Narváez-Rivas Mónica, Pham Alessandra J, Schilling M Wes, León-Camacho Manuel
Food Characterization and Quality Department, Instituto de la Grasa (C.S.I.C.), Avda. Padre García Tejero 4, 41012 Seville, Spain.
Department of Food Science, Nutrition, and Health Promotion, Box 9805, Mississippi State University, MS 39762, United States.
Talanta. 2014 May;122:58-62. doi: 10.1016/j.talanta.2014.01.040. Epub 2014 Jan 31.
A new method for the isolation and analysis of cholesterol oxidation products (COPs) using solid phase extraction (SPE) and silica columns was developed using gas chromatography-flame ion detection (GC-FID). The method comprises of saponification and liquid-liquid extraction of the unsaponifiable fraction prior to the isolation and derivatization of the COPs to trimethylsilyl ethers. The COPs used in this study are cholestane-5α-6α-epoxide, cholestane-3β-5α-6β-triol, 25-hydroxycholesterol and 5-cholesten-3β-ol-7-one. In order to identify the COPs fraction a GC-ion-trap-mass spectrometry experiment were conducted using authentic standards to verify the presence of the COPs. The method was effective at rapidly separating the COPs (25 min run). Calibration curves were linear with the LODs and LOQs bellow 0.03 and 0.07 mgkg(-1) for all cases, respectively. This methodology gave a total recovery for every compound that was used in the study. Betulin was used as an internal standard to monitor the recovery. The method was validated with a standard mixture of COPs. The method has been applied to characterize the COP fraction of subcutaneous fat from Iberian dry-cured ham. Cholestane-5α-6α-epoxide, cholestane-3β-5α-6β-triol, 25-hydroxycholesterol and 5-cholesten-3β-ol-7-one have been identified for the first time in these samples.
开发了一种使用固相萃取（SPE）和硅胶柱分离与分析胆固醇氧化产物（COPs）的新方法，并结合气相色谱 - 火焰离子检测（GC - FID）技术。该方法包括在将COPs衍生化为三甲基硅醚进行分离和衍生化之前，对不皂化物进行皂化和液 - 液萃取。本研究中使用的COPs为胆甾烷 - 5α - 6α - 环氧化物、胆甾烷 - 3β - 5α - 6β - 三醇、25 - 羟基胆固醇和5 - 胆甾烯 - 3β - 醇 - 7 - 酮。为了鉴定COPs组分，使用标准品进行了气相色谱 - 离子阱 - 质谱实验，以验证COPs的存在。该方法能够有效快速分离COPs（运行时间25分钟）。所有情况下校准曲线均呈线性，检测限（LOD）和定量限（LOQ）分别低于0.03和0.07 mgkg⁻¹。该方法对研究中使用的每种化合物都给出了总回收率。使用桦木醇作为内标来监测回收率。该方法通过COPs标准混合物进行了验证。该方法已应用于表征伊比利亚干腌火腿皮下脂肪中的COPs组分。在这些样品中首次鉴定出胆甾烷 - 5α - 6α - 环氧化物、胆甾烷 - 3β - 5α - 6β - 三醇、25 - 羟基胆固醇和5 - 胆甾烯 - 3β - 醇 - 7 - 酮。
