Determination of Cholesterol Oxidation Products in Meat Products by Saponification at Room Temperature Combined with Gas Chromatography-Quadrupole-Time-of-Flight Mass Spectrometry

In view of the diverse forms and unstable properties of cholesterol oxidation products (COPs) in meat products under specific conditions, a sample pretreatment technology, saponification at room temperature, was established for the accurate detection of 9 COPs in meat products by gas chromatography-quadrupole-time-of-flight mass spectrometry (GC-Q-TOF/MS). The results showed that the optimal pre-treatment conditions were extraction with chloroform/methanol (2:1, V/V), saponification at room temperature for 18 h, re-extraction with ethyl acetate/n-hexane (80:20, V/V), and clean-up using an aminopropyl solid-phase extraction (SPE) cartridge. The linear ranges were 0.002-0.5 μg/mL for 7α-hydroxyl cholesterol and 7β-hydroxyl cholesterol, and 0.02-0.5 μg/mL for 5β,6β-epoxide cholesterol, 5α,6α-epoxide cholesterol, 20α-hydroxy cholesterol, 3β,5α,6β-trihydroxy cholesterol, 25α-hydroxy cholesterol, 7-keto cholesterol and 27α-hydroxy cholesterol. The determination coefficients were ≥ 0.9990 for all analytes, the limits of detection (LOQ) were 0.0001-0.001 mg/kg, and the limits of quantitation (LOQ) were 0.0002-0.002 mg/kg. The method had good accuracy and high precision. The average recoveries from spiked samples were 89.7%-116.1%, with relative standard deviations (RSDs) of less than 8.7%. Compared with high temperature saponification, saponification at room temperature effectively prevented the autoxidation of cholesterol and the degradation of 7-ketocholesterol, and reduced the exogenous introduction and endogenous loss of target substances, thereby enabling the rapid and accurate detection of the 9 COPs.