A sensitive and specific LC–MS/MS method for determination of a novel antihypertensive peptide FR-6 in rat plasma and pharmacokinetic study
Yu Yang,
Xingyan Bao,
Jiangyue Ning,
Ruiyan Huang,
Yuan Liang,
Zelong Yan,
Haotian Chen,
Li Ding,
Chang Shu
Affiliations
Yu Yang
Key Laboratory of Drug Quality Control and Pharmacovigilance (China Pharmaceutical University), Ministry of Education, Nanjing, 210009, China; Department of Pharmaceutical Analysis, School of Pharmacy, China Pharmaceutical University, Nanjing, 211198, China
Xingyan Bao
Key Laboratory of Drug Quality Control and Pharmacovigilance (China Pharmaceutical University), Ministry of Education, Nanjing, 210009, China; Department of Pharmaceutical Analysis, School of Pharmacy, China Pharmaceutical University, Nanjing, 211198, China
Jiangyue Ning
Key Laboratory of Drug Quality Control and Pharmacovigilance (China Pharmaceutical University), Ministry of Education, Nanjing, 210009, China; Department of Pharmaceutical Analysis, School of Pharmacy, China Pharmaceutical University, Nanjing, 211198, China
Ruiyan Huang
Key Laboratory of Drug Quality Control and Pharmacovigilance (China Pharmaceutical University), Ministry of Education, Nanjing, 210009, China; Department of Pharmaceutical Analysis, School of Pharmacy, China Pharmaceutical University, Nanjing, 211198, China
Yuan Liang
Key Laboratory of Drug Quality Control and Pharmacovigilance (China Pharmaceutical University), Ministry of Education, Nanjing, 210009, China; Department of Pharmaceutical Analysis, School of Pharmacy, China Pharmaceutical University, Nanjing, 211198, China
Zelong Yan
Key Laboratory of Drug Quality Control and Pharmacovigilance (China Pharmaceutical University), Ministry of Education, Nanjing, 210009, China; Department of Pharmaceutical Analysis, School of Pharmacy, China Pharmaceutical University, Nanjing, 211198, China
Haotian Chen
Key Laboratory of Drug Quality Control and Pharmacovigilance (China Pharmaceutical University), Ministry of Education, Nanjing, 210009, China; Department of Pharmaceutical Analysis, School of Pharmacy, China Pharmaceutical University, Nanjing, 211198, China
Li Ding
Key Laboratory of Drug Quality Control and Pharmacovigilance (China Pharmaceutical University), Ministry of Education, Nanjing, 210009, China; Department of Pharmaceutical Analysis, School of Pharmacy, China Pharmaceutical University, Nanjing, 211198, China
Chang Shu
Key Laboratory of Drug Quality Control and Pharmacovigilance (China Pharmaceutical University), Ministry of Education, Nanjing, 210009, China; Department of Pharmaceutical Analysis, School of Pharmacy, China Pharmaceutical University, Nanjing, 211198, China; Corresponding author. Key Laboratory of Drug Quality Control and Pharmacovigilance (China Pharmaceutical University), Ministry of Education, Nanjing, 210009, China.
The investigation of peptide drugs has become essential in the development of innovative medications for hypertension. In this study, a sensitive high-performance liquid chromatography tandem mass spectrometry (HPLC-MS/MS) method was developed to determine the plasma concentration and stability of the antihypertensive peptide FR-6 in rats. An isotopically labeled peptide (with an unchanged sequence) was utilized as an internal standard (IS) for validation purposes. Subsequently, this assay was employed to examine the pharmacokinetics of different administration methods (tail vein and gavage) in Sprague Dawley (SD) rats. Extracted plasma samples underwent sample preparation through methanol protein precipitation, followed by elution of FR-6 on Wondasil C18 Superb column (4.6 × 150 mm, 5 μm), using a mobile phase consisting of formic acid (0.1%) in water (A) and formic acid (0.125%)-ammonium formate (2 mM) in methanol (B). Ion pairs corresponding to FR-6 and IS were monitored via multiple reaction monitoring (MRM) under positive ion mode: m/z 400.7 → 285.1 for FR-6 and m/z 406.1 → 295.1 for IS detection respectively. The method exhibited excellent linearity with respect to FR-6 concentrations. In addition, the inter-day and intra-day precision were 0.61–6.85% and 1.76–11.75%; the inter-day and intra-day accuracy were −7.28–0.13% and −7.20–2.28%, respectively. In conclusion, the matrix effect, extraction recovery, and stability data were validated according to FDA recommended acceptance criteria for bioanalytical methods. This validated method serves as a reliable tool for determining the concentration of antihypertensive peptide FR-6, and has been successfully applied in pharmacokinetic studies involving rats.
