RESEARCH ARTICLE Simultaneous quantitation of kavalactones in kava dry extracts: comparison of multi-standards and single standard validation approaches Juliana Veloso Ferreira | Isabella Campolina Pierotte | Gerson Antônio Pianetti | Isabela Costa César Departamento de Produtos Farmacêuticos, Faculdade de Farmácia, Universidade Federal de Minas Gerais, Belo Horizonte, MG, Brazil Correspondence Isabela Costa César, Departamento de Produtos Farmacêuticos, Faculdade de Farmácia, Universidade Federal de Minas Gerais. Belo Horizonte, MG 31270-901, Brazil. Email: isacesar@gmail.com Funding information Coordenaç~ ao de Aperfeiçoamento de Pessoal de Nível Superior – Brasil (CAPES) – Finance Code 001 Abstract Introduction: Dried extracts of Piper methysticum G. Forst, also known as kava, has been widely used due to its anxiolytic and sedative properties. In order to assure the quality of these extracts, it is essential to accurately quantify kavalactones, known as the active principle. Objectives: To develop and validate an analytical method for the simultaneous quantification of six major kavalactones (kavain, dihydrokavain, methysticin, dihydromethysticin, yangonin and demethoxyyangonin) in kava extracts, comparing multi-standards and single standard validation approaches. Material and methods: Separation was performed using a C18 column, water/ methanol/acetonitrile/2-propanol (66:07:09:18 v/v/v/v) and detection at 245 and 350 nm. A full method validation was performed, employing analytical standards for each compound. Commercial kava dried extracts were assayed and the results obtained using the method validated for six kavalactone standards were compared with those obtained when only kavain was used as standard. Results: Baseline resolution for all kavalactones was obtained in short run time (15 min). Although the total kavalactone content varied between samples, a similar distribution profile was observed. When the method validated with all six analytical standards was compared to the calibration using only kavain standard, kavalactone contents were considerably different (from 7.57 to 36.53%). Conclusion: The obtained results demonstrate the importance of a validated method using individual kavalactone standards for the effective quality control of kava extracts. In a next step, the method needs to be adapted to also include flavokavin B (FKB), as an important authentication marker to distinguish between the accepted variety “noble Kava” and the toxic “two-day Kava”. KEYWORDS kava, kavain, kavalactones, liquid chromatography, Piper methysticum G. Forst Received: 6 May 2020 Revised: 13 November 2020 Accepted: 14 November 2020 DOI: 10.1002/pca.3019 Phytochemical Analysis. 2020;1–9. wileyonlinelibrary.com/journal/pca © 2020 John Wiley & Sons, Ltd. 1