High‐performance liquid chromatography with diode array detection coupled to electrospray time‐of‐flight and ion‐trap tandem mass spectrometry to identify phenolic compounds from a Cistus ladanifer aqueous extract

Introduction – Cistus ladanifer is an aromatic shrub that is widespread in the Mediterranean region. The labdanum exudate is used in the fragrance industry and has been characterised. However, there is not enough information about the phenolic content of the raw plant, the aerial part of it being a very rich source of bioactive compounds. Objective – Characterisation of the bioactive compounds of the raw plant and its aerial parts. Methodology – High‐performance liquid chromatography with diode array and electrospray ionisation mass spectrometric detection was used to carry out the comprehensive characterisation of a Cistus ladanifer shrub aqueous extract. Two different MS techniques were coupled to HPLC: time‐of‐flight mass spectrometry and tandem mass spectrometry. Results – Many well‐known compounds present in Cistus ladanifer were characterised, such as flavonoids, phenolic acids, ellagitanins, hexahydroxydiphenoyl and derivatives, and other compounds. Conclusion – The method described simultaneously separated a wide range of phenolic compounds and the proposed characterisation of the major compounds of this extract was carried out. It is important to highlight that, to our knowledge, this is the first time that a Cistus ladanifer aqueous extract from the raw plant has been characterised. Copyright © 2009 John Wiley & Sons, Ltd.     

Type 1 IFN mediates cross-talk between innate and adaptive immunity that abrogates transplantation tolerance

TLR activation of innate immunity prevents the induction of transplantation tolerance and shortens skin allograft survival in mice treated with costimulation blockade. The mechanism by which TLR signaling mediates this effect has not been clear. We now report that administration of the TLR agonists LPS (TLR4) or polyinosinic:polycytidylic acid (TLR3) to mice treated with costimulation blockade prevents alloreactive CD8(+) T cell deletion, primes alloreactive CTLs, and shortens allograft survival. The TLR4- and MyD88-dependent pathways are required for LPS to shorten allograft survival, whereas polyinosinic:polycytidylic acid mediates its effects through a TLR3-independent pathway. These effects are all mediated by signaling through the type 1 IFN (IFN-alphabeta) receptor. Administration of IFN-beta recapitulates the detrimental effects of TLR agonists on transplantation tolerance. We conclude that the type 1 IFN generated as part of an innate immune response to TLR activation can in turn activate adaptive immune responses that abrogate transplantation tolerance. Blocking of type 1 IFN-dependent pathways in patients may improve allograft survival in the presence of exogenous TLR ligands.     

Sun‐induced fluorescence – a new probe of photosynthesis: First maps from the imaging spectrometer HyPlant

Variations in photosynthesis still cause substantial uncertainties in predicting photosynthetic CO₂ uptake rates and monitoring plant stress. Changes in actual photosynthesis that are not related to greenness of vegetation are difficult to measure by reflectance based optical remote sensing techniques. Several activities are underway to evaluate the sun‐induced fluorescence signal on the ground and on a coarse spatial scale using space‐borne imaging spectrometers. Intermediate‐scale observations using airborne‐based imaging spectroscopy, which are critical to bridge the existing gap between small‐scale field studies and global observations, are still insufficient. Here we present the first validated maps of sun‐induced fluorescence in that critical, intermediate spatial resolution, employing the novel airborne imaging spectrometer HyPlant. HyPlant has an unprecedented spectral resolution, which allows for the first time quantifying sun‐induced fluorescence fluxes in physical units according to the Fraunhofer Line Depth Principle that exploits solar and atmospheric absorption bands. Maps of sun‐induced fluorescence show a large spatial variability between different vegetation types, which complement classical remote sensing approaches. Different crop types largely differ in emitting fluorescence that additionally changes within the seasonal cycle and thus may be related to the seasonal activation and deactivation of the photosynthetic machinery. We argue that sun‐induced fluorescence emission is related to two processes: (i) the total absorbed radiation by photosynthetically active chlorophyll; and (ii) the functional status of actual photosynthesis and vegetation stress.     

Continuous administration of polyphenols from aqueous rooibos (Aspalathus linearis) extract ameliorates dietary-induced metabolic disturbances in hyperlipidemic mice

The incidence of obesity and related metabolic diseases is increasing globally. Current medical treatments often fail to halt the progress of such disturbances, and plant-derived polyphenols are increasingly being investigated as a possible way to provide safe and effective complementary therapy. Rooibos (Aspalathus linearis) is a rich source of polyphenols without caloric and/or stimulant components. We have tentatively characterized 25 phenolic compounds in rooibos extract and studied the effects of continuous aqueous rooibos extract consumption in mice. The effects of this extract, which contained 25% w/w of total polyphenol content, were negligible in animals with no metabolic disturbance but were significant in hyperlipemic mice, especially in those in which energy intake was increased via a Western-type diet that increased the risk of developing metabolic complications. In these mice, we found hypolipemiant activity when given rooibos extract, with significant reductions in serum cholesterol, triglyceride and free fatty acid concentrations. Additionally, we found changes in adipocyte size and number as well as complete prevention of dietary-induced hepatic steatosis. These effects were not related to changes in insulin resistance. Among other possible mechanisms, we present data indicating that the activation of AMP-activated protein kinase (AMPK) and the resulting regulation of cellular energy homeostasis may play a significant role in these effects of rooibos extract. Our findings suggest that adding polyphenols to the daily diet is likely to help in the overall management of metabolic diseases.     

Analysis of ven3 and ven6 reticulate mutants reveals the importance of arginine biosynthesis in Arabidopsis leaf development

Arabidopsis thaliana reticulate mutants exhibit differential pigmentation of the veinal and interveinal leaf regions, a visible phenotype that often indicates impaired mesophyll development. We performed a metabolomic analysis of one ven6 (venosa6) and three ven3 reticulate mutants that revealed altered levels of arginine precursors, namely increased ornithine and reduced citrulline levels. In addition, the mutants were more sensitive than the wild-type to exogenous ornithine, and leaf reticulation and mesophyll defects of these mutants were completely rescued by exogenous citrulline. Taken together, these results indicate that ven3 and ven6 mutants experience a blockage of the conversion of ornithine into citrulline in the arginine pathway. Consistent with the participation of VEN3 and VEN6 in the same pathway, the morphological phenotype of ven3 ven6 double mutants was synergistic. Map-based cloning showed that the VEN3 and VEN6 genes encode subunits of Arabidopsis carbamoyl phosphate synthetase (CPS), which is assumed to be required for the conversion of ornithine into citrulline in arginine biosynthesis. Heterologous expression of the Arabidopsis VEN3 and VEN6 genes in a CPS-deficient Escherichia coli strain fully restored bacterial growth in minimal medium, demonstrating the enzymatic activity of the VEN3 and VEN6 proteins, and indicating a conserved role for CPS in these distinct and distant species. Detailed study of the reticulate leaf phenotype in the ven3 and ven6 mutants revealed that mesophyll development is highly sensitive to impaired arginine biosynthesis.     

Uncovering the post-embryonic functions of gametophytic- and embryonic-lethal genes

An estimated 500-1 000 Arabidopsis (Arabidopsis thaliana) genes mutate to embryonic lethality. In addition, several hundred mutations have been identified that cause gametophytic lethality. Thus, a significant fraction of the ～25,000 protein-coding genes in Arabidopsis are indispensable to the early stages of the diploid phase or to the haploid gametophytic phase. The expression patterns of many of these genes indicate that they also act later in development but, because the mutants die at such early stages, conventional methods limit the study of their roles in adult diploid plants. Here, we describe the toolset that allows researchers to assess the post-embryonic functions of plant genes for which only gametophytic- and embryonic-lethal alleles have been isolated.