Jean De Graeve

Radioactive assay for aryl hydrocarbon hydroxylase. Improved method and biological importance

By addition of two volumes of a 1M aqueous KOH/dimethylsulfoxide (1585; v/v) mixture to the enzymatic incubation medium, it is possible to selectively extract the unmetabolized benzo(a)pyrene in hexane. Therefore, the radio-activity remaining in the water phase corresponds to all the in vitro synthesized metabolites. This isotopic method is very sensitive (2 × 10−11 moles) and is almost insensitive to the room lighting. The aryl hydrocarbon hydroxylase activities found with this method are 2,3 and 10 times higher in the liver, lung and kidney respectively compared to those obtained with the fluorimetric method.

Determination of papaverine in blood samples by gas-liquid chromatography and mass fragmentography

The measurement of papaverine in blood samples by using either a glass capillary column with a flame-ionization detector or a packed column with mass fragmentographic detection is described. The two methods permit the determination of the normal range of concentrations of papaverine in blood (2-500 ng/ml). Owing to its high specificity, mass fragmentography is greatly superior to capillary chromatography, which is sometimes subject to interferences by solvent impurities.

Fragmentation of penicillin catalysed by the exocellular DD-carboxypeptidase-transpeptidase of Streptomyces strain R61 isotopic study of hydrogen fixation on carbon 6

The slow degradation of complex EI* (the first-order rate constant k4 is of the order of 10m4 s-r) results in enzyme reactivation and in the fragmentation of the penicillin molecule. The fragments released are N-acylglycine and an unstable intermediate (Z) which in turn gives rise to N-formyl-D-penicillamine [3-51. Formation of N-acylglycine requires a double hydrolysis within the fl-lactam ring, i.e., rupture of both the amide linkage and the Cs-C6 bond. This latter reaction results in the formation of a -CH2-methylene group at Cg. This report describes isotopic studies on the mechanism of hydrogen fixation on Cb. The approach rested upon the effects of DsO on the fragmentation reaction; it made use of the fact that the protons of the methylene groups are not exchangeable.

Alkaloids from leaves of Pterotaberna inconspicua and the Kisantu hybrid problem

Abstract In the leaves of Pterotaberna inconspicua , two alkaloids with the vincadifformine skeleton have been found. One is a monomer and corresponds to 5-oxo vincadifformine; the other, kisantine, is a dimeric species ( M + 810) composed of two highly oxidized tabersonine units. These results suggest a parenthood between the Kisantu hybrid and P. inconspicua .

STEROID-16 α-HYDROXYLASE IN RAT LIVER: BIOCHEMICAL AND BIOLOGICAL PROPERTIES

Publisher Summary This chapter describes some fundamental biochemical properties of the rat liver steroid-16α-hydroxylase. To overcome the methodological difficulties in measuring steroid hydroxylase activities accurately, new and precise assays were developed for progesterone, pregnenolone, and testosterone 16α-hydroxylase. These assays are based on the release of a 16α-positioned atom of tritium into the incubation medium during the enzymatic hydroxylation. This same principle was already applied to the assay of cholesterol-7α-hydroxylase and of progesterone and pregnenolone-17α-hydroxylase. On a quantitative basis, the activity of the steroid-16α-hydroxylase seems to vary extensively not only with the steroid used as a substrate but also as a function of different physiological parameters and pharmacological treatments. The experiments described in the chapter demonstrate the decisive advantages of the tritium exchange method for assaying the microsomal steroid hydroxylases. Providing that a specifically tritiated substrate is available and that its tritium is not released by nonenzymatic reactions during the various manipulations, such methods are technically very easy to realize and allow the performance of many assays at the same time.