Whole‐cell biotransformation of oleanolic acid by free and immobilized cells of Nocardia iowensis: Characterization of new metabolites

Abstract

Whole-cell biotransformation of oleanolic acid by free and immobilized cells of Nocardia iowensis: Characterization of new metabolitesIn this study, Nocardia iowensis was used to transform oleanolic acid (OA) into oleanane derivatives. The first derivative, which was found after 24 h of cultivation, was the known and already described OA methyl ester. After 1 week, two other derivatives (oleanonic acid methyl ester and an unknown metabolite) were identified as new products of a biotransformation by N. iowensis. These oleanane metabolites were characterized by HPLC, HPLC-ESI-MS, and HPLC-1 H NMR spectroscopy. The biotransformation was performed by suspended and immobilized cells (ICs) of N. iowensis. Cells immobilized in alginate beads were used in order to prepare a continuous process. The substrate uptake of free and ICs was similar, whereas the peak area of OA methyl ester of the ICs was only about 10% of the native cells. However, the final product (oleanonic acid methyl ester) concentrations were similar in both approaches, whereas the unknown metabolite 3 was only detected transiently in the medium of ICs. Based on these results, a new biosynthetic pathway for the biotechnological production of oleanonic acid methyl ester is proposed.

Biotransformation of Oleanane and Ursane Triterpenic Acids

“…The use of resting or immobilized cells of N . iowensis DSM 45197 as biocatalysts of the OA (approximately 0.3 g/L) transformation process for 13 days resulted in the formation of methyl OA ( 91 ) as the main bioconversion product (more than 60.0%), small amounts (≤5.0%) of methyl 3-oxo-olean-12-en-28-oat ( 92 ), and metabolite 93 unidentified by the authors [ 16 ]. 3-oxo-OA ( 92 ) was shown to have pronounced antimelanoma [ 124 ], antileishmanial, and antitrypanosomal effects [ 125 ].…”

“…3-oxo-OA ( 92 ) was shown to have pronounced antimelanoma [ 124 ], antileishmanial, and antitrypanosomal effects [ 125 ]. Despite numerous successful examples to increase the efficiency of the biotransformation process by immobilizing microbial cells [ 126 ], the use of fixed Nocardia cells in alginate carriers led to a decrease in their catalytic activity, as confirmed by a 10-fold decrease in the formation of compound 91 and only a short-term occurrence of compound 93 in the culture medium [ 16 ]. The ability of Nocardia sp.…”

“…Furthermore, microbial conversion ensures specific modifications of triterpenic molecule sites that are either not modified or poorly modified by synthetic transformations [ 12 ]. Note that, among the known microbial biocatalysts, members of mycelial fungi are the most studied [ 13 , 14 , 15 ] whereas bacterial catalysts are only represented by a few gram-positive species [ 16 , 17 , 18 , 19 , 20 ]. The first papers on microbial transformation of triterpenoids were published in the 1960s [ 21 ].…”

Microbial Transformations of Plant Secondary Metabolites

“…Microbial transformations of plant metabolites mostly involve the use of free whole cells [3,19,20]. Still, there are some examples of the use of immobilized cells, such as entrapment in calcium-alginate beads of Nocardia iowensis for the conversion of oleanolic acid into oleanane derivatives, aiming at continuous operation [29] and Pseudomonas aeruginosa resting cells for hydroxylation of tyrosol [30]. Alginate was the hydrogel used for the immobilization of Rhodotorula minuta for the biotransformation of (L)-citronellal to (L)-citronellol [31].…”

“…The oleanolic acid was transformed by Nocardia iowensis into oleanolic and oleanonic acid methyl esters, regarded as potential candidates for novel pharmaceuticals due to their remarkable activities against herpes simplex virus and human immunodeficiency virus reverse transcriptase [29].…”