Suzuki Yosuke, Tanaka Ryota, Oyama Nobuhiro, Nonoshita Ko, Hashinaga Kazuhiko, Umeki Kenji, Sato Yuhki, Hiramatsu Kazufumi, Kadota Jun-Ichi, Itoh Hiroki
Department of Clinical Pharmacy, Oita University Hospital, Hasama-machi, Oita, Japan.
Department of Clinical Pharmacy, Oita University Hospital, Hasama-machi, Oita, Japan.
Clin Biochem. 2017 Dec;50(18):1228-1236. doi: 10.1016/j.clinbiochem.2017.09.011. Epub 2017 Sep 18.
Protein-free (unbound) drug concentrations have been reported to be better biomarker of pharmacodynamics compared with total drug concentrations. In this study, we developed and validated an assay for the quantification of total and free itraconazole and hydroxyitraconazole, a main metabolite with antifungal activity, in human plasma using ultra-performance liquid chromatography coupled to tandem mass spectrometry (UPLC-MS/MS).
DESIGN & METHODS: Plasma sample was ultra-filtrated for the measurement of free itraconazole and hydroxyitraconazole concentrations. The samples were prepared by solid phase extraction, and then subject to UPLC-MS/MS quantification.
The assay fulfilled the requirements of the US Food and Drug Administration (FDA) and the European Medicines Agency (EMA) guidelines for assay validation, with a lower limit of quantification of 10ng/mL for total itraconazole and hydroxyitraconazole, and 0.1 and 0.5ng/mL for free itraconazole and hydroxyitraconazole, respectively. Recovery rates of total itraconazole and hydroxyitraconazole from whole plasma ranged from 53.3% to 64.0%, and recovery rates of free itraconazole and hydroxyitraconazole from ultrafiltrated plasma ranged from 81.6% to 98.7%. Matrix effect varied between 79.1% and 109.4% for total itraconazole and hydroxyitraconazole, and between 81.3% and 99.7% for free itraconazole and hydroxyitraconazole. The assay was successfully applied to therapeutic drug monitoring of itraconazole in three patients with chronic progressive pulmonary aspergillosis or invasive pulmonary aspergillosis. Plasma free hydroxyitraconazole concentrations were 8.1-, 23.3-, and 51.1-fold higher than plasma free itraconazole concentrations in the three patients.
A method for sensitive and selective quantification of plasma total and free itraconazole and hydroxyitraconazole concentrations was developed using UPLC-MS/MS. Free hydroxyitraconazole concentration may be most important in therapeutic drug monitoring of itraconazole.
据报道，与总药物浓度相比，无蛋白（未结合）药物浓度是更好的药效学生物标志物。在本研究中，我们开发并验证了一种使用超高效液相色谱-串联质谱法（UPLC-MS/MS）定量测定人血浆中总伊曲康唑和羟基伊曲康唑（一种具有抗真菌活性的主要代谢物）以及游离伊曲康唑和羟基伊曲康唑的分析方法。
对血浆样本进行超滤以测量游离伊曲康唑和羟基伊曲康唑浓度。样品通过固相萃取制备，然后进行UPLC-MS/MS定量分析。
该分析方法符合美国食品药品监督管理局（FDA）和欧洲药品管理局（EMA）的分析方法验证指南要求，总伊曲康唑和羟基伊曲康唑的定量下限为10ng/mL，游离伊曲康唑和羟基伊曲康唑的定量下限分别为0.1ng/mL和0.5ng/mL。总伊曲康唑和羟基伊曲康唑从全血中的回收率在53.3%至64.0%之间，游离伊曲康唑和羟基伊曲康唑从超滤血浆中的回收率在81.6%至98.7%之间。总伊曲康唑和羟基伊曲康唑的基质效应在79.1%至109.4%之间，游离伊曲康唑和羟基伊曲康唑的基质效应在81.3%至99.7%之间。该分析方法已成功应用于3例慢性进行性肺曲霉病或侵袭性肺曲霉病患者的伊曲康唑治疗药物监测。这3例患者血浆中游离羟基伊曲康唑浓度分别比游离伊曲康唑浓度高8.1倍、23.3倍和51.1倍。
使用UPLC-MS/MS开发了一种灵敏且选择性地定量血浆中总伊曲康唑和游离伊曲康唑及羟基伊曲康唑浓度的方法。游离羟基伊曲康唑浓度在伊曲康唑治疗药物监测中可能最为重要。
