Lactoperoxidase-catalyzed oxidation of [4-14C]estradiol

The conversion of [4-14C]estradiol to water-soluble products by lactoperoxidase (EC 1.11.1.7) in the presence of added or generated H2O2 was studied using albumin or tyrosine as acceptor. The enzyme was able to catalyze the oxidation and binding of estradiol to albumin even in the absence of 2,4-dichlorophenol at very low concentrations of hydrogen peroxide. Other systems in which H2O2 was replaced by oxygen and Mn2+, light-sensitized riboflavin or glutathione was also shown to be active in the conversion of estradiol to water-soluble products and the effect of inhibitors on these reactions was investigated. Possible mechanisms for the peroxidase-catalyzed formation of these estradio metabolites are discussed.     

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.     

Isoprenoid biosynthesis in a marine sponge of the Amphimedon genus: incorporation studies with [1-14C] acetate, [4-14C] cholesterol and [2-14C] mevalonate

1. We present quantitative evidence from incorporation of [1-14C] acetate that the enzymes to synthesise isoprenoids are present in the marine sponge Amphimedon sp. and that efficient carotenoid synthesis takes place. 2. The de novo synthesis of b,b-carotene and (3R,3'R)-zeaxanthin may occur in a chlorophyll a-producing microalgal symbiont with subsequent aromatisation to (3R)-isoagelaxanthin by the sponge itself. 3. Amphimedon sp. contains nuclear-modified sterols derived by modification of conventional dietary sterols.     

Thermophilic Anaerobic Biodegradation of [14C]Lignin, [14C]Cellulose, and [14C]Lignocellulose Preparations

Thermophilic (55°C) anaerobic enrichment cultures were incubated with [¹⁴C-lignin]lignocellulose, [¹⁴C-polysaccharide]lignocellulose, and kraft [¹⁴C]lignin prepared from slash pine, Pinus elliottii, and ¹⁴C-labeled preparations of synthetic lignin and purified cellulose. Significant but low percentages (2 to 4%) of synthetic and natural pine lignin were recovered as labeled methane and carbon dioxide during 60-day incubations, whereas much greater percentages (13 to 23%) of kraft lignin were recovered as gaseous end products. Percentages of label recovered from lignin-labeled substrates as dissolved degradation products were approximately equal to percentages recovered as gaseous end products. High-pressure liquid chromatographic analyses of CuO oxidation products of sound and degraded pine lignin indicated that no substantial chemical modifications of the remaining lignin polymer, such as demethoxylation and dearomatization, occurred during biodegradation. The polysaccharide components of pine lignocellulose and purified cellulose were relatively rapidly mineralized to methane and carbon dioxide; 31 to 37% of the pine polysaccharides and 56 to 63% of the purified cellulose were recovered as labeled gaseous end products. An additional 10 to 20% of the polysaccharide substrates was recovered as dissolved degradation products. Overall, these results indicate that elevated temperatures can greatly enhance rates of anaerobic degradation of lignin and lignified substrates to methane and low-molecular-weight aromatic compounds.     

Synthesis, storage, and stability of [4-14C]oxaloacetic acid

A simple procedure for preparing [4-14C]oxaloacetic acid based on the reaction between [14C]HCO-3 and phosphoenolpyruvate catalyzed by phosphoenolpyruvate carboxylase is described. A simple method for preparing highly purified phosphoenolpyruvate carboxylase from maize leaves is described and the degradation of oxaloacetate under conditions of varying pH and divalent metal ion concentration is reported on. [4-14C]Oxaloacetic acid is stable for several months in 0.1 M HCl solution at -80 degrees C.     

Synthesis of [1,2-3H2]cholecalciferol and metabolism of [4-14C,1,2-3H2]- and [4-14C,1-3H]-cholecalciferol in rachitic rats and chicks

[1,2-(3)H(2)]Cholecalciferol has been synthesized with a specific radioactivity of 508mCi/mmol by using tristriphenylphosphinerhodium chloride, the homogeneous hydrogen catalyst. With doses of 125ng (5i.u.) of [4-(14)C,1-(3)H(2)]cholecalciferol the tissue distribution in rachitic rats of cholecalciferol and its metabolites (25-hydroxycholecalciferol and peak P material) was similar to that found in chicken with 500ng doses of the double-labelled vitamin. The only exceptions were rat kidney, with a very high concentration of vitamin D, and rat blood, with a higher proportion of peak P material, containing a substance formed from vitamin D with the loss of hydrogen from C-1. Substance P formed from [4-(14)C,1,2-(3)H(2)]cholecalciferol retained 36% of (3)H, the amount expected from its distribution between C-1 and C-2, the (3)H at C-1 being lost. 25-Hydroxycholecalciferol does not seem to have any specific intracellular localization within the intestine of rachitic chicks. The (3)H-deficient substance P was present in the intestine and bone 1h after a dose of vitamin D and 30min after 25-hydroxycholecalciferol. There was very little 25-hydroxycholecalciferol in intestine at any time-interval, but bone and blood continued to take it up over the 8h experimental period. It is suggested that the intestinal (3)H-deficient substance P originates from outside this tissue. The polar metabolite found in blood and which has retained its (3)H at C-1 is not a precursor of the intestinal (3)H-deficient substance P.     

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 ent-kaurenol-[17-14C], ent-kaurenal-[17-14C] and ent-kaurenoic acid-[17-14C] by germinating Hordeum distichon grains

Subcellular fractions from germinated barley embryos, chloroplast preparations and whole germinating barley grains are able to carry out the conversions ent-kaurenol → ent-kaurenal → ent-kaurenoic acid → ent-hydroxykaurenoic acid, the initial steps of the biosynthetic pathway to gibberellins. Whole grains, and chloroplasts to a slight extent, incorporate radioactivity from ent-kaurenol-[17-¹⁴C] and ent-kaurenoic acid-[17-¹⁴C] into materials with similar but distinct properties from the gibberellins GA₁, GA₃, GA₄ and GA₇.     

Metabolism of [4-14C]oestradiol by oestrogen-induced uterine peroxidase

1. An enzyme that catalyses the metabolism and binding of [4-(14)C]oestradiol to protein and to other high-molecular-weight substances in the presence of H(2)O(2) was shown to be absent from the uteri of immature rats and to be induced by physiological doses of oestrogen or pregnant-mare-serum gonadotrophin. 2. The pH optimum, stability to heat and other characteristics of the uterine enzyme system as well as its subcellular distribution were determined. 3. The increase in the ability of uterine preparations to convert [4-(14)C]oestradiol into water-soluble products as a result of oestrogen treatment was accompanied by an increase in peroxidase and NADH oxidase activities and was inhibited by actinomycin D and cycloheximide. 4. The results support the proposal that the increase in peroxidase activity after oestrogen treatment might be part of an adaptive response of the uterus permitting it to bind and inactivate oestrogens and thus limit the duration of their effect upon this target tissue.     

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.