RESUMEN
Carbohydrates regulate an inimitable spectrum of biological functions, yet successfully leveraging this therapeutic avenue continues to be frustrated by low affinities with glycan-specific proteins. A conspicuous exception is the interaction of monosialotetrahexosylganglioside (GM1) with the carbohydrate-recognition domain of cholera toxin from Vibrio cholerae: this is one of the strongest protein-carbohydrate interactions known. To establish the importance of a long-discussed key hydrogen bond between C2 of the terminal galactose of GM1 and the B subunit pentamer of cholera toxin (CTB5), the total synthesis of a selectively fluorinated GM1 epitope was conducted in 19 steps. This process of molecular editing (Oδ-H â Fδ-) strategically deletes the hydrogen bond donor while retaining the localized partial charge of the substituent. Comparison of the binding affinity of F-GM1/CTB5 with native GM1, the GM1 carbohydrate epitope, and meta-mononitrophenyl-α-galactoside (MNPG) revealed a trend that fully supports the importance of this key interaction. These NMR data suggest that F-GM1 binds in a closely similar conformation as native GM1. Crystallographic analyses of the complex also confirm that the OH â F bioisosteric exchange at C2 of the terminal galactose induces a ring conformation that eliminates key hydrogen bonds: these interactions are compensated for by inter- and intramolecular fluorine-specific interactions.
RESUMEN
In order to gain a better insight into pesticide and pollutant exposure in forests, a rapid and sensitive gas chromatography-tandem mass spectrometry (GC-MS/MS) method for the determination of 208 pesticide residues in leaves and needles has been established. The modified QuEChERS (quick, easy, cheap, effective, rugged and safe) approach uses 2 g of homogenized sample, acetonitrile and water as extraction agents, combined with citrate buffer for the following salting out step. The limits of quantification (LOQs) were determined to 0.0025-0.05 mg kg-1, respectively. Calibration curves showed a linear range between the respective LOQ and 1.0 mg kg-1 with coefficients of determination (R2) ≥ 0.99 for all analyzed pesticides. The recovery rates ranged from 69.7% to 92.0% with a relative standard deviation below 20%. The analysis of beech leaves, spruce and pine needles (each n = 3) provided a proof of concept for the developed methodology and revealed the presence of six pesticide residues (boscalid, epoxiconazole, fenpropimorph, lindane, terbuthylazine, terbuthylazine-desethyl). The results underline the strong need for systematic surveillance of the uncontrollable exposure of pesticides to nature.