Glycerol-based amphiphilic polyether mono azide third-generation dendrons were synthesized through a divergent strategy. A repetitive synthetic sequence of O-alkylation and isopropylidene deprotection reaction was adopted for the synthesis, pentamethylene was used as spacer. Dansyl chloride was attached to the focal point as a fluorophore. The photophysical property in aqueous solution of the protected dendrons showed self-association behaviour from second generation onwards even at a very low concentration (1×10-8 M). A hypsochromic shift of around 60 nm in the emission maximum, significant increase in fluorescence intensity, almost ten-fold increase in the fluorescence anisotropy and a fourfold increase in average fluorescence lifetime of dansyl moi-ety were observed on going from first generation dendron to higher generation dendrons in aqueous medium. The absence of such behavior in the corresponding unprotected, relatively hydrophilic dendritic units clearly indicated that the aggregation is due to the presence of hydrophobic isopropylidene end groups. Quenching studies of the dendrimers in aqueous solution using a hydrophilic quencher Ag+ revealed that the dansyl moiety in protected higher generations dendritic units is significantly shielded from the surroundings, which further rendered support to the fact that higher order dendrons undergo aggregation in aqueous medium. © 2010 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.