Waters beh c18 column

Wheat (Jimai 22) was provided by Crop Research Institute, Shandong Academy of Agricultural Sciences (Jinan, China). Flour (Fuqiang) was supplied by Beijing Guchuan Food Co., Ltd. (Beijing, China). The pesticide standards, including triphenyl phosphate (TPP), carbendazim, bensulfuron methyl, triazophos, chlorpyrifos, and carbosulfan (all 99% purity), were purchased from Accustandard Inc. (New Haven, CT, USA). The five pesticides (carbendazim, bensulfuron methyl, triazophos, chlorpyrifos, carbosulfan) chemical structures are shown in Figure 1. Chromatographic-grade methanol and formic acid were purchased from Mreda (Beijing, China), and 0.22 µm nylon membrane filter was purchased from Tianjin jinteng Experimental Equipment Co., Ltd. (Tianjin, China). Waters BEH C₁₈ column (100 mm × 2.1 mm, 1.7 μm) was purchased from Waters (Milford, CT, USA). QuEChERS purifier and salt package were purchased from Beijing Dima Outai Science Technology Co., Ltd. (Beijing, China).

A panel of 39 neurotransmitters including dopamine, serotonin, glutamic acid, L-glutamine, and γ-aminobutyric acid (GABA) in our 22 serum samples (case, n = 12; control, n = 10) were quantitatively detected by external calibration curves for each metabolite using AB Sciex 6500 Triple Quad LC–MS/MS at Beijing Genomics Institution (Shenzhen, China). Protein precipitation was first performed on the serum samples, aliquots of 50 μL were drawn, and four volumes of methanol were added. After thorough vortex and centrifugation, an aliquot of 180-μL supernatant was pipetted for drying. The dried residues were stored at − 80 °C until analysis. Quality control (QC) samples were performed using the same method. The chromatographic column is Waters BEH C18 column (1.7 μm, 2.1 mm × 100 mm, Waters, USA). The mobile phase consisting of 0.1% formic acid in water (A) and methanol (B) was delivered at a flow rate of 0.35 mL min⁻¹ and column temperature of 55 °C. The linear gradient elution program was as follows: 0–2 min, 2% B and 2.5–15 min, 20–80% B. Multiple reaction monitoring (MRM) was used in monitoring the results of all compounds. Differential neurotransmitters were screened as follows: VIP > 1.0, two-tailed Student’s t-test (P < 0.05), and ratio (< 0.8 or > 1.2).

The TP content of camellia seed oil was extracted by methanol–water solution (80%:20% v/v) and determined by Folin–Ciocalteu method according to the colorimetric method described previously by Delfan‐Hosseini, Nayebzadeh, Mirmoghtadaie, Kavosi, and Hosseini (2017). A calibration curve of gallic acid in methanol was carried out in the concentration ranges of 0.04–0.40 mg/ml. The results were expressed as μg gallic acid equivalent per gram of oil samples. Triplicate test was performed for each sample.
Tocopherols content of the extracted oils was determined using UPLC method with fluorescence detection. Waters liquid chromatography system equipped with a column heater, a photodiode array detector ACQ‐FLR, controlled by Waters Empower chromatographic software. In all analyses, an Acquity UPLC Waters BEH C18 column of 1.7 μm (2.1 × 50 mm) was used. The analysis was carried out at 35°C temperature under isothermal condition, and the mobile phase was composed of 100% acetonitrile. The volume of injection is 10 μl, and the flow rate was 0.5 ml/min. Using FLR to detect and quantify α‐tocopherol, the excitation wavelengths and emission wavelengths are 294 nm and 338 nm, respectively. A calibration curve of α‐tocopherol in toluene was performed in the concentration ranges of 0–300 mg/ml. Results were expressed in mg of α‐tocopherol per kilogram of oil.

Digestion of protein (250 μg per sample) from three oysters was performed according to the FASP procedure [29 (link)–31 (link)]. Protein quality was tested by a Bradford protein assay kit according to the manufacturer’s instructions. TMT labelling of peptides was performed according to a procedure described previously [32 (link)]. Mobile phase A (2% acetonitrile, adjusted to pH 10.0 using ammonium hydroxide) and B (98% acetonitrile) were used to develop a gradient elution. The lyophilised powder was dissolved in solution A and centrifuged at 12,000 g for 10 min at room temperature. The sample was fractionated using a Waters BEH C18 column (4.6 × 250 mm, 5 μm; Waters) on a Rigol L3000 HPLC system, with a column temperature of 45°C. Eluates were monitored at an absorbance wavelength of 214 nm, fractions were collected at one tube per min, and combined into 10 fractions. All fractions were dried under vacuum, then reconstituted in 0.1% (v/v) formic acid (FA) in water.