Method-Dependent Variability in Hempseed Lipidomics: Combined Influence of Extraction, Derivatisation, and Genotype on Fatty Acid Composition and Stability

Hempseed (Cannabis sativa L.) is recognised as a valuable source of nutritionally important lipids, particularly polyunsaturated fatty acids (PUFA). Despite growing interest in hempseed oil, the combined influence of extraction and derivatisation on fatty acid profiling remains poorly understood, limiting comparability and standardisation of lipidomic analyses, particularly for fibre-type hemp cultivars. This study evaluated the effects of microwave-assisted and Soxhlet extraction, combined with acidic and basic derivatisation, on fatty acid composition, lipid quality indices, and oxidative stability in Romanian fibre-type hempseed cultivars. The analysed oils were dominated by linoleic and α-linolenic acids, with favourable PUFA/SFA and ω-6/ω-3 ratios, low atherogenic and thrombogenic indices, and high hypocholesterolaemic/hypercholesterolaemic ratios, indicating excellent nutritional quality. Statistical analyses confirmed that extraction and derivatisation significantly influenced fatty acid composition and lipid quality indices, while principal component analysis identified extraction methodology as the main source of variability. Oxidative stability showed moderate correlations with fatty acid composition, suggesting that differences in lipid profiles may influence susceptibility to oxidation. Overall, the study demonstrates that Romanian fibre-type hemp cultivars are valuable sources of nutritionally favourable lipids and emphasises the need for standardised extraction and derivatisation protocols to improve the reliability and comparability of hempseed lipidomic analyses.