Self-Association of Protected Newkome-Type Second-Generation Dendrimers at Nanomolar Level Concentrations in Aqueous Solution Subhendu K. Mohanty, Shyamala Thirunavukarasu, Sundarababu Baskaran,* and Ashok K. Mishra* Department of Chemistry, Indian Institute of Technology Madras, Chennai-600036, India Received January 27, 2004; Revised Manuscript Received May 4, 2004 ABSTRACT: Protected Newkome-type second-generation dendrimers (based on Lin’s amine) were synthesized with a pyrene moiety attached to the core. The photophysical property in aqueous solution of the protected dendrimers shows self-association behavior in water. Pyrene excimer emission at 475 nm is observed in water even at very low concentrations of protected dendrimer (ca. 5 × 10 -8 M). This emission band is absent in other solvents even up to a concentration of 10 -5 M. The corresponding unprotected dendrimer does not show the pyrene excimer fluorescence. The amide of pyrene butyric acid with tert-butylamine shows the formation of excimer, albeit with very low intensity. Quenching studies on the dendrimer with hydrophilic quencher iodide anion (I - ) reveal that there is significant quenching of fluorescence intensity in the case of N-tert-butyl-4-pyren-1-ylbutyramide as compared to that of the pyrene-attached second-generation protected dendrimer. This shows that the pyrene moiety in the case of the protected dendrimer is significantly shielded from the surrounding. Introduction Higher generation dendrimers assume a globular shape and as such act like micelles 1 with the core of the dendrimer completely shielded from the environ- ment. On the other hand, lower generation dendrimers due to the partial shielding of the core from the environment reveal greater detail about the structural features of the dendrimer. Hyperbranch dendrimers 2 have been the focus of recent studies pertaining to structural morphology of dendrimers. Cardona and co- workers 3 have reported the synthesis and the photo- physical properties of Newkome-type 4 dendrimers with different fluorescent chromophores (fluorophores). A fluorophore when attached to the core provides insight into the physical properties of the molecule as a whole. Pyrene, 5 due to its long fluorescence lifetime and its characteristic excimer emission, is an ideal fluorophore for such studies. Sluch and co-workers 6 reported the excitation dynamics of poly(allylcarbosilane) dendrimers with pyrene at the core at a concentration of ca. 10 -3 M. Recently Chou and co-workers 7 reported the self- association of hyperbranched poly(sulfone-amine) den- drimers in water. This was studied by the addition of pyrene-labeled dendrimer to an aqueous solution con- taining free pyrene and monitoring the resultant exci- mer emission. A protected Newkome-type second-generation den- drimer has a hemiellipsoid structure (Figure 1) with the core being accessible to the environment. In this article, we report the self-association of such a dendrimer in water even at nanomolar concentration (5 × 10 -8 M). Experimental Section Synthesis of Dendrimers. Scheme 1 depicts the synthesis of Lin’s amine, monomers 6 and 10, starting from TRIS. Tris- (hydroxymethyl)aminomethane was treated with tert-butyl acrylate in the presence of DMSO and NaOH, resulting in the formation of the corresponding triester 6. 7 Protection of the nitrogen with benzyl chloroformate produced 7, and hydrolysis of the tert-butyl groups using formic acid resulted in the formation of the triacid 8 quantitatively. Triacid 8 when refluxed with catalytic amount of pTSA in ethanol and benzene resulted in the triethyl ester 9 in quantitative yield. Triethyl ester 9 under catalytic hydrogenation yielded the amine derivative 10. The monomers 6 and 10 were then used for the synthesis of dendrimers, tetraamide 2 and 3, respectively (Scheme 2). The triacid 8 was coupled with monomers 6 and 10 under peptide coupling conditions, resulting in the second-generation dendrimers 11 and 12, respectively. Hydrogenation of the carbamates 11 and 12 gave the corresponding free amines 13 and 14, respectively, which were then coupled to pyrenebutyric acid under similar peptide coupling conditions as was used for the preparation of the second-generation dendrimers 11 and 12 (Scheme 2). Using similar reaction conditions, monoa- mide 1 was synthesized from pyrenebutyric acid and tert- butylamine. Reagents for Synthesis. All chemicals were reagent grade. Tetrahydrofuran (THF) used was freshly distilled from sodium benzophenone. Column chromatography was performed on silica gel (100-200 mesh) while TLC was performed on aluminum-backed plates coated with 0.25 mm silica gel. N-Tris[(2-{[(tris{[2-(tert-butoxycarbonyl)ethoxy]- methyl}methyl)amino]carbonyl}ethoxy)methyl]methyl}- 4-pyren-1-ylbutyramide (Tetraamide 2). To a solution of 4-(1-pyrene)butyric acid (58 mg, 0.2 mmol) in dry THF (1 mL) was added EDCl (46 mg, 0.2 mmol), HOBt (27 mg, 0.2 mmol), and Et 3N (34 μL, 0.2 mmol). To the stirred solution at room temperature was added N-tris[(2-{[(tris{[2-(tert-butoxycarbonyl)- ethoxy]methyl }methyl)amino]carbonyl }ethoxy)methyl]- methylamine 8 (360 mg, 0.2 mmol) dissolved in dry THF (3 mL). The reaction was stirred at room temperature, and after the completion of reaction, as indicated by TLC, THF was removed under vacuum, and the reaction mixture was diluted with EtOAc (10 mL) and washed with water (5 mL), 0.5 M HCl solution (5 mL), and then with saturated solution of brine (5 mL). The organic layer was separated, dried over anhydrous Na 2SO4, and concentrated under reduced pressure to yield the crude compound, which was purified over silica gel using 50-80% gradient elution of EtOAc in hexane to yield the pure pyrene attached dendrimer, tetraamide 2 (200 mg), in 48% yield. IR (neat): 3440, 2912, 1728, 1667, 1523, 1369, * Corresponding authors. E-mail: sbhaskar@iitm.ac.in (S.B.); mishra@iitm.ac.in (A.K.M.). 5364 Macromolecules 2004, 37, 5364-5369 10.1021/ma0498215 CCC: $27.50 © 2004 American Chemical Society Published on Web 06/18/2004