Chemoenzymatic Asymmetric Synthesis of Chiral Triazole Fungicide (R)-Tebuconazole in High Optical Purity Mediated by an Epoxide Hydrolase from Rhodotorula paludigensis.

Tebuconazole is a chiral triazole fungicide used globally in agriculture as a racemic mixture, but its enantiomers exhibit significant enantioselective dissimilarities in bioactivity and environmental behaviors. The steric hindrance caused by the tert-butyl group makes it a great challenge to synthesize tebuconazole enantiomers. Here, we designed a simple chemoenzymatic approach for the asymmetric synthesis of (R)-tebuconazole, which includes the biocatalytic resolution of racemic epoxy-precursor (2-tert-butyl-2-[2-(4-chlorophenyl)ethyl] oxirane, rac-1a) by Escherichia coli/Rpeh whole cells expressed epoxide hydrolase from Rhodotorula paludigensis (RpEH), followed by a one-step chemocatalytic synthesis of (R)-tebuconazole. It was observed that (S)-1a was preferentially hydrolyzed by E. coli/Rpeh, whereas (R)-1a was retained with a specific activity of 103.8 U/g wet cells and a moderate enantiomeric ratio (E value) of 13.4, which was remarkably improved to 43.8 after optimizing the reaction conditions. Additionally, a gram-scale resolution of 200 mM rac-1a was performed using 150 mg/mL E. coli/Rpeh wet cells, resulting in the retention of (R)-1a in a 97.0% ees, a 42.5% yields, and a 40.5 g/L/d space-time yield. Subsequently, the synthesis of highly optical purity (R)-tebuconazole (>99% ee) was easily achieved through the chemocatalytic ring-opening of the epoxy-precursor (R)-1a with 1,2,4-triazole. To elucidate insight into the enantioselectivity, molecular docking simulations revealed that the unique L-shaped substrate-binding pocket of RpEH plays a crucial role in the enantioselective recognition of bulky 2,2-disubstituted oxirane 1a.

Lower prevalence of circulating invariant natural killer T (iNKT) cells in patients with acute myocardial infarction undergoing primary coronary stenting.

Invariant natural killer T cells are a unique lymphocyte subtype that can recognize lipid antigens presented by CD1d and release pro-atherogenic cytokines such as interferon-gamma. We studied the importance of iNKT cells, other lymphocyte cell types and CD11b in the peripheral blood of patients diagnosed with acute myocardial infarction (AMI) before and after primary coronary stenting. Lymphocyte population profiles and CD11b were compared between patients with AMI and healthy control subjects using flow cytometry. Both the absolute number and cell fractions of iNKT, CD3+CD4+ lymphocytes were significant lower in AMI patients than health controls. The cell fraction of NK cells was also reduced, while there was a significant increase in the cell fractions and absolute numbers of CD3+CD8+ lymphocytes, B lymphocytes and mean fluorescence intensity values of labeled CD11b. The number of iNKT cells was significantly and positively correlated with cholesterol and low-density lipoprotein levels in blood samples from AMI patients before primary coronary stenting. Logistic regression analysis demonstrated that the absolute number of iNKT cells was a significant independent predictor for restenosis during the 243 day post-operative follow-up. This study demonstrates that iNKT cell number may be a useful predictor of clinical outcome in AMI patients with primary coronary stenting.