Department of Pharmaceutics, National Institute of Pharmaceutical Education and Research (NIPER)-Guwahati, Changsari, 781101, Assam, India.
Department of Pharmaceutical Analysis, NIPER-Guwahati, Changsari, 781101, Assam, India.
Anal Methods. 2022 Feb 24;14(8):834-842. doi: 10.1039/d1ay01919h.
Ultra-performance liquid chromatography electrospray ionization tandem mass spectrometry (UPLC/ESI/MS/MS) for the concomitant quantification of active plant constituents, namely quercetin and piperine, in rat plasma was developed and validated to assess pharmacokinetics after a single oral administration. Liquid-liquid extraction technique with ethyl acetate and -hexane (1 : 1) was used, and fisetin was added as an internal standard (IS). Effective chromatographic separation of quercetin, piperine and IS was executed on a Waters Acquity BEH C column (50.0 mm × 2.1 mm, 1.7 μm) using formic acid both (0.1% w/v) in water (A) and acetonitrile (B) as the mobile phase in gradient mode. For detection purposes, positive electrospray ionization (ESI) mode was used with multiple reaction monitoring (MRM) mode for estimation using [M + H] fragment ions / 303.04 → 152.9 for quercetin, 286.12 → 201.04 for piperine and 287.01 → 136.93 for IS. The method was linear over the calibration range of 0.1-200 ng mL. The lower limit of quantification (LLOQ) of quercetin and piperine was obtained as 0.1 ng mL in rat plasma, along with negligible matrix effect and acceptable stability. Furthermore, the bioanalytical method was successfully implemented to determine the pharmacokinetic profiles of quercetin-and piperine-enriched nanostructured lipid carriers (NLCs) in rat plasma after oral administration. The enhancement in the oral bioavailability of quercetin and piperine was 20.72 and 4.67 fold, respectively, compared to their native pristine dispersions. Future exploration of the concentrations of these active constituents in human plasma and organs is feasible using this sensitive, validated UPLC/ESI/MS/MS method.
超高效液相色谱-电喷雾串联质谱法（UPLC/ESI/MS/MS）用于同时定量大鼠血浆中槲皮素和胡椒碱等植物活性成分，方法学验证后用于单次口服给药后的药代动力学评估。采用乙酸乙酯-正己烷（1:1）进行液液萃取，以非瑟酮作为内标（IS）。在 Waters Acquity BEH C 柱（50.0mm×2.1mm，1.7μm）上，以甲酸（0.1%w/v）分别作为水相（A）和乙腈（B）中的流动相，采用梯度洗脱模式实现了槲皮素、胡椒碱和 IS 的有效色谱分离。检测时，采用正电喷雾电离（ESI）模式，用多反应监测（MRM）模式对[M+H]+碎片离子/303.04→152.9 进行定量分析，用于槲皮素；286.12→201.04 用于胡椒碱；287.01→136.93 用于 IS。该方法在 0.1-200ng/mL 范围内呈线性。槲皮素和胡椒碱的定量下限（LLOQ）在大鼠血浆中分别为 0.1ng/mL，且基质效应可忽略不计，稳定性良好。此外，该方法成功用于测定口服给药后大鼠血浆中槲皮素和胡椒碱富载纳米结构脂质载体（NLCs）的药代动力学特征。与它们的原始分散体相比，槲皮素和胡椒碱的口服生物利用度分别提高了 20.72 倍和 4.67 倍。使用这种灵敏、经验证的 UPLC/ESI/MS/MS 方法，未来有望在人血浆和器官中检测这些活性成分的浓度。
