Methods for quantitative determination of microalgal lipid and fatty acids content

Microalgae represent a promising feedstock for sustainable biofuel production and high-value lipid-based bioproducts due to their high lipid productivity and rapid growth rates. Accurate and reproducible lipid quantification is essential for strain selection, process optimization, and industrial scaling. This review presents a comprehensive and critical evaluation of contemporary lipid quantification methods applied to microalgae. The methodologies are categorized into screening, quantitative, and profiling approaches, encompassing techniques such as solvent extraction, in situ and direct transesterification, colorimetric assays, spectroscopic tools (NIR, FTIR), and chromatographic techniques (GC, LC–MS/MS). Each method is evaluated across multiple performance axes, including analytical accuracy, throughput, requirement to the sample, technical complexity, and standardization potential. Results are synthesized using the comparative tables. While high-throughput screening tools (e. g., Nile Red, SPV) offer speed and easiness of using, they exhibit limitations in accuracy and reproducibility. Quantitative methods such as acid-catalyzed in situ transesterification coupled with gas chromatography demonstrate a strong balance between precision and scalability. Profiling methods, including LC–MS/MS, provide the highest molecular resolution but are cost- and labor-intensive. The review highlights the need for methodological harmonization and discusses the trade-offs associated with analytical choices in research and industry. Practical recommendations are proposed for selecting the appropriate techniques depending on application context — from early-stage screening to advanced lipidomic profiling.