E003 2 1

Total cholesterol (TC), triglyceride (TG), and total bile acid (TBA) contents in the plasma or liver tissues were all measured using assay kits (A111-1-1, A110-1-1, and E003-2-1; Nanjing Jiancheng Bioengineering Institute, China), according to the manufacturer’s instructions. Protein concentrations in tissue samples were determined using the BCA protein quantitative assay kit (Beyotime Biotechnology, Shanghai, China). TC and TG levels were expressed as mmol/g protein.

The levels of total triglyceride (TG), total cholesterol (TC), high-density-lipoprotein cholesterol (HDLC), low-density-lipoprotein cholesterol (LDLC), and total bile acid (TBA) in serum were measured by analysis kits (A110-2, A111-2, A112-1, A113-1, and E003-2-1, Jiancheng Institute of Biotechnology, Nanjing, China). Non-esterified fatty acid (NEFA) was measured by an automatic biochemical analyzer (Olympus Au5400). The concentrations of very-low-density-lipoprotein-cholesterol (VLDL-C) and lysophosphatidylcholine (LPC) were measured by enzyme-linked immunosorbent assay kits (H249, Jiancheng, Nanjing and CEK621Ge, Cloud-clone corp, Wuhan, China, respectively) according to the instruction manual.

Mice were anaesthetized by chloral hydrate, and fresh mouse serum was obtained by blood sampling from the eyes. Then, the mice were killed by exsanguination. The total bile acid concentrations in the serum was measured using an enzymatic assay kit (E003-2-1, Nanjing Jiancheng Biological Engineering Institute, Nanjing, China). The enzymatic cycling method was used to analyse the concentrations of serum total bile acid [57 (link)]. An enzymatic cycling reaction system was constructed for both the standard and tested samples. After incubation at 37 °C for 1 min, the absorbance at 405 nm (A0) was read, and then the absorbance at 405 nm (A1) was read after incubation at 37 °C for 3 min. The absorbance changes (A1 − A0) in the standard and tested samples were calculated. The total bile acid concentrations of the tested sample was obtained by the following equation: [(A1tested sample − A0tested sample)/(A1standard sample − A0standard sample)] × standard sample concentrations.