Quality evaluation of NMN produced via different synthesis methods: A multi-technique analysis using NMR, IR, GC-TCD, and HPLC-UV

Abstract Nicotinamide mononucleotide (NMN) has gained significant attention as a promising anti-aging and health-promoting supplement, driving the development of efficient and sustainable production methods. Among these methods, enzymatic synthesis has emerged as a highly specific and environmentally friendly approach. This study presents a comprehensive evaluation of NMN synthesized via two key enzymatic pathways—nicotinamide riboside kinase (NRK)-mediated and nicotinamide phosphoribosyltransferase (NAMPT)-mediated routes—along with commercially available raw NMN materials. Key quality attributes, including purity, content, moisture levels, thermal stability, and biological activity, were systematically assessed. Results show that NAMPT-synthesized NMN consistently achieves 100% purity, matching pharmaceutical-grade standards, while NRK-synthesized NMN displays batch-dependent variability, with purity ranging from 92.13% to 96.19%. Stability analysis reveals that NAMPT-derived NMN maintains high thermal stability, whereas NRK-derived NMN undergoes significant physical and chemical degradation at elevated temperatures, with nicotinamide as the primary degradation product. Additionally, bioactivity testing demonstrated that NAMPT-derived NMN retains significantly higher biological activity compared with NRK-synthesized and commercial NMN. These findings provide essential data for optimizing NMN production processes and support the establishment of industry-wide quality control standards.