Stoichiometry-Dependent Structural Defects in Low-Generation PAMAM Dendrimers

Abstract Low-generation PAMAM dendrimers synthesized via a divergent approach from EDA cores were characterized to assess stoichiometry-dependent structural defects and product heterogeneity. MALDI-TOF MS, RP-HPLC-UV, SEC, and 1H NMR were employed as complementary techniques to assign and compare defect-related signatures, including trailing-generation species, intramolecular cyclization, and intermolecularly linked dendritic byproducts and/or higher-mass linked dendritic populations. Theoretical molecular weights were estimated using a modified Carothers equation and compared with experimental descriptors, although these comparisons were interpreted as descriptive stoichiometric trends rather than quantitative validation of the theoretical treatment. MALDI-TOF MS revealed repetitive fragment intervals (Δm/z = 28, 60, 114), consistent with incomplete amidation, cyclization, and retro-Michael reactions. RP-HPLC-UV was used as an orthogonal semi-quantitative chromatographic fingerprinting technique to compare sample heterogeneity under identical analytical conditions. SEC provided apparent hydrodynamic-size information, and SEC-derived molecular-weight-related descriptors were interpreted cautiously due to the compact architecture of dendrimers and calibration with linear polymer standards, while 1H NMR gave semi-quantitative information on branched and bridged amide environments. The fraction of non-bridged species (f0) showed a trend consistent with theoretical expectations, although the limited number of stoichiometric conditions precludes rigorous statistical validation. These findings highlight the critical role of stoichiometric control in minimizing defects and reducing structural heterogeneity in low-generation PAMAM products, and emphasize the value of complementary analytical techniques for semi-quantitative assessment of dendrimer heterogeneity.