Qualitative and quantitative drug residue analyses: Chlortetracycline in white-tailed deer (Odocoileus virginianus) and supermarket meat by liquid chromatography tandem-mass spectrometry.

Chlortetracycline is (CTC) is a tetracycline antibiotic which is being in the white-tailed deer industry to improve production and animal health. In this paper, we present a method for determining chlortetracycline residues in edible white-tailed deer tissues, using liquid chromatography with heated electrospray ionization and mass spectrometry detection. The procedure involved extraction with EDTA-McIlvaine buffer at pH 4.0, followed by solid-phase extraction cleanup using a hydrophilic-lipophilic balance (HLB) cartridge. The liquid chromatography analysis was performed with heated electrospray ionization and mass spectrometry detection. The limit of quantification for the method was 2.7 ng/g and limit of detection was 0.8 ng/g. The recovery values were >78.5% for muscle, 65.1% for kidney, 63.1% for liver. Mean tissue residue concentration of chlortetracycline and it's epimer, 4-epi chlortetracycline (4-epi-CTC) at 10-day withdrawal period for kidney, liver, muscle was 122.8, 44.7 and 26.7 ng/g, respectively. Chlortetracycline tissue residue concentration at 45-day withdrawal period for kidney, liver, muscle was 19.2, 28.9 and 10.7 ng/g, respectively. Mean tissue concentration of CTC was less than the established maximum residual limit (MRL) values for bovine tissues. We have validated and successfully applied this method in the qualitative and quantification of chlortetracycline in white-tailed deer tissue samples.

Qualitative and Quantitative Drug residue analyses: Florfenicol in white-tailed deer (Odocoileus virginianus) and supermarket meat by liquid chromatography tandem-mass spectrometry.

A method for confirmation and detection of Florfenicol amine residues in white-tailed deer tissues was developed and validated in our laboratory. Tissue samples were extracted with ethyl acetate and cleaned up on sorbent (Chem-elut) cartridges. Liguid chromatography (LC) separation was achieved on a Zorbax Eclipse plus C18 column with gradient elution using a mobile phase composed of ammonium acetate in water and methanol at a flow rate of 300µL/min. Qualitative and quantitative analyses were carried out using liquid chromatography - heated electrospray ionization(HESI) and atmospheric pressure chemical ionization (APCI)-tandem mass spectrometry in the multiple reaction monitoring (MRM) interface. The limits of detection (LODs) for HESI and APCI probe were 1.8ng/g and 1.4ng/g respectively. Limits of quantitation (LOQs) for HESI and APCI probe were 5.8ng/g and 3.4ng/g respectively. Mean recovery values ranged from 79% to 111% for APCI and 30% to 60% for HESI. The validated method was used to determine white-tailed deer florfenicol tissue residue concentration 10-days after exposure. Florfenicol tissue residues concentration ranged from 0.4 to 0.6µg/g for liver and 0.02-0.05µg/g for muscle and a trace in blood samples. The concentration found in the tested edible tissues were lower than the maximum residual limit (MRL) values established by the federal drug administration (FDA) for bovine tissues. In summary, the resulting optimization procedures using the sensitivity of HESI and APCI probes in the determination of florfenicol in white-tailed deer tissue are the most compelling conclusions in this study, to the extent that we have applied this method in the evaluation of supermarket samples drug residue levels as a proof of principle.

Arsenate and perchlorate toxicity, growth effects, and thyroid histopathology in hypothyroid zebrafish Danio rerio.

Exposure to perchlorate or other thyrotoxic compounds can cause hypothyroidism in most vertebrates, and this may affect levels of endogenous antioxidants and cause oxidative stress. Arsenic also induces oxidative stress in animals by modifying the antioxidant capacity and may alter the thyroid homeostasis. Therefore, hypothyroidism may affect the toxicity of arsenate. In order to test this hypothesis, zebrafish (Danio rerio) were made hypothyroid by exposure to perchlorate, and toxicity of arsenate in hypothyroid and euthyroid fish was compared. The endpoints were LC50 and thyroid histopathology. Additionally, the recovery of thyroid histopathological indices after cessation of perchlorate exposure was determined. The current study showed that 96 h LC50 of perchlorate anion and arsenate ion to juveniles fish (37 day post-fertilization) were 2532 and 56 mg l(-1), respectively. In addition, hypothyroid fish were more sensitive to arsenate, with a 96 h LC50 of 43 mg l(-1). Growth rates were also significantly retarded by perchlorate exposure. After cessation of perchlorate exposure, there was recovery of thyroid histopathology in terms of epithelial cell height, but not colloid area or growth rate. In conclusion, perchlorate enhances arsenate toxicity to juvenile zebrafish, and the rate of thyroid recovery after cessation of perchlorate exposure depends on the endpoints examined.

Age-dependent characterization of pendrin gene expression in various tissues of deer mice.

Pendrin is a membrane transport protein which functions as the transporter of chloride, bicarbonate, formate, and iodide. In this study, we characterized pendrin gene expression in various tissues of deer mice (Peromyscus maniculatus), a sentinel wildlife species. Deer mice were euthanized at post-natal day (PND) 21 (day of weaning) and PND 45 (24 days post-weaning) for tissue collection. A deer mouse-specific partial pendrin cDNA sequence was generated, from which Taqman-specific probe and primers were designed for quantification of mRNA equivalents of pendrin gene expression using real-time polymerase chain reaction (PCR). The expression profile was standardized to glyceraldehyde-3-phosphate dehydrogenase (GAPDH). Results indicate that the pendrin gene was expressed at different levels in the different tissues of developing deer mice relative to GAPDH expression. Expression in the tissues was determined to be age-dependent. Pendrin gene was highly expressed in the kidney, lungs and reproductive tissues. PND 21 expression in the kidney and testes was significantly lower than PND 45. This study represents the first identification of differential expression of pendrin gene in various deer mouse tissues.

Reproductive effects of hexahydro-1,3,5-trinitroso-1,3,5-triazine in deer mice (Peromyscus maniculatus) during a controlled exposure study.

Contamination with hexahydro-1,3,5-trinitro-1,3,5-triazine (Royal Demolition Explosive [RDX]) has been identified at areas of explosive manufacturing, processing, storage, and usage. Thus, the potential exists for exposure to N-nitroso compounds, hexahydro-1-nitroso-3,5-dinitro-1,3,5-triazine, hexahydro-1,3-dinitroso-5-nitro-1,3,5-triazine, and hexahydro-1,3,5-trinitroso-1,3,5-triazine (TNX), formed via anaerobic transformation of RDX. Following exposure, reproductive toxicity of TNX was evaluated in three consecutive litters of deer mice (Peromyscus maniculatus). Hexahydro-1,3,5-trinitroso-1,3,5-triazine was administered ad libitum via drinking water at four doses: 0 (control), 1, 10, and 100 microg/L. Endpoints investigated included reproductive success, offspring survival, offspring weight gain, offspring organ weights, and liver TNX residues. Data from the present study indicate that TNX bioaccumulates in the liver and is associated with postpartum mortality, dose-dependent decrease in body weight from birth to weaning, and decrease in kidney weight of deer mice offspring.

Evaluation of adult quail and egg production following exposure to perchlorate-treated water.

Twenty-three adult female northern bobwhite (Colinus virginianus) quail were exposed to 0, 0.01, 0.1, and 1 mM ammonium perchlorate (AP) in drinking water for 30 d. Eggs laid in all treatment groups, including control, were collected, dated, given an identification number, and weighed. On day 30 of exposure, 10 birds were euthanized by carbon dioxide asphyxiation. Gross toxicological endpoints and thyroid histology were evaluated in 10 birds. Egg production and accumulation of perchlorate in the eggs (n = 10) and liver (n = 5) were determined. Perchlorate did not affect body or organ weights significantly; however, at 1 mM, AP caused alteration of thyroid gland morphology. Perchlorate did not affect egg production, but significant accumulation was observed in the eggs and livers of exposed birds.