Investigation on the Chemical Structure of Nonextractable Residues of the Fungicide Cyprodinil in Spring Wheat Using 13C-C1-Phenyl-Cyprodinil on 13C-Depleted Plants—An Alternative Approach to Investigate Nonextractable Residues

Abstract

¹³C-labelled cyprodinil was applied on ¹³C-depleted wheat plants with 27-fold field application rate. A control experiment applying same amounts of ¹⁴C-cyprodinil showed that main portions of the residues were detected in the cellulose (15% NER), hemicellulose (28.3% NER), and lignin fraction (23.3% NER). 16.7% were detected in water soluble polymers, 6% in both, pectin and protein fraction, and 4% in the starch containing fraction. Free cyprodinil was detectable by TLC in all fractions except lignin. A direct characterization of the residues in vivo by CP-MAS was not successful. Cell wall fractions were further analysed by liquid state NMR to determine the structure of the mobilized highly polymer/polar residues: Within lignin, where most of the residues were located at field application rate, neither intact cyprodinil nor its metabolites could not be detected. The ¹³C-label introduced was probably incorporated in the polymer as natural lignin monomers and thus are not considered as bound residues according to IUPAC definition.     

Studies on bound 14C-prometryn residues in soil and plants

A high-temperature distillation technique was developed for determining and chemically identifying the bound (nonextractable) residues of ¹⁴C-prometryn in an organic soil and plants. A considerable portion of the bound ¹⁴C residues in the incubated organic soil was identified as prometryn. These residues were absorbed by plants grown in the soil. Hono-and di-$\text{N}$-dealkylated metabolites of prometryn were present in the plant bound ¹⁴C residues and a major portion of bound residues as associated with lignin. Soil-bound ¹⁴C residues were also released from soil by microbes. The bound ¹⁴C residues in soil were associated with humin, humic acid, and fulvic acid fractions. Thermoanalytical methods were used to obtained information on the nature and location of ¹⁴C bound residues in soil and humic materials.     

Distribution of the herbicide atrazine in a microcosm with riparian forest plants

Pesticides applied on sugarcane reach the subsoil of riparian forests and probably contaminate the river water. This work was conducted to learn about the phytoremediation of atrazine and subsoil contamination using the common riparian forest species of Cecropia hololeuca Miq. and Trema micranta (L.) Blum. These plants were grown in soil microcosms where ¹⁴C-atrazine at 1/10 of the field-recommended dose was applied at the bottom of the microcosm simulating the movement from contaminated ground water to the upper soil layers and into plants. Residues of ¹⁴C-atrazine were detected in all parts of the microcosm including soil, rhizosphere and the roots in different layers of the microcosm, stem and leaves. Atrazine mineralization was higher (10.2%) in the microcosms with plants than the control microcosms without plants (1.2%). The upward movement of this pesticide from deeper to more superficial soil layers occurred in all the microcosms with plants, powered by evapotranspiration process. From the atrazine applied in this study about 45% was taken up by C. hololeuca and 35% by T. micrantha. The highest amount of radioactivity (%) was found in the fine roots and the specific radioactivity (% g^?1) showed that thick, fine roots and leaves bioaccumulate atrazine. The enhanced mineralization of atrazine as well the phytostabilization effect of the tree biomass will reduce the bioavailability of these residues and consequently decrease the hazardous effects on the environment.     

In vitro release of bound (nonextractable) 14C residues from plants by corn leaves bomogenates

Corn leaves homogenates were found to release bound (nonextractable) ¹⁴C residues from the aerial portion of matured corn plants. The 22,000 g pellet and 10,000 g supernatant fractions were the most active in releasing the bound ¹⁴C residues. The released ¹⁴C residues comprised mainly 2-OH analogues of the $\text{N}$-monodealkylated analogues of atrazine. It is suggested that the enzymatic system in plants may cause metabolic conversion of bound residues.     

Fate of atrazine and chlorpyrifos during solid state fermentation—examination of processes

Abstract

Solid state fermentation (SSF) was investigated as a means to dispose of two commonly used pesticides, chlorpyrifos (O, O‐diethyl O‐(3,5,6‐trichloro‐2‐pyridyl) phosphorothioate) and atrazine (2‐chloro‐4‐ethylamino‐6‐isopropylamino‐1,3,5‐triazine). SSF experiments were carried out in bench‐scale bioreaetors (equipped with CO₂ and volatile organic traps) containing a mixture of lignocellulosic materials and a radiolabeled pesticide. Ethyl acetate‐extractable, alkali soluble, and alkali insoluble fractions were evaluated for radioactivity following a 60‐d incubation period at 40°C. The majority of the [2, 6‐pyridyl‐¹⁴C]chlorpyrifos was associated with the ethyl acetate extract (about 74%), 17% was trapped as organic volatiles by polyurethane foam traps and < 0.5% of the chlorpyrifos was mineralized to CO². Only small amounts of the radioactivity were associated with alkali soluble (0.0003%) and alkali insoluble (0.3%) fractions. In the [¹⁴C‐U‐ring] atrazine bioreactors, very little of the radioactivity volatilized (<0.5%) and less than 0.5% was mineralized to CO₂. Approximately 57% of the applied radioactivity was associated with the ethyl acetate extract while 9% and 24% of the radioactivity was associated with the alkali soluble (humic and fulvic acids) and alkali insoluble fractions, respectively. Possible reaction mechanisms by which covalent bonds could be formed between atrazine (or metabolites) and humic substances were investigated. The issue of bound atrazine residue (alkali soluble fraction) was at least partially resolved. Oxidative coupling experiments revealed that formation of covalent bond linkages between amino substituent groups of atrazine residue and humic substances is highly unlikely.     

Effect of organic amendment on degradation and formation of bound residues of methabenzthiazuron in soil under constant climatic conditions

Abstract

The degradation of [phenyl‐U‐¹⁴C]methabenzthiazuron (MBT) and formation of bound residues in the surface soil of an orthic luvisol were studied under constant climatic conditions (20°C, 40 % of maximum water holding capacity). In two treatments (with and without preincubation in the soil) maize straw was amended at a rate of 1.5 g/100 g dry soil in addition to the application of MBT. The mineralization of uniformly labeled maize straw was studied simultaneously. In additional flasks, MBT was incubated at 0, 10 and 30°C with and without addition of maize straw.

The turnover of the amended maize straw led to an enhanced dissipation of MBT which was mainly due to the formation of bound residues. This corresponded to a higher microbial activity in the soil after straw amendment and the intensive mineralization of the radiolabeled maize straw. About 2–3 % of the applied radioactivity from the radiolabeled maize straw was measured in the soil microbial biomass 10 and 40 days after application whereas ¹⁴C from MBT was only incorporated into soil microbial biomass in the treatments with straw amendment.

Within the bound residue fractions relatively more radioactivity was measured in fulvic and humic acids after straw amendment. Increasing temperatures promoted the dissipation of MBT and the formation of bound residues in both treatments, but without amendment of maize straw these effects were far less pronounced. The laboratory scale degradation experiment led to similar results as were found in a corresponding lysimeter study. Differences that were observed could be explained by different temperature regimes of the experiments and time of aging in soil.     

The metabolic fate of 14C‐parathion by some fresh water phytoplankton and its possible effects on the algal metabolism

Abstract

The growth and total carbohydrate contents of Nostoc muscorum and Tolypothrix tenuis were greatly and significantly reduced by the application of parathion. “Chlorophyll a”, carotene biosynthesis and the rate of glucose absorption were enhanced after supplementation of parathion to the culture media of both cyanobacteria. Nitrogen released to the media, total nitrogen content and total nitrogen fixed were increased in both organisms‐ Increase in protein content was accompanied by remarkable drop in amino, peptide and ammonia fractions‐ Phosphorus uptake, RNA, DNA and total phosphorus content were accelerated to reach maximum accumulation at the highest insecticide level. In metabolism study using ¹⁴C‐labelled compound, parathion was readily degraded by Nostoc and Tolypothrix. Following ten days incubation, the aqueous fractions contained 21.1% and 18.1% of the initial activity in Nostoc and Tolypothrix respectively. TLC analysis of the hydrolytic products revealed the presence of three metabolites: p‐aminophenol, p‐nitrophenol and aminoparathion.     

Metabolism of the Nonylphenol Isomer [Ring-U-14c]-4-(3′,5′-Dimethyl-3′-Heptyl)-Phenol by Cell Suspension Cultures of Agrostemma githago and Soybean

Abstract

The biotransformation of the nonylphenol isomer [ring-U-¹⁴C]-4-(3′,5′-dimethyl-3′-heptyl)-phenol (4-353-NP, consisting of two diastereomers) was studied in soybean and Agrostemma githago cell suspension cultures. With the A. githago cells, a batch two-liquid-phase system (medium/n-hexadecane 200:1, v/v) was used, in order to produce higher concentrations and amounts of 4-353-NP metabolites for their identification; 4-353-NP was applied via the n-hexadecane phase. Initial concentrations of [¹⁴C]-4-353-NP were 1 mg L^?1 (soybean), and 5 and 10 mg L^?1 (A. githago). After 2 (soybean) and 7 days (A. githago) of incubation, the applied 4-353-NP was transformed almost completely by both plant species to four types of products: glycosides of parent 4-353-NP, glycosides of primary 4-353-NP metabolites, nonextractable residues and unknown, possibly polymeric materials detected in the media. The latter two products emerged especially in soybean cultures. Portions of primary metabolites amounted to 19–22% (soybean) and 21–42% of applied ¹⁴C (A. githago). After liberation from their glycosides, the primary 4-353-NP metabolites formed by A. githago were isolated by HPLC and examined by GC-EIMS as trimethylsilyl derivatives. In the chromatograms, eight peaks were detected which due to their mass spectra, could be traced back to 4-353-NP. Seven of the compounds were side-chain monohydroxylated 4-353-NP metabolites, while the remaining was a (side-chain) carboxylic acid derivative. Unequivocal identification of the sites of hydroxylation/oxidation of all transformation products was not possible. The main primary metabolites produced by A. githago were supposed to be four diastereomers of 6′-hydroxy-4-353-NP (about 80% of all products identified). It was concluded that plants contribute to the environmental degradation of the xenoestrogen nonylphenol; the toxicological properties of side-chain hydroxylated nonylphenols remain to be examined.     

Plant availability of bound anilazine residues in a degraded loess soil

Abstract

The relative biological availability of [benzene ring‐U‐¹⁴C] and Ctriazine‐U‐¹⁴C] anilazine for maize plants was studied in a degraded loess soil in a standardized microecosystem. The total uptake of radiocarbon in the course of the 4‐week experiment was 3.1 and 4 % respectively of the radioactivity applied if anilazine was uniformly mixed into the soil immediately before beginning the experiment. However, if anilazine was subjected to a degradation at 65 % of the maximum water holding capacity of the soil and temperatures varying daily between 16 and 27°C for 100 days before the plant experiment then the uptake was reduced to 0.4 or 0.7 % respectively. The uptake from soil with non‐extractable (bound) anilazine residues was similarly low. The mineralization rate of aged and bound anilazine residues was below 0.1 % of the radioactivity applied. Up to 2/3 of the radioactivity present in the soil after the plant experiment remained in the humic fraction.     

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.     

Biodegradation of Trifluralin in Harran Soil

Abstract

Degradation of trifluralin (α,α,α-trifluoro-2,6-dinitro-N,N-dipropyl-p-toluidine) was investigated in soils taken from three different locations at Harran region of Turkey under laboratory conditions. Surface (0–10 cm) soils, which were taken from a pesticide untreated field Gürgelen, Harran-1 and Ikizce regions in the Harran Plain, were incubated in biometer flasks for 350 days at 25°C. Ring-UL-¹⁴C-trifluralin was applied at the rate of 2 µg g^?1 with 78.7 kBq radioactivity per 100 g soil flask. Evolved ¹⁴CO₂ was monitored in KOH traps throughout the experiment. Periodically, soil sub-samples were removed and extracted by supercritical fluid extraction (SFE). Unextractable soil-bound ¹⁴C residues were determined by combustion. During the 350 days incubation period 6.6, 5.4, and 3.3% of the applied radiocarbon was evolved as ¹⁴CO₂ from the Harran-1, Gürgelen, and Ikizce soil, respectively. At the end of 350 days the SFE-extractable and bound ¹⁴C-trifluralin residues were 39.0 and 29.2% of the initially applied herbicide in Gürgelen soil. The corresponding values for Harran-1 and Ikizce soils were 36.2, 28.4% and 41.6, 18.5% respectively.     

Metabolic fate of the 14C-labeled herbicide clodinafop-propargyl in soil

The metabolic fate of ¹⁴C-phenyl-labeled herbicide clodinafop-propargyl (CfP) was studied for 28 days in lab assays using a soil from Germany (A_p horizon, silt loam, and cambisol). Mineralization amounted to 12.40% of applied ¹⁴C after 28 days showing a distinct lag phase until day 7 of incubation. Portions of radioactivity extractable by means of 0.01 M CaCl₂ solution (bioavailable fraction) decreased rapidly and were 4.41% after 28 days. Even immediately after application, only 57.31% were extracted with the aqueous solvent. Subsequent extraction using accelerated solvent extraction (ASE; acetonitrile/water 4:1, v/v) released 39.91% of applied ¹⁴C with day 0 and 26.16% with day 28 of incubation from the samples. Non-extractable portions of radioactivity thus, increased with time amounting to 11.99% (day 0) and 65.00% (day 28). A remarkable increase was observed between 14 and 28 days correlating with the distinct increase of mineralization. No correlation was found throughout incubation with general microbial activity as determined by DMSO reduction. Analysis of the CaCl₂ and ASE extracts by radio-TLC, radio-HPLC and GC/MS revealed that CfP was rapidly cleaved to free acid clodinafop (Cf), which was further (bio-) transformed; DT₅₀ values (based on radio-TLC detection of the parent compound) were far below 1 day (CfP) and about 7 days (Cf). TLC analysis pointed to 2-(4-hydroxyphenoxy)-propionic acid as further metabolite. Due to fractionation of non-extractable residues, most of the ¹⁴C was associated with fulvic and humic acids, portions in humin fractions and non-humics were moderate and low, respectively. Using a special strategy, which included pre-incubation of the soil with CfP and then mineralization of ¹⁴C-CfP as criterion, a microorganism was isolated from the soil examined. The microorganism grew using CfP as sole carbon source with concomitant evolution of ¹⁴CO₂. The bacterium was characterized by growth on commonly used carbon sources and by 16S rDNA sequence analysis. The sequence exhibited high similarity with that of Rhodococcus wratislaviensis (99.56%; DSM 44107, NCIMB 13082).     

Release of soil bound (nonextractable) residues by various physiological groups of microorganisms∗

Abstract

Soil bound ¹⁴C‐labeled residues were released by four different physiological groups of microorganisms from an organic soil treated with ¹⁴C‐ring‐labeled prometryn [2‐(methylthio) ‐4,6‐bis(isopropylamino)‐s‐triazine]. The extent to which the different microbial populations released bound ¹⁴C residues (25–30% of the total bound ¹⁴C) from the Y‐irradiated soil after 28 days incubation did not differ considerably. Analysis of the extractable material from the incubated soil showed the presence of small amounts of the parent compound, and its hydroxy and mono‐N‐dealkylated analogues. Low level of ¹⁴CO₂ (1.5–3.0% of the total bound ¹⁴C) was evolved from the microbial systems indicating ring cleavage of the released material as being a very minor reaction.     

Metabolism of the14C-labeled herbicide clodinafop-propargyl in plant cell cultures of wheat and tobacco

The metabolism of ¹⁴C-clodinafop-propargyl (CfP) was examined in cell cultures of wheat (Triticum aestivum L. cv. ‘Heines Koga II’) and tobacco (Nicotiana tabacum L.). Besides the non-transgenic tobacco culture, cultures transformed separately with cDNA of human cytochrome P450-monooxygenases (P450s) CYP1A1, CYP1A2, CYP3A4, CYP2B6 and CYP2C19 were examined. Experiments with wheat were executed in the presence and absence of safener cloquintocet-mexyl (CqM). After 48 h of incubation, only about 10% of applied ¹⁴C was found in media (both tobacco and wheat). Non-extractable residues of ¹⁴C-CfP in wheat cells were 16.54% (without CqM) and 30.87% (with CqM). In all tobacco cultures, 82.41–92.46% of applied radioactivity was recovered in cell extracts. In contrast to wheat, non-extractable residues amounted only to 1.50–2.82%. As determined by radio-thin layer chromatography (TLC) and -high-performance liquid chromatography (HPLC), the parent CfP was not found in the cell extracts of wheat; in tobacco cell extracts, only traces of CfP were detected. After a hydrolysis of assumed carbohydrate conjugates of CfP derived polar ¹⁴C-labeled compounds, TLC and HPLC analysis showed that in wheat, a more complex pattern of metabolites of CfP were observed as compared to all tobacco cultures. In hydrolysates resulting from wheat, the identity of three primary products was confirmed by means of GC-EI-MS: free acid clodinafop (Cf), hydroxy-Cf hydroxylated at the pyridinyl moiety, and 4-(5-chloro-3-fluoropyridin-2-yloxy)phenol. In hydrolysates derived from all tobacco cultures, main metabolite was Cf besides only traces of further unidentified products. Differences among the different tobacco cultures (non-transgenic, transgenic) did not emerge. According to kinetics of disappearance of primary metabolite Cf as well as formation of polar soluble products and non-extractable residues, metabolization of CfP proceeded at a noticeably higher rate in wheat cells treated with safener CqM than in cells without CqM treatment. Thus, these results indicated a stimulation of CfP's metabolism by CqM, although metabolic profiles observed in CqM treated and non-treated cells (after hydrolysis) were qualitatively similar. The findings obtained from all tobacco cultures suggested that with the exception of ester cleavage to Cf, CfP cannot be metabolized by tobacco itself or by the human P450s examined.     

Uptake of 14C-atropine and/or its transformation products from soil by wheat (Triticum aestivum var Kronjet) and their translocation to shoots

Plant uptake of toxins and their translocation to edible plant parts are important processes in the transfer of contaminants into the food chain. Atropine, a highly toxic muscarine receptor antagonist produced by Solanacea species, is found in all plant tissues and can enter the soil and hence be available for uptake by crops. The absorption of atropine and/or its transformation products from soil by wheat (Triticum aestivum var Kronjet) and its distribution to shoots was investigated by growing wheat in soil spiked with unlabeled or ¹⁴C-labeled atropine. Radioactivity attributable to ¹⁴C-atropine and its transformation products was measurable in plants sampled at 15 d after sowing (DAS) and thereafter until the end of experiment. The highest accumulation of ¹⁴C-atropine and/or its transformation products by plants was detected in leaves (between 73 and 90% of the total accumulated) with lower amounts in stems, roots, and seeds (approximately 14%, 9%, and 3%, respectively). ¹⁴C-Atropine and/or its transformation products were detected in soil leachate at 30, 60, and 90 DAS and were strongly adsorbed to soil, with 60% of the applied dose adsorbed at 30 DAS, plateauing at 70% from 60 DAS. Unlabeled atropine was detected in shoots 30 DAS at a concentration of 3.9 ± 0.1 μg kg^?1 (mean ± SD). The observed bioconcentration factor was 2.3 ± 0.04. The results suggest a potential risk of atropine toxicity to consumers.     

Metabolic fate of the 14C-labeled herbicide clodinafop-propargyl in a sediment–water system

The metabolic fate of ¹⁴C-phenyl-labeled herbicide clodinafop-propargyl (¹⁴C-CfP) was studied for 28 days in lab assays using a sediment–water system derived from a German location. Mineralization was 5.21% of applied ¹⁴C after 28 days exhibiting a distinct lag phase until day 14 of incubation. Portions of radioactivity remaining in water phases decreased at moderate rate to 18.48% after 28 days; 62.46% were still detected in water after 14 days. Soxhlet extraction of the sediment using acetonitrile released 35.56% of applied ¹⁴C with day 28, while 33.99% remained as non-extractable residues. A remarkable increase of bound ¹⁴C was observed between 14 and 28 days correlating with the distinct increase of mineralization. No correlation was found throughout incubation with microbial activity of the sediment as determined by dimethyl sulfoxide reduction. Dissolved oxygen and pH value of water phases remained almost constant for 28 days. Analyses of Soxhlet extracts of the sediment and ethyl acetate extracts of water phases by radio-TLC and radio-HPLC revealed that CfP was rapidly cleaved to free acid clodinafop (Cf), which was further (bio-) transformed. DT₅₀ values (based on radio-HPLC) were below 1 day (CfP) and slightly above 28 days (Cf). Further metabolites were not detected. Fractionation of humic and non-humic components of the sediment demonstrated that CfP's non-extractable residues were predominantly associated with fulvic acids up to 14 days of incubation (3.36%), whereas after 28 days, the majority of radioactivity was found in the humin/mineral fraction (13.30% of applied ¹⁴C). Due to high-performance size-exclusion chromatography of the fulvic acids fraction derived from assays incubated for 28 days, this portion of ¹⁴C was firmly, possibly covalently bound to fulvic acids and did not consist of CfP or Cf. Using an isolation strategy comprising preincubation of sediment with CfP and mineralization of ¹⁴C-CfP as criterion, a microorganism was isolated from the sediment examined. It grew on ¹⁴C-CfP as sole carbon source with evolution of ¹⁴CO₂. The bacterium was characterized by growth on commonly used carbon sources and 16S rDNA sequence analysis. Its sequence exhibited high similarity with that of Nocardioides aromaticivorans strain H-1 (98.85%; DSM 15131, JCM 11674).     

Fate of 14C-ethyl prothiofos insecticide in canola seeds and oils

Canola plants were treated with ¹⁴C- prohiofos under conditions simulating local agricultural practices. ¹⁴C-residues in seeds were determined at different time intervals. At harvest time about 32 % of ¹⁴C-activity was associated with oil. The methanol soluble ¹⁴C-residues accounted for 12 % of the total seed residues after further seeds extraction, while the cake contained about 49 % of the total residues. About 69 % of the ¹⁴C-activity in the crude oil could be eliminated by simulated commercial processes locally used for oil refining. Chromatographic analysis of crude and refined oil revealed the presence of the parent compound together with three metabolites which were identified as prothiofos oxon, O-ethyl phosphorothioate and O-ethyl S-propyl phosphorothioate, besides one unknown compound. While methanol extract revealed the presence of despropylthio prothiofos and O-ethyl phosphoric acid as free metabolites acid hydrolysis of the conjugated metabolites in the methanol extract yielded 2, 4-dichlorophenole which was detected by color. When rats were fed the extracted cake for 72 hours, the bound residues were found to be bioavailable. The main excretion route was via the expired air (42 %), while the ¹⁴C-residues excreted in urine and feces were 30 % and 11 %, respectively. The radioactivity detected among various organs accounted to 7.5 %.Chromatographic analysis of urine indicated the presence of prothiofos oxon, O-ethyl phosphoric acid and 2, 4-dichlorophenole as main degradation products of prothiofos in free and conjugated form.     

Comparative studies of the fate of atrazine‐14C and pentachlorophenol‐14C in various laboratory and outdoor soil‐plant systems

Abstract

Mass balance and fate of atrazine‐ ¹⁴C and pentachlorophenol‐ ¹⁴C (PCP‐ ¹⁴C) were studied in short‐term tests in a closed aerated laboratory soil‐plant system, using two concentrations in soil and two plant species, as well as under outdoor conditions for one vegetation period. In the laboratory, for both pesticides bioaccu‐mulation factors of radiocarbon taken up by the roots into plants were low. They were higher for lower (1 ppm) than for higher soil concentrations (6 ppm for atra‐zine, 4 ppm for pentachlorophenol) and varied with the plant species. Mineralization to ¹⁴CO₂ in soil was negatively related to soil concentration only for PCP‐ ¹⁴C. Conversion rates in soil including the formation of soil‐bound residues were higher for the lower concentrations of both pesticides than for the higher ones; conversion rates in plants were species‐dependent. In 14 terms of CO₂ formation and of conversion rates, PCP was less persistent in soil than was atrazine. For both pesticides, laboratory data on conversion and mineralization gave a rough prediction of their persistence in soil under long‐term outdoor conditions, whereas bio‐accumulation factors in plants under long‐term outdoor conditions could not be predicted by short‐term laboratory experiments.     

Fate of the veterinary antibiotic 14C-difloxacin in soil including simultaneous amendment of pig manure with the focus on non-extractable residues

The fate of ¹⁴C-labeled difloxacin (¹⁴C-DIF) was studied in time course experiments after application on soil (Ap horizon of silt loam) and amendment of authentic DIF containing pig manure (146 mL kg^?1; 4.17 MBq kg^?1; 0.85 mg kg^?1) or water (124 mL kg^?1; 0.42 MBq kg^?1; 0.09 mg kg^?1) for 56 and 120 days of incubation, respectively.

Mineralization of ¹⁴C-DIF was below 0.2% in both experiments after 56 days or 120 days. In the course of the experiments, portions of extractable radioactivity (Accelerated Solvent Extraction (ASE); acetonitrile-water) decreased to 19–21% depending only little on manure amendment. Non-extractable residues of ¹⁴C-DIF increased to 70–74% after 56 days and 120 days, respectively, and therefore were the main route of ¹⁴C-DIF in soil. According to radioanalytical HPLC and LC-MS/MS, only the parent compound was found in all extracts over the whole time of the experiment. According to fractionation of the non-extractable residues (NER) into particle size fractions, ¹⁴C portions were associated to the water used for fractionation, the silt and clay fractions, whereas no radioactivity was detected in the sand fraction. The majority of ¹⁴C was found within the clay fractions.

Fractionation of humic components showed that radioactivity derived from ¹⁴C-DIF was associated with humic acids, fulvic acids, humins and minerals and very little with soluble, non-humic HCl fraction. The highest portions of radioactivity were found in the fulvic acid fraction. Results obtained by size exclusion chromatography (SEC) of the purified fulvic acids were similar for every sample analyzed. One large portion of ¹⁴C co-eluted with fulvic acids of a molecular weight below 910 g mol^?1. Both fractionation methods demonstrated that the parent compound DIF or initial metabolites were rapidly integrated into humic materials and, thus, were major components of NER.     

Fate of the 14C-labeled herbicide prosulfocarb in a soil and in a sediment-water system

The fate of ¹⁴C-labeled herbicide prosulfocarb was studied in an agricultural soil and in a sediment-water system, the sediment part of which was derived from Yangtze Three Gorges Reservoir, China. Time-course studies were performed for 28 d and 49 d, respectively. Main transformation routes of ¹⁴C-prosulfocarb were mineralization to ¹⁴CO₂ and formation of nonextractable residues amounting to 12.13% and 10.43%, respectively, after 28 days (soil), and 9.40% and 11.98%, respectively, after 49 d (sediment-water system). Traces of prosulfocarbsulfoxide were detected by means of TLC, HPLC, and LC-MS; other transformation products were not found. Initial extraction of soil assays using 0.01 M CaCl₂ solution showed that the bioavailability of the herbicide was considerably low; immediately after application (0.1 d of incubation), only 4.78% of applied radioactivity were detected in this aqueous fraction. DT₅₀ values of ¹⁴C-prosulfocarb estimated from radio-TLC and -HPLC analyses were above 28 d in soil and ranged between 29 d and 49 d in the sediment-water system. Partitioning of ¹⁴C from water to sediment phase occurred with DT₅₀ slightly above 2 d. With regard to the sediment-water system, adsorption occurred with log K_oc = 1.38 (calculated from 2 day assays) and 2.35 (49 d assays). As similarly estimated from portions of ¹⁴C found in CaCl₂ extracts of the 0.1 d assays, ¹⁴C-prosulfocarb's log K_oc in soil was 2.96. With both experiments, similar portions of nonextractable radioactivity were associated with all soil organic matter fractions, i.e. nonhumics, fulvic acids, humic acids, and humin/minerals. Throughout all sample preparation, the experiments were severely impaired by losses of radioactivity especially with concentration of samples containing water in vacuo. All findings pointed to volatility of parent prosulfocarb in presence of water rather than volatility of transformation products. According to literature data, this behavior of prosulfocarb was not expected, though volatility was demonstrated under field conditions.