Oral nicotine pouches with an aftertaste? Part 1: screening and initial toxicological assessment of flavorings and other ingredients

Abstract

Nicotine pouches are oral products that deliver nicotine without containing tobacco. Previous studies mainly focused on the determination of known tobacco toxicants, while yet no untargeted analysis has been published on unknown constituents, possibly contributing to toxicity. Furthermore, additives might enhance product attractiveness. We therefore performed an aroma screening with 48 different nicotine-containing and two nicotine-free pouches using gas chromatography coupled to mass spectrometry, following acidic and basic liquid–liquid extraction. For toxicological assessment of identified substances, European and international classifications for chemical and food safety were consulted. Further, ingredients listed on product packages were counted and grouped by function. Most abundant ingredients comprised sweeteners, aroma substances, humectants, fillers, and acidity regulators. 186 substances were identified. For some substances, acceptable daily intake limits set by European Food Safety Agency (EFSA) and Joint FAO/WHO Expert Committee on Food Additives are likely exceeded by moderate pouch consumption. Eight hazardous substances are classified according to the European CLP regulation. Thirteen substances were not authorized as food flavorings by EFSA, among them impurities such as myosmine and ledol. Three substances were classified by International Agency for Research on Cancer as possibly carcinogenic to humans. The two nicotine-free pouches contain pharmacologically active ingredients such as ashwagandha extract and caffeine. The presence of potentially harmful substances may point to the need for regulation of additives in nicotine-containing and nicotine-free pouches that could be based on provisions for food additives. For sure, additives may not pretend positive health effects in case the product is used.

Oral nicotine pouches with an aftertaste? Part 2: in vitro toxicity in human gingival fibroblasts

“…Other substances in nicotine pouches could contribute to cytotoxicity and altered gene expression as concentrations are likely to rise with an increasing extraction time likewise to nicotine. Aroma substances contained in the five pouches were screened in part 1 of this project (Mallock-Ohnesorg et al 2023 ). Seven of the identified flavor compounds received a harmonized hazard classification label according to CLP: benzaldehyde, benzyl alcohol, benzyl benzoate, carvone, citral, limonene, and linalool.…”

“…Screening for unknown substances contained in nicotine pouches was performed in part 1 of this study where the procedure is described in more detail (Mallock-Ohnesorg et al 2023 ). In brief, a method using liquid–liquid extraction (LLE) and gas chromatography with mass spectrometric detection (GC/MS) was adapted from Hutzler et al ( 2014 ).…”

“…To address potential health risks that are new to nicotine pouches, this study was conducted in two parts. In part 1, 48 nicotine pouches and 2 nicotine-free pouches were assessed for their ingredients and for further substances identified by a GC–MS-based screening approach (Mallock-Ohnesorg et al 2023 ). An initial toxicological assessment was performed for the identified substances based on regulatory databases (Mallock-Ohnesorg et al 2023 ).…”

“…In part 1, 48 nicotine pouches and 2 nicotine-free pouches were assessed for their ingredients and for further substances identified by a GC–MS-based screening approach (Mallock-Ohnesorg et al 2023 ). An initial toxicological assessment was performed for the identified substances based on regulatory databases (Mallock-Ohnesorg et al 2023 ). For part 2, which is described in this manuscript, in vitro toxicity in human gingival fibroblasts (HGF-1) was assessed for five different nicotine pouches and the reference snus CRP1.1.…”

The Biological Effects of Toxic Dose Combustion Cigarettes and Heated Tobacco Products in BEAS-2B and A549 Cells

“…At present, there are many evaluation systems for the in vitro toxicological safety assessment of traditional cigarette smoke [8,9], but the research on the safety assessment system of heated tobacco products (HTP) is not yet comprehensive. At the same time, the HTP of emerging industry, which has no perfect technical regulations for the products in various countries, nor is there any unified standard in the industry.…”