Probing the Interactions of Porphyrins with Macromolecules Using NMR Spectroscopy Techniques

Abstract

Porphyrinic compounds are widespread in nature and play key roles in biological processes such as oxygen transport in blood, enzymatic redox reactions or photosynthesis. In addition, both naturally derived as well as synthetic porphyrinic compounds are extensively explored for biomedical and technical applications such as photodynamic therapy (PDT) or photovoltaic systems, respectively. Their unique electronic structures and photophysical properties make this class of compounds so interesting for the multiple functions encountered. It is therefore not surprising that optical methods are typically the prevalent analytical tool applied in characterization and processes involving porphyrinic compounds. However, a wealth of complementary information can be obtained from NMR spectroscopic techniques. Based on the advantage of providing structural and dynamic information with atomic resolution simultaneously, NMR spectroscopy is a powerful method for studying molecular interactions between porphyrinic compounds and macromolecules. Such interactions are of special interest in medical applications of porphyrinic photosensitizers that are mostly combined with macromolecular carrier systems. The macromolecular surrounding typically stabilizes the encapsulated drug and may also modify its physical properties. Moreover, the interaction with macromolecular physiological components needs to be explored to understand and control mechanisms of action and therapeutic efficacy. This review focuses on such non-covalent interactions of porphyrinic drugs with synthetic polymers as well as with biomolecules such as phospholipids or proteins. A brief introduction into various NMR spectroscopic techniques is given including chemical shift perturbation methods, NOE enhancement spectroscopy, relaxation time measurements and diffusion-ordered spectroscopy. How these NMR tools are used to address porphyrin-macromolecule interactions with respect to their function in biomedical applications is the central point of the current review.

Keywords: NMR spectroscopy; cyclodextrin; drug delivery; interaction; micelles; nucleic acids; phospholipids; polymer; porphyrin; proteins; surfactant.

Figure 1

Structures of porphyrins (I), chlorins (II), bacteriochlorins (III), corrins (IV), tetraphenylporphyrins (V), phthalocyanines (VI), texaphyrins (VII), and corroles (VIII).

Figure 2

Scheme of porphyrin ring current.

Figure 3

¹H NMR spectrum of chlorin e4 in DMSO-d₆.

Figure 4

Overview of selected NMR techniques to study porphyrin–macromolecule interactions.

Figure 5

¹H NMR spectrum (500 MHz) of DOPC SUVs in water before (bottom) and after (top) addition of a Ce6 derivative. The interaction of the chlorin with the DOPC bilayer induces a split of the DOPC choline resonances so that the outer and inner PL layers become distinguishable.

Figure 6

¹H NMR spectra of selected polymeric nanoparticles used for drug delivery in aqueous solutions, from bottom to top: DOPC SUVs, triblock copolymer (PEG-PPG-PEG) micelles, polyvinylpyrrolidone (PVP) and β-cyclodextrin (CD).

Figure 7

2D ¹H¹H NOESY (excerpt) of chlorin e6 serine amide (SerCe) associated with PVP at a SerCe/PVP molar ratio of 3:20 in phosphate buffered saline (PBS). (Reprinted with permission from: M. Hädener et al., J. Phys. Chem. B 2015, 119, 12117−12128. [163] Copyright © 2015, American Chemical Society).

Figure 8

(A) Structure of cyclodextrin and the α-1,4-d-glucopyranose unit with atom numbering; sketch of the cone-shaped structure of CDs in which the protons H(3) and H(5) point to the hydrophobic interior of the cavity (blue), the protons H(1), H(2) and (H4) point to the hydrophilic exterior (red), and the protons H(6,6′) are located at the rim of the primary face. (B) Examples of possible 2:1 inclusion complexes formed with tetraphenylporphyrins (TPPs): (I) inclusion via secondary face, (II) inclusion via primary face, opposite (anti) conformation, and (III) complex with adjacent (syn) conformation.

Figure 9

Structures of micelle forming surfactants.

Figure 10

¹H NMR spectra of CTAB micelles (A) in the presence of TPPS₄ and (B) in the presence of chlorin e6 (Ce6) at increasing molar ratios TPPS₄/Ce6 : CTAB in aqueous buffer solutions (pH = 7.2, CTAB constant concentration 40 mM) (unpublished data).

Figure 11

Structures of block copolymers forming micelles discussed in the current review.

Figure 12

(A) ¹H ¹H-NOESY spectrum of Ce4–KP in phosphate buffered saline (PBS, molar ratio 3:10); intramolecular NOE cross peaks are visible for the Ce4 meso proton resonances (marked by the blue square) and intermolecular NOEs are visible between the Ce4 meso resonances and the methyl resonance of KP (highlighted in red and indicated in the structures); (B) overlay of ¹H-DOSY spectra of 3 mM Ce4 in DMSO (shown in black), 10 mM KP in PBS (shown in blue), and Ce4–KP in PBS at a molar ratio of 3:10 (shown in red). For Ce4–KP (3:10) the DOSY spectrum and the projection spectrum are scaled up by a factor of 64 in the region between 7 and 11 ppm. T = 310 K. (Reprinted from Gjuroski et al. Journal of Controlled Release, 316, (2019), 150-167 [143], © 2019 Published by Elsevier B.V., with permission from Elsevier).