摘要
目的建立新型乙酰丙酮分光光度法,并用于测定疫苗中游离甲醛残留量。方法结合反应原理,通过正交试验分析乙酰丙酮显色液的配制因素对测定结果的影响并优化配方,通过全波长扫描确定乙酰丙酮分光光度法的测定波长,并对方法的线性和范围、准确性、精密度、检出限、耐用性和样品适用性进行了验证。结果发现乙酸铵浓度为乙酰丙酮试液的关键影响因素,确定了最优配方——乙酰丙酮显色液含乙酸铵1.0摩尔/升(mol/L),乙酸0.24%(V/V),乙酰丙酮1.0%(V/V);确定以412纳米(nm)作为检测波长。新型乙酰丙酮分光光度法在1微克/毫升(g/ml)～40g/ml的浓度范围内具有良好的线性(r2>0.9995),精密度良好,相对标准差均<5%,加标回收率均在95.0%～105.0%,检出限为0.11g/ml;样品60℃水浴5～10min,结果无显著变化;乙酰丙酮显色液可于室温保存10d内使用;含铝佐剂样品经4000g(重力加速度)离心5min,可有效去除对比色结果的干扰。结论该方法简便灵敏,结果准确可靠,可广泛适用于疫苗中游离甲醛残留量的测定。
Objective To develop a new calorimetric method with acetylacetone for the determination of residual free formaldehyde in vaccine. Methods The influence of the preparation of acetylacetone reagent on test results were analysised and optimized through orthogonal test. Detection wavelength of new method was determined by full wavelength scan. The linearity, range, precision, accuracy, detection limit, robustness and sample suitability of test method were validated. Results Ammonium acetate concentration was confirmed to be the key factor of acetylacetone reagent. Preparation of acetylacetone reagent was optimized. The reagent solution contains 1.0M of ammonium ace tare, 0.24% （V/V） of acetic acid, 1.0% （V/V） of acetyl acetone.The suitable wavelength of ultraviolet detector was confirmed to be 412nm. New method has a good linearity in the concentration range of 1 μg . m1-1 ～40μg . ml-1 （fl〉0.9995）, and a good precision （relative standard deviation not greater than 5% ）. The recovery rates of spiked samples are between 95.0% ～ 105.0%, the detection limit is 0.11μg . ml-1. The time of water-bath （60 ℃ ）did not have much effect on the detection results. The acetylacetone reagent is stable for 10 days storage at room temperature. Centrifuging at 4000g for 5min, color development reaction can both remove the adjuvants effectively. Conclusion New method is simple and sensitive with accurate and reliable results, which is suitable for the determination of residual free formaldehyde in vaccine.
出处
《中国疫苗和免疫》
CAS
2012年第2期166-171,共6页
Chinese Journal of Vaccines and Immunization
关键词
乙酰丙酮
新型分光光度法
游离甲醛
疫苗
Acctylacctone
New colorimetric method
Free formaldehyde
Vaccine
