Ceramide (N-acylsphingosine) biosynthesis has been proposed to involve introduction of the 4,5-trans-double bond of sphingosine after synthesis of dihydroceramide (i.e. N-acylsphinganine). For the first time, the in vitro conversion of dihydroceramide to ceramide has been demonstrated using rat liver microsomes and N-[1-14 C]octanoyl-D-erythro-sphinganine (st-H 2 Cer) and either NADH or NADPH as co-substrate; the apparent K m values for st-H 2 Cer and NADH were 340 and 120 M, respectively. Molecular oxygen is required for enzymatic activity, and cyanide, divalent copper, as well as antibodies raised against cytochrome b 5 are inhibitory, which suggests that this enzyme should be named dihydroceramide desaturase based on these similarities with the mechanism of ⌬ 9 -desaturase (stearoyl-CoA desaturase). Factors that influenced the activity of dihydroceramide desaturase include the alkyl chain length of the sphingoid base (in the order C 18 > C 12 > C 8 ) and fatty acid (C 8 > C 18 ); the stereochemistry of the sphingoid base (D-erythro-> L-threo-dihydroceramides); the nature of the headgroup, with the highest activity with dihydroceramide, but some (ϳ20%) activity with dihydrosphingomyelin (activity was not detected with dihydroglucosylceramide, however); and the ability to utilize alternative reductants (ascorbic acid could substitute for a reduced pyridine nucleotide, but was inhibitory at higher concentrations). Dihydroceramide desaturase was inhibited by dithiothreitol, which suggests that it might be possible to alter ceramide synthesis by varying the thiol status of hepatocytes. Consistent with this hypothesis, when rat hepatocytes were cultured in varying concentrations of N-acetylcysteine (5 and 10 mM), there was a decrease in the relative incorporation of [ 14 C]serine into [ 14 C]ceramide. These studies have conclusively established the pathway of ceramide synthesis via desaturation of dihydroceramide and have uncovered several properties of this reaction that warrant further consideration for their relevance to both sphingolipid metabolism and signaling.Over the last few years, it has become clear that ceramide plays important roles in the metabolism of cells. It is involved as a second messenger in what has become known as the sphingomyelin cycle (reviewed in Ref. 1) and, to name but a few, serves as a mediator of cellular senescence (2), apoptosis, and differentiation in many cell types (3). It was also observed that not only ceramide derived from the sphingomyelin pool found in the plasma membrane but also that from de novo synthesis is involved in the cellular responses to inducers of apoptosis (4) and differentiation (5). These results suggest an important role of the anabolic pathway of ceramides in signal transduction. It is therefore very likely that sphingolipid biosynthesis is tightly regulated. Taking into consideration that dihydroceramide does not mimic the effects of ceramide (6, 7), one possible site of regulation might be dihydroceramide desaturase. In addition, ceramide is also involve...

The synthesis and characterization of several 2,9‐diarylphenanthrolines with anthracenyl substituents in the 2‐position is described. These compounds were used to prepare a series of homoleptic and heteroleptic bis(phenanthroline)copper(I) complexes, whose cyclic voltammetry, UV/Vis and fluorescence data are investigated. From PM3 semiempirical calculations it becomes clear that two different strategies can be applied to prepare clean heteroleptic complexes.

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The preparation of several new and soluble bisphenanthrolines that are important building blocks for various supramolecular structures based on heteroleptic copper(I) complexes is described.

Dihydroceramide desaturase is responsible for the introduction of the 4,5-trans double bond into ceramide. Here, we describe the localization of this enzyme in the endoplasmic reticulum (ER) using ER-and Golgi-enriched fractions from rat liver. Furthermore, enzyme topology was studied. Mild proteolysis of ER-derived vesicles under conditions which assure membrane integrity (latency of mannose 6-phosphatase was at least 91%) resulted in an up to 90% inactivation of dihydroceramide desaturase activity. This indicates a cytosolic orientation of dihydroceramide desaturase activity in the ER membrane.