目的:建立用气相色谱法测定人类乳头瘤病毒(HPV)疫苗中3-O-脱酰基-4'-单磷酰脂质A(MPL)的含量。方法:HPV疫苗中的MPL经过与氢氧化钠-甲醇的皂化反应,以己烷进行萃取,以三氟醋酐进行衍生化,再经HP-5色谱柱(30 m×0.32 mm,0.25 μm)分离,火焰离子检测器(FID)检测,用MPL对照品和内标十五烷酸(pentadecanoic acid)进行内标法定量测定HPV疫苗中MPL含量。结果:MPL质量浓度在15~150 μg·mL-1范围内线性关系良好(r=0.999),精密度RSD为0.24%,重复性RSD为1.0%,稳定性RSD为0.37%,平均回收率为100.1%~100.5%(RSD小于2.0%)。12个批次的HPV疫苗中MPL含量范围为93.4~103.2 μg·mL-1,符合质控要求。结论:本法准确可靠,特异性强,可对HPV疫苗中MPL进行检测。
Objective: To establish an analytical method for detection the content of 3-O-deacyl-4'-monophosphoryl lipid A(MPL) adjuvant from human papillomavirus(HPV) vaccine with gas chromatography. Methods: After saponified with sodium hydroxide, extracted in hexane and derivated by trifluoroacetic anhydride, the target components were separated by HP-5 column(30 m×0.32 mm, 0.25 μm) and detected using gas chromatography-flame ionization detector(GAS-FID) and quantified by MPL standard material with the help of an internal standard pentadecanoic acid. Results: The linear relationship was good in the range of 15-150 μg·mL-1 with good linearity(r=0.999). The RSDs of precision, reproducibility and stability were 0.24%, 1.0% and 0.37% separately. The average recovery rates were in the ranges of 100.1%-100.5%, with RSD less than 2.0%.The MPL contents of 12 batches vaccines were tested in the range of 93.4-103.2 μg·mL-1, which meets the specification. Conclusion: This accurate and specific method can be applied to the MPL detection in vaccine.
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