Use of Experimental Design for the Optimization of the Production of New Secondary Metabolites by Two Penicillium Species

Abstract

A fractional factorial design approach has been used to enhance secondary metabolite production by two Penicillium strains. The method was initially used to improve the production of bioactive extracts as a whole and subsequently to optimize the production of particular bioactive metabolites. Enhancements of over 500% in secondary metabolite production were observed for both P. oxalicum and P. citrinum. Two new alkaloids, citrinalins A (5) and B (6), were isolated and identified from P. citrinum cultures optimized for production of minor metabolites.

Evaluating methods for the isolation of marine-derived fungal strains and production of bioactive secondary metabolites

“…In the case of isolation and growth of marinederived fungal strains, the effect of seawater concentration in the media is also significantly relevant, as showed by Bugni & Ireland (2004), Yang et al (2007) as well as by our previous investigations (Vita-Marques et al, 2008;Pimenta et al, 2010). In general, high sea water concentration inhibits rather than promotes the growth and secondary metabolites production.…”

“…A number of new compounds have been previously reported by us (Hernandez et al, 2000;Hernandez et al, 2004;Lira et al, 2006;Pimenta et al, 2010). We have subsequently developed a multi-screening approach in order to obtain and select bioactive extracts from marine-derived fungi culture media (Vita-Marques et al, 2008).…”

Biosynthesis of two dihydropyrrole-polyketides from a marine-derived Penicillium citrinum

“…13 C-Labeled precursors were added at the fourth day of growth, at a final concentration of 1 mg mL -1 for each precursor, using previously optimized growth conditions. 15 Separate growth experiments were performed without the addition of labeled precursors as control. Cultures were harvested after 14 days for each growth experiment using distinct 13 C-labeled precursors.…”

“…On the other hand, feeding with [U- 13 C 5 ]ornithine showed marked incorporation at the dihydropyrrole group, for which 13 C-13 C coupling constants of the dihydropyrrole moiety could be unambiguously measured: 74 Hz for C-2/C-3, 36 for C-3/C-4, 36 for C-4/C-5 and 7 for C-2/C-5 (Figures S14 to S16). Since compound 2 has been isolated as a minor derivative, 15 some of the feeding experiments gave insufficient amounts of labeled 2 to obtain reliable 13 C NMR spectra to measure the incorporation rates of [1,2-13 C 2 ]acetate and of [U- 13 C 5 ]ornithine. The results obtained evidenced that both 1 and 2 are derived from acetate, ornithine and methionine.…”

“…14 Compounds 1 and 2 display insecticidal, fungicidal activities and induce precocious metamorphosis in the milkweed bug Oncopeltus fasciatus (Insecta). 12 We have recently isolated the same compounds 1 and 2 from a marine-derived P. citrinum, 15 and became interested to establish the actual biosynthetic pathway leading to 1 and 2. This is because while pyrrole, dihydropyrrole or pyrrolidine alkaloids isolated from non-fungal sources are typically derived from proline, 16,17 ornithine, 1 or even from acetate, 16 the biosynthesis of the above mentioned dihydropyrroles isolated from fungi have not yet been investigated.…”