Polyphenols from Plant Root: An Expanding Biological Frontier

International audienc

Assessing Carnivorous Plants for the Production of Recombinant Proteins

The recovery of recombinant proteins from plant tissues is an expensive and time-consuming process involving plant harvesting, tissue extraction, and subsequent protein purification. The downstream process costs can represent up to 80% of the total cost of production. Secretion-based systems of carnivorous plants might help circumvent this problem. Drosera and Nepenthes can produce and excrete out of their tissues a digestive fluid containing up to 200 mg. L-1 of natural proteins. Based on the properties of these natural bioreactors, we have evaluated the possibility to use carnivorous plants for the production of recombinant proteins. In this context, we have set up original protocols of stable and transient genetic transformation for both Drosera and Nepenthes sp. The two major drawbacks concerning the proteases naturally present in the secretions and a polysaccharidic network composing the Drosera glue were overcome by modulating the pH of the plant secretions. At alkaline pH, digestive enzymes are inactive and the interactions between the polysaccharidic network and proteins in the case of Drosera are subdued allowing the release of the recombinant proteins. For D. capensis, a concentration of 25 μg of GFP/ml of secretion (2% of the total soluble proteins from the glue) was obtained for stable transformants. For N. alata, a concentration of 0.5 ng of GFP/ml secretions (0.5% of total soluble proteins from secretions) was reached, corresponding to 12 ng in one pitcher after 14 days for transiently transformed plants. This plant-based expression system shows the potentiality of biomimetic approaches leading to an original production of recombinant proteins, although the yields obtained here were low and did not allow to qualify these plants for an industrial platform project

A simple SDS-Page protein pattern from pitcher secretions as a new tool to distinguish Nepenthes species (Nepenthaceae)

International audiencePremise of the study - Carnivorous plants have always fascinated scientists because these plants are able to attract, capture and digest animal prey using their remarkable traps that contain digestive secretions. Nepenthes is one of the largest genera of carnivorous plants, with 120 species described thus far. Despite an outstanding diversity of trap designs, many species are often confused with each other and remain difficult to classify because they resemble pitchers or of the occurrence of interspecific hybrids. Methods - Here, we propose a new method to easily distinguish Nepenthes species based on a 1D SDS PAGE protein pattern analysis of their pitcher secretions. Intraspecific comparisons were performed between specimens growing in different environmental conditions to ascertain the robustness of this method. Key results - Our results show that, at the juvenile stage and in the absence of prey in the pitcher, an examined species is characterized by a specific and stable profile, whatever the environmental conditions. Conclusions - The method we describe here can be used as a reliable tool to easily distinguish between Nepenthes species and to help with potential identification based on the species-specific protein pattern of their pitcher secretions, which is complementary to the monograph informatio

Optimizing asteroid orbit computation for Gaia with normal points

Context. In addition to the systematic observations of known solar-system objects (SSOs), a continuous processing of new discoveries requiring fast responses is implemented as the short-term processing of Gaia SSO observations, providing alerts for ground-based follow-up observers. The common independent observation approach for the purposes of orbit computation has led to unrealistically large ephemeris prediction uncertainties when processing real Gaia data. Aims. We aim to provide ground-based observers with a cloud of sky positions that is shrunk to a fraction of the previously expected search area by making use of the characteristic features of Gaia astrometry. This enhances the efficiency of Gaia SSO follow-up network and leads to an increased rate of asteroid discoveries with reasonably constrained orbits with the help of ground-based follow-up observations of Gaia asteroids. Methods. We took advantage of the separation of positional errors of Gaia S SO observations into a random and systematic component. We treated the Gaia observations in an alternative way by collapsing up to ten observations that correspond to a single transit into a single so-called normal point. We implemented this input procedure in the Gaia S SO short-term processing pipeline and the OpenOrb software. Results. We validate our approach by performing extensive comparisons between the independent observation and normal point input methods and compare them to the observed positions of previously known asteroids. The new approach reduces the ephemeris uncertainty by a factor of between three and ten compared to the situation where each point is treated as a separate observation. Conclusions. Our new data treatment improves the sky prediction for the Gaia SSO observations by removing low-weight orbital solutions. These solutions originate from excessive curvature of observations, introduced by short-term variations of Gaia attitude on the one hand, and, as a main effect, shrinking of systematic error bars in the independent observation case on the other hand. We anticipate that a similar approach may also be utilized in a situation where observations from a single observatory dominate.Peer reviewe

Plant Milking Technology—An Innovative and Sustainable Process to Produce Highly Active Extracts from Plant Roots

We have used an original technology (Plant Milking Technology) based on aeroponic cultivation of plants associated with the gentle recovery of active ingredients from roots. Extraction of bioactive molecules was achieved by soaking the roots, still attached to the living plants, into a nontoxic solvent for a 2 h period. This nondestructive recovery process allows using the same root biomass for successive harvesting dates, in a recyclable way. We have applied this technology to Morus alba L. (mulberry tree), an emblematic tree of the Traditional Chinese Medicine (TCM). Trees were aeroponically grown in large-scale devices (100 m2) and were submitted to nitrogen deprivation to increase the content in active molecules (prenylated flavonoids). The Plant Milking technology applied to Morus alba L. allowed to produce an extract enriched in prenylated compounds (18-fold increase when compared to commercial root extract). Prenylated flavonoids (moracenin A and B, kuwanon C, wittiorumin F, morusin) presented a high affinity for the aged-associated collagenase enzyme, which was confirmed by activity inhibition. In accordance, M. alba extract presents efficient properties to regulate the skin matrisome, which is critical during skin aging. The benefits have been especially confirmed in vivo on wrinkle reduction, in a clinical study that involved aged women. Plant Milking technology is an optimal solution to produce active ingredients from plant roots, including trees, that meet both customer expectations around sustainability, as well as the need for an efficient production system for biotechnologists

Proteome analysis of digestive fluids in Nepenthes pitchers

International audienceBackground and Aims: Carnivorous plants have developed strategies to enable growth in nutrient-poor soils. For the genus Nepenthes, this strategy represents producing pitcher-modified leaves that can trap and digest various prey. These pitchers produce a digestive fluid composed of proteins, including hydrolytic enzymes. The focus of this study was on the identification of these proteins.Methods: In order to better characterize and have an overview of these proteins, digestive fluid was sampled from pitchers at different stages of maturity from five species of Nepenthes (N. mirabilis, N. alata, N. sanguinea, N. bicalcarata and N. albomarginata) that vary in their ecological niches and grew under different conditions. Three complementary approaches based on transcriptomic resources, mass spectrometry and in silico analysis were used.Key Results: This study permitted the identification of 29 proteins excreted in the pitchers. Twenty of these proteins were never reported in Nepenthes previously and included serine carboxypeptidases, α- and β-galactosidases, lipid transfer proteins and esterases/lipases. These 20 proteins display sequence signals allowing their secretion into the pitcher fluid.Conclusions:Nepenthes pitcher plants have evolved an arsenal of enzymes to digest prey caught in their traps. The panel of new proteins identified in this study provides new insights into the digestive process of these carnivorous plants

Protocol for automated kinetic investigation/optimization of the RAFT polymerization of various monomers

A standard protocol for the parallel optimization of Reversible Addition–Fragmentation Chain Transfer (RAFT) polymerization conditions using an automated synthesizer is described in this report. Experimental design based on the knowledge obtained from previous screening experiments and the literature is the most effective initial step of the High-Throughput Experimentation (HTE) cycle. In this paper, the polymerization procedure is explained step-by-step including preparation of stock solutions, inertization of the reactors and synthesizer environment, liquid transfers to or from the reactor vessels including sampling, as well as termination of the polymerization. Automated characterization techniques for the determination of monomer conversion and the molecular weight distribution of the polymers are discussed to complete the HTE cycle. Consequently, analysis of the data obtained from parallel screening of reactions and their products will result in the design of the next experimental cycle

Granulocyte colony-stimulating factor and respiratory status of critically ill neutropenic patients with hematologic malignancies

In patients with hematologic malignancies, respiratory status may deteriorate during neutropenia recovery. This multicenter, observational study aims to evaluate granulocyte colony-stimulating factor (G-CSF) impact on respiratory status in critically ill neutropenic patients. Among 1011 critically ill patients with hematologic malignancies, 288 were neutropenic and included in this study. 201 (70%) did not receive G-CSF at day 1 or 2. After propensity score matching for the probability of receiving G-CSF at day 1 or 2, there was no association between G-CSF and respiratory deterioration at day 14 (OR =1.19; 95%CI (0.57-2.51); p = .64). Additional sensitivity analysis in patients admitted for acute respiratory failure showed similar results (OR =1.34; 95%CI (0.5-3.59); p = .57). Among patients who recovered from neutropenia, 75% experienced respiratory deterioration during neutropenia recovery. This study confirms that neutropenia recovery is a situation at risk of respiratory deterioration. However, whether G-CSF is an aggravating factor cannot be supported by our results