Bioaccumulation of melamine in catfish muscle following continuous, low-dose, oral administration

In this study, catfish muscle was analyzed for melamine (MEL) and cyanuric acid (CYA) residues following experimental feeding with low doses of MEL and MEL and CYA (MEL+CYA) and with the insoluble melamine-cyanurate complex (MEL=CYA). Catfish were daily fed 0.1 mg/kg BW of MEL for 15, 28, or 42 days, 0.1 mg/kg BW of MEL+CYA for 28 days, 2.5 mg/kg BW of MEL+CYA for 14 days, or 400 mg/kg BW of MEL=CYA for 3 days. Residues in the tissue were determined by LC-MS/MS. MEL was extracted with acidic acetonitrile, followed by defatting with dichloromethane, and isolated with cation exchange solid phase extraction (SPE). For CYA analysis, fish were extracted with dilute acetic acid, defatted with hexane, and cleaned up with a graphitic carbon SPE. Catfish fed 0.1 mg/kg BW of MEL reached a maximum muscle residue concentration of 0.33 ± 0.04 mg/kg (ppm) after 28 days of continuous feeding. The same concentration was found for MEL+CYA feeding at the 0.1 mg/kg BW level for 28 days. Feeding at 2.5 mg/kg BW of MEL+CYA yielded muscle concentrations above the 2.5 mg/kg level of concern for most of the study fish. Finally, catfish fed high levels of the MEL=CYA complex (400 mg/kg BW) accumulated relatively little MEL in the muscle (0.14 ± 0.07 mg/kg) and, unlike treatment with MEL+CYA, did not form renal melamine-cyanurate crystals. Appreciable concentrations of CYA were not detected in any of the muscles tested. These studies provide data to model the bioaccumulation of triazine residues into edible fish tissue as a result of the continuous consumption of adulterated feed.     

Evaluation of hematology and serum biochemistry of cold-stunned green sea turtles (Chelonia mydas) in North Carolina, U.S.A

Hypothermia or cold-stunning is a condition in which the body temperature of an animal decreases below normal physiologic range and which has been linked to severe morbidity in sea turtles. Reports have focused on the physiologic changes caused by cold-stunning in Kemp's Ridley sea turtles (Lepidochelys kempii) and loggerhead sea turtles (Caretta caretta), but few have evaluated the green sea turtle (Chelonia mydas). This study evaluated hematologic and serum biochemical profiles of cold-stunned green sea turtles in North Carolina, USA. When compared with healthy, free-ranging juvenile green turtles from the same region, cold-stunned turtles exhibited hypoglycemia, hypocalcemia (both total and ionized calcium), hyponatremia, hypokalemia, hypoproteinemia, hypoalbuminemia, hyperphosphatemia, and elevations in uric acid and blood urea nitrogen. These findings contrast with some previously reported changes in cold-stunned Kemp's Ridley and loggerhead sea turtles. These results emphasize the importance of basing therapeutic regimens on biochemical analyses in cold-stunned sea turtles.     

Resveratrol in raw and baked blueberries and bilberries

Resveratrol in the fruits of bilberry (Vaccinium myrtillus L.), the lowbush "wild" blueberry (Vaccinium angustifolium Aiton), the rabbiteye blueberry (Vaccinium ashei Reade), and the highbush blueberry (Vaccinium corymbosum L.) were measured using a new assay based on high-performance liquid chromatography-tandem mass spectrometry (LC-MS/MS). The LC-MS/MS assay provided lower limits of detection than previous methods for resveratrol measurement, 90 fmol of trans-resveratrol injected on-column, and a linear standard curve spanning >3 orders of magnitude. The recoveries of resveratrol from blueberries spiked with 1.8, 3.6, or 36 ng/g were 91.5 +/- 4.5, 95.6 +/- 6.5, and 88.0 +/- 3.6%, respectively. trans-Resveratrol but not cis-resveratrol was detected in both blueberry and bilberry samples. The highest levels of trans-resvertatrol in these specimens were 140.0 +/- 29.9 pmol/g in highbush blueberries from Michigan and 71.0 +/- 15.0 pmol/g in bilberries from Poland. However, considerable regional variation was observed; highbush blueberries from British Columbia contained no detectable resveratrol. Because blueberries and bilberries are often consumed after cooking, the effect of baking on resveratrol content was investigated. After 18 min of heating at 190 degrees C, between 17 and 46% of the resveratrol had degraded in the various Vaccinium species. Therefore, the resveratrol content of baked or heat-processed blueberries or bilberries should be expected to be lower than in the raw fruit. Although blueberries and bilberries were found to contain resveratrol, the level of this chemoprotective compound in these fruits was <10% that reported for grapes. Furthermore, cooking or heat processing of these berries will contribute to the degradation of resveratrol.     

Tailor made plasmin substrates as potential diagnostic tool to test for mastitis

Plasmin is a protease in milk, which concentration is increased during mastitis. The aim of the study was design and characterize short, tailor made fluorogenic substrates for plasmin to see differences in plasmin activity in healthy milk compared to mastitic milk. According to the specificity matrix of plasmin, containing 125 currently known plasmin substrates, six novel plasmin specific substrates were designed and characterized. Eight other substrates were insensitive for plasmin, which was in line with our expectations. The plasmin-sensitive substrates showed no cross-reactivity with proteases from mastitogenic bacteria in vitro. A proof of concept to distinguish healthy milk from mastitic milk is described. Potentially, these substrates present a rapid alternative to the currently established methodologies to detect mastitis in milk.     

Avian whole‐brain spheroid cultures: applications in pesticide toxicity

The adult hen is the species of choice when assessing the potential of organophosphate compounds (OPs) to evoke OP‐induced delayed neuropathy (OPIDN). OPIDN has been shown to depend on the irreversible covalent binding of certain OPs to the enzyme neuropathy target esterase (NTE). Levels of this enzyme in human brain are similar to those in avian species. We have previously shown that the hen embryo whole brain spheroids are histotypically similar to adult brain and express measurable levels of acetylcholine esterase (AChE) and NTE, the key enzymes in mechanisms of OP toxicity. Single‐cell suspensions were prepared from meninges‐free 7 day in ovo hen brains. Cells were grown in serum‐free media and incubated on an orbital shaker at 37 °C in 5% carbon dioxide humidified air. Spheroids can be maintained long‐term in culture and allow the effects of repeated dose and recovery studies to be conducted. Hen embryo brain spheroids were exposed to varying concentrations of tri‐o‐cresyl phosphate, leptophos and its metabolite leptophos oxon at day 14 in vitro and the effects on the activity of AChE and NTE measured. Whilst current legislation regarding toxicity testing of OPs requires the use of animals, our in vitro model provides a potential pre‐screen for novel chemical entities and commercial OP mixture variants. Such an approach should refine and reduce the number of animals used in development of new materials. © 2000 Society of Chemical Industry     

Omphalophlebitis and osteomyelitis associated with Klebsiella septicemia in a premature foal

Gram-negative neonatal septicemia was diagnosed in a premature Standardbred colt. Clinical signs included lethargy, weakness, loss of suckle reflex, tachypnea, and injected mucous membranes. Sequelae included pneumonia, omphalophlebitis, septic arthritis, and osteomyelitis. Prepartum maternal uterine infection, premature delivery, abnormal parturition with premature fetal membrane separation, and failure of passive transfer of colostral immunoglobulins increased the foal's risk for developing sepsis. Treatment included administration of moxalactam disodium and cefadroxil. The clinical efficacy of cephalosporin antibiotics in the treatment of gram-negative sepsis is discussed.