Sod kit

Total phenolic content was measured by Folin–Ciocalteu reagent using gallic acid as a standard [7 (link)]. Total flavonoid content was measured by NaNO2-Al(NO3)3-NaOH reagent using rutin as a standard [7 (link)]. The measurement of total acid was performed by acid-base titration method [12 (link)]. The content of VC was measured by colorimetric method [12 (link)]. Measurement of SOD content was performed via SOD kit (SOD kit purchased from Nanjing Jiancheng Institute of Biological Engineering, Jiangsu, China) according to the kit instructions.
HPLC was used to analyze organic acids in RFV based on the literature [25 (link)] with slight modifications. Separation was conducted using an Agilent Z0RBAX SB-AQ column (250 mm × 4.6 mm, 5 µm, American Agilent Corporation (Santa Clara, CA, USA) with an Agilent1260 VWD detector. The relevant parameters of the mobile phase are as follows: monopotassium phosphate buffer solution (0.02 mol/L, pH 2) the ratio of methanol was 95:5, the injection volume was 10 µL and the flow rate was 0.8 mL/min, the spectra were recorded at 210 nm, the column temperature was 35 °C. The experiment was conducted three times for each sample.

Fasting blood samples on 1 day before surgery (baseline) and postoperative day 1 (POD1) were collected. The whole blood samples were then centrifuged at 3000 rpm for 10 min and the obtained serum samples were stored at −80°C. MDA level was measured by spectrophotometric method using thiobarbituric acid (TBA) and MDA kits (Jiancheng Bioengineering, Nanjing, Jiangsu, China). The determination of MDA levels relied on the reaction with TBA to generate the products ‘thiobarbituric acid reactive substances’ (TBARS) that could be measured by the method of fluorimetry (excitation at 532 nm and emission at 553 nm) or colorimetry (532 nm). The level of MDA was measured colorimetrically in this current study. SOD activity was measured by the hydroxylamine method using SOD kits (Jiancheng Bioengineering, Nanjing, Jiangsu, China). The determination of SOD activity was based on the suppression role of SOD on the superoxide anion through a dismutation reaction. C-reactive protein (CRP) was measured by the method of enzyme-linked immunosorbent assay (ELISA) using kits (R&D Systems, Minneapolis, MN, U.S.A.). The biochemical determinations were carried out in duplicates in triplicate and the mean value was recorded. Hemoglobin, white blood cell, albumin, urea and creatinine were also measured using the preoperative blood samples in the laboratory of our hospital.

The commercial AST, ALT, MDA, and SOD kits were purchased by Jiancheng Bioengineering Institute (Nanjing, China). Enzyme-linked immunosorbent assay (ELISA) tests for the detection of IL-6, IL-1β, and TNF-α were produced by Nanjing KeyGEN Biotech. Co., Ltd. (Nanjing, China). All the antibodies were provided by Cell Signaling Technology (Danvers, USA).