Effects of aging and sediment composition on hexachlorobenzene desorption resistance compared to oral bioavailability in rats.

Studies were conducted to assess the effects of black carbon, clay type and aging (1-1.5yr) on desorption and bioavailability of hexachlorobenzene (HCB) in spiked artificial sediments. Tenax (a super sorbent)-mediated desorption was used to examine the effects of these parameters on the physicochemical availability of HCB. The Tenax-mediated desorption of HCB from the four aged artificial sediments exhibited biphasic kinetics. The fast desorbing fractions ranged from 64.8% to 22.3%, showing reductions of 4.0-18.9% compared with freshly-spiked sediments. Statistical analysis on the fast desorbing fractions showed that all three treatment effects (i.e., montmorillonite clay, black carbon content, and aging) were significant. Two sediments with higher black carbon content exhibited much greater aging effects (i.e., greater reduction in fast desorbing fraction) than the other two sediments without the addition of black carbon. For both freshly-spiked and aged sediments, the desorption resistant sediment-bound HCB (i.e., slow desorbing fraction) correlated reasonably well to previously reported rat fecal elimination of HCB, which is a measure of the non-bioavailable fraction of sediment-bound HCB. A similar correlation was also observed between fast desorbing fraction and previously reported accumulation of HCB in the rat body (carcass+skin). These observations suggest that physicochemical availability, as defined by the desorption of HCB from sediments, provides a reasonable prediction of the oral bioavailability of sediment-bound HCB to rats. These results showed that montmorillonite clay, black carbon and aging reduced physicochemical availability and ultimately bioavailability of sediment-bound HCB.

Effect of organic carbon content, clay type, and aging on the oral bioavailability of hexachlorobenzene in rats.

Bioavailability of lipophilic chemicals is influenced by the physicochemical properties of soils/sediment such as particle size, pH, clay, and organic carbon content. The present study investigated the effects of sediment composition and aging on the oral bioavailability of hexachlorobenzene (HCB) in rats. Formulated sediments were prepared using various ratios of kaolinite and montmorillonite clay, sand, peat moss, and black carbon, spiked with (14)C-HCB, and orally administered to rats prior to and after one year of aging in dark at 10 degrees C. In the nonaged sediments there was a 21 to 45% reduction in the oral bioavailability of HCB when compared to the corn oil standard without any clear pattern of the impact of the sediment clay and/or organic carbon content. One year of aging resulted in statistically significant (p = 0.049) reduction in the oral bioavailability of HCB from the sediments compared to the corn oil standard and nonaged sediment indicating stronger interactions between HCB and sediment contents with aging. The mean reduction in oral bioavailability after one year of aging ranged from approximately 5 to 14% greater than that observed for nonaged sediments. The fecal elimination of the HCB-derived radioactivity from the one-year-aged sediments was much higher than the nonaged sediments, consistent with the lower absorption from the gastrointestinal tract due to lower desorption of HCB from the aged sediments. Increase in the fecal elimination and decrease in oral bioavailability of (14)C-HCB was related to the increase in clay and black carbon.

The role of type I and type II 5' deiodinases on hexachlorobenzene-induced alteration of the hormonal thyroid status.

Treatment of male Wistar rats with hexachlorobenzene (HCB) (1000 mg/kg b.w.) for 3-30 days decreases circulating levels of thyroxine (T4) but does not affect triiodothyronine (T3). Time courses were determined for 5' deiodinase type I (5' D-I) activity in thyroid, liver, and kidney and 5' deiodinase type II (5' D-II) activity in brown adipose tissue (BAT) to test the possibility that increased deiodinase activity might contribute to the maintenance of the serum T3 level. Specific 5' D-I activity was increased in the thyroid at 21 days and thereafter. No significant changes were observed in the liver, however, total 5' D-I activity in this tissue was increased at 30 days of treatment as a consequence of liver weight enhancement. HCB decreased kidney 5' D-I activity after 15 days, and BAT 5' D-II activity after 21 days of treatment. Total body 5' D-I activity was significantly increased by 30 days of HCB-treatment. HCB increased the activity of hepatic T4 uridine diphosphoglucuronosyl transferase (UDPGT) in a time-dependent manner, without changes in T3 UDPGT. We propose that increased T4 to T3 conversion in the thyroid and in the greatly enlarged liver may account for the maintenance of serum T3 concentration in hypothyroxinemic HCB-treated rats.

The aquatic vertebrate embryo as a sentinel for toxins: zebrafish embryo dechorionation and perivitelline space microinjection.

Pollution of aquatic ecosystems poses a serious threat to aquatic organisms and ultimately the entire ecosystem. Understanding how a toxin affects embryonic development is key to determining the risk a pollutant represents to the environment. Extraembryonic membranes, such as the chorion of fish eggs, provide a protective barrier between the embryo and the environment. Although the fish chorion excludes many chemical pollutants, some noxious agents can still gain access to the aquatic embryo. Therefore a monitoring system that tests the effects directly upon the embryo must be established. Although exposure to a single toxin in the laboratory can determine the concentration at which a pollutant becomes a health or environmental hazard, embryos and adults in nature are not merely affected by a single chemical, but are exposed to mixtures of different pollutants. Zebrafish (Danio rerio) and medaka (Oryzias latipes) embryos were employed for the rapid observation of the effects of single chemicals and chemical mixtures on development. Using dechorionation and a perivitelline space microinjection system, the embryos were effective sentinels for low concentrations of aquatic pollutants. The developmental effects of small quantities of toxins were observed. Embryos treated during the late gastrula stage of development with hexachlorobenzene (HCB); 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD); toluene; benzene; or mixtures of these chemicals developed cardiovascular abnormalities. The zebrafish dechorionation exposure technique, Micro Intrachorionic Zebrafish Embryo Live Laboratory test, was especially effective in testing the pollutant mixtures. Combinations of both TCDD and benzene (as well as the toluene and benzene combinations) were tested and the mixtures acted synergistically; the combinations were more toxic than either chemical by itself. Hexachlorobenzene- and TCDD-treated embryos tested positively for expression of cytochrome P450 1A indicating that the cytochrome metabolic pathways were already functional in these early embryos, and suggested that a product of the cytochrome system may be involved in HCB and TCDD pollution associated cardiovascular defects.

Liver-cell tumours in rats fed hexachlorobenzene.

All animals in a group of female Agus rats fed 100 ppm of hexachlorobenzene (HCB) in their diet for up to 90 weeks developed multiple liver-cell tumours whereas none were seen in control rats. There was no evidence of skin lesions or nervous symptoms usually seen with higher doses of HCB. A high incidence of liver-cell tumours was also seen with MRC Wistar female rats fed HCB for 75 weeks.

Dose-response relationships in hexachlorobenzene-induced porphyria.

The rate of development of hexachlorobenzene (HCB)-induced porphyria in female Wistar rats was determined using HCB dosage and porphyrin analysis protocols designed to determine factors which contribute to the delay commonly observed between initial exposure to HCB and the detection of porphyria. Measurements were made of HCB and porphyrin concentrations in the livers, kidneys, and spleens of female Wistar rats exposed continuously (up to 56 days) or for 1 day to HCB (at dietary concentrations of 1000 ppm and 100 ppm). The experiments showed that when a corn oil solution of HCB was added to the diet at a concentration of 1000 ppm, HCB accumulated rapidly in all organs, and the delay in appearance of elevated liver highly carboxylated porphyrins (HCPs) was at most 4 days (approximately 8-fold elevation of HCPs on day 4). One day of exposure to this diet was sufficient to cause elevated liver HCPs, thus showing that continuous exposure to HCB was not required to cause porphyria in this species. Solid HCB added directly to the diet (1000 ppm) resulted in less rapid HCB accumulation and less rapid development of porphyria. The experiments demonstrated that the appearance of a delay in HCB-induced porphyria in the Wistar rat is caused by the rate at which HCB is absorbed, and by using total hepatic porphyrins (rather than HCPs) as the indicator of the disorder. The experiments also showed that HCB-induced liver enlargement and neurotoxicity are not necessarily associated with the severity of porphyria.

Thyroid function and thyroxine metabolism in hexachlorobenzene-induced porphyria.

The effects of hexachlorobenzene (HCB) administration on the development of porphyria and on changes in thyroid function and thyroid hormone metabolism were examined. Female Wistar rats were treated with HCB for 1 or 8 weeks. At both treatment times liver weight was notably increased with a slight change in thyroid weight at 8 weeks. Serum thyroxine (T4) levels were depressed, whereas levels of triiodothyronine (T3) were not depressed significantly at both treatment times. One or eight weeks of HCB treatment did not alter the incorporation and distribution of [125I] into intrathyroidal aminoacids. A 50% reduction in protein bound iodine (PB[125I]) was seen in both groups of animals. HCB altered [125I]T4 metabolism in rat liver slices, increasing T4 dehalogenation. HCB administration for 1 week did not affect urinary excretion of porphyrins or their precursors, or hepatic porphyrin content. The activity of aminolaevulinate synthase was not affected, but there was a 25% and 51% inhibition in porphyrinogen carboxy-lyase (PCL) activity for the uroporphyrinogen disappearance or the coproporphyrinogen formation respectively. After 8 weeks of HCB administration the rats showed a characteristic porphyria. Our results show that HCB treatment increased hepatic thyroxine metabolism, without alterations in thyroid hormone synthesis. Serum T4 and PCL activity behaved differently in both time- and dose-dependent studies, with serum T4 being the more sensitive parameter which responded at earlier times and lower doses.

Toxicogenomics of subchronic hexachlorobenzene exposure in Brown Norway rats.

Hexachlorobenzene (HCB) is a persistent environmental pollutant with toxic effects in man and rat. Reported adverse effects are hepatic porphyria, neurotoxicity, and adverse effects on the reproductive and immune system. To obtain more insight into HCB-induced mechanisms of toxicity, we studied gene expression levels using DNA microarrays. For 4 weeks, Brown Norway rats were fed a diet supplemented with 0, 150, or 450 mg HCB/kg. Spleen, mesenteric lymph nodes (MLN), thymus, blood, liver, and kidney were collected and analyzed using the Affymetrix rat RGU-34A GeneChip microarray. Most significant (p < 0.001) changes, compared to the control group, occurred in spleen, followed by liver, kidney, blood, and MLN, but only a few genes were affected in thymus. This was to be expected, as the thymus is not a target organ of HCB. Transcriptome profiles confirmed known effects of HCB such as stimulatory effects on the immune system and induction of enzymes involved in drug metabolism, porphyria, and the reproductive system. In line with previous histopathological findings were increased transcript levels of markers for granulocytes and macrophages. New findings include the upregulation of genes encoding proinflammatory cytokines, antioxidants, acute phase proteins, mast cell markers, complements, chemokines, and cell adhesion molecules. Generally, gene expression data provide evidence that HCB induces a systemic inflammatory response, accompanied by oxidative stress and an acute phase response. In conclusion, this study confirms previously observed (immuno)toxicological effects of HCB but also reveals several new and mechanistically relevant gene products. Thus, transcriptome profiles can be used as markers for several of the processes that occur after HCB exposure.

Hexachlorobenzene-induced early changes in ornithine decarboxylase and protein tyrosine kinase activities, polyamines and c-Myc, c-Fos and c-Jun proto-oncogenes in rat liver.

Hexachlorobenzene (HCB) is a lipophilic chemical compound that is widely distributed in the environment. HCB is known to cause liver tumors in experimental animals. In the present study the in vivo effect of HCB treatment on ornithine decarboxylase (ODC) and protein tyrosine kinase (PTK) activities, free polyamine content, and c-Myc, c-Fos, and c-Jun protein levels in rat liver were investigated. HCB (1000 mg/kg body weight) increased hepatic immunodetectable c-Myc, c-Fos, and c-Jun levels after 6 h, and ODC activity and spermine and putrescine content after 18 and 24 h, while maximum stimulation of PTK activity occurred at 12 h. PTK and ODC activities varied in a dose-dependent manner. The time-course of c-Myc, c-Fos, and c-Jun protein levels was different for each proto-oncogene. They were all elevated at the second day of treatment, while only c-Fos and c-Jun remained elevated after 10 days of HCB exposure. These data jointly suggest that the increase in ODC activity may be the consequence of proto-oncogene induction. The alterations in PTK activity suggest that the growth factor signal transduction pathway may be involved in the regulation of the proto-oncogene levels or/and ODC activity. The decrease in PTK activity after the first day, even in the presence of alpha-D-Difluoromethylornithine (DFMO), an inhibitor of ODC activity, suggests that it is not regulated by polyamines. These results may be relevant to the early molecular events involved in HCB tumor promoter activity in rat liver.

The delay in polyhalogenated aromatic hydrocarbon-induced porphyria: the effect of diet preparation.

Porphyria development in female Wistar rats has been followed by dosing the animals with hexachlorobenzene (HCB) either dissolved in corn oil or as a solid mixed with the diet. It was found that the corn oil preparation resulted in much faster uptake of HCB into the liver, and much faster accumulation of liver porphyrins. Diet preparation is thus shown to be a major factor in determining whether the development of porphyria is associated with the delay phenomenon. Evidence is also presented suggesting that the neurological symptoms of porphyria are not caused by high porphyrin levels.

Modulation of hexachlorobenzene-induced hepatic porphyria by methyl isobutyl ketone in the rat.

Potential toxic interaction between hexachlorobenzene (HCB) and methyl isobutyl ketone (MiBK) was investigated using two different schedules of toxicant administration. The first schedule involved simultaneous administration of HCB (50 mg/kg/d, p.o. in 10 ml/kg corn oil at 10.00 a.m. for 5 d/wk) and MiBK (7.5 mmol/kg/d, p.o. in 10 ml/kg corn oil at 4.00 p.m. for 3 d/wk) for 6 weeks. The second schedule involved an initial dosing of 25 or 50 mg HCB/kg/d for 12 consecutive days, followed by the administration of 7.5 mmol MiBK/kg every other day for 27 days. When administered simultaneously, MiBK reduced the severity of HCB-induced porphyria, but when given sequentially after HCB accumulation, it enhanced the porphyrinogenic response. These results suggest that the effect of combined exposure to HCB and MiBK on hepatic porphyria depends on the sequence of the administration of both chemicals, and that the mechanism involved in this interaction may invoke both the induction and inhibition of specific hepatic isoenzymes by MiBK.

Studies on sex differences in excretion of sulphur derivatives of hexachlorobenzene and pentachloronitrobenzene by rats.

The appearance of sex-related differences in the excretion of sulphur derivatives of hexachlorobenzene (HCB) and pentachloronitrobenzene (PCNB) was studied in vitro and in vivo. Sexually immature rats given HCB showed initially no differences in the excretion of N-acetyl-S-(pentachlorophenyl)cysteine, but 5-8 days after weaning the urinary levels of the sulphur derivative began to increase in females until a 10-fold difference between both sexes was established. The studies in vitro and the analysis of tissues after in vivo administration of PCNB showed that conjugation with glutathione and hydrolysis of the conjugates to yield free pentachlorothiophenol do not present sex-related differences. These data tend to reinforce the view that an active renal secretory mechanism probably induced by estrogens during sexual maturation is responsible for the highly efficient excretion of sulphur derivatives of HCB and PCNB by female rats.

Hexachlorobenzene, a dioxin-like compound, disrupts auditory function in rat.

Hexachlorobenzene (HCB) is a dioxin-like compound widely distributed in the environment. In this study, we investigated the effects of HCB on the cochlea. Conscious free-moving rats were given HCB per os daily for 4 weeks at doses of 0.16, 4 or 16 mg/kg in olive oil, whereas the control group received olive oil only. The effects of HCB were evaluated at various time intervals, by measuring auditory nerve acoustic thresholds and plasma thyroid hormone concentration by radioimmunoassay. Histological evaluation involved surface preparation and scanning electron microscopy observations of cochlear hair cells. At a dose of 0.16 mg/kg, HCB induced no loss of acoustic sensitivity, whereas at 4 mg/kg, it induced cochlear sensitivity deficits at the mid-frequencies (2-16 kHz) with complete recovery once treatment was stopped. At a dose of 16 mg/kg, permanent threshold shifts were observed at all frequencies tested (from 1 to 32 kHz). Morphological studies showed no cochlear hair cell loss or alteration of stereocilia. HCB treatment reduced circulating thyroxine concentrations. Thyroidectomy had no effect on cochlear sensitivity in control animals. Thus, HCB is a potent oto-toxicant, and its ototoxicity may be independent of its thyroidal effects.

High-fat diet enhances the accumulation of hexachlorobenzene (HCB) by pregnant rats during continuous exposure to HCB.

To investigate the influence of a high-fat diet on HCB distribution and accumulation, pregnant rats in study 1 were fed a high-fat or control diet containing HCB, and, in study 2, pregnant rats were given a single HCB dose by intragastric gavage and HCB-free high-fat or control diet. In study 1, the high-fat diet group had higher HCB concentrations in fat tissues and liver than did the controls. In study 2, although the total amounts of HCB in the fat tissue and liver were greater in the high-fat diet group than in the controls, no significant differences in HCB concentration were observed between the two groups. The high-fat diet group also showed more fecal excretion of HCB. Therefore, HCB accumulation in rats fed a high-fat diet was enhanced more by continuous exposure to HCB than by administration of a single dose.

Dosing via gavage or diet for reproduction studies: a pilot study using two fat-soluble compounds-hexachlorobenzene and aroclor 1254.

The choice of a dosing route for in vivo toxicological tests is often dictated by practical constraints. Reproduction studies are particularly challenging in this regard since the determination of no-effect levels and allowable daily intakes from reproduction data encompasses exposure of the dam to the test xenobiotic prior to pregnancy, during gestation and during lactation. The fetus/infant can be exposed to the xenobiotic as well as the dam's metabolic products of the test xenobiotic during gestation and lactation. We initiated a series of two-litter, pilot reproduction studies with Sprague-Dawley and Fischer 344 rats to specifically ascertain the amount of xenobiotic and its metabolites ingested by the nursing neonate on lactation days 4, 7, 12, 17 and 21, when its dam received the xenobiotic via its diet or by gavage. The xenobiotics studied in this initial series of experiments were hexachlorobenzene (HCB) and Aroclor(R) 1254 (polychlorinated biphenyls; PCBs). The dams were dosed for 28 days, mated to untreated males and then remated approximately 2 weeks after weaning their first litter to a second untreated male. Dietary levels of 10 ppm HCB or 10 ppm PCBs, and gavage doses of 0.9 mg HCB or 0.8 mg PCBs/kg body weight/day were chosen and resulted in similar doses of HCB and PCBs per unit of the body weight of the dam during the premating period. There were no apparent toxicological effects regarding the dam nor were any of the reproduction parameters (feed consumption, dam weight, litter size, pup weight, external anomalies and day 4 viability index) significantly different from control values. Following impregnation, the body weight of the dam increased appreciably during gestation, but its feed consumption increased only slightly. During lactation, the dam's feed consumption increased markedly while its body weight increased slightly. Consequently, when dams received the xenobiotic in their diet they consumed slightly less xenobiotic per unit of body weight during gestation when compared to the gavaged dams, whereas the situation was dramatically reversed during lactation. While the greater consumption of xenobiotic by the dietary-dosed dams during lactation did result in more HCB (P

Transfer of planar and non-planar chlorobiphenyls, 4,4'-DDE and hexachlorobenzene from blood to milk and to suckling infants.

The transfer coefficients of tetra- and hexachlorobiphenyls (PCB-54; -80; -155; and -169), HCB and 4,4'-DDE between milk and blood were monitored for eight weeks in sheep previously administered with these compounds by intramuscular injection. Analyses were performed by high resolution gas chromatography. The milk/blood ratio on a fat basis was close to 1 for HCB, over 1 for 4,4'-DDE, PCB-155, and -169 and below 1 for PCB-54 and -80. It is speculated that the deviation from the ratio 1 results from the interactions of organochlorines with (lipo)proteins in blood and/or milk. In milk, the enrichment of 4,4'-DDE, PCB-155 and -169 was observed. The relative toxicity expressed by the toxic equivalent (on a fat basis) was approximately 2.5 times higher in milk than in blood.

Ultrastructural changes in liver of the rat fed hexachlorobenzene.

Changes in hepatic ultrastructure were observed in rats given hexachlorobenzene (HCB) as a component of the diet in concentrations of 5, 10, and 25 ppm for 3, 6, and 12 months. A significant change was in the cellular regions normally containing smooth endoplasmic reticulum (SER). Sometimes the SER proliferated and at other times it appeared to be replaced by quantities of storage product presumed to be glycogen. Proliferation of SER was often associated with the smaller doses of HCB and the increase in storage products with the larger doses of HCB. However, dose relationships were variable. The most significant changes were in mitochondria which became elongated or swollen. Swollen mitochondria were occasionally as large as nuclei. Usually only a fraction of the mitochondria in a cell appeared abnormal. Storage bodies 1 to 4 mum in diameter developed in some cells. These bodies were surrounded by a double membrane derived from the endoplasmic reticulum and may have been the partly digested remains of degenerated mitochondria.

Ultrastructural changes in ovarian follicles of monkeys administered hexachlorobenzene.

OBJECTIVE: To test hexachlorobenzene (HCB), an environmental pollutant, for its potential toxicity to the ovary. DESIGN: Nonhuman primates were orally administered the pollutant at doses between 0.01 and 10 mg of HCB/kg of body weight to test viability of ovarian follicles. At the end of dosing period, the monkeys received a compound that contained follicle-stimulating and luteinizing hormones to stimulate development of follicles that would be examined by electron microscopy. ANIMALS: Twenty, 6- to 13-year-old cynomolgus monkeys were randomly assigned to 5 groups. PROCEDURE: The HCB, in concentrations of 0.01, 0.1, 1.0, and 10.0 mg/kg, was orally administered with glucose in gelatin capsules for 13 weeks. Monkeys receiving capsules containing glucose only served as controls. After the 13th week, monkeys were given a compound that contained follicle-stimulating and luteinizing hormones daily during days 2 to 7 from the start of menses. On day 8 of the menstrual cycle, 5,000 IU of human chorionic gonadotropin was administered, and 35 to 38 hours later, 1 ovary from each monkey was obtained during laparotomy. Approximately 1-mm cubes of tissue from each ovary were harvested, fixed by immersion in buffered 2% glutaraldehyde, and processed for transmission electron microscopy. RESULTS: Ultrastructure of ovarian follicles was altered in the monkeys administered HCB. Lesions were observed in the follicles from monkeys given the lowest concentrations of HCB, and comprised condensed mitochondria in the developing ova and follicular cells that contained nuclei with deep indentations and abnormal accumulation of cytoplasmic lipid droplets. Alterations, such as herniation of the ooplasm, degeneration of the follicular cells, and appearance of abnormal spaces between follicular cells were observed in the follicles from monkeys of the 0.1 to 1.0 mg of HCB/kg dosage groups. The most relevant alterations were seen in the mitochondrion, an organelle that appeared to be most sensitive to the compound. Mitochondria were condensed, with abnormal intracristal spaces in the lower-dosage groups, and were markedly degenerated in the 10 mg/kg group. The effect of HCB were dose-related. CONCLUSION: The HCB is a reproductive system toxicant, and its damaging effects may be a result of augmentation of lipid peroxidation, especially in the primary follicle, which abnormally affects cellular membranes and thus, impairs their permeability.

Surface epithelium of the ovary following oral administration of hexachlorobenzene to the monkey.

Hexachlorobenzene (HCB) is a toxic and carcinogenic chemical that has been implicated in female reproductive dysfunctions, including destruction of ovarian follicles in primates. The purpose of this study was to determine the effects of HCB on ovary surface epithelium (SE). Gelatin capsules containing HCB mixed with glucose were given to 16 cynomolgus monkeys housed under controlled conditions in dosages of 0.0, 0.1, 1.0, or 10.0 mg/kg body weight daily, for 90 days; the first group served as the control. At necropsy one ovary from each animal was removed, fixed in glutaraldehyde, and processed by conventional methods for examination by transmission electron microscopy. SE from the animals in control group consisted of a single layer of squamous-to-cuboidal cells which possessed microvilli and contained cytoplasm rich in organelles; the nuclei were placed in middle of the cells. Although the types of alteration were similar in the treated groups, the degree of severity increased with increasing dose levels. In the lowest dose group (0.1 mg/kg) tested, stratification of cells was observed in some areas. Many cells were tall columnar, highly irregular in outline, and showed signs of degeneration. The nuclei had migrated toward the apical surface. Cytoplasm contained a large number of lysosomes, and numerous vesicles, which may have been swollen endoplasmic reticulum. In the 10 mg/kg group the affected cells were in advanced stages of degeneration. These observations support the evidence that HCB is a potent reproductive toxicant. Further studies are required to establish the effects of this damage on reproductive performance.

Residues in broiler chickens fed low levels of hexachlorobenzene.

Male broiler chicks (Hubbard-Hubbard) were fed graded levels (0, 0.0006, 0.006, 0.03, and 0.120 p.p.m.) of hexachlorobenzene (HCB) for 8 weeks. Gel permeation chromatography was used to prepare tissue samples from fat, heart, gizzard, leg muscle, breast muscle, kidney, and liver for analysis by gas-liquid chromatography employing electron capture detection. No treatment related trends could be determined for body or organ weights. Histopathologic examination of brain, liver, testes, pancreas, small intestine, ventriculus, spleen, kidney, lung, and heart failed to reveal lesions in either control or treated groups. A linear relationship was found between HCB accumulation in tissues and the dietary HCB level. HCB accumulation was greatest in adipose tissue followed by the heart, gizzard, leg, kidney, liver, and breast. The biomagnification of HCB in adipose tissue of broiler chickens was 11 to 18 times the concentration in the diet. For example, a concentration of 0.03 p.p.m. of HCB in the diet resulted in the accumulation of HCB in adipose tissue in excess of 0.5 p.p.m. After 7 weeks, birds were taken off rations containing HCB. After a withdrawal period of five weeks, 0.5 p.p.m. of HCB remained in adipose tissue of birds that had been fed 0.12 p.p.m. of HCB.