Comprehensive analysis of oxidative stability and nutritional values of germinated linseed and sunflower seed oil.

Germination of seeds is known to affect the nutritional composition of cold-pressed oils. This study focused on the effects of germination on the antioxidants and oxidative stability of linseed and sunflower seed oil. As hypothesized, germination led to increased antioxidant activities and tocopherol, chlorophyll and carotenoid content. Analysis revealed a 37.2 ± 3.5-fold and 11.6 ± 1.5-fold increase in polyphenol content in linseed and sunflower seed oil from germinated seeds, respectively. Using LC-HRMS/MS, profiles with up to 69 polyphenolic substances were identified in germinated seed oils for the first time. Germination promoted lipid hydrolysis, as evidenced by NMR, with overall significant decreases in triacylglycerol content leading to increased diacylglycerol and free fatty acid values. Rancimat measurements predicted a 4.10 ± 0.52-fold longer shelf-life for germinated linseed oil. This study successfully demonstrated the potential of germination to develop PUFA-rich oils with enhanced antioxidant capacity and oxidative stability.

Identification of the Brucea javanica Constituent Brusatol as a EGFR-Tyrosine Kinase Inhibitor in a Cell-Free Assay.

Inhibitors of the tyrosine kinase (TK) activity of the epidermal growth factor receptor (EGFR) are routinely used in cancer therapy. However, there is a need to discover a new TK inhibitor. This study evaluated extracts from Brucea javanica and its components for their potential as novel EGFR-TK inhibitors. The cytotoxic effect of a g aqueous extract and its fractions was assessed by MTT assays with A549 lung cancer cells. The two fractions with the highest cytotoxicity were analyzed by LC/MS and 1H NMR. Brusatol was identified as the main constituent of these fractions, and its cytotoxic and pro-apoptotic activities were confirmed in A549 cells. To elucidate the inhibitory activity of brusatol against EGFR-TK, a specific ADP-GloTM kinase assay was used. In this assay, the IC50 value for EGFR-TK inhibition was 333.1 nM. Molecular dynamic simulations and docking experiments were performed to identify the binding pocket of brusatol to be located in the intracellular TK-domain of EGFR. This study demonstrates that brusatol inhibits EGFR-TK and therefore harbors a potential as a new therapeutic drug for the therapy of EGFR-depending cancers.

Mechanisms of lipid oxidation in water-in-oil emulsions and oxidomics-guided discovery of targeted protective approaches.

Lipid oxidation is an inevitable event during the processing, storage, and even consumption of lipid-containing food, which may cause adverse effects on both food quality and human health. Water-in-oil (W/O) food emulsions contain a high content of lipids and small water droplets, which renders them vulnerable to lipid oxidation. The present review provides comprehensive insights into the lipid oxidation of W/O food emulsions. The key influential factors of lipid oxidation in W/O food emulsions are presented systematically. To better interpret the specific mechanisms of lipid oxidation in W/O food emulsions, a comprehensive detection method, oxidative lipidomics (oxidomics), is proposed to identify novel markers, which not only tracks the chemical molecules but also considers the changes in supramolecular properties, sensory properties, and nutritional value. The microstructure of emulsions, components from both phases, emulsifiers, pH, temperature, and light should be taken into account to identify specific oxidation markers. A correlation of these novel oxidation markers with the shelf life, the organoleptic properties, and the nutritional value of W/O food emulsions should be applied to develop targeted protective approaches for limiting lipid oxidation. Accordingly, the processing parameters, the application of antioxidants and emulsifiers, as well as packing and storage conditions can be optimized to develop W/O emulsions with improved oxidative stability. This review may help in emphasizing the future research priorities of investigating the mechanisms of lipid oxidation in W/O emulsion by oxidomics, leading to practical solutions for the food industry to prevent oxidative rancidity in W/O food emulsions.

Exposure to dietary fatty acids oleic and palmitic acid alters structure and mechanotransduction of intestinal cells in vitro.

Intestinal cells are continuously exposed to food constituents while adapting to peristaltic movement and fluid shear stress. Oleic acid (OA) and palmitic acid (PA) are among the most prevalent fatty acids with respect to dietary lipids. Despite the central importance of dietary lipids for a balanced diet, awareness about potential detrimental effects related to excessive consumption is increasing; this includes toxicity, metabolic deregulation, and, particularly for cancer cells, a benefit from the uptake of fatty acids related to promotion of metastasis. Expanding on this, we started elucidating the effects of OA and PA (25-500 µM) on non-transformed human intestinal epithelial cells (HCEC-1CT) in comparison to colon carcinoma cells (HCT116), with regard to the mechanosensory apparatus. Hence, intestinal cells' motility is on the one side essential to ensure adaption to peristaltic movement and barrier function, but also to enable metastatic progression. Incubation with both OA and PA (≥ 25 µM) significantly decreased membrane fluidity of HCT116 cells, whereas the effect on HCEC-1CT was more limited. Application of rhodamine-labelled PA demonstrated that the fatty acid is incorporated into the plasma membrane of HCT116, which could not be observed in the non-tumorigenic cell line. Down-streaming into the intracellular compartment, a pronounced rearrangement of actin cytoskeleton was evident in both cell lines (OA and PA; 25 and 100 µM). This was accompanied by a variation of translocation efficiency of the mechanosensitive co-transcription factor YAP1, albeit with a stronger effect seen for PA and the cancer cells. Untargeted proteomic analysis confirmed that exposure to OA and PA could alter the response capacity of HCT116 cells to fluid shear stress. Taken together, OA and PA were able to functionally modulate the mechanosensory apparatus of intestinal cells, implying a novel role for dietary fatty acids in the regulation of intestinal pathophysiology.

Effect of Encapsulation Material on Lipid Bioaccessibility and Oxidation during In Vitro Digestion of Black Seed Oil.

Different encapsulation materials might not only affect lipid hydrolysis but also lipid oxidation during in vitro digestion. Thus, this study aimed to investigate the effect of two commonly used shell materials, starch and gelatin, on the extent of lipolysis and bioaccessibility of the main and some minor lipid compounds, as well as on the oxidative status in encapsulated black seed oil (Nigella sativa) during in vitro digestion. The study was carried out using 1H nuclear magnetic resonance spectroscopy, liquid chromatography-mass spectrometry and high-performance liquid chromatography-UV. It was shown that starch increased the level of lipid hydrolysis in black seed oil during gastric in vitro digestion, while no differences were observed in the intestinal digestates between starch-encapsulated oil and gelatin-encapsulated oil. Similarly, the bioaccessibility of minor compounds (tocopherols, sterols and thymoquinone) was not influenced by the shell materials. However, regarding lipid oxidation, a 20- and 10-fold rise of free oxylipins was obtained in oils encapsulated by starch and gelatin, respectively, after intestinal in vitro digestion. This study evidenced that gelatin rather than starch should be used for the encapsulation of oils to minimize the digestion-induced formation of bioactive oxylipins.

Pre-Operative Assessment of Micronutrients, Amino Acids, Phospholipids and Oxidative Stress in Bariatric Surgery Candidates.

Obesity has been linked to lower concentrations of fat-soluble micronutrients and higher concentrations of oxidative stress markers as well as an altered metabolism of branched chain amino acids and phospholipids. In the context of morbid obesity, the aim of this study was to investigate whether and to which extent plasma status of micronutrients, amino acids, phospholipids and oxidative stress differs between morbidly obese (n = 23) and non-obese patients (n = 13). In addition to plasma, malondialdehyde, retinol, cholesterol and triglycerides were assessed in visceral and subcutaneous adipose tissue in both groups. Plasma γ-tocopherol was significantly lower (p < 0.011) in the obese group while other fat-soluble micronutrients showed no statistically significant differences between both groups. Branched-chain amino acids (all p < 0.008) and lysine (p < 0.006) were significantly higher in morbidly obese patients compared to the control group. Malondialdehyde concentrations in both visceral (p < 0.016) and subcutaneous (p < 0.002) adipose tissue were significantly higher in the morbidly obese group while plasma markers of oxidative stress showed no significant differences between both groups. Significantly lower plasma concentrations of phosphatidylcholine, phosphatidylethanolamine, lyso-phosphatidylethanolamine (all p < 0.05) and their corresponding ether-linked analogs were observed, which were all reduced in obese participants compared to the control group. Pre-operative assessment of micronutrients in patients undergoing bariatric surgery is recommended for early identification of patients who might be at higher risk to develop a severe micronutrient deficiency post-surgery. Assessment of plasma BCAAs and phospholipids in obese patients might help to differentiate between metabolic healthy patients and those with metabolic disorders.

Manganese and iron PCP pincer complexes - the influence of sterics on structure and reactivity.

The syntheses of various manganese and iron PCP pincer complexes via a solvothermal oxidative addition methodology is described. Upon reacting [Mn2(CO)10] with the ligands (P(C-Br)PCH2-iPr) (1a) and (P(C-Br)PO-iPr) (1b), Mn(I) PCP pincer complexes [Mn(PCPCH2-iPr)(CO)3] (2a) and [Mn(-PCPO-iPr)(CO)3] (2b) were obtained. Protonation of 2a with HBF4·Et2O led to the formation of [Mn(κ3P,CH,P-P(CH)PCH2-iPr)(CO)3]BF4 (3) featuring an η2-Caryl-H agostic bond. The solvothermal reaction of 1a with [Fe2(CO)9] afforded the Fe(II) PCP pincer complex [Fe(PCPCH2-iPr)(CO)2Br] (4). Treatment of 4 with Li[HBEt3] afforded the Fe(I) complex [Fe(PCPCH2-iPr)(CO)2] (5a). When using the sterically more demanding ligands (P(C-Br)PCH2-tBu) (1c) and (P(C-Br)PO-tBu)(1d) striking differences in reactivity were observed. While neither 1c nor 1d showed any reactivity towards [Mn2(CO)10], the reaction with [Fe2(CO)9] and [Fe(CO)5] led to the formation of the Fe(I) complexes [Fe(PCPCH2-tBu)(CO)2] (5b) and [Fe(PCPO-tBu)(CO)2] (5c). X-ray structures of representative complexes are provided.

Gastric Serotonin Biosynthesis and Its Functional Role in L-Arginine-Induced Gastric Proton Secretion.

Among mammals, serotonin is predominantly found in the gastrointestinal tract, where it has been shown to participate in pathway-regulating satiation. For the stomach, vascular serotonin release induced by gastric distension is thought to chiefly contribute to satiation after food intake. However, little information is available on the capability of gastric cells to synthesize, release and respond to serotonin by functional changes of mechanisms regulating gastric acid secretion. We investigated whether human gastric cells are capable of serotonin synthesis and release. First, HGT-1 cells, derived from a human adenocarcinoma of the stomach, and human stomach specimens were immunostained positive for serotonin. In HGT-1 cells, incubation with the tryptophan hydroxylase inhibitor p-chlorophenylalanine reduced the mean serotonin-induced fluorescence signal intensity by 27%. Serotonin release of 147 ± 18%, compared to control HGT-1 cells (set to 100%) was demonstrated after treatment with 30 mM of the satiating amino acid L-Arg. Granisetron, a 5-HT3 receptor antagonist, reduced this L-Arg-induced serotonin release, as well as L-Arg-induced proton secretion. Similarly to the in vitro experiment, human antrum samples released serotonin upon incubation with 10 mM L-Arg. Overall, our data suggest that human parietal cells in culture, as well as from the gastric antrum, synthesize serotonin and release it after treatment with L-Arg via an HTR3-related mechanism. Moreover, we suggest not only gastric distension but also gastric acid secretion to result in peripheral serotonin release.

Caloric restriction increases levels of taurine in the intestine and stimulates taurine uptake by conjugation to glutathione.

Our previous study indicated increased levels of taurine-conjugated bile acids (BA) in the intestine content of mice submitted to caloric restriction (CR). In the current project, we found increased levels of free taurine and taurine conjugates, including glutathione (GSH)-taurine, in CR compared to ad libitum fed animals in the mucosa along the intestine but not in the liver. The levels of free GSH were decreased in the intestine of CR compared to ad libitum fed mice. However, the levels of oxidized GSH were not affected and were complemented by the lack of changes in the antioxidative parameters. Glutathione-S transferases (GST) enzymatic activity was increased as was the expression of GST genes along the gastrointestinal tract of CR mice. In the CR intestine, addition of GSH to taurine solution enhanced taurine uptake. Accordingly, the expression of taurine transporter (TauT) was increased in the ileum of CR animals and the levels of free and BA-conjugated taurine were lower in the feces of CR compared to ad libitum fed mice. Fittingly, BA- and GSH-conjugated taurine levels were increased in the plasma of CR mice, however, free taurine remained unaffected. We conclude that CR-triggered production and release of taurine-conjugated BA in the intestine results in increased levels of free taurine what stimulates GST to conjugate and enhance uptake of taurine from the intestine.

Ingredient-Dependent Extent of Lipid Oxidation in Margarine.

This study reports the impact of margarine-representative ingredients on its oxidative stability and green tea extract as a promising antioxidant in margarine. Oil-in-water emulsions received much attention regarding factors that influence their oxidative stability, however, water-in-oil emulsions have only been scarcely investigated. Margarine, a widely consumed water-in-oil emulsion, consists of 80-90% fat and is thermally treated when used for baking. As different types of margarine contain varying additives, their impact on the oxidative stability of margarine during processing is of pressing importance. Thus, the influence of different ingredients, such as emulsifiers, antioxidants, citric acid, ß-carotene and NaCl on the oxidative stability of margarine, heated at 80 °C for 1 h to accelerate lipid oxidation, was analyzed by the peroxide value and oxidation induction time. We found that monoglycerides influenced lipid oxidation depending on their fatty acyl chain. α-Tocopheryl acetate promoted lipid oxidation, while rosemary and green tea extract led to the opposite. Whereas green tea extract alone showed the most prominent antioxidant effect, combinations of green tea extract with citric acid, ß-carotene or NaCl increased lipid oxidation in margarine. Complementary, NMR data suggested that polyphenols in green tea extracts might decrease lipid mobility at the surface of the water droplets, which might lead to chelating of transition metals at the interface and decreasing lipid oxidation.

Bitter Sensing TAS2R50 Mediates the trans-Resveratrol-Induced Anti-inflammatory Effect on Interleukin 6 Release in HGF-1 Cells in Culture.

Recent data have shown anti-inflammatory effects for trans-resveratrol (RSV) and rosmarinic acid (RA) in various immune-competent cell models through reduction of lipopolysaccharide (LPS)-induced interleukin 6 (IL-6) release. Because both compounds have been reported to taste bitter, we hypothesized an involvement of human bitter taste sensing receptors (TAS2Rs) on IL-6 release in LPS-treated human gingival fibroblasts (HGF-1). First, the bitter taste intensity of RSV and RA was compared in a sensory trial with 10 untrained panelists, of whom 90% rated a 50 ppm of RSV in water solution more bitter than 50 ppm of RA. A mean 19 ± 6% reduction of the RSV-induced bitter taste intensity was achieved by co-administration of 50 ppm of the bitter-masking, TAS2R43 antagonist homoeriodictyol (HED). Mechanistic experiments in a stably CRISPR-Cas9-edited TAS2R43ko gastric cell model revealed involvement of TAS2R43 in the HED-evoked effect on RSV-induced proton secretion, whereas the cellular response to RSV did not depend upon TAS2R43. Next, the IL-6 modulatory effect of 100 µM RSV was studied in LPS-treated immune-competent HGF-1 cells. After 6 h of treatment, RSV reduced the LPS-induced IL-6 gene expression and protein release by -46.2 ± 12.7 and -73.9 ± 2.99%, respectively. This RSV-evoked effect was abolished by co-administration of HED. Because real-time quantitative polymerase chain reaction analyses revealed a regulation of TAS2R50 in RSV with or without HED-treated HGF-1 cells, an siRNA knockdown approach of TAS2R50 was applied to verify TAS2R50 involvement in the RSV-induced reduction of the LPS-evoked IL-6 release in HGT-1 cells.

In Vitro Digestion of Grape Seed Oil Inhibits Phospholipid-Regulating Effects of Oxidized Lipids.

The intake of dietary lipids is known to affect the composition of phospholipids in gastrointestinal cells, thereby influencing passive lipid absorption. However, dietary lipids rich in polyunsaturated fatty acids, such as vegetable oils, are prone to oxidation. Studies investigating the phospholipid-regulating effect of oxidized lipids are lacking. We aimed at identifying the effects of oxidized lipids from moderately (18.8 ± 0.39 meq O2/kg oil) and highly (28.2 ± 0.39 meq O2/kg oil) oxidized and in vitro digested cold-pressed grape seed oils on phospholipids in human gastric tumor cells (HGT-1). The oils were analyzed for their antioxidant constituents as well as their oxidized triacylglycerol profile by LC-MS/MS before and after a simulated digestion. The HGT-1 cells were treated with polar oil fractions containing epoxidized and hydroperoxidized triacylglycerols for up to six hours. Oxidized triacylglycerols from grape seed oil were shown to decrease during the in vitro digestion up to 40% in moderately and highly oxidized oil. The incubation of HGT-1 cells with oxidized lipids from non-digested oils induced the formation of cellular phospholipids consisting of unsaturated fatty acids, such as phosphocholines PC (18:1/22:6), PC (18:2/0:0), phosphoserine PS (42:8) and phosphoinositol PI (20:4/0:0), by about 40%-60%, whereas the incubation with the in vitro digested oils did not affect the phospholipid metabolism. Hence, the gastric conditions inhibited the phospholipid-regulating effect of oxidized triacylglycerols (oxTAGs), with potential implications in lipid absorption.

Free Radical Scavenging Activity of Carbonyl-Amine Adducts Formed in Soybean Oil Fortified with Phosphatidylethanolamine.

Non-enzymatic browning reactions between lipid aldehydes and aminophospholipids might play an important role in the oxidative stability of cold-pressed vegetable oils. We, therefore, aimed to study the Maillard-type reaction between hexanal, a lipid oxidation product of linoleic acid, and phosphatidylethanolamine (PE (16:0/18:1)) at a ratio of 2:1 at conditions representative of the extraction of cold-pressed soybean oils (CPSBO) and determine the radical scavenging activity of the carbonyl-amine adducts with the 2,2-diphenyl-1-picrylhydrazyl (DPPH) assay. The reaction product, 2-pentyl-3,5-dibutyl-dihydropyridine, could be identified by means of LC-ESI-QTOF-MS/MS. The formation of this nitrogen-containing heterocycle significantly increased with time and temperature (p < 0.05). The products formed during the carbonyl-amine reaction between PE (16:0/18:1) and hexanal at 60 °C showed a radical scavenging activity of approximately 20% (p < 0.05). The fraction, containing 2-pentyl-3,5-dibutyl-dihydropyridine, contributed to, but was not solely responsible for, the radical scavenging activity (p < 0.05). Incubation of CPSBO fortified with PE (16:0/18:1) at 60 °C for 60 min had the strongest radical scavenging activity of 85.1 ± 0.62%. Besides 2-pentyl-3,5-dibutyl-dihydropyridine, other carbonyl-amine adducts might impact the radical scavenging activity of CPSBO as well. The oxidative stability of CPSBO might be increased by promoting the formation of carbonyl-amine reaction products, such as 2-pentyl-3,5-dibutyl-dihydropyridine.

Simultaneous Analysis of Epoxidized and Hydroperoxidized Triacylglycerols in Canola Oil and Margarine by LC-MS.

The progress of lipid oxidation in foods is evaluated by measuring the peroxides and their scission products. However, hydrogen abstraction-independent pathways are not considered by commonly applied methods despite the known reactivity of epoxides toward biomolecules. Herein, a novel liquid chromatography tandem-mass spectrometry method was developed to detect hydroperoxidized and epoxidized triacylglycerols (TAGs) without derivatization or hydrolyzation of food samples. Epoxidized TAGs could be detected in refined canola oil at concentrations of 96.8 ± 2.08 µM, while only 5.77 ± 0.04 µM hydroperoxidized TAGs could be determined. In contrast to canola oil, margarine was more resistant to lipid oxidation since generation of epoxidized TAGs could only be marginally enhanced from 21.7 ± 0.48 to 28.8 ± 0.64 µM in margarine after treatment at 180 °C for 60 min, as also reflected by a peroxide value of 0.80 ± 0.00 mequiv O2/kg, which remained unchanged. The new method allows the assessment of food safety by the simultaneous measurement of hydroperoxidized and epoxidized TAGs without hydrolysis and laborious sample preparation.

Iron PCP Pincer Complexes in Three Oxidation States: Reversible Ligand Protonation To Afford an Fe(0) Complex with an Agostic C-H Arene Bond.

In the current investigation, the reaction of Fe2(CO)9 with the ligand precursor 2-chloro-N1,N3-bis(diisopropylphosphanyl)-N1,N3-diethylbenzene-1,3-diamine (P(C-Cl)PNEt- iPr) (1) was investigated. When a suspension of Fe2(CO)9 and 1 in CH3CN was transferred in a sealed microwave glass vial and stirred for 18 h at 110 °C the complex [Fe(PCPNEt- iPr)(CO)2Cl] (2) was obtained. In an attempt to prepare the hydride Fe(II) complex [Fe(PCPNEt- iPr)(CO)2H] (3), 2 was reacted with 1 equiv of Li[HBEt3] in THF. Instead of ligand substitution, this complex underwent a one electron reduction which led to the formation of the low-spin d7 Fe(I) complex [Fe(PCPNEt- iPr)(CO)2] (4). Exposure of a benzene solution of 4 to NO gas (1 bar) at room temperature affords the diamagnetic complex [Fe(PCPNEt- iPr)(CO)(NO)] (5). This is the first iron PCP nitrosyl complex. Protonation of 5 with HBF4·Et2O affords the cationic Fe(0) complex [Fe(κ3 P,CH,P-P(CH)PNEt- iPr)(CO)(NO)]BF4 (6) which features an η2-Caryl-H agostic bond. Even with relatively weak bases such as NEt3 the agostic C-H bond can be deprotonated with reformation of the starting material 5. Therefore, protonation of 5 is completely reversible.

Synthetic iron complexes as models for natural iron-humic compounds: Synthesis, characterization and algal growth experiments.

A series of monomeric and dimeric FeIII complexes with O,O-; O,N-; O,S-coordination motifs has been prepared and characterized by standard analytical methods in order to elucidate their potential to act as model compounds for aquatic humic acids. Due to the postulated reduction of iron in humic acids and following uptake by microorganisms, the redox behavior of the models was investigated with cyclic voltammetry. Most of the investigated compounds showed iron reduction potentials accessible to biological reducing agents. Additionally, observed reduction processes were predominantly irreversible, suggesting that subsequent reactions can take place after reduction of the iron center. Also the stability of the synthesized complexes in pure water and artificial seawater was monitored from 24h up to 21days by means of UV-Vis spectrometry. Several complexes remained stable even after 21days, showing only partially precipitation but some of them showed changes in UV-Vis spectra already after 24h which were connected to protonation/deprotonation processes as well as redox processes and degradation of the complexes. The ability to act as an iron source for primary producers was tested in algal growth experiments with two marine algae species Chlorella salina and Prymnesium parvum. Some of the compounds showed effects on the algal cultures, which are comparable with natural humic acids and better as for the samples kept under ideal conditions. Those findings help to understand which functional groups of humic acids could be responsible for the reversible iron binding and transport in aquatic humic substances.

Evaluation of Palm Oil as a Suitable Vegetable Oil for Vitamin A Fortification Programs.

Fortification programs are considered to be an effective strategy to mitigate vitamin A deficiency in populations at risk. Fortified vegetable oils rich in polyunsaturated fatty acids were shown to be prone to oxidation, leading to limited vitamin A stability. Thus, it was hypothesized that fortified oils consisting of mainly saturated fatty acids might enhance the stability of vitamin A. Mildly (peroxide value: 1.0 meq O2/kg) and highly (peroxide value: 7.5 meq O2/kg) oxidized palm oil was stored, after fortification with 60 International Units/g retinyl palmitate, in 0.5 L transparent polyethylene terephthalate bottles under cold fluorescent lighting (12 h/day) at 32 °C for 57 days. An increase of the peroxide value by 15 meq O2/kg, which was also reflected by a decrease of α-tocopherol congener by 15%-18%, was determined independent of the initial rancidity. The oxidative deterioration of the highly oxidized palm oil during storage was correlated with a significant 46% decline of the vitamin A content. However, household storage of mildly oxidized palm oil for two months did not induce any losses of vitamin A. Thus, mildly oxidized palm oil may be recommended for vitamin A fortification programs, when other sources of essential fatty acids are available.

Inhibition of topoisomerase II by phase II metabolites of resveratrol in human colon cancer cells.

SCOPE: The cytotoxic and genotoxic potential of phase II metabolites of resveratrol (RSV) was investigated in human colon cells with special emphasis on human topoisomerase (TOP) II. METHODS AND RESULTS: Cell-free screening of topoisomerase II (TOPII) inhibition by the decatenation assay showed inhibitory potential for RSV (≥200 µM) and for the first time for the three human phase II metabolites RSV-3-sulfate (≥200 µM), RSV-3-glucuronide (≥100 µM) and RSV-disulfate (≥100 µM). Conjugation at the C4'-position (RSV-4'-sulfate and RSV-4'-glucuronide) resulted in loss of the inhibitory potential in this assay. Cell-based experiments with RSV and the most abundant metabolite in humans, RSV-3-Sulf, revealed no TOP poisoning in HT29 and Caco-2 cells up to 250 µM. Further, the phase II metabolite exhibited only minor effects in the comet assay and showed negligible cytotoxic effects and apoptotic potential after 1 and 24 h incubation. Fluorescence microscopy and HPLC-DAD analysis identified cellular uptake of RSV and of RSV-3-Sulf although to a lesser extent when compared to RSV. Furthermore, within the cells fractional deconjugation of RSV-3-Sulf to the parent compound was observed. CONCLUSION: Sulfate- and glucuronide-phase II metabolites might contribute to the genotoxic potential of RSV by inhibition of TOPII activity. By deconjugation at the target site RSV-3-Sulf might serve as a pool of the parent compound.

Effect of Copper on Fatty Acid Profiles in Non- and Semifermented Teas Analyzed by LC-MS-Based Nontargeted Screening.

Unsaturated fatty acids are well-known precursors of aroma compounds, which are considered important for green tea quality. Due to the known copper-induced oxidation of unsaturated fatty acids and the broad variability of the amount of copper present in tea infusions, this paper investigates the influence of copper, added at a nontoxic concentration (300 µM) to non- and semifermented teas, on the degradation of fatty acids and fatty acid hydroperoxides thereof. The abundance of fatty acids in green and oolong tea was determined by means of a nontargeted approach applying high-resolution MS/MS. As a result, most of the fatty acids in green and oolong tea were already oxidized prior to copper addition. Addition of 300 µM CuSO4 to the oolong tea sample resulted in a decrease of 13-hydroperoxy-9Z,11E-octadecadienoic acid, an important flavor precursor, from 0.12 ± 0.02 to 0.05 ± 0.01 µM (p = 0.035), and other oxidized fatty acids decreased as well. However, copper-induced degradation of oxidized fatty acids was less pronounced in green tea compared to oolong tea, most likely due to the formation of copper complexes with low-molecular-weight compounds as evidenced by electron paramagnetic resonance spectroscopy.

Sulfated and Glucuronated trans-Resveratrol Metabolites Regulate Chemokines and Sirtuin-1 Expression in U-937 Macrophages.

The natural anti-inflammatory compound resveratrol (RES) is metabolized upon ingestion. After dietary-scale doses, plasma concentrations of sulfated and glucuronated metabolites in humans exceed those of RES. The aim of this in vitro study was to assess the effect of physiological concentrations (1 µM) of the most abundant RES metabolites (RES-3-O-sulfate, R3S; RES-disulfates, RdS; RES-3-O-glucuronide, R3G; RES-4'-O-glucuronide, R4G) on genes and proteins involved in immune cell chemotaxis and inflammation (IL-8, MIP-1b, MCP-1, CCR1, CCR2, CXCR2, SIRT1) in a cell model of lipopolysaccharide (LPS)-activated U-937 macrophages. Levels of MCP-1 mRNA were comparably decreased after 3 h of treatment with R3S and RdS by -24.7 ± 5.51 and -28.7 ± 19.2%, respectively. LPS-induced MCP-1 protein release was reduced after 3 h of treatment by R3S (-20.8 ± 13.9%) and RdS (-25.7 ± 8.29%). After a 9 h treatment, RdS also inhibited IL-8 and MIP-1b protein release by -22.9 ± 3.57 and -20.1 ± 7.00%, respectively. Glucuronides showed differential effects after 6 h of treatment, with R4G up-regulating mRNA of MIP-1b (24.5 ± 14.8%) and R3G and R4G down-regulating CXCR2 surface protein compared to cells treated with LPS alone, by -5.33 ± 4.18 and -15.2 ± 5.99%, respectively. On the contrary, R3G and R4G up-regulated SIRT1 mRNA by 22.7 ± 17.9 and 22.8 ± 16.9%, respectively, in LPS-stimulated U-937 macrophages, showing anti-inflammatory properties. In conclusion, sulfated RES metabolites show an interesting beneficial potential for attenuating inflammatory immune processes.