Radix Paeoniae Rubra and Radix Paeoniae Alba Attenuate CCl4-induced acute liver injury: an ultra-performance liquid chromatography-mass spectrometry (UPLC-MS) based metabolomic approach for the pharmacodynamic study of Traditional Chinese Medicines (TCMs)

Abstract

Metabolomics has been frequently used in pharmacodynamic studies, especially those on traditional Chinese medicine (TCM). Radix Paeoniae Alba and Radix Paeoniae Rubra are popularly used in TCM, and both have hepatoprotective effects. In this study, a CCl(4)-induced acute liver injury rat model was established and confirmed by the observed serum aminotransferase activities. The metabolomics approach was applied to study the influence of Radix Paeoniae Alba and Radix Paeoniae Rubra on the metabolic changes in rats with acute liver injury. The partial least-squares-discriminant analysis (PLS-DA) of rat serum and their ultra-performance liquid chromatography-mass spectrometry (UPLC-MS) fingerprints allowed discrimination of controlled, acute liver injury-model rats after administration of the two types of TCMs. The time-dependent PLS-DA plots showed that the changes in the metabolic patterns of the rats, which were administered with the TCMs, had stabilized within 2 h after they received the intraperitoneal CCl(4) injection. The results indicated the protective effect of TCMs against liver injury. Several potential biomarkers were detected and identified, which included creatine, deoxycholic acid, choline, 5-methylenetetrahydrofolate, folic acid, and glycocholic acid. The physiological significance of these metabolic changes was discussed.

Figure A1

UPLC-MS total ion chromatogram of the serum samples from (A) the CCl₄-induced acute liver injury rat models (CCl₄); (B) Radix Paeoniae Rubra (RPR)-treated group; (C) Radix Paeoniae Alba (RPA)-treated group, and (D) the control (VEH) group after 24 h of CCl₄ or olive oil injection.

Figure A2

UPLC-MS-selected ion recording chromatogram of the blank serum spiked with a mixture of 500 ng/mL bile acid standards (1, TUDCA; 2, THDCA; 3, GUDCA; 4, TCA; 5, GCA; 6, TCDCA; 7, TDCA; 8, UDCA; 9, HDCA; 10, GCDCA; 11, GDCA; 12, CA; 13, TLCA; 14, GLCA; 15, CDCA; 16, DCA; 17, LCA).

Figure S3

Heat map of bile acids in serum from rats of the CCl₄, RPR, RPA, and VEH groups collected at 24 h after intraperitoneal injection of CCl₄. (The concentrations of the bile acids are represented by colors ranging from light green to dark red. LCA, GCA, TCA, TCDCA, TDCA, and TLCA were significantly altered among groups and showed the recovery potential of the TCM-treated groups).

Figure 1

Partial least squares-discriminant analysis (PLS-DA) scores plot of the olive oil-treated controls (VEH) and the CCl₄-induced liver injury rat model (CCl₄) group at 24 h after the intraperitoneal injection of CCl₄, which was generated by: (A) metabolomic fingerprinting analysis data and (B) bile acid profiling analysis data (▲ VEH group, CCl₄ group).

Figure 2

Trajectory analysis of (A) the CCl₄-induced liver injury rat model group; (B) Radix Paeoniae Rubra (RPR)-treated group, and (C) Radix Paeoniae Alba (RPA)-treated group (▲ 1 h before injection, 0.5 h after injection, 1 h after injection, 2 h after injection, 4 h after injection, and 6 h after injection).

Figure 2

Trajectory analysis of (A) the CCl₄-induced liver injury rat model group; (B) Radix Paeoniae Rubra (RPR)-treated group, and (C) Radix Paeoniae Alba (RPA)-treated group (▲ 1 h before injection, 0.5 h after injection, 1 h after injection, 2 h after injection, 4 h after injection, and 6 h after injection).

Figure 3

PLS-DA scores plot of rat serum samples from all the four groups, which were collected at 24 h after the intraperitoneal injection of CCl₄, as generated from the bile acid profiling analysis data (▲ VEH group, RPA group, RPR group, CCl₄ group).