Water extrustion in the trap bladders ofUtricularia vulgaris

In the trap bladder ofUtricularia vulgaris, increase in sucrose concentrations in bladder lumen fluid decreased resetting rate. Addition of 350 mM sucrose to lumen fluid stopped the resetting. Therefore, water seems to move down the water potential gradient between the lumen and the arm cells of bifid trichomes, which are the site of inlet in the water pathway. Application of dinitrophenol, sodium azide, KCN, monoiodoacetic acid or pentachlorophenol in lumen fluid much reduced the water outflow. Temperature coefficient of bladder resettings was about 2. No effect of darkness on resetting rate was found. These facts show that the resetting requires energy supplied from respiration and there exists an active ion transport mechanism somewhere in the water pathway. No effect on the resetting was seen upon immersing the bladder in 700 mM surcose solution. In the capital cells of the pavement epithelium in its outer and middle zones, which are the site of outlet in water pathway, membrance potential and resistance were lower than those in other cells. These facts indicate that bulk flow of the cell sap from the capital cells to the outside takes place by intracellular hydrostatic pressure.     

Functional characteristics of traps of aquatic carnivorous Utricularia species

Firing and resetting of traps in aquatic Utricularia species are associated with water flow and trap volume changes. In this study, trap thickness was used as a measure of water flow and was monitored automatically using an electronic position sensor. The basic characteristics of mechanically stimulated firing and resetting were measured in isolated traps from 13 aquatic Utricularia species and in two trap size categories in Utricularia reflexa. This allowed to study the relationship between these trap functions and trap thickness and length (criteria of trap size). Additionally, the characteristics of spontaneous firings (without any mechanical stimulation) were compared for U. reflexa traps fed or denied prey during a 1-day period. On the absolute scale, the 13 Utricularia species differed considerably in their firing and resetting rates. Significant interspecific differences were also found in the magnitude of firing (in total 3.7–4.2 times) and resetting rates (10–24 times) per unit trap thickness or length. Overall, traps of Utricularia australis, Utricularia stellaris and Utricularia inflata showed the greatest firing and resetting rates. The relative magnitude of firing per unit trap thickness or length showed a highly significant negative correlation with both trap thickness and length and the same also held for the relative resetting rates. Smaller and narrower traps are thus relatively more effective at trap firing and resetting than larger traps. Neither firing nor resetting characteristics were significantly different between unfed and prey-fed traps of U. reflexa and this was also true for the occurrence of spontaneous firings. A strict linear resetting rate, without any lag-period, was found during the first 3 min after trap firing in U. reflexa. This suggests that water is pumped out of the trap continuously and probably recirculates. Given the concepts of spontaneous firing and water recirculation, an ecological model based on the literature data has been devised to quantify the daily N and P gain from the ambient water by Utricularia traps devoid of animal prey. The model shows that the total N and P content estimated in the trap fluid is too high to be accumulated from only the ambient water. This implies that the prey-free traps do not take up N or P from the trap fluid but rather exude a quantity of these nutrients to the fluid to support the microbial community. Therefore, the trap microorganisms behave more as parasites than commensals and represent an additional ecological cost for trap maintenance.     

The comparison of mechanically stimulated and spontaneous firings in traps of aquatic carnivorous Utricularia species

Firing and resetting of traps in aquatic Utricularia species are associated with water flows and trap volume changes. In this study, trap thickness was used as a measure of water flow and was monitored automatically using an electronic position sensor. Isolated traps from three aquatic Utricularia species were monitored over the course of 1-2 days to verify spontaneous firings (without any mechanical stimulation) and describe their basic characteristics. Isolated traps of three Utricularia species were initially fired by mechanical stimulation and allowed to naturally reset within a period of 24-48 h. Within this resting period, spontaneous firings were found in the traps of all species and in two trap age categories of U. vulgaris. The timing of spontaneous firings was found to be irregular. Spontaneous firings ranged between 0.29 and 2.4 during the 24-h resting period and the mean time between two spontaneous firings was highly variable within each species (319-891 min). There was no quantitative difference between spontaneous and mechanically stimulated firings of the traps. Spontaneous firings could explain how phytoplankton or detritus enters traps even when no prey species are present.     

Simple rules based on pile slope are used in the self organization of sand pile formation by <Emphasis Type="Italic">Pheidole oxyops</Emphasis> ants

We tested the hypothesis that slope influences where worker ants deposit excavated soil on piles near the nest entrance. We predicted that ants will deposit their load near the top of a pile where the slope changes from upward to downward, to prevent material rolling back towards the entrance. We tested this hypothesis by studying five natural colonies of Pheidole oxyops ants at a field site at S?o Sim?o, Brazil. At this site, each colony was dumping sandy soil excavated from its underground nest in a crescent-shaped pile c. 13 cm from and perpendicular to the nest entrance. Each nest was given an experimental sand pile of symmetrical curved cross section on a plywood platform that could be tilted 15 degrees up or down. From videos, the locations where individual ants dumped their soil loads were measured in relation to the inner (position = 0) and outer (position = 1) edges of the pile. When the platform was tilted down the ants deposited their loads significantly closer to the inner edge (0.458 ± 0.007) than when not tilted (0.530 ± 0.006). When the platform was tilted up the ants deposited their loads significantly further from the inner edge (0.626 ± 0.006) than when not tilted (0.522 ± 0.006). These results support the hypothesis that ants use pile slope in deciding where to dump their load. A similar rule is probably used in other ant species that place excavated soil in steep piles near the nest entrance. Received 5 February 2007; revised 10 June 2007; accepted 9 October 2007.     

Methionine biosynthesis in Saccharomyces cerevisiae: mutations at the regulatory locus ETH2. I. Genetic data

Summary Evidence has been obtained that sodium azide is an inhibitor of cell division in wild-type and aziA strains of Salmonella typhimurium. The bacteria grown in media containing sodium azide and glucose formed long filaments. It has been found that sodium azide had a stronger inhibitory effect on DNA synthesis than on cell mass increase. When filaments produced by azide action were transferred to azide-free medium very rapid increase in DNA content was observed during the first 45 min. After this time, when relative DNA content was increased the rate of DNA synthesis was reduced and cell divisions reappeared.Inhibitory effect of azide on DNA biosynthesis in vitro was observed with toluenized cells of S. typhimurium. Only ATP-dependent radioactive dTMP incorporation into DNA was affected by sodium azide. It had no effect on the incorporation in the absence of ATP.Mutant aziC was isolated in S. typhimurium by scoring for clones with normal cell division in the presence of sodium azide. Azide had much less effect on DNA biosynthesis in vivo and in vitro in aziC cells as compared with isogenic controls.This work was supported by the Polish Academy of Sciences within the project 09.3.1., and by the U.S. Public Health Service, grant No. 05-032-1.     

Selection of biochemical mutants of Aspergillus niger with enhanced extracellular ribonuclease production

Combined with u.v. irradiation and the nitrosoguanidine method, selection of biochemical mutants resistant to metabolic inhibitors (2-deoxy-D-glucose, antimycin A, sodium orthovanadate and sodium azide) was a very efficient method for improvement of ribonuclease production by Aspergillus niger. Resistance to sodium azide produced highest RNase production, greatest frequency of positive mutation and shortest sporulation time. The most active strain, Aspergillus niger SA-13-20 resistant to sodium azide, was obtained, which had a 433% increase in RNase production in comparison with the parent strain and had good subculturing stability. Its dynamic characters were similar to those of the parent strain.     

Trapping and sex pheromone-mediated flight and landing behaviour of maleCtenopseustis obliquana

A sex pheromone-baited delta trap was found to be inefficient at eliciting landing and entering of the trap by maleCtenopseustis obliquana. The inefficiency of the delta trap related to turbulence altering the pheromone plume and the concomitant effect on the flight manoeuvres of male moths. In the wind tunnel, high proportions of males flew upwind and landed on the sides, outside, of the trap, but only a relatively small proportion of these males entered the trap and contacted the sticky surface. When males approached the delta trap, they tended to fly in wide zigzags (i.e., large inter-track reversal distances) and at an altitude near the top of the trap, where the trap was relatively narrow in width (compared to the bottom). Thus, these flight manoeuvres largely precluded males from entering the trap. Greater numbers of male moths entered the trap when: (i) the front barriers of the delta trap were removed, (ii) pheromone dosage was increased to 300 μg, and (iii) the trap design was changed to a rectangular one. The first two changes appeared to influence the flight manoeuvres of males (who appeared to fly with narrower inter-track reversal distances), while the third change apparently did not affect the glight manoeuvres of males, but rather allowed more males to enter the trap because of the greater area of the entrance. The low trap catches of maleC. obliquana in the field are also a consequence of the glue which is currently used. After contact with this glue most males are able to escape, flying off the sticky surface and losing contact with the pheromone. A field trial found that delta traps with another glue caught more than three times the number of males of the related tortricid moth,Epiphyas postvittana, than delta traps with the currently used glue.     

Altered control of glutamate dehydrogenases in ornithine utilization mutants of Pseudomonas aeruginosa

Two classes of ornithine-nonutilizing (oru) mutants of Pseudomonas aeruginosa PAO were investigated. Strains carrying the oru-310 mutation were entirely unable to grow on l-ornithine as the only carbon and nitrogen source and were affected in the assimilation of a variety of nitrogen sources (e.g., amino acids, nitrate). The oru-310 mutation caused changes in the regulation of the catabolic NAD-dependent glutamate dehydrogenase; this enzyme was no longer inducible by glutamate but instead could be induced by ammonia. The oru-310 locus was cotransducible with car-9 and tolA in the 10 min region of the chromosome. An oru-314 mutant was severely handicapped in ornithine medium but could grow when a good carbon source was added; the mutant also showed pleiotropic growth effects related to nitrogen metabolism. The oru-314 mutation affected the regulation of the anabolic NADP-dependent glutamate dehydrogenase, which was no longer repressed by glutamate but showed normal derepression in the presence of ammonia. The oru-314 locus was mapped by transduction near met-9011 at 55 min. Both oru mutants could grow on l-glutamate, l-proline, or l-ornithine amended with 2-oxoglutarate, albeit slowly. We speculate that insufficient 2-oxoglutarate concentrations might account, at least in part, for the Oru^- phenotype of the mutants.     

Characterization of Foam- and Emulsion-stabilizing Functions of Enzymatically Modified Proteins with Surfactancy

A papain-catalyzed reaction involving covalent incorporation of l-leucine n-alkyl ester is available for producing an enzymatically modified protein (EMP) with surfactancy [Agric. Biol. Chem., 45, 1621 (1981)]. In the present work we used gelatin as a starting material and incorporated l-leucine n-hexyl ester to produce a whippable EMP and l-leucine n-dodecyl ester to produce an emulsifiable EMP. A foam system formed with the whippable EMP was much stabler than that formed with sodium dodecylsulfate. The emulsifiable EMP also gave a much stabler oil-in-water emulsion than Tween-80 did. The stability of the emulsion formed with EMP was not affected by the presence of NaCl at a very high concentration. The observed foam and emulsion stabilities were well explained by the data for decreased mobility of the involved water protons. These results may indicate that EMP molecules, when arranged at the air/water or oil/water interface, can bind a part of the water to form thick barriers which prevent the air or oil particles from coalescing.     

Organization of mature external glands on the trap and other organs of the bladderwortUtricularia monanthos

Summary The mature dome-shaped glands which cover the outer surfaces of the trap, leaves, anchor and runner stolons inU. monanthos are described using conventional and some high voltage transmission electron microscopy. The glands occur as scattered ordinary external glands and as a compact clump of vestibule glands at the entrance to the doorway. Each gland rests on a basal epidermal cell and consists of a single pedestal and terminal cell. Vestibule and leaf glands differ slightly from the other glands mainly in the structure of the outer wall of the terminal cell. Nuclear crystals are prominent and the cytoplasm of the pedestal and terminal cells contains tubular structures usually aggregated near the nucleus. The pedestal cell is a transfer cell with short wall protuberances on the outer wall, conspicuous mitochondria and a heavily impregnated lateral wall.The terminal cell often has an outer wall that is greatly thickened and a protoplast that may degenerate early. In the most developed cells the protoplast remains active for a long period and the outer wall is differentiated into several layers. The outermost layer is cuticularized consisting of an open meshwork of deposits. In leaf glands a local polysaccharide mass is usually developed within the cuticularized region. The inner non-impregnated region of the outer wall may show four layers. In vestibule glands fewer layers are present and the wall shows prominent lamellations. Some ordinary external glands differentiate a sponge-like substructure within the inner wall.The ultrastructure and function of the glands are discussed. We support the concept that mature external glands are responsible for secreting water, with those on traps being particularly active during the resetting of the organ. Our work provides a structural basis for recent suggestions by other workers that the mechanism of secretion probably involves establishing a standing osmotic gradient within the gland.     

Decolorization of Victoria blue by the white rot fungus, <Emphasis Type="Italic">Phanerochaete chrysosporium</Emphasis>

Synthetic textile dyes are among the most dangerous chemical pollutants released in industrial wastewater streams. Recognizing the importance of reducing the environmental impact of these dyes, the ability of the white rot fungus Phanerochaete chrysosporium to decolorize various textile dyes was investigated. This fungus decolorized 6 of the 14 structurally diverse dyes with varying efficiency (between 14% and 52%). There was no discernable pattern of decolorization even among dyes of the same chemical class, suggesting that attack on the dyes is relatively non-specific. Among the three dyes which showed >40% decolorization, Victoria Blue B (VB) was chosen for further analysis because the ability of the fungus to decolorize VB was nearly independent over a relatively broad concentration range. Blocking lignin peroxidase (LiP) and manganese peroxidase (MnP) production by the fungus did not substantially affect VB decolorization. Inhibition of laccase production by adding various inhibitors to shaken cultures reduced VB decolorization significantly suggesting a role for laccase in VB decolorization. When sodium azide and aminotriazole were used to inhibit endogenous catalase and cytochrome P-450 oxygenase activities, there was 100% and 70% reduction in VB decolorization, respectively. Adding benzoate to trap hydrogen peroxide-derived hydroxyl radicals resulted in 50% decolorization of VB. Boiling the extracellular fluid (ECF) for 30 min resulted in approximately 50% reduction in VB decolorization. Collectively, these data suggest that laccase, and/or oxygenase/oxidase and a heat-stable non-enzymatic factor, but not Lip and MnP, play a role in VB decolorization by P. chrysosporium.     

Purification and characterization of laccase from <Emphasis Type="Italic">Pycnoporus sanguineus</Emphasis> and decolorization of an anthraquinone dye by the enzyme

The white rot fungus Pycnoporus sanguineus produced high amount of laccase in the basal liquid medium without induction. Laccase was purified using ultrafiltration, anion-exchange chromatography, and gel filtration. The molecular weight of the purified laccase was estimated as 61.4 kDa by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The enzyme oxidized typical substrates of laccases including 2,2′-azino-bis(3-ethylbenzthiazoline-6-sulfonate), 2,6-dimethoxyphenol, and syringaldazine. The optimum pH and temperature for the purified laccase were 3.0 and 65°C, respectively. The enzyme was stable up to 40°C, and high laccase activity was maintained at pH 2.0–5.0. Sodium azide, l-cysteine, and dithiothreitol strongly inhibited the laccase activity. The purified enzyme efficiently decolorized Remazol Brilliant Blue R in the absence of added redox mediators. The high production of P. sanguineus laccase as well as its decolorization ability demonstrated its potential applications in dye decolorization.     

Uptake of amino acids by actidione-treated yeast cells

The active uptake ofl-aspartic acid, glycine andl-lysine by actidione-treated cells ofSaccharomyces cerevisiae was found to be inhibited by anaerobic conditions in the absence of a source of energy, only facilitated diffusion persisting. Similarly, metabolic inhibitors (iodoacetamide, sodium fluoride and potassium sorbate) inhibited the uptake very substantially. 2,4-Dinitrophenol and sodium azide appeared to inhibit the movement of the transport carrier itself, while uranyl ions showed a complex interaction pattern, ranging from inhibition at concentrations of 10^?6–10^?4 m, to stimulation at concentrations of 3×10^?4–10^?3 m, to pronounced inhibition at higher concentrations. The uptake was pH-dependent with optima forl-aspartic acid near pH 4, for glycine near pH 5, forl-lysine near pH 6.5.     

Different nitrogen sources and growth responses of Spirulina platensis in microenvironments

Spirulina platensis was cultivated, in comparative studies, using several sources of nitrogen. The standard source used (sodium nitrate) was the same as that used in the synthetic medium Zarrouk, whereas the alternative nitrogen sources consisted of ammonium nitrate, urea, ammonium chloride, ammonium sulphate or acid ammonium phosphate. The initial nitrogen concentrations tested were 0.01, 0.03 and 0.05 M in an aerated photobioreactor at 30 °C, with an illuminance of 1900 lux, and 12 h-light/12 h-dark photoperiod over a period of 672 h. Maximum biomass was produced in medium containing sodium nitrate (0.01–0.03–0.05 M), followed by ammonium nitrate (0.01 M) and urea (0.01 M). The final biomass concentrations were 1.992 g l^–1 (0.03 M sodium nitrate), 1.628 g l^–1 (0.05 M sodium nitrate), 1.559 g l^–1 (0.01 M sodium nitrate), 0.993 g l^–1 (0.01 M ammonium nitrate) and 0.910 g l^–1 (0.01 M urea). This suggested that it is possible to utilize nitrogen sources other than sodium nitrate for growing S. platensis, in order to decrease the production costs of scaled up projects.     

Role of aquatic vegetation coverage on hypoxia and sunfish abundance in bays of a eutrophic reservoir

We studied the relation between aquatic vegetation coverage, summer dissolved oxygen and density of sunfishes (Lepomis spp.) in 10 shallow bays of a eutrophic reservoir. The bays ranged 5.2–15.7 ha in area, 0.6–1.3 m mean depth and 6–91% vegetation area coverage. Over the 10 bays mean dissolved oxygen concentration ranged 5.8–9.0 mg l^-1 in open water at least 20 m away from the vegetation, 0.8–8.1 mg l^-1 at the vegetation-water edge and 0.5–7.7 mg l^-1 in dense vegetation. Dissolved oxygen concentrations were inversely related to vegetation coverage. In open water, dissolved oxygen levels were consistently higher than 8 mg l^-1 when vegetation coverage was <20% of the bay and decreased to about 6 mg l^-1 at coverages near 80%. At the vegetation-water edge and within dense vegetation, dissolved oxygen levels dropped rapidly as vegetation coverage increased to 20% of the bay; when vegetation reached about 50% coverage, dissolved oxygen remained near 1.5 mg l^-1at the vegetation-water edge, but oxygen dropped below 1 mg l^-1in dense vegetation. Scarce vegetation harbored high Lepomis relative abundance (fish per m² of vegetation) whereas extensive vegetation harbored low relative abundance, both contributing little to absolute abundance (total fish in all vegetation); however, intermediate coverage offered a combination of mid-level fish relative abundance that together with mid-level plant coverage translated into high absolute fish abundance. We suggest this response is related to hypoxia, and where aquatic vegetation is extensive, the effect of vegetation on hypoxia and water quality in general may influence fish populations in a way similar to that often attributed to reduced foraging efficiency and increased competitive interactions.     

The effects of folic acid and nitric oxide synthase inhibition on coronary flow and oxidative stress markers in isolated rat heart

The aim of this study was to assess the effects of folic acid on coronary flow and oxidative stress markers with or without non-specific inhibition of nitric oxide synthase by l-NAME in isolated rat hearts. The hearts of male Wistar albino rats (n = 12, age 8 weeks, body mass 180–200 g) were retrograde perfused according to the Langendorff technique at gradually increased constant perfusion pressure (40–120 cmH₂O). Coronary flow and markers of oxidative stress: nitrite outflow, superoxide anion production, and index of lipid peroxidation (by measuring thiobarbituric acid reactive substances) in coronary effluent were calculated. The experiments were performed during control conditions and in presence of folic acid (100 μM) alone or folic acid (100 μM) plus l-NAME (30 μM). Control values of coronary flow varied in range from 4.37 ± 0.10 ml/min/g wt at 40 cmH₂O to 12.05 ± 0.42 ml/min/g wt at 120 cmH₂O. Nitrite outflow varied from 1.68 ± 0.17 nmol/min/g wt at 40 cmH₂O to 3.56 ± 0.17 nmol/min/g wt at 120 cmH₂O and was parallel with coronary perfusion pressure-coronary flow curve. Folic acid significantly increased coronary flow (40–120 cmH₂O, 5.63 ± 0.10 ml/min/g wt and 15.2 ± 0.42 ml/min/g wt, respectively) and was accompanied by significant increase in nitrite outflow (2.28 ± 0.29 nmol/min/g wt at 40 cmH₂O to 6.66 ± 0.50 nmol/min/g wt at 120 cmH₂O). In addition, folic acid significantly decreased superoxide anion production especially at upper coronary perfusion pressure values (60% at 120 cmH₂O) and increased index of lipid peroxidation (37.16% at 120 cmH₂O), respectively. Folic acid plus l-NAME did not change control values of coronary flow significantly. However, folic acid plus l-NAME increased nitrite outflow especially at upper coronary perfusion pressure values (43.05% at 120 cmH₂O) and did not change significantly superoxide anion production or index of lipid peroxidation versus control values, respectively. The results clearly showed that on isolated rat hearts at gradually increased constant perfusion pressure, folic acid increased coronary flow, increased nitrite outflow, decreased superoxide anion production, and increased index of lipid peroxidation. These effects were reversed or blocked by l-NAME thus demonstrating mediation or at least participation of NO in the mechanism of the folic acid-induced effects.     

Production of a thermostable extracellular lipase by Kluyveromyces marxianus

Kluyveromyces marxianus was grown in submerged culture in a complex medium with several potential inducers of lipolytic activity (triacylglycerols, fatty acids). The highest extracellular lipolytic enzyme production (about 80 U ml^–1 in 3 d) was obtained when the medium was supplemented with 2 g urea l^–1 plus 5 g tributyrin l^–1. Addition of surfactants (1 g l^–1) did not improve production. The lipase had a high thermal stability in aqueous solution (73% residual activity after 9 d at 50 °C, 16 min half-life time at 100 °C). It was also stable at acidic pH and showed good tolerance to organic solvents (70% residual activity after 2 d in n-hexane of cyclohexane).     

Peroxidase from Bacillus sp. VUS and its role in the decolorization of textile dyes

Peroxidase was purified by an ion exchange chromatography followed by gel filtration chromatography from dye degrading Bacillus sp. strain VUS. The optimum pH and temperature of the enzyme activity was 3.0 and 65°C, respectively. This enzyme showed more activity with n-propanol than other substrates tested viz. xylidine, 3-(3,4-dihydroxy phenyl) Lalanine (L-DOPA), hydroxyquinone, ethanol, indole, and veratrole. Km value of the enzyme was 0.076 mM towards n-propanol under standard assay conditions. Peroxidase was more active in presence of the metal ions like Li²⁺, Co²⁺, K²⁺, Zn²⁺, and Cu²⁺ where as it showed less activity in the presence of Ca²⁺ and Mn²⁺. Inhibitors like ethylenediamine tetraacetic acid (EDTA), glutamine, and phenylalanine inhibited the enzyme partially, while sodium azide (NaN₃) completely. The crude as well as the purified peroxidase was able to decolourize different industrial dyes. This enzyme decolourized various textile dyes and enhanced percent decolourization in the presence of redox mediators. Aniline was the most effective redox mediator than other mediators tested. Gas chromatography-Mass spectrometry (GC-MS) confirmed the formation of 7-Acetylamino-4-hydroxy-naphthalene-2-sulphonic acid as the final product of Reactive Orange 16 indicating asymmetric cleavage of the dye.     

Purification and characterization of veratryl alcohol oxidase from Comamonas sp. UVS and its role in decolorization of textile dyes

In the present work, we have purified veratryl alcohol oxidase (VAO) enzyme from Comamonas UVS to evaluate its potential to decolorize textile dyes. VAO was purified (13.9 fold) by an ion exchange followed by the size exclusion chromatography. Molecular weight of the VAO was estimated to be about 66 kDa by SDS-PAGE. The optimum pH and temperature of oxidase were 30°C and 65°C, respectively. VAO showed maximum activity with n-propanol among the various substrates (n-propanol, veratryl alcohol, L-dopa, tryptophan, etc.). Under standard assay conditions, K_m value of the enzyme was 2.5 mM towards veratrole. The enzyme activity was completely inhibited by 0.5 mM sodium azide. L-cysteine, dithiothreitol, and the metal chelator, EDTA had a slight inhibitory effect. The purified enzyme was able to decolorize textile dyes, Red HE7B (57.5%) and Direct Blue GLL (51.09%) within 15 h at 40 μg/mL concentration. GC-MS analysis of the metabolites suggested oxidative cleavage and desulphonation of these dyes.     

Biotransformation of [ring-U-14C]4-n-nonylphenol by Agrostemma githago cell culture in a two-liquid-phase system

The biotransformation of [¹⁴C]4-n-nonylphenol (5 mg l^–1; 10 mg l^–1) by Agrostemma githago cell suspensions was studied using a batch two-liquid-phase system (medium/n-hexadecane 200:1, v/v). The highly lipophilic 4-n-nonylphenol was applied via n-hexadecane phase. After 7 d of incubation, more than 85% of applied 4-n-nonylphenol was absorbed by the cells, and 40% was transformed to 10 side-chain monohydroxylated metabolites (two with additional double bond at side-chain). The primary metabolites were analyzed by GC-EIMS. In the cells, the monohydroxylated products and residual 4-n-nonylphenol were present as glycosides. The method proved to be suitable for the production of primary metabolites of 4-n-nonylphenol on a larger scale for identification purposes and for metabolic profiling of the compound.