Vito Virgina De, Saba Alessandro, Lee Hong-Ki, Owen Helen, Poapolathep Amnart, Giorgi Mario
Department of Veterinary Medicine, University of Sassari, Via Vienna 2, 07100 Sassari, Italy.
Department of Surgical, Medical, Molecular Pathology and Critical Area, University of Pisa, Italy.
J Pharm Biomed Anal. 2016 Jan 25;118:251-258. doi: 10.1016/j.jpba.2015.11.004. Epub 2015 Nov 7.
Grapiprant, a novel pharmacologically active ingredient, acts as a selective EP4 receptor antagonist whose physiological ligand is prostaglandin E2 (PGE2). It is currently under development for use in humans and dogs for the control of pain and inflammation associated with osteoarthritis. The aim of the present study was to develop an easy and sensitive method to quantify grapiprant in canine plasma and to apply the method in a canine patient. Several parameters, both in the extraction and detection method were evaluated. The final mobile phase consisted of ACN:AcONH4 (20 mM) solution, pH 4 (70:30, v/v) at a flow rate of 1 mL/min. The elution of grapiprant and IS (metoclopramide) was carried out in isocratic mode through a Synergi Polar-RP 80A analytical column (150 mm × 4.6 mm). The best excitation and emission wavelengths were 320 and 365 nm, respectively. Grapiprant was extracted from the plasma using CHCl3, which gave a recovery of 88.1 ± 10.22% and a lower limit of quantification (LLOQ) of 10 ng/mL. The method was validated in terms of linearity, limit of detection (LOD), LLOQ, selectivity, accuracy and precision, extraction recovery, stability, and inter-laboratory cross validation, according to international guidelines. The chromatographic runs were specific with no interfering peaks at the retention times of the analyte and IS, as confirmed by HPLC-MS experiments. In conclusion, this was a simple and effective method using HPLC-FL to detect grapiprant in plasma, which may be useful for future pharmacokinetic studies.
格拉匹普兰特是一种新型药理活性成分，作为选择性EP4受体拮抗剂，其生理配体为前列腺素E2（PGE2）。目前正在研发用于人和犬，以控制与骨关节炎相关的疼痛和炎症。本研究的目的是开发一种简便、灵敏的方法来定量犬血浆中的格拉匹普兰特，并将该方法应用于犬类患者。对提取和检测方法中的几个参数进行了评估。最终流动相由乙腈：醋酸铵（20 mM）溶液组成，pH 4（70:30，v/v），流速为1 mL/min。格拉匹普兰特和内标（甲氧氯普胺）通过Synergi Polar-RP 80A分析柱（150 mm×4.6 mm）以等度洗脱模式进行洗脱。最佳激发波长和发射波长分别为320和365 nm。使用氯仿从血浆中提取格拉匹普兰特，回收率为88.1±10.22%，定量下限（LLOQ）为10 ng/mL。根据国际指南，该方法在线性、检测限（LOD）、LLOQ、选择性、准确度和精密度、提取回收率、稳定性以及实验室间交叉验证方面进行了验证。如HPLC-MS实验所证实，色谱运行具有特异性，在分析物和内标的保留时间处没有干扰峰。总之，这是一种使用HPLC-FL检测血浆中格拉匹普兰特的简单有效方法，可能对未来的药代动力学研究有用。
