<![CDATA[Yeast-derived single-cell oils (SCOs) are promising renewable lipid sources for biodiesel and high-value bioproducts. Among oleaginous yeasts, Zygosaccharomyces siamensis AP1, isolated from Indonesian wild bee honey, demonstrates significant potential for lipid production; however, further improvement is needed for industrial-scale applications. This study aimed to enhance lipid accumulation in Z. siamensis AP1 via ultraviolet (UV) mutagenesis, followed by selective screening with cerulenin and ethanol/hydrogen peroxide (H₂O₂) stress. The yeast underwent repeated UV exposure to achieve 60% lethality. The surviving cells were then screened on media with increasing concentrations of cerulenin (5–160 µmol/L) or ethanol/H₂O₂ (2–4% v/v, 1–2 mmol/L). Lipid-producing mutants were characterized for growth kinetics, lipid content, and fatty acid profiles. Among the 43 mutants obtained, the cerulenin-resistant strain CR15 exhibited superior performance, reaching 28.6% lipid content after 96 hours, an increase from 24.6% at 72 hours. At the same, the wild type and the oxidative-stress mutant (MT19) showed reduced lipid accumulation over time. The wild-type lipid content decreased from 22.86% at 72 hours to 18.53% at 96 hours, while lipid content from strain MT19 decreased from 24.64% at 72 hours to 20.09% at 96 hours. Gas chromatography analysis revealed that CR15 produced a diverse fatty acid profile with both even- (arachidonic acid (16%), eicosadienoic acid (5.88%), gondoic acid (18.70%), linoleic acid (26.63%), palmitoleic acid (11.89%) and odd-chain (margaric acid (14.46%)) fatty acids, suggesting mutations in fatty acid synthase (FAS) enzyme function. In contrast, MT19 primarily produced linoleic acid (45.75%) as an adaptive response to oxidative stress. These results indicate that UV mutagenesis coupled with cerulenin selection can effectively enhance lipid biosynthesis and alter fatty acid composition in Z. siamensis. The CR15 mutant represents a promising candidate for advancing sustainable microbial oil production. Further transcriptomic analysis is suggested to clarify the genetic basis of these metabolic changes.]]>
