Influence of ß-Carotene and Lycopene on Oxidation of Ethyl Linoleate in One- and Disperse-Phased Model Systems.

The induction period (IP) of ethyl linoleate stressed at 60 °C was monitored via the formation of hydroperoxides. The addition of lycopene (1% w/w) increased the IP from 7.0 to 10.0 h to prove the strong antioxidative potential in contrast to ß-carotene with pro-oxidative effects (IP: 6.0 h), both showing strong scavenging activity under fast degradation. When peroxidation was induced by singlet oxygen, both carotenoids effectively inhibited the formation of hydroperoxides, with quenching activity only observed at low singlet oxygen concentrations, while scavenging still dominated. Thus, carotenoids did not interact with the introduced singlet oxygen but rather with the radical intermediates of fat oxidation. These experiments were then transferred to lecithin-based micelles more related to biological systems, where singlet oxygen was generated in the outer aqueous phase. Lycopene and ß-carotene delayed or inhibited lipid peroxidation depending on concentration. In this setup, ß-carotene showed exclusively quenching activity, while lycopene was additionally degraded to about 70%.

Oxidative Fragmentation of Aspalathin Leads to the Formation of Dihydrocaffeic Acid and the Related Lysine Amide Adduct.

In the present study, the degradation of C-glucosidic dihydrochalcone aspalathin as the major phenolic compound in rooibos (Aspalathus linearis) was investigated. Analyses by gas chromatography-mass spectrometry of aqueous aspalathin-lysine incubations after silylation showed the formation of dihydrocaffeic acid [3-(3,4-dihydroxyphenyl)-propionic acid] under oxidative conditions as a novel degradation product up to 10 mol %. High-performance liquid chromatography analyses revealed the concurrent formation of the dihydrocaffeic acid lysine amide at about 30-fold lower concentrations, which was unequivocally verified by synthesis of an authentic reference standard. The amide was also verified in aspalathin-protein incubations after enzymatic hydrolysis by high-performance liquid chromatography-tandem mass spectrometry analyses. Thus, the covalent interaction of phenolic plant compounds with proteins under mild conditions (ambient temperatures and neutral pH) was confirmed for the first time. Acid and free amide were also quantitated in rooibos teas with significantly higher values in fermented varieties. The mechanism of formation was clarified to be initiated by singlet oxygen and to include a rearrangement-fragmentation mechanism with 1,2,3,5-tetrahydroxybenzene as the counterpart.

Characterization and Quantitation of Steryl Glycosides in Solanum melongena.

Glycosylated plant sterols or steryl glycosides (SGs) are a small group of glycolipids occurring ubiquitously in plants. In contrast to free sterols, they are insufficiently characterized concerning structural variety, quantity, and biological function. In particular, the type of sugar usually attached to the C-3 hydroxy function of the respective sterol is poorly studied. Eggplants ( Solanum melongena) are rich in phytochemicals including SGs. In the present work, the unique glycosylation pattern was investigated by a highly selective LC-MS/MS method that allowed quantitation of the glucosides and galactosides of the most common sterols: cholesterol, ß-sitosterol, campesterol, and stigmasterol. The quantitatively most important structure was ß-sitosteryl ß-d-glucopyranoside, with 54.5 mg/kg fresh weight of total fruit (365.3 mg/kg dry weight) followed by stigmasteryl ß-d-glucopyranoside and campesteryl ß-d-glucopyranoside. Analyses were performed in different tissues of eggplants (i.e., exocarp and outer mesocarp vs the remaining inner part). Steryl galactosides were determined in eggplants for the first time at significantly lower concentrations by a factor of 100. Furthermore, the rare SG ß-sitosteryl ß-d-cellobioside (3-ß-sitosteryl ß-d-glucopyranosyl-(1→4)-ß-d-glucopyranoside) was detected in eggplants for the first time. Finally, UV irradiation induced the formation of the vitamin D glucosides 7-dehydrocholesteryl ß-d-glucopyranoside and cholecalciferyl ß-d-glucopyranoside at very low levels.

Structural and Sensory Characterization of Novel Sesquiterpene Lactones from Iceberg Lettuce.

Lactuca sativa var. capitate (iceberg lettuce) is a delicious vegetable and popular for its mild taste. Nevertheless, iceberg lettuce is a source of bitter substances, such as the sesquiterpene lactones. Chemical investigations on the n-butanol extract led to the isolation of three novel sesquiterpene lactones. All compounds were isolated by multilayer countercurrent chromatography followed by preparative high-performance liquid chromatography. The structures were verified by means of spectroscopic methods, including NMR and mass spectrometry techniques. For the first time 11ß,13-dihydrolactucin-8-O-sulfate (jaquinelin-8-O-sulfate) was structurally elucidated and identified in plants. In addition, the sesquiterpene lactones cichorioside B and 8-deacetylmatricarin-8-O-sulfate were identified as novel ingredients of iceberg lettuce. Further flowering plants in the daisy family Asteraceae were examined for the above three compounds. At least one of the compounds was identified in nine plants. The comparison between the lettuce butt end and the leaves of five types of the Cichorieae tribe showed an accumulation of the compounds in the butt end. Further experiments addressed the impact of sesquiterpene lactones on color formation and bitter taste.

Vitamin D3 supplementation: Response and predictors of vitamin D3 metabolites - A randomized controlled trial.

BACKGROUND & AIMS: Large parts of the population are insufficiently supplied with vitamin D, in particular when endogenous synthesis is absent. Therefore many health care providers recommend the use of vitamin D supplements. The current study aimed to investigate the efficacy of an once-daily oral dose of 20 µg vitamin D3 to improve the vitamin D status and to evaluate predictors of response. METHODS: The study was conducted as a double-blind, randomized, placebo-controlled parallel trial from January till April 2013. In total, 105 subjects (20-71 years) were allocated to receive either a vitamin D3 supplement (20 µg/d) or a placebo for 12 weeks. Circulating levels of vitamin D3 metabolites such as the 25(OH)D3 and the 24,25(OH)2D3, and biomarkers of calcium and phosphate metabolism were quantified. RESULTS: The 25(OH)D3 serum concentrations in the placebo group decreased from 38 ± 15 nmol/L at baseline to 32 ± 14 nmol/L and 32 ± 13 nmol/L at weeks 8 and 12 of the study, respectively (p < 0.01). In the vitamin D3 group, the serum 25(OH)D3 concentration increased from 38 ± 14 nmol/L at baseline to 70 ± 15 nmol/L and 73 ± 16 nmol/L at weeks 8 and 12 of vitamin D3 supplementation (p < 0.001), respectively. As a result, 94% of the vitamin D3-supplemented participants reached 25(OH)D3 concentrations of ≥50 nmol/L and thereof 46% attained 25(OH)D3 levels of ≥75 nmol/L until the end of the study. The extent of the 25(OH)D3 increase upon vitamin D3 supplementation depended on 25(OH)D3 baseline levels, age, body weight and circulating levels of triglycerides. In contrast to 25(OH)D3, the response of 24,25(OH)2D3 to the vitamin D3 treatment was affected only by baseline levels of 24,25(OH)2D3 and age. CONCLUSIONS: The average improvement of 25(OH)D3 levels in individuals who received 20 µg vitamin D3 per day during the winter months was 41 nmol/L compared to individuals without supplementation. As a result almost all participants with the vitamin D3 supplementation attained 25(OH)D3 concentrations of 50 nmol/L and higher. The suitability of 24,25(OH)2D3 as a marker of vitamin D status needs further investigation. Clinical trial registration number at clinicaltrials.gov: NCT01711905.

Lycopene inhibits the isomerization of ß-carotene during quenching of singlet oxygen and free radicals.

The present study aimed to investigate the influence of singlet oxygen and radical species on the isomerization of carotenoids. On the one hand, lycopene and ß-carotene standards were incubated with 1,4-dimethylnaphthalene-1,4-endoperoxide that produced singlet oxygen in situ. (13Z)- and (15Z)-ß-carotene were preferentially generated at low concentrations of singlet oxygen, while high concentrations resulted in formation of (9Z)-ß-carotene. The addition of different concentrations of lycopene led to the same isomerization progress of ß-carotene, but resulted in a decreased formation of (9Z)-ß-carotene and retarded degradation of (all-E)-ß-carotene. On the other hand, isomerization of ß-carotene and lycopene was induced by ABTS-radicals, too. As expected from the literature, chemical quenching was observed especially for lycopene, while physical quenching was preferred for ß-carotene. Mixtures of ß-carotene and lycopene resulted in a different isomerization progress compared to the separate ß-carotene model. As long as lycopene was present, almost no isomerization of ß-carotene was triggered; after that, strong formation of (13Z)-, (9Z)-, and (15Z)-ß-carotene was initiated. In summary, lycopene protected ß-carotene against isomerization during reactions with singlet oxygen and radicals. These findings can explain the pattern of carotenoid isomers analyzed in fruits and vegetables, where lycopene containing samples showed higher (all-E)/(9Z)-ß-carotene ratios, and also in in vivo samples such as human blood plasma.

Lettucenin sesquiterpenes contribute significantly to the browning of lettuce.

Wound-induced changes in the composition of secondary plant compounds cause the browning of processed lettuce. Cut tissues near the lettuce butt end clearly exhibit increased formation of yellow-brown pigments. This browning reaction is typically been attributed to the oxidation of polyphenols by the enzyme polyphenol oxidase (PPO). However, in our previous study on Iceberg lettuce, we showed that, besides the enzymatic polyphenol browning, other reactions must be involved in the formation of colored structures. With the present study for the first time, we isolated yellow sesquiterpenes by multilayer countercurrent chromatography (MLCCC), followed by preparative high-performance liquid chromatography (HPLC). Further analyses by nuclear magnetic resonance (NMR) and mass spectrometry (MS) techniques identified lettucenin A and three novel derivatives. We call these compounds lettucenins A1, B, and B1. Color-dilution analyses revealed these lettucenins as key chromophores in the browning of Iceberg lettuce. A time formation curve showed the accumulation of lettucenins A and B within 40 h after cutting. Thereafter, these structures were degraded to unknown colored compounds. Lettucenin A was verified in five varieties of Lactuca. In contrast to that, lettucenin A was present only at trace levels in five varieties of Cichorium. Therefore, lettucenin A might be used as a chemosystematic marker of the genus Lactuca.

Isolation of phenolic compounds from iceberg lettuce and impact on enzymatic browning.

Enzymatic browning is generally reported as the reaction between phenolic substances and enzymes. The quality of iceberg lettuce is directly linked to this discoloration. In particular, the color change of lettuce stems considerably reduces consumer acceptance and thus decreases sales revenue of iceberg lettuce. Ten phenolic compounds (caffeic acid, chlorogenic acid, phaseolic acid, chicoric acid, isochlorogenic acid, luteolin-7-O-glucuronide, quercetin-3-O-glucuronide, quercetin-3-O-galactoside, quercetin-3-O-glucoside, and quercetin-3-O-(6″-malonyl)-glucoside) were isolated from Lactuca sativa var. capitata by multilayer countercurrent chromatography (MLCCC) and preparative high-performance liquid chromatography (HPLC). In addition, syringin was identified for the first time in iceberg lettuce. This polyphenolic ingredient was previously not mentioned for the family of Cichorieae in general. The purity and identity of isolated compounds were confirmed by different NMR experiments, HPLC-DAD-MS, and HR-MS techniques. Furthermore, the relationship between discoloration of iceberg lettuce and enzymatic browning was thoroughly investigated. Unexpectedly, the total concentration of phenolic compounds and the activity of polyphenol oxidase were not directly related to the browning processes. Results of model incubation experiments of plant extract solutions led to the conclusion that in addition to the typical enzymatic browning induced by polyphenol oxidases, further mechanisms must be involved to explain total browning of lettuce.

Oxidation of the dihydrochalcone aspalathin leads to dimerization.

Aspalathin and nothofagin are typical ingredients of unfermented rooibos (Krafczyk, N.; Glomb, M. A. J. Agric. Food Chem. 2008, 56, 3368). During oxidation these dihydrochalcones were degraded to higher molecular weight browning products under aerated and nonenzymatic conditions. In the early stages of browning reactions aspalathin formed two dimers. These two compounds were unequivocally established as atropisomers stemming from oxidative A to B ring coupling. Multilayer countercurrent chromatography (MLCCC) and preparative high-performance liquid chromatography (HPLC) were applied to obtain pure substances. The purity and identity of isolated dimers were confirmed by different NMR experiments, HPLC-DAD-MS, and HR-MS. In parallel to the formation of chromophores during the fermentation of black tea, the formation of aspalathin dimers implies an important mechanistic channel for the generation of color during the processing of rooibos.

Structure-antioxidant relationship of flavonoids from fermented rooibos.

Rooibos tea (Aspalathus linearis) contains different bioactive phenolic compounds such as dihydrochalcones, flavonols, flavanones, flavones, and flavanols. Flavonoids isolated from rooibos were subjected to different in vitro assays: Trolox equivalent antioxidant activity, LDL oxidation and Fremy's salt assays to determine the total antioxidant activity (TAA). Assay results were compared, and the structure-antioxidant relationship was investigated. A decoupled LDL oxidation test was established with the objective of having an assay adapted more to an in vivo situation. The different in vitro methods were coupled offline to HPLC-DAD. Results from these coupled offline methods were compared to the TAA to assess the usefulness of the coupling setup.