Large Volume Direct Injection Ultra-High Performance Liquid Chromatography-Tandem Mass Spectrometry-Based Comparative Pharmacokinetic Study between Single and Combinatory Uses of Carthamus tinctorius Extract and Notoginseng Total Saponins

Abstract

The combination of Carthamus tinctorius extract (CTE) and notoginseng total saponins (NGTS), namely, CNP, presents a synergistic effect on myocardial ischemia protection. Herein, comparative pharmacokinetic studies between CNP and CTE/NGTS were conducted to clarify their synergistic mechanisms. A large volume direct injection ultra-high performance liquid chromatography-tandem mass spectrometry (LVDI-UHPLC-MS/MS) platform was developed for sensitively assaying the multi-component pharmacokinetic and in vitro cocktail assay of cytochrome p450 (CYP450) before and after compatibility of CTE and NGTS. The pharmacokinetic profiles of six predominantly efficacious components of CNP, including hydroxysafflor yellow A (HSYA); ginsenosides Rg₁ (GRg₁), Re (GRe), Rb₁ (GRb₁), and Rd (GRd); and notoginsenoside R₁ (NGR₁), were obtained, and the results disclosed that CNP could increase the exposure levels of HSYA, GRg₁, GRe, GRb₁, and NGR₁ at varying degrees. The in vitro cocktail assay demonstrated that CNP exhibited more potent inhibition on CYP1A2 than CTE and NGTS, and GRg₁, GRb₁, GRd, quercetin, kaempferol, and 6-hydroxykaempferol were found to be the major inhibitory compounds. The developed pharmacokinetic interaction-based strategy provides a viable orientation for the compatibility investigation of herb medicines.

Keywords: Carthamus tinctorius extract; comparative pharmacokinetic study; compatibility mechanism; large volume direct injection; notoginseng total saponins.

Figure 1

The chemical structures of the main components contained in the combination of Carthamus tinctorius extract and notoginseng total saponins (CNP).

Figure 2

Connectivity sketch of the six-port/two-channel switching valve controlling the large volume direct injection ultra-high performance liquid chromatography–tandem mass spectrometry (LVDI-UHPLC-MS/MS) system. At the loading phase, the valve was maintained at A-channel. The sample delivered from the auto-sampler was captured onto a pre-guard column. Meanwhile, the pumps delivered the mobile phase at a high flow rate. Then, the valve was automatically switched to the B-channel to trigger the elution phase. The trapped components were flushed from the pre-guard column into an analytical column.

Figure 3

Representative multiple reaction monitoring (MRM) chromatograms of target analytes in rat plasma in a positive mode: (A) blank plasma; (B) blank plasma spiked with six chemical standards and internal standards (IS); (C) plasma sample collected at 2 h following oral administration of extract CNP (Carthamus tinctorius extract (CTE) 50 mg/kg + notoginseng total saponins (NGTS) 60 mg/kg) to rats.

Figure 4

Mean plasma concentration-time profiles of the six analytes in rats after oral administration of CTE, NGTS, and CNP. Each point represents the mean ± SD (n = 6).

Figure 5

Representative MRM chromatograms of all probe metabolites and two ISs in the incubated rat microsomal sample with no substrate cocktails: (A) blank microsomal sample, (B) blank microsomal sample spiked with seven chemical standards and two ISs monitored in a polarity switching mode.