Non-targeted discovery of high-value bio-products in Nicotiana glauca L: a potential renewable plant feedstock.

The evaluation of plant-based feedstocks is an important aspect of biorefining. Nicotiana glauca is a solanaceous, non-food crop that produces large amounts of biomass and is well adapted to grow in suboptimal conditions. In the present article, compatible sequential solvent extractions were applied to N. glauca leaves to enable the generation of enriched extracts containing higher metabolite content comparing to direct leaf extracts. Typically, between 60 to 100 metabolite components were identified within the fractions. The occurrence of plant fatty acids, fatty acid alcohols, alkanes, sterols and terpenoids was detected by gas liquid chromatography-mass spectrometry (GC-MS) and metabolite identification was confirmed by comparison of physico-chemical properties displayed by available authentic standards. Collectively, co-products such waxes, oils, fermentable sugars, and terpenoids were all identified and quantified. The enriched fractions of N. glauca revealed a high level of readily extractable hydrocarbons, oils and high value co-products. In addition, the saccharification yield and cell wall composition analyses in the stems revealed the potential of the residue material as a promising lignocellulosic substrate for the production of fermentable sugars. In conclusion a multifractional cascade for valuable compounds/commodities has been development, that uses N. glauca biomass. These data have enabled the evaluation of N. glauca material as a potential feedstock for biorefining.

Autophagy modulates the metabolism and growth of tomato fruit during development.

Although autophagy is a conserved mechanism operating across eukaryotes, its effects on crops and especially their metabolism has received relatively little attention. Indeed, whilst a few recent studies have used systems biology tools to look at the consequences of lack of autophagy in maize these focused on leaf tissues rather than the kernels. Here we utilized RNA interference (RNAi) to generate tomato plants that were deficient in the autophagy-regulating protease ATG4. Plants displayed an early senescence phenotype yet relatively mild changes in the foliar metabolome and were characterized by a reduced fruit yield phenotype. Metabolite profiling indicated that metabolites of ATG4-RNAi tomato leaves just exhibited minor alterations while that of fruit displayed bigger difference compared to the WT. In detail, many primary metabolites exhibited decreases in the ATG4-RNAi lines, such as proline, tryptophan and phenylalanine, while the representative secondary metabolites (quinic acid and 3-trans-caffeoylquinic acid) were present at substantially higher levels in ATG4-RNAi green fruits than in WT. Moreover, transcriptome analysis indicated that the most prominent differences were in the significant upregulation of organelle degradation genes involved in the proteasome or chloroplast vesiculation pathways, which was further confirmed by the reduced levels of chloroplastic proteins in the proteomics data. Furthermore, integration analysis of the metabolome, transcriptome and proteome data indicated that ATG4 significantly affected the lipid metabolism, chlorophyll binding proteins and chloroplast biosynthesis. These data collectively lead us to propose a more sophisticated model to explain the cellular co-ordination of the process of autophagy.

Adenine Nucleotide and Nicotinamide Adenine Dinucleotide Measurements in Plants.

As the principal co-factors of many metabolic pathways, the measurement of both adenine nucleotides and nicotinamide adenine dinucleotide provides important information about cellular energy metabolism. However, given their rapid and reversible conversion as well as their relatively low concentration ranges, it is difficult to measure these compounds. Here, we describe a highly sensitive and selective ion-pairing HPLC method with fluorescence detection to quantify adenine nucleotides in plants. In addition, nicotinamide adenine dinucleotide is a crucially important redox-active substrate for multiple catabolic and anabolic reactions with the ratios of NAD+ /NADH and NADP+ /NADPH being suggested as indicators of the general intracellular redox potential and hence metabolic state. Here, we describe highly sensitive enzyme cycling-based colorimetric assays (with a detection limit in the pmol range) performed subsequent to a simple extraction procedure involving acid or base extraction to allow the measurement of the cellular levels of these metabolites. © 2020 The Authors. Basic Protocol 1: Preparation of plant material for the measurement Basic Protocol 2: Measurement of ATP, ADP, and AMP via HPLC Basic Protocol 3: NAD+ /NADP+ measurements Basic Protocol 4: NADH/NADPH measurements Basic Protocol 5: Data analysis and quality control approaches.