Rapid and sensitive method for measuring norepinephrine, dopamine, 5-hydroxytryptamine and their major metabolites in rat brain by high-performance liquid chromatography. Differential effect of probenecid, haloperidol and yohimbine on the concentrations of biogenic amines and metabolites in various regions of rat brain.

A rapid and sensitive method has been developed for the simultaneous determination of norepinephrine, dopamine, 5-hydroxytryptamine and their respectively metabolites 3-methoxy-4-hydroxyphenylglycol, homovanillic acid, 3,4-dihydroxyphenylacetic acid, and 5-hydroxyindoleacetic acid in discrete brain regions of rats. The supernatants of tissue homogenates were injected directly into a reversed-phase liquid chromatography system, coupled with electrochemical detection. Each of these compounds gave a linear response over the range 5.5-200 ng/ml cerebellar homogenate (0.11-4.0 ng on column). Analytical recoveries of these compounds, added to the homogenate, were essentially complete when compared with standards dissolved in perchloric acid. The average between-assay coefficients of variation for all these compounds were lower than 6.9% over the range 5.5-200 ng/ml. The within-assay coefficients of variation were lower than 9.7%, measured at 5.5 or 23.6 ng/ml. With the present test parameters and mobile phase conditions, all compounds were readily oxidized at 0.8 V vs. a Ag/AgCl reference electrode. The method was applied to an analysis of the differential activity of biogenic amines in the rat striatum, hypothalamus, and hippocampus, produced by probenecid, haloperidol and yohimbine.

Bovine serum albumin: characterization of a fatty acid binding site on the N-terminal peptic fragment using a new spin-label.

A new spin-label, 4-(L-glutamo)-4'-[(1-oxy-2,2,5,5-tetramethyl-3L-pyrrolidinyl )amino]-3, 3'-dinitrodiphenyl sulfone, is shown to bind to one high-affinity binding site on bovine serum albumin (K = 5 X 10(4) M-1, n = 1). Analysis of the binding of the spin-label to the amino-terminal half (peptic fragment PB) and the carboxy-terminal half (peptic fragment PA) of BSA, and their complex (PA-PB), indicates that the spin-label binds to a long-chain fatty acid binding site located on PB. The usefulness of the novel specificity of the spin-label in characterizing this binding site is discussed.