Novel Urinary Biomarkers of Orange Juice Consumption, Interindividual Variability, and Differences with Processing Methods.

Untargeted metabolomics identified urinary biomarkers able to discriminate between the intake of fresh hand-squeezed and industrially processed orange juices. Processing led to an upregulation in the excretion of hydroxy-polymethoxyflavone sulfates, abscisic acid, and sinapic acid 4'-glucuronide. The demethylated polymethoxyflavone metabolites were produced with a significant interindividual variability suggesting that they could originate from gut microbiota metabolism. No correlation between the excretion levels of flavanone and polymethoxyflavone metabolites was observed, showing that gut microbiota metabolism differences could be behind the interindividual variability. Subjects with a high excretion level of hesperetin conjugates could be low or high polymethoxyflavone excretors. Flavanone phase II metabolites were primarily glucuronides, while those of demethylated polymethoxyflavones were mainly sulfates. A comparative study with the available demethylated polymethoxyflavone standards suggested that the metabolites produced in humans could be tentatively 4'-hydroxy- and/or 3'-hydroxy-polymethoxyflavone sulfates. This study is the first to describe the bioavailability and metabolism of citrus juice polymethoxyflavones in humans.