Toxicological evaluation of a fish oil concentrate containing Very Long Chain Fatty Acids.

Very long chain fatty acids (VLCFA) have a chain length ≥24 carbons. Fish contain low levels of these fatty acids. A commercial oil called EPAX® Evolve 05 with an up-concentration of VLCFAs of approximately 10 times, has been developed as a dietary supplement by Epax Norway AS. A series of toxicological studies were performed using mice and rats to determine the safety and toxicity of repeat dosing with a gavage administered VLCFA formulation. The results suggest transient lipid accumulation in kidneys and liver. Lipid accumulation was seen with the test item and with the soya control but was not dose related. Liver and kidney lipid accumulation, whilst present in 14- day repeat dose study, was absent in a 90-day rat study. No treatment-effect was seen in urine analysis in any of the studies. No treatment-related effects were seen with a functional observation battery, ophthalmological examination, haematology, urine analysis, oestrus cycle, thyroid hormones, organ weight, or histopathology. In the 90-day study the liver enzymes ALP, AST and ALT were statistically significantly increased with test item but within control values. There were no associated histological findings in the liver suggesting there was no toxic effect and the normalisation of values for all liver enzymes in the recovery groups suggests an adaptive response rather than a prevailing toxic response. The no-observed-adverse-effect level (NOAEL) was determined as 1200 mg VLCFA/kg b.w./day.

Safety assessment of fish oil green extraction and in vivo acute toxicity evaluation.

Sardine co-products can represent an interesting source of bioactive compounds, such as polyunsaturated fatty acids and in particular omega-3. This study aimed to investigate extraction of oil from sardine co-products by enzymatic hydrolysis using two proteases: commercial Alcalase and protease Bb from a local fungal strain (P2) of Beauveria bassiana, which overproduces proteases. Despite a higher degree of hydrolysis (41.34%) than Alcalase (24.28%), protease Bb allowed the extraction of approximately the same oil content. Resulting oil from both processes had the same fatty acid profile. Interestingly, the all-produced oil displayed an attractive w6/w3 ratio, an indicator of nutritional quality, of the order of 0.16. The safety of the generated oils was also assessed by treating two groups of Wistar rats with the fish oil administered by oral gavage at the doses (30 mg/kg and 300 mg/kg body weight) for 14 days using olive oil as a vehicle. Compared to controls used, both treated groups showed no statistically significant differences. Consequently, the acute oral toxicity evaluated by hematological, biochemical, and histological studies showed the safety of the oil generated using B. bassiana protease.

Toxicity of oxidized fish oil in pregnancy: a dose-response study in rats.

Fish oil (FO) supplements are consumed during pregnancy to increase dietary omega-3. However, FO is often oxidized past recommended limits. In rats, a large dose of highly oxidized FO substantially increased newborn mortality, but the effects of human-relevant doses of less oxidized oil are unknown. A dose-response study in rats was conducted to estimate the safe level of oxidation during pregnancy. Sprague-Dawley rat dams were mated, then individually housed and provided with a gel treatment on each day of pregnancy. Treatment groups differed only in the FO content of the gel; control (no oil), PV5, PV10, and PV40 [0.05 mL of FO oxidized to a peroxide value (PV) of 5, 10, or 40 meq/kg], or PV40(1 mL) (1 mL of PV40). A subset of dams was culled on gestational day 20 to enable sampling, and the remainder were allowed to give birth. Newborn mortality was recorded. Offspring were sampled on postnatal days 2 and 21, and dams on day 21. There were no signs of unwellness during pregnancy. However, there was markedly increased neonatal mortality affecting the PV40(1 mL) (12.8%) and PV40 (6.3%) groups, but not the control, PV5, or PV10 groups (1%-1.4%). Dietary-oxidized FO altered the expression of placental genes involved in antioxidant pathways and the production of free radicals. Highly oxidized FO was toxic in rat pregnancy leading to a marked increase in mortality even at a human-relevant dose. We observed no toxic effects of FOs with PV ≤10 meq/kg, suggesting that this is an appropriate maximum limit.

Skipjack tuna (Katsuwonus pelamis) eyeball oil exerts an anti-inflammatory effect by inhibiting NF-κB and MAPK activation in LPS-induced RAW 264.7 cells and croton oil-treated mice.

The effect of tuna eyeball oil (TEO) on lipopolysaccharide (LPS)-induced inflammation in macrophage cells was investigated. TEO had no cytotoxicity in cell viability as compared to the control in LPS induced RAW 264.7 cells. TEO reduced the levels of NO and pro-inflammatory cytokines by up to 50% in a dose-dependent manner. The expression of NF-κB and MAPKs as well as iNOS and COX-2 proteins was reduced by TEO, which suggests that its anti-inflammatory activity is related to the suppression of the NF-κB and MAPK signaling pathways. The rate of formation of ear edema was reduced compared to that in the control at the highest dose tested. In an acute toxicity test, no mice were killed by TEO doses of up to 5000mg/kg body weight during the two week observation period. These results suggested that TEO may have a significant effect on inflammatory factors and be a potential anti-inflammatory therapeutic.

Toxicological evaluation of arachidonic acid (ARA)-rich oil and docosahexaenoic acid (DHA)-rich oil.

The safety of DHA-rich oil from Schizochytrium sp. and ARA-rich oil from Mortierella alpina was separately evaluated by testing for gene mutations, clastogenicity, and aneugenicity, and by conducting 28-day and 90-day dietary studies in Wistar rats. The results of all genotoxicity tests were negative. The 28-day and 90-day studies involved dietary exposure to 1000, 2500, and 5000 mg per kg bw of the DHA-rich and ARA-rich oils and two control diets: water and corn oil (vehicle control). There were no treatment-related effects of either the DHA-rich or ARA-rich oils on clinical observations, body weight, food consumption, behavior, hematology, clinical chemistry, coagulation, urinalysis parameters, or necropsy findings. Increases in cholesterol and triglyceride levels were considered related to a high oil diet and non-adverse. The no observable adverse effect level (NOAEL) for both the DHA-rich and ARA-rich oils was 5000 mg per kg bw, the highest dose tested. The results confirm that these oils possess toxicity profiles similar to those of other currently marketed oils and support the safety of DHA-rich oil from Schizochytrium sp. and ARA-rich oil from Mortierella alpina for their proposed uses in food.

Effect of different degrees of hydrogenated fish oil on intestinal carcinogenesis in Min/+ mice.

UNLABELLED: Intake of trans fatty acids from hydrogenated fish oils has been related to increased risk of coronary heart diseases. The possible effect on colorectal carcinogenesis is unclear. MATERIALS AND METHODS: Multiple intestinal neoplasia (Min/+) mice were fed one of four experimental diets: either raw fish oil (FO), low (LHFO)-, high (HHFO)- or fully-hydrogenated fish oil (FFHO), from 0 to 9 weeks of age. The number and size of intestinal tumors were recorded. RESULTS: There was no difference between the intervention groups in the numbers of developed intestinal tumors. The tumor size was statistically significantly lower in HHFO vs. the FO-group in male Min/+ mice. The HHFO and FHFO groups had lower weight gain than did the FO group (p=0.008 and p=0.04, respectively), but gender differences, due to effect of dietary intervention on weight gain, were found in Min/+ mice. CONCLUSION: When compared with raw fish oil, different degrees of hydrogenation of the fish oil had no effect on intestinal carcinogenesis in Min/+ mice.

Safety assessment of EPA-rich triglyceride oil produced from yeast: genotoxicity and 28-day oral toxicity in rats.

The 28-day repeat-dose oral and genetic toxicity of eicosapentaenoic acid triglyceride oil (EPA oil) produced from genetically modified Yarrowia lipolytica yeast were assessed. Groups of rats received 0 (olive oil), 940, 1880, or 2820 mg EPA oil/kg/day, or fish oil (sardine/anchovy source) by oral gavage. Lower total serum cholesterol was seen in all EPA and fish oil groups. Liver weights were increased in the medium and high-dose EPA (male only), and fish oil groups but were considered non-adverse physiologically adaptive responses. Increased thyroid follicular cell hypertrophy was observed in male high-dose EPA and fish oil groups, and was considered to be an adaptive response to high levels of polyunsaturated fatty acids. No adverse test substance-related effects were observed on body weight, nutritional, or other clinical or anatomic pathology parameters. The oil was not mutagenic in the in vitro Ames or mouse lymphoma assay, and was not clastogenic in the in vivo mouse micronucleus test. In conclusion, exposure for 28 days to EPA oil derived from yeast did not produce adverse effects at doses up to 2820 mg/kg/day and was not genotoxic. The safety profile of the EPA oil in these tests was comparable to a commercial fish oil.

Safety assessment of EPA-rich oil produced from yeast: Results of a 90-day subchronic toxicity study.

The safety of eicosapentaenoic acid (EPA) oil produced from genetically modified Yarrowia lipolytica yeast was evaluated following 90 days of exposure. Groups of rats received 0 (olive oil), 98, 488, or 976 mg EPA/kg/day, or GRAS fish oil or deionized water by oral gavage. Rats were evaluated for in-life, neurobehavioral, anatomic and clinical pathology parameters. Lower serum cholesterol (total and non-HDL) was observed in Medium and High EPA and fish oil groups. Lower HDL was observed in High EPA and fish oil males, only at early time points. Liver weights were increased in High EPA and Medium EPA (female only) groups with no associated clinical or microscopic pathology findings. Nasal lesions, attributed to oil in the nasal cavity, were observed in High and Medium EPA and fish oil groups. No other effects were attributed to test oil exposure. Exposure to EPA oil for 90 days produced no effects at 98 mg EPA/kg/day and no adverse effects at doses up to 976 mg EPA/kg/day. The safety profile of EPA oil was comparable to that of GRAS fish oil. These results support the use of EPA oil produced from yeast as a safe source for use in dietary supplements.

Evaluación de la toxicidad aguda oral y de la actividad antimicrobiana de una mezcla de aceite de hígado de tiburones de Cuba / Assessment of the oral acute toxicity and the antimicrobial activity of an oily mixture from shark's liver of Cuba

Se evaluó la toxicidad aguda oral y la actividad antimicrobiana de una mezcla de aceites de hígado de tiburón, de las especies Rhincodon typu (tiburón ballena) y Galeocerdo cuvier (tiburón tigre), que habitan en zonas aledañas a las costas del litoral norte occidental de Cuba, para su posterior uso farmacéutico, debido a que presenta un alto contenido de vitaminas y de ácidos grasos, que le confieren actividad antioxidante y antiinflamatoria. El estudio de la toxicidad aguda oral demostró que la mezcla de aceites de hígado de tiburones, no provocó alteraciones macroscópicas en los órganos extraídos, ni síntomas tóxicos severos, ni mortalidad de ninguno de los animales empleados en el estudio a la dosis de 20 mL/kg. Los resultados del estudio de la actividad antimicrobiana demostraron una ligera actividad bacteriostática frente a K pneumoniae; además una actividad antifúngica frente a Microsporum canis; y resistencia frente a C albicans y T mentagrophytes a las concentraciones evaluadas

The total acute toxicity and the antimicrobial activity of an oil mixtures from shark liver of Rhicodon typu (whale-shark) and Galeocerdo cuvier (tigger-shark) was assessed in species leaving in the adjacent costs of Cuban northern coastal for its subsequent pharmaceutical use due to its high content of vitamins and fatty acids and its antioxidant and anti-inflammatory activity. Study of oral acute toxicity demonstrated that oil mixture of shark liver hasn't macroscopic alterations in removed organs, severe toxic symptoms and on mortality of any animals used in study at 20 mL/kg dose. Study results of antimicrobial activity showed a slight bacteriostatic activity against K pneumoniae and an antifungal activity against Microsporum canis, and a resistance against C albicans and T mentagrophytes at assessed concentrations

Dietary n-3 polyunsaturated fatty acids enhance metastatic dissemination of murine T lymphoma cells.

Epidemiological investigation and animal studies have shown that dietary n-3 PUFA prevent the development and progression of certain types of cancer. However, conflicting results have been reported by the few studies that focused on the effect of dietary n-3 PUFA on the development of metastases. In the present study, we investigated the metastatic dissemination of murine T lymphoma lines with different metastatic potential transplanted into mice fed a fish oil diet, compared with mice fed a maize oil diet. Transplantation of highly metastatic S11 cells into animals fed a fish oil diet induced a large lymphomatoid infiltration in the spleen, associated with an eight-fold increase in spleen weight, compared with normal animals on the same diet. In contrast, only a limited increase in spleen weight was found in animals transplanted with S11 cells while fed a maize oil diet. No significant increase in spleen weight was found in animals transplanted with low-metastatic 164T2 cells regardless of whether they were fed a fish oil or a maize oil diet. At the end of experiment, an overt cachexia was shown by animals fed a fish oil diet transplanted with S11 cells, but not by those transplanted with 164T2 cells. The particularly high pro-metastatic effect of dietary n-3 PUFA on S11 cells rules out the generalisation that dietary n-3 PUFA inhibit tumour growth and progression.

Differential induction of cytochrome P450 1A1 and 1B1 mRNA in primary cultured bovine hepatocytes treated with TCDD, PBDD/Fs and feed ingredients.

This study investigates the dose-dependent expression of CYP1A1 and CYP1B1 in primary cultured bovine hepatocytes exposed to TCDD, several polybrominated dibenzo-p-dioxins and furans (PBDD/Fs) congeners and fish oil used as animal feed ingredients to identify their dioxin-like potentials. Hepatocytes were isolated from calf liver, cultured and treated for 24h with the target compounds or extracts. Quantitative real time polymerase chain reaction analysis (qRT-PCR) showed that relative mRNA levels for CYP1A1 and CYP1B1 exhibited a dose-dependent induction by TCDD. The EC(50) of the TCDD concentration for CYP1A1 expression was approximately 4-fold less than that of CYP1B1. The estimated dioxin-like toxic potential of PBDD/Fs could be ranked in the following order: 2,3,7,8-TBDD>1,2,3,7,8-PBDF>2,3,4,7,8-PBDF>1,2,3,6,7,8-HBDD. A good correlation was also observed in HRGC/HRMS-derived TEQs in fish oil samples and relative CYP1A1 mRNA induction in bovine hepatocytes treated with purified fish oil extracts. The data suggested that quantification of biomarker regulations in primary cultured hepatocytes could represent an effective tool for both the screening and study of various chemical entities in larger animals.

Acute toxicity of the water-soluble fraction of spent lubricating oil on the African catfish Clarias gariepinus.

Static tests were employed to assess the acute toxicity of the water-soluble fraction (WSF) of spent automotive lubricating oil (of mixed SAE grades) on Clarias gariepinus, a freshwater fish commonly cultured in Nigeria. Median lethal concentrations (LC50) of the WSF were found to decrease as a function of exposure time from 690+/-21 (after 24 h) to 513+/-58 mg/l (after 96 h). The characteristics of the WSF such as mean acidity (pH 6.6), turbidity (40 NTU), total dissolved solids (TDS; 40 mg/l) and significantly reduced (P<0.05) dissolved-oxygen (DO) values (1.44 mg/l) were not compliant with existing standards set for discharged effluents. The solubility of the detected straight-chain aliphatics ranked as C14>C16>C32>C18>C28; that of the simple aromatics was ortho-xylene>para-xylene; and that of the polyaromatic hydrocarbons (PAHs) was acenaphthylene>9H-fluorene>naphthalene>anthracene>phenanthrene>chrysene>benzo[k]fluoranthene>benzo[a]pyrene>benzo[b]fluoranthene, most of which being serious carcinogens. These oil constituents and the overall physico-chemical properties of the WSF are expected to act synergistically on the test organism (C. gariepinus), eliciting the quantal responses observed. The toxicity of the WSF points to the base constituent, oxidative degradation, and mechano-chemical reactions associated with aged crankcase oils. These oils, therefore, should definitely no longer be disposed into water streams or landscape, not even at sub-lethal concentrations, because of the inherent toxicity of their soluble fractions and the associated danger of bioaccumulation.

Assessment of carcinogenic potential of repeated fish fried oil in mice.

Our prior studies have shown that single topical treatment of repeated fish fried oil extract (RFFE), containing various polycyclic aromatic hydrocarbons (PAHs), to the dorsal epidermis of mice caused enhancement of DNA damage along with higher expression of p53 and p21WAF1 proteins and cell-cycle arrest. In the present study carcinogenic potential of repeated fish fried oil (RFFO) and RFFE was assessed. Single topical application of RFFO (100 microL/animal) and RFFE (100-500 microg/animal) to Swiss albino female mice resulted in significant induction (1.8- to 7.4-fold) of ornithine decarboxylase activity. Twice weekly topical application of methylcholanthrene (MCA) for 24 wk or single topical application of 7,12-dimethylbenzanthracene (DMBA) or RFFO or RFFE, as initiator followed by twice weekly application of 12-O-tetradecanoyl phorbol myristate acetate (TPA) as promoter for 24 wk, resulted in development of skin papillomas after 6, 7, 18, and 9 wk, respectively. The cumulative number of tumors in MCA, DMBA/TPA, RFFE (200 microg)/TPA, and RFFE (500 microg)/TPA groups were 276, 168, 34, and 58 after 24 wk while negligible or minimal initiating activity was noticed in RFFO/TPA group. No tumors were found in animals either given twice weekly topical application of RFFO or a single initiating dose of DMBA followed by twice weekly application of RFFO. Histopathology of skin of animals treated with RFFE/TPA showed marked proliferation of epidermal layers along with abnormal mitosis and multinucleated tumor appearance. Skin of animals in groups RFFO/TPA and DMBA/RFFO showed sloughing and regeneration of epidermal layers, oedema along with proliferation of fibroblasts. Histochemical localization of gamma-glutamyl transpeptidase was found to be substantially higher in skin of mice treated with RFFO/TPA and RFFE/TPA. Animals treated with RFFO/TPA, DMBA/RFFO, and RFFE/TPA resulted in significant induction of cutaneous aryl hydrocarbon hydroxylase (AHH) (421-432%), ethoxyresorufin-O-deethylase (252-316%), and glutathione S-transferase (133-245%) activities. Animals treated with RFFO/TPA, DMBA/RFFO, and RFFE/TPA led to significant reduction in glutathione content (39-44%) with a concomitant increase in lipid peroxidation (254-492%). Animals treated with RFFO/TPA and RFFE/TPA led a significant decrease in catalase (43-69%) and superoxide dismutase (20-31%) activities while glutathione reductase activity was found to be diminished (23-51%) in RFFO, RFFO/TPA, DMBA/RFFO, and RFFE/TPA treated groups. These results suggest that RFFE possess skin tumor initiating activity and that it may have weak promoting activity as well, which may involve free radicals.

Murine strain differences in 8-hydroxy-deoxyguanosine formation in hepatic DNA induced by oxidized lard and dietary oils.

Difference of 8-hydroxy-deoxyguanosine (8-OH-dG) formation in liver DNA in C3H/HeN and in C57BL/6 mice--fed oxidized lard and dietary oils (soybean and sardine)--was investigated. The blank levels of 8-OH-dG were higher in C3H/HeN mice (highly sensitive to liver tumorigenesis) than in C57BL/6 mice (resistant strain). The level of 8-OH-dG increased much more in C3H/HeN mice than in the C57BL/6 mice fed by oxidized lard and dietary oil treatment. Feeding oxidized lard and dietary oils increased 8-oxo-guanine DNA glycosylase I (OGG1) and mRNA 8-oxo-dGTPase in C57BL/6 mice. On the other hand, no appreciable change of mRNA in the C3H/HeN mice was observed. The formation differences of 8-OH-dG from the two murine strains fed with oxidized lard and dietary oils may be associated with the different mRNA levels in the DNA repair enzymes because the mRNA levels in the DNA repair enzymes were much lower in C3H/HeN mice than in C57BL/6 mice.

Induction of hepatic cytochrome P450 isozymes, benzo(a)pyrene metabolism and DNA binding following exposure to polycyclic aromatic hydrocarbon residues generated during repeated fish fried oil in rats.

In the present study the effect of repeated fish fried oil (RFFO) and its extract (RFFE) on hepatic cytochrome P450 (CYP) isozymes, benzo(a)pyrene (BP) metabolism and DNA adduct formation was undertaken. HPLC analysis of RFFO showed the presence of several polycyclic aromatic hydrocarbons. CYP in microsomes from control and RFFO-treated animals showed a peak at 450 nm; however, a shift of 2 nm in the SORET region along with significant induction was observed in microsomes prepared from 3-methylcholanthrene (MC)- and RFFE-treated animals. Activities of hepatic ethoxyresorufin-O-deethylase, methoxyresorufin-O-deethylase, aryl hydrocarbon hydroxylase and erythromycin-N-demethylase were found to be significantly (P < 0.05) induced following exposure of RFFE, whereas none of these enzymes were altered in RFFO-treated group. Immunoblot analysis revealed that RFFE and MC were potent inducers of CYP1A1, 1A1/2 and 3A1 isozymes, where as RFFO showed no change in these protein levels. RT-PCR analysis showed induction of cDNA of CYP1A1 and CYP3A1 by RFFE treatment. Hepatic microsomes prepared from RFFE exposed animals enhanced BP metabolism with a concomitant increase in the relative proportion of BP 7,8-diol. Hepatic microsomes prepared from animals pretreated with RFFE and MC significantly enhanced the binding of [(3)H]-BP to calf thymus DNA. The overall results suggest that exposure to RFFE may induce hepatic CYP isozymes thereby producing enhanced reactive metabolites with a potential to bind with DNA that may result in cancer.

Subchronic toxicity of baltic herring oil and its fractions in the rat (III) bone tissue composition and dimension, and ratio of n-6/n-3 fatty acids in serum phospholipids.

Changes in total bone mineral density determined by the bone-ash method were recently demonstrated in rats, exposed to Herring oil from the contaminated southern part of the Baltic Sea. In the present study more detailed analysis of bone structure and biomechanics was performed and obtained results were evaluated in the context of dietary factors, such as polyunsaturated fatty acids, vitamin D and vitamin A. Baltic Sea herring oil was fractionated into one relatively pollutant-free fraction (F1), and two fractions with pronounced enrichment of pollutants (F2 and F3). Female Sprague-Dawley rats were fed diets supplemented with Baltic Herring oil, its fractions, Nordic Sea capelin oil or soy oil. Femur was scanned with peripheral quantitative computed tomography (pQCT) and also tested by a mechanical compression analysis. Polyunsaturated fatty acids, vitamin A and D were analysed in serum. Rats fed the high dose of herring oil exhibited shorter femur length with decreased diaphyseal cortical bone mineral density, as well as lowered metaphyseal cross-sectional area compared to the soy oil group. Rats fed the high dose of F1 diet had increased cortical and decreased trabecular area, and higher total and trabecular bone mineral density. Rats fed the low dose of F2 diet showed similar changes associated with increased maximum load and energy absorption in compression test of the femoral metaphysis. In summary, our findings in changes of bone geometry and density could not be linked to any isolated exposure parameter, suggesting synergistic or antagonistic effects of several components of the test diets.

Subchronic toxicity of Baltic herring oil and its fractions in the rat I: Fractionation and levels of organohalogen pollutants.

Baltic herring (Clupea harengus) oil was extracted and fractionated. To examine the contribution to toxicity and biological effects of different halogenated organic pollutants, the herring oil and the fractions were mixed into pelleted food and given to Sprague-Dawley female rats at three levels, corresponding to a human intake of 1.6, 8.2 and 34.4 kg fish per week. Herring oil, its fractions, as well as liver tissues from exposed rats, were analyzed for: eight chlorinated biphenyls, all 2,3,7,8-substituted chlorinated dibenzo-p-dioxins and dibenzofurans, hexachlorocyclohexanes, hexachlorobenzene, 1,1,1-trichloro-2,2-bis(4-chlorophenyl)ethane (DDT), DDT-metabolites, three brominated diphenylethers as well as extractable organically bound chlorine and halogenated fatty acids. A bioassay (EROD) was used for measuring the dioxin-like enzyme induction activity. Nordic Sea lodda (Mallotus villosus) oil was used as a nutritionally equivalent control, with much lower levels of halogenated organic pollutants. A full toxicological subchronic examination is reported in the following paper (Stern et al. 2002). In this study, we report that the fractionation procedure resulted in a substantial reduction of most of the pollutants in the triacylglycerol fraction, and a pronounced enrichment of most of the pollutants into the two other fractions. However, all contaminants were present at some levels in all of the fractions. The concentrations of organohalogens found in this study were representative for Baltic herring during the mid-1990s. Rat liver tissue showed similar residue patterns as the diet, with the exception of chlorinated dibenzo-p-dioxin and dibenzofuran congeners that had a higher liver retention than pesticides, chlorinated biphenyls and brominated diphenylethers.

Subchronic toxicity of Baltic herring oil and its fractions in the rat II: Clinical observations and toxicological parameters.

This study aimed to increase the knowledge about the toxicity of fish-derived organohalogen pollutants in mammals. The strategy chosen was to separate organohalogen pollutants derived from Baltic herring (Clupea harengus) fillet, in order to obtain fractions with differing proportions of identified and unidentified halogenated pollutants, and to perform a subchronic toxicity study in rats, essentially according to the OECD guidelines, at three dose levels. Nordic Sea lodda (Mallotus villosus) oil, with low levels of persistent organohalogen pollutants, was used as an additional control diet. The toxicological examination showed that exposure to Baltic herring oil and its fractions at dose levels corresponding to a human intake in the range of 1.6 to 34.4 kg Baltic herring per week resulted in minimal effects. The spectrum of effects was similar to that, which is observed after low-level exposure to pollutants such as chlorinated dibenzo-p-dioxins and dibenzofurans (CDD/F) and chlorinated biphenyls, despite the fact that these contaminants contribute to a minor part of the extractable organically bound chlorine (EOCI). The study confirmed previous findings that induction of hepatic ethoxyresorufin deethylase (EROD) activity takes place at daily intake levels 0.15 ng fish-derived CDD/F-TEQs/kg body weight. The study also demonstrated that hepatic vitamin A reduction takes place at somewhat higher daily exposure levels, i.e. 0.16-0.30 ng fish-derived CDD/F-TEQs/kg body weight. Halogenated fatty acids, the major component of EOCI, could not be linked to any of the measured effects. From a risk management point of view, the study provides important new information of effect levels for Ah-receptor mediated responses following low level exposure to organohalogen compounds from a matrix relevant for human exposure.

Acute and chronic toxicity study of fish oil and garlic combination.

The purpose of this study is to evaluate the acute and chronic toxicology of oral intake of fish oil (omega-3 fatty acid) and garlic combination food supplements. These supplements were proven to have beneficial effects on the lipid profile. Therefore, it is important to evaluate the potential long-term effects of fish oil and garlic combination supplements on the biochemistry of organ structure and function. The hypothesis to be tested was that acute and chronic high-dose supplements of fish oil and garlic may not adversely affect organ histology but may influence certain metabolic activities. A double-blind, placebo-controlled study was carried out using 28 Sprague Dawley rats separated into a placebo group (16 rats) and a supplement group (12 rats). The supplement group received the ingredients in chow inserts at a dosage that was equivalent to three times the maximum safe daily dosage for fish oil and the usual daily dosage for garlic (the maximum safe daily dosage recommended by the United States Food And Drug Administration for a 70-kg human is a total of 3 g/day intake of EPA and HDA omega-3 fatty acids from conventional and dietary sources. The usual daily garlic usage is garlic powder = 1200 mg). The study was conducted over a period of 12 months with evaluations performed at baseline, 2 months, 6 months, and 12 months. Results confirm the expected acute triglyceride, total cholesterol and LDL suppression at these higher dosages in the supplement group. Acutely and chronically, there were no differences in external appearance, level of activity, daily food consumption, blood cell count, kidney function, thyroid function, prothrombin time (PT), and activated partial prothrombin time (PTT), which remained within normal ranges in the supplement group. Organ histology remained unchanged. Although during the chronic toxicity period the triglyceride and LDL suppression persisted, it was noted that total cholesterol and HDL levels increased. The increase in cholesterol and HDL in the supplement group during chronic toxicity periods is simultaneous with loss of suppression of plasma levels of other liver function marker enzymes, ALT and AST, which are not involved in cholesterol synthesis. This possibly suggests that other liver enzymes involved in cholesterol synthesis, such as HMG-co A reductase, follow a similar escape from suppression.

Subchronic toxicity of fish oil concentrates in male and female rats.

There are an overwhelming number of reports indicating the beneficial effects of fish oil supplements in human and animal nutrition. The purpose of this study, second in a series, was to evaluate the effects, particularly those that may be harmful, of high-dose, long-term consumption of fish oil concentrates (FOC) using male and female rats. One hundred and twenty male and 120 female rats were gavaged daily with oils and oil mixtures in a volume equal to 0.5% body weight (5 mL/kg/d) for 13 weeks. The administered oils were corn oil, pure menhaden oil (MO), pure MaxEPA fish oil or different mixtures of corn oil with MO. The stability and the homogeneity of the dosing solutions were tested under study conditions. The animals received isocaloric and isonitrogenous diets throughout. Food and pure water were supplied ad libitum. At the end of the in-life phase of the study, the animals were anaesthetized with CO2 and humanely killed by exsanguination. Blood and other tissues were prepared for various clinical, histopathological and laboratory tests. Some beneficial effects of FOC, such as reduction in total serum cholesterol, in rats were confirmed. However, we also observed a significant reduction in absolute amount of serum HDL and a significant increase in relative liver and spleen weights in both sexes with the high dose of FOC. High doses of FOC (5 mL/kg/d) reduced serum iron and vitamin E concentrations. A reduction in osmotic fragility of RBC as well as an increase in RBC deformity were also observed in rats treated with high doses of FOC. These rats showed a significant overall increase in WBC count. We conclude that in rats, subchronic consumption of high levels of FOC can be beneficial but may also be harmful because of induction of clinical abnormalities including increased red cell deformity, increased relative liver and spleen weights, and reduced serum HDL, iron and vitamin E concentrations.