Classification of commercial cultivars of Humulus lupulus L. (hop) by chemometric pixel analysis of two dimensional nuclear magnetic resonance spectra

The development of fast and effective spectroscopic methods that can detect most compounds in an untargeted manner is of increasing interest in plant extracts fingerprinting or profiling projects. Metabolite fingerprinting by nuclear magnetic resonance (NMR) is a fast growing field which is increasingly applied for quality control of herbal products, mostly via 1D ¹H NMR coupled to multivariate data analysis. Nevertheless, signal overlap is a common problem in ¹H NMR profiles that hinders metabolites identification and results in incomplete data interpretation. Herein, we introduce a novel approach in coupling 2D NMR datasets with principal component analysis (PCA) exemplified for hop resin classification. Heteronuclear multiple bond correlation (HMBC) profile maps of hop resins (Humulus lupulus) were generated for a comparative study of 13 hop cultivars. The method described herein combines reproducible metabolite fingerprints with a minimal sample preparation effort and an experimental time of ca. 28 min per sample, comparable to that of a standard HPLC run. Moreover, HMBC spectra provide not only unequivocal assignment of hop major secondary metabolites, but also allow to identify several isomerization and degradation products of hop bitter acids including the sedative principal of hop (2-methylbut-3-en-2-ol). We do believe that combining 2D NMR datasets to chemometrics, i.e. PCA, has great potential for application in other plant metabolome projects of (commercially relevant) nutraceuticals and or herbal drugs.     

Retrograde Transport: Two (or More) Roads Diverged in an Endosomal Tree?

Some proteins and lipids traffic from the plasma membrane to the trans Golgi network (TGN)/Golgi apparatus and the endoplasmic reticulum, via the retrograde transport route. Endosomes are an obligatory through station. Whether early, recycling and late endosomes all hand off material to the TGN have remained a matter of debate. In this review, we give a short historical overview on how retrograde transport was discovered and explored. We then summarize and critically discuss data that have been put forward in favour of the existence of trafficking interfaces between each of the different endocytic localizations and the TGN. We finally point out some conceptual and technological challenges that will have to be met to establish definite conclusions for each of these scenarios.     

Endocytosis and toxicity of clostridial binary toxins depend on a clathrin‐independent pathway regulated by Rho‐GDI

Clostridial binary toxins, such as Clostridium perfringens Iota and Clostridium botulinum C2, are composed of a binding protein (Ib and C2II respectively) that recognizes distinct membrane receptors and mediates internalization of a catalytic protein (Ia and C2‐I respectively) with ADP‐ribosyltransferase activity that disrupts the actin cytoskeleton. We show here that the endocytic pathway followed by these toxins is independent of clathrin but requires the activity of dynamin and is regulated by Rho‐GDI. This endocytic pathway is similar to a recently characterized clathrin‐independent pathway followed by the interleukin‐2 (IL2) receptor. We found indeed that Ib and C2II colocalized intracellularly with the IL2 receptor but not the transferrin receptor after different times of endocytosis. Accordingly, the intracellular effects of Iota and C2 on the cytoskeleton were inhibited by inactivation of dynamin or by Rho‐GDI whereas inhibitors of clathrin‐dependent endocytosis had no protective effect.     

Complexation of β-lactoglobulin fibrils and sulfated polysaccharides

Fibrils of β-lactoglobulin, formed by heating at pH 2, were titrated with a sulfated polysaccharide (κ-carrageenan) to determine the morphology and mechanism of complex formation at low pH. Structural information on the resultant complexes was gathered using transmission electron microscopy, atomic force microscopy, Doppler electrophoresis, and small-angle neutron scattering. Electrophoresis demonstrated that the carrageenan complexed with protein fibrils until reaching a maximum complexation efficiency at a protein/polysaccharide (r) weight ratio of 5:3. Neutron scattering and microscopy indicated an increasing formation of spherical aggregates attached along the protein fibrils with increases in the carrageenan concentration. These globular particles had an average diameter of 30 nm. Small-angle neutron scattering of these complexes could be accurately described by a form factor corresponding to multistranded twisted ribbons with spherical aggregates along their contour length, arranged in a necklace configuration.     

The deceptive nature of UVA tanning versus the modest protective effects of UVB tanning on human skin

The relationship between human skin pigmentation and protection from ultraviolet (UV) radiation is an important element underlying differences in skin carcinogenesis rates. The association between UV damage and the risk of skin cancer is clear, yet a strategic balance in exposure to UV needs to be met. Dark skin is protected from UV-induced DNA damage significantly more than light skin owing to the constitutively higher pigmentation, but an as yet unresolved and important question is what photoprotective benefit, if any, is afforded by facultative pigmentation (i.e. a tan induced by UV exposure). To address that and to compare the effects of various wavelengths of UV, we repetitively exposed human skin to suberythemal doses of UVA and/or UVB over 2 weeks after which a challenge dose of UVA and UVB was given. Although visual skin pigmentation (tanning) elicited by different UV exposure protocols was similar, the melanin content and UV-protective effects against DNA damage in UVB-tanned skin (but not in UVA-tanned skin) were significantly higher. UVA-induced tans seem to result from the photooxidation of existing melanin and its precursors with some redistribution of pigment granules, while UVB stimulates melanocytes to up-regulate melanin synthesis and increases pigmentation coverage, effects that are synergistically stimulated in UVA and UVB-exposed skin. Thus, UVA tanning contributes essentially no photoprotection, although all types of UV-induced tanning result in DNA and cellular damage, which can eventually lead to photocarcinogenesis.     

Usefulness of alkaline hydrogen peroxide oxidation to analyze eumelanin and pheomelanin in various tissue samples: application to chemical analysis of human hair melanins

Eumelanin and pheomelanin in tissue samples can be specifically measured as the markers pyrrole-2,3,5-tricarboxylic acid (PTCA) and 4-amino-3-hydroxyphenylalanine after acidic permanganate oxidation and hydroiodic acid hydrolysis, respectively. Those degradation methods, although widely applied, are not easily performed in most laboratories. To overcome this difficulty, we developed alkaline H(2)O(2) oxidation in 1 M K(2)CO(3) that produces, in addition to the eumelanin marker PTCA, thiazole-2,4,5-tricarboxylic acid (TTCA) and thiazole-4,5-dicarboxylic acid (TDCA) as markers for pheomelanin and pyrrole-2,3-dicarboxylic acid (PDCA) as a marker for 5,6-dihydroxyindole-derived eumelanin. Those four degradation products can be easily separated by HPLC and analyzed with ultraviolet detection. The alkaline H(2)O(2) oxidation method is simple, reproducible and applicable to all pigmented tissues. Its application to characterize eumelanin and pheomelanin in human hair shows that PTCA and TTCA serve as specific markers for eumelanin and pheomelanin, respectively, although some caution is needed regarding the artificial production of TTCA from eumelanic tissue proteins.     

Acute stress alters auditory selective attention in humans independent of HPA: a study of evoked potentials

Background

Acute stress is a stereotypical, but multimodal response to a present or imminent challenge overcharging an organism. Among the different branches of this multimodal response, the consequences of glucocorticoid secretion have been extensively investigated, mostly in connection with long-term memory (LTM). However, stress responses comprise other endocrine signaling and altered neuronal activity wholly independent of pituitary regulation. To date, knowledge of the impact of such “paracorticoidal” stress responses on higher cognitive functions is scarce.We investigated the impact of an ecological stressor on the ability to direct selective attention using event-related potentials in humans. Based on research in rodents, we assumed that a stress-induced imbalance of catecholaminergic transmission would impair this ability.

Methodology/Principal Findings

The stressor consisted of a single cold pressor test. Auditory negative difference (Nd) and mismatch negativity (MMN) were recorded in a tonal dichotic listening task. A time series of such tasks confirmed an increased distractibility occuring 4–7 minutes after onset of the stressor as reflected by an attenuated Nd. Salivary cortisol began to rise 8–11 minutes after onset when no further modulations in the event-related potentials (ERP) occurred, thus precluding a causal relationship.This effect may be attributed to a stress-induced activation of mesofrontal dopaminergic projections. It may also be attributed to an activation of noradrenergic projections. Known characteristics of the modulation of ERP by different stress-related ligands were used for further disambiguation of causality. The conjuncture of an attenuated Nd and an increased MMN might be interpreted as indicating a dopaminergic influence. The selective effect on the late portion of the Nd provides another tentative clue for this.

Conclusions/Significance

Prior studies have deliberately tracked the adrenocortical influence on cognition, as it has proven most influential with respect to LTM. However, current cortisol-optimized study designs would have failed to detect the present findings regarding attention.