Size Exclusion Chromatography Method for Purification of Nicotinamide Mononucleotide (NMN) from Bacterial Cells

Over 12% of the world’s health resources are spent on treating diabetes, as high blood glucose is the third cause of mortality worldwide. Insulin resistance is the basis of the most common form of diabetes: type 2 diabetes. Recent animal studies report successful attempts at reversing type 2 diabetes by the administering of the NAD+ precursor nicotinamide mononucleotide (NMN). However, the current high price of this molecule urges for more efcient and cost-efective production methods. This work proposes a method for purifying NMN by Size Exclusion Chromatography (SEC) on silica with a covalently attached coating of poly(2-hydroxyethyl aspartamide) (PolyHEA) stationary phase using an isocratic elution with a denaturing mobile phase (50mM formic acid) from a complex molecular mixture such as a fermentation broth. The eluted peaks were identifed by UV-Vis analysis and confrmed with ESI+ mass spectrometry and a HPLC reversed-phase method. The proposed SEC method is simple, patent-free, directly applicable for industrial production with a minimum scale up efort. The need for multiple chromatographic steps is eliminated and the lysate fltration and clarifcation steps are simplifed. Substantial reduction in NMN production costs and increased purity of NMN to the level suitable for usage in humans are expected.