Rationale Over the years, many techniques have been developed for honey adulteration detection. The several methods based on the AOAC have been admitted to the in-house method confirmed by Iranian Standard Organization (ISIRI). Besides the unprecedented fraud in honey, the scientific requirements to apply high-tech procedures. Therefore, unraveling these discrepancies requires comparisons between the elderly and novel methods. Methods The conventional methods determined different parameters such as reducing sugars before hydrolysis, sucrose amount, fructose to glucose ratio, Proline, and hydroxy methyl furfural. Applying novel IRMS technique determined δ13C values for glucose with high quality and quantity. The honey carbohydrates, such as glucose, fructose, disaccharide, and trisaccharide, were assessed using LC/IRMS. Results Based on these findings regarding the conventional and IRMS methods for testing samples. According to our results, the conventional method successfully passed 18 to 20 samples by ISIRI standards. However, the IRMS method showed different results, with only 2 out of 20 samples being original and pure without adulteration. Conclusion We collected twenty samples from various regions and examined them using two methods to determine their authenticity. The first method used is the C13 isotope and isotope ratio mass spectrometry (IRMS), while the second method is based on conventional methods using ISIRI. After analyzing the results with Prism software, we found a statistically significant difference between the confidence level of the IRMS method and conventional methods, with a p-value less than 0.05.

Abstract Recently, food fraud has been observed in all aspects more than in the past and one of the most common impurities has been identified in honey. Industries try to produce fake honey to make more profit. Recently, honey has been adulterated in more sophisticated ways that are difficult to detect by conventional methods. In this study, a new isotope ratio mass spectrometry method was used in comparison with traditional chemistry methods based on the national standards of Iran (ISIRI). twenty different honey samples were collected from other regions. These samples have been examined using two methods to determine authenticity. The first one is related to the C13 isotope and using isotope ratio mass spectrometry (IRMS) and the second one is conventional methods based on ISIRI. The results have been analyzed by Prism software, and based on this, the difference between the confidence level of IRMS method and conventional methods was statistically significant and the p-value was less than 0.05.