A Candidate for Multitopic Probes for Ligand Discovery in Dynamic Combinatorial Chemistry

Abstract

Multifunctionalized materials are expected to be versatile probes to find specific interactions between a ligand and a target biomaterial. Thus, efficient methods to prepare possible combinations of the functionalities is desired. The concept of dynamic combinatorial chemistry (DCC) is ideal for the generation of any possible combination, as well as screening for target biomaterials. Here, we propose a new molecular design of multitopic probes for ligand discovery in DCC. We synthesized a new Gable Porphyrin, GP1, having prop-2-yne groups as a scaffold to introduce various functional groups. GP1 is a bis(imidazolylporphyrinatozinc) compound connected through a 1,3-phenylene moiety, and it gives macrocycles spontaneously and quantitatively by strong imidazole-to-zinc complementary coordination. Some different types of functional groups were introduced into GP1 in high yields. Formation of heterogeneous macrocycles composed of GP1 derivatives having different types of substituents was accomplished under equilibrium conditions. These results promise that enormous numbers of macrocycles having various functional groups can be provided when the kinds of GP components increase. These features are desirable for DCC, and the present system using GP1 is a potential candidate to provide a dynamic combinatorial library of multitopic probes to discover specific interactions between a ligand and a biomaterial.

Keywords: amphiphilic; complementary coordination; copper(I)-catalyzed alkyne-azide cycloaddition (CuAAC); dynamic combinatorial chemistry (DCC); dynamic combinatorial library (DCL); gel permeation chromatography (GPC); multifunctionalized material; supramolecular macrocycle; zinc porphyrin.

Figure 1

An image of advanced multitopic DCC systems. Recombination of five kinds of self-assembled cyclic hexamers gives various heterogeneous cyclic hexamers.

Figure 2

An imidazole zinc porphyrin (ImZnP) and its self-assembled dimer.

Figure 3

A Gable porphyrin (GP) and its self-assembled pentamer and hexamer.

Figure 4

Molecular models of self-assembled (left top and bottom) pentamer and (right top and bottom) hexamer of GP1s prepared on Materials Studio^®. Gray: carbon, blue: nitrogen, dark blue: zinc, white: hydrogen atoms. Substituents on meso-positions are omitted for clarity.

Figure 5

Structure of Gable porphyrin GP1.

Figure 6

Possible coordination isomers (conformational isomers) of self-assembled pentamers and hexamers composed of Gable porphyrin (GP). Molecular models of a 5mer (Out-In/Out-Out/In-In = 5/0/0, clockwise) and a 6mer (6/0/0) are presented in Figure 4.

Figure 7

TrisPor gives two possible conformational isomers, which can be assigned in ¹H-NMR spectra.

Scheme 1

Preparation of compounds 5 and 6. Compounds 7 and 8 were detected in the crude mixture of 5, and they were not isolated.

Scheme 2

Preparation of free-base Gable porphyrin GP1Fb.

Scheme 3

Preparation of zinc porphyrin 5Zn and formation of its self-assembly dimer.

Scheme 4

Preparation of zinc Gable porphyrin GP1.

Figure 8

¹H-NMR (400 MHz, CDCl₃) of free-base porphyrin 5 and coordination dimer of zinc porphyrin 5Zn.

Figure 9

¹H-NMR (400 MHz, CDCl₃) spectra of (a) GP1Fb and (b) GP1.

Scheme 5

Introduction of various substituents into the imidazole moieties (moiety) on GP1 and 5Zn by CuAAA. Their molecular weights (M.W.) are shown with their structures.

Scheme 6

Introduction of a PEG8 moiety into 5Zn by CuAAA.

Figure 10

¹H-NMR (400 MHz, CDCl₃) spectra of (top) 5Zn and (bottom) PEG8-5Zn.

Figure 11

MALDI-TOF MS spectra of (a) GP 1, (b) PEG8-GP 1, (c) PY-GP1, (d) BA-GP1, and (e) F9GP-1 on JSM-S3000 (JEOL) with spiral mode. Matrix: DCTB, [M + Na]⁺ species were observed. Insets: their expanded spectra.

Figure 12

(a) Fluorescence (ex. 564 nm) and (b) UV-Vis absorption spectra of 5Zn in (plain) 100% acetonitrile and (dotted) a mixture of 80:20 vol/vol of water and acetonitrile. (c) Fluorescence quantum yields of 5Zn in a mixture of water and acetonitrile.

Figure 13

(a) Fluorescence (ex. 564 nm) and (b) UV–vis absorption spectra of PEG8-5Zn in (plain) 100% acetonitrile and (dotted) 100% ion exchange water. (c) Fluorescence quantum yields of PEG8-5Zn in a mixture of water and acetonitrile.

Figure 14

(a) GPC charts of (black) before and (red) after reorganization of PEG8-GP1. (column; TOSOH TSKgel G4000H_HR (exclusion limit; 400 kDa), eluent; CHCl₃:THF=95:5, flow rate; 1.0 mL/min, detection; 565 nm) (b) UV-vis spectra obtained on PDA detector at retention time (RT) 9.1 min in the GPC after reorganization. In this Figure, characteristic Soret bands, 448 and 446 nm, corresponding to 6mer and 5mer are overlapped, and they are difficult to be distinguished.

Figure 15

GPC charts of reorganized (a) GP1, (b) PY-GP1, and (c) PEG8-GP1. (column; TOSOH TSKgel G4000H_HR×2 (exclusion limit; 400,000 Da), eluent; CHCl₃:THF=95:5, flow rate; 1.0 mL/min, detection; 565 nm).

Figure 16

(a) UV-vis spectra of PEG8-GP1 overlaid before the peak maximum (17. 8 min) in Figure 15c. (b) Overlaid after the peak maxmum in Figure 15c. The maximum wavelength is shifted from 448 to 446 nm in (b).

Figure 17

GPC charts of PEG8-GP1 reorganized at (a) 0 °C, (b) 25 °C, (c) 40 °C, and (d) 60 °C. (column; TOSOH TSKgel G4000H_HR×2 (exclusion limit; 400,000 Da), eluent; CHCl₃:THF = 95:5, flow rate; 1.0 mL/min, detection; 565 nm).

Figure 18

GPC charts of reorganized (a) PY-GP1, (b) PEG8-GP1, (c) as prepared, and (d) reconstituted samples of 1:1 mixture of PY-GP1 and PEG8-GP1. (column; TOSOH TSKgel G4000H_HR×2 (exclusion limit; 400,000 Da), eluent; CHCl₃:THF = 95:5, flow rate; 1.0 mL/min, detection; 565 nm).

Figure 19

(a) UV-vis spectra of PY-GP1 and PEG8-GP1 overlaid before the peak maximum (17.8 min) in Figure 18d. (b) Overlaid after the peak maximum in Figure 18d. The maximum wavelength is constant at 446 nm in both (a) and (b).

Figure 20

Five UV-vis absorption spectra normalized at 446 nm are overlaid. Original data were obtained on GPC analysis as shown in Figure 18d. At 350 nm, the five spectra of RT: 17.70, 17.80, 17.91, 18.10, and 18.18 min can be discriminated from the bottom to the top. Pyrene moiety is mainly observed in the rage of 250–350 nm. Increasing the intensities at 350 nm indicates that the ratio of the PY-GP1 component increased compared with that of PEG8-GP1 in the cyclic pentamers.

Figure 21

Possible heterogeneous cyclic structures composed of 1:1 ratio of PY-GP1 and PEG8-GP1, (a) 3:3 micelle-like cyclic hexamer. All of the pyrene moieties are gathered inside, whereas PEG groups exist in the outside. Not observed. (b) A mixture of 3:2 and 2:3 of cyclic pentamer.

Figure 22

¹⁹F-NMR spectra (376.32 MHz, CDCl₃, 298 K) of (a) F9-N3 and (b) dimer of F9-5Zn.

Figure 23

¹⁹F-NMR spectra (376.32 MHz, CDCl₃, 298 K) of F9-GP1. (a) before and (b) after reorganization.

Figure 24

¹⁹F-NMR spectra (376.32 MHz, CDCl₃, 298 K) of (a) reorganized sample of F9-GP1, and (b) reconstituted sample composed of F9-GP1 and PEG8-GP1 (1:1).