Identification of new bile alcohols, 5 beta-cholestane-3 alpha,7 alpha,24,26-tetrol, 5 beta-cholestane-3 alpha,7 alpha,25,26-tetrol, and 5 beta-cholestane-3 alpha,7 alpha,26,27-tetrol in human gallbladder bile

The nature of cholestanetetrols present as the glucurono-conjugates in human gallbladder bile was studied. Glucurono-conjugated bile alcohols were isolated by ion exchange chromatography and, after enzymatic hydrolysis, were fractionated by reversed phase partition chromatography to give a fraction containing tetrahydroxy bile alcohols which was analyzed by gas-liquid chromatography and mass spectrometry. Along with the three previously identified bile alcohols, 5 alpha- and 5 beta-cholestane-3 alpha, 7 alpha, 12 alpha,24-tetrols, and 5 beta-cholestane-3 alpha, 7 alpha, 12 alpha,26-tetrol, three new cholestanetetrols, possessing two hydroxyl groups in the ring system and two in the side chain, were detected in the tetrahydroxy bile alcohol fraction. These new bile alcohols were identified as 5 beta-cholestane-3 alpha, 7 alpha,24,26-tetrol, 5 beta-cholestane-3 alpha, 7 alpha,25,26-tetrol, and 5 beta-cholestane-3 alpha, 7 alpha,26,27-tetrol by direct comparison of their gas-liquid chromatographic behaviors and mass spectral data with those of authentic standards prepared from chenodeoxycholic acid by partial synthesis.     

Formation of giant protoplasts from protoplasts of Pleurotus cornucopiae by the cell wall lytic enzyme

Summary When purified protoplasts of Pleurotus cornucopiae IFO9614 were incubated with a mixture of cell wall lytic enzymes, they were found to increase their size. Their average diameter increased from 4.3 m to 31 m after 65 h incubation at 24° C. The presence of cellulase ONOZUKARS in the enzyme mixture had a significant effect on the formation of giant protoplasts. Regeneration frequency of giant protoplasts in a medium containing 0.5 M sucrose was 3.5%, approximately six times that of normal protoplasts.     

Synthesis of four diastereoisomers at carbons 24 and 25 of 3 alpha,7 alpha,12 alpha,24-tetrahydroxy-5 beta-cholestan-26-oic acid, intermediates of bile acid biosynthesis

The synthesis of four stereoisomers at C-24 and C-25 of 3 alpha,7 alpha,12 alpha,24-tetrahydroxy-5 beta-cholestan-26-oic acid is described. Pyridium chlorochromate oxidation of 3 alpha,7 alpha,12 alpha-triacetoxy-5 beta-cholan-24-ol (II) prepared from cholic acid (I) afforded 3 alpha,7 alpha,12 alpha-triacetoxy-5 beta-cholan-24-al (III) which was converted to a mixture of the four stereoisomers (IV-VII) by a Reformatsky reaction with ethyl DL-alpha-bromopropionate followed by alkaline hydrolysis. Separation of these isomers (IV-VII) was achieved by silica gel column chromatography, and subsequent reversed-phase partition column chromatography. The configurations at C-24 were elucidated by conversion of each isomer into (24R)- or (24S)-5 beta-cholestane-3 alpha,7 alpha,12 alpha,24-tetrol (XII or XI) by Kolbe electric coupling, the C-24 configurations of which were determined by modified Horeau's method and 13C-nuclear magnetic resonance spectroscopy. The stereochemistries at C-25 were deduced by comparison of IV-VII with the products of the hydroboration followed by oxidation with alkaline hydrogen peroxide of (24E)-3 alpha,7 alpha,12 alpha-trihydroxy-5 beta-cholest-24-en-26-oic acid (XIII).     

In Vitro Phosphorylation of Proteins in IAA-Treated Mesocotyls in Zea mays

Five-mm sections of elongation zones of Zea mesocotyls wereincubated for designated periods with various concentrationsof IAA. In vitro protein phosphorylation in the soluble fraction(85,000 x g supernatant) prepared from the sections was analyzedby sodium dodecyl sulfate-polyacrylamide gel electrophoresis.The phosphorylation of proteins in the soluble fraction thathad been prepared from sections incubated for 20 min in thepresence of 10{small tilde}^s M IAA was greater than that inthe sections incubated for 20 min without IAA. The amount ofphosphorylation of proteins per protein became higher when higherconcentrations increased (10{small tilde}⁸—10{small tilde}⁵M).The growth of sections incubated in the presence of 10{smalltilde}⁸ M IAA or higher concentrations was greater than thatof sections incubated in the absence of IAA. The promotion ofgrowth by IAA was greater at higher concentrations of IAA. Proteinsin the soluble fraction, prepared from sections incubated for20 min in the presence of 10{small tilde}⁵ M IAA, were phosphorylatedin the presence of either 10 fM cAMP, 10 µM cGMP, 100µM W-7, 100 µM W-5, 20 µM H-7 or 20 µMHA1004. The calmodulin antagonist, W-7, and the inhibitor ofprotein kinase C, H-7, inhibited the phosphorylation of proteinsstimulated by incubation with IAA. These results suggest thatIAA promotes cell elongation via protein phosphorylation thatdepends on calmodulin-dependent protein kinase and protein kinaseC. (Received November 29, 1995; Accepted May 20, 1996)     

Is there an energy conservation “system” in brain that protects against the consequences of energy depletion?

A poorly understood marked decrease (circa 50% of control) in local cerebral glucose utilization is caused by sublethal doses of NaCN. The decrease is global, occurring in essentially all brain regions and is entirely reversible within hours, leaving no obvious pathology. This event is not unique to NaCN in so far as a strikingly similar pattern of decreased glucose utilization occus with some other toxins. Nor can it be attributed to a direct action of NaCN since local application by microdialysis to the striatum produces a global depression. These results imply that some widely distributed system or substance is involved. We speculate the existence of a system possibly related to the reticular activating system that senses a fall in energy production and acts globally to make cells quiescent and thus would give some protection from excitotoxic driven damage.Special issue dedicated to Dr. Sidney Ochs.     

Visual prey discrimination of queen and worker ants by a generalist lizard

Laboratory experiments were conducted to test the hypothesis that widelyforaging generalist lizard,Eumeces okadae, visually discriminates palatable queen ants from unpalatable worker ants. The workers ofCamponotus japonicus andLasius niger were rejected on sight, while the queens of both species were eaten with little prior chemical examination by tongue flicks or licks. Comparison of the lizards' responses towards the workers and wingless queens of similar size indicated that neither body size nor presence or absence of wings accounted for difference in responses toward the 2 ant castes. The lizards probably discriminated different ant castes by body proportions.     

Identification of 3 alpha, 6 alpha, 7 alpha, 12 alpha-tetrahydroxy-5 beta-cholestanoic acid in Zellweger's syndrome

In order to confirm the occurrence of 3 alpha, 6 alpha, 7 alpha, 12 alpha-tetrahydroxy-5 beta-cholestanoic acid in Zellweger's syndrome, the nature of tetrahydroxycholestanoic acids present in a patient with this disease was studied. Urinary bile acids were extracted with a Sep-pak C18 cartridge and methylated after alkaline hydrolysis. The methyl esters were purified by silica gel column chromatography, and the methyl tetrahydroxycholestanoate fraction was analyzed by gas liquid chromatography-mass spectrometry. Along with already known side chain hydroxylated derivatives of 3 alpha, 7 alpha, 12 alpha-trihydroxy-5 beta-cholestanoic acid, 3 alpha, 7 alpha, 12 alpha, 24- and 3 alpha, 7 alpha, 12 alpha, 26-tetrahydroxy-5 beta-cholestanoic acids, three nuclear hydroxylated derivatives of 3 alpha, 7 alpha, 12 alpha-trihydroxy-5 beta-cholestanoic acid were found. One of them was identified as 3 alpha, 6 alpha, 7 alpha, 12 alpha-tetrahydroxy-5 beta-cholestanoic acid by direct comparison with the authentic standard which was chemically synthesized from 3 alpha, 6 alpha, 7 alpha, 12 alpha-tetrahydroxy-5 beta-cholanoic acid by side chain elongation.     

REPRODUCTIVE ISOLATION BY HOST SPECIFICITY IN A PAIR OF PHYTOPHAGOUS LADYBIRD BEETLES

Crossing experiments and food-choice tests show that two sympatric species of phytophagous ladybird beetles, Epilachna niponica and E. yasutomii, are reproductively isolated by host-plant specificity. Adult beetles selected their natural hosts when given choices, though some accepted the host of the other species when no choice was offered. In each species, survival of larvae to the second instar was significantly higher on their own host plant. No evidence for sexual isolation, gametic isolation, hybrid inviability, or reduced hybrid fertility was detected. Reproductive isolation by host specificity is an important prerequisite for certain models of sympatric speciation. Although the present example supports the plausibility of such models, an allopatric origin of host-plant specificity cannot be discounted.     

Potential Distribution and Ionic Concentration at the Bean Root Surface of the Growing Tip and Lateral Root Emerging Points

The electrical potential distribution has been measured preciselyaround the root surface of the bean sprout Vigna mungo (L) Hepper.A large negative potential well was found at the growth portionof the root tip. Also, in the matured region of the root, wefound a negative potential well at an unspecified position inspite of the fact that nothing was detected on the smooth surface.A lateral root emerge was found to have initiated after 15–20hours just at the position corresponding to the potential well.With the expectation that these potentials can be elucidatedbased on the transport of ions which are released or absorbedby the root as a result of cell activity, we precisely measuredthe concentrations of major ion species (K⁺, H⁺, and Cl^–)around the root. The theoretical potential distribution curvesobtained by putting all the concentration data into the Henderson'sEquation for a liquid junction (diffusion) potential coincidedwell with the experimental curves. (Received October 24, 1994; Accepted March 24, 1995)