氨基糖苷类抗生素分析分析分析资料.ppt

* * 氨基糖苷类抗生素分析 药学院药物分析教研室 【大纲要求】 1、掌握氨基糖苷类抗生素代表药物的体内过程 2、熟悉代表药物的体内分析方法 SM GM 主要代表药物 SisM AK ◆ ◆ ◆ 治疗范围窄 6～20μmol/l 毒副作用较强，血药浓度高于30μmol /L 耳毒性和肾毒性 T1/2较长，动物食品残留检测 主要药动学特点 Process in vivo ■ ■ ■ T1/2 与身体健康状况密切相关 Concentration 血药浓度较低、几乎完全从肾脏排泄 Tissue distribution 主要分布于细胞外液、可通过胎盘屏障，不易通过血脑屏障 Characteristic ■ ■ ■ Nitrogen atom Alkali -OH group water soluble polarity Non-chromaphore for UV detection 旋光特性－含有多个氨基糖，一般为右旋 结合的苷键可发生水解 Analytical methods Immune analysis IA UV or fluorescence analysis Chromatography analysis LC/MS analysis High-performance liquid chromatography ◆ On-line separation and detection ◆ Pre/Post-column derivation ◆ HPLC-MS/MS Preparation of the biological sample Derivation Analytical conditions 条件摸索 ◆ ◆ ◆ Protein precipitation PP CH3CN CH3OH HClO4 Liquid-liquid extraction LLE 衍生化后进行 Solid-phase extraction SPE Normal-phase or ion-exchange Solid-phase micro-extraction SPME 衍生化后进行 Column switching Chemical Derivation UV derivation reagent Fluorescence derivation reagent 邻苯二醛 OPA 丹酰氯 DNS-Cl 茚三酮 2,4,6-三硝基苯磺酸 TNBS 1-氟-2,4-二硝基苯 FDNB Optimization suitable derivation conditions Reaction solvent and pH value Methanol or ethanol 含醇量50％为好 pH=8.5~10.5 ■ ■ Concentration of derivation reagent 对OPA ：0.1 μg/ml；其他则无影响 ■ Reaction time and temperature 20、30、60℃→60、40、10 min。OPA-β-MP在30和60℃时可产生干扰的荧光物质，需用CHCl3等提取后在测定 OPA-β-MP RP-HPLC Chromatogram Pre-column derivation Sis M Determination of GM using post-column derivation by RP-HPLC in plasma Analytical conditions: Analytical column: μBondapak C18 Mobile phase : CH3CN-H2O Flow rate:1.5 ml/min Derivation reagent : OPA Flow rate of derivation reagent : 0.8 ml/min Temperature: 50℃ Detection: fluorescence Optimization suitable conditions Protein precipitation methanol 10000 rpm Sample preparation Accuracy and precision Recovery Linear range Limit detection LOQ LOD Evaluated data Derivation reagent Mobile phase Injection value Chromatographic column Reactor Detector Recorder Waste Pump Workstation