Contents lists available at ScienceDirect Biocatalysis and Agricultural Biotechnology journal homepage: www.elsevier.com/locate/bab Biocatalysts screening of Papaver bracteatum flora for thebaine transformation to codeine and morphine Nazanin Ataee a , Jamshid Fooladi b, ⁎ , Mohammad Hasan Namaei c , Hassan Rezadoost d , Fateme Mirzajani e a Department of Microbiology, The Faculty of Biological Science, Alzahra University, Vanak, Tehran, Iran b Department of Biotechnology, The Faculty of Biological Science, Alzahra University, Vanak, Tehran, Iran c Birjand Hepatitis Research Center, Birjand University of Medical Sciences, Ghafary Ave., Birjand, Iran d Department of Phytochemistry, Medicinal Plants and Drugs Research Institute, Shahid Beheshti University, G.C., Tehran, Iran e Department of Nanobiotechnology, Protein Research Center (PRC), Shahid Beheshti University, G.C., Tehran, Iran ARTICLE INFO Keywords: Biotransformation Thebaine HPTLC LC/ESI-MS Papaver bracteatum ABSTRACT The aim of this study was to find biocatalyst which uses thebaine and extract of different parts of Papaver bracteatum to synthesize morphine alkaloids. The thebaine-resistant strains were obtained from microbial flora of different parts of P.bracteatum. They were purified and treated utilizing different concentrations of thebaine. Those that can grow at a concentration of over 500 μg/ml were chosen for the biotransformation experiments. Biotransformation experiments were carried out utilizing selected cells in the medium containing thebaine and/ or the extract of P.bracteatum; the products of such biotransformation were extracted and the profiles of metabolites were evaluated using HPTLC, and LC/ESI-MS methods. Thereafter, the effective isolate for thebaine transformation was characterized by physiological, biochemical and biomolecular methods. The results show that among 67 isolates, 12 strains were selected using the HPTLC screening as candidates that can transform thebaine into codeine and morphine. Among them, 5 strains were identified to transform plant extract, among which, using LC/ESI-MS, a candidate was selected and identified as Bacillus sp. FAR. It can be concluded from this study, that this microbial flora candidate can transform thebaine into important narcotic drugs and it will be a valuable step in biotechnology. 1. Introduction Morphine alkaloids are important class of pharmaceutical sub- stances because of their powerful analgesic (Bruce et al., 1990; Lister et al., 1999; Niknam et al., 2010), antitussive and narcotic antagonist characteristics (Kyslíková et al. 2013). Papaver somniferum (opium poppy), a traditional source of morphine alkaloids (Kyslíková et al. 2013; Nyman, 1978), with secondary metabolites accumulate at low level in plant as a sole commercial resource (Nakagawa et al., 2011; Nyman, 1978). Syntheses of these compounds were difficult and time consuming due to their complexity and strict regulation of biosynthesis morphine alkaloids pathways (Nakagawa et al., 2011). Finding an appropriate way to produce the natural pharmacological compounds is preferable because it results in more purified and qualified substances. (Bruce et al., 1995; Nakagawa et al., 2011; Rathbone and Bruce, 2002; Rinner and Hudlicky, 2012). Notwithstanding the efforts of chemists for the chemical synthesis of morphine alkaloids, these compounds are still produced by isolation from the opium poppy (Boonstra et al., 2001). An alternative approach is biotransformation; via the use of enzyme or whole cell biocatalysts, this approach has benefits over the conventional chemical processes (Bruce et al., 1995), including non- extreme pH and temperature, low levels of toxic waste products (Rathbone and Bruce, 2002), high chemo, regio and enantiosis selectivity under ecologically compatible conditions (Brunati et al., 2004). Transformation of morphine alkaloids utilizing plant cells and tissue cultures has been reported (Corchete and Yeoman, 1989; Furuya et al., 1984; Tam et al., 1982; Wilhelm and Zenk, 1997). Unfortunately, the amounts of desired metabolites produced by plant in-vitro cultures are usually lower than the content in intact plant (Muffler et al., 2011; Rao et al., 1999). In addition to plant, micro- organisms have ultimate potential to operate selective biochemical transformations (Rathbone and Bruce, 2002). There are many reports on the production of morphine alkaloids from fungi and bacteria including the genus Trametes (Boonstra et al., 2001; Bruce et al., 1990; Hailes and Bruce, 1993; Kunz et al., 1985; Kyslíková et al., 2013; Long http://dx.doi.org/10.1016/j.bcab.2016.12.006 Received 21 September 2016; Received in revised form 29 November 2016; Accepted 5 December 2016 ⁎ Corresponding author. E-mail address: jfooladi@alzahra.ac.ir (J. Fooladi). Biocatalysis and Agricultural Biotechnology 9 (2017) 127–133 Available online 09 December 2016 1878-8181/ © 2016 Published by Elsevier Ltd. MARK