Safety evaluation of a high lipid Whole Algalin Flour (WAF) from Chlorella protothecoides.

Microalgae such as Chlorella spp. have a long history of use in human food. A high lipid Whole Algalin Flour (WAF) composed of dried milled Chlorella protothecoides was evaluated for subchronic toxicity and genotoxic potential. Likelihood of food allergy potential was also evaluated by human repeat-insult patch test. In the subchronic study, rats were fed dietary levels of 25,000, 50,000 or 100,000 ppm WAF in feed for 93-94 days. No mortalities occurred. No treatment-related effects were identified for general condition, body weight, food consumption, ophthalmology, urinalysis, hematology, clinical chemistry, gross pathology, organ weights, and histopathology. Although statistically significant effects were noted for several endpoints, none was test-substance related. The no-observed-adverse-effect level (NOAEL) for WAF was based on consumption of the 100,000 ppm diet, the highest dietary concentration tested, and was 4807 mg/kg bw/d in male rats and 5366 mg/kg bw/d in female rats. Additionally, WAF (≤ 5000 µg/plate) was not mutagenic in Salmonella typhimurium or Escherichia coli tester strains nor did WAF induce a clastogenic response in bone marrow from mice given a single oral dose (2000 mg/kg bw). Further, WAF did not elicit skin sensitization in a repeat-insult dermal patch test which indicates little potential for food allergy.

Polyclonal hypergammaglobulinemia and autoantibody production induced by vaccination in farmed Atlantic salmon.

The introduction of oil-adjuvanted vaccines in salmon aquaculture made large-scale production feasible by reducing the impact of infections. Vaccines given intraperitoneally (ip) contain oil adjuvant such as mineral oil. However, in rodents, a single ip injection of adjuvant hydrocarbon oil induces lupus-like systemic autoimmune syndrome. We have recently reported that autoimmune disease in farmed salmon, characterized by production of various autoantibodies, immune complex glomerulonephritis, liver thrombosis, and spinal deformity, are previously unrecognized side effects of vaccination. In the present study, we examined whether vaccination-induced autoantibody production in farmed Atlantic salmon is a mere result of polyclonal B-cell activation. Sera were collected from 205 vaccinated and unvaccinated Atlantic salmon (experimental, 7 farms) and wild salmon. Total IgM levels and autoantibodies to salmon blood cell (SBC) extract in sera were measured by ELISA and the relationship between hypergammaglobulinemia and autoantibody production was analyzed. Comparison of endpoint titers vs levels/units using a single dilution of sera in detection of autoantibodies to SBC showed near perfect correlation, justifying the use of the latter for screening. Both total IgM and anti-SBC antibodies are increased in vaccinated salmon compared with unvaccinated controls, however, they do not always correlate well when compared between groups or between individuals, suggesting the involvement of antigen-specific mechanisms in the production of anti-SBC autoantibodies. The primary considerations of successful vaccine for aquaculture are cost-effectiveness and safety. Vaccination-induced autoimmunity in farmed Atlantic salmon may have consequences on future vaccine development and salmon farming strategy. Evaluation for polyclonal hypergamamglobulinemia and autoimmunity should be included as an important trait when vaccine efficacy and safety are evaluated in future.

Vaccination-induced systemic autoimmunity in farmed Atlantic salmon.

Over half of the salmon consumed globally are farm-raised. The introduction of oil-adjuvanted vaccines into salmon aquaculture made large-scale production feasible by preventing infections. The vaccines that are given i.p. contain oil adjuvant such as mineral oil. However, in rodents, a single i.p. injection of adjuvant hydrocarbon oil induces lupus-like systemic autoimmune syndrome, characterized by autoantibodies, immune complex glomerulonephritis, and arthritis. In the present study, whether the farmed salmon that received oil-adjuvanted vaccine have autoimmune syndrome similar to adjuvant oil-injected rodents was examined. Sera and tissues were collected from vaccinated or unvaccinated Atlantic salmon (experimental, seven farms) and wild salmon. Autoantibodies (immunofluorescence, ELISA, and immunoprecipitation) and IgM levels (ELISA) in sera were measured. Kidneys and livers were examined for pathology. Autoantibodies were common in vaccinated fish vs unvaccinated controls and they reacted with salmon cells/Ags in addition to their reactivity with mammalian Ags. Diffuse nuclear/cytoplasmic staining was common in immunofluorescence but some had more specific patterns. Serum total IgM levels were also increased in vaccinated fish; however, the fold increase of autoantibodies was much more than that of total IgM. Sera from vaccinated fish immunoprecipitated ferritin and approximately 50% also reacted with other unique proteins. Thrombosis and granulomatous inflammation in liver, and immune-complex glomerulonephritis were common in vaccinated fish. Autoimmunity similar to the mouse model of adjuvant oil-induced lupus is common in vaccinated farmed Atlantic salmon. This may have a significant impact on production loss, disease of previously unknown etiology, and future strategies of vaccines and salmon farming.

Erythrocyte membrane fatty acid content in infants consuming formulas supplemented with docosahexaenoic acid (DHA) and arachidonic acid (ARA): an observational study.

In this observational study, we compared erythrocyte membrane fatty acids in infants consuming formula supplemented with docosahexaenoic acid (DHA) and arachidonic acid (ARA) with those consuming other types of milks. In 110 infants who were participants in a cohort study of otherwise healthy children at risk for developing type 1 diabetes, erythrocytes were collected at approximately 9 months of age, and fatty acid content was measured as a percentage of total lipids. Parents reported the type of milk the infants consumed in the month of and prior to erythrocyte collection: infant formula supplemented with ARA and DHA (supplemented formula), formula with no ARA and DHA supplements (non-supplemented formula), breast milk, or non-supplemented formula plus breast milk. Membrane DHA (4.42 versus 1.79, P < 0.001) and omega-3 fatty acid (5.81 versus 3.43, P < 0.001) levels were higher in infants consuming supplemented versus non-supplemented formula. Omega-6 fatty acids were lower in infants consuming supplemented versus non-supplemented formula (26.32 versus 29.68, P = 0.023); ARA did not differ between groups. Infants given supplemented formula had higher DHA (4.42 versus 2.81, P < 0.001) and omega-3 fatty acids (5.81 versus 4.45, P = 0.008) than infants drinking breast milk. In infants whose mothers did not receive any dietary advice, use of supplemented formula is associated with higher omega-3 and lower omega-6 fatty acid status.

A comparative study of Florida strains of Cylindrospermopsis and Aphanizomenon for cylindrospermopsin production.

The toxin cylindrospermopsin (CYN) is produced by a variety of cyanobacterial genera. One of these, Cylindrospermopsis raciborskii, is generally assumed to be the source of CYN in lakes and rivers in Florida, USA. However, in this study, none of the eight Florida isolates of this species tested contained the genetic determinants involved in toxin production nor did they produce CYN. We show for the first time that Aphanizomenon ovalisporum isolated from a pond in this state has the genes putatively associated with CYN production. Analysis by liquid chromatography with mass spectrometric detection (LC/MS) revealed that it produced CYN in the range of 7.39-9.33 microg mg(-1) freeze-dried cells. 16S rDNA sequences of this strain showed 99.6% and 99.9% identity to published A. ovalisporum and Anabaena bergii 16S sequences, respectively. These results help to explain the general lack of a defined relationship between the abundance of C. raciborskii in freshwater ecosystems of Florida and observed concentrations of CYN. The latter observation raises the potential that previous reports of CYN may be coincidental with unrecorded presence of another CYN-producing species.

Debromoaplysiatoxin in Lyngbya-dominated mats on manatees (Trichechus manatus latirostris) in the Florida King's Bay ecosystem.

Proliferation of the potentially toxic cyanobacterium, Lyngbya, in Florida lakes and rivers has raised concerns about ecosystem and human health. Debromoaplysiatoxin (DAT) was measured in concentrations up to 6.31 microg/g wet weight lyngbyatoxin A equivalents (WWLAE) in Lyngbya-dominated mats collected from natural substrates. DAT was also detected (up to 1.19 microg/g WWLAE) in Lyngbya-dominated mats collected from manatee dorsa. Ulcerative dermatitis found on manatees is associated with, but has not been proven to be caused by DAT.

Doxycycline levels in preocular tear film of horses following oral administration.

OBJECTIVE: To determine the concentration of doxycycline in preocular tear film following oral administration in horses as a possible therapeutic modality for infectious and keratomalacic equine keratitis. PROCEDURE: Eight broodmares without ocular disease from a Thoroughbred breeding facility were included in this study. Each mare received 20 mg/kg of doxycycline by mouth once daily in the morning for five consecutive days. Tears were collected 1 h after doxycycline administration starting on day one of administration and continuing for 10 consecutive days. Doxycycline levels in the tears were measured using liquid chromatography with tandem mass spectrometric detection (LC-MS/MS). RESULTS: Doxycycline was present in the tears of each mare at low microg/mL levels with the highest concentration appearing on the third to fifth days (8.21-9.83 microg/mL). Doxycycline levels had fallen below quantifiable ranges by day 10. No systemic side-effects were noted in any of the horses included in this study. CONCLUSIONS: Oral doxycycline is present in preocular tear film of normal horses with noninflamed eyes and may be useful as treatment in equine ulcerative keratomalacia. The oral dose listed was tolerated well by the horses in this study. The drug levels attained at 20 mg/kg once daily orally of doxycycline may aid in the treatment of corneal ulceration in horses, but further study is warranted.

Comparison of the effects of caffeine and doxapram on respiratory and cardiovascular function in foals with induced respiratory acidosis.

OBJECTIVE: To determine and compare the effects of caffeine and doxapram on cardiorespiratory variables in foals during isoflurane-induced respiratory acidosis. ANIMALS: 6 clinically normal foals (1 to 3 days old). PROCEDURES: At intervals of > or = 24 hours, foals received each of 3 IV treatments while in a steady state of hypercapnia induced by isoflurane anesthesia (mean +/- SD, 1.4 +/- 0.3% endtidal isoflurane concentration). After assessment of baseline cardiorespiratory variables, a low dose of the treatment was administered and variables were reassessed; a high dose was then administered, and variables were again assessed. Sequential low- and high-dose treatments included doxapram (loading dose of 0.5 mg/kg, followed by a 20-minute infusion at 0.03 mg/kg/min and then 0.08 mg/kg/min), caffeine (5 mg/kg and 10 mg/kg), and saline (0.9% NaCl) solution (equivalent volumes). RESULTS: Administration of doxapram at both infusion rates resulted in a significant increase in respiratory rate, minute ventilation, arterial blood pH, PaO(2), and arterial blood pressure. These variables were also significantly higher during doxapram administration than during caffeine or saline solution administration. There was a significant dose-dependent decrease in PaCO(2) and arterial bicarbonate concentration during doxapram treatment. In contrast, PaCO(2) increased from baseline values after administration of saline solution or caffeine. The PaCO(2) value was significantly lower during doxapram treatment than it was during caffeine or saline solution treatment. CONCLUSIONS AND CLINICAL RELEVANCE: Results indicated that doxapram restored ventilation in a dose-dependent manner in neonatal foals with isoflurane-induced hypercapnia. The effects of caffeine on respiratory function were indistinguishable from those of saline solution.

Omega-3 polyunsaturated fatty acid intake and islet autoimmunity in children at increased risk for type 1 diabetes.

CONTEXT: Cod liver oil supplements in infancy have been associated with a decreased risk of type 1 diabetes mellitus in a retrospective study. OBJECTIVE: To examine whether intakes of omega-3 and omega-6 fatty acids are associated with the development of islet autoimmunity (IA) in children. DESIGN, SETTING, AND PARTICIPANTS: A longitudinal, observational study, the Diabetes Autoimmunity Study in the Young (DAISY), conducted in Denver, Colorado, between January 1994 and November 2006, of 1770 children at increased risk for type 1 diabetes, defined as either possession of a high diabetes risk HLA genotype or having a sibling or parent with type 1 diabetes. The mean age at follow-up was 6.2 years. Islet autoimmunity was assessed in association with reported dietary intake of polyunsaturated fatty acids starting at age 1 year. A case-cohort study (N = 244) was also conducted in which risk of IA by polyunsaturated fatty acid content of erythrocyte membranes (as a percentage of total lipids) was examined. MAIN OUTCOME MEASURE: Risk of IA, defined as being positive for insulin, glutamic acid decarboxylase, or insulinoma-associated antigen-2 autoantibodies on 2 consecutive visits and still autoantibody positive or having diabetes at last follow-up visit. RESULTS: Fifty-eight children developed IA. Adjusting for HLA genotype, family history of type 1 diabetes, caloric intake, and omega-6 fatty acid intake, omega-3 fatty acid intake was inversely associated with risk of IA (hazard ratio [HR], 0.45; 95% confidence interval [CI], 0.21-0.96; P = .04). The association was strengthened when the definition of the outcome was limited to those positive for 2 or more autoantibodies (HR, 0.23; 95% CI, 0.09-0.58; P = .002). In the case-cohort study, omega-3 fatty acid content of erythrocyte membranes was also inversely associated with IA risk (HR, 0.63; 95% CI, 0.41-0.96; P = .03). CONCLUSION: Dietary intake of omega-3 fatty acids is associated with reduced risk of IA in children at increased genetic risk for type 1 diabetes.

Dietary exposure of largemouth bass to OCPs changes expression of genes important for reproduction.

Dieldrin and p,p'-DDE are ubiquitous contaminants known to act as endocrine disruptors, causing impaired development and reproduction in fish and wildlife. In order to elucidate the mechanisms by which dieldrin and p,p'-DDE cause endocrine disruption in largemouth bass (Micropterus salmoides), fish were exposed subchronically through the diet to both contaminants. Following 120 days of exposure, p,p'-DDE decreased estradiol in females, but increased 11-ketotestosterone in both sexes. Dieldrin on the other hand, decreased estradiol and 11-ketotestosterone in both sexes. Both pesticides also altered steady state mRNA expression levels of a set of genes chosen to represent three possible mechanisms of endocrine disruption: (1) direct interaction with soluble sex steroid receptors, (2) biosynthesis of endogenous sex hormones, and (3) metabolism of endogenous hormones. p,p'-DDE acted as a weak estrogen, increasing the expression of vitellogenin and estrogen receptor alpha in the liver. p,p'-DDE also altered the expression of genes involved in the synthesis of endogenous hormones as well as their metabolism. Dieldrin, on the other hand, only altered expression of vitellogenin and not estrogen receptor alpha. Dieldrin also altered the expression of genes involved in hormone synthesis and metabolism, and it dramatically lowered plasma hormone levels. Both pesticides targeted expression of genes involved in all three modes of action, suggesting that they each have multiple modes of action.

Organochlorine concentrations, reproductive physiology, and immune function in unique populations of freshwater Atlantic stingrays (Dasyatis sabina) from Florida's St. Johns River.

Within the past decade, reproductive and health disorders have been reported to occur in unique populations of Atlantic stingrays (Dasyatis sabina) inhabiting certain components of Florida's St. Johns River. Since these irregularities are consistent with the alleged effects of organochlorine (OC) contaminant exposure in other Florida wildlife, the goal of this study was to examine possible associations between OC concentrations and reproduction and/or immune function in stingrays from this river system. Liver concentrations of 30 OC pesticides/pesticide metabolites and total polychlorinated biphenyls (PCBs) were measured and compared in D. sabina collected from four central Florida lakes of the St. Johns River: Lake George, Lake Harney, Lake Jesup, and Lake Monroe. Reproductive biology, serum testosterone and 17beta-estradiol concentrations, and circulating white blood cell counts were examined and compared in stingrays from lakes that were determined to contain low (Lake George), intermediate (Lake Monroe), and high (Lake Jesup) levels of pesticide contamination, based on the results of liver OC assessments. Successful breeding occurred in Lake Jesup stingrays, indicating that the degree of OC accumulation in these animals is not high enough to cause reproductive impairment. However, elevated serum steroid concentrations and white blood cell counts were observed in Lake Jesup stingrays, suggesting that endocrine and immune function may be altered in these animals due to OC exposure and/or other, as yet unknown, ecological factors. Inconsistencies in the reproductive success of Lake Monroe stingrays were observed, confirming earlier reports of reproductive complications in this sub-population. Based on these findings, previous occurrences of reproductive failure in St. Johns River stingrays may be due to environmental factors other than OC exposure.

Neurological disease in wild loggerhead sea turtles Caretta caretta.

Beginning in October 2000, subadult loggerhead sea turtles Caretta caretta showing clinical signs of a neurological disorder were found in waters off south Florida, USA. Histopathology indicated generalized and neurologic spirorchiidiasis. In loggerhead sea turtles (LST) with neurospirorchiidiasis, adult trematodes were found in the meninges of the brain and spinal cord of 7 and 3 affected turtles respectively, and multiple encephalic intravascular or perivascular eggs were associated with granulomatous or mixed leukocytic inflammation, vasculitis, edema, axonal degeneration and occasional necrosis. Adult spirorchiids were dissected from meningeal vessels of 2 of 11 LST brains and 1 of 10 spinal cords and were identified as Neospirorchis sp. Affected LST were evaluated for brevetoxins, ciguatoxins, saxitoxins, domoic acid and palytoxin. While tissues from 7 of 20 LST tested positive for brevetoxins, the levels were not considered to be in a range causing acute toxicosis. No known natural (algal blooms) or anthropogenic (pollutant spills) stressors co-occurred with the turtle mortality. While heavy metal toxicosis and organophosphate toxicosis were also investigated as possible causes, there was no evidence for their involvement. We speculate that the clinical signs and pathologic changes seen in the affected LST resulted from combined heavy spirorchiid parasitism and possible chronic exposure to a novel toxin present in the diet of LST.

Determination of steroidal estrogens in flushed dairy manure wastewater by gas chromatography-mass spectrometry.

There is a critical need to accurately measure the concentrations of natural steroidal estrogens in flushed dairy manure wastewater (FDMW) to assess any potential risk of waterway contamination resulting from land application. Estrogens are a concern because low concentrations (10-100 ng L-1) in water can adversely affect aquatic vertebrate species such as fish, turtles, and frogs by disrupting the normal function of their endocrine systems. The objective of this study was to develop a sample preparation method that permits the quantification of four natural steroidal estrogens (17alpha-estradiol, 17beta-estradiol, estrone, and estriol) in FDMW by gas chromatography-mass spectrometry (GC-MS). Solid-phase extraction with graphitized carbon black was used for the bulk extraction of estrogens from FDMW and additional sample purification was accomplished with C-18. The sample preparation method allowed estrogens to be detected accurately by GC-MS in FDMW. Spiked recovery experiments indicated that the method is satisfactory for measuring the estrogens of interest in FDMW with average recovery of >90%. As expected in FDMW, characterization of the estrogen profile revealed a large abundance of 17alpha-estradiol relative to 17beta-estradiol and estrone. Estriol was not detected in FDMW. The methodology developed in this research helps provide an analytical foundation for the quantification of steroidal estrogens in FDMW by GC-MS.

Pharmacokinetics of azathioprine following single-dose intravenous and oral administration and effects of azathioprine following chronic oral administration in horses.

OBJECTIVE: To determine pharmacokinetics of azathioprine (AZA) and clinical, hematologic, and serologic effects of i.v. and oral administration of AZA in horses. ANIMALS: 6 horses. PROCEDURE: In study phase 1, a single dose of AZA was administered i.v. (1.5 mg/kg) or orally (3.0 mg/kg) to 6 horses, with at least 1 week between treatments. Blood samples were collected for AZA and 6-mercaptopurine (6-MP) analysis 1 hour before and at predetermined time points up to 4 hours after AZA administration. In study phase 2, AZA was administered orally (3 mg/kg) every 24 hours for 30 days and then every 48 hours for 30 days. Throughout study phase 2, blood samples were collected for CBC determination and serum biochemical analysis. RESULTS: Plasma concentrations of AZA and its metabolite, 6-MP decreased rapidly from plasma following i.v. administration of AZA, consistent with the short mean elimination half-life of 1.8 minutes. Oral bioavailability of AZA was low, ranging from 1% to 7%. No horses had abnormalities on CBC determination or serum biochemical analysis, other than 1 horse that was lymphopenic on day 5 and 26 of daily treatment. This horse developed facial alopecia from which 1 colony of a Trichophyton sp was cultured; alopecia resolved within 1 month after the study ended. CONCLUSIONS AND CLINICAL RELEVANCE: Overall, no adverse effects were observed with long-term oral administration of AZA to horses, although 1 horse did have possible evidence of immunosuppression with chronic treatment. Further investigation of the clinical efficacy of AZA in the treatment of autoimmune diseases in horses is warranted.

Distinctive patterns of autoimmune response induced by different types of mineral oil.

Although mineral oils are generally considered nontoxic and have a long history of use in humans, the mineral oil Bayol F (incomplete Freund's adjuvant, IFA) and certain mineral oil components (squalene and n-hexadecane) induce lupus-related anti-nRNP/Sm or -Su autoantibodies in nonautoimmune mice. In the present study, we investigated whether medicinal mineral oils can induce other types of autoantibodies and whether structural features of hydrocarbons influence autoantibody specificity. Female 3-month-old BALB/c (16-45/group) mice each received an i.p. injection of pristane (C19), squalene (C30), IFA, three medicinal mineral oils (MO-F, MO-HT, MO-S), or PBS. Sera were tested for autoantibodies and immunoglobulin levels. Hydrocarbons were analyzed by gas chromatography/mass spectrometry. IFA contained mainly C15-C25 hydrocarbons, whereas MO-HT and MO-S contained C20-C40, and MO-F contained C15-C40. Pristane and n-hexadecane were found in IFA (0.17% and 0.10% w/v, respectively) and MOs (0.0026-0.027%). At 3 months, pristane and IFA induced mainly IgG2a, squalene IgG1, and MOs IgG3 and IgM in sera. Anti-cytoplasmic antibodies were common in mice treated with MO-F, as well as those treated with pristane, squalene, and IFA. Anti-ssDNA and -chromatin antibodies were higher in MO-F and MO-S than in untreated/PBS, squalene-, or IFA-treated mice, suggesting that there is variability in the induction of anti-nRNP/Sm versus -chromatin/DNA antibodies. The preferential induction of anti-chromatin/ssDNA antibodies without anti-nRNP/Sm/Su by MO-S and MO-F is consistent with the idea that different types of autoantibodies are regulated differently. Induction of autoantibodies by mineral oils considered nontoxic also may have pathogenetic implications in human autoimmune diseases.

Endothelin 1 levels in the aqueous humor of dogs with glaucoma.

PURPOSE: Endothelin 1 is a small peptide that is involved in regulation of intraocular pressure and modulation of ocular circulation. To investigate the role of endothelin 1 in canine glaucoma, the authors measured aqueous humor levels of endothelin 1 in healthy dogs and in dogs with hypertensive glaucoma. METHODS: Aqueous humor samples were obtained with general anesthesia from the eyes of healthy dogs (n = 5) and dogs with hypertensive glaucoma (n = 10). Measurements were made by enzyme immunoassay for endothelin 1. RESULTS: The endothelin 1 aqueous humor range was 1.12 - 3.63 pg/mL for healthy dogs and 1.97 - 14.97 pg/mL for glaucomatous dogs. The healthy and glaucomatous canine endothelin 1 aqueous levels (mean +/- SD) were 2.33 +/- 0.90 and 8.11 +/- 5.03 pg/mL, respectively. A two-way analysis of variance indicated that this difference was significant (P = 0.0084). The effect of age on endothelin 1 levels was not significant (P = 0.6283). The large variability found within the glaucomatous group could be explained by the degree of damage of the retina (P = 0.0006). There was no significant correlation between intraocular pressure and endothelin 1 aqueous humor levels within the glaucomatous group (P = 0.29). CONCLUSIONS: The aqueous humor of dogs with hypertensive glaucoma contains significantly higher levels of endothelin 1 than that of healthy dogs.

Predicting maternal body burdens of organochlorine pesticides from eggs and evidence of maternal transfer in Alligator mississippiensis.

Few data exist regarding maternal-embryonal transfer of organochlorine pesticides (OCPs) in reptiles. The objective of the present study was to evaluate maternal transfer of OCPs in American alligators (Alligator mississippiensis) from low-, intermediate-, and high-OCP-exposure sites. Overall, total OCP burdens ranged from less than 0.8 ppb in blood to more than 44,000 ppb in abdominal adipose tissue (wet wt concentrations). Lipid-adjusted ratios of maternal adipose burdens (total OCPs) to yolk burdens were close to one (0.94 +/- 0.31:1), suggesting that animals were in steady state and that OCPs in eggs originated from adipose lipids. In contrast, lipid-adjusted muscle and liver OCP burdens were greater than yolk OCP burdens, suggesting that lipids in muscle were not utilized during oogenesis and that nonlipid liver tissue sequesters OCPs. Predictive equations were derived for several tissues and several OCP analytes with r2 values ranging from 0.40 to 0.99 (p < 0.05). We suggest that yolk burdens are predictive of maternal tissue burdens for certain tissues and OCPs and that certain OCPs are maternally transferred in the American alligator. Furthermore, we suggest that future studies should investigate the applicability of these predictive equations for assessing maternal exposure in other crocodilian species.

Dietary safety of cycloastragenol from Astragalus spp.: subchronic toxicity and genotoxicity studies.

Extracts, teas, and other preparations of Astragalus roots (e.g., Radix Astragali) are historically recognized traditional medicines and foods. Cycloastragenol (CAG), a bioactive triterpene aglycone from Astragalus root extracts, is being developed as a modern dietary ingredient. To this end, studies assessing subchronic toxicity and genotoxic potential were conducted. In the subchronic study with recovery component, rats ingested 0, 40, 80, or 150 mg/kg/d CAG by oral gavage for ⩾91 consecutive days. No treatment-related mortalities occurred and no cardiac effects were identified. Although several endpoints among those monitored (i.e., clinical observations, body weight, food consumption, ophthalmology, urinalysis, hematology, clinical chemistry, gross pathology, organ weights, or histopathology) exhibited statistically significant effects, none was adverse. The oral no-observed-adverse-effect level (NOAEL) for CAG was >150 mg/kg/d in male and female rats. CAG (⩽5000 µg/plate) did not induce mutagenicity in Salmonella typhimurium or Escherichia coli tester strains. Although the in vitro chromosome aberration assay gave a moderately positive response (likely due to poor solubility) for one intermediate concentration (1.50mM) with metabolic activation, responses were negative in all other test groups. Finally, in the in vivo micronucleus assay no clastogenicity was observed in peripheral erythrocytes from mice administered 2000 mg/kg CAG by intraperitoneal injection.

Safety evaluation of oleic-rich triglyceride oil produced by a heterotrophic microalgal fermentation process.

Numbers of macro- and microalgae have been used as food sources in various cultures for centuries. Several microalgae are currently being developed as modern food ingredients. The dietary safety of oleic-rich microalgal oil produced using a heterotrophic fermentation process was assessed in a 13-week feeding trial in rats with genotoxic potential evaluated using in vitro and in vivo assays. In the genotoxicity assays, the test oil was not mutagenic in Salmonella typhimurium or Escherichia coli tester strains (⩽5000µg/plate) with or without metabolic activation. Further, no clastogenic response occurred in chromosome aberration assays in the bone marrow of mice administered a single intraperitoneal dose (2000mg/kg). In the subchronic study, rats consumed feed containing 0, 25,000, 50,000 or 100,000ppm oleic-rich oil for 90days. No treatment-related mortalities or adverse effects occurred in general condition, body weight, food consumption, ophthalmology, urinalysis, hematology, clinical chemistry, gross pathology, organ weights or histopathology. Although several endpoints exhibited statistically significant effects, none were dose-related or considered adverse. Taking all studies into consideration, the NOAEL for the oleic-rich oil was 100,000ppm, the highest concentration tested and equivalent to dietary NOAELs of 5200mg/kg bw/day and 6419mg/kg bw/day in male and female rats, respectively.