Malays. J. Anal. Sci. Volume 29 Number 5 (2025): 1583

Research Article

Determination of trace ethanol levels in kombucha by GC–MS for halal verification

Wahyu Utami^* and Maryam Dzakiyah Muthiatulloh

Laboratory of Pharmaceutical Chemistry, Faculty of Pharmacy, Universitas Muhammadiyah Surakarta, Sukoharjo, Jawa Tengah 57169, Indonesia

^*Corresponding author: wahyu.utami@ums.ac.id

Received: 19 May 2025; Revised: 20 August 2025; Accepted: 28 September 2025; Published: 16 October 2025

Abstract

Ethanol, a by-product of the fermentation process in kombucha, is a crucial factor in assessing halal compliance, particularly in Muslim-majority countries. This study employed gas chromatography-mass spectrometry (GC–MS) with direct injection to quantify ethanol levels in kombucha, thereby providing high selectivity and specificity. The analysis employed a GC system featuring an Rtx-1MS column with a split ratio of 10:1, in conjunction with an MS system operating in scan mode across a mass range of 20.0 to 100.0 amu. Method validation was performed in accordance with the Association of Official Analytical Collaboration (AOAC) International guidelines, ensuring adherence to the Standard Method Performance Requirements (AOAC SMPR 2016.001) for ethanol determination in kombucha. The parameters assessed included selectivity, specificity, linearity, precision, accuracy, and the limits of detection and quantification (LOD/LOQ). The method demonstrated excellent linearity (R² > 0.990) over an ethanol concentration range of 0.05% to 3.2%. Furthermore, it exhibited a precision of 2.55% relative standard deviation (RSD), an accuracy ranging from 97% to 99%, and a limit of quantification (LOQ) of 0.001% alcohol by volume (ABV). The method met all the AOAC Standard Method Performance Requirements. Analysis of two kombucha products available on Indonesian e-commerce platforms revealed ethanol levels that exceeded halal standards. For instance, the Indonesian Council of Ulama (MUI) specifies a maximum limit of 0.5% ABV, while the Department of Islamic Development Malaysia (JAKIM) sets a threshold of 1% ABV.

Keywords: ethanol, kombucha, GC–MS, halal verification, method validation

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