Frequency-dependent modulation of functional single cell oils in Rhodotorula sp. and Aspergillus flavus under alternating current stimulation

Microbial lipid metabolism can be modulated by non-invasive physical stimuli, offering a sustainable route to tailor functional biomaterials. Here, we report for the first time the frequency-dependent modulation of single cell oils (SCOs) in the oleaginous yeast Rhodotorula sp. and the filamentous fungus Aspergillus flavus through alternating current (AC) electrostimulation during exponential and stationary growth phases. Cultures were exposed to AC frequencies ranging from 100 Hz to 1 MHz under nitrogen-limited conditions, and lipid yield, fatty acids (FAs) composition, and ultrastructural changes were assessed via gravimetric analysis, GC-MS, FTIR spectroscopy, and TEM imaging. Low-frequency stimulation (100 Hz-1 kHz) significantly enhanced lipid accumulation (up to 2.7-fold above controls) with enrichment in oleic acid, a monounsaturated fatty acid (MUFA). Mid-frequency exposure (10-100 kHz) favored MUFAs accumulation at early stages but later suppressed unsaturation, implying transient desaturase modulation. High-frequency treatment (1 MHz) selectively enriched polyunsaturated fatty acids (PUFAs), particularly linoleic acid (ω-6), during prolonged cultivation, linking oxidative stress responses to lipid remodeling. TEM revealed frequency-specific ultrastructural adaptations, including lipid droplet enlargement, membrane invaginations, and autophagic activity. Together, these results demonstrate that AC frequency acts as a tunable "metabolic switch" capable of enhancing both the quantity and quality of microbial lipids. This strategy offers strong potential for scalable production of biofunctional lipids relevant to biofuels, nutraceuticals, biomedical, and advanced biomaterial applications, which aligns with various United Nations Sustainable Development Goals (SDGs) including zero hunger (SDG 2), good health and well-being (SDG 3), and affordable and clean energy (SDG 7).