Metabolism, tissue disposition, and excretion of 1,2-bis(2,4,6-tribromophenoxy)ethane (BTBPE) in male Sprague-Dawley rats

A single oral dose of [14C] 1,2-bis(2,4,6-tribromophenoxy)ethane (BTBPE) was administered to conventional and bile-duct cannulated male Sprague-Dawley rats. Tissue disposition, excretion and metabolism was determined. BTBPE is a low-volume brominated flame retardant used in resins or plastics, and toxicity data in peer-reviewed journals is extremely limited. BTBPE was fairly insoluble in lipophilic solutions, which made dose preparation difficult. The great majority of 14C (>94%) was excreted in the feces of both groups of rats at 72 h, and tissue retention was minimal. Lipophilic tissues contained the highest concentrations of BTBPE, e.g. thymus, adipose tissue, adrenals, lung, and skin. Metabolites were excreted in the urine, bile and feces, but at a very low level. Fecal metabolites were characterized as monohydroxylated, monohydroxylated with debromination, dihydroxylated/debrominated on a single aromatic ring, monohydroxylated on each aromatic ring with accompanying debromination, and cleavage on either side of the ether linkage to yield tribromophenol and tribromophenoxyethanol. Despite a limited quantity of stable metabolites extractable in the feces, non-extractable 14C levels were relatively high (39% of the 0-24 h fecal 14C), which suggested that BTBPE could be metabolically activated in the rat and covalently bound to fecal proteins and/or lipids. It was concluded that limited absorption and metabolism of BTBPE would occur by ingestion in mammals.     

The metabolic fate of N-isopropyl-N-phenyloxamic acid in the rat and the milk goat.

Rats excreted the 14C from a single oral dose of N-isopropyl-N-[14C]phenyloxamic acid [I, a soil metabolite from 2-chloro-N-isopropylacetanilide (propachlor)] in approximately equal quantities in the urine (49.2%) and feces (48.2%). A milking goat given daily oral doses of [14C]-I (1 mg of I three times daily) excreted more 14C in the feces (56.6%) than it excreted in the urine. From both species, I accounted for 97 to 100% of the urinary 14C, and all of the 14C that was extractable from the feces (73 to 75% of the 14C in feces was extractable with methanol). Goat milk samples collected 16 hr after the last dose contained no detectable 14C. Tissue residues of 14C were determined.     

Fate and residues of 14C‐chloramphenicol in laying chickens 1

Abstract

Studies were conducted to determine the metabolic fate of chloramphenicol (CAP) in White Leghorn using the ^l4C‐labelled compound. In one experiment birds were administered orally via intra‐crop, a single dose of 100 mg (equivalent to 66 mg kg^‐1 body weight) of CAP containing 14 μCi ¹⁴C‐CAP, and its absorption, elimination and distribution in plasma were recorded. Orally dosed ¹⁴C‐compound was rapidly absorbed, efficiently distributed in plasma and eliminated in excreta (>70% in 5 hr). After 5 h, CAP equivalent residues in tissues were lower than 15 μg g^‐1 for this treatment. In a second experiment birds were given intra‐crop dose of either 0.5 or 5 mg of CAP (each dose contained 2.5 μCi ¹⁴C‐CAP) daily for five consecutive days followed by a seven day withdrawal period and elimination of ¹⁴C in excreta and eggs was monitored. More than 95% of the administered ¹⁴C was eliminated within the first 24 h after dosing. Radiocarbon (¹⁴C) was deposited preferentially in yolks compared to albumen or other tissues. Residues declined when feeding was stopped. Various metabolites were isolated and identified by a combination of TLC, LC, and LC‐MS. The main metabolic route of CAP in laying hens appears to be the glucuronidation. Cleavage of the dichloroacetate moiety was only a minor route.     

Rat urinary metabolites from O,O-diethyl-O-(3,5,6-trichloro-2-pyridyl) phosphorothioate.

Rats metabolized single oral doses of O,O-diethyl-O(3,5,6-trichloro-2-pyridyl-2,6-14C) phosphorothioate to at least six radiolabeled urinary metabolites. The urine contained about 90 percent of the dose. Three of these metabolites were identified as the glucuronide of 3,5,6-trichloro-2-pyridinol (80% the urinary 14C), a glycoside of 3,5,6-trichloro-2-pyridinol (4%), and 3,5,6-trichloro-2-pyridinol (12%).     

Studies on the origin of the methylsulfonyl-containing metabolites from propachlor.

Metabolites in which the chlorine from propachlor has been replaced by a cysteine group or a methylsulfonyl group [-S(O2) CH3] are present in the urine of rats dosed orally with propachlor. In the present study, urine from rats given single oral doses of 35S-labeled cysteine conjugate of propachlor contained metabolites having the methylsulfonyl groups labeled with 35S. No metabolites containing 14C-labeled methylsulfonyl groups were isolated from urine of rats given single oral doses of the cysteine conjugate of propachlor in which the cysteine group was uniformly labeled with 14C. These findings show that the cysteine conjugate of propachlor is the source of sulfur in the methylsulfonyl-containing metabolites. Therefore, we suggest that a C-S lyase present in the animal cleaves the cysteine conjugate of propachlor and thus allows further metabolism of the sulfur to a methylsulfonyl moiety.     

Metabolism of crufomate [(4-tert-butyl-2-chlorophenyl methyl methylphosphormidate)] by the rat.

Fifteen metabolites of crufomate (4-tert-butyl-2-chlorophenyl methyl methylphosphoramidate, I) were identified in the excreta from rats given single oral doses of I. Compound I was not detected in either the urine or the feces. The metabolic reactions observed were N-and O-demethylation, oxidations of the t-butyl moiety, replacement of the H-N-CH3 with an OH moiety, hydrolysis of the phosphoramidate moiety to yield the phenol, conjugation with glucuronic acid, and combinations of these reactions. No ring dehalogenation or ring substitution was observed.     

Studies on the origin of the methylsulfonyl‐containing metabolites from propachlor

Abstract

Metabolites in which the chlorine from propachlor has been replaced by a cysteine group or a methylsulfonyl group [‐S(O₂) CH₃] are present in the urine of rats dosed orally with propachlor. In the present study, urine from rats given single oral doses of ³⁵S‐labeled cysteine conjugate of propachlor contained metabolites having the methylsulfonyl groups labeled with S. No metabolites containing ¹⁴C‐labeled methylsulfonyl groups were isolated from urine of rats given single oral doses of the cysteine conjugate of propachlor in which the cysteine group was uniformly labeled with ¹⁴C. These findings show that the cysteine conjugate of propachlor is the source of sulfur in the methylsulfonyl‐containing metabolites. Therefore, we suggest that a C‐S lyase present in the animal cleaves the cysteine conjugate of propachlor and thus allows further metabolism of the sulfur to a methylsulfonyl moiety.     

14C-dimethoate residues in olive oil during oil processing

An olive tree was treated twice in the field with 14C-dimethoate (237.7 muCi, 2.4 g) and 14C residues were determined in the olive fruits at harvest. The fruits were crushed and pressed to extract the crude oil, then refined by neutralization, bleaching and deodorization. The crude oil contained 14.1% of the total 14C in the olive fruits. Neutralization resulted in a reduction of 14C by about 50% of the total 14C residues in oil. Bleaching and deodorization processes further reduced the 14C residues and the refined oil contained 31.6% (which corresponds to 4.4% of 14C residues of the total 14C in olive fruits) of the total 14C in the crude oil. Industrially extracted crude oil was fortified with 14C-dimethoate at 1.8 mg kg-1 (0.02 muCi) level and subjected to the same refining process. A sharp decrease in the amount of 14C was observed by neutralization and the amount of 14C remaining in the refined oil was about 7.3% of the total 14C in the crude fortified oil. The data suggest that the 14C residues in the aged and the fortified oil amples were not of the same nature. The terminal 14C residue in the refined oil obtained from the field experiment did not contain dimethoate and/or its oxon.     

Dendroremediation of trinitrotoluene (TNT) Part 2: Fate of radio-labelled TNT in trees

BACKGROUND, AIM AND SCOPE: Problems of long-term existence of the environmental contaminant 2,4,6-trinitrotoluene (TNT) and necessities for the use of trees ('dendroremediation') in sustainable phytoremediation strategies for TNT are described in the first part of this paper. Aims of the second part are estimation of [14C]-TNT uptake, localisation of TNT-derived radioactivity in mature tree tissues, and the determination of the degree of TNT-degradation during dendroremediation processes. METHODS: Four-year-old trees of hybrid willow (Salix spec., clone EW-20) and of Norway spruce (Picea abies) were cultivated in sand or ammunition plant soil (AP-soil) in wick supplied growth vessels. Trees were exposed to a single pulse application with water solved [U-14C]-TNT reaching a calculated initial concentration of 5.2 mg TNT per kg dry soil. Two months after application overall radioactivity and extractability of 14C were determined in sand/soil, roots, stem-wood, stem-bark, branches, leaves, needles, and Picea May sprouts. Root extracts were analysed by radio TLC. RESULTS: 60 days after [14C]-TNT application, recovered 14C is accumulated in roots (70% for sand variants, 34% for AP-soil variant). 15-28% of 14C remained in sand and 61% in AP-soil. 3.3 to 14.4% of 14C were located in aboveground tree portions. Above-ground distribution of 14C differed considerably between the angiosperm Salix and the gymnosperm Picea. In Salix, nearly half of above-ground-14C was detected in bark-free wood, whereas in Picea older needles contained most of the above-ground-14C (54-69%). TNT was readily transformed in tree tissue. Approximately 80% of 14C was non-extractably bound in roots, stems, wood, and leaves or needles. Only quantitatively less important stem-bark of Salix and Picea and May shoots of Picea showed higher extraction yields (up to 56%). DISCUSSION: Pulse application of [14C]-TNT provided evidence for the first time that after TNT-exposure, in tree root extracts, no TNT and none of the known metabolites, mono-amino-dinitrotoluenes (ADNT), diaminonitrotoluenes (DANT), trinitrobenzene (TNB) and no dinitrotoluenes (DNTs) were present. Extractable portions of 14C were small and contained at least three unknown metabolites (or groups) for Salix. In Picea, four extractable metabolites (or groups) were detected, where only one metabolite (or group) seemed to be identical for Salix and Picea. All unknown extractables were of a very polar nature. CONCLUSIONS: Results of complete TNT-transformation in trees explain some of our previous findings with 'cold analytics', where no TNT and no ADNT-metabolites could be found in tissues of TNT-exposed Salix and Populus clones. It is concluded that 'cold' tissue analysis of tree organs is not suited for quantitative success control of phytoremediation in situ. RECOMMENDATIONS AND OUTLOOK: Both short rotation Salicaceae trees and conifer forests possess a dendroremediation potential for TNT polluted soils. The degradation capacity and the large biomass of adult forest trees with their woody compartments of roots and stems may be utilized for detoxification of soil xenobiotics.     

Synthesis of [13C]- and [14C]-labeled phenolic humus and lignin monomers

Natural phenolic monomers are ubiquitous in the environment and are involved in the stabilization of atmospheric carbon and the transformation of xenobiotics. Investigations on the stabilization of phenolic carbons and their environmental fate are hampered by the unavailability of commercial [13C]- and [14C]-labeled phenols. Here we report the complete chemical synthesis of the lignin and humus structural monomers p-coumaric, ferulic, and caffeic acids, p-hydroxybenzaldehyde, protocatechualdehyde, vanillin, catechol, and guaiacol, uniformly [13C]- or [14C]-labeled in the aromatic ring, starting from commercially available [U-ring-13C]- or [U-ring-14C]-labeled phenol. The synthesis of these compounds involved selective ortho-hydroxylation of the aromatic ring, Friedel-Crafts alkylation, and Knoevenagel condensation. [U-ring-13C]- or [U-ring-14C]-p-coumaric acid was synthesized via p-hydroxybenzaldehyde with a 75% yield with respect to phenol. Synthesis of [U-ring-13C]- or [U-ring-14C]-ferulic acid, consisting of six single steps via guaiacol and vanillin, had an overall yield of up to 45%. Uniformly ring-labeled caffeic acid was synthesized either via catechol and protocatechualdehyde in five single steps, yielding [U-ring-14C]-caffeic acid with a 37% yield, or via guaiacol, vanillin, and ferulic acid in seven steps, yielding [U-ring-13C]-caffeic acid with an 18% yield. Ferulic acid, [14C]-labeled at beta-C of the propenoic side chain, was synthesized from [2-14C]-malonic acid under Knoevenagel conditions with a 67% yield with respect to malonic acid. Demethylation of the [beta-14C]-ferulic acid with BBr3 in CH3CN resulted in [beta-14C]-caffeic acid with a 62% yield. All [U-ring-13C]-labeled phenolic products were analyzed by 13C nuclear magnetic resonance (13C-NMR) spectroscopy and gas chromatography-mass spectrometry (GC-MS).     

Rat urinary metabolites from O,O‐diethyl‐O‐(3, 5, 6‐trichloro‐2‐pyridyl) phosphorothioate

Abstract

Rats metabolized single oral doses of O,O‐diethy1–0‐(3,5,6‐trichloro‐2‐pyridyl‐2,6‐¹⁴C) phosphorothioate to at least six radiolabeled urinary metabolites. The urine contained about 90 percent of the dose. Three of these metabolites were identified as the glucuronide of 3,5,6‐trichloro‐2‐pyridinol (80% the urinary ^l4C), a glycoside of 3,5,6‐trichloro‐2‐pyridinol (4%), and 3,5,6‐trichloro‐2‐pyridinol (12%).     

14C-carbaryl residues in hazelnut

A hazelnut ocak (shrub growing form) in the field in Black Sea region of Turkey was treated with commercial carbaryl insecticide spiked with 14C-carbaryl. Three months later, the harvested hazelnuts were separated into husk, shell, and kernel components, then homogenized and analyzed. The total and unextractable (bound) 14C-residues were determined by combustion and the extractable 14C-residues were obtained by extracting the samples with methanol. Concentrated extracts were first analyzed by thin layer chromatography (TLC). The extracts were also subjected to a series of liquid-liquid extraction procedures for clean-up and the final extracts were analyzed by high performance liquid chromatography (HPLC). Crude hazelnut oil was also extracted with hexane and analyzed for total 14C-residue. A total of 1.3% of applied radioactivity was recovered from the total nut harvested, with 0.04%, 0.06%, and 1.2% present in shell, kernel, and husk, respectively. The results show that the inedible husk and shell contained 95.7% 14C, whereas the edible kernel contained 4.3% of the total 14C recovered. The terminal 14C-residue in hazelnut kernel and oil did not contain carbaryl and/or its metabolite naphthol.     

14C‐dimethoate residues in olive oil during oil processing

Abstract

An olive tree was treated twice in the field with ¹⁴C‐dimethoate (237.7 (μCi, 2.4 g) and ¹⁴C residues were determined in the olive fruits at harvest. The fruits were crushed and pressed to extract the crude oil, then refined by neutralization, bleaching and deodorization. The crude oil contained 14.1% of the total ¹⁴C in the olive fruits. Neutralization resulted in a reduction of ¹⁴C by about 50% of the total ¹⁴C residues in oil. Bleaching and deodorization processes further reduced the ¹⁴C residues and the refined oil contained 31.6% (which corresponds to 4.4% of ^I4C residues of the total ¹⁴C in olive fruits) of the total ¹⁴C in the crude oil. Industrially extracted crude oil was fortified with ¹⁴C‐dimethoate at 1.8 mg kg^‐1 (0.02 μCi) level and subjected to the same refining process. A sharp decrease in the amount of ¹⁴C was observed by neutralization and the amount of ¹⁴C remaining in the refined oil was about 7.3% of the total ^l4C in the crude fortified oil. The data suggest that the ¹⁴C residues in the aged and the fortified oil amples were not of the same nature. The terminal ¹⁴C residue in the refined oil obtained from the field experiment did not contain dimethoate and/or its oxon.     

Bioavailability to rats of bound [14C] pirimiphos-methyl in stored wheat.

Stored wheat treated with radiolabelled pirimiphos-methyl (0-2-diethyl-amino-6-methyl-pyrimidin-4-yl 0,0-dimethyl phosphorothioate) formed bound (nonextractable) 14C residues. Supercritical fluid extraction, gas chromatography and mass spectrometric techniques were used to identify and quantitate the 14C bound residues in wheat grains. The amount of bound 14C residues present after 28 weeks of storage was about 9.9% of the applied radioactivity. Pirimiphos-methyl accounted for 80% of the bound residue. Grain-bound residues were fed to rats for 5 days. After a total period of 8 days a substantially large percentage of the administered bound 14C residues (72.9%) was eliminated in urine while feces contained only 17.9%. Bound pirimiphos-methyl in wheat grain was metabolized in rats by processes involving hydrolysis, N-dealkylation and 0-demethylation. The results indicate that wheat-bound residues of pirimiphos-methyl are highly bioavailable to the rat and may possess a toxicological potential as manifested by a significant reduction in body weight gain.     

Viable skin efficiently absorbs and metabolizes bisphenol A

Skin contact has been hypothesized to contribute to human exposure to bisphenol A (BPA). We examined the diffusion and metabolism of BPA using viable skin models: human skin explants and short-term cultures of pig ear skin, an alternative model for the study of the fate of xenobiotics following contact exposure. ¹⁴C-BPA [50-800 nmol] was applied on the surface of skin models. Radioactivity distribution was measured in all skin compartments and in the diffusion cells of static cells diffusion systems. BPA and metabolites were further quantified by radio-HPLC. BPA was efficiently absorbed in short-term cultures, with no major difference between the models used in the study [viable pig ear skin: 65%; viable human explants: 46%; non-viable (previously frozen) pig skin: 58%]. BPA was extensively metabolized in viable systems only. Major BPA metabolites produced by the skin were BPA mono-glucuronide and BPA mono-sulfate, accounting together for 73% and 27% of the dose, in pig and human, respectively. In conclusion, experiments with viable skin models unequivocally demonstrate that BPA is readily absorbed and metabolized by the skin. The trans-dermal route is expected to contribute substantially to BPA exposure in human, when direct contact with BPA (free monomer) occurs.     

14C]Pentachlorophenol mineralization in the rice rhizosphere with established oxidized and reduced soil layers

Flooded soils with rooted aquatic macrophytes have adjacent aerobic and anaerobic zones at the soil-water interface and rhizosphere where many common soil constituents undergo sequential oxidation and reduction reactions. To investigate whether pentachlorophenol (PCP) mineralization would also be enhanced under these conditions, a laboratory study was conducted to determine the conversion of [14C]PCP to 14CO2, 14CH4 and [14C]humic substances in soil microcosms with established aerobic-anaerobic zones at the soil-water interface and rice (Oryza sativa) rhizosphere. Contrary to what was expected, PCP was least rapidly degraded in rhizosphere-soil microcosms that contained the most extensive amounts of aerobic-anaerobic interfaces (63% PCP loss in 82 d) and was most rapidly degraded in soil microcosms that lacked redox interfaces in the soil profile (94% PCP loss in 82 d). Decreased PCP mineralization in the presence of aerobic-anaerobic interfaces was explained by (i) lack of sufficient O2 for aerobic PCP mineralization, due to the oxidation of other soil constituents in aerobic zones, and (ii) lack of an adequate supply of electron equivalents for reductive dechlorination of PCP, due to the reduction of other alternate electron acceptors in anaerobic zones. It was concluded that PCP mineralization is inhibited in flooded soils that contain extensive amounts of aerobic-anaerobic interfaces, due to redox cycling of other soil constituents that occur in these zones.     

Artificial sweat enhances dermal transfer of chlorpyrifos from treated nylon carpet fibers

The dermal transfer and absorption of 14C-ring-chlorpyrifos from nylon carpet fibers was measured in skin penetration-evaporation cells with excised pig skin. Prior to application, synthetic sweat was applied to skin in half of the cells. Radioactivity was measured in receptor fluid, dermis, epidermis, tape stripping samples, and vapor trap samples during a 24-h period. The sum of radiolabel recovered from the dermis and receptor fluid represented the absorbed dose. There was no significant difference (p > 0.05) in percutaneous absorption between cells that received the synthetic sweat application and "dry" cells (1.3 +/- 0.3% of applied dose). There was significantly more (p < 0.05) radiolabel recovered from tape stripping (5.4 +/- 2.1 vs. 2.8 +/- 0.6%) and in the epidermis (4.5 +/- 0.8 vs. 3.1 +/- 0.3%) from cells that received the synthetic sweat application, which indicated synthetic sweat facilitated transfer of chlorpyrifos from a treated substrate to the skin surface. The measured value for percutaneous absorption of chlorpyrifos agreed with the value predicted from an empirical model previously developed for nitro compound-containing soil.     

Behaviour of [ring-C] hydroxy-simazine in a parabraun soil

[Ring-¹⁴C] hydroxy-simazine was incubated in a parabraun soil for 102 days at 22°C and 65 % of the maximum water-holding capacity. Soil samples were analyzed according to two different extraction methods on the day of application as well as after 13, 62 and 102 days. Furthermore, the influence of air-drying the soil specimens on the extraction yield was also studied. Hydroxy-simazine was converted to ¹⁴CO₂ only up to 0.2 % of the applied radioactivity. A rapid bonding of the radioactivity to the organic mass of the soil was observed, especially in the fulvic acid and humin fractions. 62 days after beginning conversion these two fractions already altogether contained up to 85 % of the labelled ring carbon applied. Whereas in the humic acids only a maximum of 4.5 % was discovered.     

Bioavailability and toxicological potential of lentil-bound residues of malathion in rats.

Lentil grains treated with malathion and stored under laboratory conditions for 12 months formed bound residues. Bioavailability and the effects of lentil-bound residues of malathion in rats were studied. The amount of bound residues in lentils treated with 14C-malathion at 10 ppm and 50 ppm gradually increased to 9.52% and 13.01% of the initially applied doses after 12 months. When rats were fed these 14C-bound residues, radioactivity excreted in urine accounted for 34.49% of the administered dose. In feces, 26.15% of given dose was methanol-extractable while 18.67% was determined as nonextractable. Various tissues including liver, kidney, fat and lungs contained 8.93% while radioactivity in expired air (14CO2) was low (1.51%). The results indicate that lentil-bound malathion residues are highly bioavailable to rats. Analysis of the lentil material containing bound residues indicated that the main compound was malathion. Lentil-bound malathion residues were administered to albino rats at 0.95 and 6.51 ppm in the feed for 3 months. Body weights were determined during and at the end of the experiment. Terminal organ weights were also determined and a number of blood chemistry parameters were examined. A significant reduction in serum cholinesterase activity and an increase in blood urea nitrogen and in white cell count suggest a toxocological potential of the bound residues.     

Release of substituents from phenolic compounds during oxidative coupling reactions

Phenolic compounds originating from plant residue decomposition or microbial metabolism form humic-like polymers during oxidative coupling reactions mediated by various phenoloxidases or metal oxides. Xenobiotic phenols participating in these reactions undergo either polymerization or binding to soil organic matter. Another effect of oxidative coupling is dehalogenation, decarboxylation or demethoxylation of the substrates. To investigate these phenomena, several naturally occurring and xenobiotic phenols were incubated with various phenoloxidases (peroxidase, laccase, tyrosinase) or with birnessite (delta-MnO(2)), and monitored for chloride release, CO(2) evolution, and methanol or methane production. The release of chloride ions during polymerization and binding ranged between 0.2% and 41.4%. Using the test compounds labeled with 14C in three different locations (carboxyl group, aromatic ring, or aliphatic chain), it was demonstrated that 14CO(2) evolution was mainly associated with the release of carboxyl groups (17.8-54.8% of the initial radioactivity). Little mineralization of 14C-labeled aromatic rings or aliphatic carbons occurred in catechol, ferulic or p-coumaric acids (0.1-0.7%). Demethoxylation ranged from 0.5% to 13.9% for 2,6-dimethoxyphenol and syringic acid, respectively. Methylphenols showed no demethylation. In conclusion, dehalogenation, decarboxylation and demethoxylation of phenolic substrates appear to be controlled by a common mechanism, in which various substituents are released if they are attached to carbon atoms involved in coupling. Electron-withdrawing substituents, such as -COOH and -Cl, are more susceptible to release than electron-donating ones, such as -OCH(3) and -CH(3). The release of organic substituents during polymerization and binding of phenols may add to CO(2) production in soil.