Changes of plasma L-arginine levels in spontaneously hypertensive rats under induced hypotension

Nicardipine, a dihydropyridine type calcium channel blocker, was infused at two flow-rates into spontaneously hypertensive (SH) and control normotensive Wistar-Kyoto (WKY) rats (young, 6-week-old and adult, 23-week-old, n = 5) under pentobarbital anesthesia, to cause hypotension. Mean arterial blood pressure and the concentrations of plasma amino acids and norepinephrine (NE) were measured before infusion and at each step of the infusion. The reduction in blood pressure caused by nicardipine induced a decrease in plasma L-arginine concentration in both young and adult SH rats, this effect being larger in adult rats. There was no significant change in plasma levels of L-arginine in age-matched WKY rats. The concentration of other amino acids did not change in both rat strains. On the contrary, there was an increase in plasma NE concentration in both SH and WKY rats after infusion with nicardipine. Plasma L-arginine concentration showed a good inverse correlation with the logarithm of plasma NE concentration in SH and WKY rats and the correlation was expressed as Y = -alpha log(X) + m (Y, plasma L-arginine concentration (nmol/mL); X, plasma NE concentration (pmol/mL); alpha, a slope; and m, an intercept). alpha, 43.0 and 4.35 for 23-week-old SH and WKY rats, respectively, and 17.0 and 4.0 for 6-week-old SH and WKY rats, respectively. The present data together with previous data suggest a direct noradrenergic stimulation of the synthesis of nitric oxide (NO) from L-arginine. The findings also indicate an impairment of the L-arginine metabolism or pools in SH rats compared with WKY rats. The deficiency of L-arginine increases with the age of SH rats and could be related to the development and maintenance of hypertension due to inefficient production of NO.     

Synthetic oligopeptides as isoelectric point markers for capillary isoelectric focusing with ultraviolet absorption detection

Sixteen peptides (trimers to hexamers) were designed for use as a set of pI markers for capillary isoelectric focusing (CIEF). Each peptide contains one tryptophan residue for detection by UV absorption and other amino acid residues having ionic side chains, which are responsible for focusing to its pI. The pIs of these peptides were determined by slab-gel IEF using commercial carrier ampholytes. The focused peptides in the gel were detected by absorption measurement at 280 nm using a scanning densitometer and the pH gradient was determined by measuring the pH of the gel using an oxidized metal membrane electrode. The pI values of the peptides ranged from 3.38 to 10.17. The obtained values agreed well with the predicted ones, which were calculated based on amino acid compositions, with root mean square differences of 0.15 pH unit. The peptides were detected at 280 nm as very sharp peaks when separated by CIEF. The pI values of some standard proteins were redetermined by CIEF by using this set of peptide pI markers and the values agreed closely with those reported previously. The sharp focusing, stability, high purity and high solubility of these synthetic pI markers should facilitate the profiling of a pH gradient in a capillary and the determination of the pI values of proteins.     

Absolute stereochemistry of citrinadins a and B from marine-derived fungus

[Structure: see text]. Citrinadin A (2) is a pentacyclic indolinone alkaloid isolated from the cultured broth of a fungus, Penicillium citrinum, which was separated from a marine red alga. The absolute stereochemistry of the pentacyclic core in 2 and its new congener, citrinadin B (1), was elucidated by analysis of the ROESY spectrum for the chlorohydrin derivative (3) of 1 as well as comparison of the electronic circular dichroism (ECD) spectra for 1 and 2 with those of known spirooxiindole alkaloids. On the other hand, the absolute configuration at C-21 bearing an epoxide ring was assigned as S by comparison of the vibrational circular dichroism (VCD) spectra of 1 with those of model compounds 2S- and 2R-2,3-epoxy-3,3-dimethyl-1-phenylpropan-1-one (4a and 4b, respectively).     

High-performance affinity chromatography of plasmin and plasminogen on a hydrophilic vinyl-polymer gel coupled with p-aminobenzamidine

p-Aminobenzamidine was covalently attached via a spacer moiety to a microparticulate hydrophilic vinyl-polymer gel (Toyopearl HW65S) and this affinity adsorbent was used for the separation of plasmin and plasminogen by high-performance affinity chromatography. Toyopearl HW65S was alkylated with chloroacetylglycylglycine in dimethyl sulphoxide using methylsulphinyl carbanion as a catalyst, then p-aminobenzamidine was coupled to the carboxyl group of glycylglycine to form an acid amide bond. A column packed with the adsorbent retained both plasmin and plasminogen. Plasminogen was eluted with 6-aminohexanoic acid, a haptenic compound for the lysine-binding sites of plasminogen. For the elution of plasmin, the coexistence of 6-aminohexanoic acid and leupeptin (a competitive inhibitor for plasmin) was necessary. The results indicate a two-site interaction of plasmin with the immobilized ligand, i.e., at the lysine-binding sites and the catalytic site. Fluorometric detection of eluted protein and on-line assay of plasmin activity using a fluorogenic substrate, peptidylmethylcoumarylamide, revealed that effective chromatographic separation of the enzyme could be achieved with high sensitivity (10 micrograms) within 1 h.     

Separation of stereoisomers of some terpene derivatives by capillary gas chromatography-mass spectrometry and high-performance liquid chromatography using beta-cyclodextrin derivative columns

Gas chromatographic separations of the stereoisomers of menthol derivatives, important intermediates in the synthesis of physiologically active natural products, were carried out on several substituted beta-cyclodextrin (CD) columns, including per-O-methyl-beta-cyclodextrin (PME-beta-CD), heptakis(2,3-di-O-acetyl-6-tert-butyldimethylsilyl)-beta-CD (DIAC-6-TBDS-beta-CD), and heptakis(2,3-di-O-methyl-6-tert-butyldimethylsilyl)-beta-CD (DIME-6-TBDS-beta-CD) as chiral stationary phases (CSPs). With the DIME-6-TBDS-beta-CD column, a separation of the Z- and E-isomers of methylidenementhol was accomplished; no separation was achieved with the other columns. The stereoisomers of methylidenementhol and the corresponding tert-butyldimethylsilyl (TBS) ether were separated on both the beta-CD and the heptakis(2,3,6-tri-O-methyl)-beta-cyclodextrin (TME-beta-CD) columns by high-performance liquid chromatography (HPLC) with a mobile phase involving acetonitrile and H(2)O. For the separation of the Z- and E-isomers of methylidenementhol, the TME-beta-CD column was superior. In contrast, the beta-CD column was preferable in the case of the corresponding TBS ether.     

The molecular structure of chlorothiomethoxymethyl-bis(triphenylphosphine)palladium(II) methylenechloride solvate at −160°C

The precise molecular structure of [PdCl(CH₂SCH₃)(PPh₃)₂] has been determined from three-dimensional X-ray diffraction data collected at ?160°C. The CH₂Cl₂ solvated crystal ([PdCl(CH₂SCH₃)(PPh₃)₂ · CH₂Cl₂]) belongs to the monoclinic system, space group P2₁/n, with four formula units in a cell of dimensions: a 14.973(3), b 15.333(3), c 17.377(3) Å and β 115.77(1)° at ?160°C. The structure was solved by the conventional heavy atom method and refined by the least-squares procedure to R = 0.035 for observed reflections. The geometry around the palladium atom is square-planar. The phosphorus atoms of the two triphenylphosphine ligands are mutually trans. The CH₂SCH₃ group is bonded to the palladium atom only through the PdC σ-bond and the sulfur atom is not bonded to the metal atom (PdC(1) 2.061(3), SC(1) 1.796(3), SC(2) 1.817(5), Pd?S 2.973(1) Å, PdC(1)S 100.64(14)° and C(1)SC(2) 101.28(18)°). The structure is in contrast to that of [PdCl(CH₂SCH₃)(PPh₃)], in which both the carbon and sulfur atoms of the CH₂SCH₃ group are bonded to the palladium atom.