Structural Modulation of Chromic Response: Effects of Binding-Site Blocking in a Conjugated Calix[4]pyrrole Chromophore.

Herein, we modulate the chromic response of a highly colored tetrapyrrole macrocycle, namely, tetrakis(3,5-di-tert-butyl-4-oxocyclohexadien-2,5-yl)porphyrinogen (OxP) by structural modification. N-Benzylation at the macrocyclic nitrogen atoms leads to stepwise elimination of the two calix[4]pyrrole-type binding sites of OxP and serial variation of the chromic properties of the products, double N-benzylated Bz2OxP and tetra N-benzylated Bz4OxP. The halochromic (response to acidity) and solvatochromic (response to solvent polarity) properties were studied by using UV/Vis spectroscopy and NMR spectroscopy in nonpolar organic solvents. Titration experiments were used to generate binding isotherms to elucidate their binding properties with difluoroacetic acid. Differences in the halochromic properties of the compounds allowed construction of a colorimetric scale of acidity in nonpolar solvents, as the compounds in the series OxP, Bz2OxP, and Bz4OxP are increasingly difficult to protonate but maintain their propensity to change color upon protonation. The concurrent effects of binding-site blocking and modulation of acidity sensitivity are important new aspects for the development of colorimetric indicators.

Introduction

Variation in the color of a dye or pigment due to changes in its immediate environment is referred to as chromism. It is a key feature that has led to sensing applications, perhaps most importantly involving acid–base equilibria1, 2 (e.g. halochromism of pH indicators). It has also been exploited in the delineation of solvent polarity (i.e. solvatochromism) and for the construction of polarity scales such as the ET(30)3 and Z scales.4, 5 Several other chromic phenomena exist for which organic pigment molecules change color in response to an applied stimulus, including light,6 pressure,7 and temperature.8 Chromic events are typically reversible adding to their attractiveness for various applications. For example, acid–n class="Chemical">base reactions can be monitored according to reversible color changes that occur during forward and backward titration procedures of usually colorless analytes. Classically, many analytical procedures that rely on halochromism have been performed in aqueous solutions, although there are several reported methods applicable in nonpolar solvents9, 10, 11 sometimes involving visualization of an acid–base reaction by using a dye of appropriate structure.

The term halochromism (sometimes acidochromism), originally coined by Baeyer12 and recently resurrected by Reichardt,13 describes color changes that some dyes undergo in response to interactions with acid, base, or salt ions. Halochromic molecules (e.g. n class="Chemical">phenolphthalein, a component of Universal Indicator)14 interact with ions or are protonated (deprotonated) by acids (bases), which is usually accompanied by the appearance of new electronic absorption bands caused by variation in the conjugated structure of the halochromic dye. Color variation results from the different relative fractions of the halochromic dye in its different states of electronic conjugation. In contrast, solvatochromism occurs if the electronic ground and excited states of a chromophore are stabilized to different extents by solvents of differing polarities. This affects the energy difference between the ground and excited states, which results in shifts in the electronic absorption maxima (hypso‐ or bathochromic shift). A typical solvatochromic substance is Reichardt's dye,13 the behavior of which is the basis for the ET(30) polarity scale. Despite its impressive solvatochromic properties, Reichardt's dye loses its solvatochromism upon protonation, albeit reversibly.13, 15

Tetrapyrrole macrocycles compose a widely studied class of organic molecules that includes the n class="Chemical">porphyrins and other highly colored pigments known for their incorporation into biological photosynthetic and catalytic systems.16 Free‐base porphyrins (i.e. those not containing a central chelated metal cation) are known to exhibit chromic properties, including halochromism, due to protonation at the central imino nitrogen atoms,17, 18, 19 and aggregachromism20, 21 (due to excitonic coupling interactions in stacked aggregates). Although typical free‐base porphyrins (e.g. meso‐tetraphenylporphyrin) exhibit strong coloration and their electronic structures have been intensively investigated,22 it is perhaps surprising that they have not been studied in depth for their solvatochromic properties. This is because, notwithstanding their chromism in acidic media (halochromism), they tend to exhibit only small hypso‐ and bathochromic shifts in their absorption maxima in different solvents, leading to only weak variation in hue.23 Metalloporphyrins can exhibit stronger solvatochromism due to interactions between chelated metal cations and a polar coordinating solvent.24, 25 Furthermore, structural changes (e.g. N‐confusion)26, 27 have been found to lead to significant solvatochromism, whereas the inherent synthetic flexibility of porphyrins can be used to design more complicated solvatochromic systems.28 Thus, although free‐base porphyrins themselves have limited appeal as solvatochromic dyes, there are several classes of more complex derivatives, including N‐confused porphyrins, that fulfil the requirements of strong color variation simply on the basis of solvent polarity.

In previous work,9 we used an oxoporphyrinogen dye (i.e. n class="Chemical">OxP, a conjugated calix[4]pyrrole,29, 30, 31, 32, 33 Figure 1 a) to monitor acid–base reactions in nonpolar solvents and found that it could be incorporated into dye films for application as an acidity indicator by similar means to those applied for Universal Indicator. The OxP molecular structure can be variously modified by alkylation at its macrocycle core nitrogen atoms (Figure 1),34, 35, 36 and we were intrigued to assess the influence of N‐alkylation on the chromic properties of the OxP chromophore, in particular, in relation to a perceived reduction in flexibility of the macrocycle caused by steric crowding at its core. We chose the simple N21,N23‐bis(4‐bromobenzyl) (Bz, Figure 1 b) and N21,N22,N23,N24‐tetrakis(4‐bromobenzyl) (Bz, Figure 1 c) derivatives of OxP (4‐bromobenzyl derivatives are more soluble than the corresponding non‐brominated derivatives). Differences in their substitution‐dependent geometries as obtained from X‐ray crystallography are shown in Figure 1.

Figure 1

Chemical structures (left) and X‐ray crystal structures (right) of a) oxoporphyrinogen (OxP), b) bis(4‐bromobenzyl)oxoporphyrinogen (Bz), c) tetrakis(4‐bromobenzyl)oxoporphyrinogen (Bz) used in this work. Dihedral angles between opposing pyrrole groups are shown. X‐ray crystal structure in panels b, c are of the non‐brominated derivatives.[37] Accessible calix[4]pyrrole‐type binding sites are denoted by arrows.

Chemical structures (left) and X‐ray crystal structures (right) of a) oxoporphyrinogen (n class="Chemical">OxP), b) bis(4‐bromobenzyl)oxoporphyrinogen (Bz), c) tetrakis(4‐bromobenzyl)oxoporphyrinogen (Bz) used in this work. Dihedral angles between opposing pyrrole groups are shown. X‐ray crystal structure in panels b, c are of the non‐brominated derivatives.[37] Accessible calix[4]pyrrole‐type binding sites are denoted by arrows.

Herein, we study protonation and the related chromic behavior (i.e. halochromism) of these OxP derivatives in nonpolar solvent/acid mixtures. We show that N‐alkylation of n class="Chemical">OxP at its core nitrogen atoms substantially modulates chromic sensitivity to acid content over a broad range of volume fractions (vol. f. or ϕ) from 10−7 (in the case of OxP) to nearly 1 (for Bz). We also investigate the solvatochromic properties of OxP derivatives in polar/nonpolar solvent mixtures in the absence of acid. Halo‐ and solvatochromic effects are rationalized in terms of stepwise protonations and solvating interactions, respectively.

Results and Discussion

Halochromism of OxP

Previously, we reported the colorimetric response of OxP to n class="Chemical">trifluoroacetic acid (TFA),9 for which color variation upon the addition of acid was caused by the presence of di‐ and tetraprotonated species. In that case, fourfold protonation at the hemiquinonoid carbonyl sites was proven by direct observation of the intensity of the C−O−H infrared vibration. Here, we reanalyzed the UV/Vis data from that study from the point of view of its halochromic and solvatochromic properties. Spectra and analyses are shown in Figure 2. Singular value decomposition38, 39, 40, 41 (SVD) of the spectra indicates the presence of three absorbing species (Figure S1 in the Supporting Information, for details about the implementation of SVD, see the Supporting Information), which is confirmed by the presence of isosbestic points in both consecutive sets of spectral changes (Figure 2 a, b). The spectra of di‐ and tetraprotonated species (Figure 2 c) were obtained, respectively, as the endpoints of each set of spectral changes (Figure 2 a, b). We find that all experimentally observed UV/Vis spectra S obs(ϕ) are essentially simple linear combinations of spectra of the three individual absorbing species, OxP, OxP, and OxP, with acid volume fraction (ϕ) dependent coefficients, which can be expressed by the following equation [Eq. (1)]:

Figure 2

UV/Vis titration of OxP (10−5  m in CH2Cl2) with TFA on the basis of the data of Shundo et al.9 a, b) First and second sets of spectral changes occurring during titration. c) Spectra of individual absorbing species identified as the start and endpoints of each spectral change (bold lines in panels a, b). d) Relative fractions of absorbing species obtained from SVD by fitting to a 1:4 binding model (solid line); coefficients obtained by DSIS decomposition (solid circles). The same data in linear scale can be seen in Figure S14 a. e) Chemical structures and RGB colors of absorbing species (colors calculated from the spectra shown in panel c). The RGB coordinates are: OxP (R=177, G=92, B=170), OxP (R=212, G=152, B=166), and OxP (R=151, G=186, B=160). Assignment of the chemical structures (including tautomeric processes) is given in Figure 11.

UV/Vis titration of OxP (10−5  m in n class="Chemical">CH2Cl2) with TFA on the basis of the data of Shundo et al.9 a, b) First and second sets of spectral changes occurring during titration. c) Spectra of individual absorbing species identified as the start and endpoints of each spectral change (bold lines in panels a, b). d) Relative fractions of absorbing species obtained from SVD by fitting to a 1:4 binding model (solid line); coefficients obtained by DSIS decomposition (solid circles). The same data in linear scale can be seen in Figure S14 a. e) Chemical structures and RGB colors of absorbing species (colors calculated from the spectra shown in panel c). The RGB coordinates are: OxP (R=177, G=92, B=170), OxP (R=212, G=152, B=166), and OxP (R=151, G=186, B=160). Assignment of the chemical structures (including tautomeric processes) is given in Figure 11.

Figure 11

Redistribution of charge upon protonation for a) OxP and b–d) Bz.

in which S OxP, S , and S are the spectra of the individual species n class="Chemical">OxP, OxP, and OxP, respectively, and f OxP(ϕ), f (ϕ), and f (ϕ) are the relative molar fractions of the corresponding individual species with constraints of non‐negativity and obeying the unity condition, f OxP(ϕ)+f (ϕ)+f (ϕ)=1. We refer to this spectral separation process as “decomposition into the spectra of individual species” (DSIS). The result of DSIS is shown in Figure 2 d (solid circles), for which the f •(ϕ) (subscript “•” denotes any individual species) values are plotted as a function of ϕ. The quality of DSIS is excellent, as seen in Figure S2. In the presence of a solvatochromic shift, decomposition on the basis of Equation (1) is not possible (e.g. for spectra of Reichardt's dye, see Figures S12 and S13). Therefore, there is no observable solvatochromic shift for OxP under these conditions, and color variation is caused by the presence of a few different conjugated species having different UV/Vis spectra. Thus, variation of the acid volume fraction ϕ affects the relative fraction of individual absorbing species but does not influence excitation energy levels, and OxP exhibits typical halochromism in TFA/CH2Cl2 solvent mixtures.

Subsequently, we attempted to describe the interaction of OxP with n class="Chemical">TFA in terms of a host–guest binding model (in the frame of SVD, see Section S2 in the Supporting Information). Saturation of the OxP color changes is reached at low concentrations of TFA (0.002 vol. f. in CH2Cl2) so that equations governing the chemical equilibria can be used without corrections for activity. A 1:4 OxP:TFA stoichiometry binding model (for details about derivation of the binding isotherms see Section S1 in the Supporting Information) was adopted to obtain the equilibrium binding constants listed in Table 1. The resulting dependence (on ϕ) of the relative fractions of individual species is shown in Figure 2 d (solid lines). The very good match of the fit can be seen in Figures S1 d and S2. The diprotonated form is stabilized by binding of two counteranions (on the top and on the bottom) at the central NH groups and by a tautomeric process.42 The excellent quality of the data fit also indicates the low stability of any mono‐ and triprotonated OxP species, which are essentially absent due to the ease of further protonation (analogous with tetraphenylporphyrin monocation43, 44, 45). On the basis of this analysis the chemical structures including the changes in conjugation of the individual species are given in Figure 2 e.

Table 1

Stepwise binding constants[a] (decadic logarithm of numeric values of K n are shown; all K n have unit m −1) and corresponding cooperativities[b] determined by fitting of the NMR and UV/Vis titration experiments.

	DFA/TFA		Camphorsulfonic acid
	NMR	UV/Vis	NMR	UV/Vis
	OxP
log K 1	4.5–8.7[c]	2.3–6.0[d]	4.0–5.0	N/A
log K 1 K 2	8.0‐16[c]	13–15[d]	10–12	N/A
log K 3	2.5–4.0[c]	<1.0[d]	<5.0	N/A
log K 3 K 4	4.0–6.0[c]	3.6–5.0[d]	2.3–2.5	N/A
log α 12	—	6.2[d]	—	N/A
log α 34	—	>2.2[d]	—	N/A

[a] Binding constants for processes at higher acid vol. f. could not be obtained because of the solubility threshold of camphorsulfonic acid and the unknown activity coefficients of the species in solution. Due to the nature of the binding processes and the models used, only the range of the binding constants could be determined. Also, due to the high cooperativity, overall binding constants (i.e. K 1 K 2, K 3 K 4) were used instead of stepwise. Details about the binding models used are shown in the Supporting Information. [b] Cooperativity α 12=4 K 2/K 1 was determined at the geometric mean of the minimum and maximum possible values of the binding constant, K 1. Cooperativity α 34 could only be restricted by using the formula α 34>4 (K 3 K 4)min/K 3 max 2 (where subscripts min and max indicate minimum value of K 3 K 4 product and maximum value of K 3, respectively), as there was no lower boundary for K 3. Only UV/Vis measurements were used due to higher accuracy. [c] Measured with DFA. [d] Measured with TFA.

[a] Binding constants for processes at higher acid vol. f. could not be obtained because of the solubility threshold of camphorsulfonic acid and the unknown activity coefficients of the species in solution. Due to the nature of the binding processes and the models used, only the range of the binding constants could be determined. Also, due to the high cooperativity, overall binding constants (i.e. K 1 K 2, K 3 K 4) were used instead of stepwise. Details about the binding models used are shown in the Supporting Information. [b] Cooperativity α 12=4 K 2/K 1 was determined at the geometric mean of the minimum and maximum possible values of the binding constant, K 1. Cooperativity α 34 could only be restricted by using the formula α 34>4 (K 3 K 4)min/K 3 max 2 (where subscripts min and max indicate minimum value of K 3 K 4 product and maximum value of K 3, respectively), as there was no lower boundary for K 3. Only UV/Vis measurements were used due to higher accuracy. [c] Measured with n class="Chemical">DFA. [d] Measured with TFA.

Halochromism of Bz2OxP

Bz differs from OxP in that only single n class="Chemical">counteranion stabilization is allowed due to its double N‐alkylation and consequent effective blocking of one of the two binding sites (Figure 1 b). For this reason, a stable monoprotonated Bz form is readily formed. UV/Vis titration experiments of Bz with difluoroacetic acid (DFA), which was used because of its nonexchanging hydrogen utilized in NMR spectroscopy analysis, reveal three consecutive spectral changes (Figure 3 a–c) and consequently four individual species (confirmed by SVD, see Figure S3). These have clear pseudosaturation points so that the spectra of the individual protonated species can be easily extracted at the respective endpoints (Figure 3 d). Assignment of the two‐ and fourfold protonation products as individual spectra is based on their significant similarities to S and S (see Figure 4 c, d). Assignment of the UV/Vis spectrum due to Bz is supported by fitting of its binding constant (Table 1) and by NMR spectroscopy analysis (see Figure 5 and Figures S17 and S18). Similar to the OxP case, the linear combination of these four absorption spectra of the individual species accounts for all other experimentally observed spectra. This can be expressed in the following equation in which the terms have analogous meanings as in Equation (1) [Eq. (2)]:

Figure 3

UV/Vis titration of Bz (7×10−6  m in CDCl3) with DFA. a–c) Three consecutive spectral changes occurring during titration. d) Spectra of individual absorbing species identified as start and endpoints of each spectral change (bold lines in panels a–c). e) Relative fraction of absorbing species obtained from SVD by fitting first spectral change with a 1:1 binding model (solid line); coefficients obtained by DSIS decomposition (solid circles). The same data in linear scale can be seen in Figure S14 b. f) Chemical structures and colors of absorbing species (colors calculated from the spectra shown in panel d). The RGB coordinates are: Bz (R=217, G=129, B=171), Bz (R=204, G=137, B=197), Bz (R=157, G=169, B=206), and Bz (R=147, G=196, B=192). Assignment of the chemical structures (including tautomeric processes) is given in Figure 11.

Figure 4

Spectra of neutral and protonated species of OxP, Bz, and Bz in the a) neutral state and in the b) monoprotonated, c) diprotonated, and d) tetraprotonated forms. Spectra of neutral species were normalized to unity.

Figure 5

Demonstration of rotation of the Bz hemiquinonoid groups due to increasing hydroxyphenyl character after protonation. a) There is no spectral change in the tert‐butyl NMR signals upon varying the temperature in the absence of acid. b) Upon protonation, chemical exchange between both tert‐butyl signals arises. c) The most plausible explanation is rotation of the whole meso‐substituent group.

UV/Vis titration of Bz (7×10−6  m in CDCl3) with n class="Chemical">DFA. a–c) Three consecutive spectral changes occurring during titration. d) Spectra of individual absorbing species identified as start and endpoints of each spectral change (bold lines in panels a–c). e) Relative fraction of absorbing species obtained from SVD by fitting first spectral change with a 1:1 binding model (solid line); coefficients obtained by DSIS decomposition (solid circles). The same data in linear scale can be seen in Figure S14 b. f) Chemical structures and colors of absorbing species (colors calculated from the spectra shown in panel d). The RGB coordinates are: Bz (R=217, G=129, B=171), Bz (R=204, G=137, B=197), Bz (R=157, G=169, B=206), and Bz (R=147, G=196, B=192). Assignment of the chemical structures (including tautomeric processes) is given in Figure 11.

Spectra of neutral and protonated species of OxP, Bz, and Bz in the a) neutral state and in the b) monoprotonated, c) diprotonated, and d) n class="Species">tetraprotonated forms. Spectra of neutral species were normalized to unity.

The results of DSIS decomposition are shown in Figure 3 e (solid circles), in which f •(ϕ) (subscript “•” denotes any individual species) values are plotted as a function of ϕ. The excellent match of DSIS (Figure S4) confirms that Bz is also a halochromic dye.

We again attempted to describe the interactions of Bz with DFA in terms of a host–guest binding model. The first spectral change (up to vol. f. ϕ=0.001 of n class="Chemical">DFA) was fitted by using a 1:1 Bz:DFA binding model (Figure 3 e, solid lines), see Table 1. Details of the fitting procedure can be found in Figures S3 d and S4. Quantities of acid greater than vol. f. ϕ=0.001 change the polarity of the solvent so that activity coefficients cannot be approximated as concentrations. Consequently, a simple binding model is not applicable for higher protonated states of Bz beyond monoprotonation. NMR spectroscopy confirms protonation at the carbonyl oxygen atoms of the hemiquinonoid groups, and this leads to their increasing hydroxyphenyl character. This enables rotation of these groups about their meso‐position bonds due to increased single‐bond character, as can be seen in Figure 5. In addition, the phenolic type OH group around δ=6.4 ppm can also be observed in Figures S17, S18, and S22 b. The chemical identities of the individual species and their respective colors are given in Figure 3 f.

Chromism of Bz4OxP

Of the compounds studied, Bz is the least susceptible to protonation due to complete N‐alkylation and consequent blockage of both central binding sites so that no potentially stabilizing ion‐pair interactions with counteranions are allowed. UV/Vis titration with n class="Chemical">DFA (Figure 6 a, b) shows three sets of spectral changes. The first one (up to ϕ=0.01) is a slight bathochromic shift due to hydrogen bonding of acid molecules to the carbonyl oxygen atoms (Figure 6 a, green arrow) without significant variation in the shape of the spectrum, which was not observed in the previous cases. This effect is attributed to solvatochromism. The corresponding solvatochromic shift in the absorbance maximum is less than 10 nm (Figure 6 e). We already showed that hydrogen bonding with water in tetrahydrofuran was accompanied by a similar spectral variation in the case of both di‐ and tetra‐N‐substituted oxoporphyrinogens.46 We ascribe the second and third sets of spectral changes (Figure 6 a, b) to protonation at the hemiquinonoid carbonyl atoms. We could again easily obtain spectra of mono‐ and diprotonated species as pseudosaturation points during titration (Figure 6 c). Assignments were made on the basis of comparisons with individual species of the other oxoporphyrinogens (Figure 4 b, c). For vol. f. ϕ>0.01 (i.e. halochromic behavior), we used a formula similar to that used in the previous cases for DSIS decomposition [Eq. (3)]:

Figure 6

UV/Vis titration of Bz (5×10−6  m in CDCl3) with DFA. a, b) Three consecutive spectral changes occurring during titration. c) Spectra of individual absorbing species identified as start and endpoints of each spectral change (bold lines in panels a, b). d) Relative fraction coefficients obtained by DSIS. The same data in linear scale can be seen in Figure S14 c. e) Solvatochromic shift of absorbance maxima for ϕ<0.01 (hatched area in panel d). f) Chemical structures and colors of absorbing species (colors calculated from the spectra shown in panel c). The RGB coordinates are: Bz (R=244, G=165, B=173), Bz H‐bond (R=225, G=135, B=184), Bz (R=195, G=168, B=194), and Bz (R=121, G=195, B=213). Assignment of the chemical structures (including tautomeric processes) is given in Figure 11.

UV/Vis titration of Bz (5×10−6  m in CDCl3) with n class="Chemical">DFA. a, b) Three consecutive spectral changes occurring during titration. c) Spectra of individual absorbing species identified as start and endpoints of each spectral change (bold lines in panels a, b). d) Relative fraction coefficients obtained by DSIS. The same data in linear scale can be seen in Figure S14 c. e) Solvatochromic shift of absorbance maxima for ϕ<0.01 (hatched area in panel d). f) Chemical structures and colors of absorbing species (colors calculated from the spectra shown in panel c). The RGB coordinates are: Bz (R=244, G=165, B=173), Bz H‐bond (R=225, G=135, B=184), Bz (R=195, G=168, B=194), and Bz (R=121, G=195, B=213). Assignment of the chemical structures (including tautomeric processes) is given in Figure 11.

Thus, for ϕ>0.01, we obtained relative fractions of individual absorbing species (see Figure 6 d). As in the previous cases, the quality of DSIS is excellent (Figure S6). SVD performed for ϕ>0.01 confirmed the presence of three individual species in this concentration range (Figure S5). NMR titration of Bz with n class="Chemical">DFA reveals a significant shift in the pyrrole β‐proton resonance (Figure 7 and for chemical shifts as a function of acid volume fraction, see Figure S19c), which strongly suggests that positive charges are accommodated at the central nitrogen atoms of Bz (Figure 6 f).

Figure 7

Titration of Bz (8×10−4  m in CDCl3) with DFA. Asterisk (*) denotes impurities contained in acid. Some regions of the spectra are magnified for clarity. The factor of magnification is given above each region. See Figure S19 for the chemical shifts of the respective resonances during the titration.

Titration of Bz (8×10−4  m in CDCl3) with n class="Chemical">DFA. Asterisk (*) denotes impurities contained in acid. Some regions of the spectra are magnified for clarity. The factor of magnification is given above each region. See Figure S19 for the chemical shifts of the respective resonances during the titration.

As shown in Figure 4, the spectra of the corresponding mono‐, di‐, and tetraprotonated species of each n class="Chemical">OxP derivative are similar in form, which we exploited to assign the degrees of protonation of each absorbing species. According to Table 1, OxP is easy to protonate twice and is stabilized by complexation through H‐bonding of two counteranions at the central NH sites. Tetraprotonation is attained at about vol. f. ϕ=0.001 of TFA. There exists strong positive cooperativity between the first and second protonation steps and between the third and fourth protonation steps (Table 1), so that spectra due to mono‐ and triprotonated states are not observable (similar to the case for double protonation of free‐base porphyrins).43 Bz can be easily monoprotonated because of stabilization by one counteranion at the NH site. Full tetraprotonation is not reached until the vol. f. of acid is close to 1. Similarly, the presence of strong cooperativity between the third and fourth protonations prevents direct observation of a triprotonated state during the UV/Vis titration experiment. Finally, Bz is very difficult to protonate partly due to the lack of possible stabilization by counteranion binding and partly to steric crowding at the core of the molecule, which obstructs redistribution of the conjugation pathway for sharing of charge over electronegative heteroatoms (i.e. N and O, see Figure 1 c). Complete protonation of Bz is not possible even in neat acid. If we consider all the data relating to halochromism of these OxP derivatives, it is evident that N‐alkylation makes protonation more difficult and concurrently shifts the range of colorimetric sensitivity to higher acid vol. f. To illustrate this potentially useful point, an overview of the color variation of these OxP derivatives is shown in Figure 8.

Figure 8

Colorimetric response of OxP (with TFA), Bz, and Bz (with DFA) over the entire concentration range of the acids. Colors shown were obtained by interpolation from the actual measured spectra (Figures 2, 3, and 6), and the scales are overlaid with photographs of the sample cuvettes (from the titration experiments) at the appropriate vol. f.

Colorimetric response of OxP (with n class="Chemical">TFA), Bz, and Bz (with DFA) over the entire concentration range of the acids. Colors shown were obtained by interpolation from the actual measured spectra (Figures 2, 3, and 6), and the scales are overlaid with photographs of the sample cuvettes (from the titration experiments) at the appropriate vol. f.

Mechanism of Halochromism

The OxP halochromic dye system is rather unique from the point of view of its mode of action. Most simple dye systems operate, whether solvatochromic or halochromic, on the basis of solvent polarity or protonation by mechanisms mentioned earlier, respectively. Conceptually, exceptions can be made on the basis of molecular design usually involving introduction of some complexing moiety to introduce specificity of analyte interaction to the dye.13, 47 In this context, for n class="Chemical">OxP, halochromicity is strongly affected by the presence of the π‐conjugated calix[4]pyrrole‐type binding site, which introduces the possibility of counteranion binding and consequent stabilization of protonated species through ion‐pair formation.

In the series OxP, Bz, and Bz, N‐alkylation of the n class="Chemical">pyrrole groups effectively sequentially masks the binding site(s), disfavoring protonation by removing one source of stabilization (counteranion binding) and also by sterically restricting variation in the conjugated structure of the chromophore, which might otherwise stabilize protonated species. In fact, delocalization of charge in the OxP system may be one of the factors allowing the existence of the highly charged tetracation. In Figures 2 e, 3 f, and 6 f, we have assigned the structures on the basis of formal placement of charge at the central nitrogen atoms. For lower degrees of protonation, these charges may be shared over all electronegative atoms, albeit with preference for the central nitrogen atoms. This feature is supported by NMR spectroscopy (Figures 9 and 10; for other acid titrations, see Figures S16–18), which shows that pyrrole proton resonances are strongly shifted downfield. The UV/Vis spectra offer further information about the state of the dyes under acidic conditions. For example, for OxP, double protonation results in red solutions due to tautomerization of the molecule to a porphodimethene form (Figure 11 a; also supported by NMR spectroscopy, see Figures 9 and S16), resulting in an apparent splitting of the broad OxP absorption band.48 This is due to the emergence of conjugated phenolic (at λ=465 nm) and quinonoid (at λ=540 nm) π–π* absorptions (Figure 4 c). These bands indicate the relative presence of these two types of moieties in the respective structures. For doubly protonated OxP, which has been forced to exist in its tautomeric form by protonation, the intensities of these bands correspond to the abundance of the phenol and quinonoid groups. Tetracationic species of OxP and Bz are notable in that they have only weak contributions from the quinonoid band with prevalence of phenolic groups (Figure 4 d). On the other hand, Bz has only a very weak contribution of the λ=465 nm band even in neat acid (Figure 4 c). Thus, the effect of N‐alkylation is twofold: first it masks the binding site, which prevents counteranion binding; second, it effectively prevents reorganization of the chromophore through tautomerization even under conditions of high acidity. Examples of how delocalization might be depicted are shown for some of the compounds in Figure 11. As shown in Figure 11 a, doubly protonated OxP gains stabilization through tautomerization (in addition to the stabilization it gains from anion‐pair interactions). Figure 11 b shows how the positive charge introduced by protonation at the oxygen atom of Bz is transmitted to a macrocycle nitrogen atom. Figure 11 c, d reveals that charge residing on the central nitrogen atoms can be effectively delocalized.

Figure 9

Titration of OxP (0.002 m in CDCl3) with DFA. Free and protonated/complexed species are denoted by full and empty symbols, respectively. Red dotted arrows denote splitting of tert‐butyl signal due to symmetry breaking after protonation and anion binding. Some regions of the spectra are magnified for clarity. The factor of magnification is given above each region. See Figure S19 for the chemical shifts of the respective resonances during the titration.

Figure 10

Titration of Bz (7×10−4  m in CDCl3) with DFA. Free and protonated/complexed species are denoted by full and empty symbols, respectively. Asterisk (*) denotes impurities contained in acid. Some regions of the spectra are magnified for clarity. The factor of magnification is given above each region. See Figure S19 for the chemical shifts of the respective resonances during the titration.

Titration of OxP (0.002 m in n class="Chemical">CDCl3) with DFA. Free and protonated/complexed species are denoted by full and empty symbols, respectively. Red dotted arrows denote splitting of tert‐butyl signal due to symmetry breaking after protonation and anion binding. Some regions of the spectra are magnified for clarity. The factor of magnification is given above each region. See Figure S19 for the chemical shifts of the respective resonances during the titration.

Titration of Bz (7×10−4  m in CDCl3) with n class="Chemical">DFA. Free and protonated/complexed species are denoted by full and empty symbols, respectively. Asterisk (*) denotes impurities contained in acid. Some regions of the spectra are magnified for clarity. The factor of magnification is given above each region. See Figure S19 for the chemical shifts of the respective resonances during the titration.

The broad, long wavelength band around λ=800 nm in OxP is associated with guest binding at the n class="Chemical">pyrrole NH groups in OxP systems, and for example, it shifts sequentially to λ=740 nm for Bz and then to λ=690 nm for Bz (see Figure 4 c) in their doubly protonated states. We assign this shift to reduction in conjugation due to the increasing non‐coplanarity of the moieties caused by increasing steric hindrance at the macrocyclic core.

Some of the changes caused by protonation (i.e. halochromism) in the OxP series bear analogy with the solvatochromic n class="Chemical">merocyanine dyes (e.g. Brooker's merocyanine),4, 5, 49, 50 for which electron density is distributed over electronegative atoms depending on the prevailing polarity, with zwitterionic and neutral states being stabilized in polar and nonpolar solvents, respectively. For OxP, initial protonation at the carbonyl oxygen atoms leads to a tautomerization‐like process, for which positive charge is relocated and accommodated largely at the macrocyclic nitrogen atoms. Pyrrole nitrogen atoms cannot be directly protonated unless OxP is in its tautomerized porphodimethene form, which contains imino nitrogen atoms (this form can be stabilized by interactions with a polar solvent, see Figure 13 a and discussion below). In contrast, tautomerization in Bz and Bz is precluded by N‐alkyl groups, so that initial protonation must occur at the carbonyl oxygen atom. Protonation‐induced tautomerization in OxP is now a relatively well‐understood phenomenon. However, solvent‐polarity‐induced tautomerization of OxP is not a well‐studied feature.

Figure 13

Suggested structures of individual species of oxoporphyrinogens in chloroform/DMF solution: a) OxP and b) Bz.

Solvatochromic Effects

To examine any solvatochromic properties of the OxP derivatives, we investigated their behavior in n class="Chemical">dimethylformamide (DMF)/chloroform binary mixtures (Figure 12). All three compounds show solvatochromism (similar to the first spectral change for the Bz–DFA interaction, Figure 6 a) manifested as a bathochromic shift, as shown in Figure 12 a, e, g (green arrow). The central NH binding sites of OxP and Bz enable interaction with DMF, resulting in other spectral changes (two changes for OxP and one change for Bz), see Figure 12 b, c, e. Hydrogen bonding between OxP derivatives and solvents including water and carbonyl‐containing solvents (e.g. acetone)34 is known and can be described for OxP and Bz, as shown in Figure 13. For OxP (Figure 12 a), at low vol. f. of DMF, the bathochromic shift is followed at higher vol. f. DMF by a second spectral change, which we associate with a tautomeric process similar to that occurring during its protonation (according to a comparison of Figure 12 b red line with Figure 2 a blue line). As OxP undergoes further spectral changes at increasing volume fractions of DMF, it appears that the balance of polarity and solvent binding is important in the tautomerization of OxP 51 [as seen in Figure 12 b, c, for which the assigned porphodimethene tautomeric form, Figure 13 a (form 2), is only stable around 1.4×10−2 vol. f. of DMF]. The proposed individual absorbing species of OxP are shown in Figure 13 a. Interestingly, the tautomer form of OxP can only accommodate a single DMF guest through H‐bonding at the macrocycle, as the tautomer also possesses imino‐type nitrogen atoms that cannot bind DMF. Phenol groups in the tautomer may H‐bond with DMF, although this interaction ought to be weakened by steric effects of the tert‐butyl groups. At increasing DMF vol. f., the calix[4]pyrrole form is preferred, as binding of two DMF guests stabilizes that form. For Bz, despite some variation in its UV/Vis spectrum during titration with DMF (Figure 12 e), it does not undergo any tautomeric processes denoted by the lack of the absorbance maximum at λ=465 nm (Figure 4 c). We assign the second spectral change (Figure 12 e, red line) to binding of DMF at the macrocyclic core (see Figure 13 b, form 2). Similarly, there are no further spectral changes for Bz from the point of view of tautomerism (Figure 12 g). The solvatochromic shift of the spectral maximum of Bz is shown in Figure S14 f.

Figure 12

Solvatochromism of the investigated OxP derivatives in a binary mixture of chloroform/DMF; spectra taken from zero DMF concentration up to saturation. a–c) DMF‐concentration‐dependent consecutive spectral changes of OxP. d) Isolated spectra of individual species from the starting and endpoint spectra, as obtained from panels a–c. e) Spectral changes of Bz and f) corresponding isolated individual absorption spectra. g) Spectral change of Bz and h) corresponding isolated individual absorption spectra.

Solvatochromism of the investigated OxP derivatives in a binary mixture of n class="Chemical">chloroform/DMF; spectra taken from zero DMF concentration up to saturation. a–c) DMF‐concentration‐dependent consecutive spectral changes of OxP. d) Isolated spectra of individual species from the starting and endpoint spectra, as obtained from panels a–c. e) Spectral changes of Bz and f) corresponding isolated individual absorption spectra. g) Spectral change of Bz and h) corresponding isolated individual absorption spectra.

Suggested structures of individual species of oxoporphyrinogens in n class="Chemical">chloroform/DMF solution: a) OxP and b) Bz.

As in the case of protonation, we were able to isolate spectra of individual absorbing species as endpoints of spectral variations (Figure 12 d, f, h). Although pure solvatochromic changes cannot, in principle, be decomposed by using DSIS (as mentioned before for Reichardt's dye), other spectral changes (tautomeric and binding processes of OxP and Bz) can be analyzed by this method (Figures S7–S10 and Figure S14 d, e in linear scale). An overview of the color variation of the n class="Chemical">OxP derivatives for the whole DMF concentration range can be seen in Figure 14.

Figure 14

Colorimetric response of OxP derivatives over the entire concentration range of DMF. Colors shown were obtained by interpolation from the actual measured spectra (Figure 12), and the scales are overlaid with photographs of sample cuvettes (from the titration experiments) at the appropriate vol. f.

Colorimetric response of OxP derivatives over the entire concentration range of n class="Chemical">DMF. Colors shown were obtained by interpolation from the actual measured spectra (Figure 12), and the scales are overlaid with photographs of sample cuvettes (from the titration experiments) at the appropriate vol. f.

Conclusions

In summary, we present tetrakis(3,5‐di‐tert‐butyl‐4‐oxocyclohexadien‐2,5‐yl)porphyrinogen (n class="Chemical">OxP) and its N21,N23‐bis(4‐bromobenzyl) and N21,N22,N23,N24‐tetrakis(4‐bromobenzyl) derivatives (Bz and Bz, respectively) as a series of halochromic dyes that operate as colorimetric indicators over a very broad range of acid contents in nonpolar media. This was permitted by up to fourfold protonation of the OxP derivatives. The maximum protonation level was tuned by N‐alkylation, which effectively shifted the sensitivity (colorimetric range) to higher acid contents, as protonation of the molecules was increasingly obstructed. This was manifested as a gradual change in color over a broad range of acid contents (from volume fractions of 10−7 to nearly 1) over the series OxP, Bz, and then Bz. The mechanism of operation involves stabilization by counteranion binding, which enables colorimetric detection of small amounts of acid (less than 10−6  m). For Bz, stabilization by a counteranion was not permitted, so that its response was limited to higher acid contents (from volume fractions of 0.1). Our analysis was supported by UV/Vis spectrophotometric measurements, for which the spectra of the individual components were isolated, and also by NMR spectroscopy.

In the case of the series OxP, Bz, and Bz, our molecular design concept was initially n class="Chemical">based on binding‐site blocking and its effect on colorimetric response. It was predicted that the lack of binding sites would lead to a gradual negation of any colorimetric response. However, in the case of acids, we were surprised to discover the retention of response to acid analytes, even in the completely blocked derivative Bz. A further advantage of this OxP‐based indicator system is that the compounds have similar structures and properties, perhaps facilitating their application as blends of OxP derivatives and avoiding any mismatches in, for example, their solubilities. We believe that the concurrent effects of binding‐site blocking and modulation of acidity sensitivity are important new aspects for the development of colorimetric indicators for various applications.

Experimental Section

Chemicals

OxP and its derivatives were prepared by a previously reported method.29, 51, 52 We used n class="Chemical">silver‐foil‐stabilized deuterated chloroform (CDCl3) as the solvent and camphorsulfonic acid in powder form and liquid difluoroacetic acid as reagents (Sigma–Aldrich). Difluoroacetic acid (DFA) was chosen instead of commonly used trifluoroacetic acid (TFA) due to the presence of a nonexchanging proton and its associated 1H NMR signal. Samples containing oxoporphyrinogen derivatives and excess amounts of DFA were stored in the dark to avoid photoinduced chemical changes.

Camphorsulfonic acid (n class="Chemical">CSA) is poorly soluble in chloroform (≈0.02 m). Difluoroacetic acid and chloroform (in the presence of the OxP derivative) are miscible for arbitrary volume fractions.

Instrumentation

UV/Vis absorption spectra were recorded by using Jasco J‐820, Shimadzu UV‐3600, and Hitachi U‐2910 spectrophotometers (1 cm or 1 mm quartz cells with screw caps were used). NMR spectroscopy was performed by using Bruker Avance III HD 500 and JEOL AL300BX spectrometers. n class="Chemical">Tetramethylsilane was used as the internal standard. Titration experiments were performed by the addition of stock solution (acid dissolved in chloroform) into the sample by using a microsyringe. All measurements were performed at 25 °C unless otherwise specified.

Data‐Processing methods

The conversion of the measured UV/Vis spectra into the corresponding RGB values was conducted by using the Spectral calculator.53 Processing of UV/Vis data by using singular value decomposition (SVD) and decomposition into the spectra of individual species (DSIS) were performed according to the technical details given in the Supporting Information. Also, binding models (isotherms) used for UV/Vis and NMR spectroscopy data are discussed in detail in the Supporting information.

Conflict of interest

The authors declare no conflict of interest.

As a service to our authors and readers, this journal provides supporting information supplied by the authors. Such materials are peer reviewed and may be re‐organized for online delivery, but are not copy‐edited or typeset. Technical support issues arising from supporting information (other than missing files) should be addressed to the authors.

Supplementary

Click here for additional data file.