衍生化GC法测定重组乙型肝炎疫苗中游离甲醛的含量  

Determination of free formaldehyde in recombinant hepatitis B vaccine by derivatization capillary GC

作　　者：纪宏[1] 马迅[1] 祁进[1] 刘晶[1] 王威[1] 孔爱英[2] 

机构地区：[1]北京市药品检验所,北京100035 [2]总后卫生部药品仪器检验所,北京100166

出　　处：《沈阳药科大学学报》2015年第4期289-293,共5页Journal of Shenyang Pharmaceutical University

摘　　要：目的建立衍生化毛细管气相色谱法测定重组乙型肝炎疫苗[酿酒酵母或中国仓鼠卵巢细胞（Chinese hamster ovary,CHO）]中游离甲醛含量。方法应用2,4-二硝基苯肼（2,4-dinitrophenylhydrazine,2,4-DNPH）衍生重组乙型肝炎疫苗中的游离甲醛后,环己烷萃取衍生物,HP-5毛细管色谱柱分离,电子捕获检测器测定,外标法定量。色谱条件：初始温度为150℃,保持1 min,再以20℃·min-1的速率升温至250℃,保持10 min;检测器温度为350℃,尾吹60 m L·min-1;进样口温度为300℃,分流比为50∶1;载气为N2;进样量为1μL。结果游离甲醛质量浓度在1.01~30.3 mg·L-1内,与峰面积呈良好的线性关系,回归方程为A=3 968.5ρ-284.1（r=0.999 6）。低、中、高3个浓度的平均加样回收率为99.1%,RSD值为2.87%。结论衍生化毛细管气相色谱法可用于重组乙型肝炎疫苗中游离甲醛含量的质量控制。Objective To develop and verify a derivatization capillary gas chromatography method for deter- mination of free formaldehyde in the final product of recombinant hepatitis B vaccine （ Saccharomyces cere- visiae or CHO cell）. Methods The free formaldehyde in recombinant hepatitis B vaccine was derivatized by 2,4-dinitrophenylhydrazine （DNPH）. The derivatives were extracted by cyclohexane, analyzed with HP-5 capillary gas chromatography column and detected with electron capture detector. It＇s quantity was deter- mined with external standard method. Chromatographic conditions were as follows：the initial temperature was set at 150 ℃and kept for 1 min,then elevated to 250 ℃ at 20 ℃.min-1 ,holding for 10 min;the detector temperature was 350 ℃, end blowing at 60 mL. min- ~ ;the inlet temperature was 300 ℃, the split ratio was 50：1 ;N2 was chosen as carrier gas,the injection volume was 1 p,L. Results The free formaldehyde in the vaccine showed a good linear relationship to peak area within a concentration range of 1.01-30. 3 mg·L-1 , the regression equation was ：A = 3968.5p - 284. 1 （ r = 0. 999 6 ）. The average recovery of three concentration level using the established method was 99. 1%, RSD value was 2. 87 % （ n = 9 ）. Conclusions The derivatiza- tion capillary gas chromatography method for determination of free formaldehyde in recombinant hepatitis B vaccine （ saccharomyces cerevisiae or CHO cell） is developed, which is simple, precise, stable, reproducible and accurate, and might be used for determination of free formaldehyde in recombinant hepatitis B vaccine.

关 键 词：重组乙型肝炎疫苗 游离甲醛 毛细管气相色谱法 2 4-二硝基苯肼衍生化 

分 类 号：R917[医药卫生—药物分析学]