Peter Michelsen

Diastereomeric separation of natural glycero derivatives as their 1-(1-naphthyl)ethyl carbamates by high-performance liquid chromatography

Abstract Diastereomeric separations of several natural glycero derivatives as their 1-(1-naphthyl)ethyl carbamates have been performed by high-performance liquid chromatography (HPLC). The HPLC diasteromeric separations of monoalkyl-, dialkyland diacyl- sn -glycerols are discussed. By this method it was also possible to prepare derivatives and perform enantiomeric analysis in the nanomole range. The expected stuctures of the various samples were established by mass spectrometric studies of HPLC eluates.

Syntheses and chiroptical properties of some derivatives of 1-thioglycerol

Abstract A modified synthesis of 3-thio-sn-glycerol which leads to a product of high optical purity is described. The purity was demonstrated by the use of a chiral shift reagent. 3-S-Acetyl- and 3-S-oleyl-3-thio-sn-glycerol as well as 3-acyl derivatives of 3-thio-sn-glycerol have been synthesized. The ORD and CD curves of these compounds as well as of some other derivatives of 3-thio-sn-glycerol were analyzed and discussed. The CD curves of the triacyl derivatives show a positive effect at 260 nm and a negative effect at 230–250 nm.

Induction of human basophil histamine release by a novel protein kinase C activator, sn-1,2-isopropylidene-3-decanoyl-glycerol (IpOCOC9): partial characterization of secretagogue characteristics.

Human leukocytes were found to release histamine at exposure for the synthetic glyceride derivative sn‐1,2‐isopropylidene‐3‐decanoyl‐glycerol (IpOCOC9). The following characteristics for the IpOCOC9‐induced basophil histamine release were recorded. A. In the order of 25% of the cellular histamine content was extruded at 206 μmol/l and 45% at 690 μmol/l of the compound, respectively. B. Removal of extracellular Ca2+ variably affected IpOCOC9‐triggered release. C. The presence of N‐ethylmaleimide (10 μmol/l) or p‐bromophenacylbromide (10 μmol/l) markedly reduced IpOCOC9‐induced histamine release. D. The time course of the release triggered by IpOCOC9 was intermediate to those characterizing the release triggered by 4β‐phorbol 12‐myristate 13‐acetate (PMA) and by formyl‐methionyl‐leucyl‐phenylalanine (FMLP). E. Cells desensitized to IgE‐receptor‐mediated stimulation were hyperresponsive to stimulation with IpOCOC9. F. Cells treated with a low concentration of 2‐deoxyglucose were not hyperresponsive to IpOCOC9. These data show that IpOCOC9, a PMN/leukocyte protein kinase C stimulator, acts as a non‐cytotoxic secretagogue for human basophils with a mode of action which in some, but not all respects, mimics that of PMA. In particular, IpOCOC9‐triggered release resembles that reported by other authors for hyperosmolar triggering of release by mannitol.

Chiroptical properties of S-alkyl derivatives of 1-thioglycerol

Abstract Several racemic and optically active S -alkyl derivatives of 1-thioglycerol have been synthesized. From 3- S -tetradecyl-3-thio- sn -glycerol the dioleoyl and diacetyl esters were prepared. 1,2-Dioleoyl-3- S -tetradecyl-3-thio- sn -glycerol was converted into the corresponding sulphoxide and sulphone. The chiroptical properties of these compounds were investigated by ORD and CD. Such measurements can be used in studies on the metabolism of enantiomeric acylglycerols.

Highly sensitive determination and characterization of intact cellular ester-linked phospholipids using liquid chromatography-plasma spray mass spectrometry

Liquid chromatographic class separations of common cellular phospholipids combined with plasma spray ionization of the effluents were investigated. Comparison with true thermospray ionization involving ammonium acetate buffering revealed a gain in total ionization in the plasma spray of a factor of approximately 10 using a cation-exchange column and a solvent mixture consisting of acetonitrile-methanol-water (400:100:15, v/v). Plasma spray ionization studies of bovine brain polyphosphoinositides interrelated by the phosphate content in the inositol moiety showed almost identical monoglyceride and diglyceride ion clusters, indicating possibilities of studying the biochemical turnover of such phospholipids. Plasma spray ionization liquid chromatography-mass spectrometry of bacterial membrane phospholipids (Pseudomonas fluorescens) revealed possibilities of obtaining indications of individual fatty acid compositions from the spectra of the phosphatidylinositol and phosphatidylethanolamine fractions present. Conventional gas chromatographic fatty acid analysis agreed with the direct mass spectrometric structure elucidations. Interestingly, the two phospholipid classes had different relative fatty acid compositions with a significantly higher degree of cyclic fatty acids in the phosphatidyl ethanolamines. Plasma spray ionization yielded linear dose-response curves for both the monoglyceride and diglyceride fragment signals in the selected-ion monitoring mode. The detection limit for the monoglyceride and diglyceride species of phosphatidylcholine under the chromatographic and mass spectrometric conditions used was found to be in the picogram range.

Synthesis and chiroptical properties of neutral ether lipids

Abstract A variety of neutral ether lipids was synthesized. A method for the synthesis of 1,3-O-dialkyl-sn-glycerols was developed which involves selective alkylation of 3-O-alkyl-sn-glycerols. The ORD and CD curves of the various glyceryl ethers and their esters were analyzed. The correlation between the CD sign of the acyl residue and its position in the glycerol derivative was clarified.

Modulation of human leukocyte histamine release by sn‐1,2‐isopropylidene‐3‐decanoyl‐glycerol and decanoic acid cyclopentyl methylester in comparison with effects of synthetic diacylglycerols and a phorbol ester

Previous studies have shown that the glyceride derivative, sn‐1,2 ‐isopropylidene‐3‐decanoyl‐glycerol (IpOCOC9), can trigger human leukocyte histamine release. Approximately 25 % of the total cellular histamine content is extruded in the presence of 206 # of IpOCOC9; at 69 μM, however, the secretagogue action of the compound is marginal. The characteristics of the release induced by IpOCOC9 are closely similar to those reportedly recorded at hyperosmolar triggering of basophils with mannitol, and in many respects they also mimic those observed at phorbol ester‐induced histamine release. The compound decanoic acid cyclopentyl methylester (DACPME), a structural analogue of IpOCOC9, fails to induce histamine release. IpOCOC9, but not DACPME, stimulates human polymorphonuclear leukocyte cytosolic Ca2+− and phospholipid‐dependent his‐tone III‐S kinase activity (unpublished observations). The secretagogue action of IpOCOC9, has therefore tentatively, at least partly, been attributed to a direct protein kinase C activation. In the present studies, we examined the influence of IpOCOC9 and DACPME on histamine release triggered by an ensuing exposure to anti‐IgE, the calcium ionophore A23187, formyl‐methionyl‐leucyl‐phenylalanine (EMLP), or 4β‐phor‐bol 12‐myristate 13‐acetate (PMA). It is shown that IpOCOC9‐treatment of cells results in either enhancement or reduction of the release induced by anti‐IgE or by A23187, whereas FMLP‐induced release is consistently reduced and PMA‐induced release consistently enhanced by such a treatment. Treatment of cells with DACPME enhances but does not reduce anti‐IgE‐triggered release, whereas EMLP‐induced release is not affected. Pretreatment of the cells with other putative protein kinase C activators like PMA, sn‐1‐oleoyl‐2‐acetyl‐glycerol (OAC), 1,2‐dioctanoyl‐glycerol (DiCg) or the glycerol derivative sn‐1,2‐diacetyl‐3‐decanoyl‐glycerol (DiC2OCOC9) affects secretagogue‐induced basophil histamine release according to specific patterns similar to but not identical with those recorded for IpOCOC9 and DACPME. Thus, e.g., DiC2OCOC9 consistently reduces but does not enhance anti‐IgE‐triggered release. These data show that limited structural changes of IpOCOC9 may qualitatively affect its modulating properties in the human basophil histamine release system.

Highly sensitive diastereoisomeric analysis of secondary alcohols and short-chain 1,2-diacylglycerols by capillary gas chromatography—mass spectrometry

Diastereomeric separation of short-chain 1,2-diacylglycerols has been carried out by high-resolution gas chromatography. A new versatile chiral reagent, R∼(+)-2-phenylselenopropionic acid, was investigated and compared with α-methoxy-α-trifluoromethylphenylacetic acid and α-methoxy-α-methylpentafluorophenylacetic acid as diastereomeric esters of 2-octanol. By use of the mass fragmentographic technique, it was shown that diastereomeric analysis can be performed in the femtomole range.

Digestion and absorption of a sulphoxide analogue of triacylglycerol in the rat

The stereochemistry of fat digestion and absorption was studied by feeding a triacylglycerol analogue to rats with a thoracic duct cannula. The analogue, rac-1,2-dioleoyl-3-S-tetradecyl-3-thioglycerol-S-oxide was chosen since its enantiomers exhibited high rotation in optical rotary dispersion (ORD) and circular dichroism (CD). In the chyle, triacylglycerol was the major lipid but X-1,2-diacyl-3-S-tetradecyl-3-thioglycerol-S-oxide constituted 8% of lipid weight. It was resolved by thin-layer chromatography (TLC) into two diastereomers. Each of the diastereomers were analyzed for the proportions of 1-thio-sn-glycerol/3-thio-sn-glycerol isomers by ORD and CD. The 1-thio-sn-glycerol isomers dominated for both compounds indicating that they were enriched during the absorption processes, since a racemic compound was fed. The stereospecificities are probably exerted by acyltransferase(s) during chyle lipid synthesis. The methods used will be valuable tools in studies on the metabolism of enantiomeric glycerides and also for characterization of naturally occurring sulphur-containing lipids.

Digestion and absorption of trioleoyl-thioglycerol in the rat.

A triacylglycerol analogue, rac-1,2-di-O-oleoyl-3-S-oleoyl-3-thioglycerol, was fed to rats and chyle acylglycerols were analyzed. Triacylglycerol was the dominating chyle lipid but X-triacyl-1-thioglycerol constituted approx. 6% of total chyle lipids. Its identity was verified by ultraviolet and mass spectra and its stereochemical structure by ORD and CD. The proportions of triacyl-1-thio-sn-glycerol/triacyl-3-thio-sn-glycerol were 63/37 and 78/22 in two experiments. Possible reasons for this stereospecificity are discussed. The study shows that the stereochemical configuration of lipids isolated from biological material can be assessed by ORD and CD.