Five 3β, 6α-dihydroxysterols in organ-pipe cactus

The sterol diol fraction from the lipids of organ-pipe cactus, Stenocereus thurberi, was separated into five compounds: macdougallin, peniocerol, cyclostenol, stenocereol and thurberol. The last three compounds have not been described before. All compounds were characterized by physical and spectroscopic properties.     

Active potassium transport by rabbit descending colon epithelium

Summary Previous studies of rabbit descending colon have disagreed concerning potassium transport across this epithelium. Some authors reported active K⁺ secretion underin vitro short-circuited conditions, while others suggested that K⁺ transport occurs by passive diffusion through a highly potassium-selective paracellular route. For this reason, we re-examined potassium fluxes across the colon in the presence of specific and general metabolic inhibitors. In addition, electrochemical driving forces for potassium across the apical and basolateral membranes were measured using conventional and ion-sensitive microelectrodes. Under normal conditions a significant net K⁺ secretion was observed (J _net ^K =–0.39±0.081 eq/cm²hr) with⁴²K fluxes, usually reaching steady-state within approximately 50 min following isotope addition. In colons treated with serosal addition of 10^–4 m ouabain,J _sm ^K was lowered by nearly 70% andJ _ms ^K was elevated by approximately 50%. Thus a small but significant net absorption was present (J _net ^K =0.12±0.027 eq/cm²hr). Under control conditions, the net cellular electrochemical driving force for K⁺ was 17 mV, favoring K⁺ exit from the cell. Cell potential measurements indicated that potassium remained above equilibrium after ouabain, assuming that passive membrane permeabilities are not altered by this drug. Net K⁺ fluxes were abolished by low temperature.The results indicate that potassium transport by the colon may occur via transcellular mechanisms and is not solely restricted to a paracellular pathway. These findings are consistent with our previous electrical results which indicated a nonselective paracellular pathway. Thus potassium transport across the colon can be modeled as a paracellular shunt pathway in parallel with pump-leak systems on the apical and basolateral membranes.     

Quantitation and interaction of glycosaminoglycans with Alcian blue in dimethyl sulfoxide solutions.

An improved method is described for the quantitation of glycosaminoglycans separatedon cellulose acetate, stained with Alcian blue, and dissolved in a dimethyl sulfoxide solution. Standard curves are shown for all eight glycosaminoglycans. It is shown that absorption at the Alcian blue orthochromatic E_max is depressed under conditions which favor formation of dye-glycosaminoglycan complexes. The interaction between Alcian blue and the eight glycosaminoglycans was studied in dimethyl sulfoxide solutions of varying composition. It was shown that the extent of complex formation depends both on the glycosaminoglycan and the composition of the dimethyl sulfoxide solution. A dimethyl sulfoxide solution which contains 0.094 m H₂SO₄ is described which maximizes dye-glycosaminoglycan dissociation and thus the absorbance. Also, an improved staining method is described which improves dye uptake by the glycosaminoglycans and consequently increases the sensitivity of glycosaminoglycan quantitation.     

Investigation of the pH dependence of proton uptake by porcine heart mitochondrial malate dehydrogenase upon binding of NADH

The pH dependence of proton uptake upon binding of NADH to porcine heart mitochondrial malate dehydrogenase (l-malate: NAD⁺ oxidoreductase, EC 1.1.1.37) has been investigated. The enzyme has been shown to exhibit a pH-dependent uptake of protons upon binding NADH at pH values from 6.0 to 8.5. Enzyme in which one histidine residue has been modified per subunit by the reagent iodoacetamide (E. M. Gregory, M. S. Rohrbach, and J. H. Harrison, 1971, Biochim. Biophys. Acta253, 489–497) was used to establish that this specific histidine residue was responsible for the uptake of a proton upon binding of NADH to the native enzyme. It has also been established that while there is no enhancement of the nucleotide fluorescence upon addition of NADH to the iodoacetamide-modified enzyme, NADH is nevertheless binding to the modified enzyme with the same stoichiometry as with native enzyme. The data are discussed in relation to the involvement of the essential histidine residue in the catalytic mechanism of “histidine dehydrogenases” recently proposed by Lodola et al. (A. Lodola, D. M. Parker, R. Jeck, and J. J. Holbrook, 1978, Biochem. J.173, 597–605) and the catalytic mechanism of “malate dehydrogenases” recently proposed by L. H. Bernstein and J. Everse (1978, J. Biol. Chem.253, 8702–8707).     

Effects of inorganic amendments (N,P and K) to soil on the rhizosphere microflora of antirrhinum plants infected withVerticillium dahliae Kleb

Summary A study of the inorganic amendments (N, P and K) to soil, and their effect on the rhizosphere microflora, as well as their relation to the control of wilt of antirrhinum plants caused byVerticillium dahliae Kleb. was done. Ammonium sulphate was the only chemical found to be significantly inhibitory toV. dahliae in vitro. Soil amendments (NPK) affected the rhizosphere microorganisms of the antirrhinum plants. Higher concentration of the chemicals were phytotoxic. It was further observed that ammonium sulphate, and the combined chemicals (NPK 25%) in soil delayed the senescence in healthy plants, suggests that chemical fertilisers affected the host plants directly. Addition of ammonium sulphate (0.25%), calcium nitrate (0.25%, 0.5%) combined NPK (0.25%) to soil caused considerable reduction in disease severity. It is assumed that this reduction may be caused by the (1) fungitoxic nature of the chemicali.e. ammonium sulphate, (2) antagonistic environment for the pathogen in the rhizosphere was boostedi.e. where calcium nitrate was added as soil amendments and (3) reduction in disease severity in soil-amended with combined NPK, may be due to the fact that antagonistic actinomycete population was boosted in the rhizosphere.     

Characteristics of androgen binding in rat adrenal tissue using [3H]methyltrienolone (R1881) as tracer

A high level of binding of [3H]methyltrienolone (R1881 = 17beta-hydroxy-17alpha-methyl-estra-4, 9, 11-trien-3-one) was found in cytosol prepared from adrenals of castrated male rats. Binding of [3H]R1881 was of high affinity (DK = 6.2 nM) and highly specific for androgens. The [3H]R1881 complex migrates at 7-9S on sucrose gradients in low ionic strength buffer and at 4-5S in buffer containing 0.4M KC1. All binding studies have been performed in parallel with rat ventral prostate and adrenal cytosol. The present data suggest the presence of an androgen binding component in rat adrenal tissue.     

Radioimmunoassay of 2-hydroxyestrone

Under the protection of ascorbic acid a 2-hydroxyestrone bovine serum albumin conjugate was prepared containing intact 2-hydroxyestrone as determined by gas chromatographymass spectrometry. Using this antigen highly specific antibodies were raised in rabbits. Cross-reactivity for 2-hydroxyestradiol and 2-hydroxyestriol was 26 and 4.5%, respectively. An assay procedure of 2-hydroxyestrone in human plasma is described. Using special precautions the assay allows the determination of 2-hydroxyestrone in plasma samples of women (50–95 pg/ml), pregnant women (105–220 pg/ml), men (45–65 pg/ml) and children (20–40 pg/ml).     

The heterocytotoxicity of human serum. II. Role of natural antibody and of the classical and alternative complement pathways.

Mouse lymphoid cells and tumor cell lines were employed as target cells for the investigation of the mechanism of heterocytotoxicity in human serum samples. It was shown that the heterocytotoxic effects were due to two differing mechanisms. Cytotoxicity was mediated in part, by activation of the alternative complement pathway on target cell membrane, a process which was antibody-independent. A second mechanism of cytotoxicity was induced by natural antibodies to a target cell, which probably mediated activation of the classical complement pathway. These data may shed light on the frequently observed cytotoxicity in mammalian sera for various target cells.     

Resolution of optical isomers by chiral high-performance liquid chromatography: separation of dihydrodiols and tetrahydrodiols of benzo[a]pyrene and benz[a]anthracene

A competitive enzyme-linked immunoassay (CELIA) for human serum angiotensin-1-converting enzyme (ACE) was developed. The sensitivity was amplified by using a secondary antibody and an avidin biotin-conjugated horseradish peroxidase complex as the enzyme-labeled reagent. This configuration was compared to three other configurations for an indirect CELIA, and was found to be the most sensitive. A sensitivity of 39 ng/ml ACE was achieved with intraassay and interassay coefficients of variation of 6.3 and 8%, respectively. CELIA will detect ACE in human serum without interference from either pharmacological or endogenous ACE inhibitors. In normal human volunteers, ACE values obtained using CELIA correlated well with values obtained by enzymatic assay.