Decasubstituted Pillar[5]arene Derivatives Containing L-Tryptophan and L-Phenylalanine Residues: Non-Covalent Binding and Release of Fluorescein from Nanoparticles

Abstract

Sensitive systems with controlled release of drugs or diagnostic markers are attractive for solving the problems of biomedicine and antitumor therapy. In this study, new decasubstituted pillar[5]arene derivatives containing L-Tryptophan and L-Phenylalanine residues have been synthesized as pH-responsive drug nanocarriers. Fluorescein dye (Fluo) was loaded into the pillar[5]arene associates and used as a spectroscopic probe to evaluate the release in buffered solutions with pH 4.5, 7.4, and 9.2. The nature of the substituents in the pillar[5]arene structure has a huge influence on the rate of delivering. When the dye was loaded into the associates based on pillar[5]arene derivatives containing L-Tryptophan, the Fluo release occurs in the neutral (pH = 7.4) and alkaline (pH = 9.2) buffered solutions. When the dye was loaded into the associates based on pillar[5]arene with L-Phenylalanine fragments, the absence of release was observed in every pH evaluated. This happens as the result of different packing of the dye in the structure of the associate. This fact was confirmed by different fluorescence mechanisms (aggregation-caused quenching and aggregation-induced emission) and association constants. It was shown that the macrocycle with L-Phenylalanine fragments binds the dye more efficiently (lgK_a = 3.92). The experimental results indicate that the pillar[5]arene derivatives with amino acids fragments have a high potential to be used as a pH-responsive drug delivery devices, especially for promoting the intracellular delivering, due to its nanometric size.

Keywords: L-Phenylalanine; L-Tryptophan; dye release; fluorescein; nanoparticles; pillar[5]arene; self-assembly.

Figure 1

Planar pS- and pR-conformers of methoxypillar[n]arenes (n = 5–10).

Scheme 1

Synthetic route for preparation of pillar[5]arenes 2, 3.

Figure 2

Graphs of cell viability for (a) MCF-7 and (b) PC-3 lines treated with compounds 2 and 3 obtained by colorimetric MTT test.

Figure 3

UV–vis spectra of mixtures of Fluo (1 × 10⁻⁴ M) with (a) pillar[5]arene 2 and (b) pillar[5]arene 3. Macrocycles concentrations were varied from 3 × 10⁻⁵ M to 3 × 10⁻⁴ M. Arrows mean hypochromic effect with increasing concentration of pillar[5]arenes.

Figure 4

(a) Fluorescence spectra of Fluo (1 × 10⁻⁵ M) at different pillar[5]arene 2 concentrations (5 × 10⁻²–1.5 × 10⁻⁴ M); (b) fluorescence spectra of Fluo (1 × 10⁻⁵ M) at different pillar[5]arene 3 concentrations (5 × 10⁻²–1.5 × 10⁻⁴ M).

Figure 5

Schematic representation of the formation of various associates based on synthesized macrocycles 2, 3, and Fluo, exhibiting different fluorescence mechanisms.

Figure 6

(a) Fragment of the 2D ¹H–¹H NOESY NMR spectrum of 2+Fluo in 1:1 molar ratio (CD₃OD–d₄, 298 K, 400 MHz); (b) scheme of interaction between macrocycle 2 and Fluo.

Figure 7

Top 3 scoring poses of fluorescein docked to the different conformations of (a) 2. Dashed lines represent hydrogen bonding (yellow), electrostatic contacts between charged atoms (purple), π–stacking (blue). (b) The first complex from (a) with 2 shown using a space-filling model.

Figure 8

TEM image of aggregates formed by macrocycle 2 and Fluo (a); 3 and Fluo (b) at 1:1 ratio (EtOH–H₂O, 1 × 10⁻⁴ M).

Figure 9

Electronic absorption spectra of the system 2+Fluo in (a) phosphate buffer solution (pH = 7.4); (b) sodium tetraborate buffer (pH = 9.2) (C₂ = 1 × 10⁻⁵ M, C_Fluo = 1 × 10⁻⁵ M).