Noninvasive Nicotine Quantitation and Flavor Differentiation of <i>on!</i> Nicotine Pouches Using Raman Spectroscopy: A Direct, Reliable, and Preparation-Free Approach

Nicotine pouches are a growing category in the smoke-free market, serving as an alternative to traditional tobacco products due to their noncombustible nature. Liquid and gas chromatography techniques coupled to various detectors were proven to be effective for determining nicotine content in tobacco products, including nicotine pouches. However, they require multiple steps and labor-intensive procedures for analysis. In this study, we developed a novel Raman spectroscopic method for accurately determining nicotine content in nicotine pouch products using a wide-area illumination scheme providing a 6 mm laser illumination diameter and a long focal length. Using chemometric tools, we analyzed 15 nicotine pouch products from a single commercially available brand (<i>on!</i> nicotine pouches), encompassing five flavor variantsOriginal, Mint, Wintergreen, Citrus, and Berryacross three nicotine levels (2, 4, and 8 mg). Partial least-squares regression was used to determine the nicotine content. The method accuracy was found to be 99.0-102.3% for all 4 and 8 mg nicotine pouch products, including all five flavor variants. For 2 mg nicotine pouches, method accuracy was calculated to be 93.1-108.8%, except for the Berry flavor due to the more significant fluorescence interference in lower nicotine content. Furthermore, the Raman spectra were analyzed using the <i>k</i>-nearest neighbor, allowing the classification of nicotine pouches into five groups based on their flavor variants. The results underscore the use of Raman spectroscopy as a rapid, direct, and noninvasive technique for product characterization, enabling accurate determination of nicotine content in nicotine pouches and facilitating their classification into distinct flavor-based groups.