In vitro studies of iron bioavailability

The bioavailability of iron from foods is ultimately determined by interactions between iron and other components in the digestive milieu. Perhaps the most important factor is the concentration of Fe2+ during transit through the duodenum. During in vitro simulations of human digestion it is possible to probe the concentration of Fe2+, the rate of Fe2+ formation, and total iron concentration using ferrous chromogens. It is crucial, of course, that the chromogen not interfere with the redox reactions occurring during digestion. Accordingly, ferrozine was examined with regard to its ability to reduce complexes Fe3+, alter rates of Fe3+ production, detect Fe2+ present in the digestive mixture and differentiate the effects of chelating and reducing agents in the mobilization of iron from pinto beans. The chromogen was found to be free from apparent artefacts and to be a sensitive and reproducible probe of the state of iron in digestive mixtures.     

Polarized electric field effects on the regulation of succinate dehydrogenase activity in amphibian muscle and liver: kinetic study

Electropolarity treatment (0.8 V/DC/Cm) was given to the gastrocnemius muscle of Bufo melanostictus every day for 5 min. for 5 days, and kinetic study of succinate dehydrogenase (SDH) in muscle and liver was conducted with different effectors - sodium malonate, ethylene diamine tetra acetic acid (EDTA), calcium chloride (CACl2) and sodium citrate. Of the four modulators tested, the malonate and EDTA inhibited while sodium citrate and CACl2 activated the enzyme. The significance of the modulation in SDH activity to different extents was discussed.     

Radioimmunoassay of polypeptide hormones using immunochemically coated plastic tubes

A method has been developed for immobilisation of antisera on fresh plastic tubes through an immunochemical bridge. This type of immobilisation has been shown to be more consistent than direct adsorption on plastic. Such immunochemically coated antisera on plastic tube has been used in the development of a noncentrifugation radioimmunoassay. This assay system has been found to be technically as sound as the conventional method.     

An in vitro demonstration of peroxisome proliferation and increase in peroxisomal β-oxidation system mRNAs in cultured rat hepatocytes treated with ciprofibrate

Using the normal adult rat hepatocytes, plated on rat tail collagen-coated dishes and fed a chemically defined medium, we demonstrate here that ciprofibrate at 0.1 mM concentration, increases significantly the mRNA levels of fatty acyl-CoA oxidase, enoyl-CoA hydratase/3-hydroxyacyl-CoA dehydrogenase bifunctional protein, and thiolase (the three enzymes of the β-oxidation system), and causes peroxisome proliferation. Increase in mRNA levels of these genes was evident within 1 h and was maximal 24 h after the addition of ciprofibrate. In hepatocytes cultured in the absence of ciprofibrate, the basal levels of these enzymes were low and further declined with time. Concomitant treatment of hepatocytes with cycloheximide did not inhibit or superinduce the mRNA levels, indicating that this induction may represent a primary (direct) effect of this compound on the expression of these genes and does not apparently involve short-lived repressor protein(s).     

Isolation and identification of Pseudomonas spp. from Schirmacher Oasis, Antarctica.

Ten cultures of Pseudomonas spp. were established from soil samples collected in and around a lake in Antarctica. Based on their morphology, biochemical and physiological characteristics, and moles percent G + C of their DNA, they were identified as P. fluorescens, P. putida, and P. syringae. This is the first report on the identification of Pseudomonas spp. from continental Antarctica.     

Effect of lithium on regulation of two molecular forms of Na+,K(+)-ATPase in rat brain

Effects of lithium in vivo and in vitro on the two molecular forms of Na+,K(+)-ATPase in rat brain were investigated. Inhibition by strophanthidin, affinity to monovalent cations and cellular localization of the enzyme were used to differentiate the two molecular forms. K+ dependent p-nitrophenylphosphatase activity and strophanthidin inhibition studies revealed selective increase in the activity of low affinity form but not high affinity form of the enzyme following lithium treatment. Na+ sensitivity of neither forms of Na+,K(+)-ATPase was changed but K+ sensitivity of low affinity form was increased due to lithium. Lithium showed biphasic effects on low affinity form of the enzyme; activation at low concentration and inhibition at high concentration. The results suggest that lithium in vivo regulates the concentration of extra cellular potassium by selectively acting at K+ site of low affinity form of the enzyme (astroglial) but not on high affinity form (neuronal enzyme) and leading to changes in neuronal depolarization.     

Bacterial RNA isolation with one hour centrifugation in a table-top ultracentrifuge.

A procedure for the rapid preparation of cesium-chloride purified RNA from E. coli and the cyanobacterium Synechococcus sp. PCC7942 is described. Cells are lysed in modified sucrose, Triton X-100, EDTA, Tris buffer with phenol/chloroform. The cleared lysate is extracted further with phenol/chloroform and RNA is peleted by centrifugation through a 5.7 M CsCl cushion. High quality RNA can be prepared in three hours using this procedure.     

Expression of glutathione peroxidase I gene in selenium-deficient rats.

We have characterized a cDNA pGPX1211 encoding rat glutathione peroxidase I. The selenocysteine in the protein corresponded to a TGA codon in the coding region of the cDNA, similar to earlier findings in mouse and human genes, and a gene encoding the formate dehydrogenase from E. coli, another selenoenzyme. The rat GSH peroxidase I has a calculated subunit molecular weight of 22,155 daltons and shares 95% and 86% sequence homology with the mouse and human subunits, respectively. The 3'-noncoding sequence (greater than 930 bp) in pGPX1211 is much longer than that of the human sequences. We found that glutathione peroxidase I mRNA, but not the polypeptide, was expressed under nutritional stress of selenium deficiency where no glutathione peroxidase I activity can be detected. The failure of detecting any apoprotein for the glutathione peroxidase I under selenium deficiency and results published from other laboratories supports the proposal that selenium may be incorporated into the glutathione peroxidase I co-translationally.