Possible Obesogenic Effects of Bisphenols Accumulation in the Human Brain.

Evidence of bisphenols' obesogenic effects on humans is mixed and inconsistent. We aimed to explore the presence of bisphenol A (BPA), bisphenol F (BPF) and chlorinated BPA (ClBPA), collectively called the bisphenols, in different brain regions and their association with obesity using post-mortem hypothalamic and white matter brain material from twelve pairs of obese (body mass index (BMI) >30 kg/m2) and normal-weight individuals (BMI <25 kg/m2). Mean ratios of hypothalamus:white matter for BPA, BPF and ClBPA were 1.5, 0.92, 0.95, respectively, suggesting no preferential accumulation of the bisphenols in the grey matter (hypothalamic) or white matter-enriched brain areas. We observed differences in hypothalamic concentrations among the bisphenols, with highest median level detected for ClBPA (median: 2.4 ng/g), followed by BPF (2.2 ng/g) and BPA (1.2 ng/g); similar ranking was observed for the white matter samples (median for: ClBPA-2.5 ng/g, BPF-2.3 ng/g, and BPA-1.0 ng/g). Furthermore, all bisphenol concentrations, except for white-matter BPF were associated with obesity (p < 0.05). This is the first study reporting the presence of bisphenols in two distinct regions of the human brain. Bisphenols accumulation in the white matter-enriched brain tissue could signify that they are able to cross the blood-brain barrier.

Ethyl Acetate Extract of Origanum vulgare L. ssp. hirtum Prevents Streptozotocin-Induced Diabetes in C57BL/6 Mice.

Type 1 diabetes (T1D) is an autoimmune disease that develops as a consequence of pancreatic ß-cell death induced by proinflammatory mediators. Because Origanum vulgare L. ssp. hirtum (Greek oregano) contains antiinflammatory molecules, we hypothesized that it might be beneficial for the treatment of T1D. An ethyl acetate extract of oregano (EAO) was prepared from the leaves by a polar extraction method. Phytochemical composition was determined by liquid chromatography-UV diode array coupled to ion-trap mass spectrometry with electrospray ionization interface (LC/DAD/ESI-MS(n) ). In vitro immunomodulatory effect of EAO was estimated by measuring proliferation (MTT) or cytokine secretion (ELISA) from immune cells. Diabetes was induced by multiple low doses of streptozotocin (MLDS) in male C57BL/6 mice and EAO was administered intraperitoneally for 10 d. Determination of cellular composition (flow cytometry) and cytokine production (ELISA) was performed on 12th d after diabetes induction. EAO suppressed the function of both macrophages and lymphocytes in vitro. In vivo, EAO treatment significantly preserved pancreatic islets and reduced diabetes incidence in MLDS-challenged mice. Besides down-modulatory effect on macrophages, EAO reduced the number of total CD4(+) and activated CD4(+) CD25(+) T cells. Furthermore, EAO affected the number of T helper 1 (Th1) and T helper 17 (Th17) cells through downregulation of their key transcription factors T-bet and RORγT. Because EAO treatment protects mice from development of hyperglycemia by reducing proinflammatory macrophage/Th1/Th17 response, this plant extract could represent a basis for future diabetes therapy.

Methanolic extract of Origanum vulgare ameliorates type 1 diabetes through antioxidant, anti-inflammatory and anti-apoptotic activity.

Type 1 diabetes (T1D), an autoimmune inflammatory disorder, develops as a consequence of pancreatic ß-cell destruction and results in hyperglycaemia. Since current T1D therapy mainly involves insulin replacement, the aim of the present study was to evaluate the therapeutic potential of Origanum vulgare L. ssp. hirtum (Greek oregano) leaf extract rich in biophenols for the treatment of T1D. The phytochemical profile of methanolic oregano extract (MOE) and aqueous oregano extract (AOE) was determined by liquid chromatography/electrospray ion-trap tandem MS (LC/DAD/ESI-MSn), while their main compounds were quantified by HPLC with diode array detection. After establishing their potent in vitro antioxidant activity, the extracts were administered to C57BL/6 mice treated with multiple low doses of streptozotocin for diabetes induction. While prophylactic AOE therapy had no impact on diabetes induction, MOE reduced diabetes incidence and preserved normal insulin secretion. In addition, MOE scavenged reactive oxygen and nitrogen species and, therefore, alleviated the need for the up-regulation of antioxidant enzymes. MOE treatment specifically attenuated the pro-inflammatory response mediated by T helper 17 cells and enhanced anti-inflammatory T helper 2 and T regulatory cells through the impact on specific signalling pathways and transcription factors. Importantly, MOE preserved ß-cells from in vitro apoptosis via blockade of caspase 3. Finally, rosmarinic acid, a predominant compound in MOE, exhibited only partial protection from diabetes induction. In conclusion, acting as an antioxidant, immunomodulator and in an anti-apoptotic manner, MOE protected mice from diabetes development. Seemingly, there is more than one compound responsible for the beneficial effect of MOE.

1H-NMR as a structural and analytical tool of intra- and intermolecular hydrogen bonds of phenol-containing natural products and model compounds.

Experimental parameters that influence the resolution of 1H-NMR phenol OH signals are critically evaluated with emphasis on the effects of pH, temperature and nature of the solvents. Extremely sharp peaks (Δν1/2≤2 Hz) can be obtained under optimized experimental conditions which allow the application of 1H-13C HMBC-NMR experiments to reveal long range coupling constants of hydroxyl protons and, thus, to provide unequivocal assignment of the OH signals even in cases of complex polyphenol natural products. Intramolecular and intermolecular hydrogen bonds have a very significant effect on 1H OH chemical shifts which cover a region from 4.5 up to 19 ppm. Solvent effects on -OH proton chemical shifts, temperature coefficients (Δδ/ΔT), OH diffusion coefficients, and nJ(13C, O1H) coupling constants are evaluated as indicators of hydrogen bonding and solvation state of phenol -OH groups. Accurate 1H chemical shifts of the OH groups can be calculated using a combination of DFT and discrete solute-solvent hydrogen bond interaction at relatively inexpensive levels of theory, namely, DFT/B3LYP/6-311++G (2d,p). Excellent correlations between experimental 1H chemical shifts and those calculated at the ab initio level can provide a method of primary interest in order to obtain structural and conformational description of solute-solvent interactions at a molecular level. The use of the high resolution phenol hydroxyl group 1H-NMR spectral region provides a general method for the analysis of complex plant extracts without the need for the isolation of the individual components.

Olive leaf extracts are a natural source of advanced glycation end product inhibitors.

Advanced glycation end products (AGEs), which are readily formed and accumulated with sustained hyperglycemia, contribute to the development of diabetic complications. As a consequence, inhibition of AGE formation constitutes an attractive therapeutic/preventive target. In the current study, we explored the phytochemical composition and the in vitro effect of two different olive leaf extracts (an aqueous and a methanolic) on AGE formation. The methanolic olive leaf extract inhibited fluorescent AGE formation in a bovine serum albumin (BSA)-ribose system, whereas the aqueous extract had no effect in both BSA-fructose and BSA-ribose systems. The phytochemical profile was investigated with liquid chromatography-ultraviolet-visible (UV-Vis) diode array coupled to electrospray ionization multistage mass spectrometry (LC/DAD/ESI-MS(n)). Quantification of the major phenolic compounds was performed with high performance liquid chromatography with UV-Vis diode array detection and nuclear magnetic resonance spectroscopy. Among the major phenolic components (luteolin, hydroxytyrosol, luteolin-4'-O-ß-D-glucopyranoside, luteolin-7-O-ß-D-glucopyranoside, and oleuropein), luteolin and luteolin-4'-O-ß-D-glucopyranoside were assigned as potent inhibitors of AGE formation. The extraction procedure greatly affects the composition and therefore the anti-glycation potential of olive leaves.

Rapid and direct low micromolar NMR method for the simultaneous detection of hydrogen peroxide and phenolics in plant extracts.

A rapid and direct low micromolar ¹H NMR method for the simultaneous identification and quantification of hydrogen peroxide and phenolic compounds in plant extracts was developed. The method is based on the highly deshielded ¹H NMR signal of H2O2 at ∼10.30 ppm in DMSO-d6 and the combined use of picric acid and low temperature, near the freezing point of the solution, in order to achieve the minimum proton exchange rate. Line widths of H2O2 below 3.8 Hz were obtained for several Greek oregano extracts which resulted in a detection limit of 0.7 µmol L⁻¹. Application of an array of NMR experiments, including 2D ¹H-¹³C HMBC, spiking of the samples with H2O2, and variable temperature experiments, resulted in the unequivocal assignment of H2O2 precluding any confusion with interferences from intrinsic phenolics in the extract.

Unprecedented ultra-high-resolution hydroxy group (1)H NMR spectroscopic analysis of plant extracts.

A general method is demonstrated for obtaining ultra-high resolution in the phenolic hydroxy group 1H NMR spectroscopic region, in DMSO-d6 solution, with the addition of picric acid. Line-width reduction by a factor of over 100 was observed, which resulted in line-widths ranging from 1.6 to 0.6 Hz. This unprecedented resolution, in combination with the shielding sensitivity of the hydroxy group absorptions to substituent effects at least up to 11 bonds distant and the application of 2D 1H-13C HMBC techniques, allows the unequivocal structure analysis of natural products with phenolic hydroxy groups in complex plant extracts.