Hplc gradient grade

Standards of flavonoids (scutellarin, baicalin, baicalein, wogonin, wogonoside, luteolin, chrysin), verbascoside, sugars (glucose, fructose, xylose, galactose, rhamnose, sucrose and cellobiose) and organic acids (quinic, malic, ascorbic, succinic, citric, α-keto-glutaric, oxalic and tartaric), methanol and acetonitrile (HPLC gradient grade) were purchased from Sigma-Aldrich (Darmstadt, Germany). Water was obtained from Milipore Milli-Q Gradient water purification system (Barnstead, US).

The total phenolic content was established by using the Folin–Ciocalteu reagent following a little modified method [48 (link)]. The sub-extracts, extracts were evaporated and then dissolved with 500 μL of 70% methanol (HPLC-Gradient grade, Sigma–Aldrich, Darmstadt, Germany VWR chemicals) at 70 °C. The mixtures were centrifuged at 3500 g for 10 min, and the supernatants were collected in separate tubes. The pellets were re-extracted under identical conditions. Supernatants were combined and used for total phenolic assay. For the total phenolic assay, 20 μL of extract was dissolved into 2 mL of distilled water. Two hundred microliters of dissolved extract were mixed with 1 mL of Folin–Ciocalteu reagent (previously diluted tenfold with distilled water) and kept at 25 °C for 3–8 min; 0.8 mL of sodium bicarbonate (75 g L⁻¹) solution was added to the mixture. After 60 min at 25 °C, absorbance was measured at 765 nm. The results were expressed as gallic acid equivalents. The measurement of total phenolics was done under absorbance at 765 nm with the use of a Jenway UV/Vis 6405 spectrophotometer (Jenway, Chelmsford, UK). The total phenolic contents were determined from the linear equation of a standard curve prepared with gallic acid. The results were calculated as mg of gallic acid equivalent per g dry weight of extract.

C118P standard compound (C118 phosphate, chemical purity is 97.3%), C118P injection (C118P content is 95.6%) and C118 (chemical purity is 95.9%) were manufactured by Nanjing Sanhome Pharmaceutical Co. Ltd. (Nanjing, China). Colchicine (chemical purity is 98.8%, as an internal standard) was purchased from Meryer (Shanghai) Chemical Technology Co., Ltd (Shanghai, China). Methanol and acetonitrile were purchased from Merck (HPLC gradient grade, LiChrosolv GG, Darmstadt, Germany). Milli-Q water was generated from Milli-Q system (Gradient A1, Millipore Inc, Bedford, MA, USA). Pooled human liver microsomes (HLMs) were prepared from eleven Mongolian donors aged from 24 to 38 and were purchased from RILD Research Institute for Liver Diseases Co. Ltd. (Shanghai, China). HPLC grade or the best grade commercially available was used in our study for all other chemicals.

Antibiotic standards for this study were sourced from Shanghai Yuanye Bio-Technology Co., Ltd. in Shanghai, China. The investigation focused on four categories of antibiotics within nine distinct sections of the yellow croaker fish. These categories included QNs, SAs, MLs, and TLs, comprising a total of 42 different compounds, as listed in Table 1. All standard stock solutions were stored in opaque containers at a temperature of −20 °C, with fresh stock solutions being prepared anew every three months. The solvent used was methanol of HPLC-gradient grade, procured from Merck KGaA in Darmstadt, Germany. Ultra-pure water was generated by utilizing the Milli-Q Integral Water Purification System produced in Billerica, MA, USA. Ethylenediaminetetraacetic acid disodium salt (Na2EDTA), of HPLC-gradient grade and with a minimum purity of 99.0%, formic acid of HPLC-gradient grade and with a minimum purity of 98%, and ammonia solution of chromatography grade with a minimum purity of 25% were all acquired from Aladdin in Shanghai, China.