Previous reports had described satisfactory halogenations on the sugar moiety mediated by fluorinase and chlorinase (O’Hagan et al., 2002; Eustaquio et al., 2008). However, these enzymes cannot introduce the halogen into the base moiety. Biosynthesis of purine nucleoside analogues by transglycosylation has been extensively studied (Sinisterra et al., 2010). However, there have been few reports about obtaining pyrimidine nucleosides halogenated on the base moiety using whole cells. In

all cases, conversion rates were < 50% (Pal & Nair, 1997). Microorganism immobilization is a good way to carry out the bioprocess under preparative conditions. Cell entrapment techniques are the most widely used for whole cell immobilization (Trelles et al., 2004). The main advantages of PS-341 this methodology are high operational stability, easy upstream separation, and bioprocess scale-up feasibility. The aim of this study was to obtain 5-halogenated 2′-deoxynucleosides with potential antitumoral activity using a smooth, cheap, and environmentally friendly methodology. We have been able to develop a bioprocess for 5-fluoro and 5-chloro-2′-deoxyuridine HTS assay production using immobilized Aeromonas salmonicida ATCC 27013. Nucleosides and nucleobases were purchased from Sigma Chem. Co. (Brazil). Culture media compounds were obtained from Britania S.A. (Argentine). Chemicals were from Sigma Chem. Co. and Britania S.A. HPLC

grade solvents used were from Sintorgan S.A. (Argentine). Most of the microorganisms were kindly supplied by the ‘Colección Española de Cultivos Tipo (CECT)’, Universidad de Valencia (Spain). Microorganisms were grown until stationary phase in LB medium (5 g L−1 meat extract, 10 g L−1 peptone, and 5 g

L−1 NaCl in deionized water adjusted to pH 7). Cells were harvested by centrifugation for 10 min at 17 500 g, were then washed once with potassium phosphate buffer (30 mM, pH 7), and finally recentrifuged and stored at 4 °C until use. A taxonomic screening with bacterial strains was performed using the following ADP ribosylation factor genera: Aeromonas (10), Bacillus (8), Citrobacter (3), Chromobacterium (1), Enterobacter (6), Escherichia (7), Klebsiella (2), Micrococcus (3), Serratia (4), Proteus (7), and Xanthomonas (4). All microorganisms assayed were non pathogenic for humans. The reaction to select the microorganisms was performed with 1 × 1010 CFU, 10 mM 5-fluorouracil and 2.5 mM thymidine or uridine in 1 mL of potassium phosphate buffer (30 mM, pH 7). Reactions were performed at 30 °C and 200 r.p.m. Samples were taken at 1, 3, 6, and 24 h and centrifuged at 17 500 g during 5 min. Reactions were performed at 30 °C with 1 × 1010 CFU, 2.5 mM 5-fluorouracil and 10 mM thymidine at different phosphate concentration (20–40 mM), pH values (6–8), and shaking speed (100–300 r.p.m.). Reactions were carried out with 1 × 1010 CFU, 2.