Efficient Enzymatic Preparation of Flavor Esters in Water

A straightforward biocatalytic method for the enzymatic preparation of different flavor esters starting from primary alcohols (e.g., isoamyl, n-hexyl, geranyl, cinnamyl, 2-phenethyl, and benzyl alcohols) and naturally available ethyl esters (e.g., formate, acetate, propionate, and butyrate) was developed. The biotransformations are catalyzed by an acyltransferase from Mycobacterium smegmatis (MsAcT) and proceeded with excellent yields (80-97%) and short reaction times (30-120 min), even when high substrate concentrations (up to 0.5 M) were used. This enzymatic strategy represents an efficient alternative to the application of lipases in organic solvents and a significant improvement compared with already known methods in terms of reduced use of organic solvents, paving the way to sustainable and efficient preparation of natural flavoring agents

Evaluation of pre-analytical factors affecting plasma DNA analysis.

Pre-analytical factors can significantly affect circulating cell-free DNA (cfDNA) analysis. However, there are few robust methods to rapidly assess sample quality and the impact of pre-analytical processing. To address this gap and to evaluate effects of DNA extraction methods and blood collection tubes on cfDNA yield and fragment size, we developed a multiplexed droplet digital PCR (ddPCR) assay with 5 short and 4 long amplicons targeting single copy genomic loci. Using this assay, we compared 7 cfDNA extraction kits and found cfDNA yield and fragment size vary significantly. We also compared 3 blood collection protocols using plasma samples from 23 healthy volunteers (EDTA tubes processed within 1 hour and Cell-free DNA Blood Collection Tubes processed within 24 and 72 hours) and found no significant differences in cfDNA yield, fragment size and background noise between these protocols. In 219 clinical samples, cfDNA fragments were shorter in plasma samples processed immediately after venipuncture compared to archived samples, suggesting contribution of background DNA by lysed peripheral blood cells. In summary, we have described a multiplexed ddPCR assay to assess quality of cfDNA samples prior to downstream molecular analyses and we have evaluated potential sources of pre-analytical variation in cfDNA studies

Flow-based enzymatic synthesis of melatonin and other high value tryptamine derivatives: a five-minute intensified process

To increase the uptake of biocatalytic processes by industry, it is essential to demonstrate the reliability of enzyme-based methodologies directly applied to the production of high value products. Here, a unique, efficient, and sustainable enzymatic platform for the multi-gram synthesis of melatonin, projected to generate around 1.5 billion U.S. dollars worldwide by 2021, and its analogues was developed. The system exploits the covalent immobilization of MsAcT (transferase from Mycobacterium smegmatis) onto agarose beads increasing the robustness and longevity of the immobilized biocatalyst. The fully-automated process deriving from the integration between biocatalysis and flow chemistry is designed to maximize the overall yields (58-92%) and reduce reaction times (5 min), overcoming the limitation often associated with bioprocesses and bridging the gap between lab scale and industrial production

Complete genome sequence of the epidemic and highly virulent CTX-M-15-producing H30-Rx subclone of Escherichia coli ST131

We report the complete genome sequence, including five complete plasmid sequences, of Escherichia coli ST131 isolate JJ1886. The isolate was obtained in 2007 in the United States from a patient with fatal urosepsis and belongs to the virulent, CTX-M-15-producing H30-Rx sublineage

Lipase mediated enzymatic kinetic resolution of phenylethyl halohydrins acetates: A case of study and rationalization

Racemic phenylethyl halohydrins acetates containing several groups attached to the aromatic ring were resolved via hydrolysis reaction in the presence of lipase B from Candida antarctica (Novozym\uae 435). In all cases, the kinetic resolution was highly selective (E > 200) leading to the corresponding (S)-\u3b2-halohydrin with ee > 99 %. However, the time required for an ideal 50 % conversion ranged from 15 min for 2,4-dichlorophenyl chlorohydrin acetate to 216 h for 2-chlorophenyl bromohydrin acetate. Six chlorohydrins and five bromohydrins were evaluated, the latter being less reactive. For the \u3b2-brominated substrates, steric hindrance on the aromatic ring played a crucial role, which was not observed for the \u3b2-chlorinated derivatives. To shed light on the different reaction rates, docking studies were carried out with all the substrates using MD simulations. The computational data obtained for the \u3b2-brominated substrates, based on the parameters analysed such as NAC (near attack conformation), distance between Ser-O and carbonyl-C and oxyanion site stabilization were in agreement with the experimental results. On the other hand, the data obtained for \u3b2-chlorinated substrates suggested that physical aspects such as high hydrophobicity or induced change in the conformation of the enzymatic active site are more relevant aspects when compared to steric hindrance effects

Clinical and Mucosal Immune Correlates of HIV-1 Semen Levels in Antiretroviral-Naive Men.

Background. This study was done to characterize parameters associated with semen human immunodeficiency virus (HIV)-1 ribonucleic acid (RNA) viral load (VL) variability in HIV-infected, therapy-naive men. Methods. Paired blood and semen samples were collected from 30 HIV-infected, therapy-naive men who have sex with men, and 13 participants were observed longitudinally for up to 1 year. Human immunodeficiency virus RNA, bacterial load by 16S RNA, herpesvirus (Epstein-Barr virus and cytomegalovirus [CMV]) shedding, and semen cytokines/chemokines were quantified, and semen T-cell subsets were assessed by multiparameter flow cytometry. Results. Semen HIV RNA was detected at 93% of visits, with \u3e50% of men shedding high levels of virus (defined as \u3e5000 copies/mL). In the baseline cross-sectional analysis, an increased semen HIV VL correlated with local CMV reactivation, the semen bacterial load, and semen inflammatory cytokines, particularly interleukin (IL)-8. T cells in semen were more activated than blood, and there was an increased frequency of Th17 cells and γδ-T-cells. Subsequent prospective analysis demonstrated striking interindividual variability in HIV and CMV shedding patterns, and only semen IL-8 levels and the blood VL were independently associated with semen HIV levels. Conclusions. Several clinical and immune parameters were associated with increased HIV semen levels in antiretroviral therapy-naive men, with induction of local proinflammatory cytokines potentially acting as a common pathway