目的: 建立顶空气相色谱法测定3型肺炎球菌结合物原液的乙腈残留量。方法: 采用Agilent Technologies DB-WAX毛细管色谱柱(30 m×0.53 mm,1.00 μm),载气为N2,色谱柱流量为4.0 mL·min-1,分流比为10∶1,起始柱温为50 ℃,保持5 min,以10 ℃·min-1的速率升温至200 ℃,保持15 min,检测器温度为220 ℃。恒温炉温度为80 ℃、平衡时间为30 min。结果: 系统适用性试验中理论板数不低于5 000,分离度不低于1.5;不高于5 μg·mL-1的CDAP对3型肺炎球菌结合物原液中乙腈残留量检测没有干扰;乙腈的检测限为0.12 μg·mL-1,定量限为0.5 μg·mL-1;3型肺炎球菌结合物原液中乙腈的回收率在108.3%~111.7%;重复性的RSD在0.40%~3.7%,中间精密度的RSD在1.7%~4.6%;线性回归方程为Y=3 039X-493.7,r2=0.999 8,线性范围为0.2~32 μg·mL-1。分别在改变色谱柱流量±1.0 mL·min-1,检测器温度±10 ℃,分流比5∶1的条件下考察3型肺炎球菌结合物原液中乙腈相对误差在-3.8%~3.5%,分离度(>1.5)均符合要求。4种工序中乙腈含量范围分别为2 825~3 164 μg·mL-1、2 737~3 164 μg·mL-1、未检出和0.283 μg·mL-1、未检出。结论: 采用顶空气相色谱法测定3型肺炎球菌结合物原液中乙腈残留量,方法快捷,操作简单,结果可靠。
Objective: To establish a headspace gas chromatography method for the determination of acetonitrile residue in bulk of serotype 3 pneumococcal conjugate. Methods: Agilent Technologies DB-WAX capillary column (30 m×0.53 mm, 1.00 μm) was used with N2 as carrier gas. The flow rate of the column was 4.0 mL·min-1, the split ratio was 10∶ 1, and the initial column temperature was 50 ℃ for 5 min. The temperature was heated to 200 ℃ at the rate of 10 ℃·min-1 and kept at 220 ℃ for 15 min. The temperature of the constant temperature furnace was 80 ℃ and the balance time was 30 min. Results: In the system suitability test, the number of theoretical plates was not less than 5 000 and the separation degree was not less than 1.5. CDAP not higher than 5 μg·mL-1 did not interfere with the determination of acetonitrile residue in bulk of serotype 3 pneumococcal conjugate. The limits of detection and quantification of acetonitrile were 0.12 μg·mL-1 and 0.5 μg·mL-1, respectively. The recoveries of acetonitrile in bulk of serotype 3 pneumococcal conjugate was ranged from 108.2%-111.7%. The RSD of repeatability was 0.40%-3.7%, and RSD of intermediate precision was 1.7%-4.6%. The linear regression equation was Y=3 039X-493.7, r2=0.999 7, the linear range was 0.2-32 μg·mL-1. The recoveries of acetonitrile in bulk of serotype 3 pneumococcal conjugate changing the chromatographic conditions sequentially: column flow 3.0 mL·min-1, column flow 5.0 mL·min-1, detector temperature 210 ℃, detector temperature 230 ℃ and shunt ratio 5∶1. Acetonitrile of measurement result relative error was -3.8%-3.5%, and the separation degree(>1.5) met the requirements. The content ranges of acetonitrile in the four processes were 2 825-3 164 μg·mL-1, 2 737-3 164 μg·mL-1, not detected and 0.283 μg·mL-1, not detected. Conclusion: Headspace gas chromatography is a rapid, simple and reliable method for the determination of acetonitrile residue in bulk of serotype 3 pneumococcal conjugate.
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ChP 2020. Vol Ⅲ[S].2020: 511,684
