Comparative Assessment of Derivatization Methods on Quantification of 4-Alkyl Branched-Chain Fatty Acids in Mutton

This study systematically evaluated five derivatization methods for the gas chromatography-mass spectrometry (GC-MS) analysis of three key 4-alkyl branched-chain fatty acids (4-Me-8:0, 4-Et-8:0, and 4-Me-9:0) associated with mutton flavor compounds in mutton adipose tissue. Among the tested methods, sulfuric acid-methanol methylation demonstrated superior performance in terms of sensitivity, recovery, anreproducibility. The method exhibited linearity ( R² > 0.990) across a concentration range of 1–40 µg/mL, with detection limits (LOD) and quantification limits (LOQ) of 1.24–2.18 µg/kg and 4.15–7.28 µg/kg, respectively. Spike recovery rates ranged from 77.73% to 108.48%. The precision (%RSD) and accuracy were maintained below 3.40% and ± 5.45%, respectively.Even after adding a mixture of fatty acids containing isomers, the precision (% RSD) and accuracy remained below 11.08% and ± 10.03%, respectively. This optimized protocol establishes a reliable analytical framework for quantifying flavor-associated 4-alkyl-branched fatty acids, advancing research on mutton flavor chemistry and quality control.