators such as S. maccormicki, probably due their high trophic position and higher concentrations of PFAAs in the Northern Hemisphere compared to the Southern Hemisphere. Based on stable isotopes correlations, our results suggest that the trophic position (δ15N) and the foraging area (δ13C and δ34S) influence PFAAs concentrations in Antarctic seabirds. Our results point to the possibility that long-distance migratory birds may have as bio-vectors in the transport of pollutants, including PFCAs, in Antarctic environments, although this must be further confirmed in future studies using a mass balanced approach, such as extractable organofluorine (EOF).

A rapid method for the detection and quantification of legacy and emerging per- and polyfluoroalkyl substances (PFAS) in bird feathers using UPLC-MS/MS.

The bioaccumulation and toxicity of per- and polyfluoroalkyl substances (PFAS) have raised scientific and public concern in recent decades, leading to regulatory measures for some PFAS (e.g. perfluorooctane sulfonate (PFOS) and perfluorooctanoic acid (PFOA)). In addition, the discovery of new PFAS alternatives in the environment has led to growing concern about the presence of numerous other PFAS that are used unrestricted. Feathers have been successfully applied as non-destructive indicators for various contaminants, mostly metals and persistent organic pollutants (POPs), whereas their suitability as an indicator for PFAS is still discussed. Previous studies on PFAS in feathers have focused primarily on perfluoroalkyl sulfonic acids (PFSAs) and perfluoroalkyl carboxylic acids (PFCAs); analytical methods for other groups of PFAS or PFAS alternatives in feathers are still lacking. Hence, this study aimed to develop a rapid, sensitive and reliable analytical method for determining a broad range of PFAS (N = 32) in feathers, using liquid chromatography-tandem mass spectrometry (LC-MS/MS). An extraction duration of 24 h was found to be sufficient to extract the majority of PFAS from the feathers. The extraction recovery of the internal standards ranged on average from 68% (PFBA) to 97% (PFOS). The spike recovery was within an acceptable range of at least 70% for most of the target analytes and the precision was often > 80%. A further extract clean-up using weak anion exchange (WAX) solid phase extraction (SPE), was proven unnecessary, as it resulted in a similar or lower spike recovery, and, as a consequence, a lower precision and higher quantification limit. The analytical method allows detection of low PFAS concentrations in a low quantity of matrix (i.e. small feathers). The developed LC-MS/MS method was validated and shown to be a fast, sensitive and reliable method for determining a broad range of legacy and emerging PFAS in feathers.

Effects of the UV filter, oxybenzone, adsorbed to microplastics in the clam Scrobicularia plana.

Microplastics (MPs) lipophilic nature and widespread distribution raises concerns due to their increasing presence in the marine environment and their ability to adsorb organic contaminants, as being potential vehicles for transport and potential source of accumulation of organic contaminants by marine organisms. The organic UV-filter, oxybenzone (BP-3) is a constituent of sunscreens and personal care products, entering the marine environment either by direct contact with swimmers or by wastewater effluents. In this study the ecotoxicological effects of exposure to low-density polyethylene (LDPE) microplastics with and without adsorbed BP-3 were investigated in the peppery furrow shell clam, Scrobicularia plana. LDPE microplastics with a size range of 11-13 µm were previously contaminated with an environmentally relevant concentration of BP-3 (82 ng g-1). S. plana individuals were exposed to a concentration of 1 mg L-1 of microplastics with and without BP-3 adsorbed in a water-sediment exposure system for 14 days. Clams were sampled at the beginning of the experiment and after 3, 7, and 14 days of exposure. Multiple biomarkers were analysed to investigate the effect of exposure in different clam tissues, gills, digestive gland, and haemolymph. Antioxidant (superoxide dismutase, catalase, glutathione peroxidase) and biotransformation (glutathione-S-transferases) enzyme activities, oxidative damage (lipid peroxidation), genotoxicity (single and double strand DNA breaks), and neurotoxicity (acetylcholinesterase activity) were assessed along with two biomarker indexes to assess the overall health status. Results indicate that after 7 days of exposure MPs with adsorbed BP-3 induced oxidative stress and damage, when compared to exposure to virgin MPs and control treatments. Neurotoxic effects were also noted in MPs with adsorbed BP-3 after 14 days exposure, while some evidence points to increased genotoxicity with exposure time. Overall results indicate that gills were more affected by exposure to microplastics than digestive gland and that biomarkers alterations are apparently more related to the toxicity of BP-3 adsorbed than virgin MPs alone.

Development and validation of an extraction method for the analysis of perfluoroalkyl substances (PFASs) in environmental and biotic matrices.

Although long chained PFASs have been phased-out in several countries, their persistence in the environment and bioaccumulative potential cause the environmental and biotic concentrations to remain high, highlighting the need to further monitor these pollutants. Currently several methods are used for the quantification of perfluoroalkyl substances (PFASs) in biological matrices including different ways to correct for recovery losses, each with its specific pros and contras. With this paper we aim to re-evaluate current methodologies and to create an updated new analytical guideline that is applicable for both abiotic and biotic matrices. The developed LC/MS/MS method was validated and shown to be specific, selective, linear, robust and sensitive. Reliable results could still be obtained 6 days after extraction. The recoveries varied, depending on the matrix, between 1% and 100%, but nevertheless, a high accuracy was obtained even at the lowest recoveries. A reduction of sample mass could significantly increase method recoveries and therefore it is highly recommended to take less matrix. We confirmed that using the ISTD closest in terms of functional group and carbon chain length is a suitable method for the quantification of PFASs that lack a corresponding ISTD. The newly described method was, depending on the matrix, similar in terms of sensitivity and reliability compared to a frequently used method and could be used simultaneously in future monitoring studies. Therefore, we recommend to select the purification method based on the target analytes as well as the sample matrix. CAPSULE: The newly described method was similar in terms of sensitivity and reliability compared to a frequently used method and a selection of purification methods should be based on the target analyte and sample matrix.