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Nigericin, one of the main ionophoric polyethers produced by various <i>Streptomyces</i> strains, presents relevant biological activities including antibacterial and recently studied antitumor properties. This work describes the influence of different culture conditions on the production of this metabolite by <i>Streptomyces</i> sp. SF10, isolated from a semi-arid soil sample collected at Ch&#233;lia Mountain, in Khenchela (Northeastern Algeria) and identified as <i>Streptomyces youssoufiensis</i>. The extracts from the strain, cultured in a solid state or submerged fermentation conditions, using several carbon sources at different pH values, in the presence or absence of iron (II) sulfate and in co-culture with other <i>Streptomyces</i> species, were analyzed using a high-performance liquid chromatography (HPLC) system equipped with an evaporative light scattering detector (ELSD). The best culture conditions provided a concentration of nigericin of 0.490 &#177; 0.001 mg/mL in the extract. The HPLC-ELSD method, optimized here for the quantitative detection of nigericin, can find wider applications in the analysis of several other metabolites characterized by a similar polycyclic polyether structure or, more generally, by the lack of significant chromophores in their molecular structure.