Determination of Ethanol Content in Kombucha Products by Gas Chromatography with Flame Ionization Detection: A...

Background: Kombucha is a nonalcoholic, fermented tea beverage that has recently received negative attention because of documented concentrations of ethanol in excess of allowable limits of ≥0.5% alcohol by volume (ABV). Objective: Our previously reported headspace GC with flame-ionization detection (GC-FID) method was adopted by the AOAC Expert Review Panel as First Action Official Method^SM 2016.12 in September 2016 based on published single-laboratory validation study results. This paper describes the corresponding multilaboratory study using this method to further validate its performance parameters. Methods: Four laboratories participated in the study and received practice samples, test samples, reference standards, and detailed protocols. Eight kombucha samples sent out to laboratories were randomly assigned sample numbers and were blinded in terms of content and identity. Each laboratory analyzed all samples using the GC-FID method and reported their results. Results: Cochran's C-test and single and double Grubbs' tests were used to identify and remove outliers. Horwitz ratio values for all samples were between 0.5 and 1.7. As per the Standard Method Performance Requirements (SMPRs^®), all samples within the analytical range of 0.1-2.0 ABV% had RSD_R values <6%. Conclusions: The results from this study demonstrate the evaluated GC-FID method meets the SMPR requirements and is fit for purpose for detecting ethanol in kombucha products. Highlights: Kombucha, a nonalcoholic, fermented beverage, has been found to contain ≥0.5% ABV. First Action Official Method 2016.12, a headspace GC-FID method for determining ethanol in kombucha, is supported by a multilaboratory study.