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The monitoring of Advanced Oxidation/Reduction Processes (AO/RPs) for the evaluation of the yield and mechanisms of decomposition of perfluorinated compounds (PFCs) is often a more difficult task than their determination in the environmental, biological or food samples with complex matrices. This is mostly due to the formation of hundreds, or even thousands, of both intermediate and final products. The considered AO/RPs, involving free radical reactions, include photolytic and photocatalytic processes, Fenton reactions, sonolysis, ozonation, application of ionizing radiation and several wet oxidation processes. The main attention is paid to the most commonly occurring PFCs in the environment, namely PFOA and PFOS. The most powerful and widely exploited method for this purpose is without a doubt LC/MS/MS, which allows the identification and trace quantitation of all species with detectability and resolution power depending on the particular instrumental configurations. The GC/MS is often employed for the monitoring of volatile fluorocarbons, confirming the formation of radicals in the processes of C‒C and C‒S bonds cleavage. For the direct monitoring of radicals participating in the reactions of PFCs decomposition, the molecular spectrophotometry is employed, especially electron paramagnetic resonance (EPR). The UV/Vis spectrophotometry as a detection method is of special importance in the evaluation of kinetics of radical reactions with the use of pulse radiolysis methods. The most commonly employed for the determination of the yield of mineralization of PFCs is ion-chromatography, but there is also potentiometry with ion-selective electrode and the measurements of general parameters such as Total Organic Carbon and Total Organic Fluoride. The presented review is based on about 100 original papers published in both analytical and environmental journals., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Biotransformation of 6:2 FTOH [F(CF2)6CH2CH2OH] by the white-rot fungus, Phanerochaete chrysosporium, was investigated in laboratory studies. 6:2 FTOH is a raw material increasingly being used to replace products that can lead to long-chain perfluoroalkyl carboxylic acids (PFCAs, ≥ 8 carbons). During a product's life cycle and after final disposal, 6:2 FTOH-derived compounds may be released into the environment and potentially biotransformed. In this study, P. chrysosporium transformed 6:2 FTOH to perfluorocarboxylic acids (PFCAs), polyfluorocarboxylic acids, and transient intermediates within 28 days. 5:3 Acid [F(CF2)5CH2CH2COOH] was the most abundant transformation product, accounting for 32-43 mol % of initially applied 6:2 FTOH in cultures supplemented with lignocellulosic powder, yeast extract, cellulose, and glucose. PFCAs, including perfluoropentanoic (PFPeA) and perfluorohexanoic (PFHxA) acids, accounted for 5.9 mol % after 28-day incubation. Furthermore, four new transformation products as 6:2 FTOH conjugates or 5:3 acid analogues were structurally confirmed. These results demonstrate that P. chrysosporium has the necessary biochemical mechanisms to drive 6:2 FTOH biotransformation pathways toward more degradable polyfluoroalkylcarboxylic acids, such as 5:3 acid, with lower PFCA yields compared to aerobic soil, sludge, and microbial consortia. Since bacteria and fungi appear to contribute differently toward the environmental loading of FTOH-derived PFCAs and polyfluorocarboxylic acids, wood-rotting fungi should be evaluated as potential candidates for the bioremediation of wastewater and groundwater contaminated with fluoroalkyl substances.

6:2 FTI [F(CF2)6CH2CH2I] is a principal industrial raw material used to manufacture 6:2 FTOH [F(CF2)6CH2CH2OH] and 6:2 FTOH-based products and could enter aerobic environments from possible industrial emissions where it is manufactured. This is the first study to assess 6:2 FTI aerobic soil biotransformation, quantify transformation products, and elucidate its biotransformation pathways. 6:2 FTI biotransformation led to 6:2 FTOH as a key intermediate, which was subsequently biotransformed to other significant transformation products, including PFPeA [F(CF2)4COOH, 20 mol % at day 91], 5:3 acid [F(CF2)5CH2CH2COOH, 16 mol %], PFHxA [F(CF2)5COOH, 3.8 mol %], and 4:3 acid [F(CF2)4CH2CH2COOH, 3.0 mol %]. 6:2 FTI biotransformation also led to a significant level of PFHpA [F(CF2)6COOH, 16 mol % at day 91], perhaps via another putative intermediate, 6:2 FTUI [F(CF2)6CH ═ CHI], whose molecular identity and further biotransformation were not verified because of the lack of an authentic standard. Total recovery of the aforementioned per- and polyfluorocarboxylates accounted for 59 mol % of initially applied 6:2 FTI by day 91, in comparison to 56 mol % when soil was dosed with 6:2 FTOH, which did not lead to PFHpA. Thus, were 6:2 FTI to be released from its manufacture and undergo soil microbial biotransformation, it could form PFPeA, PFHpA, PFHxA, 5:3 acid, and 4:3 acid in the environment.

6:2 FTOH and 8:2 FTOH [FTOHs, F(CF2)nCH2CH2OH, n = 6, 8] are the principal polyfluorinated raw materials used to manufacture FTOH-based products, which may be released to WWTPs during their product life cycle. For the first time, anaerobic biotransformation of FTOHs and key biotransformation intermediates in WWTP digester sludge under methanogenic conditions was investigated. 6:2 FTOH was transformed to 6:2 FTCA, [F(CF2)6CH2COOH, 32-43 mol %], 6:2 FTUCA [F(CF2)5CF═CHCOOH, 1.8-8.0 mol %], and 5:3 acid [F(CF2)5CH2CH2COOH, 18-23 mol %] by day 90 and day 176 in two separate studies. 8:2 FTOH was transformed by day 181 to 8:2 FTCA (18 mol %), 8:2 FTUCA (5.1 mol %), and 7:3 acid (27 mol %). 6:2 and 8:2 FTOH anaerobic biotransformation led to low levels of perfluorohexanoic acid (PFHxA, ≤0.4 mol %) and perfluorooctanoic acid (PFOA, 0.3 mol %), respectively. 6:2 FTUCA anaerobic biotransformation led to a newly identified novel transient intermediate 3-fluoro 5:3 acid [F(CF2)5CFHCH2COOH] and 5:3 acid, but not 5:2 sFTOH [F(CF2)5CH(OH)CH3] and α-OH 5:3 acid [F(CF2)5CH2CH(OH)COOH], two precursors leading to PFPeA (perfluoropentanoic acid) and PFHxA. Thus, FTOH anaerobic biotransformation pathways operated by microbes in the environment was likely inefficient at shortening carbon chains of FTOHs to form PFCAs (perfluorinated carboxylic acids). These results imply that anaerobic biotransformation of FTOH-based products may produce polyfluorinated acids, but is not likely a major source of PFCAs detected in anaerobic environmental matrices such as anaerobic digester sludge, landfill leachate, and anaerobic sediment under methanogenic conditions.

ω-3 fatty acid desaturase is a key enzyme for the biosynthesis of ω-3 polyunsaturated fatty acids via the oxidative desaturase/elongase pathways. Here we report the identification of three ω-3 desaturases from oomycetes, Pythium aphanidermatum, Phytophthora sojae, and Phytophthora ramorum. These new ω-3 desaturases share 55 % identity at the amino acid level with the known Δ-17 desaturase of Saprolegnia diclina, and about 31 % identity with the bifunctional Δ-12/Δ-15 desaturase of Fusarium monoliforme. The three enzymes were expressed in either wild-type or codon optimized form in an engineered arachidonic acid producing strain of Yarrowia lipolytica to study their activity and substrate specificity. All three were able to convert the ω-6 arachidonic acid to the ω-3 eicosapentanoic acid, with a substrate conversion efficiency of 54-65 %. These enzymes have a broad ω-6 fatty acid substrate spectrum, including both C18 and C20 ω-6 fatty acids although they prefer the C20 substrates, and have strong Δ-17 desaturase activity but weaker Δ-15 desaturase activity. Thus, they belong to the Δ-17 desaturase class. Unlike the previously identified bifunctional Δ-12/Δ-15 desaturase from F. monoliforme, they lack Δ-12 desaturase activity. The newly identified Δ-17 desaturases could use fatty acids in both acyl-CoA and phospholipid fraction as substrates. The identification of these Δ-17 desaturases provides a set of powerful new tools for genetic engineering of microbes and plants to produce ω-3 fatty acids, such as eicosapentanoic acid and docosahexanoic acid, at high levels.

The polyfluorinated carboxylic acids 5:3 acid (C(5)F(11)CH(2)CH(2)CO(2)H) and 7:3 acid (C(7)F(15)CH(2)CH(2)CO(2)H) are major products from 6:2 FTOH (C(6)F(13)CH(2)CH(2)OH) and 8:2 FTOH (C(8)F(17)CH(2)CH(2)OH) aerobic biotransformation, respectively. The 5:3 and 7:3 acids were dosed into domestic WWTP activated sludge for 90 d to determine their biodegradability. The 7:3 acid aerobic biodegradability was low, only 1.7 mol% conversion to perfluoroheptanoic acid (PFHpA), whereas no transformation was observed previously in soil. In stark contrast, 5:3 acid aerobic biodegradability was enhanced 10 times in activated sludge compared to soil. The 5:3 acid was not activated by acyl CoEnzyme A (CoA) synthetase, a key step required for further α- or ß-oxidation. Instead, 5:3 acid was directly converted to 4:3 acid (C(4)F(9)CH(2)CH(2)CO(2)H, 14.2 mol%) and 3:3 acid (C(3)F(7)CH(2)CH(2)CO(2)H, 0.9 mol%) via "one-carbon removal pathways". The 5:3 acid biotransformation also yielded perfluoropentanoic acid (PFPeA, 5.9 mol%) and perfluorobutanoic acid (PFBA, 0.8 mol%). This is the first report to identify key biotransformation intermediates which demonstrate novel one-carbon removal pathways with sequential removal of CF(2) groups. Identified biotransformation intermediates (10.2 mol% in sum) were 5:3 Uacid, α-OH 5:3 acid, 5:2 acid, and 5:2 Uacid. The 5:2 Uacid and 5:2 acid are novel intermediates identified for the first time which confirm the proposed pathways. In the biodegradation pathways, the genesis of the one carbon removal is CO(2) elimination from α-OH 5:3 acid. These results suggest that there are enzymatic mechanisms available in the environment that can lead to 6:2 FTOH and 5:3 acid mineralization. The dehydrogenation from 5:3 acid to 5:3 Uacid was the rate-limiting enzymatic step for 5:3 acid conversion to 4:3 acid., (Copyright © 2012 Elsevier Ltd. All rights reserved.)

In vitro metabolism of 1,2,3,3,3-pentafluoropropene (PFP) was investigated in the present study. PFP was metabolized via cytochrome P450-catalyzed oxidative dehalogenation in liver microsomes and glutathione transferase (GST)-catalyzed conjugation in liver microsomes and cytosol. Two oxidation products, 2,3,3,3-tetrafluoropropionaldehyde (TPA) and 3,3,3-trifluoropyruvaldehyde (TFPA), and two GSH conjugates, S-(2,3,3,3-tetrafluoropropenyl)-GSH (TFPG) and S-(1,2,3,3,3-pentafluoropropyl)-GSH (PFPG) were identified. Enzyme kinetic parameters for the formation of TFPA, TFPG, and PFPG were obtained in male and female rat, mouse, dog, and human liver microsomes and cytosol and were confirmed using freshly isolated male rat hepatocytes. For the TFPA pathway, dog microsomes exhibited much larger K(m) values than rat, mouse, and human microsomes. Sex differences in the rates of metabolism within a given species were minor and generally were less than 2-fold. Across the species, liver microsomes were the primary subcellular fraction for GSH S-conjugation and the apparent reaction rates for the formation of TFPG were much greater than those for PFPG in liver microsomes. PFPG was unstable and had a half-life of approximately 3.9 h in a phosphate buffer (pH 7.4 and 37°C). The intrinsic clearance values for the formation of TFPA were much greater than those for the formation of GSH S-conjugates, suggesting that cytochrome P450-mediated oxidation is the primary pathway for the metabolism of PFP at relatively low PFP concentrations. Because saturation of the GST-mediated reactions was not reached at the highest possible PFP concentration, GSH S-conjugation may become a much more important pathway at higher PFP concentrations (relative to the K(m) for TFPA).

A quantitative method for fluoride determination using RP HPLC and UV detection was developed. The method is based on the reaction of fluoride with triphenylhydroxysilane in highly acidic conditions and extraction of the reaction product into n-heptane. Chromatographic conditions as well as the derivatization parameters were optimized. LOQ for fluoride ion was evaluated as 0.25 μM (4.75 ppb) and linear range of response up to 75 μM (1.42 ppm) was obtained. The method was developed as a part of a procedure designed for the determination of total organic fluorine in natural waters, using SPE on a carbon sorbent and sodium biphenyl reagent for the defluorination reaction. LOD for a model compound, perfluorooctanoic acid, calculated for the complete procedure with 2000-fold preconcentration, is 20 ppt (n=4). Initial results show feasibility of total organic fluorine determination for environmental purposes.

The gram-negative bacterium Proteus mirabilis can exist in either of two cell types, a vegetative cell characterized as a short rod and a highly elongated and hyperflagellated swarmer cell. This differentiation is triggered by growth on solid surfaces and multiple inputs are sensed by the cell to initiate the differentiation process. These include the inhibition of flagellar rotation, the accumulation of extracellular putrescine and O-antigen interactions with a surface. A key event in the differentiation process is the upregulation of FlhD(2)C(2), which activates the flagellar regulon and additional genes required for differentiation. There are a number of genes that influence FlhD(2)C(2) expression and the function of these genes, if known, will be discussed in this review. Additional genes that have been shown to regulate gene expression during swarming will also be reviewed. Although P. mirabilis represents an excellent system to study microbial differentiation, it is largely understudied relative to other systems. Therefore, this review will also discuss some of the unanswered questions that are central to understanding this process in P. mirabilis.

The first studies to explore 6-2 fluorotelomer alcohol [6-2 FTOH, F(CF(2))(6)CH(2)CH(2)OH] aerobic biodegradation are described. Biodegradation yields and metabolite concentrations were determined in mixed bacterial culture (90d) and aerobic soil (180d). 6-2 FTOH primary degradation half-life was less than 2d in both. The overall mass balance in mixed bacterial culture (day 90) was approximately 60%. At day 90, the molar yield was 6% for 6-2 FTA [F(CF(2))(6)CH(2)COOH], 23% for 6-2 FTUA [F(CF(2))(5)CFCHCOOH], 16% for 5-2 sFTOH [F(CF(2))(5)CHOHCH(3)], 6% for 5-3 acid [F(CF(2))(5)CH(2)CH(2)COOH], and 5% for PFHxA [F(CF(2))(5)COOH]. The overall mass balance in aerobic soil was approximately 67% (day 180). At day 180, the major terminal metabolites were PFPeA, [F(CF(2))(4)COOH, 30%], PFHxA (8%), PFBA [F(CF(2))(3)COOH, 2%], and 5-3 acid (15%). A new metabolite 4-3 acid [F(CF(2))(4)CH(2)CH(2)COOH] accounted for 1%, 6-2 FTOH for 3%, and 5-2 sFTOH for 7%. Based on 8-2 FTOH aerobic biodegradation pathways, PFHxA was expected in greatest yield from 6-2 FTOH degradation. However, PFPeA was observed in greatest yield in soil, suggesting a preference for alternate degradation pathways. Selected metabolites were also studied in aerobic soil. 5-3 Acid degraded to only 4-3 acid with a molar yield of 2.3%. 5-2 sFTOH degraded to PFPeA and PFHxA, and 5-2 FT Ketone [F(CF(2))(5)COCH(3)] degraded to 5-2 sFTOH, suggesting that 5-2 sFTOH is the direct precursor to PFPeA and PFHxA. Another new metabolite, 5-3 ketone aldehyde [F(CF(2))(5)COCH(2)CHO] was also identified in mixed bacterial culture. The formation of PFBA, PFPeA, and 4-3 acid indicates that multiple -CF(2)- groups in 6-2 FTOH were removed during microbial biodegradation., (Copyright 2009 Elsevier Ltd. All rights reserved.)

Fluorotelomer alcohols (FTOHs) are raw materials used in the manufacture of polymeric and surfactant products. Based on previous findings from single oral dosing in rats with radiolabeled 8-2 FTOH, glutathione (GSH) depletion and/or the presence of perfluorinated/polyfluorinated acids and aldehyde metabolites was hypothesized to account for the hepatocellular lesions observed in male rats from a 90-day subchronic oral dosing study. Further, the reported nephropathy in female rats from the subchronic experiment was hypothesized to have been initiated by a thiol metabolite produced by degradation of GSH conjugates. In the current investigation, the kinetics of 8-2 FTOH and its metabolites along with liver GSH status were evaluated in the rat following daily oral dosing with 8-2 FTOH for 45 days at 5 and 125 mg/kg/day. Liver GSH stores 1-2h after dosing were unaffected, suggesting that GSH depletion is not likely a relevant mode of action in the liver. The tissue metabolite data indicate that the liver toxicity mode of action is likely associated with elevated levels of perfluoroalkyl acids found in males, since other polyfluorinated metabolites and 8-2 FTOH were present in livers from female rats at comparable or higher levels. Detection of the N-acetyl cysteine conjugate of the unsaturated parent telomer alcohol in urine from female rats and not male rats provides some evidence to support the mechanistic basis for the observed kidney effects. Further, the increasing levels of perfluorooctanoic acid (PFOA) in plasma from female rats over the 45-day dosing phase, while unexpected, may reflect an increased net absorption of 8-2 FTOH, slow elimination of intermediates in the metabolic pathway between 8-2 FTOH and PFOA, or altered kidney clearance. The results of this study have enhanced our understanding of 8-2 FTOH kinetics and metabolism and potential modes of action in the rat, which will guide the design of future studies for FTOHs and our need to define the mechanistic basis for the observed effects.

The biodegradation pathways and metabolite yields of [3-(14)C] 8-2 fluorotelomer alcohol [8-2 FTOH, F(CF(2))(7)(14)CF(2)CH(2)CH(2)OH) in aerobic soils were investigated. Studies were conducted under closed (static) and continuous headspace air flow to assess differences in degradation rate and metabolite concentrations in soil and headspace. Aerobic degradation pathways in soils were in general similar to those in aerobic sludge and bacterial culture. (14)C mass balance was achieved in soils incubated for up to 7 months. Up to 35% (14)C dosed was irreversibly bound to soils and was only recoverable by soil combustion. The average PFOA yield was approximately 25%. Perfluorohexanoic acid (PFHxA) yield reached approximately 4%. (14)CO(2) yield was 6.8% under continuous air flow for 33 days. Three metabolites not previously identified in environmental samples were detected: 3-OH-7-3 acid [F(CF(2))(7)CHOHCH(2)COOH], 7-2 FT ketone [F(CF(2))(7)COCH(3)] and 2H-PFOA [F(CF(2))(6)CFHCOOH]. No perfluorononanoic acid (PFNA) was observed. The formation of 2H-PFOA, PFHxA, and (14)CO(2) shows that multiple -CF(2)- groups were removed from 8-2 FTOH. 7-3 Acid [F(CF(2))(7)CH(2)CH(2)COOH] reached a yield of 11% at day 7 and did not change thereafter. 7-3 Acid was incubated in aerobic soil and did not degrade to PFOA. 7-2 sFTOH [F(CF(2))(7)CH(OH)CH(3)], a transient metabolite, was incubated and degraded principally to PFOA. 7-3 Acid may be a unique metabolite from 8-2 FTOH biodegradation. The terminal ratio of PFOA to 7-3 acid ranged between 1.8-2.5 in soils and 0.6-3.2 in activated sludge, sediment, and mixed bacterial culture. This ratio may be useful in evaluating environmental samples to distinguish the potential contribution of 8-2 FTOH biodegradation to PFOA observed versus PFOA originating from other sources.

Biodegradation of fluorinated polymers is of interest to assess them as a potential source of perfluorocarboxylates (PFCAs) in the environment. A fluoroacrylate polymer product test substance was studied in four aerobic soils over two years to assess whether the fluorotelomer alcohol (FTOH) side chains covalently bonded to the polymer backbone may be transformed to form PFCAs. The test substance itself was not directly measured; instead, nine analytes were determined to evaluate biodegradation. Terminal biotransformation products measured included perfluorooctanoate (PFO), perfluorononanoate (PFN), perfluorodecanoate (PFD), perfluoroundecanoate (PFU), and pentadecafluorodecanoate (7-3 acid). The molar concentration of 8-2 fluorotelomer alcohol (8-2 FTOH) in the test substance, fluoroacrylate polymer and residual unreacted raw materials and impurities ("residuals") were compared with the molar concentrations of the terminal biotransformation products for mass balance and kinetic assessments. Over the two year time frame of the experimental study, the fluoroacrylate polymer showed a slight extent of potential biodegradation under the experimental conditions of the study. A biodegradation half-life of 1200-1700 years was calculated for the fluoroacrylate polymer based on the rate of formation of PFO in aerobic soils. When the degradation rates of the fluoroacrylate polymer and residuals were applied to estimated total historic fluoroacrylate polymer production, use and disposal, the biodegradation of fluoroacrylate polymer and residuals is calculated to contribute less than 5 tonnes of PFO per year globally to PFCAs present in the environment.

The detection of perfluorinated organic compounds in the environment has generated interest in their biological fate. 8-2 Fluorotelomer alcohol (8-2 FTOH, C(7)F(15)CF(2)CH(2)CH(2)OH), a raw material used in the manufacture of fluorotelomer-based products, has been identified in the environment and has been implicated as a potential source for perfluorooctanoic acid (PFOA) in the environment. In this study, the in vitro metabolism of [3-(14)C] 8-2 FTOH and selected acid metabolites by rat, mouse, trout, and human hepatocytes and by rat, mouse, and human liver microsomes and cytosol were investigated. Clearance rates of 8-2 FTOH in hepatocytes indicated rat > mouse > human >/= trout. A number of metabolites not previously reported were identified, adding further understanding to the pathway for 8-2 FTOH metabolism. Neither perfluorooctanoate nor perfluorononanoate was detected from incubations with human microsomes. To further elucidate the steps in the metabolic pathway, hepatocytes were incubated with 8-2 fluorotelomer acid, 8-2 fluorotelomer unsaturated acid, 7-3 acid, 7-3 unsaturated acid, and 7-2 secondary fluorotelomer alcohol. Shorter chain perfluorinated acids were only observed in hepatocyte and microsome incubations of the 8-2 acids but not from the 7-3 acids. Overall, the results indicate that 8-2 FTOH is extensively metabolized in rats and mice and to a lesser extent in humans and trout. Metabolism of 8-2 FTOH to perfluorinated acids was extremely small and likely mediated by enzymes in the microsomal fraction. These results suggest that human exposure to 8-2 FTOH is not expected to be a significant source of PFOA or any other perfluorocarboxylic acids.

Considering recent reports on widespread occurrence and concerns about perfluoroalkyl substances (PFAS) in environmental and biological systems, analysis of these compounds have gained much attention in recent years. Majority of analyte-specific methods are based on a LC/MS/MS or a GC/MS detection, however many environmental or biological studies would benefit from a total organic fluorine (TOF) determination. Presented work was aimed at developing a method for TOF determination. TOF is determined as an amount of inorganic fluoride obtained after defluorination reaction conducted off-line using sodium biphenyl reagent directly on the sorbent without elution of retained analytes. Recovered fluoride was analyzed using flow-injection system with either fluorimetric or potentiometric detection. The TOF method was tested using perfluorocarboxylic acids (PFCA), including perfluorooctanoic acid (PFOA), as model compounds. Considering low concentrations of PFAS in natural samples, solid-phase extraction as a preconcentration procedure was evaluated. Several carbon-based sorbents were tested, namely multi-wall carbon nanotubes, carbon nanofibres and activated carbon. Good sorption of all analytes was achieved and defluorination reaction was possible to carry out directly on a sorbent bed. Recoveries obtained for PFCAs, adsorbed on an activated carbon sorbent, and measured as TOF, were 99.5+/-1.7, 110+/-9.4, 95+/-26, 120+/-32, 110+/-12 for C4, C6, C8, C10 and C12-PFCA, respectively. Two flow systems that would enable the defluorination reaction and fluoride determination in a single system were designed and tested.

LC with MS detection was optimized for sensitive and selective analysis of main classes of natural dyes used in ancient times for dyeing textiles -- red anthraquinoids, yellow flavonoids, and known degradation products of flavonols -- hydroxybenzoic acids. Fragmentation patterns of both negative and positive molecular ions for the above mentioned compounds were investigated. Three acquisition modes of MS analysis: scanning, SIM, and multiple reaction monitoring (MRM) in both positive and negative ion modes were optimized and compared with each other and with the UV-Vis diode-array detection. Even though in the applied chromatographic system formic acid was used in the mobile phase, SIM in the negative ion mode was the most selective and sensitive detection for all the investigated compounds when both mixtures of standards and analysis of extracts from archeological samples were concerned, with one exception -- alizarin, for which MS detection in positive ion mode was more sensitive. Detection limits obtained with MS detection for all investigated compounds except quinizarin were lower than the ones obtained with the diode-array UV-Vis detection, making MS detection the most suitable tool for the analysis of natural dyes and their degradation products in extracts from archeological samples.

Reversed-phase HPLC determination of the herbicide MCPA and its products of radiolytic degradation has been optimized. The radiolytic degradation was carried out using gamma-irradiation and was optimized in terms of irradiation dose and pH of irradiated MCPA solution. Decomposition of 100 ppm MCPA in pure solutions required irradiation with a 3 kGy dose. The main products of irradiation in the dose range up to 10-kGy were various phenolic compounds and carboxylic acids. The developed method was applied for treatment of industrial waste from production of MCPA. The 10-kGy dose was needed for decomposition of 500 ppm of MCPA in the industrial waste samples; however, the presence of stoichiometric amount of hydrogen peroxide in the irradiated waste allowed a 50% reduction of the gamma-irradiation dose. Despite complete decomposition of MCPA in the industrial waste, in order to reduce the toxicity of irradiated waste, measured by the Microtox bioluminescence test, higher than a 10 kGy irradiation dose was needed.

Perfluorinated carboxylic acids (PFCAs) belong to anthropogenic fluoroorganic compounds that have been detected in the natural environment and living organisms including humans. A capillary zone electrophoretic method with indirect UV detection using 2,4-dinitrobenzoic acid (2,4-DNBA) as a chromophore probe has been developed for analysis of PFCAs (C6-C12) in water. Optimal analyte resolution and detection sensitivity was obtained with 50 mM Tris solution of pH 9.0 and 50% methanol as a background electrolyte (BGE). The baseline separation of C6-C12 PFCAs was obtained within 20 min with detection limits in the range from 0.6 to 2.4 ppm.

The methodology for the determination of perfluorooctanoate (C(7)F(15)COO-, PFO), fluorotelomer alcohols (FTOHs: 6-2, 8-2, and 10-2), perfluorooctyl iodide (PFOI), and 8-2-8 fluorotelomer alcohol ester in complex fluorotelomer-based commercial products has been demonstrated and validated. Sample preparation procedures allowing determination of residual levels of these compounds were developed. The analytes were detected either by LC/MS/MS (PFO), LC/MS (FTOHs), or GC/MS (PFOI, 8-2-8 ester). The methods were validated by investigating the recoveries of analytes spiked at multiple levels to authentic sample matrices. The recoveries generally were between 70 and 130%. The limits of detection were in sub-microg/g range and the limits of quantitation were in the mug/g range. The methods were applied to fluorotelomer-based raw materials and fluorotelomer-based surfactants and polymeric products and represent methods useful for the determination of higher carbon chain length homologs as well.

Fluorotelomer alcohols (FTOHs) are important polyfluorinated raw materials that belong to the general category of perfluoroalkyl substances (PFAS). PFAS, including perfluoroalkyl carboxylates (PFCAs) and perfluoroalkyl sulfonates, have recently attracted considerable attention because they are persistent and found globally in the environment. FTOHs are precursors that may degrade in the environment to PFCAs. The development of analytical methods for determination FTOHs in environmental samples is necessary to determine the environmental presence of FTOHs. This work presents the development and validation of a liquid chromatography/tandem mass spectrometry (LC/MS/MS) method for the determination of FTOHs (6-2, 8-2, 10-2) in aqueous samples. Chromatographic conditions were optimized in order to obtain focused FTOH chromatographic peaks. The mobile phase and mass spectrometric conditions were optimized to enable formation of deprotonated FTOH molecules in the negative ion electrospray mode. Two extraction methods were investigated using acetonitrile and methyl tert-butyl ether (MTBE). These methods were validated for a range of environmental water samples fortified with FTOHs at three different levels. Both extraction methods resulted in recoveries from 70 to 120%. Detection limits of FTOHs were estimated to be approximately 0.09 ng/mL for LC/MS/MS detection. An LC/MS method was also developed for FTOH determination with an estimated 1.2 ng/mL limit of detection. Various sample storage scenarios were investigated. It was determined that the aqueous samples of FTOHs are best preserved by storing them frozen in sealed vials with aluminum foil lined septa., (Copyright 2006 John Wiley & Sons, Ltd.)

There is increasing scientific interest to understand the environmental fate of fluorotelomer alcohols (FTOHs) and fluorotelomer-based products which may break down to FTOHs. Both are expected to enter aqueous waste streams, which would be processed in a wastewater treatment plant and therein subject to microbial biodegradation. We investigated the biodegradation of 3-14C, 1H,1H,2H,2H-perfluorodecanol [CF3(CF2)6(14)CF2CH2CH2OH, 14C-8-2 FTOH] in mixed bacterial culture and activated sludge. 14CO2 and 14C-organic volatiles in the headspace of the sealed bottles and bottles with continuous air flow were analyzed up to 4 months. After sample extraction with acetonitrile, 14C-labeled biotransformation products (metabolites) were quantified by LC/ARC (on-line liquid chromatography/ accurate radioisotope counting) and identified by quadrupole time-of-flight (Q-TOF) mass spectrometry and GC/MSD (mass selective detector). Three metabolites not yet reported in the literature have been identified as CF3(CF2)6(14)CHOHCH3 (7-2 sFTOH), CF3(CF2)6(14)CH=CHCOOH (7-3 unsaturated acid or 7-3 u acid), and CF3(CF2)6(14)CH=CHCONH2 (7-3 u amide) along with five previously reported metabolites [CF3(CF2)6(14)CF2CH2CHO (8-2 FTAL), CF3(CF2)6 (14)CF2CH2COOH (8-2 acid), CF3(CF2)6(14)CF=CHCOOH (8-2 u acid), CF3(CF2)6(14)CH2CH2COOH (7-3 acid), and CF3(CF2)6(14)COOH (PFOA)]. No CF3(CF2)6(14)CF2COOH (14C-PFNA) was observed, indicating that alpha-oxidation does not take place. It was found that strong adsorption to the activated sludge and subsequent transformation, even under continuous air flow, greatly reduced partitioning of 8-2 FTOH or any transformation products to air. CF3(CF2)4COOH (PFHA; perfluorohexanoic acid) was observed and increased in mixed bacterial culture over 28 days and accounted for about 1% of the initial 14C-8-2 FTOH concentration from day 28 to day 90. 14CO2 accounted for 1% of initial 14C in activated sludge with continuous air flow at day 1 and increased over time. In closed bottles, 14CO2 in the headspace of activated sludge medium increased to 12% of the available 14C over 135 days with periodic addition of ethanol, as compared to 3% when no additional ethanol was added. These results show that replenishment of organic carbon enhanced microbial mineralization of multiple--CF2--groups in the fluorocarbon chain of 14C-8-2 FTOH. At day 90 the net increase of fluoride ion in the mixed bacterial culture was 93 microg L(-1), equivalent to 12% of total mineralization (destruction) of the 14C-8-2 FTOH. These results demonstrate that perfluorinated carbon bonds of 14C-8-2 FTOH are defluorinated and mineralized by microorganisms under conditions which may occur in a wastewater treatment plant, forming shorter fluorinated carbon metabolites.

A selective and sensitive method for analysis of perfluorooctanoic acid (PFOA) in human serum and plasma, utilizing liquid chromatography tandem mass spectrometry (LC-MS/MS), has been developed and thoroughly validated to satisfy strict FDA guidelines for bioanalytical methods. A simple, automated sample preparation procedure, involving extraction of the target analyte with acetonitrile on protein precipitation media in a 96-well plate format was developed, allowing efficient handling of large numbers of samples. The proposed method uses the calibration standards prepared in a surrogate matrix (rabbit serum or plasma) and (13)C-labeled PFOA as the internal standard to account for matrix effects, instrument drift, and extraction efficiency. Human serum and plasma could not be used for matrix matching of calibration standards as endogenous levels of PFOA observed in the control human serum and plasma significantly exceeded the targeted lower limit of quantitation (LLOQ) of the method. Precision and accuracy of the method were demonstrated by analysis of rabbit serum and plasma control samples fortified at 0.5, 5, and 40 ng/mL PFOA and human serum and plasma fortified at 1.0, 5.0, 40 ng/mL PFOA. The LLOQ of 0.5 ng/mL PFOA was experimentally demonstrated for rabbit and human serum and plasma. Within-day precision and accuracy, short-term stability, freeze-thaw stability, equivalence of response between PFOA and APFO (the ammonium salt of PFOA), and dilution of concentrated samples were also investigated. The results of the validation experiments comply with the precision and accuracy limits defined by the FDA guidance document: "Guidance for Industry, Bioanalytical Method Validation", May 2001.

Methods were developed to quantify the amount of perfluorooctanoic acid (PFOA) extracted from textile and carpet samples through contact with water, methanol, and sweat and saliva simulants using LC/MS/MS. The limit of quantitation (LOQ) for samples extracted in water and sweat simulant is 1 ppb (ng PFOA (g sample)(-1)) while the limits of quantitation for samples extracted in saliva simulant and methanol were 3 ppb and 2.5 ppb, respectively. Method validation results are provided for a polyester control textile sample that was extracted in water on two different days by different analysts, which gave an overall recovery of 103% and standard deviation of 5.3% for 30 analyses. However, for routine application of these methods to a large number of sample sets differing in chemical and physical compositions, a complete validation for each sample type is not practical or possible since control samples for fortifications are often not available. Instead, suitable analytical methods and acceptance criteria are described which ensure accurate PFOA quantitation in each of the solvent extract types. During routine use of these methods, post-extraction spike recoveries for the different sample types and solvents are 100 +/- 15% using a dual isotopically labeled (13)C-PFOA internal standard to correct for matrix effects. A comparison of extraction solvent versus time using a wrist action shaker for textile and carpet samples demonstrates that the total extractable amount of PFOA is similar for each of the solvent types. However, as expected the rate of extraction in water and simulants is significantly less than that of methanol. Finally, a comparison of 2 h and 24 h wrist action shaker extractions with a 1.5 h pressurized fluid extraction (PFE) in methanol reveals that the 24 h wrist action shaker yields the highest results. The 2 h wrist action shaker results are similar to those of the 1.5 h PFE extraction.

A capillary electrophoretic method with UV detection for separation and quantitation of perfluorocarboxylic acids (PFCAs) from C6-PFCA to C12-PFCA has been developed. The optimization of measurement conditions included the choice of the most appropriate type and concentration of buffer in the background electrolyte (BGE), as well as the type and the content of an organic modifier. The optimal separation of investigated PFCAs was achieved with 50 mM phosphate buffer and 40% isopropanol in the BGE using direct UV detection. The optimum wavelength for direct UV detection was optimized at 190 nm. For indirect detection, several chromophores were studied. Five mM 3,5-Dinitrobenzoic acid (3,5-DNBA) in 20 mM phosphate buffer BGE and indirect UV detection at 280 nm gave the optimal detection and separation performance for the investigated PFCAs. The possibility of on-line preconcentration of solutes by stacking has been examined for indirect detection. The detection limits (LODs) determined for direct UV detection ranged from 2 microg/mL for C6-PFCA to 33 microg/mL for C12-PFCA. The LODs obtained for indirect UV detection were comparable to those obtained for direct UV detection.

This study investigated the biodegradation potential of 3-(14)C,1H,1H,2H,2H-perfluorodecanol [CF3(CF2)6(14)CF2CH2CH2OH, 14C-labeled 8-2 telomer B alcohol or 14C-labeled 8-2 TBA] by diluted activated sludge from a domestic wastewater treatment plant under aerobic conditions. After sample extraction with acetonitrile, biotransformation products were separated and quantified by LC/ARC (on-line liquid chromatography/accurate radioisotope counting) with a limit of quantification about 0.5% of the 14C counts applied to the test systems. Identification of biotransformation products was performed by quadrupole time-of-flight mass spectrometry. Three transformation products have been identified: CF3(CF2)6(14)CF2CH2COOH (8-2 saturated acid); CF3(CF2)6(14)CF=CHCOOH (8-2 unsaturated acid); and CF3(CF2)6(14)COOH (perfluorooctanoic acid, PFOA), representing 27, 6.0, and 2.1% of the initial 14C mass (14C counts applied) after 28 days, respectively. A transformation product, not yet reported in the literature, has also been observed and tentatively identified as CF3(CF2)6(14)CH2CH2COOH (2H,2H,3H,3H-perfluorodecanoic acid); it accounted for 2.3% of the mass balance after 28 days. The 2H,2H,3H,3H-perfluorodecanoic acid is likely a substrate for beta-oxidation, which represents one of the possible pathways for 8-2 telomer B alcohol degradation. The 8-2 saturated acid and 8-2 unsaturated acid cannot be directly used as substrates for beta-oxidation due to the proton deficiency in their beta-carbon (C3 carbon) and their further catabolism may be catalyzed by some other still unknown mechanisms. The 2H,2H,3H,3H-perfluorodecanoic acid may originate either from the major transformation product CF3(CF2)6(14)CF2CH2COOH or from other unidentified transformation products via multiple steps. Approximately 57% of the starting material remained unchanged after 28 days, likely due to its strong adsorption to the PTFE (poly(tetrafluoroethylene)) septa of the test vessels. No CF3(CF2)6(14)CF2COOH (perfluorononanoic acid) was observed, indicating that alpha-oxidation of CF3(CF2)6(14)CF2CH2COOH did not occur under the study conditions. Several 14C-labeled transformation products that have not yet been identified (each less than 1% of the mass balance) were also observed and together accounted for 7% of the total 14C mass balance after 28 days. It is not clear whether these unidentified transformation products were resulting from further metabolism of 8-2 saturated acid or 8-2 unsaturated acid. The results suggest that perfluorinated acid metabolites such as perfluorooctanoic acid account for only a very small portion of the transformation products observed. Also, the observed volatility and bioavailability of 14C-labeled 8-2 TBA for microbial degradation was markedly decreased as a result of the presence of a strongly adsorbing matrix such as PTFE in the experimental systems. It is apparent that the biological fate of 8-2 telomer B alcohol is determined by multiple degradation pathways, with neither beta-oxidation nor any other enzyme-catalyzed reactions as a single dominant (principal) mechanism under the study conditions.

Liquid chromatography (LC) combined with ultraviolet-visible (UV-Vis) and mass spectrometric (MS) detection was utilized to study the chemical components present in extracts of natural dyes originating from fiber samples obtained from Coptic textiles from Early Christian Art Collection of National Museum in Warsaw. Chromatographic retention, ionization, UV-Vis and mass spectra of twenty selected dye compounds of flavanoid-, anthraquinone- and indigo-types were studied. Most of the investigated compounds could be ionized by positive and negative ion electrospray ionization. Difficulties with the ionization by electrospray were experienced for indigotin and brominated indigotins, but these were ionized by atmospheric pressure chemical ionization. Mass spectrometric detection, utilizing different scanning modes of a triple quadrupole mass spectrometer, combined with the UV-Vis detection was demonstrated to be a powerful approach to detection and identification of dyes in the extracts of archeological textiles. Using this approach the following compounds were identified in the extracts of Coptic textiles: luteolin, apigenin, rhamnetin, kaempferol, alizarin, purpurin, xanthopurpurin, monochloroalizarin, indirubin, and so the type of dye that was utilized to dye the textiles could be identified. Detection capabilities for several dye-type analytes were compared for the UV-Vis and mass spectrometric detection. The signal-to-noise ratios obtained for luteolin, apigenin, and rhamnetin were higher for the MS detection for most of the examined sample extracts. Purpurin, alizarin, and indirubin showed similar signal-to-noise ratios for UV-Vis and mass spectrometric detection.

An improved method for coupling capillary electrophoresis (CE) with condensation nucleation light scattering detection (CNLSD) is described. The method employs an electrospray aerosol source followed by aerosol particle neutralization within a weak plasma established by a polonium-210 alpha emitter. Sensitive, universal detection of underivatized proteins, peptides, and amino acids is demonstrated. The system performance was significantly affected by the operational mode of the electrospray source, characterized by the physical appearance of the droplet at the end of the electrospray capillary. With a so-called pulsating mode, relatively low backgrounds were obtained with 10 mM ammonium acetate or 10 mM ammonium acetate/10 mM ammonia CE buffers using one diffusion screen, allowing detection of proteins at single microgram per milliliter levels and amino acids and peptides at submicrogram per milliliter levels. Linearity of response, expressed as peak height or peak area; mass limits of detection (LODs) for proteins, peptides, and amino acids at the picogram level, corresponding to femtomole levels of peptides and amino acids or subfemtomole levels of proteins; and better detectability compared to UV absorbance at 214 and 200 nm were demonstrated. The separation efficiencies obtained with the CE-electrospray-CNLSD system are much higher than those obtained for a previously reported pneumatic nebulizer-based system and in the range of approximately 20,000-160,000 plates/m using the pulsating electrospray mode. With careful adjustment of the electrospray voltage, a different operating mode, called the silver bullet mode, could be established for which higher signals, lower background and background noise, and higher separation efficiencies were observed compared to the pulsating mode. The lower background levels observed with the silver bullet mode eliminated the need for the use of a diffusion screen for background control with the buffer employed. With the silver bullet mode without a diffusion screen, linear response and LODs at the 15 ng/mL level, corresponding to subpicogram or 1-2 fmol levels of underivatized peptides and amino acids, and plate numbers in the range of 65,000-220,000 plates/m were estimated.

We describe two means for interfacing condensation nucleation light scattering detection to capillary electrophoresis (CE). With the first method, a fused-silica capillary was used for the separation and the CE was grounded through a Nafion membrane that also connected the system to a microconcentric pneumatic nebulizer. Limits of detection (LODs) for underivatized amino acids were at the low microgram per milliliter level, and separation efficiencies were ∼9 times lower than the optimum predicted for these species based on the injection plug width and axial dispersion by diffusion. LODs were limited by background nonvolatiles resulting from dissolution of fused silica at the high pHs used for the separations. An alternate system employed PEEK capillaries which acted as the separation capillary and also as the inner nebulizer capillary. In this case, the exit end of the capillary was coated with conductive paint which extended to the tip of the nebulizer, was in contact with the CE buffer, and was grounded to complete the CE circuit. Response was nonlinear and the separation efficiency of this system was somewhat lower than that for the Nafion membrane system. Response as peak heights for all of the amino acids and peptides studied was nearly identical on a mass basis. With this system, much lower background signals were obtained, and as a result, LODs for underivatized amino acids and peptides were below the 1 μg/mL level, corresponding to less than 10 pg or less than 100 fmol injected. Both systems were fairly simple, effective means to generate aerosols with the low flows of CE and should be applicable to interfacing of other aerosol-based detectors with CE.