AOAC 2018 Methods

(p) LC vials and caps. (q) Chromatographic column.— Supelco Ascentis Express C18, 2.0 µm, 150 x 2.1 mm, Part No. 50814-U. C. Materials and Reagents (a) Methanol (MeOH). —HPLC grade. (b) Water (H 2 O). —LC-MS grade or deionized. (c) Acetonitrile (ACN). —HPLC grade (LC-MS grade is required if MS detection is used). (d) Ethanol (EtOH). —HPLC grade. (e) Ammonium formate (NH 4 HCO 2 ). —LC-MS grade. (f) Formic acid (FA). —LC-MS grade. D. Reference Standards Commercial reference standards were obtained in the form of solutions at a concentration of 1 mg/mL in methanol. Standards listed in Appendix 1 were purchased from Cerilliant (Round Rock, TX, USA). E. Preparation of Reagent Solutions and Standards Note: Different volumes may be prepared provided that the concentrations remain the same. All standard solutions should be stored at -20°C. (a) LC Mobile Phase A (20 mM aqueous NH 4 HCO 2 , pH 3.2). —Weigh 1.26 ± 0.01 g of NH 4 HCO 2 into an appropriate reservoir, add 1000 mL of H 2 O. Mix thoroughly. Adjust pH value of the mobile phase A to 3.2 using FA and mix thoroughly. (b) LC Mobile phase B (ACN). —Transfer 1000 mL of ACN to a suitable glass vessel. (c) Mixed Stock Standard Solution (10 µg/mL). —Combine 100 µL of individual analyte solutions into a 10-mL volumetric flask. Dilute to volume with MeOH. (d) Calibration Standard Solutions. —Dilute the Mixed Stock Standard Solution with MeOH using the dilution scheme in Table 2018.11A to prepare various calibration standard solutions in 2-mL amber LC vials. Vortex to mix. Table 2018.11A. Preparation of cannabinoid calibration standard solutions Calibration level Final concentration (µg/mL) Volume of Mixed Stock Standard Solution (µL) Volume of MeOH (µL)

1 2 3 4 5

0.5 1.0 2.5 5.0

50

950 900 750 500

100 250 500

10.0

1000

0

F. Preparation of Test Samples (a) Homogenization and Storage of Samples. —A minimum of 5 g of representative sample should be taken for homogenization (procedures described in the document Guidance