The proposed formation of 3-hydroxy-4,5-dimethyl-2(5H)-furanone (sotolone) from 4-hydroxy-l-isoleucine (1) and the corresponding lactone 3-amino-4,5-dimethyl-3,4-dihydro-2(5H)-furanone (2) by thermally induced oxidative deamination was corroborated. The formation of sotolone was studied in model systems by reacting 1 or 2 with different carbonyl compounds in a phosphate buffer at pH 5 at 100 °C for 1 h. The amount of sotolone was quantified by stable isotope dilution assays using 13C2-labeled sotolone as internal standard and GC-MS operating in the selected ion monitoring mode. In general, α-ketoaldehydes were found to be more reactive than α-diketones. Methylglyoxal gave rise to about 64 μg sotolone per mg 1 (7.4 mol %) compared to less than 1 μg (<0.1 mol %) when reacted with 2,3-pentanedione. Using 2 as the starting material, the yields were increased to 274 μg (35.9 mol %) and 5.4 μg (0.7 mol %), respectively. The optimum pH of the reaction with HIL was 5, representing the best compromise between the lactonization step and the amino−carbonyl reaction. Significant amounts of sotolone were generated only at temperatures higher than 70 °C. The yield increased over a period of 10 h to about 210 μg/mg 1 (23.8 mol %). The Strecker degradation of 1, resulting in 3-hydroxy-2-methylbutanal, was a competitive reaction to the formation of sotolone. Keywords: 3-Hydroxy-4,5-dimethyl-2(5H)-furanone (sotolone); 4-hydroxy-l-isoleucine; oxidative deamination; Strecker degradation; quantification by isotope dilution assay; model system studies; synthesis

A systematic investigation of the human metabolism of hydroxycinnamic acid conjugates was carried out. A set of 24 potential human metabolites of coffee polyphenols has been chemically prepared, and used as analytical standards for unequivocal identifications. These included glucuronide conjugates and sulfate esters of caffeic, ferulic, isoferulic, m-coumaric and p-coumaric acids as well as their dihydro derivatives. A particular focus has been made on caffeic and 3,4-dihydroxyphenylpropionic acid derivatives, especially the sulfate conjugates, for which regioselective preparation was particularly challenging, and have so far never been identified as human metabolites. Ten out of the 24 synthesized conjugates have been identified in human plasma and/or urine after coffee consumption. A number of these conjugates were synthesized, characterized and detected as hydroxycinnamic acid metabolites for the first time. This was the case of dihydroisoferulic acid 3'-O-glucuronide, caffeic acid 3'-sulfate, as well as the sulfate and glucuronide derivatives of 3,4-dihydroxyphenylpropionic acid.

Two headspace techniques based on mass spectrometry detection (MS), electronic nose, and solid phase microextraction coupled to gas chromatography-mass spectrometry (SPME-GC/MS) were evaluated for their ability to differentiate various infant formula powders based on changes of their volatiles upon storage. The electronic nose gave unresolved MS fingerprints of the samples gas phases that were further submitted to principal component analysis (PCA). Such direct MS recording combined to multivariate treatment enabled a rapid differentiation of the infant formulas over a 4 week storage test. Although MS-based electronic nose advantages are its easy-to-use aspect and its meaningful data interpretation obtained with a high throughput (100 samples per 24 h), its greatest disadvantage is that the present compounds could not be identified and quantified. For these reasons, a SPME-GC/MS measurement was also investigated. This technique allowed the identification of saturated aldehydes as the main volatiles present in the headspace of infant milk powders. An isotope dilution assay was further developed to quantitate hexanal as a potential indicator of infant milk powder oxidation. Thus, hexanal content was found to vary from roughly 500 and 3500 microg/kg for relatively non-oxidized and oxidized infant formulas, respectively.

A new method for the quantification of N-(l-deoxy-D-fructos-l-yl)-glycine (DFG) was developed based on isotope dilution fast atom bombardment tandem mass spectrometry using 13C-labelled DFG as an internal standard. This method, which requires neither derivatization nor clean-up of the samples, was used to study the degradation of DFG under different conditions (time and pH). It was found that the decomposition of DFG at 90°C was favoured at pH 7 compared to pH 6. The higher stability of DFG at pH 6 was due to 1,2enoIization leading to the relatively stable 3-deoxglucosone.

Herein, the first enantioselective total synthesis of a number of biologically relevant (-)-epicatechin conjugates is described. The success of this synthesis relied on (i) optimized conditions for the stereospecific cyclization step leading to the catechin C ring; on (ii) efficient conjugation reactions; and on (iii) optimized deprotection sequences. These standard compounds have been subsequently used to elucidate for the first time the pattern of (-)-epicatechin conjugates present in four different human biological fluids following (-)-epicatechin absorption.