Antiatherogenic effects of ellagic acid and urolithins in vitro.

Atherosclerosis, one of the leading causes of death worldwide, is characterized by impaired endothelial function and lipid metabolism, among other factors. Ellagitannins are a class of phenolic compounds that may play a role in cardiovascular health. This work aimed to study the potential atheroprotective effects of urolithins, ellagitannin-derived gut microbiota metabolites, on different key factors in atherosclerosis development: the ability of monocytes to adhere to endothelial cells and the uptake and efflux of cholesterol by macrophages. The biotransformations urolithins undergo in peripheral cells were also evaluated. Results indicated that some urolithins and ellagic acid were able to reduce the adhesion of THP-1 monocytes to human umbilical vein endothelial cells (HUVECs) and the secretion of a cellular adhesion molecule (sVCAM-1) and pro-inflammatory cytokine (IL-6). Urolithin C, a combination of urolithins A and B, and ellagic acid also decreased the accumulation of cholesterol in THP-1-derived macrophages, but they were not able to promote cholesterol efflux. The analysis of cell media by UHPLC-ESI-MS(n) indicated urolithins and ellagic underwent extensive metabolism, with sulfate and methyl conjugation. This evidence indicates that atherosclerotic processes may be attenuated by urolithins, but future human intervention trials are required to establish if is translated in vivo.

Multiresidue pesticide analysis of tuber and root commodities by QuEchERS extraction and ultra-performance liquid chromatography coupled to tandem mass spectrometry.

A simple, rapid, and reliable multiresidue method to determine 84 pesticides in potato and carrot samples by ultra-performance liquid chromatography coupled to MS/MS has been developed and fully validated for routine analysis according to ISO/IEC 17025:2005. The method makes use of a buffered Quick, Easy, Cheap, Effective, Rugged, and Safe (QuEChERS) sample preparation procedure based on a single extraction with acidified acetonitrile, followed by partitioning with salts. Chromatographic conditions were optimized in order to achieve a rapid separation in the multiple reaction monitoring mode. Performance characteristics of the method, including an estimation of measurement uncertainty using validation data, are reported for both matrixes. Calibration curves were linear from 0.010 to 0.150 mg/kg for most compounds. The LOD and LOQ were 0.006 and 0.010 mg/kg, respectively, except for fluorocloridone, fluquinconazol, and hexitiazox, which were 0.030 and 0.050 mg/kg, respectively. Recoveries obtained were in the range 70-116%, with intraday precision values < or = 20% RSD and interday precision values < or = 25% RSD at two different concentration levels. The overall uncertainty of the method was estimated at two concentrations as being lower than 34% in all cases. The method has been applied to the analysis of 70 vegetable samples, and imidacloprid and linuron were the pesticides most frequently found in potato and carrot commodities, respectively.

Quality control evaluation of nutraceutical products from Ginkgo biloba using liquid chromatography coupled to high resolution mass spectrometry.

Analysis of 11 commercial nutraceutical products obtained from ginkgo has been performed using ultra-high performance liquid chromatography coupled to single-stage Orbitrap high resolution mass spectrometry (UHPLC-Orbitrap-MS). The main phytochemicals present in these samples were detected and quantified, utilizing a database containing 65 compounds. Phytochemicals were extracted using a mixture of an aqueous solution of methanol:water (80:20, v/v) in two sequential solid-liquid extractions. Adequate validation parameters were obtained. The validated compounds exhibited suitable linearity with determination coefficients (R(2)) higher than 0.99, and intra and inter-day precision were lower than 17 and 22%, respectively. Limits of detection (LODs) and quantification (LOQs) were calculated, ranging from 2 to 10 µg L(-1), except for myricetin (LOD, 150 µg L(-1) and LOQ, 300 µg L(-1)). Results indicate that the amount of terpenoids greatly varies among samples, ranging from 1133 (C7) to 12706 mg kg(-1) (C11). This emphasizes the importance of improve quality control in ginkgo-based products. Moreover, retrospective analysis allowed the detection of some undesirable substances as ginkgolic acid in the samples evaluated.

Influence of storage conditions in the evolution of phytochemicals in nutraceutical products applying high resolution mass spectrometry.

An adequate knowledge about possible transformation of bioactive compounds in nutraceutical products during long storage is important in order to know potential modifications of this type of compounds. In this study, one year monitoring was performed in different types of nutraceutical products based on natural extracts (green tea, soy, royal jelly and grapes) observing the appearance of new bioactive compounds, which were not detected at the initial conditions, as well as the decrease of some of the detected compounds. To determine these transformation products, an analytical procedure based on high resolution mass spectrometry (Exactive-Orbitrap analyzer) was applied. It was noted that transformation products were detected after 3 months of storage in green tea and soy products, while 6 months were necessary to observe transformation products in royal jelly.

Whole Rye Consumption Improves Blood and Liver n-3 Fatty Acid Profile and Gut Microbiota Composition in Rats.

BACKGROUND: Whole rye (WR) consumption seems to be associated with beneficial health effects. Although rye fiber and polyphenols are thought to be bioactive, the mechanisms behind the health effects of WR have yet to be fully identified. This study in rats was designed to investigate whether WR can influence the metabolism of n-3 and n-6 long-chain fatty acids (LCFA) and gut microbiota composition. METHODS: For 12 weeks, rats were fed a diet containing either 50% WR or 50% refined rye (RR). The WR diet provided more fiber (+21%) and polyphenols (+29%) than the RR diet. Fat intake was the same in both diets and particularly involved similar amounts of essential (18-carbon) n-3 and n-6 LCFAs. RESULTS: The WR diet significantly increased the 24-hour urinary excretion of polyphenol metabolites-including enterolactone-compared with the RR diet. The WR rats had significantly more n-3 LCFA-in particular, eicosapentanoic (EPA) and docosahexanoic (DHA) acids-in their plasma and liver. Compared with the RR diet, the WR diet brought significant changes in gut microbiota composition, with increased diversity in the feces (Shannon and Simpson indices), decreased Firmicutes/Bacteroidetes ratio and decreased proportions of uncultured Clostridiales cluster IA and Clostridium cluster IV in the feces. In contrast, no difference was found between groups with regards to cecum microbiota. The WR rats had lower concentrations of total short-chain fatty acids (SCFA) in cecum and feces (p<0.05). Finally, acetate was lower (p<0.001) in the cecum of WR rats while butyrate was lower (p<0.05) in the feces of WR rats. INTERPRETATION: This study shows for the first time that WR consumption results in major biological modifications-increased plasma and liver n-3 EPA and DHA levels and improved gut microbiota profile, notably with increased diversity-known to provide health benefits. Unexpectedly, WR decreased SCFA levels in both cecum and feces. More studies are needed to understand the interactions between whole rye (fiber and polyphenols) and gut microbiota and also the mechanisms of action responsible for stimulating n-3 fatty acid metabolism.

Identification and quantification of phytochemicals in nutraceutical products from green tea by UHPLC-Orbitrap-MS.

A method has been developed and validated for the simultaneous detection and quantification of phytochemicals in nutraceutical products obtained from green tea. For that purpose, ultra-high performance liquid chromatography coupled to single-stage Orbitrap high resolution mass spectrometry (UHPLC-Orbitrap-MS) has been used. A database containing 37 compounds has been used for the detection and identification of the target compounds. The developed methodology was based on solid-liquid extraction, using a mixture of methanol:H2O (80:20, v/v, pH 4), followed by dilution (10 times) with a mixture of ammonium acetate:methanol (50:50, v/v). Chromatographic conditions were optimised and full scan accurate mass data acquisition using electrospray ionisation in positive and negative ion mode was used. Moreover, all-ion fragmentation mode was used to get information of fragment ions, and they were used for identification purposes. The developed method was validated, obtaining repeatability (intra-day) and inter-day precision values (expressed as relative standard deviation, RSD) lower than 16% and 20%, respectively. Lower limits were also evaluated and limits of detection (LODs) ranged from 1 to 50 µg L(-1), while limits of quantification (LOQs) ranged from 2 to 150 µg L(-1). Recovery was performed at five levels and it ranged from 70% to 109%. Finally, this method was used to evaluate the phytochemical content in 10 samples (tablets or capsules), showing concentrations of (+)-catechin, (-)-epicatechin, gallic acid, (-)-gallocatechin and quercetin-3-O-rutinoside, ranging from 258 (C6) to 10,729 (C6) mg kg(-1).

Identification and quantification of the main isoflavones and other phytochemicals in soy based nutraceutical products by liquid chromatography-orbitrap high resolution mass spectrometry.

The specific phytochemicals composition of soy nutritional supplements is usually not labelled. Hence, 12 dietary supplements were analyzed in order to detect and identify the main phytochemicals present in these samples, using a database containing 60 compounds. Ultra-high performance liquid chromatography coupled to single-stage Orbitrap high resolution mass spectrometry (UHPLC-Orbitrap-MS) has been used. Two consecutive extractions, using as extraction solvent a mixture of methanol:water (80:20, v/v), were employed, followed by two dilutions (10 or 100 times depending on the concentration of the components in the sample) with a mixture of an aqueous solution of ammonium acetate 30mM:methanol (50:50, v/v). The method was validated, obtaining adequate recovery and precision values. Limits of detection (LODs) and quantification (LOQs) were calculated, ranging from 2 to 150µgL(-1). Isoflavones were the predominant components present in the analyzed supplements with values higher than 93% of the total amount of phytochemicals in all cases. The aglycones (genistein, daidzein, glycitein and biochanin A) as well as their three conjugated forms, ß-glucosides (genistin, daizin and glycitin) were detected and quantified, being daidzein the isoflavone detected at higher concentration in 8 out of 12 samples reported, with values ranging from 684 to 35,970mgkg(-1), whereas biochanin A was detected at very low concentrations, ranging from 18 to 50mgkg(-1). Moreover, other phytochemicals as flavones, flavonols, flavanones and phenolic acids were also detected and quantified.

Fast analysis of polyphenols in royal jelly products using automated TurboFlow™-liquid chromatography-Orbitrap high resolution mass spectrometry.

This study describes the development of a novel, simple and fast analytical method for the detection and quantification of polyphenols in royal jelly products, using an in-house database containing more than 50 compounds. The extraction method consisted of sample dilution, followed by a fast on-line system composed of turbulent flow chromatography (TurboFlow™) coupled to liquid chromatography (LC)-Exactive-Orbitrap analyzer. The total run time was 18min, including automated extraction, analytical chromatography and re-equilibration. The method was validated obtaining limits of quantification (LOQ) ranging from 10 to 150µg/kg. The linearity range was up to 2000µg/L and determination coefficients (R2) were higher than 0.994. Adequate recoveries were obtained at three concentration levels (500, 1000 and 2000µg/kg). This method was applied to the analysis of nine samples and the concentration of polyphenols ranged from 14 (apigenin) to 18,936µg/kg (ferulic acid).