Assessing and influencing the fractional contribution of erythrocyte-bound 59Fe to individual 59Fe tissue content in murine 59Fe distribution studies.

BACKGROUND: Murine proteins of iron homoeostasis are frequently manipulated to investigate the mechanisms of iron-distribution and their toxicological consequences. Beyond subtracting erythrocyte-bound 59Fe of the residual blood content determined for each tissue (subtraction method), procedures are needed to determine 59Fe distribution in murine models of, e.g. inflammation or diabetes that cause local hyperaemia and changes in microcirculation. AIM: Two new methods were developed to correct total 59Fe tissue content individually for erythrocyte-bound 59Fe-labelled haem iron. METHODS: Iron-deficiency and iron-overload was induced in male C57BL6 mice by feeding of respective diets. Distribution of 59Fe between different tissues was determined 24h, 14, and 28 days after intravenous injection of 59Fe trace amounts. Haem-bound 59Fe was separated from non-haem 59Fe in homogenates from all tissues by dispersion in a mix of lipophilic cyclohexanone and hydrophilic H3PO4 (separation method). Moreover, the reduction of 59Fe-labelled tissue blood content was determined in all organs after in vivo saline perfusion via the left ventricle (perfusion method). RESULTS AND DISCUSSION: 59Fe-labelled non-haem iron determined by the separation method was not significantly different from values determined by the subtraction method, except for the iron-deficient spleen 14 and 28 days after 59Fe injection when the separation method yielded approximately 20% higher values. Approximately 20% of 59Fe-labelled haem iron spilled over into the hydrophilic phase. The impact of this error decreases in parallel to 59Fe radioactivity in the residual tissue blood content: thus, it is higher in iron-deficient mice which accumulate more 59Fe in their erythrocytes than iron-adequate and iron-rich mice. For the same reason this type of error is more marked after long distribution periods and in organs with high residual blood content. Saline perfusion via the left ventricle reduced total blood content in mice to less than 10%. Liver (95%) and duodenum (94%) showed the highest removal of blood while it is lowest in spleen (66%) and lungs (69%). CONCLUSIONS: The separation and the perfusion method can be used to correct the impact of erythrocyte-bound haem iron individually. A margin of error below 10% was determined for all organs except for spleen, lungs, and fat. Both methods can be applied sequentially to obtain satisfactory results in spleen, lungs, and fat.

Different behaviour of 63Ni and 59Fe during absorption in iron-deficient and iron-adequate jejunal rat segments ex vivo.

Nickel exhibits low oral toxicity. It shares the absorptive pathways for iron, though there are substantial quantitative differences in handling of both metals. To analyse these differences more closely, jejunal segments from iron-deficient and iron-adequate rats were luminally perfused ex vivo with 59Fe and 63Ni at six different concentrations (1-500 micromo1/l) under steady state conditions. 59Fe over-all absorption increased 2.0-4.6-fold in iron-deficiency at luminal concentrations between 1 and 100 micromol/l, while 63Ni absorption increased to a much lower extent (2.6-fold at 1 micromol/l and 1.5-fold at higher luminal concentrations). Moreover, there was a 5-7-fold higher concentration for 63Ni in the jejunal tissue than in the absorbate at luminal concentrations above 50 micromol/l which was not observed at 1 micromol 63Ni/l and not for 59Fe. 63Ni tissue load showed a linear and a saturable fraction. In iron-deficiency the saturable 63Ni fraction increased 4-fold as compared to only 1.5-fold increments for 59Fe. Moreover, a substantially higher share of 63Ni was retained in the jejunal tissue at high as compare to low luminal concentrations after perfusion had been continued without luminal radioactivity. This was not found for 59Fe and suggests a concentration-dependent block of 63Ni export across the enterocytes' basolateral membrane. To explain these results one may speculate that 63Ni may bind more tightly to tissue ligands than 59Fe due to the higher thermodynamic and kinetic stability of nickel complexes. In particular, nickel may bind to a basolateral population of metal carriers and block its own basolateral transfer in a concentration-dependent manner. Tight 63Ni binding to non-specific jejunal ligands is responsible for the unaltered high linear fraction of jejunal 63Ni load in iron-deficient and iron-adequate segments. Binding of 63Ni to food and tissue ligands in the small intestine may, thus, be a likely explanation for the low oral nickel toxicity.

109Cd accumulation in the calcified parts of rat bones.

A recent epidemiological study showed an increased risk for bone fractures after chronic low-level cadmium exposure. This finding agrees with those of cadmium accumulation in rat bones after chronic oral exposure which reduced the mechanical strength of the bones. There are indications that ossicular cadmium uptake may be higher during growth and may contribute over proportion to life long cadmium accumulation in the skeleton. The present study investigates this hypothesis in 59 male Sprague-Dawley rats. 109Cd distribution showed no differences after intravenous (i.v.) administration of different doses (0.02-2.00 micromol 109Cd/kg body weight) and at different time points after injection (3 and 10 days). Iron-deficiency had no impact on 109Cd distribution, neither during growth nor in adult animals. Age, however, showed an impact on cadmium distribution. Hepatic 109Cd accumulation was significantly higher in adult rats while 109Cd distribution in the bones as well as 109Cd concentration in cortical and trabecular bone tissue was significantly higher during growth. No difference in 109Cd uptake was found between femur epiphysis and diaphysis after one-dose i.v. application, which is in contrast to earlier results after chronic oral cadmium administration to rats. This difference may be explained by a different saturation for cadmium uptake in these two bone sections. Cadmium exposure during growth, thus, seems to contribute considerably to cumulative ossicular cadmium accumulation over a lifetime and possibly to cadmium-derived bone fragility in advanced age.

Time course of arsenite-induced copper accumulation in rat kidney.

Oral administration of inorganic arsenic has been shown to lead to an accumulation of copper in the kidneys of rats and guinea pigs. However, nothing is known about the characteristics and mechanisms of this organ-specific renal copper accumulation. Many heavy metals accumulate in the kidney, either after environmental or occupational exposure. An additional accumulation of any other trace metals, even essential ones, may therefore be critical for that organ. This prompted us to follow the course of the renal copper accumulation. Rats were given daily subcutaneous doses of sodium arsenite for 12 d. Each second day, three rats were killed by exsanguination and the liver, kidneys, and blood removed and analysed for As, Cu, and other trace elements by atomic emission spectrometry. Results indicate that arsenic and copper accumulate in the kidney cortex synchroneously over time. Arsenic also accumulated in the liver and red blood cells (RBC). Copper levels in the RBC and liver as well as copper excretion into the urine were unaffected. After terminating arsenite administration, there was a slow decline in tissue levels of both arsenic and copper, a phenomenon which was parallel for both metals. Because the copper level in the liver was not affected, it is concluded from this study that renal processes and not hepatic or biliary mechanisms might be responsible for the renal copper accumulation. Furthermore, the strong linear correlation (r = 0.85) between arsenic and copper levels in the kidney during and after arsenite administration suggests a functional relationship between arsenic and copper with respect to their retention in the kidney.

Use of gamma-spectrometry for simultaneous determination of 210Pb, 73As, 109Cd, 203Hg and 59Fe distribution and excretion in rats at low doses.

gamma-Spectrometry permits the identification and quantification of different gamma-isotopes in the same aliquot. To estimate the sensitivity and discriminative power of a comparably small and inexpensive 8% germanium detector, we determined the detection limits for simultaneously applied 210Pb, 73As, 109Cd, 203Hg and 59Fe. The concentration of Fe and of each of the four potential environmental contaminants was determined in aliquots from all organs and tissues 10 days after simultaneous i.v. administration (2 micromol/kg body weight) to adult and growing iron-deficient and iron-adequate rats. Relating these values to the total size of each organ permitted to derive a whole body distribution pattern for all five isotopes in each individual animal. Cumulative renal and faecal excretion values were determined during the 10 day distribution period to calculate the half-lives for both excretory pathways for all five isotopes simultaneously. Distribution and excretion values corresponded well to literature data. Extrapolation of the results showed that the detector would be sensitive enough to discriminate and quantify the five metals at human dietary exposure levels. The results recommend to use gamma-spectrometry to investigate kinetic aspects of interactions between toxic and essential trace metals, because the method reduces the number of required animals drastically.

Food viscosity as determinant for adaptive growth responses in rat intestine: long-term feeding of different hydroxyethyl celluloses.

Carbohydrate gelling agents can be regarded as being representative for the soluble and viscous fractions of dietary fibre. Their dietary concentration affects the consistency of the ingested food as well as the dilution of nutrients and energy. By feeding hydroxyethyl cellulose (HEC) differing in molecular mass, and thus in its viscosity properties, only the consistency of the diet was modified. Three HEC (of low (LV), medium (MV) and high viscosity (HV)) were employed in a 6-week feeding study with female rats to evaluate the effect of the viscosity on adaptive responses of intestinal growth variables. Each of the HEC was added in three increasing concentrations (8, 16, and 32%, w/w) to a fibre-free control diet to yield nine test groups besides a fibre-free and an additional, fibre-rich, cereal-based control group. Except for the highest concentration of the high viscosity product (32% HV-HEC), the dilution of the energy density of the diet was almost completely compensated by an increased food intake. With the same exception, energy utilisation was not impaired and, therefore, body-weight gains in the test groups were not significantly different from that in the control. Most other changes, e.g. increases in small intestinal length, mucosal DNA content, caecal and colonic weight, not only depended on the dietary concentration but also on the viscosity of HEC in a manner that either increasing the viscosity at a given dietary concentration or increasing the dietary concentration at a given viscosity led to the same results. These findings clearly prove the important role of the viscosity of the lumen content, as a mere physico-chemical factor, in determining adaptative growth responses in the intestinal tract of rats.

Effect of loperamide on mucosal guanylyl cyclase activity in rat jejunum following Escherichia coli heat-stable toxin-induced fluid accumulation.

Loperamide has antidiarrhoeal activities against secretagogues with different mechanisms of action. Besides its opioid-like effect on intestinal motility and secretion it might exhibit additional antisecretory properties which may not be completely elucidated yet. Direct effects of loperamide on mucosal guanylyl cyclase have never been observed. We therefore investigated the effect of loperamide on intestinal fluid transport altered by heat-stable Escherichia coli enterotoxin which acts by stimulating mucosal guanylyl cyclase. Net fluid movement was determined during a 1 hr incubation period in ligated jejunal loops of anaesthetised female Wistar rats. Transport rates of net fluid movement were calculated from the loop contents measured gravimetrically at the beginning and the end of the experiments. Addition of heat-stable Escherichia coli enterotoxin to the luminal solution resulted in a net secretion of water which was significantly reversed into net absorption by loperamide. The specific activity of the particulate guanylyl cyclase was determined in mucosal scrapings of the jejunum without and with the addition of heat-stable Escherichia coli enterotoxin and/or loperamide. Additions of loperamide of up to 10 micromol/l did not change guanylyl cyclase activity. We conclude that the effect of loperamide counteracting heat-stable Escherichia coli enterotoxin induced changes of intestinal fluid transport does not involve a direct effect on guanylyl cyclase.

Active transport inhibition in rat small intestine by amphiphilic amines: an in vitro study with various local anaesthetics.

In the present investigation with rings of everted rat small intestine, amphiphilic amines such as local anaesthetics (e.g. lidocaine, procaine, tolycaine) were employed to study their effects on intestinal absorption of methyl alpha-D-glucoside, L-leucine, D-fructose, and 2-deoxy-D-glucose. All the amphiphilic amines tested, except for benzocaine, significantly inhibited Na(+)-dependent active uptake of methyl alpha-D-glucoside and L-leucine while leaving uptake of D-fructose (facilitated diffusion) and 2-deoxy-D-glucose (passive diffusion) unaffected. Increasing concentrations of lidocaine in the incubation medium inhibited the uptake of methyl alpha-D-glucoside (IC(50) approximately 3.5 mmol/L) and L-leucine (IC(50) approximately 6 mmol/L) in a dose-dependent manner. Complete reversibility of the inhibitory effect could only be achieved at short-term incubations (

Species differences in arsenic-mediated renal copper accumulation: a comparison between rats, mice and guinea pigs.

1. Administration of arsenite leads to an accumulation of copper in the rat kidney. Owing to the high retention of arsenic in the erythrocytes, however, the rat is considered to possess special toxicokinetics of arsenic and is therefore considered less comparable with other species in this respect. 2. Therefore, we compared the effect of dietary arsenite in mice and guinea pigs with that in rats. Each species was divided into four groups of animals according to the diets fed which contained increasing concentrations of sodium arsenite (NaAsO2; 0, 10, 30 and 60 mg As/kg of diet). Animals were killed after 1, 2 and 3 weeks. Tissues were sampled and analyzed for arsenic and other trace metals (Cu, Fe, Zn and Mn). 3. Compared to controls with copper levels of about 10 microg Cu/g wet wt. in the renal cortex, dietary administration of arsenite up to 60 mg As/kg of diet for 3 weeks to rats increased cortical levels to 65 microg Cu/g wet wt. An increase of renal copper levels similar to that in rats, was only observed in guinea pigs but not in mice. Renal copper accumulation in guinea pigs was time- and concentration-dependent as in rats. Feeding a diet with 60 mg As/kg for 3 weeks increased cortical copper levels from about 6 - 40 microg Cu/g wet wt. Renal copper levels in mice as well as other trace metal levels in guinea pigs and mice were not essentially altered by dietary arsenite. 4. The study shows that the renal copper-arsenic interaction is not restricted to the rat. Since in rats and guinea pigs, but not in mice, arsenic accumulated in the kidney rather similarly, a common mechanism is suggestive. As it was previously shown in rats that only inorganic arsenic is involved in this interaction, a rapid conversion of the inorganic form into methylated metabolites as in mice may diminish the extent of the renal copper accumulation whereas the lack of, or a less efficient, methylation as in guinea pigs or rats increases it.

Effect of dental materials on gluconeogenesis in rat kidney tubules.

The effect of dental composite components triethyleneglycoldimethacrylate (TEGDMA) and hydroxyethylmethacrylate (HEMA) as well as mercuric chloride (HgCl(2)) and methylmercury chloride (MeHgCl) on gluconeogenesis was investigated in isolated rat kidney tubules. From starved rats kidney tubules were prepared and isolated by digestion with collagenase. Every 10 min up to 60 min 1-ml samples were drawn from the cell suspension for quantitating the glucose content. Glucose formation in controls was 3.3 +/- 0.2 nmol/mg. per min (mean +/- SEM, n=21). Relative rates of glucose formation were obtained by expressing individual rates as a percentage of the corresponding control. X-Y concentration curves (effective concentration, EC) of the substances were calculated by fitting a four-parametric sigmoid function to the relative rates of glucose formation at various test concentrations. At the end of the incubation period cell viability was assessed by trypan blue exclusion. Cell viability decreased within the 60 min interval from 90 to approx. 80% (controls), <25 (HEMA), <20 (TEGDMA), <10 (MeHgCl), and <10% (HgCl(2)). Values of 50% effective concentration (EC(50)) were calculated from fitted curves. EC(50) values were (mmol; mean +/- SEM; n=4): HEMA, 17.7 +/- 2.9; TEGDMA, 1.8 +/- 0.2; MeHgCl, 0.018 +/- 0.0005; and HgCl(2), 0. 0016 +/- 0.0005. The toxic effect of HgCl(2) was approximately 1000 or 10 000 higher than that of the dental composite components TEGDMA or HEMA, respectively.

Kinetic analysis of 59Fe movement across the intestinal wall in duodenal rat segments ex vivo.

Duodenal segments from iron-deficient and iron-adequate rats were luminally perfused ex vivo with solutions containing 1, 10, 50, 100, 200 and 500 micromol 59Fe/l. When duodenal tissue load and mucosal-to-serosal transport had reached a steady state, perfusion was continued without luminal 59Fe supply. Mobilization of 59Fe from the duodenal tissue into the serosally released absorbate followed first-order rate kinetics, which permitted calculation of the asymptotic maximum, the rate constant, and the initial mobilization rate for tissue-to-absorbate transfer. There was no evidence for adaptation of 59Fe tissue binding in iron-deficient segments. 59Fe tissue-to-absorbate transfer increased in proportion to the mobilizable fraction of recently absorbed iron in the tissue, which is indicative of simple diffusion or carrier-mediated transport below saturation. Regulation of the mucosal uptake step appears to determine the mobilizable 59Fe fraction and thus the adaptation of the overall iron absorption process to the demand. Iron retention in the duodenal tissue and iron transfer from here into the body appear not to be either regulated or rate limited.

Longitudinal pattern of enzymatic and absorptive functions in the small intestine of rats after short-term exposure to dietary cadmium chloride.

In vitro and in situ findings suggest an impairment of digestive and absorptive functions in the small intestine by enteral cadmium salts. In the rat, diets with up to 1 mmol Cd/kg are well tolerated, however, so that the impairment might not be this drastic or compensated by adaptive changes. To elucidate whether small intestinal functions are altered, we studied the effect of dietary cadmium on the longitudinal pattern of mucosal enzymes and the in vitro uptake of methyl alpha-D-glucoside in the small intestine of female rats. Three groups of rats were employed, a control group and two groups receiving dietary CdCl2 either at 0.3 or 1.0 mmol Cd/kg of diet. Rats were killed after 1 week of feeding. The entire small intestine was removed, rinsed with ice-cold saline and divided into 12 segments of equal length. Mucosal scrapings from each segment were used to measure mucosal cadmium levels, sucrase, lactase, alkaline phosphatase, glycylleucine-hydrolase, and diamine oxidase activities. Sugar uptake was determined in vitro in all segments using everted rings tissue accumulation method. Although cadmium levels in the mucosa were high (>100 ng Cd/mg protein or >100 micromol Cd/kg WW) most enzyme activities were only slightly changed. When significant decreases in activity were detected, they were only observed in the proximal small intestine. Sugar uptake was also impaired only in proximal segments, the maximal transport capacity was reduced by approximately 20%. These findings suggest that cadmium even at dietary levels of 1 mmol/kg do not lead to a drastic impairment of digestive and absorptive functions in the small intestine and that in the rat presently observed, mostly proximal impairments are easily compensated by unaltered distal functions. Certainly, absorption of micronutrients, for which an impaired proximal function cannot be compensated, e.g. iron, might be critical in this respect.

Iron supplementation.

Iron deficiency affects approx. 20% of the world population. Due to predominantly vegetarian diets that reduce the bioavailability of food iron drastically, deficiency states are most widely distributed in developing countries. In addition, iron demand is increased by blood losses and by fast growth which increases the risk of iron deficiency in infants, young adolescents, and in menstruating and pregnant women. The symptoms of iron deficiency include impaired physical and intellectual performance. Iron supplementation may help to break the vicious cycle between inadequate nutrition and poverty. Fortification programs have to consider social and health aspects, including provision against iron overload. Excess iron stores may promote cancer and increase the cardiovascular risk, though the latter is a subject of current debate. The best approach to control such risks is individual iron supplementation geared to the demand by adequate laboratory controls. However, this approach is too costly for general application in developing countries. Food-iron fortification has successfully reduced iron deficiency in many trials and, in comparison, is much cheaper. As iron deficiency is widely distributed in most developing countries, the risk of inducing iron overload in the general population is low. Genetically determined diseases that may lead to siderosis, such as hereditary haemochromatosis or thalassaemia major, show a limited geographic and ethnic distribution. Such subgroups can be largely avoided by targeting food-iron fortification to infants, young adolescents, or pregnant women. Food vehicle and iron compound have to be matched in order to optimise iron bioavailability and to avoid rancidity in food, spoiling its taste and odour. The fortification of salt, sugar and spice mixtures or of bakery products with a short shelf-life are valid approaches to this end. Alternatively, haem iron can be used to fortify cereal-based food staples in developing countries such as tortillas or chappaties. Thus, a variety of options is available to solve the technical problems of food iron fortification. However, optimal solutions have to be tailored to the individual situation in each country.

Small-intestinal absorption of cadmium and the significance of mucosal metallothionein.

1 Although food intake is among the most important routes of Cd exposure, not many details are known about the intestinal absorption mechanisms of Cd. In this respect Cd is representative of most other nonessential, merely toxic metals. 2 Based on a concept of two distinguishable steps, intestinal absorption of Cd is characterized by high accumulation within the intestinal mucosa and a low rate of diffusive transfer into the organism. 3 After uptake into the mammalian organism, Cd is sequestered into hepatic metallothionein (MT). It is assumed that hepatic Cd-MT then gradually redistributes Cd to the kidney, which is the main target organ for chronic Cd toxicity. 4 When feeding low levels of dietary CdCl2, however, Cd accumulates preferentially in the kidney and to a lesser degree in the liver, a distribution pattern also found after intravenous and peroral administration of the Cd-MT complex itself. As dietary Cd induces intestinal MT, intestinal Cd-MT complexes could be at least partly responsible for the renal accumulation of dietary Cd. 5 For this mechanism, however, serosal release of mucosal Cd-MT is required. In fact, in vitro findings in rats reveal a concentration-dependent release of intestinal MT to the serosal side of the small intestine. These results indicate that endogenous intestinal MT may deliver Cd-MT to other inner organs, thus contributing to the preferential renal accumulation of ingested Cd.

State of iron repletion and cadmium tissue accumulation as a function of growth in young rats after oral cadmium exposure.

To check the hypothesis that adequate dietary iron supplementation reduces cadmium retention and cadmium-induced anaemia during fast growth, three different dietary iron concentrations (6 mg/kg = iron-deficient; 55 mg/kg = marginal iron supply; 180 mg/kg = luxurious iron supply) were offered to growing rats. Four groups of rats at different age (44 days approximately = 150 +/- 6 g, 49 days approximately = 180 +/- 3 g, 57 days approximately = 220 +/- 4 g, and 84 days approximately = 295 9 g) received a diet with 55 mg Fe/kg which is a marginal iron-supply during growth. Six animals in each age group were exposed to 10 mg Cd/l as CdCl2 in the drinking water for 1 week; six animals in each age group received no cadmium. In the youngest and oldest groups additional 6 animals were exposed to the same cadmium dose but received an iron-deficient (6 mg Fe/kg) and an iron-adequate diet (180 mg Fe/kg) together with corresponding controls. The state of iron repletion was monitored by the tissue iron content in liver, kidney, and duodenum as well as by the concentrations of haemoglobin, plasma iron and plasma transferrin. The youngest animals showed the highest percent weight increases. Cadmium administration influenced neither growth rates nor food and water intake. At a dietary iron content of 55 mg/kg, iron repletion was negatively correlated to growth while the cadmium content in liver and kidney showed a positive correlation. At fast growth, a dietary iron content of 6 mg/kg lead to iron-deficiency anaemia and high cadmium retention. At all dietary iron concentrations, cadmium retention as well as the cadmium-related reduction in haemoglobin concentration was significantly higher at fast growth. Adequate dietary iron supplementation reduced cadmium retention and cadmium-induced anaemia significantly. Thus, the delicate balance between iron supply and the increased iron demand during growth can be disturbed within one week by a daily cadmium intake as low as 0.7-1.3 mg Cd/kg body weight.

Arsenic-copper interaction in the kidney of the rat: influence of arsenic metabolites.

The present study was an attempt to investigate whether the renal accumulation of Cu observed in the kidneys of rats and guinea-pigs exposed to arsenite (As-III) was an effect of arsenite alone or also shared by its metabolites--arsenate (As-V), dimethylarsinic acid and monomethylarsonic acid. The four arsenic compounds were administered subcutaneously and separately to rats for 12 days in increasing doses. Kidney, liver and blood were taken and analysed for As, Cu and other trace elements by atomic emission spectrometry. Results indicate that administration of As-V leads to renal Cu accumulation similar to that observed on administration of As-III and that the accumulation in both cases is dependent on the dose of arsenic, although higher doses of As-V were required to achieve renal Cu levels comparable to that of As-III. A constant molar As:Cu ratio independent of arsenic dose was obtained in the kidney. Dimethylarsinic acid did not affect renal Cu levels at all. Administration of monomethylarsonic acid led to a slight increase in renal Cu levels which did not increase further in spite of increased doses of monomethylarsonic acid. It is concluded from these studies that neither the metabolic transformation of inorganic arsenic to its methylated products nor its metabolites (dimethylarsinic acid and monomethylarsonic acid) caused the observed renal Cu accumulation, rather, the inorganic form of As, either in the trivalent or pentavalent form is responsible.

Small-intestinal transfer mechanism of prunasin, the primary metabolite of the cyanogenic glycoside amygdalin.

1. The small-intestinal transfer of prunasin (D-mandelo-nitrile-beta-D-glucoside), the primary metabolite of amygdalin which is not absorbed in the small intestine as such, was studied in rat jejunum and ileum in vitro. 2. As shown by high pressure liquid chromatography, prunasin is transferred essentially intact across the intestinal wall, without cleavage of the glycosidic bond and thus no formation of benzaldehyde or cyanide during the mucosal passage. 3. Only the jejunal transfer of prunasin followed saturation kinetics (vmax = 1.6 mumol cm-1 min-1; KT = 460 mumol l-1) and exhibited a clearsodium-ion dependence. As indicated by the temperature dependence, only the jejunal mucosa-to-serosa transfer and the corresponding tissue uptake of prunasin required apparently high activation energies. Transfer in the terminal ileum showed diffusion characteristics. 4. Jejunal methyl alpha-D-glucoside transfer was inhibited by the presence of prunasin. Furthermore, the tissue uptake of methyl alpha-D-glucoside in rat jejunum was competitively inhibited by prunasin. 5. The results indicate that prunasin is absorbed unmetabolised in the jejunum of the rat via the transport system of glucose.

Influence of hydroxycarboxylic acids on the water solubility of various bismuth compounds.

In an equilibrium dialysis assay (bismuth being determined by atomic absorption spectrometry) a constant amount of bismuth (Bi, CAS 7440-69-9) (final maximum conc. 50 mumol Bi/l) was dialyzed against solutions with increasing concentrations of the chelators (0-25 mmol/l). At pH 5, 50% of Bi(III) nitrate was soluble in solutions with 0.3, 6.3, 13.4 and 14.6 mmol/l of edetic acid (EDTA), citric, tartaric and malic acid, respectively. At the highest concentration applied, lactic acid kept 7% of bismuth in solution. Without any chelator, bismuth was found to be essentially insoluble (limit of detection: approx. 4 nmol Bi/l). A concentration-dependent increase in solubility was found also for the other bismuth compounds; the sequence of the solubilizing capacity of the chelators was the same as for Bi(III) nitrate. Lowering the pH to 3 generally increased and raising the pH to 7, decreased the solubility of bismuth.