Kinetic resolution of a drug precursor by Burkholderia cepacia lipase immobilized by different methodologies on superparamagnetic nanoparticles
Introduction
The use of lipases as catalysts is contributing to the rise of the exciting and rapidly growing area of chiral organic synthesis [1], [2], [3], [4], [5], [6], [7], [8], [9], [10]. As a matter of fact, research in this area is pursuing the discovery of new efficient enzymes, new target compounds, and also of new convenient solid supports, capable of sustaining the enzymatic activity in organic media with minimum loss. In particular, the immobilization of enzymes on magnetic particles is viewed as a very attractive strategy, for allowing their separation and recovery from the reaction media by the simple use of a magnet [11], [12], [13], [14], [15]. On the other hand, the reusability of the immobilized enzymes represents an outstanding green-chemistry approach, reducing the cost and amount of such expensive biocatalysts. It should be noticed that superparamagnetic nanoparticles exhibit a single magnetic domain in their nanocrystalline structures, and this is responsible for their exceptional magnetization properties in the presence of a magnetic field. A suitable superparamagnetic species is based on magnetite (γ-Fe3O4). Large surface area, high mass transference and rapid response to external magnetic fields are remarkable characteristics ennobling this type of nanometric support or carrier. In addition, enzyme immobilization on superparamagnetic nanoparticles can permit the recovery of the biocatalyst without spending energy, by using permanent magnets.
Recently, Burkholderia cepacia lipase (BCL, also known as Pseudomonas cepacia lipase) has been employed as biocatalyst in a variety of synthetic applications, performing reactions, such as hydrolysis [16], [17], [18], enantioselective acylation [19] and transesterification [20], [21], [22], [23]. In view of the wide usefulness of P. cepacia lipase, the possibility of exploring its recyclability using superparamagnetic nanoparticles may be quite rewarding. For this reason, a detailed study focusing on the immobilization of this lipase on superparamagnetic nanoparticles was carried out, aiming its application in the kinetic resolution of the chiral drug precursor, (RS)-2-bromo-1-(phenyl)ethanol. This compound is envisaged as a common building block to the chiral compounds Fluoxetine, Tomoxetine and Nisoxetine [24], [25], [26], which are among the most important pharmaceuticals for the treatment of psychiatric disorders (depression, anxiety, alcoholism) and also metabolic problems (obesity and bulimia).
Section snippets
Results and discussion
Immobilization of enzymes at superparamagnetic nanoparticles involves specific molecular interactions at the surface, and can induce some structural reorganization, thus changing the behavior of the catalytic sites. Therefore, a comparative study was carried out in this work, by immobilizing the lipase according to three different procedures. Initially, magnetite (Fe3O4) nanoparticles (MagNP) were prepared by the co-precipitation method, mixing Fe3+ and Fe2+ ions in NaOH solution, under an
General methods
Amano Lipase PS from B. cepacia was purchased from Sigma–Aldrich. Solvents were purified by standard procedures. Some reagents are commercially available and were used without further purification. Thin-layer chromatography (TLC) was performed using pre-coated plates (Aluminum foil, Silica Gel 60 F254 Merck, 0.25 mm). Silica gel (0.035–0.070 mm, Acros) was used for column chromatography. GC analyses were performed in a Shimadzu GC-17A instrument with a FID detector, using hydrogen as a carrier
Acknowledgements
The authors thank FAPESP and CNPq for financial support. The authors acknowledge the support from FAPESP, CNPq, and PETROBRAS, and Professor Pedro K. Kyohara (Institute of Physics, Univ. S. Paulo) for obtaining TEM images of APTS-MagNP.
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