Charles J. W. Brooks

Isolation and identification of cholesterol α-oxide and other minor sterols in human serum

The isolation and identification of cholesterol α-oxide, coprostanol, β-sitosterol, cholest-4-en-3-one and cholesta-4, 6-dien-3-one from human serum are reported. Compounds were isolated by thin layer chromatography and were identified by gas liquid chromatography and gas chromatography-mass spectrometry (GC-MS). Data for standard sterols are also reported. The possible origins of these minor components and the significance of their presence are discussed.

Lipids of human atheroma Part 4. Characterisation of a new group of polar sterol esters from human atherosclerotic plaques

Abstract Lipid extracts from plaques of grossly diseased human aortas were fractionated by silicic acid column chromatography. Examination of selected fractions by thin-layer chromatography revealed the presence of a number of esters with polarity intermediate between that of the triglycerides and cholesterol. Acetylation of this group of compounds decreased their polarity, thus suggesting the presence of free hydroxyl groups. Alkaline hydrolysis established that the main neutral component was cholesterol, accompanied by smaller amounts of 26-hydroxycholesterol, 7β- and 7α-hydroxycholesterol and a 24-hydroxycholesterol. The polarity of the cholesterol esters was due to their acyl moieties: three of the constituent acids have been identified as two isomeric 9-hydroxyoctadeca-10,12-dienoic acids and a 13-hydroxyoctadeca-9,11-dienoic acid. The polar sterol esters constitute a new group of lipids associated with human atheroma. The possible role of peroxidised linoleates in the formation of these compounds is discussed.

Lipids of human atheroma: VIII. Oxidised derivatives of cholesteryl linoleate

Abstract Cholesterol linoleate hydroperoxides, isolated from the lipids of advanced atherosclerotic plaques of human aortas obtained during post mortem examination, have been shown to comprise a mixture of 9- and 13-hydroperoxides similar to those produced by autoxidation. Corresponding 9- and 13-keto-octadecadienoates, found in variable amounts in plaque lipids, may be satisfactorily separated by preparative thin-layer chromatography after selective reduction of the accompanying hydroperoxides with SnCl 2 . The two classes of oxidised linoleate were characterised in the form of the diols produced by reduction with LiA1 2 H 4 , using combined gas chromatography-mass spectrometry. Gas-liquid chromatography of the drimanoate esters of methyl 13-hydroxystearate derived from arterial hydroxycholesterol esters indicated a 1:1 ratio of the 13−( S )- and 13−( R )-derivatives.

Lipids of human atheroma Part 5. The occurrence of a new group of polar sterol esters in various stages of human atherosclerosis

Abstract A considerable amount of cholesterol esters of 9- and 13-hydroxyoctadecadienoic acids was detected in the more advanced stages of human aortic atheroma. Esters of this class were not detectable in “early” lesions, but formed an increasing proportion of the total extractable lipid material with increasing severity of atheroma. The possible significance of this correlation is discussed.

Application of Cholesterol oxidase in the analysis of steroids

Abstract Cholesterol oxidase has been used in the qualitative analysis of model mixtures of hydroxylic steroids. Selective oxidation of Δ 5 -3β-ols and 5α-3β-ols of the sterol series has been accomplished: the resulting ketones were completely separable by gas-liquid chromatography. Steroids with the cholane and pregnane types of side-chain were oxidised more slowly than cholesterol and those of the androstane group were almost unattacked. Steroids with modified sterol side-chains (C-24 alkylated sterols, hydroxycholesterols, sapogenins) were satisfactory substrates.

Studies of urinary metabolites of 2-(4-isobutylphenyl)propionic acid by gas-liquid chromatography-mass spectrometry

Abstract Human urinary metabolites of the oral anti-rheumatic drug RS-2(4-isobutylphenyl)propionic acid (“ibuprofen”) have been further characterised by gas chromatograph-mass spectrometry of appropriate derivatives. Gas-phase analytical resoltion of diasteromeric amides formed with R-(−)-α-phenylethylamine showed that the excreted drug was enriched in the (−)-enantiomer. Four other metabolites resulted from oxidative transformations of the isobutyl group; the stuctures of two of these had previously been reported by other workers. Retention index values (for OV-1, OV-17 and QF-1 phases) are recorded, together with salient mass spectrometric data.

Gel filtration in lipophilic solvents using hydroxyalkoxypropyl derivatives of sephadex

Abstract Liquid-gel chromatography involving a lipophilic derivative of Sephadex has been investigated in two solvent systems. The separations studied involve gel filtration mechanisms, and elution data are quoted for eighty compounds. Analysis of column performance characteristics has indicated high efficiency, good loading capacity and quantitative recovery of sample with little ‘tailing’. The potential application of these techniques to purification and group separation has been evaluated in respect of both neutral and polar lipid fractions.

Sesquiterpenoid phytoalexins from suspended callus cultures of Nicotiana tabacum

Abstract Treatment of suspended callus cultures of Nicotiana tabacum with commercial cellulase elicited four principal stress metabolites including the phytoalexin capsidiol and a second eremophilane-type diol, shown on the basis of chemical and spectroscopic evidence to be 4-epieremophil-9-ene-11 /gx, 12-diol (without assignment of absolute configuration). This diol appears to be structurally identical with debneyol isolated from N. debneyi (see accompanying paper). Among minor metabolites were an isomer and a dehydro-analogue of the diol. GC/MS of cyclic derivatives (boronates and di-t-butylsilylene derivatives) of vicinal diols was useful for their detection and characterisation. The remaining two major metabolites appeared to be phytuberol and phytuberin.

Elicitation of capsidiol accumulation in suspended callus cultures of capsicum annuum

Abstract Capsidiol was elicited in suspended callus cultures of Capsicum annuum in response to commercial cellulase (ex Trichoderma viride), or pectinase (ex Aspergiltus niger), or a sterile extract from Gliocladium deliquescens. Amounts of capsidiol up to 2.9 mg per 100 ml of culture were accumulated in response to the G. deliquescens extract. Capsidiol was the preponderant phytoalexin produced in the cultures: minor congeners were present at levels below 0.1% of the amounts of capsidiol.

Isomers of 24-ethylidenecholesterol: Gas chromatographic and mass spectrometric characterization

Abstract The C-28 isomers of 24-ethylidenecholesterol, Δ 5 -avenasterol and fucosterol, are characterized by gas chromatography and mass spectrometry. Similar results are quoted for one isomer of 24-ethylidenelophenol (citrostadienol) and 24-ethylidene-Δ 7 -cholesten-3β-ol (Δ 7 -avenasterol).