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Rapid analysis of neomycin in cochlear perilymph of guinea pigs using disposable SPE cartridges and high performance liquid chromatography-tandem mass spectrometry
Journal of Chromatography B ( IF 3 ) Pub Date : 2018-06-26 , DOI: 10.1016/j.jchromb.2018.06.055
Mian Zu , Jinan Jiang , Hui Zhao , Shili Zhang , Yan Yan , Shiwei Qiu , Shuolong Yuan , Jiawei Han , Yue Zhang , Weiwei Guo , Shiming Yang

Irreversible hearing loss induced by aminoglycoside in human through local or systemic administration route negatively impacts quality of life. The aim of this work was to develop and validate an analytical method suitable for the detection and quantification of neomycin in cochlear perilymph of guinea pig after local application. The SupelMIP SPE column was used for the pre-treatment of matrix. Chromatographic separation was conducted by a reversed phase ODS column (100 × 2.1 mm, 3 μm) at 40 °C in gradient mode with 0.2‰ (v/v) HFBA in water and 0.2‰ (v/v) HFBA in acetonitrile as mobile phase, at a flow rate of 0.30 mL/min, with retention time of 3.50 and 3.62 min for internal standard tobramycin and analyte neomycin, respectively. The MS was performed with positive ionization mode, with data acquisition in Multi Reaction Monitor (MRM) mode. This method was proved to be specific, accurate (97.1–115% of nominal values) and precise (CV% < 15%). Calibration curves for matrix matched standard of neomycin ranged from 1.25 to 200 μg/mL, with LOD and LLOQ of 0.625 and 1.25 μg/mL in blank matrix. The matrix effect was corrected to (−0.1) - 1.33 by adding internal standard. The relative SPE recovery values were ≥98.9% in low, medium and high QC samples. Neomycin in matrix proved to be stable under room temperature - and −20 °C, or under three freeze-thawing cycles, or under processing as well. Finally, the proposed method was successfully applied to a toxicokinetics study of neomycin in perilymph after round window membrane (RWM) administration, which was in accordance with threshold shift of auditory brainstem response (ABR) test related to hearing loss.



中文翻译:

一次性SPE柱和高效液相色谱-串联质谱法快速分析豚鼠耳蜗外周血中的新霉素

氨基糖苷通过局部或全身给药途径在人体内引起的不可逆性听力损失会对生活质量产生负面影响。这项工作的目的是开发和验证一种适用于豚鼠耳蜗周围淋巴局部应用后新霉素检测和定量的分析方法。SupelMIP SPE色谱柱用于基质的预处理。色谱分离是通过反相ODS柱(100×2.1 mm,3μm)在40°C下以梯度模式进行的,其中水中0.2‰(v / v)HFBA和乙腈中0.2‰(v / v)HFBA作为流动相内标妥布霉素和分析物新霉素的保留时间分别为0.30 mL / min和3.50 min和3.62 min。MS以正电离模式执行,并在多反应监测器(MRM)模式下进行数据采集。事实证明,该方法是特定的,准确的(标称值的97.1–115%)和精确的(CV%<15%)。新霉素基质匹配标准品的校准曲线范围为1.25至200μg/ mL,空白基质中的LOD和LLOQ为0.625和1.25μg/ mL。通过添加内标将基体效应校正为(-0.1)-1.33。在低,中和高质控样品中,相对固相萃取的相对回收率值≥98.9%。基质中的新霉素被证明在室温--20°C,三个冷冻-解冻循环或加工过程中均稳定。最后,该方法已成功应用于圆窗膜(RWM)给药后外周淋巴中新霉素的毒物动力学研究,这符合与听力损失相关的听性脑干反应(ABR)测试的阈值漂移。精确(标称值的97.1–115%)和精确(CV%<15%)。新霉素基质匹配标准品的校准曲线范围为1.25至200μg/ mL,空白基质中的LOD和LLOQ为0.625和1.25μg/ mL。通过添加内标将基体效应校正为(-0.1)-1.33。在低,中和高质控样品中,相对固相萃取的相对回收率值≥98.9%。基质中的新霉素被证明在室温--20°C,三个冷冻-解冻循环或加工过程中均稳定。最后,该方法已成功应用于圆窗膜(RWM)给药后周围淋巴中新霉素的毒物动力学研究,这符合与听力损失相关的听觉脑干反应(ABR)测试的阈值漂移。精确(标称值的97.1–115%)和精确(CV%<15%)。新霉素基质匹配标准品的校准曲线范围为1.25至200μg/ mL,空白基质中的LOD和LLOQ为0.625和1.25μg/ mL。通过添加内标将基体效应校正为(-0.1)-1.33。在低,中和高质控样品中,相对固相萃取的相对回收率≥98.9%。基质中的新霉素被证明在室温--20°C,三个冷冻-解冻循环或加工过程中均稳定。最后,该方法已成功应用于圆窗膜(RWM)给药后周围淋巴中新霉素的毒物动力学研究,这符合与听力损失相关的听觉脑干反应(ABR)测试的阈值漂移。新霉素基质匹配标准品的校准曲线范围为1.25至200μg/ mL,空白基质中的LOD和LLOQ为0.625和1.25μg/ mL。通过添加内标将基体效应校正为(-0.1)-1.33。在低,中和高质控样品中,相对固相萃取的相对回收率值≥98.9%。基质中的新霉素被证明在室温--20°C,三个冷冻-解冻循环或加工过程中均稳定。最后,该方法已成功应用于圆窗膜(RWM)给药后周围淋巴中新霉素的毒物动力学研究,这符合与听力损失相关的听觉脑干反应(ABR)测试的阈值漂移。新霉素基质匹配标准品的校准曲线范围为1.25至200μg/ mL,空白基质中的LOD和LLOQ为0.625和1.25μg/ mL。通过添加内标将基体效应校正为(-0.1)-1.33。在低,中和高质控样品中,相对固相萃取的相对回收率值≥98.9%。基质中的新霉素被证明在室温--20°C,三个冷冻-解冻循环或加工过程中均稳定。最后,该方法已成功应用于圆窗膜(RWM)给药后周围淋巴中新霉素的毒物动力学研究,这符合与听力损失相关的听觉脑干反应(ABR)测试的阈值漂移。通过添加内标将基体效应校正为(-0.1)-1.33。在低,中和高质控样品中,相对固相萃取的相对回收率值≥98.9%。基质中的新霉素被证明在室温-20°C,三个冷冻-融化循环或加工过程中均稳定。最后,该方法已成功应用于圆窗膜(RWM)给药后周围淋巴中新霉素的毒物动力学研究,这符合与听力损失相关的听觉脑干反应(ABR)测试的阈值漂移。通过添加内标将基体效应校正为(-0.1)-1.33。在低,中和高质控样品中,相对固相萃取的相对回收率值≥98.9%。基质中的新霉素被证明在室温--20°C,三个冷冻-解冻循环或加工过程中均稳定。最后,该方法已成功应用于圆窗膜(RWM)给药后周围淋巴中新霉素的毒物动力学研究,这符合与听力损失相关的听觉脑干反应(ABR)测试的阈值漂移。或正在处理中。最后,该方法已成功应用于圆窗膜(RWM)给药后周围淋巴中新霉素的毒物动力学研究,这符合与听力损失相关的听觉脑干反应(ABR)测试的阈值漂移。或正在处理中。最后,该方法已成功应用于圆窗膜(RWM)给药后周围淋巴中新霉素的毒物动力学研究,这符合与听力损失相关的听觉脑干反应(ABR)测试的阈值漂移。

更新日期:2018-06-26
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