Vassilios M. Kapoulas

Detection of virgin olive oil adulteration with refined oils by second-derivative spectrophotometry

Abstract Weak absorption maxima of conjugated tetraenoic systems (285–315 nm) can be quantitated by second-derivative spectrophotometry. The distance of two consecutive extremes (maximum-minimum) of the second-derivative reflection at 315 nm, measured in units of absorptivity (ΔK 315 ) shows the most characteristic differences between virgin olive oil and refined olive, olive kernel and seed oils. The ranges of ΔK 1% 315 values were found to be 0·008–0·015 and 0·010–0·030 for virgin olive oils of the last and older crops, respectively, whereas ΔK 1% 315 values of refined olive and olive kernel oils exceed 0·450 and 0·990, respectively. It is demonstrated that 5% adulteration of virgin olive oil by refined oils may be detected effectively by this technique.

Phosphono-sphingomyelins. I: Synthesis of ceramide (trimethyl) aminoethyl phosphonate

Abstract The synthesis of the phosphono-analogue of sphingomyelin is described. The N -acyl-D-erythro-sphingosyl-1-( N , N , N -trimethyl-2-aminoethyl) phosphonate was obtained by phosphonylation of N -acyl-3- O -benzoyl-D-erythro-sphingosine with (2-bromoethyl)phosphonic acid chloride and triethylamine, subsequent quaternisation with anhydrous trimethylamine and benzene at 55–60°C for four days, and finally, consecutive removal of the protective group by mild alkaline hydrolysis. Comparison of the CD spectra of both, natural sphingomyelin and its phosphono-analogue, confirmed that their structures and configurations were identical.

In vivo metabolism of platelet-activating-factor (1-O-alkyl-2-acetyl-sn-glycero-3-phosphocholine) by the protozoan Tetrahymena pyriformis

The metabolism of exogenous platelet-activating-factor was studied in the protozoan Tetrahymena pyriformis in vivo. When the cells are exposed to 1.10(-6) M PAF, the molecule is rapidly metabolized to 1-O-alkyl-2-acyl(long chain)-GPC, a major component of the protozoan membranes. The appearance of lyso-PAF from the first minutes even in low levels provides evidence that deacetylation is an intermediate step. After incubation for 30 min, transformation to aminoethyl phosphonolipids is also observed. The fate of PAF in concentrations 1.5.10(-11) M or 1.10(-8) M PAF, was the same. An amount of PAF depending on the external PAF concentration remained intact in the cell even after 1 h incubation. Our results suggest that the easily cultured protozoan can be a useful model for studying PAFs metabolism.

Lipids of Pinus halepensis pollen

Abstract The total lipids of Pinus halepensis pollen were separated into individual classes of neutral and polar lipids and the components of each class were identified and determined quantitatively. Free fatty acids, waxes and triacylglycerols were found as the main constituents of neutral lipids and phosphatidylcholine and phosphatidylethanolamine of polar lipids. Glycerylether derivatives were detected in neutral and polar lipid fractions. Free and esterified volatile fatty acids were also found in pollen and its neutral lipid fraction.

Semisynthetic Preparation of 1-O-hexadecyl-2-acetyl-sn-glyceryl-3- phosphorylcholine (Platelet Activating Factor)

Abstract 1-O-hexadecyl-2-acetyl-sn-glyceryl-3-phosphorylcholine (platelet activating factor, PAF), (1.9 μmol) was prepared from the total lipid extract of the protozoan Tetrahymena pyriformis 9 × 107 cells. The procedure involved mild alkaline hydrolysis of the total lipids, followed by acetylation and purification of the product by preparative TLC and HPLC . The yield was 60 % with respect to the content of 1-O-alkyl-2-acyl-sn-glyceryl-3-phosphorylcholine in the total lipids, determined after preparative TLC . The alkyl side chain of the semisynthetic PAF was composed of hexadecyl residue. Our product was identified as PAF according to its biological activity, the chromatographic behaviour on TLC and HPLC , the physicochemical properties and the behaviour under treatment with PLA2 and Lipase from Rhizopus arrhizus. The above procedure is proposed as a facile, inexpensive and convenient method.

Electrocardiographic alterations induced by AGEPC in Wistar rats in relation to its hypotensive and hematologic effects

AGEPC administration into Wistar rats caused no remarkable thrombocytopenia, slight decrease of the percent count of PMNs in whole blood accompanied by anequal leukocytopenia and a transient increase in hematocrit, due to fluid extraversion. Apart from the dramatic fall in blood pressure caused by AGEPC, relatively sinus bradycardia was recorded at doses over 6 micrograms/kg b.w. S-T segment elevation, mainly evident in II, III and AVF leads, was also recorded within the first minutes after AGEPC administration, at doses over 1 microgram/kg b.w. At lethal doses, various degrees of A-V block resulting in complete A-V block with idioventricular rhythm, or injury pattern resulting in ventricular fibrillation or ventricular flutter, were recorded. At sublethal doses no arrhythic manifestations were recorded, while S-T segment elevation upward inversion became gradually normal.

Ion transport and regulation ofTetrahymena pyriformis in isotonic and hypotonic media

SummaryThe ion and volume regulatory mechanisms ofTetrahymena pyriformis were studied in normal or hypotonic nutrient media and in isotonic inorganic media with different Na/K ratios, in conjunction with the effects of a general metabolic inhibitor (low temperature) and a specific inhibitor (iodoacetate). For K two mechanisms of active influx were found: The first is sensitive to IAc and seems to be the basic mechanism for the maintenance of the Ki/Ko gradient. The second is sensitive to cooling and related to the function of the contractile vacuole; it is also responsible for the high intracellular levels of K. The passive K efflux seems to be a basic factor for volume regulation, together with the contractile vacuole. It increases in hypotonic media and this seems to be related to structural changes of the membranes occurring in hypotonic media. For Na two mechanisms of active transport were also found: One for active Na efflux with highKm, which is associated with the contractile vacuole and another, for active Na influx with lowKm, which is inhibited by high levels of intracellular K.The electrochemical potentials of Na and K and the membrane potential (Cl Nernst potential) were also studied in isotonic inorganic media. The membrane is negatively polarized, except if Nao<5 mM when it becomes positive. In normal conditions, Na is transported outwards actively and leaks passively, while K is transported inwards actively and leaks 56 times more rapidly than Na ions.A model for the overall transport and regulation of ions inTetrahymena is proposed.

Study of the glycogenolytic action of platelet activating factor in Tetrahymena pyriformis

1. A novel action of AGEPC on non-inflammatory cells was revealed, namely the ability to stimulate glycogenolysis in Tetrahymena pyriformis cells. 2. The glycogenolytic effect of AGEPC seems to be dependent on Ca2+ transport and regulation, thus the effects are completely inhibited by Verapamil and partially by EGTA. 3. The influence of Propranolol, Labetalol, Atenolol and Theophylline in the glycogenolytic effect of AGEPC are also studied. 4. Our findings suggest that the AGEPC promoted glycogenolysis in Tetrahymena through a mechanism distinct from that of catecholamines.

Uptake of platelet-activating factor (1-O-alkyl-2-acetyl-sn-glyceryl-3-phosphorylcholine) by Tetrahymena pyriformis

Abstract 1. 1. Tetrahymena pyriformis rapidly takes up large amounts of alkyl-acetyl-GPC (PAF) without changing its mobility, shape or size. 2. 2. Time course experiments indicated that maximal uptake occurs after 1.5 and 7 min respectively, when alkyl-acetyl-GPC is added in methanol or in 2.5 mg BSA/ml saline solution. Thereafter, a continuous deacetylation of the substrate is observed. 3. 3. The study of alkyl-acetyl-GPC concentration effect on the uptake revealed that two routes of uptake, a saturable and a non-saturable one occur. Saturation is obtained for concentrations ranging from 5.7 to 7.8 × 10 9 molecules/cell . 4. 4. During the conversion of alkyl-acetyl-GPC to alkyl-acyl(long chain)-GPC, the non-saturable uptake increases.

Optimization of Conditions for Accurate Phosphonate and Total Phosphorus Assay on Lipid Samples, in Conjunction with Thin-Layer Chromatography

The range of inorganic acid normalities for maximum color formation of the phosphomolybdenum- blue complex (under heating) increases by elevating the ammonium molybdate concentration, and at a ratio of molybdate molarity/acid normality equal to 10, there is maximum color development at any acid normality in the range 1-4 ɴ with either HClO4 or H2SO4 (or their mixtures). On the basis of these features a revised method is described for the accurate determination of phosphonate-P percent of total-P. on lipid extracts and on TLC bands. The color at the final step, in both cases, is developed under the same conditions of molybdate, HClO4 and H2SO4 concentrations, thus avoiding possible errors produced by the use of two separate calibration curves.