Rapid GC-FID-Based Profiling of Fatty Acids from Plant Leaf Tissue via Trans Methylation

Fatty acids (FAs) are key components of plant metabolism with significant implications for nutrition, bio fortification, and industrial applications. Accurate profiling of FA composition is therefore essential, particularly for high-throughput screening of plant tissues. However, conventional lipid extraction and analysis methods are often labor-intensive and require large amounts of biomass, limiting their applicability for small tissue samples. Here, we describe a rapid, sensitive, and reproducible protocol for fatty acid profiling from minimal quantities of plant leaf tissue. The workflow involves efficient total lipid extraction using a chloroform–methanol system, followed by trans-methylation to generate fatty acid methyl esters (FAMEs), and subsequent identification and quantification using gas chromatography–flame ionization detection (GC-FID). The method is optimized for micro-scale samples (50–100 mg fresh weight) and enables reliable detection of major fatty acid species, particularly C18 fatty acids, without the need for internal standards. This protocol provides a streamlined approach for fatty acid analysis, offering improved sensitivity, reduced sample requirement, and compatibility with routine laboratory workflows. It is well-suited for applications in plant phenotyping, metabolic studies, and crop improvement programs requiring rapid and efficient lipid profiling.