Validation of the in vitro gastrointestinal (IVG) method to estimate relative bioavailable lead in contaminated soils.

The effect of the dosing vehicle (e.g., dough) on the ability of an in vitro gastrointestinal (IVG) method to predict relative bioavailable Pb associated with soil ingestion was evaluated. Bioaccessible Pb determined by the IVG method was compared with relative bioavailable Pb measured from dosing trials using juvenile swine for 18 contaminated soils ranging from 1270 to 14200 mg Pb kg(-1). Bioaccessible Pb was measured in the IVG gastric extraction (GE) and intestinal extraction (IE) solutions. Mean bioaccessible Pb values were 32.2% for GE without dough, 23.0% for GE with dough, 1.06% for IE without dough, and 0.56% for IE with dough. It is possible that phytic acid associated with the dough addition decreased bioaccessible Pb. In vivo relative bioavailable Pb ranges for different swine tissues were 1 to 87% for blood, 0 to 110% for liver, 1 to 124% for kidney, and 0.04 to 94% for bone. Strong linear relationships between IVG GE Pb with dough (r > 0.76, P < 0.0002), IVG IE Pb with dough (r > 0.56, P < 0.015), and IVG GE Pb without dough (r > 0.81, P < 0.0001) and in vivo bioavailable Pb as estimated with blood, kidney, liver, and bone were found. Inexpensive in vitro methods may be useful in providing an estimate of the variability in relative bioavailable Pb at a single study site. The IVG method can be used to estimate relative bioavailable Pb, As, and Cd in contaminated soil.

Chemical extraction methods to assess bioavailable arsenic in soil and solid media.

Soil ingestion by children is an important pathway in assessing public health risks associated with exposure to arsenic-contaminated soils. Soil chemical methods are available to extract various pools of soil arsenic, but their ability to measure bioavailable arsenic from soil ingestion is unknown. Arsenic extracted by five commonly used soil extractants was compared with bioavailable arsenic measured in vivo by immature swine (Sus scrofa) dosing trials. Fifteen contaminated soils that contained 233 to 17 500 mg kg(-1) arsenic were studied. Soil extractants were selected to dissolve surficially adsorbed and/or readily soluble arsenic (water, 1 M sodium acetate, 0.1 M Na2HPO4/0.1 M NaH2PO4) and arsenic in Fe and Mn oxide minerals (hydroxylamine hydrochloride, ammonium oxalate). The mean percent of total arsenic extracted was: ammonium oxalate (53.6%) > or = hydroxylamine hydrochloride (51.7%) > phosphate (10.5%), acetate (7.16%) > water (0.15%). The strongest relationship between arsenic determined by soil chemical extraction and in vivo bioavailable arsenic was found for hydroxylamine hydrochloride extractant (r = 0.88, significant at the 0.01 probability level). Comparison of the amount of arsenic extracted by soil methods with bioavailable arsenic showed the following trend: ammonium oxalate, hydroxylamine hydrochloride > in vivo > phosphate, acetate > water. The amount of arsenic dissolved in the stomach (potentially bioavailable) is between surficially adsorbed (extracted by phosphate or acetate) and surficially adsorbed + nonsurficial forms in Fe and Mn oxides (extracted by hydroxylamine hydrochloride or ammonium oxalate). Soil extraction methods that dissolve some of the amorphous Fe, such as hydroxylamine hydrochloride, can be designed to provide closer estimates of bioavailable arsenic.

In vitro lead bioaccessibility and phosphate leaching as affected by surface application of phosphoric acid in lead-contaminated soil.

Phosphate treatment of lead-contaminated soil may be a cost-effective remedial alternative for in situ stabilizing soil Pb and reducing Pb toxicology to human. The leaching behaviors of the P added to soil surface and the effect on subsurface Pb bioaccessibility must be addressed for this remedial technology to be acceptable. A smelter-contaminated soil containing an average of 2,670 mg Pb kg(-1), collected from the Jasper County Superfund Site located in Jasper County, Missouri, was surface treated with 10 g P kg(-1) as phosphoric acid (H(3)PO(4)). Following a simulated column leaching and 90-day treatment of field plots, respectively, bioaccessible Pb, P, and pH in soil profile were measured. Surface treatment using H(3)PO(4) effectively stabilized soil Pb and reduced leachable Pb and the bioaccessibility. Phosphate leached into deeper profile significantly lowered bioaccessible Pb in subsurface. Reduction of Pb bioaccessibility increased as a linear function of increasing soil P. Although surface H(3)PO(4) treatment resulted in an enhanced leaching of added P and may increase potential risk of surface and groundwater pollution, the P leaching under field conditions is very limited. Lime addition following the treatment may reduce the leachability of added P and further immobilize soil Pb.

Validation of X-ray fluorescence-measured Swine femur lead against atomic absorption spectrometry.

The aim of this study was to apply the technique of (109)Cd-based K-shell X-ray fluorescence (XRF) bone lead measurements to swine femurs and to validate the concentrations obtained therefrom against an independent chemical measurement of bone lead: atomic absorption spectrometry (AAS). The femurs ranged in lead concentration from 1.0 to 24.5 microg of lead per gram of ashed bone, as measured by AAS. On average, XRF overestimated AAS-measured femur lead by 2.6 microg/g [95% confidence interval (CI), 1.1-4.0 microg/g], approximately 2 microg/g poorer than that observed in studies of human tibiae. Measurements of swine femur and, by extension, of nonhuman bones may require adjustment of the XRF spectrum peak extraction method.

Lead immobilization using phosphoric acid in a smelter-contaminated urban soil.

Transformation of soil lead (Pb) to pyromorphite, a lead phosphate, may be a cost-effective remedial strategy for immobilizing soil Pb and reducing Pb bioavailability. Soil treatment using phosphoric acid (H3PO4) was assessed for its efficacy to reduce Pb solubility and bioaccessibility. Soil containing 4,360 mg of Pb kg(-1), collected from a smelter-contaminated site in Joplin, MO, was reacted with 1,250, 2,500, 5,000, and 10,000 mg of P kg(-1) as H3PO4. The reaction was followed by measurements of Pb bioaccessibility, solubility products, and microprobe analyses. Soluble Pb concentration in the soil decreased with increasing H3PO4 addition. Adding 10,000 mg of P kg(-1) reduced bioaccessible Pb by 60%. The logarithm of bioaccessible Pb decreased as a linear function of increasing H3PO4 addition with an R2 of 0.989. A higher soil/solution ratio was required to extract bioaccessible Pb after the treatment. Microprobe analyses showed that the Pb particles contained P and Cl after the reaction, and the spectra generated by the wavelength-dispersive spectrometer were similar to those of synthetic chloropyromorphite. Lead solubility in the P-treated soil was less than predicted for hydroxypyromorphite [Pbs(PO4)3-OH] and greater than predicted for chloropyromorphite [Pbs(PO4)3Cl]. The P treatment caused approximately 23% redistribution of soil Pb from the clay and silt size fractions to the sand fraction. Soil treatment with H3PO4 resulted in the formation of a compound similar to chloropyromorphite and reduced bioaccessibility of soil Pb, which may have a potential as an in situ technique for Pb-contaminated soil remediation.

Sensitivity and specificity of serum copper determination for detection of copper deficiency in feeder calves.

OBJECTIVE: To determine the relationship between serum and liver copper concentrations and evaluate serum copper determination for diagnosis of copper deficiency in juvenile beef calves. DESIGN: Cross-sectional study. ANIMALS: 105 juvenile beef calves. PROCEDURE: Copper concentrations were measured in paired liver and serum samples from 6- to 9-month-old beef calves. Regression models that predicted liver copper concentration as a function of serum copper concentration were developed. Sensitivity and specificity of serum copper concentration for detection of low liver copper concentration were determined, using a range of serum copper concentrations as test endpoints. Positive and negative predictive values were calculated. RESULTS: The association between serum and liver copper concentrations was significant; however, regression models accounted for only a small portion of the variation in liver copper concentrations. For a serum copper concentration endpoint of 0.45 microg/g, sensitivity and specificity for detection of low liver copper concentration were 0.53 and 0.89, respectively. Positive and negative predictive values of serum copper concentration for detection of low liver copper concentration ranged from 0.37 to 0.85 and 0.63 to 0.94, respectively. CONCLUSIONS AND CLINICAL RELEVANCE: Regression models are inappropriate for predicting copper status as a function of serum copper concentration. Serum copper concentration is fairly specific for detection of low liver copper concentration but only marginally sensitive when serum copper concentration of 0.45 microg/g is used as a test endpoint. The value of serum copper concentration as a diagnostic indicator depends on prevalence of copper deficiency.

Anatomic and physiologic reference values in least shrews (Cryptotis parva).

BACKGROUND AND PURPOSE: The least shrew is an established animal model for reproductive and pharmacologic research. Biologic reference data are needed to assess animal health status and provide a rationale for use of novel statistical programs to evaluate the effects of orally administered substances in toxicologic and pharmacologic studies. METHODS: Organ weights, blood biochemical and hematologic values, and food and water consumption data were collected from 50-day-old shrews after two weeks' consumption of a standard feline diet. RESULTS: In general, data correlated well with values reported for other mammalian species. Plasma phosphorus concentration was high. There was a significant difference in food and water consumption per gram of body weight between shrews at lower and upper (+/- 1 SD) weight ranges for the study. The 3.2-g animals consumed 27% more food per gram of body weight than did the 5.0-g animals. CONCLUSIONS: The high phosphorus concentration was attributed to hemolysis resulting from the axillary cut method of blood sample collection. The small size of the shrew allowed demonstration of the Kleiber effect within a +/- 1 SD weight range in a single species. The phenomenon necessitates the use of statistical methods other than the typical tests establishing the significance of the differences between the means of groups for oral toxicologic and pharmacologic studies.

Metal toxicosis in horses.

The ubiquity and stability of metals in the environment make them unique as a pollutant or an essential dietary component. Metals are neither created nor destroyed by chemical processes but are redistributed in the environment. In combination with other elements, metal compounds and alloys are essential materials of the contemporary world. Inappropriate use or distribution in the environment leads to adverse health effects on all biologic systems, including horses. Gastrointestinal upset is a common feature of acute toxicosis with metals in general. Among the metals discussed, arsenic and inorganic mercury have a propensity to do severe damage to the gut. Deposition of cadmium on forage is the source most likely to intoxicate horses. This subchronic to chronic problem in horses is manifest as disease of the musculoskeletal system and kidneys. Iron-containing hematinics are widely used in racetrack horses and occasionally result in hepatopathy when excessive doses are administered. Lead continues to be recognized as the most significant environmental metal pollutant. Poisoning is encountered routinely in humans and animals. Of the animal species of veterinary concern, lead-poisoned horses are not a frequent encounter. Lead-intoxicated horses show signs of peripheral neuropathy (laryngeal hemiplegia), intermittent colic, and mild anemia. Acute mercury poisoning sometimes occurs from the common use of mercury-containing blistering agents, with most clinical findings related to acute renal failure. Chronic excessive intake of zinc by horses is uncommon but devastating in rapidly growing foals. The mechanism of chronic zinc toxicosis is coupled to the induced copper deficiency. The condition is a disease of cartilage in the articular and growth physes.

Subclinical copper accumulation in llamas.

A 9-year-old, intact male llama with mild ataxia and generalized malaise of 1 month's duration was euthanized following clinical evaluation. Excessive liver copper concentrations were found in the llama and also in clinically normal herdmates. This case documents multiple animals with increased hepatic stores from standard diets and mineral supplements.

Developmental toxicity of purified fumonisin B1 in pregnant Syrian hamsters.

The purpose of this study was to characterize the developmental toxicity of fumonisin B1 (FB1), a mycotoxin produced by Fusarium moniliforme, on fetal Syrian hamsters. Fusarium moniliforme has been associated with a variety of diseases in animals and esophageal cancer in humans. Purified FB1 causes leukoencephalomalacia in horses and is hepatocarcinogenic in rats. Fumonisin B1 has been associated with fetal toxicity in rats and mice and has been suggested to be involved in reproductive failure in pregnant sows. Results from a preliminary developmental toxicity study using an aqueous extract of F. moniliforme corn-culture material in hamsters suggested that FB1 was a developmental toxicant. These results were verified using purified FB1. Six groups of ten time-mated female Syrian hamsters were dosed with 0.0-18 mg kg(-1) day(-1) of FB1 by gavage on days 8-12 of gestation and euthanized on day 15. Live fetuses were weighed and examined for gross external and internal abnormalities and skeletal anomalies. Purified fumonisin B1 was shown to cause dose-dependent fetal death and delayed fetal development without causing fetal abnormalities.

Dietary Fusarium moniliforme culture material induces in vitro tumor necrosis factor-alpha like activity in the sera of swine.

Sera obtained from a group of pigs (n = 5) fed a diet amended with fumonisin containing Fusarium moniliforme culture material was used to determine the levels of Tumor Necrosis Factor-Alpha (TNF) activity by a functional bioassay utilizing the TNF sensitive WEHI 140 mouse fibrosarcoma cell line. Two pigs developed signs consistent with pulmonary edema which was confirmed by pathologic examination in only one pig. Significant, time dependent increases in TNF-like activity were observed in all pigs during the five days of the trial. Another group of pigs (n = 5) was given a defined daily dose of the same culture material by gastric intubation. Two pigs developed fulminant pulmonary edema and sharp increases in TNF activity were observed during the 3 days of the trial in all pigs. In both cases the activity was not abrogated by addition of a neutralizing anti-human TNF monoclonal antibody suggesting that other factors may have been responsible for these effects, possibly the increased levels of sphingoid bases in the serum. Since the pig has become an important model in the study of TNF mediated endotoxic shock, these studies illustrate the relevance of certifying the absence of this important mycotoxin from corn based animal diets, specially if functional assays are used to monitor the activity of TNF in serum.

Bioavailability of lead to juvenile swine dosed with soil from the Smuggler Mountain NPL Site of Aspen, Colorado.

Bioavailability of lead (Pb) has become an issue in quantifying exposure of sensitive populations and, where necessary, establishing cleanup levels for contaminated soil. Immature swine were used as a model for young children to estimate the degree to which Pb from two fully characterized composite samples from the Smuggler Mountain Superfund Site in Aspen, Colorado may be bioavailable to resident children. The composite soils contained 14,200 and 3870 micrograms Pb/g of soil. Relative and absolute enteric bioavailabilities of Pb in soil (oral dose groups of 75,225, and 675 micrograms Pb/kg body wt/day) were estimated by comparison with an orally administered soluble Pb salt (lead acetate = PbAc2.3H2O) (dose groups of 0, 75, and 225 micrograms Pb/kg body wt/day) and an intravenously administered aqueous solution of Pb (100 micrograms Pb/kg/ day) from the same trihydrate salt administered daily for 15 days to 50 juvenile swine. The biological responses (area under the blood Pb concentration-time curve, and the terminal liver-, kidney-, and bone-lead concentrations) produced by Pb from PbAc2.3H2O and lead-contaminated soils were determined. This study revealed Pb from soil containing 14,200 micrograms Pb/g of soil had a bioavailability relative to Pb from PbAc (RBA), ranging from 56% based on the area under the blood lead concentration-time curve (AUC) versus dose, to 86% based on calculations from liver-Pb loading versus dose. Similarly, Pb from soil containing 3870 micrograms Pb/g of soil had an RBA ranging from 58% based on the AUC versus dose, to 74% based on calculations from liver- and kidney-Pb loading versus dose. Bioavailability of Pb in soils may be more or less than EPA's default RBA of 60%, therefore, measuring site-specific RBAs provides a basis for improved exposure and risk assessment.

Acute arsenic toxicosis in five horses.

Five adult horses presented with acute clinical signs of watery diarrhea, excessive salivation, muscle tremors, ataxia, and depression. Four died within 24 hours and the fifth was euthanatized approximately 48 hours after onset of clinical signs. Necropsy finds in two of the horses included hyperemia of gastric mucosa, intestines filled with green to black watery fluid, and multifocal to coalescing, hemorrhagic 1.0-2.0-cm-diameter ulcers of the mucosa of the cecum and large colon. Histopathologic changes in the cecum and large colon consisted of mucosal necrosis and ulceration, vascular thrombosis, necrosis of submucosal blood vessels, and infiltration by mixed mononuclear inflammatory cells and neutrophils. Arsenic toxicosis was suspected. The owner had not been feeding the horses any grain; however, a mixture of grain and pink powder was found in the pasture. Liver arsenic concentrations in the two horses were 14.0 and 11.0 ppm, a sample of renal cortex contained 108 ppm arsenic, and the grain/powder mixture found in the pasture was positive for arsenic at > 3,000 ppm. kidney lead concentrations were 6.5 and 4.2 ppm. Results were consistent with lead arsenate or lead arsenite poisoning.

Use of orally administered succimer (meso-2,3-dimercaptosuccinic acid) for treatment of lead poisoning in dogs.

OBJECTIVE: To determine whether succimer (meso-2,3-dimercaptosuccinic acid) would be effective in reducing blood lead concentration in dogs with naturally acquired lead poisoning and whether treated dogs would develop clinically important adverse effects. DESIGN: Prospective case series. ANIMALS: 13 dogs with moderate-to-high blood lead concentrations (39 to 120 micrograms/dl) and clinical signs of lead poisoning. PROCEDURE: Dogs were treated with succimer (10 mg/kg of body weight, PO, q 8 h) for 10 days. Blood and urine samples were analyzed for lead concentration before, during, and after treatment with succimer. RESULTS: Mean blood lead concentrations on days 0, 3, 7, and 20 were 70.6, 32.7, 16.8, and 27.6 micrograms/dl, respectively. Mean blood lead concentrations decreased 53.6, 76.2, and 60.9% from pretreatment value on days 3, 7, and 20, respectively. Mean urine lead concentrations on days 0, 3, 7, and 20 were 70.0, 485.4, 254.3, and 28.3 micrograms/dl, respectively. CLINICAL IMPLICATIONS: In dogs with naturally acquired lead poisoning, succimer administered orally for 10 days effectively reduced blood lead concentrations and eliminated clinical signs of lead poisoning.

Developmental effects of fumonisin B1 in mice.

Developmental and toxic effects of aqueous extracts of F. moniliforme culture material containing known levels of fumonisin B1 were recently reported in mice and included maternal hepatotoxicity and lethality, maternal body weight gain reduction, increased embryonic resorptions, reduced offspring body weights, and fetal malformations including cleft palate, hydrocephalus, malformed ribs and incomplete digital and sternal ossification. These studies also suggested that the effects of the fungal extract on the mouse offspring may be mediated via maternal effects. The contribution of fumonisin B1 (FB1), a major toxic metabolite of F. moniliforme, in the induction of these effects was evaluated in this study by administering 0 to 100 mg pure FB1/kg of body weight on gestational days (GD) 7 through 15 to pregnant Charles River CD1 mice and assessing maternal health and fetal development till the end of gestation. Doses of 25 mg/kg or higher of pure FB1 induced maternal liver lesions (mostly necrotic changes), associated with ascites and increased hepatocytic nuclear diameter. Fumonisin doses of 50 mg/kg or higher also resulted in significantly increased maternal ALT on GD12, and reduced offspring bodyweights on GD18. Increased resorptions and decreased numbers of live offspring were only evident at 100 mg FB1/kg body weight. Offspring exhibited dose-dependent increase in the incidence and severity of hydrocephalus of both the lateral and third ventricles at doses of 25 mg/kg or higher. Doses of 25 mg/kg or higher also increased the sphinganine/sphingosine (Sa/So) ratios in maternal but not fetal livers. These results suggest that FB1 may be a developmental toxicant accounting for most but not all earlier reported effects of F. moniliforme culture extract. Association of FB1 effects on the offspring with maternal hepatoxicity and with alteration of Sa/So ratio in maternal but not fetal liver supported the earlier claim that FB1 effects on the mouse offspring are mediated by maternal hepatotoxicity.

Pharmacokinetics of pyropheophorbide-a-hexyl ether in the dog.

BACKGROUND AND OBJECTIVES: Pyropheophorbide-a-hexyl ether (HPPH) is a new compound being investigated for use as a photosensitizer for photodynamic therapy; however, the pharmacokinetics are not known for any of the target species likely to be treated with this drug. The objective of this study was to determine the pharmacokinetic parameters of this drug prior to institution of a clinical trial in canine patients with various cancers. STUDY DESIGN, MATERIALS AND METHODS: HPPH (0.3mg/kg i.v.) was administered to 12 dogs and blood samples were drawn at intervals for 24 hours and plasma HPPH concentrations were determined. Pharmacokinetic parameters were calculated for each dog. RESULTS: No evidence of toxicity was noted in any dog. The mean half-life was calculated to be 26.98 +/- 2.35 hrs. The mean clearance was 5.061 +/- 0.214 ml/hr/kg. The mean volume of distribution of the central compartment was 0.069 +/- 0.003 L/kg, and the mean steady state volume of distribution was 4.47 +/- 0.25 L/kg. CONCLUSION: The conclusion is that 0.3 mg/kg HPPH injected intravenously resulted in measurable plasma levels for 24 hrs, and resulted in no detectable adverse reactions.

Liver disease in cattle induced by consumption of moldy hay.

Normally innocuous forages are sporadically associated with hepatogenous photosensitization outbreaks at certain times of the year or when grown and harvested during unusual environmental conditions, such as periods of excessive rainfall. Allegations of livestock illness following consumption of such moldy hays are associated with clinical syndromes uncharacteristic of known forage-related diseases, suggesting that unidentified toxin(s) may be responsible. This study was instigated by field observations of hepatogenous photosensitization in cattle fed alfalfa-grass forage. To document the toxic nature of the hay, large bales of hay (450 kg) were fed, ad libitum, to 3 groups of 2 calves each. Elevated serum liver enzymes provided evidence of hepatobiliary disease. Gamma glutamyl transferase activities in serums of the calves sustained at least a 10-fold increase above baseline during the feeding trials. Histologic examination of liver biopsies and postmortem sections revealed mild periportal fibrosis and biliary hyperplasia. Culture material from 12 fungal isolates from the hay failed to induce liver disease in calves.

Chronic effects of dietary fumonisin on the heart and pulmonary vasculature of swine.

Fumonisins, mycotoxins produced in Fusarium moniliforme-infected corn, are hepatotoxic in several species and induce hepatocarcinomas in laboratory rats, leukoencephalomalacia in equine species, and fulminant pulmonary edema in swine. To date, the effects of fumonisin on pigs has dealt solely with the high levels required to induce the dramatic development of acute fulminant, and usually lethal, pulmonary edema. Less sensational, but equally important, are the chronic effects of fumonisin on swine health. In the first trial, six gilts were fed, ad libitum, a formulated complete ration containing 100 mg fumonisin B1 (FB1)/kg for 10 days followed by a ration containing 190 mg FB1/kg for up to 83 days. In the second trial, five gilts were fed, ad libitum, rations containing 150-170 mg FB1/kg for up to 210 days. The concentration of fumonisin B2 (FB2) was 31 +/- 2% that of the FB1. Weight-matched controls were selected for cardiac parameter comparison. Pigs fed fumonisin for at least 93 days developed nodular hyperplasia of the liver. Evidence from the microscopic examination of heart and lungs and gravimetric measurements of cardiac tissues from four pigs exposed to dietary fumonisins for 6 months suggests that the pulmonary vasculature is a target of chronic exposure. This study provides additional evidence linking the pathogenesis of fumonisin-induced lesions in a variety of organ systems of various animal species to a vascular mechanism.