A sensitive and specific method is described for quantifying various cholesterol oxidation products in foodstuffs, including 7 beta-hydroxycholesterol, cholesterol-alpha-epoxide, cholestane-triol, 7-ketocholesterol and 25-hydroxycholesterol. A chloroform-methanol extract of the food was fractionated over two successive silica columns. Two fractions containing different classes of oxysterols were then analyzed as trimethylsilyl derivatives by capillary gas liquid chromatography, using on-column injection and a temperature gradient from 70 to 200 degrees C. The detection limit was about 0.5 microgram/g dry weight for egg yolk powder. Fresh egg yolk contained only 1.2 micrograms/g of total oxides per g dry weight, showing that artifactual oxidation during the procedure was minimal. Recovery of 5 pure oxysterols added to egg yolk at levels of 6.5 and 10 micrograms/g was between 93 and 102%. In commercial egg yolk and whole egg powder stored for one year, total amounts of oxysterols ranging from 21 to 137 micrograms/g dry weight were found. In duplicates of mixed Dutch diets, total amounts ranged from 3.6 to 6.2 micrograms/g dry weight. Duplicates containing mostly fried and baked foods did not have higher levels than duplicates in which foods had been prepared by boiling or left raw. We conclude that a normal mixed diet provides only minor amounts of cholesterol oxidation products.

The coffee diterpene cafestol occurs in both robusta and arabica beans. It is present in unfiltered coffee brews and raises serum concentrations of cholesterol, triacylglycerols, and alanine aminotransferase in humans. The effects are linear with the cafestol dose. Unfiltered coffee also contains the related compound kahweol, which occurs only in the major coffee strain arabica. The activity of kahweol is unknown. In a randomized, double-blind crossover study, we gave 10 healthy male volunteers either pure cafestol (61-64 mg/d) or a mixture of cafestol (60 mg/d) and kahweol (48-54 mg/d) for 28 d. Relative to baseline values, cafestol raised mean (+/-SEM) total serum cholesterol concentrations by 0.79 +/- 0.14 mmol/L (31 +/- 5 mg/dL), low-density-lipoprotein (LDL) cholesterol by 0.57 +/- 0.13 mmol/L (22 +/- 5 mg/dL), fasting triacy-glycerols by 0.65 +/- 0.12 mmol/L (58 +/- 11 mg/dL), and alanine aminotransferase by 18 +/- 2 U/L (all P < 0.01). Relative to cafestol alone, the mixture of cafestol plus kahweol increased total cholesterol by another 0.23 +/- 0.16 mmol/L (9 +/- 6 mg/dL) (P = 0.08), LDL cholesterol by 0.23 +/- 0.16 mmol/L (9 +/- 6 mg/dL) (P = 0.09), triacylglycerols by 0.09 +/- 0.10 mmol/L (8 +/- 9 mg/dL) (P = 0.20), and alanine aminotransferase by 35 +/- 11 U/L (P = 0.004). Thus, the effect of cafestol on serum lipid concentrations was much larger than the additional effect of kahweol, and the hyperlipidemic potential of unfiltered coffee mainly depends on its cafestol content. Both cafestol and kahweol raised alanine aminotransferase concentrations, and their hyperlipidemic effect thus seems not to be coupled with their effect on liver cells.

Trans fatty acids in foods are usually analyzed by gas-liquid chromatography (GLC) of fatty acid methyl esters (FAME). However, this method may produce erroneously low values because of insufficient separation between cis and trans isomers. Separation can be optimized by preceding silver-ion thin-layer chromatography (Ag-TLC), but this is laborious. We have developed an efficient method for the separation of 18-carbon trans fatty acid isomers by combining GLC of FAME with GLC of fatty acid 4,4-dimethyloxazoline (DMOX) derivatives. We validated this method against conventional GLC of FAME, with and without preceding Ag-TLC. Fatty acid isomers were identified by comparison with standards, based on retention times and mass spectrometry. Analysis of DMOX derivatives allowed the 13t, 14t, and 15t isomers to be separated from the cis isomers. The combination of the GLC analyses of FAME and DMOX derivatives gave results comparable with those obtained by GLC of FAME after preceding Ag-TLC, while saving about 100 h of manpower per 25 samples. It allowed the identification and quantitation of 11 trans and 8 cis isomers and resulted in 25% higher values for total C 18:1 trans, compared with the analysis of FAME alone. The combination of DMOX and FAME analyses, as applied to the analysis of 14 foods that contained ruminant fat and partially hydrogenated vegetable and fish oils, indicated that the most common isomers were 11t in ruminant fats, 9t in partially hydrogenated fish fats, and either 9t or 10t in partially hydrogenated vegetable fats. The combination of GLC analyses of FAME and DMOX derivatives of fatty acids improves the quantitation of 18-carbon fatty acid isomers and may replace the laborious and time-consuming Ag-TLC. JAOCS 75, 977-985 (1998).

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Abstract. de Roos B, Meyboom S, Kosmeijer-Schuil TG, Katan MB (Wageningen Agricultural University, Wageningen, The Netherlands). Absorption and urinary excretion of the coffee diterpenes cafestol and kahweol in healthy ileostomy volunteers. J Intern Med 1998; 244: 451-60.Objectives. To determine the absorption and urinary excretion of the cholesterol-raising coffee diterpenes cafestol and kahweol in humans. Subjects and design. Nine healthy ileostomists consumed a dose of one, two or three cups of Frenchpress coffee together with a standardized breakfast on three separate days in random order. Subsequently, ileostomy effluent was collected for 14 h and urine for 24 h. Stability of cafestol and kahweol was also assessed under simulated gastrointestinal tract conditions. Main outcome measures. Absorption of diterpenes, stability of diterpenes during incubation with gastrointestinal fluids, and urinary excretion of diterpenes. Results. Corrected mean absorptions expressed as percentages of the amount consumed and the amount entering the duodenum were 67 and 88%, respectively, for cafestol, and 72 and 93%, respectively, for kahweol. We found losses of diterpenes during incubation in vitro with gastric juice (cafestol, 24%; kahweol, 32%), during storage with ileostomy effluent (cafestol, 18%; kahweol, 12%), and during freeze-drying (cafestol, 26%; kahweol, 32%). Mean excretion of glucuronidated plus sulphated conjugates in urine was 1.2% of the ingested amount for cafestol and 0.4% of the ingested amount for kahweol. Conclusions. About 70% of the ingested cafestol and kahweol is absorbed in ileostomy volunteers. Possibly, undetected metabolites are present in ileostomy effluent, resulting in lower absorption percentages. Only a small part of the diterpenes is excreted as a conjugate of glucuronic acid or sulphate in urine. Therefore, these compounds are extensively metabolized in the human body.