Phenolic compounds from leaves of lingonberry (Vaccinium vitis-idaea L.), bilberry (Vaccinium myrtillus L.), and the natural hybrid of bilberry and lingonberry (Vaccinium x intermedium Ruthe L., hybrid bilberry) were identified using LC/TOF-MS and LC/MS/MS after extraction from the plant material in methanol in an ultrasonicator. The phenolic profiles in the plants were compared using the LC/TOF-MS responses. This is the first thorough report of phenolic compounds in hybrid bilberry. In total, 51 different phenolic compounds were identified, including flavan-3-ols, proanthocyanidins, flavonols and their glycosides, and various phenolic acid conjugates. Of the identified compounds, 35 were detected in bilberry, 36 in lingonberry, and 46 in the hybrid. To our knowledge, seven compounds were previously unreported in Vaccinium genus and many of the compounds are reported for the first time from bilberry and lingonberry.

Phenolic compounds from the lingonberry (Vaccinium vitis-idaea) were identified using LC-TOFMS, LC-MS/MS, and NMR experiments. The compounds were extracted from the plant material using methanol in an ultrasonicator and further isolated and purified using solid-phase extraction and preparative liquid chromatographic techniques. A total of 28 phenolic compounds were at least tentatively identified, including flavonols, anthocyanidins, catechins and their glycosides, and different caffeoyl and ferulic acid conjugates. This is apparently the first report of coumaroyl-hexose-hydroxyphenol, caffeoyl-hexose-hydroxyphenol, coumaroyl-hexose-hydroxyphenol, quercetin-3-O-alpha-arabinofuranoside, kaempferol-pentoside, and kaempferol-deoxyhexoside in the plant, and the flavonol acylglycosides quercetin-3-O-[4' '-(3-hydroxy-3-methylglutaroyl)]-alpha-rhamnose and kaempferol-3-O-[4' '-(3-hydroxy-3-methylglutaroyl)]-alpha-rhamnose are presented here for the first time ever. In addition, more detailed structure in comparison to earlier reports is described for some compounds previously known to exist in lingonberry.

ABSTRACT:HepaRG cells possess the unique property to differentiate in vitro and to express various functions of mature hepatocytes, including the major cytochromes P450 (P450s). In the present study, we carefully analyzed mRNA expression and activity of the major P450s and their responsiveness to three prototypical inducers, phenobarbital, rifampicin, and omeprazole, in differentiated HepaRG cell cultures over a 4-week period after low and high seeding. Only minor differences were observed in P450 activities when measured by two cocktails of probe substrates, probably related to the choice and/or concentration of substrates. Similar results were obtained from the two cell seeding conditions. Expression and activities of several P450s were dimethyl sulfoxidedependent. However, basal P450 expression and activities as well as their responsiveness to the prototypical inducers were well maintained over the 4-week period, and a good correlation was observed between transcript levels and corresponding activities. Thus, CYP1A2, CYP2B6, and CYP3A4 were found to accurately respond to their respective prototypical inducers, i.e., omeprazole, phenobarbital, and rifampicin. Likewise, basal expression of several phase II enzymes, transporters, and nuclear receptors, and response to inducers were also well preserved. More genes were found to be induced in HepaRG cells than in primary human hepatocytes, and no marked variation was noticed between the different passages. Taken together, these data support the conclusion that HepaRG cells represent a promising surrogate to primary human hepatocytes for xenobiotic metabolism and toxicity studies.

Both clopidogrel and ticlopidine significantly inhibited the CYP2B6-catalyzed bupropion hydroxylation. Patients receiving either clopidogrel or ticlopidine are likely to require dose adjustments when treated with drugs primarily metabolized by CYP2B6.

Reactive metabolites are believed to be one of the main reasons for unexpected drug-induced toxicity issues, by forming covalent adducts with cell proteins or DNA. Due to their high reactivity and short lifespan they are not directly detected by traditional analytical methods, but are most traditionally analyzed by liquid chromatography/tandem mass spectrometry (LC/MS/MS) after chemical trapping with nucleophilic agents such as glutathione. Here, a simple but very efficient assay was built up for screening reactive drug metabolites, utilizing stable isotope labeled glutathione, potassium cyanide and semicarbazide as trapping agents and highly sensitive ultra-performance liquid chromatography/time-of-flight mass spectrometry (UPLC/TOFMS) as an analytical tool. A group of twelve structurally different compounds was used as a test set, and a large number of trapped metabolites were detected for most of them, including many conjugates not reported previously. Glutathione-trapped metabolites were detected for nine of the twelve test compounds, whereas cyanide-trapped metabolites were found for eight and semicarbazide-trapped for three test compounds. The high mass accuracy of TOFMS provided unambiguous identification of change in molecular formula by formation of a reactive metabolite. In addition, use of a mass defect filter was found to be a usable tool when mining the trapped conjugates from the acquired data. The approach was shown to provide superior detection sensitivity in comparison to traditional methods based on neutral loss or precursor ion scanning with a triple quadrupole mass spectrometer, and clearly more efficient detection and characterization of reactive drug metabolites with a simpler test setup.

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Bilberry (Vaccinium myrtillus) represents one of the richest flavonoid sources among plants. Flavonoids play variable, species-dependent roles in plant defences. In bilberry, flavonoid metabolism is activated in response to solar radiation but not against mechanical injury. In this paper, the defence reaction and biosynthesis of phenolic compounds of bilberry was studied after infection by a fungal endophyte (Paraphaeosphaeria sp.) and a pathogen (Botrytis cinerea). The defence response of bilberry was faster against the endophyte than the pathogen. All flavonoid biosynthesis genes tested were activated by each infection. Biosynthesis and accumulation of phenolic acids, flavan-3-ols and oligomeric proanthocyanidins were clearly elevated in both infected samples. Infection by the pathogen promoted specifically accumulation of epigallocatechin, quercetin-3-glucoside, quercetin-3-O-α-rhamnoside, quercetin-3-O-(4"-HMG)-R-rhamnoside, chlorogenic acid and coumaroyl quinic acid. The endophyte-infected plants had a higher content of quercetin-3-glucuronide and coumaroyl iridoid. Therefore, accumulation of individual phenolic compounds could be specific for each infection. Quantity of insoluble proanthocyanidins was the highest in control plants, suggesting that they might act as storage compounds and become activated by degradation upon infection.