Application of COI Gene-Based Molecular Analysis for Verifying Honey Authenticity and Detecting Trace Residues in Vegan Food Products

Honey is a natural bee product with confirmed health-promoting properties, the quality and authenticity of which are of key importance from a consumer’s perspective. However, the demand for honey is affected by the problem of its adulteration. Moreover, despite its numerous taste and health benefits, honey may be an undesirable product for some groups of consumers, such as people with allergies or vegans. This work aimed to develop a sensitive molecular test enabling the unambiguous detection of honey adulteration and the identification of its trace amounts in food products. The test was based on the analysis of a fragment of the cytochrome c oxidase gene subunit I using real-time PCR with SYBR®Green dye and melting curve analysis. The key parameter of the analysis was the melting temperature, which in the case of natural honey was within a narrow range of 74.34–75.38 °C (for its dilutions, 71.10–77.00 °C). The developed method demonstrated high repeatability and sensitivity, enabling the detection of honey presence even at a level of 0.1%. To products labelled as vegan, Tm analysis effectively distinguished samples containing trace amounts of honey from those that were truly vegan. The procedure used is simple, highly repeatable, and effective even in the case of processed products. The developed method can be successfully used to control the quality and authenticity of honey, meeting the requirements of V-Label certification.