Long-term distribution and metabolism of [1-14C]dolichol injected intravenously into rats

We have previously shown that [1-14C]dolichol mixed in vitro with rat serum and injected intravenously is rapidly cleared from the circulation and appears primarily in the liver. One day after injection the liver accounted for 80% of the isotope in whole animals, whereas after 130 days it represented only 50%. During the 130 days the specific radioactivity (dpm/g liver) decreased by more than 20-fold. In contrast, the spleen retained at 130 days 85% of the radioactivity initially present and its specific radioactivity decreased by only a factor of two. At this time small amounts of isotope were also found in carcass (internal organs removed), gastrointestinal tract and contents, and lungs. Trace amounts of radioactivity were extractable from testes and kidneys, while the heart and brain were essentially free of radioactivity. At all times after injection nearly all the radioactivity present in all tissues was still associated with dolichol. Only trace amounts of [1-14C]dolichyl fatty acyl ester and no [1-14C]phosphorylated derivatives of dolichol were present in the liver and spleen removed 156 days postinjection. Fractionation of liver between 1h and 93 days after injection suggested that [1-14C]dolichol becomes associated primarily with a lysosome-enriched fraction. The accumulation of [1-14C]dolichol in this and other subcellular compartments involved both an inward and outward flow of radioactivity, suggesting that deposition of dolichol in lysosomes is not a one-way terminal process.     

Distribution, metabolism and excretion of [1-14C]dolichol injected intravenously into rats.

[1-14C]Dolichol mixed in vitro with rat serum and injected intravenously into rats was rapidly cleared from the circulation in a manner consistent with a two-compartment model. About 80% of the radioactivity recovered from animals killed after 1 day was in the liver, with smaller amounts being found in lung, carcass (internal organs removed), gastrointestinal tract and contents, and spleen. The kidneys, testes and heart contained little radioactivity, and the brain did not appear to take up any [1-14C]dolichol. The half-life for the turnover of radioactivity from [1-14C]dolichol in tissues varied considerably, being 2 days for the lung, 17 for liver and about 50 days for the carcass. After 1 day, and also after 4 and 21 days, most of the radioactivity in all tissues was as [1-14C]dolichol and as [1-14C]dolichyl fatty acyl ester, although a small amount of incorporation of [1-14C]dolichol radioactivity into phospholipids was also observed. Faeces collected over the first 4 days after injection contained 13% of the [1-14C]dolichol dose, but urine and expired air contained only small amounts of radioactivity. Radioactivity in faeces was nearly all as unchanged [1-14C]dolichol and as [1-14C]dolichyl fatty acyl ester. The [1-14C]dolichol remaining in liver after 21 days appeared to be in a pool (possibly lysosomes) where most of it was not subject to excretion.     

In vitro oxidation of [U-14C] palmitate, [1-14C] and [6-14C] glucose in newborn and adult rats and pigs

Studies have been made on the intensity of oxidation of [U-14C]-palmitate, [1-14C]- and [6-14C]-glucose by slices of the liver and skeletal muscles of new-born, 1-day, 5-day and adult Wistar rats and domestic pigs. It was found that the level of 14CO2 production from these substrates is higher in tissues of rats than in those of pigs. At early stages of ontogenesis, in tissues of both species intensive oxidation of glucose is observed together with oxidation of fatty acids. In the course of ontogenetic development, the intensity of glucose utilization significantly decreases, whereas the level of fatty acid catabolism remains relatively unaffected.     

Metabolism of [1-14C]glyoxylate, [1-14C]-glycollate, [1-14C]glycine and [2-14C]-glycine by homogenates of kidney and liver tissue from hyperoxaluric and control subjects

1. The metabolism of [1-(14)C]glyoxylate to carbon dioxide, glycine, oxalate, serine, formate and glycollate was investigated in hyperoxaluric and control subjects' kidney and liver tissue in vitro. 2. Only glycine and carbon dioxide became significantly labelled with (14)C, and this was less in the hyperoxaluric patients' kidney tissue than in the control tissue. 3. Liver did not show this difference. 4. The metabolism of [1-(14)C]glycollate was also studied in the liver tissue; glyoxylate formation was demonstrated and the formation of (14)CO(2) from this substrate was likewise unimpaired in the hyperoxaluric patients' liver tissue in these experiments. 5. Glycine was not metabolized by human kidney, liver or blood cells under the conditions used. 6. These observations show that glyoxylate metabolism by the kidney is impaired in primary hyperoxaluria.     

Absorption and distribution of sodium [2-14C]barbital in tissues of normal and dystrophic mice

The absorption and distribution of [2-14C]barbital after oral administration was studied in various tissues, including skeletal muscle, of normal and dystrophic mice. There appeared to be a more rapid gastric emptying in the mutant homozygote as reflected in lower levels of the drug recuperated from the gastrointestinal tract. This resulted in initially higher plasma and tissue concentrations of barbital in the dystrophic mice. Two hours after oral administration, this kinetic profile was reversed so that less barbital remained in the tissues of the dystrophic mouse. The tissue:plasma concentration ratios were consistently, but not significantly, higher in all tissues of the dystrophic animals. Analysis of the half-life of the drug in both groups suggests that there is an increase in the distribution volume of barbital in the dystrophic mice. The phenomenon of more rapid absorption of the barbiturate seems to be more consistent as the symptoms of the disease progress. The altered absorption and disposition of barbital in various tissues of the dystrophic mouse support the concept that a generalized multisystemic disorder may be crucial to the pathogenesis of murine muscular dystrophy, in contradistinction to a purely myogenic origin.     

Pathways of Utilization of [1-14C] Glucose and [6-14C] Glucose in Slices of Peas

Slices of ripening seeds of the pea (Pisum sativum) were suppliedwith [1-¹⁴C] G and [6-¹⁴C] G, and the S.A. was determined ofthe respirod carbon dioxide, pyruvate, and the acids of theT.C.A.C. as well as that of the individual carbon atoms of citrateand malate. The possibility that there exist active and inactive pools ofthe T.C.A.C. acids in the pea is considered and, for most ofthe acids, rejected. The results cannot be explained on the bais of the T.C.A.C.because the S.A. of the carbon dioxide liberated was some tentimes higher than could have come from the malate via the T.C.A.C.,too much ¹⁴C accumulated in the cycle acids to have come frompyruvate by the operation of the T.C.A.C., and the patterrnof label in citrate and malate was different from that expected. An alternative explanation is put forward based on the oxidationof glucose by the P.P.P. and movement of ¹⁴C by a series ofrapid isotope exchange reactions.     

Bioconversion of alpha-[14C]zearalenol and beta-[14C]zearalenol into [14C]zearalenone by Fusarium roseum 'Gibbosum'

Cultures of Fusarium roseium 'Gibbosum' on rice were treated with [14C]zearalenone, alpha[14C]zearalenol, or beta-[14C]zearalenol to determine whether a precursor-product relationship exists among these closely related fungal metabolites. Culture extracts were purified by silica gel column chromatography and fractionated by high-pressure liquid chromatography, and the level of radioactivity was determined. Within 7 days, the beta-[14C]zearalenol was converted to zearalenone, and no residual beta-[14C]zearalenol was detectable. Most of the alpha-[14C]zearalenol added was also converted into zearalenone with 14 days. In cultures treated with [14C]zearalenone, no radioactivity was noted in any other components.     

Biological fate of [14C]-1-nitropyrene in rats following intragastric administration

1-Nitropyrene (1-NP), a weak carcinogen associated with diesel exhaust particles, has previously been detected in workplace atmospheres with in-use diesel engines and in the general environment. In order to gain insight in its biological fate, a single dose of [14C]-1-NP (27.6 microCi, 750 mg/kg body weight, b.w.) was administered intragastrically to rats and the presence of metabolites in blood and tissue homogenates, and radioactivity associated with blood proteins and tissue DNA, were studied. Early peak levels of radioactivity observed in blood and tissue homogenates indicated a rapid absorption of [14C]-1-NP from the gastrointestinal tract. Metabolite patterns observed in plasma, liver and kidney homogenates strongly suggested an important role of the intestinal microflora in the enterohepatic recirculation, but not in nitroreduction of 1-NP prior to absorption from the gastrointestinal tract. This might explain the low levels of radioactivity associated with blood proteins, since 1-nitrosopyrene, a product of nitroreduction of 1-NP, is likely to be involved in protein binding. Levels of radioactivity associated with plasma proteins were approximately four times higher than the levels of radioactivity associated with hemoglobin (401.0 and 84.1 pmol/g protein per micromol 1-NP kg b.w., respectively, at 24 h). Maximal 25% of the associated radioactivity was released following mild alkaline hydrolysis of either hemoglobin or plasma proteins. 1-Aminopyrene was the only released compound after hydrolysis of hemoglobin. In addition to 1-aminopyrene, two more polar unidentified metabolites were detected following hydrolysis of plasma proteins. Association of radioactivity with DNA was highest in the liver at the first moments of observation (7.4 pmol 14C Eq./mg DNA per micromol 1-NP kg b.w.), but decreased rapidly to levels lower than observed for kidney DNA (max. 3.0 pmol 14C Eq./mg DNA per micromol 1-NP kg b.w. at 24 h). In lungs 8-50 times less radioactivity was associated with DNA than observed in the liver and kidneys. The results of this study show, that 1-NP undergoes an extensive and complex biotransformation in vivo, resulting in a variety of metabolites present in blood and tissue homogenates and a diversity of blood protein adducts. Concentrations of plasma metabolites, blood protein adducts and DNA adducts were rather low. In addition, previous studies also showed relatively low concentrations of metabolites present in urine. Therefore, sensitive and selective methods will be needed in order to evaluate the biological fate of 1-NP, associated with diesel exhaust particles, in humans.     

Uptake of [2-14C]abscisic acid and distribution of 14C in apple embryos

Pyrus malus L. var. Golden delicious embryos were incubated with (±)-[2-¹⁴C]abscisic acid (ABA) (10^-5 M, 355 kBq mol^-1). After incubations of various durations, the radioactivity was measured in whole embryos, cotyledons, and embryonic axes.With either 48-h or 16-d incubation periods, the uptake of [¹⁴C]ABA depended upon the mode of culture used. The lowest values corresponded to the absorption by the embryonic axis, the highest to the absorption by the distal parts of the two cotyledons. The cotyledons accumulated the main part of the radioactivity under all conditions. Dormant and almost completely after-ripened embryos cultivated for 4 d showed no significant differences in the radioactivity uptake for identical modes of culture. There was a linear relationship between exogenous ABA concentrations (0.5 to 3.10^-5 M) and ABA uptake for embryos cultivated for 4 d with the distal parts of the cotyledons immersed in the medium.Abbreviations ABA abscisic acid. RM, RM⁺, C/2 M, and CM are different modes of embryo cultures: embryonic axis immersed alone (RM), together with the proximal parts of the cotyledons (RM⁺); distal parts of the cotyledons immersed alone (CM); embroyo flat on the medium, the root and the external surface of one cotyledon being in contact with the medium (C/2 M) - PP proximal parts of the cotyledons - DP distal parts of the cotyledons     

Biosynthesis of [14C]zearalenone from [1-14C]acetate by Fusarium roseum 'Gibbosum'.

Addition of [1-14C]acetate or [1,2-14C]acetate to actively growing cultures of Fusarium roseum 'Gibbosum' on rice yielded zearalenone with a specific activity ranging between 1.63 and 46.5 microCi/mmol.     

Bioconversion of alpha-[14C]zearalenol and beta-[14C]zearalenol into [14C]zearalenone by Fusarium roseum 'Gibbosum'.

Cultures of Fusarium roseium 'Gibbosum' on rice were treated with [14C]zearalenone, alpha[14C]zearalenol, or beta-[14C]zearalenol to determine whether a precursor-product relationship exists among these closely related fungal metabolites. Culture extracts were purified by silica gel column chromatography and fractionated by high-pressure liquid chromatography, and the level of radioactivity was determined. Within 7 days, the beta-[14C]zearalenol was converted to zearalenone, and no residual beta-[14C]zearalenol was detectable. Most of the alpha-[14C]zearalenol added was also converted into zearalenone with 14 days. In cultures treated with [14C]zearalenone, no radioactivity was noted in any other components.     

Oxidation of [1-14C]palmitoyl-CoA, [1-14C]acetyl-CoA and [2-14C]pyruvate in rabbit adrenal, liver and heart mitochondria under normal conditions and in acute stress

The acute immobilized stress was studied for its effect on oxidation rate of [1-14C]palmitoyl-CoA, [1-14C]acetyl-CoA and [2-14C]pyruvate in mitochondria of the adrenals, liver and heart of rabbits. The stress effect on the energy metabolism of adrenals is associated with an increase of the rate of CO2 formation from pyruvate and with a decrease of the rate of CO2 formation from palmitoyl-CoA. Intensified oxidation of all substrates is observed in the heart mitochondria. The processes of beta-oxidation are more active in the liver. The data obtained evidence for differences in the mechanisms of energy metabolism reconstruction under acute stress in tissues with different functional specialization.