ISOTOPIC CHARACTERIZATION OF CREATINE SUPPLEMENTS FOR AUTHENTICITY EVALUATION

Creatine (α-methyl guanidinoacetic acid) is a widely used supplement due to its role in the rapid resynthesis of ATP during high-intensity efforts. In the market, supplements present formulation variations that reflect different raw material origins and production processes. In this context, evaluating the consistency between the declared composition and the isotopic profiles is relevant to characterize products and understand the diversity among brands. Carbon is sensitive to the addition of plant-derived carbohydrates (C3 or C4), while hydrogen and oxygen isotopes can reflect synthesis pathways and industrial conditions, allowing differentiation of creatine formulations even without a direct link to environmental factors. The objective of this study was to isotopically characterize creatine supplements to assess their authenticity in relation to the declared composition. A total of 47 commercial samples of creatine were analyzed, including pure products, pre-workouts, Creapure® versions, and formulations containing maltodextrin. Samples were dried in a vacuum oven and, subsequently, for analysis, aliquots of 200 to 250 μg were weighed into silver capsules for deuterium and oxygen and into tin capsules for carbon, using an analytical balance with a resolution of 1 μg (MX5, Mettler Toledo, Switzerland). Isotopic analyses were performed using an isotope ratio mass spectrometry system (EA Isolink-Delta V, Thermo Scientific), which determined the isotopic ratios (δ¹³C, δ²H, and δ¹⁸O) and the elemental abundances (%H and %O) of carbon, hydrogen, and oxygen. The results showed that δ¹³C was effective in differentiating samples with declared maltodextrin, with values ranging from -31.74 to -23.23 mUr for those without it and from -17.13 to -15.25 mUr for those containing maltodextrin, due to its non-synthetic plant origin, generating distinct carbon and oxygen signatures compared to pure creatine. The %O ranged from 26.20 to 31.47% in pure formulations and from 36.78 to 39.87% in those with maltodextrin, also allowing their separation. The δ²H revealed characteristic patterns for Creapure® (ranging from -64.67 to -59.42 mUr) and pre-workout samples (−23.01 mUr), while %H values ranged from 5.81 to 6.68%, indicating sample standardization. The δ¹⁸O showed little differentiation among the analyzed sample types. The isotopic analysis of the three elements proved effective for identifying adulteration based on different creatine formulations, allowing the evaluation of consistency with the declared ingredients and providing insight into the isotopic diversity present in commercial supplements.