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Selected AbstractsInfluence of in-stream diel concentration cycles of dissolved trace metals on acute toxicity to one-year-old cutthroat trout (Oncorhynchus clarki lewisi)ENVIRONMENTAL TOXICOLOGY & CHEMISTRY, Issue 12 2007David A. Nimick Abstract Extrapolating results of laboratory bioassays to streams is difficult, because conditions such as temperature and dissolved metal concentrations can change substantially on diel time scales. Field bioassays conducted for 96 h in two mining-affected streams compared the survival of hatchery-raised, metal-näive westslope cutthroat trout (Oncorhynchus clarki lewisi) exposed to dissolved (0.1-,m filtration) metal concentrations that either exhibited the diel variation observed in streams or were controlled at a constant value. Cadmium and Zn concentrations in these streams increased each night by as much as 61 and 125%, respectively, and decreased a corresponding amount the next day, whereas Cu did not display a diel concentration cycle. In High Ore Creek (40 km south of Helena, MT, USA), survival (33%) after exposure to natural diel-fluctuating Zn concentrations (range, 214,634 ,g/L; mean, 428 ,g/L) was significantly (p = 0.008) higher than survival (14%) after exposure to a controlled, constant Zn concentration (422 ,g/L). Similarly, in Dry Fork Belt Creek (70 km southeast of Great Falls, MT, USA), survival (75%) after exposure to diel-fluctuating Zn concentrations (range, 266,522 ,g/L; mean, 399 ,g/L) was significantly (p = 0.022) higher than survival (50%) in the constant-concentration treatment (392 ,g/L). Survival likely was greater in these diel treatments, both because the periods of lower metal concentrations provided some relief for the fish and because toxicity during periods of higher metal concentrations was lessened by the simultaneous occurrence each night of lower water temperatures, which reduce the rate of metal uptake. Based on the present study, current water-quality criteria appear to be protective for streams with diel concentration cycles of Zn (and, perhaps, Cd) for the hydrologic conditions tested. [source] Fusion of farnesyldiphosphate synthase and epi -aristolochene synthase, a sesquiterpene cyclase involved in capsidiol biosynthesis in Nicotiana tabacumFEBS JOURNAL, Issue 14 2002Maria Brodelius A clone encoding farnesyl diphosphate synthase (FPPS) was obtained by PCR from a cDNA library made from young leaves of Artemisia annua. A cDNA clone encoding the tobacco epi -aristolochene synthase (eAS) was kindly supplied by J. Chappell (University of Kentucky, Lexington, KY, USA). Two fusions were constructed, i.e. FPPS/eAS and eAS/FPPS. The stop codon of the N-terminal enzyme was removed and replaced by a short peptide (Gly-Ser-Gly) to introduce a linker between the two ORFs. These two fusions and the two single cDNA clones were separately introduced into a bacterial expression vector (pET32). Escherichia coli was transformed with the expression vectors and enzymatically active soluble proteins were obtained after induction with isopropyl thio-,- d -thiogalactoside. The recombinant enzymes were purified using immobilized metal affinity chromatography on Co2+ columns. The fusion enzymes produced epi- aristolochene from isopentenyl diphosphate through a coupled reaction. The Km values of FPPS and eAS for isopentenyl diphosphate and farnesyl diphosphate, respectively, were essentially the same for the single and fused enzymes. The bifunctional enzymes showed a more efficient conversion of isopentenyl diphosphate to epi -aristolochene than the corresponding amount of single enzymes. [source] High-dose intravenous paraoxon exposure does not cause organophosphate-induced delayed neuropathy (OPIDN) in mini pigsJOURNAL OF APPLIED TOXICOLOGY, Issue 4 2001G. Petroianu Abstract Organophosphorus compounds are inhibitors of serine hydrolases. Some of these compounds produce, in addition to their high acute toxicity, a more persistent effect: organophosphate-induced delayed neuropathy (OPIDN). The putative molecular entity whose inhibition is thought to be responsible for OPIDN is the neuropathy target esterase (NTE). Although in vitro NTE is resistant to paraoxon (PX), occasional case reports have associated PX with OPIDN. To assess clinically whether or not high-dose i.v. PX causes OPIDN in mini pigs, 14 mini pigs were anaesthesized, intubated and mechanically ventilated. In a first set of experiments eight pigs received 1 mg PX kg,1 body weight (BW) dissolved in alcohol. Two control animals received alcohol in a corresponding amount. After infusion of PX, survival of the animals during the acute phase of intoxication was achieved by intensive-care support, using appropriate drugs and fluids according to a pre-established protocol. The mini pigs were extubated 1036 ± 363 min later (mean ± SD). The pigs were observed prior to PX application and for 6 weeks thereafter for any abnormalities and/or signs of OPIDN, such as leg weakness, ataxia and paralysis. Observations were graded on a scale for three categories (position, motor deficiency, reaction), with a maximal cumulative score of 9. In a second set of experiments (four additional pigs) larger PX doses were used (3, 9, 27 and 81 mg kg,1 BW). After recovering from general anaesthesia/surgery, within 2 weeks all animals reached the initial score on the scale. It can be concluded that high-dose i.v. PX exposure does not induce OPIDN in mini pigs during the 6-week observation period. Copyright © 2001 John Wiley & Sons, Ltd. [source] Direct analysis of single leaf disks for chemopreventive glucosinolatesPHYTOCHEMICAL ANALYSIS, Issue 3 2002Qiaomei Wang Abstract Natural isothiocyanates, produced during plant tissue damage from methionine-derived glucosinolates, are potent inducers of mammalian phase 2 detoxification enzymes such as quinone reductase (QR). A greatly simplified bioassay for glucosinolates based on induction and colorimetric detection of QR activity in murine hepatoma cells is described. It is demonstrated that excised leaf disks of Arabidopsis thaliana (ecotype Columbia) can directly and reproducibly substitute for cell-free leaf extracts as inducers of murine QR, which reduces sample preparation to a minimum and maximizes throughput. A comparison of 1 and 3,mm diameter leaf disks indicated that QR inducer potency was proportional to disk circumference (extent of tissue damage) rather than to area. When compared to the QR inducer potency of the corresponding amount of extract, 1,mm leaf disks were equally effective, whereas 3,mm disks were 70% as potent. The QR inducer potency of leaf disks correlated positively with the content of methionine-derived glucosinolates, as shown by the analysis of wild-type plants and mutant lines with lower or higher glucosinolate content. Thus, the microtitre plate-based assay of single leaf disks provides a robust and inexpensive visual method for rapidly screening large numbers of plants in mapping populations or mutant collections and may be applicable to other glucosinolate-producing species. Copyright © 2002 John Wiley & Sons, Ltd. [source] Use of a plant-derived enzyme template for the production of the green-note volatile hexanalBIOTECHNOLOGY & BIOENGINEERING, Issue 3 2003Frank Schade Abstract Hexanal is a key organoleptic element of green-note that is found in both fragrances and flavors. We report a novel process for the production of hexanal using immobilized enzyme templates extracted from different plant sources in combination with hollow-fiber ultrafiltration for in situ separation. Enzyme templates, known to be responsible for the synthesis of hexanal from linoleic acid (18:2), were isolated from naturally enriched tissues including carnation petals, strawberry and tomato leaves. These templates were immobilized in an alginate matrix and used as a biocatalyst in a packed-bed bioreactor. Continuous product recovery was achieved using a hollow-fiber ultrafiltration unit. The effects of pH, reaction temperature, and substrate and enzyme concentrations were studied and their effects on hexanal generation identified and optimized. Utilizing optimized conditions, hexanal production 112-fold higher than endogenous steady-state levels in a corresponding amount of plant tissue could be achieved over a 30-minute period. Based on the reactor studies, product inhibition also appears to be an important factor for bioreactor-based hexanal production. © 2003 Wiley Periodicals, Inc. Biotechnol Bioeng 84: 265,273, 2003. [source] | |||||||||||||