Gaby Schmitt

An interlaboratory study of TEX86 and BIT analysis of sediments, extracts, and standard mixtures

Two commonly used proxies based on the distribution of glycerol dialkyl glycerol tetraethers (GDGTs) are the TEX86 (TetraEther indeX of 86 carbon atoms) paleothermometer for sea surface temperature reconstructions and the BIT (Branched Isoprenoid Tetraether) index for reconstructing soil organic matter input to the ocean. An initial round-robin study of two sediment extracts, in which 15 laboratories participated, showed relatively consistent TEX86 values (reproducibility +/- 3-4 degrees C when translated to temperature) but a large spread in BIT measurements (reproducibility +/- 0.41 on a scale of 0-1). Here we report results of a second round-robin study with 35 laboratories in which three sediments, one sediment extract, and two mixtures of pure, isolated GDGTs were analyzed. The results for TEX86 and BIT index showed improvement compared to the previous round-robin study. The reproducibility, indicating interlaboratory variation, of TEX86 values ranged from 1.3 to 3.0 degrees C when translated to temperature. These results are similar to those of other temperature proxies used in paleoceanography. Comparison of the results obtained from one of the three sediments showed that TEX86 and BIT indices are not significantly affected by interlaboratory differences in sediment extraction techniques. BIT values of the sediments and extracts were at the extremes of the index with values close to 0 or 1, and showed good reproducibility (ranging from 0.013 to 0.042). However, the measured BIT values for the two GDGT mixtures, with known molar ratios of crenarchaeol and branched GDGTs, had intermediate BIT values and showed poor reproducibility and a large overestimation of the true (i.e., molar-based) BIT index. The latter is likely due to, among other factors, the higher mass spectrometric response of branched GDGTs compared to crenarchaeol, which also varies among mass spectrometers. Correction for this different mass spectrometric response showed a considerable improvement in the reproducibility of BIT index measurements among laboratories, as well as a substantially improved estimation of molar-based BIT values. This suggests that standard mixtures should be used in order to obtain consistent, and molar-based, BIT values.

Na+‐Independent Binding of [3H]GABA and [3H]Muscimol to Subcellular Particles of Neural Primary Cultures and Whole Brain

y-AMINOBUTYRIC acid (GABA) is bound to particulate preparations of the vertebrate CNS by highaffinity processes in the absence of added Na+, and it is believed that such processes represent interactions of GABA and its synaptic receptors (e.g. Enna and Snyder, 1975: reviews by DeFeudis, 1977 and Johnston, 1978). In order to examine this hypothesis further, the binding of [RH]GABA and of the potent GABA-agonist, [3H]muscimol, to subcellular particles of whole brain and of neurone-enriched and astroblast cultures have been compared.

High-affinity binding of [3H]muscimol to subcellular particles of a neurone-enriched culture of embryonic rat brain

High-affinity, Na+-independent binding of [3H]muscimol (KB approximately equal to 1.6 x 10(-8) M; Bmax approximately equal to 0.14 nmol/g pellet) occurred to a frozen-thawed particulate fraction of 74-h-old neurone-enriched cultures prepared from the cerebra of 12-13-day-old rat embryos. This finding provides evidence that GABA-receptors exist on cultured neurones which contain only a few synaptic connections.

Effect of cyclohexenonic long chain fatty alcohols on neurite outgrowth. Study on structure-activity relationship

Abstract Four series of long chain fatty alcohols bearing a cyclohexenone moiety in addition to a ω-alkanol side chain were synthesized using “Umpolung” reactivity strategy. Their effect on neurite outgrowth was evaluated by means of fetal rat neurons in culture. The length of the ω-hydroxy side chain is a crucial factor for biological activity.

Substrate Specificity of [3H] Muscimol Binding to a Particulate Fraction of a Neuron-enriched Culture of Embryonic Rat Brain

Abstract: The effects of some GABA analogues and some drugs on the binding of [3H]muscimol (3.08 nM) to thoroughly washed subcellular particles prepared from a neuron‐enriched culture of embryonic rat brain were examined using Na+‐free Tris‐citrate medium and a centrifugation method. Competition for [3H]muscimol binding sites by excess(10−5 M) unlabelled GABA provided estimates of “specific” binding. In accord with in vivo neuropharmacological studies on GABA receptors and with in vitro studies on cerebral membrane preparations, [3H]muscimol binding was potently inhibited by muscimol itself (IC50, 2.5 nM), GABA (1C50, 43 nM), isoguvacine (IC50, 61 nM), and 3‐aminopropanesulphonic acid (IC50, 160 nM), and less potently inhibited by the GABA antagonist bicuculline methobromide (IC50, 800 nM). δ‐ Aminovaleric acid (IC50, 2.6 μM), the glycinelp‐alanine antagonist strychnine (IC50, 6.6 μM), and the predominantly glial GABA uptake inhibitors β‐alanine (IC50, 23 μM) and p‐proline (IC50, 66 μM) also inhibited [3H]muscimol binding. Other inhibitors of Na+‐dependent GABA uptake, (±)‐nipecotic acid, L‐ 2,4‐diaminobutyric acid, and guvacine, as well as picrotoxinin, were relatively inactive as inhibitors of [3H]muscimol binding (IC50≥ 1 mM). In addition to revealing that GABA receptors are present on neuronal membranes before the formation of most synapses, this binding of [3H]muscimol that occurs to neuronal, but not to glial, membranes might be useful as a “neuronal marker” and for the further characterization and isolation of GABA receptors.

A convergent synthesis of 2′-o-methyl uridine

Abstract A convergent synthesis of 2′-O-methyl uridine (1) is described. The key steps in our synthesis are: (1) a facile obtention of the 2′-O-methyl sugar synthon using totally selective and efficient methylation conditions; (2) a stereoselective high-yield condensation with an uracil derivative, yielding the desired β-form.

Synthesis of optically active diastereomers of a nonproteic neurotrophic mimetic

Abstract The four diastereomers 3 , elongated analogues of perhydroretinol, are synthesized starting from both optically pure 3-bromo-2-methyl-1-propanol enantiomers. All exhibit neurotrophic activity on cultured neuronal cells derived from fetal rat cerebral hemispheres.

Preferential phosphorylation at the primary alcohol of non-protected thymidine or carbohydrates

Abstract A preferential phosphorylation at the primary alcohol of non-protected thymidine or carbohydrates by the hydrogen phosphonate method is described. Thus, 6-(mannopyrannosyl), 6-(glucopyrannosyl) or 5′-(thymidinyl) phosphodiesters were obtained in 35% to 80% yields.

Synthesis of the phosphodiester of 3β(7β-hydroxycholesterol) and of 5′(3′deoxy, 3′azido-thymidine).

Abstract In order to enhance the therapeutic efficacy of azido-thymidine (AZT) by improving its pharmacokinetic properties and to try to target its action to the lymphocytes, the phosphodiester of 3β(7β-hydroxycholesterol) and of 5′(3′deoxy, 3′azido-thymidine) 1 was synthesized using two different techniques of phosphorylation : the phosphoramidite and the hydrogen phosphonate methodologies. Preliminary results show an in vitro anti-HIV activity.

Synthesis of phosphoric acid diesters of 7β-hydroxycholesterol and of carbohydrates

Abstract In order to enhance the antitumor efficacy of 7β-hydroxycholesterol by targetting its action to defined organs, the phosphoric acid esters of 3(7β-hydroxycholesteryl) and of 6(galactopyranosyl) 1 or 6(mannopyranosyl) 2 were synthesized by the phosphoramidite method (with protected C-1, 2, 3, 4 hydroxyl groups for the carbohydrates). As the protection of the sugars increased the length of the synthesis, we decided to use the hydrogen-phosphonate methodology which leads to a selective phosphorylation at the primary alcohol of carbohydrates and avoids the use of protected carbohydrates. Compound 2 was synthesized in good yield. However compound 1, probably due to steric hindrance, could not be obtained by this second method.