Bioavailability of iron from oral ferric polymaltose in humans.

The bioavailability or iron from iron(III)hydroxide polymaltose complex (ferric polymaltose, Fe-PM) was studied in human volunteers with normal or depleted iron stores as well as in patients with iron deficiency anemia. From an oral iron dose of 100 mg neutron activated Fe-PM, starved subjects with depleted iron stores absorbed significantly less (p < 0.003) 59Fe (3.91 +/- 2.24%, mean +/- SD) as compared to the reference, aqueous 59Fe(II) ascorbate solution (13.8 +/- 6.19%). Using non-radiolabeled, commercial Fe-PM no postabsorptive serum iron increase was found after oral Fe-PM (100 mg Fe dosage) in a group of 7 patients with haemorrhagic or posthaemorrhagic iron deficiency anemia. In addition, almost no haemoglobin increase was observed in 9 patients during a 4-weeks treatment period when given Fe-PM (100-300 mg Fe/d) on empty stomach, whereas subsequent treatment with ferrous sulfate (100-200 mg Fe/d) was therapeutically effective (0.15-0.23 g/dl Hb-increase/d). When given 100 or 300 mg Fe/d Fe-PM together with meal, 3 out of 6 patients showed a higher iron utilization rate (3.4-11.9%/d) than given without meal (0.5-7.5%/d). In vitro incubation studies demonstrated that Fe-PM is very stable at neutral pH. A small release of iron from the high molecular weight complex was found only at low pH (< 2). However, high amounts of ionic iron were measured in the reaction tubes after incubating solutions of Fe-PM together with ascorbic acid. This finding could explain the somewhat higher bioavailability of Fe-PM when given with vitamin C containing meals.(ABSTRACT TRUNCATED AT 250 WORDS)

Liver iron quantification: studies in aqueous iron solutions, iron overloaded rats, and patients with hereditary hemochromatosis.

For the noninvasive liver iron quantification by MRI in human iron overload diseases, fundamental proton relaxation mechanisms were studied in aqueous solutions with ferritin and other iron compounds, in experimentally iron overloaded rats, and in patients with iron overload diseases. MR-relaxation rates as a function of iron concentrations in the range of 0-7.5 mg Fe/g aqueous iron solutions, 0-5.4 mg Fe/g rat liver in vivo, and 0.16-4.9 mg Fe/g human liver in vivo were determined from multi- and sets of single-spin echo sequences (1.5 T imager). As predicted by theory, transverse relaxation rates (1/T2) in aqueous iron solutions, in liver tissue of rats, and in human liver tissue increased linearly with the iron concentration. A preliminary calibration for the liver iron quantification by MRI was performed from in vivo measurements of liver 1/T2-relaxation rates and liver iron quantification by atomic absorption spectroscopy in biopsies from 13 patients. With the single spin-echo method, precise in vivo liver iron quantification in humans also above 2.0 mg Fe/g liver tissue (T2 < 15 ms) should be accomplished on any imager with shortest spin-echo time available, at least TE < 20 ms.

205Bi/206Bi cyclotron production from Pb-isotopes for absorption studies in humans.

Pb(p, xn) thick target excitation functions were measured in the energy range 10-38 MeV in order to optimize the production of isotopically pure radiobismuth from natPb, 206Pb, and 207Pb. Additionally, the decay of Po-isotopes from deuteron irradiation of natural bismuth (209Bi) was exploited for radiobismuth production. 205Bi was produced from 206Pb at 20 MeV with only 2% of 206Bi at 4 weeks post irradiation. Bismuth compounds as used in the treatment of peptic ulcer were labeled with 205Bi for absorption studies in animals and subjects.

Performance of Helicobacter pylori acid extract and urease enzyme-linked immunosorbent assays in relation to 14C-urea breath test.

The 14C-urea breath test has been shown to be a reliable non-invasive method to detect the presence or absence of H. pylori infection. Alternatively, a number of techniques have been devised to detect circulating antibodies against H. pylori in serum, the most commonly used being enzyme-linked immunosorbent assays (ELISA). In the present study we compared the value of two ELISA antigen preparations, an acid glycine extract and a urease preparation, in relation to the results achieved in a 14C-urea breath test. Seventy-five gastroenterology outpatients were screened for the presence of H. pylori infection using the urea breath test. At the same time serum specimens were obtained. Thirty-seven patients had a positive breath test, i.e. they expired more than 2% of the oral 14C test dose within 60 min. Using the breath test as reference, sensitivity and specificity for the acid extract were 89.2% and 84.2% respectively, and for the urease ELISA 81.1% and 89.5%. Agreement between the two ELISAs was found in 82.7%, overall agreement between all three tests was observed in 77.3%. All three tests were found to be useful for monitoring therapy directed against H. pylori.

Non-transferrin-bound-iron in serum and low-molecular-weight-iron in the liver of dietary iron-loaded rats.

1. The feeding of 0.5% (3,5,5-trimethylhexanoyl)ferrocene (TMH-ferrocene) in rats resulted in a severe and progressive liver siderosis (total liver iron, 30 mg/g liver wet weight, after 30 weeks). 2. High concentrations of an iron-rich ferritin (up to 250 mg/l) were detected in serum of heavily iron-loaded rats forming a large fraction of non-transferrin-bound-iron (5000 micrograms/dl in maximum). 3. Ferritin and not haemosiderin was the major iron storage protein in the liver. 4. The total liver iron concentration (from 0.4 to > 30 mg Fe/g wet wt) but not the cytosolic low-molecular-weight-iron fraction (from 0.5 to 2.5 microM) was extremely increased during iron-loading.

Metabolism of iron from (3,5,5-trimethylhexanoyl)ferrocene in rats. A dietary model for severe iron overload.

The feeding of diets enriched with (3,5,5-trimethylhexanoyl)ferrocene (TMH-ferrocene) has been shown recently to produce a severe experimental iron overload in rats and has been considered as an adequate animal model for hereditary haemochromatosis in humans. We synthesized three 59Fe-labelled ferrocene compounds with different lipophilic characters (ferrocene, TMH-ferrocene, and 1,1'-bis(3,5,5-trimethylhexanoyl)ferrocene [(TMH)2-ferrocene]) and studied the metabolism of iron from these compounds in comparison with the hydrophilic ferrous sulphate in rats with iron deficiency, and normal and increased iron stores. The bioavailability of iron from TMH-ferrocene (whole body retention, 48% from a 5 mg Fe dose) was twice as high as from ferrocene and six times higher than from (TMH)2-ferrocene and ferrous sulphate. In contrast to the well-known iron salts (ferrous sulphate), the intestinal absorption of TMH-ferrocene iron was independent from the dose (1 or 5 mg Fe) and similar in iron-deficient and iron-loaded rats, indicating that the intestinal absorption of the TMH-ferrocene is not regulated by the body iron stores. After intestinal absorption, TMH-ferrocene iron in the portal blood is transported to the liver independently from transferrin. In contrast to absorbed ferrocene, iron from TMH-ferrocene is almost completely released from the hydrocarbon moiety within the liver. Depending on the body iron stores, TMH-ferrocene iron is then incorporated preferentially into haemoglobin (iron-deficient rats) or added to the iron stores in the liver (iron-loaded rats). A transient storage of the 59Fe-label in fat tissue was observed only from oral ferrocene but not from TMH-ferrocene. Due to the outstandingly high bioavailability of TMH-ferrocene, the chronic feeding of this compound resulted in a fast and progressive iron overload in rats (liver iron: 16.9 mg Fe/g wet weight after 10 weeks of feeding a diet containing 0.5% TMH-ferrocene), and can be regarded as the best characterized and most useful animal model for severe hepatocellular iron overload in humans.

Noninvasive liver-iron quantification by computed tomography in iron-overloaded rats.

RATIONALE AND OBJECTIVES: The benefit of computed tomography (CT) for the noninvasive determination of liver-iron concentration in human iron-overload diseases is a controversy in the literature. To study the sensitivity of CT for liver-iron quantification under experimental conditions, the authors measured single- and dual-energy CT numbers in vivo in the livers of iron-overloaded rats. METHODS: Thirty-five rats were subjected to an iron-rich diet for various periods, from 1 to 20 weeks, then scanned by single- and dual-energy CT. CT absorption was correlated to liver-iron content, which was determined by wet ashing and spectrophotometry. RESULTS: Whereas a good correlation (r = 0.99 at 96 kV; r = 0.95 at 125 kV) between CT numbers and liver-iron concentration was found, CT was insensitive to low concentrations of iron. Dual-energy CT scanning results showed greater scattering in liver-iron quantification compared with single-energy CT. CONCLUSIONS: In rats, the sensitivity of single- and dual-energy CT is too low to quantify liver iron in the diagnostically most relevant region of mild liver siderosis (1-3 mg iron/g wet weight [w.wt]).

Bismuth absorption from 205Bi-labelled pharmaceutical bismuth compounds used in the treatment of peptic ulcer disease.

The absorption of bismuth from five 205Bi-labelled pharmaceutically used bismuth compounds was studied in man. From single oral doses of all compounds under investigation only less than 0.1% bismuth was absorbed and excreted with the urine. A significantly higher absorption was observed from the colloidal bismuth subcitrate (0.042% of the dose) and the basic bismuth gallate (0.038%) than from the basic bismuth salicylate, nitrate, and aluminate (0.005-0.002%). No retention of bismuth in the whole body was found from the single dose experiment. The biologic fast-term half-lives of absorbed bismuth were calculated to be 0.12 and 1.5 days.

Iron overload of the liver by trimethylhexanoylferrocene in rats.

Iron-deficient female Wistar rats were fed a diet, which contained 0.5% trimethylhexanoylferrocene, over a 56-week period. This dietary iron loading resulted in a progressive siderosis and enlargement of the liver with a maximum iron content of 947.0 +/- 148.0 mg (vs. 0.07 +/- 0.04 mg in iron deficiency) and a maximum organ weight of 39.4 +/- 6.6 g (vs. 6.9 +/- 1.4 g in iron-deficient control rats). Up to 43 weeks, whole liver iron rose by increase in iron concentration (max. 28.0 +/- 6.1 mg/g wet weight, w.w.) as well as by enlargement of the organ. Afterwards whole liver iron increased solely by ongoing hepatomegaly. At the commencement of iron loading, stainable iron was almost exclusively stored by hepatocytes equally throughout all areas of the liver lobule. Later, the distribution of iron-loaded hepatocytes became strikingly periportal, and, in addition, Kupffer cells as well as sinus-lining endothelia began to store iron. Animals with a liver iron concentration of more than 10.4 +/- 0.75 mg/g w.w. showed no further increase in ferritin and haemosiderin within hepatocytes. Iron-burdened Kupffer cells/macrophages, however, accumulated permanently, hereby forming intrasinusoidal and portal siderotic nodules and areas. First signs of liver damage such as necrosis of single hepatocytes and mild fibrosis began at a liver iron concentration of 14.7 +/- 1.4 mg/g w.w. With advancement of iron loading, focal necrosis of hepatocytes and iron-burdened macrophages took place, and significant perisinusoidal as well as portal fibrosis developed. Cirrhosis, however, the final stage of impairment in iron overload of the liver in humans, could not be induced in this animal model up to now.

Bioavailability of bismuth from 205Bi-labelled pharmaceutical oral Bi-preparations in rats.

The bioavailability of 205Bi from various 205Bi-labelled pharmaceutical oral bismuth preparations was studied in rats. The intestinal absorption, calculated from 205Bi whole body retention and accumulated 205Bi urinary excretion, was small in general, but significantly higher (0.26-0.33% of dose) from oral bismuth citrates (basic bismuth citrate, colloidal bismuth subcitrate) as compared to basic bismuth nitrate, salicylate, gallate, and bismuth aluminate (0.04-0.11% of dose). After oral administration, the retained bismuth was mainly accumulated in the kidney, followed by bone, red blood cells and the lung. The whole body retention, faecal and urinary excretions of 205Bi were described by a three-compartment model. Biological 205Bi half-lives of 10, 36 and 295 h were derived in rats.

Inhibition of intestinal absorption and decorporation of radiocaesium in humans by hexacyanoferrates(II).

The effect of hexacyanoferrate(II) preparations, KFe[Fe(CN)6], (KFeHCF) anol Fe4[Fe(CN)6]3, (FeHCF) on intestinal radiocaesium absorption was studied in two male volunteers. The 134Cs absorption was decreased from 100 to 3-10% when 500-1000 mg KFeHCF or FeHCF were administered 10 min before the 134Cs-labelled test meal. However, when HCF was administered simultaneously with the test meal, the 134Cs absorption was decreased to only 38-63%. The biological half-time of previously absorbed 134Cs was reduced from 106 (73) to 44 (46) days by daily administration of 3 times 0.5 g KFeHCF. The 134Cs dose conversion factors lie below the values recommended by IRCP 30, indicating that the IRCP model represents a cautious description of the Cs biokinetics in our study.

Efficacy of different hexacyanoferrates(II) in inhibiting the intestinal absorption of radiocaesium in rats.

The inhibitory effect of various oral doses of different hexacyanoferrate(II) compounds (HCF) and the influence of the time interval of HCF-administration on intestinal 134Cs-absorption was studied in rats. Optimum inhibition was obtained by administration of HCF together with or 2 min before oral 134Cs loading. Using appropriate low amounts (0.1-0.5 mg) of the different HCF compounds, the inhibitory effect increased in the sequence KZnHCF less than KCuHCF less than FeHCF less than KCoHCF = KNiHCF less than NH4FeHCF = KFeHCF. Oral administration of 5 mg (0.5 mg) of KFeHCF, together with 134CsCl loading, reduces 134Cs-absorption from 41% (control) to 0.8% (2.8%). Zinc-, copper-, cobalt, and nickel hexacyanoferrates(II), despite showing a high caesium sorption capacity in vitro, were less effective in rats and are not suited for in vivo application, also because they may produce toxic side effects. As a consequence, the orally administered colloidal-soluble iron(III) hexacyanoferrates(II) (NH4Fe[Fe(CN)6] and KFe[Fe(CN)6]) have to be considered as the most valuable countermeasure against radiocaesium absorption for humans and domestic animals in the case of a severe nuclear accident in the future. Manganese oxide, a non-hexacyanoferrate(II) compound with known in vitro caesium binding capacity, showed no inhibitory effect on radiocaesium absorption in rats.

Bioavailability of iron and cyanide from 59Fe- and 14C-labelled hexacyanoferrates(II) in rats.

"Soluble" (KFe(III)[Fe(II)(CN)6]) and "insoluble Prussian blue" (Fe(III)4[Fe(II)(CN)6]3 labelled with 59Fe either in the ferric (Fe(III)) or ferro (Fe(II)) position and 14C in the cyanide group were synthesized and administered intraperitoneally or orally to adult female rats with normal body iron stores. Following i.p. injection of KFe[Fe(CN)6], the colloidal complex is disintegrated into ferric iron and hexacyanoferrate(II) anion almost completely. About 96% of the ferric iron was retained in the body. Nearly 90% of both ferrous iron and cyanide were excreted with the urine within 7 days after i.p. injection, indicating that most of the undissociated hexacyanoferrate(II) anion ([Fe(CN)6]4-) was excreted through the kidney. Only 9% of the ferrous iron from [Fe(CN)6]4- was found mainly in carcass, liver and gut. As the 59Fe/14C-ratios in organs were found close to 1.0, the dissociation of the hexacyanoferrate(II) anion can only be small in vivo. No detectable 14CO2-activity (less than 0.01%) was monitored in the breath of rats after i.p. injection of the 14C-labelled KFe[Fe(CN)6], also indicating that no significant amounts of cyanide were released after parenteral administration. After oral administration of the soluble and insoluble Prussian blue, 0.3-0.7% of the ferric iron was absorbed and retained mainly in carcass, liver and blood. Only 0.06-0.18% of the ferrous iron was absorbed and mostly excreted with the urine (0.05-0.15%), so that only 0.01-0.03% of the oral ferrous 59Fe was retained in the body after 7-10 days. Very small fractions of 14C-label from the 14CN-group of the soluble and insoluble hexacyanoferrate(II) were observed in the exhaled air (0.04-0.08% of the oral dose). From the 14CO2-exhalation, the 14C-urine excretion and the distribution of iron in blood and organs it can be concluded that the hexacyanoferrate(II) moiety disintegrated only to a small extent in the intestinal tract after oral administration. From a dose of 36 mg hexacyanoferrate(II)/kg, an amount of free (non-complex bound) cyanide can be calculated which is in maximum two orders of magnitude below the LD100-level. Thus, the very low bioavailability of iron and cyanide from hexacyanoferrate(II) compounds after oral application is demonstrated in rats. In the case of a severe nuclear accident, appropriate doses of "soluble" and "insoluble" Prussian blue can be used as safe and effective antidote against radiocaesium contamination.

Ferrioxamine and its hexadentate iron-chelating metabolites in human post-desferal urine studied by high-performance liquid chromatography and fast atom bombardment mass spectrometry.

Three iron-containing fractions were detected by high-performance liquid chromatography (HPLC) on a reverse-phase column in the 24-h urine of two patients with hereditary hemochromatosis following the injection of deferoxamine mesylate (Desferal). These fractions have virtually identical absorption spectra in the visible range, with a broad maximum around 430 nm. Molecular weight determination of these fractions was performed by fast atom bombardment mass spectrometry (FAB-MS), which gave intense ion signals for the protonated molecular ions of the intact iron chelates, namely, at m/z 614 for ferrioxamine (FOA; Mr 613), at m/z 629 for metabolite I (FOA-MI; Mr 628), and at m/z 601 for metabolite II (FOA-MII; Mr 600). The molecular weight of FOA-MI is compatible with deamination of the terminal amino function and oxidation of the adjacent carbon atom to a carboxyl group; the molecular weight of FOA-MII is compatible with loss of a C2H4 unit from FOA-MI by beta oxidation. Quantification of iron in post-Desferal urine samples either by atomic absorption spectrometry (AAS) or by HPLC leads to results which are identical within experimental error. In ten subsequent 12-h urine samples of a patient under therapy (subcutaneous infusion of Desferal), the following distribution of urinary iron was found: FOA-MI, 58.4 +/- 4.7% (arithmetic mean +/- SD); FOA, 33.2 +/- 4.9%; FOA-MII, 8.4 +/- 1.7%. Addition of 2 mM ethylenediaminetetraacetic acid (EDTA) to the chromatographic solvents was used as a stability test for FOA and its two metabolites MI and MII.(ABSTRACT TRUNCATED AT 250 WORDS)

Bioavailability of iron and cyanide from oral potassium ferric hexacyanoferrate(II) in humans.

After oral administration of 500 mg KFe[Fe(CN)6] labelled with 59Fe either in the ferric or ferrous position and with 14C in the cyanide group only 0.22% of the FeII and less than 0.04% of the FeIII were absorbed in three male volunteers. Only 2 mg non-complex bound 14C-labelled cyanide (0.03 mg CN-/kg body wt) were absorbed from 500 mg [14C]KFeHCF, which is about a factor of 20-100 below the lethal dose in humans (0.5-3.5 mg CN-/kg body wt). Therefore, iron(III) hexacyanoferrates(II) can be considered as safe antidotes, i.e. for inhibiting the intestinal absorption of radiocaesium or for accelerating the excretion of already absorbed 134/137Cs in the case of a severe nuclear accident.

Intestinal absorption of iron from 59Fe-labelled hexacyanoferrates(II) in piglets.

The intestinal absorption of 59Fe and 14C from hexacyanoferrates(II) was studied in piglets. KFeIII[FeII (CN)6] (I) and FeIII4[FeII(CN)6]3 (II) were labelled with 59Fe both in the Fe(III)-position (outside the complex anion, a) or in the Fe(II)-position (hexacyanoferrate anion, b). Labelling of the Fe(III)-position resulted in a 59Fe-absorption of 1.47% (Ia) and 1.34% (IIa), as judged by the 59Fe whole-body-retention measurement 14 days after oral administration. Even smaller amounts, 0.20% from Ib or 0.15% from IIb of the 59Fe-dose were absorbed and retained from the hexacyanoferrates(II) labelled in the Fe(II)-position. No 14CO2 was detected in the expired air of piglets after oral application of Fe4[59Fe(14CN)6]3, indicating that the amount of incorporated free cyanide ions can only be extremely small or even nil.

Iron absorption in man calculated from erythrocyte incorporation of the stable isotope iron-54 determined by fast atom bombardment mass spectrometry.

The methodology of precise isotope abundance determinations of erythrocyte iron by fast atom bombardment mass spectrometry and signal averaging is established. For the determination of the 54Fe/56Fe ratio a relative precision of 0.5% and an absolute precision of 0.03% is achieved. After oral loading with 54Fe-enriched samples in the range between 5 and 25 mg per subject, the 14-day erythrocyte incorporation of 54Fe has been determined in five individuals, namely, two adults, two children, and one infant. In the two adults, the oral dose of 54Fe was simultaneously labeled with a trace amount of carrier-free 59Fe. In these double-isotope loading tests, a good agreement was observed between the absorption data determined on the basis of whole body retention of 59Fe and on the basis of the 54Fe erythrocyte incorporation. The stable isotope methodology applied allows measurement of the iron absorption using highly enriched 54Fe at a dose of 25 mg for an adult or at a dose of 5 mg for infants of about 1 year of age.

Prevention of enteral radiocesium absorption by hexacyanoferrates(II) in piglets.

The efficacy of different hexacyanoferrates(II) in preventing the enteral absorption of 134Cs was studied in piglets. As compared to the controls, oral application of 134Cs together with KFe[Fe(CN)6], NH4Fe[Fe(CN)6], or Fe4[Fe(CN)6]3 resulted in a strong reduction of the 134 Cs-uptake by more than 97%. The decrease in enteral absorption depends on the dose of administered hexacyanoferrate(II), whereas differences between the compounds under study were small. The biological half-life of 134Cs in non-hexacyanoferrate(II) treated piglets was 21.6 +/- 3.3 days (mean +/- SD).

Intestinal absorption of 59Fe from neutron-activated commercial oral iron(III)-citrate and iron(III)-hydroxide-polymaltose complexes in man.

Commercial oral iron preparations (drops) containing trivalent iron either as a citrate (C) or hydroxide-polymaltose complex (HP) were labelled with 59Fe in the thermal neutron flux of a research reactor. No measurable differences were observed between the original commercial preparation and the neutron-activated samples. In an intraindividual comparison oral doses of 100 mg 59Fe were administered as an aqueous 59Fe(II)-ascorbate solution (= reference), C (-59Fe) and HP (-59Fe) to starved subjects with normal and depleted iron stores. Two weeks later the whole body retention of absorbed 59Fe was measured and used for the calculation of 59Fe-absorption. Subjects with normal Fe-stores absorbed means a +/- SEM +/- SD = 8.53 +/- 0.29 +/- 1.2% of the 59Fe from the aqueous 59Fe(II)-ascorbate solution, 1.58 +/- 0.12 +/- 0.49 of the C-59Fe and 0.81 +/- 0.06 +/- 0.27% of the HP-59Fe. Subjects with depleted Fe-stores absorbed means a +/- SD = 17 +/- 3% of the 59Fe from the aqueous 59Fe(II)-ascorbate solution and only 2.4 +/- 1% from the HP-59Fe. The relative bioavailabilities were reduced from 100% (reference = Fe(II)-ascorbate) to 19% for the C-Fe and to 9.5% for the HP-Fe in subjects with normal Fe-stores. In subjects with depleted iron stores the relative bioavailability was decreased from 100 to 14% for the HP-Fe. Oral iron preparations with a relative bioavailability less than 10% or less than 30% are considered to be therapeutically ineffective or insufficient, respectively.

The NIH-shift in the in vivo hydroxylation of ring-deuterated L-phenylalanine in man.

Oral loading with L-[ring-2H5]phenylalanine has been performed at a dose of 25 mg/kg for detection of heterozygotes for classic phenylketonuria. Using three differently labeled batches of ring-deuterated L-phenylalanine, quantitative analysis of deuterium-labeled L-phenylalanine and L-tyrosine in plasma revealed different label distributions. Three different reaction mechanisms for the 4-hydroxylation of L-phenylalanine to L-tyrosine were used as the basis for model calculations of the transformation of the L-phenylalanine label distribution into that of L-tyrosine. The best agreement between observed and calculated distributions was found for the mechanism involving a migration of the 4-substituent into the 3- or 5-position (NIH-shift), followed by a random loss of the 4-/3- or the 4-/5-substituent from this intermediate structure.