Efficient resolution of profen ethyl ester racemates by engineered Yarrowia lipolytica Lip2p lipase

Enzyme-catalyzed enantiomer discrimination is still a great challenge for the development of industrial pharmaceutical processes. For the resolution of ibuprofen, naproxen and ketoprofen racemates, three major anti-inflammatory drugs, only lipases from Candida rugosa present a high selectivity if solvent and surfactant use is discarded. However, their catalytic activities are too low. In the present work, we demonstrate that the lipase Lip2p from the yeast Yarrowia lipolytica has a higher catalytic activity than C. rugosa lipases to hydrolyze the ethyl esters of ibuprofen, naproxen and ketoprofen, but its selectivity is not sufficient [E = 52 (S); 11 (S) and 1.5 (R) respectively]. The enantioselectivity was further improved by site-directed mutagenesis, targeted at the substrate binding site and guided by molecular modelling studies. By investigating the binding modes of the (R)- and (S)-enantiomers in the active site, two amino acid residues located in the hydrophobic substrate binding site of the lipase, namely residues 232 and 235, were identified as crucial for enantiomer discrimination and enzyme activity. The (S) enantioselectivity of Lip2p towards ethyl ibuprofen esters was rendered infinite (E ? 300) by replacing V232 by an A or C residue. Substitution of V235 by C, M, S, or T amino acids led to a great increase in the (S)-enantioselectivity (E ? 300) towards naproxen ethyl ester. Finally, the variant V232F enabled the efficient kinetic resolution of ethyl ketoprofen ester enantiomers [(R)-enantiopreference; E ? 300]. In addition to the increase in selectivity, a remarkable increase in velocity by 2.6, 2.7 and 2.5 times, respectively, was found for ibuprofen, naproxen and ketoprofen ethyl esters.     

Wiener’s test for super-Brownian motion and the Brownian snake

We prove a Wiener-type criterion for super-Brownian motion and the Brownian snake.If F is a Borel subset of ℝ ^d and x ∈ ℝ ^d , we provide a necessary and sufficientcondition for super-Brownian motion started at δ _x to immediately hit the set F. Equivalently, this condition is necessary and sufficient for the hitting time of F by theBrownian snake with initial point x to be 0. A key ingredient of the proof isan estimate showing that the hitting probability of F is comparable, up to multiplicative constants,to the relevant capacity of F. This estimate, which is of independent interest, refines previous results due to Perkins and Dynkin. An important role is played by additivefunctionals of the Brownian snake, which are investigated here via the potentialtheory of symmetric Markov processes. As a direct application of our probabilisticresults, we obtain a necessary and sufficient condition for the existence in a domain D of a positivesolution of the equation Δ; u = u ² which explodes at a given point of ∂ D. Received: 5 January 1996 / In revised form: 30 October 1996     

Enhancement of poly(vinyl ester) solubility in supercritical CO2 by partial fluorination: the key role of polymer-polymer interactions

An enhancement of poly(vinyl ester) solubility in supercritical carbon dioxide (sc-CO(2)) can be achieved by decreasing the strength of the polymer-polymer interactions. To demonstrate this, a library of statistical copolymers of vinyl acetate and vinyl trifluoroacetate was synthesized by RAFT/MADIX polymerization with varying compositions at a given number-average molecular weight. These copolymers exhibited unprecedentedly low cloud-point pressures in sc-CO(2) at 40 °C compared with previously reported poly(vinyl esters). Surface tension measurements combined with a computational approach evidenced the prominent role played by polymer-polymer interactions.     

CO<Subscript>2</Subscript>-Expanded Alkyl Lactates: A Physicochemical and Molecular Modeling Study

With the perspective of finding alternative benign media for various applications, this paper presents a study of the physicochemical behavior of some members of the alkyl lactate family when expanded by CO₂. Experimental and molecular modeling techniques have been used to determine and/or predict relevant physicochemical properties of these systems such as swelling, Kamlet–Taft parameters {polarity/polarizability (π*) and proticity or hydrogen-bond donator ability (α), dielectric constants and solubility parameters}. To complete the study of these properties, sigma profiles of the three lactates molecules have been obtained by performing quantum mechanical and phase equilibria calculations of CO₂/alkyl lactate systems by using the Peng–Robinson equation of state.