**3. Uv-vis spectra of porphyrins**

90 Macro to Nano Spectroscopy

Fig. 4. Two-step one-flask room-temperature synthesis of porphyrins.

and meso-position determine the water or solvent solubility of porphyrins.

Fig. 5. Porphyrin numeration.

Its structure supports a highly stable configuration of single and double bonds with aromatic characteristics that permit the electrophilic substitution reactions typical of aromatic compounds such as halogenation, nitration, sulphonation, acylation, deuteration, formylation. Although this, in the porphyrins there are two different sites on the macrocycle where electrophylic substitution can take place with different reactivity (Milgrom, 1997): positions 5, 10, 15 e 20, called *meso* and positions 2, 3, 7, 8, 12, 13, 17 and 18, called -pyrrole positions (Fig. 5). The first kind of compounds are widely present in natural products, while the second have no counterpart in nature and were developed as functional artificial models. The activation of these sites depends of the porphyrins electronegativity that can be controlled by the choice of the metal to coordinate to the central nitrogen atoms. For this, the introduction of divalent central metals produces electronegative porphyrin ligands and these complexes can be substituted on their *meso*-carbon. On the other hand, metal ions in electrophylic oxidation states (e.g. Sn IV) tend to deactivate the *meso*-position and activate the pirrole to electrophylic attack. The chemical characteristics of substituents in -pyrrole It was recognized early that the intensity and colour of porphyrins are derived from the highly conjugated -electron systems and the most fascinating feature of porphyrins is their characteristic UV-visible spectra that consist of two distinct region regions: in the near ultraviolet and in the visible region (Fig. 6).

It has been well documented that changes in the conjugation pathway and symmetry of a porphyrin can affect its UV/Vis absorption spectrum (Gouterman, 1961; Whitten et al. 1968; Smith, 1976; Dolphin, 1978; Nappa & Valentine, 1978; Wang et al. 1984; Rubio et al. 1999).

The absorption spectrum of porphyrins has long been understood in terms of the highly successful "four-orbital" (two highest occupied orbitals and two lowest unoccupied \* orbitals) model first applied in 1959 by Martin Gouterman that has discussed the importance of charge localization on electronic spectroscopic properties and has proposed the fourorbital model in the 1960s to explain the absorption spectra of porphyrins (Gouterman, 1959; Gouterman, 1961).

Fig. 6. UV-vis spectrum of porphyrin with in insert the enlargement of Q region between 480-720 nm.

According to this theory, as reported in Figure 7, the absorption bands in porphyrin systems arise from transitions between two HOMOs and two LUMOs, and it is the identities of the metal center and the substituents on the ring that affect the relative energies of these transitions. The HOMOs were calculated to be an a1u and an a2u orbital, while the LUMOs were calculated to be a degenerate set of eg orbitals. Transitions between these orbitals gave rise to two excited states. Orbital mixing splits these two states in energy, creating a higher energy state with greater oscillator strength, giving rise to the Soret band, and a lower energy state with less oscillator strength, giving rise to the Q-bands.

The electronic absorption spectrum of a typical porphyrin (Fig. 6) consists therefore of two distinct regions. The first involve the transition from the ground state to the second excited state (S0 S2) and the corresponding band is called the Soret or B band. The range of

The Use of Spectrophotometry UV-Vis for the Study of Porphyrins 93

When porphyrinic macrocycle is protonated or coordinated with any metal, there is a more symmetrical situation than in the porphyrin free base and this produces a simplification of

Neglecting the overall charge of the macrocycle, a monomeric free-base porphyrin H2-P in aqueous solution can add protons to produce mono H3-P+ and dications H4-P2+ at very low pHs, or loose protons to form the centrally monoprotic H-P- at pH about 6 or aprotic P2 species at pH ≥ 10 (Fig. 8). These chemical forms of porphyrin may exist in equilibrium, depending upon the pH of the solution and can be characterized from the change of the electronic absorption spectrum. The change in spectra upon addition of acid or basic substances can generally be attributed to the attachment or the loss of protons to the two imino nitrogen atoms of the pyrrolenine-like ring in the free-base (Gouterman, 1979; Giovannetti et al, 2010). The N-protonation induced a red-shifts that are consistent with

frontier molecular orbital calculations for protonated porphyrins (Daniel et al., 1996).

Fig. 8. Typical Uv-vis specrtum of dianion P2- (pH about 10) monoprotic H-P-

Spectrophotometric titration was employed for determining the acid dissociation constants over the inter pH range and change in absorbance with pH can be attributed to the following acid dissociation reactions of porphyrins. Upon addition of acid the spectral pattern of porphyrins changes from the four Q-band spectrum, indicating *D*2h symmetry for free-base porphine, to a two Q-band spectrum for the formation of dications H4-P2+ (Fig. 8 c), indicating *D*4h symmetry, characteristic of porphyrin coordinated to a metal ion through the

and dication H4-P2+ porphyrin (pH about 1).

(pH about 6)

Q bands pattern for the formation of two Q bands.

**4. The equilibrium of porphyrins** 

absorption is between 380-500 nm depending on whether the porphyrin is - or *meso*substituted. The second region consists of a weak transition to the first excited state (S0 S1) in the range between 500-750 nm (the Q bands). These favourable spectroscopic features of porphyrins are due to the conjugation of 18 - electrons and provide the advantage of easy and precise monitoring of guest-binding processes by UV-visible spectroscopic methods (Yang et al. 2002; Gulino et al., 2005; Di Natale et al. 2000; Paolesse & D'Amico, 2007) CD, (Scolaro et al. 2004; Balaz et al. , 2005) fluorescence, (Zhang et al., 2004; Zhou et al., 2006) and NMR spectroscopy (Shundo et al., 2009; Tong et al., 1999).

Fig. 7. Porphyrin HOMOs and LUMOs. (A) Representation of the four Gouterman orbitals in porphyrins. (B) Drawing of the energy levels of the four Gouterman orbitals upon symmetry lowering from *D*4*h* to *C*2V. The set of eg orbitals gives rise to Q and B bands.

The relative intensity of Q bands is due to the kind and the position of substituents on the macrocycle ring. Basing on this latter consideration, porphyrins could be classified as *etio*, *rhodo*, *oxo-rhodo* e *phyllo* (Prins et al 2001).

When the relative intensities of Q bands are such that IV > III > II > I, the spectrum is said *etio-type* and porphyrins called *etioporphyrins*. This kind of spectrum is found in all porphyrins in which six or more of the -positions are substituted with groups without electrons, *e.g.*, alkyl groups. Substituent with -electrons, as carbonyl or vinyl groups, attached directly to the -positions gave a change in the relative intensities of the Q bands, such that III > IV > II > I. This is called *rhodo-type* spectrum (*rhodoporphyrin*) because these groups have a "reddening" effect on the spectrum by shifting it to longer wavelengths. However, when these groups are on opposite pyrrole units, the reddening is intensified to give an *oxo-rhodo-type* spectrum in which III > II > IV > I. On the other hand, when *meso*positions are occupied, the *phyllo-type* spectrum is obtained, in which the intensity of Q bands is IV > II > III > I (Milgron 1997).

While variations of the peripheral substituents on the porphyrin ring often cause minor changes to the intensity and wavelength of the absorption features, protonation of two of the inner nitrogen atoms or the insertion/change of metal atoms into the macrocycle usually strongly change the visible absorption spectrum.

When porphyrinic macrocycle is protonated or coordinated with any metal, there is a more symmetrical situation than in the porphyrin free base and this produces a simplification of Q bands pattern for the formation of two Q bands.
