Age-dependent changes in rat liver microsomal and mitochondrial NADPH-dependent lipid peroxidation.

Purified outer membrane proteins O-8 and O-9 were able to bind to the peptidoglycan sacculi in sodium dodecyl sulfate solution. Binding was stimulated by lipopolysaccharide, that of protein O-9 being stimulated more remarkably. Proteins which had been heated in sodium dodecyl sulfate solution did not bind to the peptidoglycan sacculi even in the presence of lipopolysaccharide, while heated lipopolysaccharide stimulated the binding of non-heated proteins. The removal by pronase of the lipoprotein covalently bound to the peptidoglycan sacculi did not change the protein binding ability of the sacculi.     

REGIONAL CHANGES IN CEREBRAL GABA CONCENTRATION AND CONVULSIONS PRODUCED BY d AND BY l-ALLYLGLYCINE

Abstract— The convulsant action of allylglycine (2-amino-4-pentenoic acid) is due to the metabolic conversion of allylglycine to 2-keto-4-pentenoic acid, a more potent glutamic acid decarboxylase inhibitor and more potent convulsant than the parent compound. We report regional changes in cerebral GABA concentration in rats after administration of d - and l -allylglycine. d -Allylglycine (3.75 mmol/kg) induced convulsions in 95–115 min, characterised by repeated clonic limb movements and rapid rotation around the head to tail axis. GABA concentrations were only reduced in cerebellum and ponsmedulla during the pre and post-convulsive periods. The localised reduction of GABA concentration is consistent with the enzymic conversion of d -allylglycine to 2-keto-4-pentenoic acid catalysed by cerebral d -amino acid oxidase, an enzyme known to be localised to the hind brain and spinal cord. l -allylglycine (1.2mmol/kg i.p.) induced convulsions in 65 -90 min, characterised by violent running followed by tonic flexion and extension. During the pre-convulsive period, GABA concentrations were reduced in all brain areas studied except the globus pallidus and ventral midbrain. The widespread decreases in GABA concentration suggest that the enzyme(s) which catalyse the conversion of l -allylglycine to 2-keto-4-pentenoic acid are widely distributed within the brain.     

Prevention of thromboxane B2-induced hepatocyte plasma membrane bleb formation by certain prostaglandins and a proteinase inhibitor

Isolated hepatocytes incubated in the presence of thromboxane B2 developed many plasma membrane blebs which are a characteristic feature of toxic or ischaemic cell injury. When hepatocytes were incubated in the presence of both thromboxane B2 and the non-lysosomal proteinase inhibitor, leupeptin, were also well protected from the formation of blebs. This implies that thromboxane B2 is able to activate non-lysosomal proteinases which appear to attack certain cytoskeletal proteins. The data presented are consistent with thromboxane B2 acting as an intermediary in a proposed mechanism of cell injury and death in which elevated cytosolic free Ca2+ levels activate phospholipase A2 and the arachidonic acid cascade.     

Enzymic lipid peroxidation in the microsomal fraction of rat brain

Abstract: An enzymic lipid peroxidation system has been demonstrated in the microsomal fraction of rat brain and the requirements and optimal conditions for assay determined. The involvement of NADPH-cytochrome c reductase was demonstrated in vesicles reconstituted with lipids extracted from the brain microsomal fraction. Further characterization of the system made use of substances shown to inhibit the liver microsomal system. α-Tocopherol was shown to be an effective inhibitor of lipid peroxidation in the brain microsomal system, whereas Na₂SO₃ had no effect, which is indicative that free radical transfer occurs only in the hydrophobic regions. Neither superoxide dismutase nor catalase inhibited lipid peroxidation. The implications of an NADPH-cytochrome c reductase-dependent lipid peroxidation system that is not linked to a drug hydroxylation system and appears to differ from the liver microsomal system in a number of other ways are discussed.     

Carbon-13 NMR studies on [4-13C] uracil labelled E. coli transfer RNA1(Val1).

In this paper we describe carbon-13 nuclear magnetic resonance results on 13C-enriched purified transfer RNAI(VAL) from from E. coli SO-187, a uracil requiring auxotroph. The organism was grown on uracil 90% 13C-enriched at the carbonyl C4 position. Transfer RNAI(Val) was purified from bulk tRNA by sequential chromatography on columns of BD cellulose, DEAE-Sephadex A-50 and reverse gradient sepharose 4B. Dihydrouridine, 4-thiouridine, and uridine 5-oxyacetic acid located at discrete positions in the polymer backbone were tentatively assigned in the highly resolved 25 MHz 13C-spectra. Chemical shift versus temperature plots reveal differential thermal perturbation of the ordered solution structure, evident in the large dispersion (ca 3-4 ppm) of the uridine C4 resonances. Over the range 26-68 degrees C, V in the anticodon displays the largest downfield shift. Whereas several uridine residues rapidly shift downfield between 50-68 degrees, one moves upfield beginning at 37 degrees. The results are qualitatively compared with proton NMR analysis of the three dimensional structure.     

An Embedding Method for Histochemical Studies of Undecalcified Skeletal Growth Plate

We have used glycol methacrylate to study undecalcified skeletal growth plate and subchondral bone. Minor modifications of the original technique including dehydration in glycol methacrylate vacuum infiltration and polymerization in the cold make it quite suitable for embedding of such tissues. Moreover, specimens can be processed quickly and the morphologic and biochemical integrity of the tissue retained so that histochemical procedures can be readily applied. Collagen, glycosaminoglycan, glycogen, lipid, calcium and the activity of alkaline and acid phosphatase were localized. This technique appears to be very useful for studying skeletal tissues.     

P nuclear magnetic resonance spectroscopy of lipopolysaccharides from pseudomonas aeruginosa

Intact lipopolysaccharide antigens isolated from seven different immunotypes of Pseudomonas aeruginosa have been examined by ³¹P-NMR spectroscopy. These macromolecular complexes contain phosphorus covalently attached to the carbohydrate residues present in the lipid A moiety and the ‘core’ oligosaccharide region. The spectral signals for various ortho- and pyro-phosphoric esters were observed. All phosphate groups appeared to be mono-esterified. Certain shifts characteristic for phosphate diester groups, observed in lipopolysaccharide complexes from other Gram-negative bacteria, were absent. Furthermore, no evidence was found to indicate that phosphate groups are involved in the covalent linkage of individual lipopolysaccharide complexes to form dimers or trimers.     

31P nuclear magnetic resonance spectroscopy of lipopolysaccharides from pseudomonas aeruginosa

Intact lipopolysaccharide antigens isolated from seven different immunotypes of Pseudomonas aeruginosa have been examined by ³¹P-NMR spectroscopy. These macromolecular complexes contain phosphorus covalently attached to the carbohydrate residues present in the lipid A moiety and the ‘core’ oligosaccharide region. The spectral signals for various ortho- and pyro-phosphoric esters were observed. All phosphate groups appeared to be mono-esterified. Certain shifts characteristic for phosphate diester groups, observed in lipopolysaccharide complexes from other Gram-negative bacteria, were absent. Furthermore, no evidence was found to indicate that phosphate groups are involved in the covalent linkage of individual lipopolysaccharide complexes to form dimers or trimers.     

Synthesis of C-substituted triazoles,isoxazoles, and pyrazoles by 1,3-dipolar cycloaddition to acetylenic sugars

Addition of phenyl azide to 3,5-di-O-acetyl-6,7-dideoxy-1,2-O-isopropylidene-β-l-idio-hept-6-ynofuranose (1) and subsequent saponification gave a 4-substituted 1-phenyl-1,2,3-triazole derivative (3) whose optical rotatory dispersion (o.r.d.) curve was positive. The α-d-gluco analog (5) of 1 similarly gave the 5-epimer (7) of 3; its o.r.d. curve was negative. Both 3 and 7 were degraded to the known 1-phenyl-1,2,3-triazole-4-carboxaldehyde. Similarly, addition of 2,4,6-trimethylbenzonitrile N-oxide to 1 or 5 gave the corresponding, crystalline 3-mesitylisoxazoles as single products; ¹³C-n.m.r. spectroscopy was used to establish the orientation of addition. Related 3-mesitylisoxazoles (11 and 13) were obtained from 1,2:3,4-di-O-isopropylidene-d-glycero-α-d-galacto-oct-7-ynopyranose (10) and its l-glycero 6-epimer (12), respectively; 11 showed the expected, large levorotation, and the 6-epimer 13 was also levorotatory. Benzonitrile (N-phenyl)imine, prepared in situ from 1-(α-chlorobenzylidene)-2-phenylhydrazine and base, did not react with 10 (or its 6-epimer 12), but did react with the 6-keto analog to give a 5-substituted 1,3-diphenyl-1,2-diazole.