Chronic diseases affecting the central nervous system (CNS) like Alzheimer’s or Parkinson’s disease typically develop with advanced chronological age. Yet, aging at the metabolic level has been explored only sporadically in humans using biofluids in close proximity to the CNS such as the cerebrospinal fluid (CSF). We have used an untargeted liquid chromatography high-resolution mass spectrometry (LC-HRMS) based metabolomics approach to measure the levels of metabolites in the CSF of non-neurological control subjects in the age of 20 up to 74. Using a random forest-based feature selection strategy, we extracted 69 features that were strongly related to age (page 

AbstractClassical terpenoid biosynthesis involves the cyclization of the linear prenyl pyrophosphate precursors geranyl-, farnesyl-, or geranylgeranyl pyrophosphate (GPP, FPP, GGPP) and their isomers, to produce a huge number of natural compounds. Recently, it was shown for the first time that the biosynthesis of the unique homo-sesquiterpene sodorifen by Serratia plymuthica 4Rx13 involves a methylated and cyclized intermediate as the substrate of the sodorifen synthase. To further support the proposed biosynthetic pathway, we now identified the cyclic prenyl pyrophosphate intermediate pre-sodorifen pyrophosphate (PSPP). Its absolute configuration (6R,7S,9S) was determined by comparison of calculated and experimental CD-spectra of its hydrolysis product and matches with those predicted by semi-empirical quantum calculations of the reaction mechanism. In silico modeling of the reaction mechanism of the FPP C-methyltransferase (FPPMT) revealed a SN2 mechanism for the methyl transfer followed by a cyclization cascade. The cyclization of FPP to PSPP is guided by a catalytic dyad of H191 and Y39 and involves an unprecedented cyclopropyl intermediate. W46, W306, F56, and L239 form the hydrophobic binding pocket and E42 and H45 complex a magnesium cation that interacts with the diphosphate moiety of FPP. Six additional amino acids turned out to be essential for product formation and the importance of these amino acids was subsequently confirmed by site-directed mutagenesis. Our results reveal the reaction mechanism involved in methyltransferase-catalyzed cyclization and demonstrate that this coupling of C-methylation and cyclization of FPP by the FPPMT represents an alternative route of terpene biosynthesis that could increase the terpenoid diversity and structural space.

AbstractCocoa fermentation plays a crucial role in producing flavor and bioactive compounds of high demand for food and nutraceutical industries. Such fermentations are frequently described as a succession of three main groups of microorganisms (i.e., yeast, lactic acid, and acetic acid bacteria), each producing a relevant metabolite (i.e., ethanol, lactic acid, and acetic acid). Nevertheless, this view of fermentation overlooks two critical observations: the role of minor groups of microorganisms to produce valuable compounds and the influence of environmental factors (other than oxygen availability) on their biosynthesis. Dissecting the metabolome during spontaneous cocoa fermentation is a current challenge for the rational design of controlled fermentations. This study evaluates variations in the metabolic fingerprint during spontaneous fermentation of fine flavor cocoa through a multiplatform metabolomics approach. Our data suggested the presence of two phases of differential metabolic activity that correlate with the observed variations on temperature over fermentations: an exothermic and an isothermic phase. We observed a continuous increase in temperature from day 0 to day 4 of fermentation and a significant variation in flavonoids and peptides between phases. While the second phase, from day four on, was characterized for lower metabolic activity, concomitant with small upward and downward fluctuations in temperature. Our work is the first to reveal two phases of metabolic activity concomitant with two temperature phases during spontaneous cocoa fermentation. Here, we proposed a new paradigm of cocoa fermentation that considers the changes in the global metabolic activity over fermentation, thus changing the current paradigm based only on three main groups of microorganism and their primary metabolic products.

The global demand for fine-flavour cocoa has increased worldwide during the last years. Fine-flavour cocoa offers exceptional quality and unique fruity and floral flavour attributes of high demand by the world\'s elite chocolatiers. Several studies have highlighted the relevance of cocoa fermentation to produce such attributes. Nevertheless, little is known regarding the microbial interactions and biochemistry that lead to the production of these attributes on farms of industrial relevance, where traditional fermentation methods have been pre-standardized and scaled up. In this study, we have used metagenomic approaches to dissect on-farm industrial fermentations of fine-flavour cocoa. Our results revealed the presence of a shared core of nine dominant microorganisms (i.e. Limosilactobacillus fermentum, Saccharomyces cerevisiae, Pestalotiopsis rhododendri, Acetobacter aceti group, Bacillus subtilis group, Weissella ghanensis group, Lactobacillus_uc, Malassezia restricta and Malassezia globosa) between two farms located at completely different agro-ecological zones. Moreover, a community metabolic model was reconstructed and proposed as a tool to further elucidate the interactions among microorganisms and flavour biochemistry. Our work is the first to reveal a core of microorganisms shared among industrial farms, which is an essential step to process engineering aimed to design starter cultures, reducing fermentation times, and controlling the expression of undesirable phenotypes.