Brown S A, Newkirk D R, Hunter R P, Smith G G, Sugimoto K
Texas A&M University, Department of Veterinary Physiology and Pharmacology, College Station 77843-4466.
J Assoc Off Anal Chem. 1990 May-Jun;73(3):479-83.
Sodium hydroxide digestion of unhomogenized kidney and skeletal muscle for 20 min at 70 degrees C was a superior method for extracting gentamicin from tissues, compared with simple homogenization, trichloroacetic acid precipitation of homogenized tissue, and sodium hydroxide digestion of homogenized tissue. Fluorescence polarization immunoassay was used to quantitate gentamicin. Sodium hydroxide digestion of unhomogenized tissue allowed for the recovery of 90.0 +/- 5.9% (means +/- SD) from renal cortex and 79.9 +/- 3.5% from skeletal muscle. The limit of sensitivity was 17.4 ng/g kidney tissue, 15.8 ng/g digested muscle, and 39.0 ng/g digested heart. The within-assay coefficient of variation (CV) at 100 ng/g kidney was 9.2%; at 500 ng/g kidney, the CV was 2.5%; and at 2000 ng/g kidney, the CV was 1.5%. The between-assay coefficient of variation was less than 7.5% for all concentrations from kidney, and the 99% confidence interval at 100 ng/g kidney was 71.7-112.4 ng gentamicin/g kidney. The within-assay coefficient of variation (CV) at 100 ng/g muscle was 15%; at 500 ng/g muscle, the CV was 2.6%; and at 2000 ng/g muscle, the CV was 2.3%. The between-assay coefficient of variation was less than 15% for all concentrations from muscle, and the 99% confidence interval at 100 ng/g muscle was 72.5-136.8 ng gentamicin/g muscle. Gentamicin-free milk could be distinguished from milk containing gentamicin concentrations of 10 ng/mL milk with 95% confidence, and from milk containing concentrations of 30 ng gentamicin/mL milk with 99% confidence. Quantitative results at or below the tolerance level can be obtained within 90 min of sample acquisition using these extraction and assay methods.
与简单匀浆、匀浆组织的三氯乙酸沉淀以及匀浆组织的氢氧化钠消化相比，在70摄氏度下对未匀浆的肾脏和骨骼肌进行20分钟的氢氧化钠消化是从组织中提取庆大霉素的更好方法。采用荧光偏振免疫分析法对庆大霉素进行定量。未匀浆组织的氢氧化钠消化法从肾皮质的回收率为90.0±5.9%（均值±标准差），从骨骼肌的回收率为79.9±3.5%。灵敏度极限为肾组织17.4纳克/克、消化后的肌肉15.8纳克/克、消化后的心脏39.0纳克/克。在肾组织浓度为100纳克/克时，测定内变异系数（CV）为9.2%；在肾组织浓度为500纳克/克时，CV为2.5%；在肾组织浓度为2000纳克/克时，CV为1.5%。对于来自肾脏的所有浓度，测定间变异系数均小于7.5%，在肾组织浓度为100纳克/克时，99%置信区间为71.7 - 112.4纳克庆大霉素/克肾脏。在肌肉浓度为100纳克/克时，测定内变异系数（CV）为15%；在肌肉浓度为500纳克/克时，CV为2.6%；在肌肉浓度为2000纳克/克时，CV为2.3%。对于来自肌肉的所有浓度，测定间变异系数均小于15%，在肌肉浓度为100纳克/克时，99%置信区间为72.5 - 136.8纳克庆大霉素/克肌肉。不含庆大霉素的牛奶与含庆大霉素浓度为10纳克/毫升牛奶的区分置信度为95%，与含庆大霉素浓度为30纳克/毫升牛奶的区分置信度为99%。使用这些提取和测定方法，在采集样品后90分钟内即可获得等于或低于耐受水平的定量结果。
