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2
*
Department of Biological Sciences and
College of Veterinary Medicine, Mississippi State University, MS 39762;
**
Otsu Nutraceuticals Research Institute, Otsuka Pharmaceutical Company, Shiga 520–002 Japan; and
Scientific Affairs, Pharmavite Corporation, Valencia, CA 91355
2To whom correspondence should be addressed. E-mail: rkb1{at}ra.msstate.edu.
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ABSTRACT |
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 TOP ABSTRACT INTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONLITERATURE CITED |
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KEY WORDS: • mercury • fiber • pesticides • accumulation • absorption • excretion • mice
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INTRODUCTION |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONLITERATURE CITED |
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–5
). NDO also change the densities and relative proportions of different bacterial groups in the small and large intestine of laboratory mice (6
). The higher densities of lactic acid–producing bacteria (LAB) resulting from diets supplemented with NDO, other prebiotics or probiotics are considered to provide various health benefits, including immunomodulation (7
), and to enhance resistance to enteric and systemic infections with pathogens (8
, our unpublished data). Diets supplemented with NDO enhance absorption of calcium in animal models (9
–11
) and humans (12
–14
) and also increase the capacity of the small intestine to transport glucose (15
,16
). The numerous and diverse health benefits have led to the development and marketing of various NDO as dietary supplements, despite some concerns that the health benefits have not been adequately established (17
)
However, do diets supplemented with NDO and other fiber sources provide only benefits? It can be predicted that by increasing the size and absorptive capacities of the small and large intestine, NDO would increase the absorption and accumulation of environmental contaminants. Contrary to this prediction, wheat bran reduces mercury retention in mice (18
), and the viscous indigestible polysaccharides sodium alginate and guar gum reduce the accumulation of pentachlorobenzene by rats, apparently by increasing fecal excretion (19
). Similarly, feeding rats diets with fibers from rice bran, corn, soybean, spinach and burdock increased excretion of polychlorinated dibenzofurans, polychlorinated dibenzo-p-dioxins and polychlorinated biphenyls (20
,21
). A polysaccharide (rhamnogalacturonan-II) is able to form complexes with selected cations and may be useful for reducing the absorption and increasing the elimination of dietary lead (22
). Despite these encouraging findings, there is little known about the complex relations among NDO, changes in the GIT bacteria and intestinal absorption, and accumulation and elimination of environmental contaminants.
The present study tested the hypothesis that diets supplemented with NDO that are not digested by the host but can be fermented by the resident bacteria, will result in a larger GIT with more absorptive capacities and will thereby increase the risk of accumulating environmental contaminants after chronic, low level exposure. This was accomplished by feeding mice diets with different levels of NDO from various sources for 6–8 wk before they were exposed to one of two model contaminants (mirex and methylmercury). The mice were simultaneously exposed to retinol to determine whether the NDO affected the absorption, elimination and accumulation of a lipid-soluble nutrient in a similar manner. Additional mice fed the same diets were used for measurements of small intestine dimensions, in vitro rates of absorption by the small intestine for the two contaminants, retinol, glucose and for routine microbiology of the contents of the small and large intestine.
Mirex (dodecachlorooctahydro-1,3,4-metheno-2H-cyclobuta[cd]pentalene) is an environmentally stable and lipophilic insecticide that accumulates and persists in tissues of laboratory rodents, especially adipose tissue (23
,24
) and causes hepatic carcinomas (25
). Methylmercury is an environmental pollutant of widespread concern. Methylmercury is lipophilic, highly toxic to the central nervous system (26
) and is a serious health hazard because of teratogenecity (27
). Both compounds are poorly metabolized, if at all, by eucaryotic cells, which allowed us to study accumulation and excretion without having to account for metabolic transformations by the host that can complicate interpretations.
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MATERIALS AND METHODS |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONLITERATURE CITED |
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All aspects of the research using animals were approved by the Mississippi State University Institutional Animal Care and Use Committee and were performed in facilities accredited by the American Association for the Accreditation of Laboratory Animal Care. Mice of the Swiss-Webster strain were purchased from a commercial supplier (Charles River Laboratories, Wilmington, MA). In light of age and gender influences on toxicokinetics (28
–30
), the study used female mice obtained at 32–35 d of age. Upon arrival, the mice were housed (5 per cage) in a controlled environment (22°C, 50% relative humidity, with lighting from 0700 to 1900 h). For the first week after arrival the mice were fed a commercial diet (Lab diet, PMI Feeds, St. Louis, MO). The mice were fed to excess and had constant access to water during the acclimation and experimental periods.
The cages were assigned randomly to two control and five experimental diets based on the AIN 76A (31
) formulation with fiber added at 0 or 100 g/kg dry diet (Table 1
). One control diet contained a form of cellulose (solkafloc) that is poorly fermented by the GIT bacteria and for this study, was not considered to be a NDO. The second control diet lacked a source of fiber (fiber free). The five experimental diets had the cellulose replaced entirely with the NDO [4G-ß-D-galactosylsucrose (lactosucrose)(Ensuiko suger refining, Yokohama, Japan), inulin (Raftiline HP; Orafti, Tienen, Belgium), polydextrose (Danisco culter Japan, Tokyo, Japan), indigestible dextrin (Fibersol 2; Matsutani chemical industry, Itami, Japan) or a 1:1 combination of indigestible dextrin + inulin]. The diets were fed for 6–8 wk to allow the gastrointestinal tract and the resident bacteria to adapt. Body mass of individual mice was recorded every 2 wk and food consumption by each cage was recorded every week. Each diet was fed to a total of 60 mice; 50 mice were used to measure accumulation of mirex, methylmercury and retinol, and 10 were used to study bacteriology and measure rates of absorption by the small intestine.
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After the 6- to 8-wk feeding period, groups of 25 mice from each diet were exposed for 2 wk to mirex or methylmercury. The doses were estimated from previous studies of laboratory mice to be 2.4% of the reported 50% lethal dose for mirex (32
) and 7.8% for methylmercury (33
). The levels of exposure were considered to be nontoxic, but relevant to possible environmental exposure. The contaminants were administered to the mice using "creamy" (smooth) peanut butter as the vehicle, which is readily eaten by mice.
Accumulation of the contaminants was quantified by quantifying tissue concentrations of radiolabeled compounds. Each mouse exposed to mirex was given 0.4 g of peanut butter each day to which was added 103.2 nmol of 14C mirex/g (11.1 kBq/g; Sigma Chemical, St Louis, MO) and 72.5 nmol/g unlabeled mirex (gift of Dr. Earl Alley, Mississippi State University). Mice exposed to methylmercury were fed the same amount of peanut butter with 14.7 nmol/g 14C labeled (11.1 kBq/g; American Radiolabeled Compounds, St. Louis, MO) and 18.7 nmol/g unlabeled methylmercury (Aldrich Chemical, Milwaukee, WI). Each contaminant was given in combination with supplemental retinol [12.3 nmol/(mouse · d)] to determine whether any changes in accumulation of the contaminants would be matched by a lipid-soluble nutrient. The retinol was added to the peanut butter as 5.3 nmol/g 3H-labeled (11.1 kBq/g; New England Nuclear, Boston, MA) and 30.7 nmol/g unlabeled all-trans retinol (Sigma Chemical); this doubled the levels normally present in the AIN 76A diet. For the 2-wk period, the mice received a total dose of 6.22 kBq/mouse for the radiolabeled contaminants and retinol.
During the last 2 d of the exposure period, each cage of mice was transferred to a metabolic cage for quantitative collection of urine and feces. After recording mass and volume, the feces and urine were stored at -20°C until analyzed within 2 wk of collection.
Bacteriology.
The diets were fed for 8–10 wk to the additional groups of 10 mice before they were used to study the resident bacteria. These mice were also fed 0.4 g of peanut butter per mouse each day for the final 2 wk, but were not exposed to the contaminants or the additional retinol. After the mice were killed by CO2 asphyxiation, the entire postgastric alimentary canal was removed, the associated mesentery was cut, and the lengths of the small and large intestine were recorded. Segments of 5 cm that contained digesta were removed from the middle of the small and large intestine and were immediately transferred to an anaerobic chamber. The cecum was removed from the large intestine segment, opened longitudinally and the contents were removed by rinsing with Ringer. The dry mass of the cecum was recorded after drying for 2–3 d at 60°C.
After the small and large intestine segments were placed in the anaerobic chamber, each was opened longitudinally and the contents were removed and mass recorded. The mucosa was then removed from both segments by gentle scraping using a sterile glass microscope slide. The samples of contents and mucosa were homogenized, serially diluted in reduced yeast broth and plated using an autoplater (Model 4000, Spiral Biotech, Gaithersburg, MD). Total anaerobes were enumerated using Center for Disease Control anaerobe blood agar (Beckton-Dickinson, Cockeysville, MD), total aerobes on tryptic soy agar with 5% sheep blood (Beckton-Dickinson), enterics on MacConkey II agar (Beckton-Dickinson) and lactobaccilli on lactobacilli-selective agar (33
). Aerobic bacteria were cultured in atmospheric conditions for 2–3 d, whereas the anaerobe plates were kept in the anaerobic chamber for 4–5 d. All plates were incubated at 37°C. Enterics and lactobacilli were identified by gram staining, colony morphology, aerotolerance and the Crystal System (Beckton-Dickinson). Densities were normalized to colony forming units per gram wet mass of the samples.
Measuring rates of absorption.
The everted sleeve method was used to measure rates of absorption (34
). Briefly, additional segments were taken from the proximal (originating 
10 cm distal to the pyloric sphincter) and distal (10 cm proximal to the ileocolonic junction) small intestine of the mice used to study bacteriology and placed in cold (2–4°C), aerated (95% O2 and 5% CO2) mammalian Ringers. From both segments, four sleeves of 1 cm length were mounted on stainless steel rods 1.5 or 2 mm in diameter. The sleeves were kept in cold, aerated Ringers during and after the mounting preparation. Measurements of absorption began 45 min after death. After the four tissues from each segment were equilibrated to 37°C for 4 min they were suspended for 2 min in Ringers that was aerated and stirred (1200 rpm) and contained tracer concentrations of either radiolabeled mirex (14C), methylmercury (14C), retinol (3H) or D-glucose (14C). The glucose solution also contained 50 mmol/L unlabeled D-glucose that was added by isosmotic replacement of NaCl. Tracer concentrations of 3H-labeled L-glucose were added to the glucose solution for correction of D-glucose in the extracellular fluid and passively absorbed. Tracer concentrations of 14C polyethylene glycol were added to the mirex and methylmercury solutions, whereas 3H polyethylene glycol was added to the solution with retinol for correction of labeled isotope in the extracellular fluid. Tissues exposed to the glucose solution were rinsed for 20 s in cold Ringers. After the incubation, the tissues were carefully blotted to remove adherent fluid, removed from the rods and placed in tared vials. Wet mass was recorded, the tissues were oxidized, and the associated radioactivity collected and quantified by liquid scintillation counting. Rates of absorption were calculated and normalized to wet tissue mass. Values for glucose represent carrier-mediated transport via the Na-dependent transporter, sodium-glucose cotransporter 1, whereas those for the contaminants and retinol represent the sum of carrier-mediated processes and carrier-independent passive absorption.
Accumulation of contaminants and retinol.
After the 2-wk exposure to the contaminants and retinol, the mice were deprived of food for 5–8 h and killed by CO2 asphyxiation. Immediately after death, the postgastric alimentary canal and associated mesentery, liver, gall bladder, brain, and kidney were removed for analysis. The mesentery was removed from the small and large intestine after which the two segments were separated, the respective lengths were recorded, the contents were removed by gently squeezing each segment with a forceps and the wet mass of the large intestine was recorded. The wet mass of the mesentery, other organs and contents of the large intestine was recorded. The collected tissues were stored at -20°C until samples of 50–1000 mg were oxidized (Model 307, Packard Instrument, Meriden, CT). The 14C was recovered in Carbosorb E and Permafluor E (Packard) and the 3H was collected in Monophase S (Packard). Radioactivity was determined by liquid scintillation counting (Tri-Carb 2100TR, Packard). Samples of the peanut butter were similarly analyzed. Accumulations of the contaminants and retinol were expressed as concentrations (nmol/g tissue) and as total organ accumulation (nmol; product of concentration times organ mass).
Chemicals.
The [14C (U)]-mirex and methyl[14C] mercury (II) chloride were purchased from Sigma and American Radiolabeled Compounds, respectively. Perkin-Elmer (New England Nuclear) provided the [11,12-3H (N)]-all trans retinol, D-[14C (U)]-glucose, L-[1-3H (N)]-glucose, [1, 2-14C]-polyethylene glycol and [1, 2-3H]-polyethylene glycol. All other chemicals were purchased from Sigma and were of the highest purity available.
Statistics.
Values presented in figures and tables are means ± SEM. One-way ANOVA was used to search for diet effects. Bacterial densities were log transformed for the analysis. When a significant diet effect was detected, differences among diet groups were identified using Duncan’s multiple range test. Student’s t tests were used to compare contaminants in feces and large intestine within diet groups. For all tests, P < 0.05 was used as the critical level of significance. The statistical analyses were performed using the Statistical Analysis System, Version 7.0 (SAS Institute, Cary, NC).
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RESULTS |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONLITERATURE CITED |
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Cumulative food consumption throughout the experimental period, body mass at death and growth during the feeding period did not vary among the experimental diet groups (Table 2
), nor were differences detected between mice exposed to mirex and methylmercury (data not shown). The small and large intestine of mice fed the diets with NDO were longer than those of mice fed the diet lacking fiber. Values for mice fed the diet with cellulose were intermediate; they were longer than those from mice fed the fiber-free diet, and for most comparisons, shorter than those of mice fed the diets containing NDO. Small intestine mass did not vary among groups, but the large intestine and cecum were heavier in mice fed diets with NDO compared with the cellulose and fiber-free groups, which did not differ. Among the four NDO groups, the dry mass of cecal tissue was greater for mice fed inulin. Large intestine mass was greater for mice fed polydextrose compared with those fed the other diets.
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. Mesenteric mass was heavier in mice fed the fiber-free diet, due to what appeared visually to be a greater accumulation of fat (not quantified). Gall bladder mass for mice fed the diets with NDO was heavier than those of mice fed the cellulose and fiber-free diets (P < 0.05 for pooled data), with individual differences detected for inulin and inulin with indigestible dextrin. Urine volume and fecal mass.
The volume of urine collected during the final 2 d of exposure did not differ among groups (Table 3
). Mass of the 2-d fecal collection was higher for mice fed the diets with fiber compared with the fiber-free diet, and was heaviest for mice fed the cellulose diet. At death, the mass of contents in the large intestine was greatest for mice fed the diets with inulin and polydextrose, with the lowest values recorded in the fiber-free group.
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Densities of the groups of bacteria studied were higher in the contents of the large intestine compared with the small intestine (P < 0.01), but a diet and site interaction was not detected (data not presented). Mice fed the control diet with cellulose had lower densities of anaerobes, aerobes and lactobacilli than those fed the five experimental diets with NDO (P < 0.05). Mice fed the fiber-free control diet also had lower densities of anaerobes (P < 0.05 for comparison with the five experimental NDO diets), but aerobe and lactobacilli densities were comparable. A diet effect was not detected for the densities of enterics.
Rates of absorption by the small intestine.
Rates of carrier-mediated glucose transport [nmol/(mg · min)] by the proximal half of the small intestine did not differ among diet groups (data not presented). Glucose transport in the distal half of the small intestine was similar for mice fed the diets with NDO, and the pooled average [1.64 nmol/(mg · min)] was higher than values for mice fed the diets with cellulose (0.99) or lacking fiber (0.83). However, diet did not affect total capacities (nmol/min) of the small intestine to transport glucose.
Overt diet differences were not detected for absorption rates and capacities for either contaminants or retinol (sum of possible carrier-mediated uptake plus carrier-independent influx). However, mirex absorption by the distal half of the small intestine of mice fed the diets with fiber [7.78 pmol/(mg · min) ± 2.02] was lower than in mice fed the fiber-free diet (21.07 ± 9.36; P < 0.05). Methylmercury absorption by the distal half of the small intestine from mice fed the diets with fiber (1.01 ± 0.08) was lower than for the fiber-free group (2.23 ± 0.68; P < 0.05), but a fiber influence was not detected for the proximal half of the small intestine.
Retinol absorption by the proximal half of the small intestine of mice fed polydextrose was greater than in those fed the other diets (P = 0.005), and tended also to be higher for the distal half (P = 0.08). As a result, total capacities to absorb retinol were greater for mice fed polydextrose compared with the other groups (P = 0.006).
Tissue accumulation of environmental contaminants and retinol
Mirex.
Concentrations of mirex detected in the tissues (nmol/g tissue) and total organ accumulation (nmol) did not differ among the diet groups, with the exception of higher mirex concentrations in the gall bladder of mice fed the diet with cellulose (Table 4
).
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) did not differ among diet groups for the brain, mesentery, kidney, liver and small intestine. Mice fed the diet with cellulose had higher methylmercury concentrations in the gall bladder and large intestine. Concentrations in the large intestines of mice fed the fiber-free and cellulose diets (4.0 ± 0.2 nmol/g) were higher than for mice fed the diets with the NDO (3.5 ± 0.1 nmol/g; P < 0.05). Similarly, the fiber-free and cellulose diets resulted in small intestine concentrations (7.6 ± 0.2 nmol/g) that tended to be higher than for the other diets (7.1 ± 0.1 nmol/g; P < 0.10).
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or to methylmercury (Table 5)
. However, when organ mass was considered, total retinol accumulation by the large intestines of mice fed the diet with polydextrose (0.14 ± 0.01 nmol/entire large intestine) exceeded values for the other diets (0.11 ± 0.003; P < 0.0001). Interestingly, when the seven tissues were considered collectively, total retinol accumulation by mice exposed to methylmercury (55.1 ± 0.81 nmol for the seven diets) was lower than in mice exposed to mirex (70.6 ± 0.9 nmol; P < 0.05). The difference between contaminants was significant for the small intestine, liver and mesentery. Urine and fecal excretion of environmental contaminants and retinol
Mirex.
Urinary excretion of mirex was not affected by the experimental diets (data not shown). However, the concentration of mirex in the contents of the large intestine at the time of death was higher for mice fed the diet with inulin (Table 6
). Mirex levels in the 2-d fecal collections were higher compared with the contents of the large intestine (P < 0.05) and tended to be higher for mice fed lactosucrose compared with the other treatments (P = 0.057 by ANOVA). Total mirex excretion, based on the sum of the 2-d fecal and urinary losses, did not differ among diet groups.
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). Concentrations of methylmercury were higher in the feces than in the contents of the large intestine (P < 0.05). The combined 2-d urinary and fecal losses of methylmercury did not differ among groups.
Retinol.
Diet differences were not detected for urine concentrations of retinol for mice exposed to both contaminants (data not shown). However, retinol concentrations in the contents of the large intestine generally were greater in mice fed inulin compared with those fed the other diets, with the difference significant for the mice coexposed to mirex (Table 6)
. Fecal concentrations and the combined losses of retinol in the urine and feces generally were lower for mice fed polydextrose, and significantly so for those exposed to methylmercury (P < 0.05).
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DISCUSSION |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONLITERATURE CITED |
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,35
,36
), but even more so in the gall bladder (present study). Mirex is preferentially deposited in adipose and hepatic tissue (24
, present study), and after deposition in the body, there is very little excretion (23
,24
). Except for tissues associated with the alimentary canal, the different amounts (0 vs. 100 g/kg) and sources of fiber did not affect accumulation (nmol/g) by individual tissues for either environmental contaminant after 2 wk of exposure. Despite a lack of diet differences for total accumulation of mirex and retinol (nmol; calculated by summing the products of organ concentration times mass for each of the intact tissues), total methylmercury accumulation was lower when mice were fed diets with the NDO, as reported previously for wheat bran (18
).
Dietary fiber is critical for maintaining normal GIT structure and functions (37
,38
), and fermentable forms influence nutrient and mineral absorption in humans and animal models (9
–18
,39
). Fermentation of NDO and other fibers increases lumenal concentrations of short-chain fatty acids (SCFA), notably butyrate (40
), which is reported to stimulate growth of the small intestine (41
,42
) and the associated mucosa (43
). The longer small intestine and increased dimensions of the large intestine and cecum of mice fed the diets with NDO compared with those fed the fiber-free diet are consistent with these findings and other reports (44
). Moreover, the changes in the chemical characteristics of the lumenal environment caused by bacterial fermentation (e.g., lower pH, accumulation of SCFA) may influence characteristics of the mucosa and the availability of solutes.
Carrier systems have not been described for mirex and methylmercury. Although intestinal absorption of both compounds is considered to be passive and concentration dependent, methylmercury is absorbed rapidly and almost completely (36
,45
). This led to the expectation that rates of absorption per unit intestine would not vary among diets and the hypothesis that mice with larger intestines due to eating a diet with NDO would accumulate more mirex and methylmercury. Instead, mice fed the diets with NDO had total small intestine absorption capacities for mirex (14 ± 1.2 nmol/min; pooled results for the six diets with fiber) and methylmercury (2.1 ± 0.2 nmol/min) that were lower than those for mice fed the fiber-free diet (33.5 and 3.6 nmol/min, respectively; P < 0.05). Of particular interest is that the differences were caused by lower rates of absorption per unit mass in the distal intestine for methylmercury (P < 0.05 for each of the 5 NDO diets compared with the fiber-free diet) and for mirex (P < 0.05; pooled results for the 5 NDO-containing diets compared with the fiber-free diet; individual comparisons were not significant). The mechanism responsible for the lower rates of mirex and methylmercury absorption in the distal small intestine of mice fed diets with fiber is uncertain.
Although retinol is a lipid-soluble vitamin, a carrier-mediated process of absorption may be involved (46
). Unlike the contaminants, an effect of diet on absorption was not detected, even in the distal small intestine. Similarly, a diet effect was not detected for glucose transport in any region. Interestingly, in vivo accumulation of retinol by the seven tissues (not including the feces) of mice exposed to methylmercury (27.2 ± 0.4% of the total retinol ingested during the 2-wk exposure; pooled data for all diets) was less than for mice exposed to mirex (34.9 ± 0.4%; P < 0.001). This is consistent with inhibition of transport processes by mercury and the greater urinary losses of retinol of mice exposed to methylmercury [0.101 ± 0.003 µg/(d · mouse); pooled data for all diets] compared with mice exposed to mirex (0.089 ± 0.004; P < 0.05).
Dietary fiber can lower in vivo absorption of environmental contaminants by reducing GIT residence time (20
) and by adsorption processes (47
). Because NDO are metabolized almost completely by the GIT bacteria (1
), adsorbed contaminants would become available as fermentation proceeded. However, because the majority of bacterial fermentation occurs in the large intestine, contaminants adsorbed by fiber would be less available for absorption in the small intestine.
The best-known responses to NDO are the those of the fecal bacteria of animals and humans, with the increased densities of LAB of particular interest because of the purported health benefits (1
–5
). Changes in the bacterial assemblages influence the metabolic characteristics and architecture of the GIT mucosa (48
,49
). Similar to our previous study (6
,50
), the GIT of mice fed the diets with NDO tended to harbor higher densities of anaerobes compared with those fed the fiber-free and cellulose diets. The higher total densities of bacteria, hence biomass, in response to the NDO may represent a larger "sink" for environmental contaminants. Adsorption of contaminants onto bacteria, although variable among bacterial species and contaminants (51
), would decrease availability to the host. Therefore, diets that promote higher rates of proliferation and greater densities of bacteria with the greatest capacities to bind contaminants should decrease availability to the host. Because of the increasing proximal-to-distal gradient for bacterial densities, this mechanism for reducing contaminant availability and accumulation would be of greater importance in the distal small intestine and the large intestine compared with the proximal small intestine.
The numerous and extensive metabolic capacities of the bacteria and the GIT mucosa and liver of the host influence the recycling, disposal and accumulation of drugs and environmental contaminants (18
,52
–54
) and have been implicated in cancer protection (55
). For example, demethylation of methylmercury to the poorly absorbed inorganic form by members of the GIT bacteria increases fecal excretion (56
). Increasing the fiber content of the diet by adding 0–30% wheat bran increases demethylation of methylmercury, apparently by changing the metabolic characteristics of the GIT bacteria, and reduces the percentage of the mercury remaining in the large intestine from 54 to 26% (18
). Correspondingly, supplementing the diet with NDO reduced accumulation of methylmercury by the large intestine more so than cellulose, with the exception of indigestible dextrin. The importance of bacterial metabolism is also evident from the reduced fecal excretion and increased tissue deposition of methylmercury when mice are treated with antibiotics to reduce bacterial densities, hence metabolic capacities (57
). These findings validate the contention that changes in the populations and metabolic characteristics of the resident bacteria induced by diet, antibiotics or other means can alter the bioavailability and toxicity of some environmental contaminants, such as methylmercury (45
). The activities of the xenobiotic metabolizing enzymes associated with the mucosa are highest in the jejunum (54
) and based on findings for the liver enzymes may be responsive to changes in the GIT bacteria (58
). Transformation reactions, by the bacteria or host tissues, are apparently less important in modulating the bioavailability and toxicity of mirex and other contaminants (59
).
As an alternative approach to search for diet effects on organ accumulation, ratios were calculated for each tissue using the concentrations of each contaminant relative to retinol. These ratios varied among the seven tissues and for both contaminants, indicating differential deposition. The ratio for the accumulation of mirex relative to retinol in the livers of mice fed the fiber-free and cellulose diets (1.26 ± 0.01; pooled data) was higher than in mice fed the diets with NDO (1.14 ± 0.01; P < 0.05). Mirex:retinol ratios were also higher for the small and large intestinal tissues of mice fed the diet with cellulose. Interestingly, the methylmercury:retinol ratios were higher for the brains of mice fed cellulose, lower for mice fed the diets with the NDO and intermediate for mice fed the fiber-free diet. Collectively, these findings show that tissue deposition varies among lipid-soluble compounds and provide evidence that NDO may result in greater retention of nutrients compared with contaminants. However, the specific responses vary among contaminants, tissues and sources of fiber.
There is obvious interest in diets that increase the elimination of ingested or accumulated contaminants. Because the mice continued to be exposed to the contaminants during the 2-d collection period, fecal levels for both contaminants and retinol represent the sum of unabsorbed and excreted contaminant and retinol. As a result, fecal concentrations overestimate actual excretion and it is not possible to determine conclusively whether excretion was influenced by diet. Despite this limitation, the differences among diets for contaminant levels in the contents of the large intestine coincide with the enhanced fecal excretion of environmental contaminants, such as polychlorinated dibenzofurans and polychlorinated dibenzo-p-dioxins, polychlorinated biphenyl, pentachlorobenzene and lead when laboratory rodents are fed diets supplemented with various sources of fiber (19
–22
). It is not known why comparable differences were not seen for the 2-d collections of feces.
The urine concentrations for the contaminants and retinol were not indicative of a diet effect. Similar to the fecal measurements, urine concentrations represent the sum of contaminant and retinol that were eliminated after deposition plus what was absorbed, but not deposited. However, ratios for the concentration of mirex in the urine relative to the feces for mice fed the fiber-free and cellulose diets (0.022 ± 0.005; for pooled data) were higher than those for mice fed the diets with NDO (0.014 ± 0.001; P < 0.03). These ratios indicate that the majority of mirex was excreted via the feces and suggest that the routes of elimination were responsive to diet composition. Urine to fecal ratios for methylmercury were higher than those for mirex, and values for mice fed the fiber-free and cellulose diets (0.580 ± 0.016) tended to be lower than the average for the other diets (0.810 ± 0.063; P = 0.07). These findings indicate that urinary elimination is more important for methylmercury compared with mirex and can be enhanced by diets containing NDO. The ratios also indicated the NDO did not increase or alter the proportion of retinol eliminated in the feces or urine.
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ACKNOWLEDGMENTS |
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FOOTNOTES |
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3 Abbreviations used: GIT, gastrointestinal tract; LAB, lactic acid–producing bacteria; NDO, nondigestible oligosaccharides; SCFA, short-chain fatty acids.
Manuscript received 12 April 2001. Initial review completed 16 May 2001. Revision accepted 24 September 2001.
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LITERATURE CITED |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONLITERATURE CITED |
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