Importance of Molecular Symmetry for Enantiomeric Excess Recognition by NMR

Recently prochiral solvating agents (pro-CSA) became a spotlight for the detection of enantiopurity by NMR. Chemical shift non-equivalency in achiral hosts introduced by the presence of chiral guest yields observable resonance signal splitting () correlating to the enantiomeric excess (e.e.). In this work, symmetry is our lens to explain porphyrin-based supramolecular receptors’ activity in a chiral environment. Based on extensive NMR analyses of the atropisomeric receptors, host symmetry is shown to be affected by porphyrin nonplanarity and further desymmetrized in the presence of a chiral guest. We have formulated a simple, symmetry-based protocol that can be used to identify pro-CSA candidates. As such, the exposed porphyrin inner core (N–H), with its strong hydrogen bond abilities, for the first time, has been exploited in enantiomeric composition analysis. Our approach in e.e. detection by N–H signals appearing in a previously underutilized region of the spectrum (below 0 ppm.), shows chemical shifts (the e.e. dependent splitting ) three times more sensitive to enantiomeric compositions than previously reported systems. The findings are complemented by extensive 2D NMR studies, including the first reporting of e.e. dependent  in nonhydrogen NMR, and supporting by density functional theory (DFT)

Among the numerous stereodiscrimination methods, nuclear magnetic resonance (NMR) spectroscopy continues to be one of the leading tools for determining the enantiomeric purity of chiral molecules. [1] However, enantiomers cannot be distinguished in an achiral environment as NMR active nuclei are isochronous. Usually, some external source of chirality [1d] is introduced in the form of a covalently bonding chiral derivatization agent (CDA), a non-covalently interacting solvating agent (CSA), or even self-induced recognition of enantiomers (SIRE) [1a] to convert the enantiomeric systems into diastereomeric ones. Recently, a new type of NMR spectroscopic detection of enantiomeric excess (e.e.) using prochiral solvating agents (pro-CSA) was introduced by Hill and co-workers. [2] In principle, in the event of attractive noncovalent physicochemical interactions, the chiral information of a guest can be transferred to an achiral host and detected as the splitting of the NMR signals. The key example of pro-CSA, N,N'-disubstituted oxoporphyrinogen (Bz2oxP) exhibits a linear response between the e.e. value and the magnitude of -proton splitting (∆) in 1 H NMR ( Figure 1a). [3] Due to N-alkylation of the Bz2oxP core, the system cannot be protonated and hence suffers serious sensitivity issues compared to unmodified oxP. However, the inevitable prototropic tautomerism and macrocyclic inversions obstruct the potential applications of oxP as a pro-CSA. [2a,4] Porphyrins, as prospective pro-CSA candidates for e.e. detection, have also been investigated. [5] While 5,10,15, is not affected by the disadvantageous tautomeric processes, as opposed to oxP, the necessary use of depressed temperatures for the e.e. detection limits the analysis to explicit solvents with a low freezing point (e.g. CDCl3) and analyte solubility during the screening (e.g. precipitation).
Frequently, the use of pro-CSA's 1 H NMR spectra for chiral analysis is severely hampered due to the numerous scalar couplings and overlapping signals that lead to analytical difficulties. [6] As the majority of organic molecule resonances appear between 0-14 ppm in the 1 H NMR scale, [7] it is desirable that the e.e. monitoring with pro-CSA would be in a distinct, wellseparated region. One of the most unique characteristics of porphyrins is the closed-loop of electrons (ring current) exhibiting large magnetic anisotropy under an applied magnetic field. While peripheral macrocycle signals relate to the typical organic resonances, the nuclei positioned within the loop experience a strong shielding effect when subjected to an external magnetic field and resonate below 0 ppm in the 1 H NMR scale. [8] Once the highly conjugated system is disrupted (e.g., in oxopophyrinogens, calix [4]pyrroles), the anisotropic shielding effect of the inner core system is lost, resulting in downfield shifting of the corresponding inner core signals.
The attractive features of the metal-free (free base) porphyrin inner core has lately drawn attention in the fields of catalysis [9] , sensing [10] , supramolecular assemblies [11] , and absolute configuration determination. [12] Typically, the imine and amine units of porphyrins are not involved in intermolecular interactions due to the planar nature of the macrocycle. However, the existing methods of ring puckering by steric strain [13] can cause a degree of outwards orientation of the inner pyrrolic entities, making these positions more basic [14] and accessible to substrates. [10] Even though porphyrins adopt a saddle-shaped 3D conformation [15] creating an 'active center' in the core, only the saddle-deformation alongside chiral guest interactions is not enough to drive the inner N-H signal to split during the 1 H NMR e.e. analysis. For example, Bz2oxP has a saddle shape and belongs to the C2v point-group notation with two mirror planes diagonally dividing all pyrroles ( Figure 1a). The symmetrical nature of Bz2oxP does not permit the e.e. discrimination using the inner core. However, the N-H signal shifts to the lower field of the spectrum due to the non-covalent interactions nevertheless remain isochronous. [3] Figure 1. Top: representation of pro-CSA's (blue above the plane, redbelow); Middle: with symmetry elements (Mirror plane σ and rotation axis Cn); Bottom: the key units used for e.e. detection by 1 H NMR; a) Bz2oxP highlighting -H splitting; [3] b) newly designed 22-P receptor system with chiral discrimination by N-H; c) All possible P atropisomers with corresponding point groups, N-H signals, and magnitude of splitting; see more detail in Figure S1.
Here we report the first example of e.e. detection using porphyrin inner core N-H resonances. We have designed P [5,10,15,20-tetrakis(2-aminiumphenyl) -2,3,7,8,12,13,17,18octaethylporphyrin] as a receptor system ( Figure 2a) exploiting three main molecular engineering strategies: 1) steric overcrowding to obtain a saddle-shaped macrocycle while retaining the porphyrin conjugation [13] and exposing the inner pyrrolic units for host-guest interactions; 2) peripheral donating groups creating a lock-and-key [10c] comparable system to encapsulate chiral analytes in the porphyrin lattice and allow the detailed NMR analysis at room temperature. [10d] 3) formation of atropisomers based on the orientation of peripheral groups [16] to have the ultimate control of the symmetry elements in pro-CSA. [17] In our previous study, we have highlighted the selective nature of host P for guests containing sulfonate or phosphonate motifs. [10d] The analyte interacts directly with the inner ring system and generates static and well-resolved NMR spectral lines. As previously mentioned, the depressed temperatures can also offer slow exchange rates for potential detection of e.e. [5] However, the aim of the following studies is the development of a readily available and highly effective analytical tool for room-temperature measurements. Therefore, (±)-10-camphorsulfonic acid (10CSA) bearing the sulfonic moiety and stereogenic centers was selected as a chiral guest in the present study.
Operating with enantiopure 10CSA(S or R) four distinct scenarios with four different P atropisomers were observed and subsequently rationalized by the symmetry operations found in P (Figure 1c). [17] In the α4-Pꞏ10CSA(S or R) complex, the inner core remains isochronous, due to the C2v point-group notation with a two-fold symmetry axis and two mirror planes passing through the pyrroles. The identical situation previously reported by Hill and coworkers in Bz2oxP pinpoints the interactions with inner N-H, however, without the e.e. discrimination due to the C2v symmetry ( Figure 1a). [3] The 22-P atropisomer with Cs symmetry features a single well-defined mirror plane dividing two pyrrolic units which preserve its achiral nature, hence allowing it to be classified as pro-CSA. The lack of other symmetry elements in 22-P allows the N-H protons to become anisochronous in a chiral environment, making chiral discrimination possible (with the highest magnitude of splitting (max) of 0.653 ppm at 100% e.e.) ( Figure 1b). The α3β-P atropisomer belongs to the C1 point-group, as it contains no symmetry elements, making the system chiral. Thus, eight signals are observed with enantiopure 10CSA due to diastereomer formation (SS-and SR-or RR-and RS-) ( Figure S1). While the e.e. detection is possible with α3β-P ( Figure S2), the practical use of such system falls short mainly due to three dominating factors: 1) the high number of inner core system signals hampers direct e.e. interpretation; 2) the magnitude of max (~0.39 ppm) is lowest of the three atropisomers with inner core splitting making it the least sensitive system; 3) the concentration of α3β-P is required to be significantly higher than that of other systems due to a large number of resonance signals and their comparatively lower intensities. On the other hand, αβαβ-P which belongs to the S4 point group has four equivalent protons located in the principal axis. While it has no mirror planes, the inversion center situated between the pyrrole units allows the inner core protons to split in equal proportions (above and below the plane) upon interaction with a chiral analyte. A single isochronous N-H signal of αβαβ-P-10CSA(S or R) becomes the inner system (N-H red/green) was found to be more than threefold greater than those of other regions (0.653 ppm).
The origin of the chemical shift non-equivalence lies deep within the concept of prochirality. [19] The desymmetrization of 22-P atoms in a single step by weak interactions with a chiral guest proves to be particularly useful for the e.e. determination. To illustrate 10CSA interactions with α2β2-P, a conformational search was performed using the α2β2-P[SO4 2-][HSO4 − ]4 structure for building starting geometries. Corresponding noncovalent interactions of the major conformer are illustrated in Figure S14. When racemic and non-racemic mixtures of 10CSA were applied in the system at constant concentrations it was found that ∆σ of the o-ArH, CH3, and N-H peaks rely on respective % of the e.e. value (Figure 2c). At the racemate point, the isochronous profile of 22-P is restored since the chiral information is transmitted in equal proportions from both the chiral components. Since the ∆σmax value of the inner core system is substantially higher than that of other regions, the resolution, of which e.e. can be detected, is considerably enhanced. Astonishingly, at as low as 2% e.e., two distinct N-H resonance singlets (∆σ 0.022 ppm.) can clearly be identified, while the other regions show only a broadening of the signals. Plotting the differences in the chemical shifts of split peaks against the % e.e. values revealed a linear dependency with the R 2 values being above 0.997 and the inner N-H fitting R 2 = 0.9994 (Figure 2d). The linear fit of the plots is a fundamental property in unlocking the easy calibration of the referenced systems for quick detection of the e.e. value (a detailed example shown in SI; Figure S10-S12). Moreover, spatially distant neighboring protons from N-H offer another important feature. Sharp and well-isolated singlets do not suffer from any vicinal scalar J-couplings or roofing effects underlining the simplicity in tracking chiral compositions. Overall, monitoring changes of this inner core system in a model chiral environment demonstrates a powerful tool for easy and sensitive detection of enantiomeric compositions. The magnitude of non-hydrogen ∆σ relies on the spatial positions, distances, and interactions with chiral guests. [3] The further the stereogenic center of a chiral guest from the host molecule, the weaker the chirality transfer is. Interestingly, this principle was previously well-defined in the porphyrin-based hostguest chirogenic systems by using circular dichroism spectroscopy. [20] The dependence of the non-equivalency to the chiral guest location can be illustrated by 13 C and 15 N NMRs. (For detailed comparison of non-hydrogen resonances see Table S1). When 22-Pꞏ10CSA(S) was compared to racemic 22-Pꞏ10CSA(SR), most of the macrocyclic ring system exhibited ∆σmax > 0.3 ppm with the central two nitrogen atoms having ∆σmax = 1.67 ppm ( Figure 3). Nevertheless, due to the greater distance from the active site, most of the phenyl ring resonance signals remained isochronous. Despite this, two particularly different scenarios were portrayed by the o-Ar-13 C NMR signals. The ∆σmax between 15 6 and 20 6 positions yielded excellent separation (~1.3 ppm), whereas the 5 6 and 10 6 imposed only marginal ∆σmax (0.04 ppm). A closer examination of the crystal structure of α2β2-P[SO4 2-][HSO4 − ]4 revealed a closer distance between C15 6 and C20 6 (~6.429 Å) than between C5 6 and C10 6 (~9.311 Å), subsequently forming a narrow channel for the chiral guest to occupy ( Figure 3). Moreover, the calculated chemical shifts of non-hydrogen atoms in 22-Pꞏ10CSA(R) using the GIAO-B3LYP/6-311++G**//BP86-D3BJ/def-SVP method and SMD solvent model correlated well with the splitting patterns observed experimentally (for more information check SI, Table S8). Of note, in 1 H NMR the 15 6 and 20 6 positions had similar ∆σmax. Hence, while hydrogens of the o-Ar group were more likely to form weak interactions with the chiral compounds, the corresponding ∆σmax value of carbon resonances hinges on the spatial arrangements and proximity to the guest. Comparison of the 22-Pꞏ10CSA(S and SR) splitting resonance signals to other atropisomeric species is detailed in SI (Table S2-S3).
To conclude, the point groups were found to play a fundamental role in adjusting supramolecular receptor systems for e.e. determinations by the NMR method. Four atropisomers containing different point group notations were thoroughly investigated by NMR with (S and R) camphorsulphonic acid pinpointing the 22 rotamer as the most sensitive receptor for chirality detection. It was found that the ∆σmax value of N-H signals can reach 0.653 ppm, a three-fold greater splitting than any known pro-CSA. Such enhanced sensitivity towards the chiral components allows for readily available and exceptionally detailed enantiomeric excess detection at room temperature by NMR. Deposition

COMMUNICATION
Just like a screwdriver turning screws, symmetry elements can be adjusted by fine-tuning the orientation of rotationally restricted side-groups in supramolecular receptors. In this research we highlight the fundamental role of symmetry in chiral reporting by NMR. Newly designed porphyrins with exposed inner core N-H system can respond to a chiral guest with exceptionally sensitive enantiomeric excess detection.

Normal-structural decomposition (NSD):
The NSD method, as developed by Shelnutt and coworkers, [1] was used to delineate, quantify, and illustrate the various distortions modes present in the tetrapyrrole macrocycles. Analysis was performed with the NSD online interface, available at https://www.sengegroup.eu/nsd. [2] Single crystal X-ray crystallography: Crystals were grown following the protocol developed by Hope, liquid-liquid diffusion of CHCl3 and MeOH with H2SO4. [3] Diffraction data were collected on a Bruker APEX 2 DUO CCD diffractometer using Incoatec IμS Cu-Kα (λ = 1.54178 Å) radiation. Crystal was mounted on a MiTeGen MicroMount and collected at 100(2) K using an Oxford Cryosystems Cobra low-temperature device. Data were collected using omega and phi scans and were corrected for Lorentz and polarization effects using the APEX software suite. [4] Data were corrected for absorption effects using the multi-scan method (SADABS). [5] Using Olex2, the structure was solved with the XT structure solution program, using the intrinsic phasing solution method and refined against │F 2 │ with XL using least-squares minimization. [6] If electron density was not sufficient, the C and N bound H atoms were placed in their expected calculated positions and refined using a riding model: N-H = 0.88 Å, C-H = 0.95-0.98 Å, with Uiso (H) = 1.5Ueq (C) for methyl H atoms and 1.2Ueq (C,N). Details of data refinements can be found in Table S4. All images were prepared using Olex2. [6a] In the structure α2β2-P[SO4 2-][HSO4 − ]4, two phenyl rings at C5 and C10 and one ethyl group are modelled over two locations using DFIX, SIMU, SADI restrains and EADP constraints. In terms of counter anion, only the SO4 2group is not disordered, while all other HSO4 − groups were modelled disordered using rigid groups. Some hydrogen atoms were placed geometrically to compensate for close contacts, the remaining hydrogens could not be located on the disordered HSO4 − moieties but were added to the formula to make the formula weight correct. Multiple disordered and partially occupied H2O molecules are modelled in the structure using SIMU and ISOR restrains. The weighting scheme was manually adjusted to ensure the goodness of fit was reasonable.
Deposition number 2143572 contain the supplementary crystallographic data for this paper. These data are provided free of charge by the joint Cambridge Crystallographic Data Centre and Fachinformationszentrum Karlsruhe http://www.ccdc.cam.ac.uk/structures. All calculations were done with Gaussian16 Rev. B.01. [7] A conformational search for α2β2-Pꞏ10CSA(R) was performed using a crystal structure of α2β2-P[SO4 2-][HSO4 − ]4 supramolecules as a starting point for building porphyrin/camphorsulfonic acid supramolecules, followed by optimization in acetonitrile. The geometry optimization and frequencies calculations were performed using BP86 [8] -D3BJ [9] /def2-SVP [10] -the method which showed a good agreement with the experimental data reported in our previous works. [11] To include acetonitrile effects the SMD [12] continuum solvent model was used, where molecular surface was represented as Solvent Accessible Surface (SAS) and the Bondi atomic radii were used. A ground state was characterized by absence of imaginary frequencies and more accurate electronic energies were calculated using the BP86-D3BJ/def2-TZVPP [10] and SMD model. During conformational search twenty start geometries converged into eight conformers corresponding to the ground state (Table S5) with one major conformer A making up 90%. The geometry were visualized using GaussView 6.1. [13] and is illustrated in figure S14.
The NMR shielding tensors were calculated at the GIAO [14] -B3LYP [15] /6-311++G** [14a, 16] level of theory using the SMD continuum solvent model to include acetonitrile effects. To calculate the 13 C chemical shifts, a scaling factor of 1.0228 [17] and a reference point TMS (180.7 ppm) were used , for calculation of the 15 N chemical shifts NH3 was used as a reference point (253.70 ppm) (Table S6 -S8). Population analysis was done using the BP86-D3BJ/def2-TZVPP//BP86-D3BJ/def2-SVP level of theory in acetonitrile and NBO [18] approach in acetonitrile. Non-covalent interactions were also analyzed using the SMD (acetonitrile), the BP86-D3BJ/def2-SVP level of theory and AIMAll program version 19.10.12 [19] In Text Supplementary Material NMR Investigation of the P-10CSA Atropisomers Figure S1. 1 H and 1 H-15 N HSQC NMR spectra obtained for 0% and 100% e.e. Pꞏ10CSA solutions (20 equivalents) in d3-acetonitrile. On the right side, a graphical illustration of Pꞏ10CSA atropisomers with corresponding point group notations, inner core system protons are highlighted in different colors correlating to the arrows marked in 1 H NMR spectra.

5
We performed 1 H NMR and 1 H-15 N HSQC analyses of all P atropisomers with racemic mixtures and enantiopure 10CSA(S or R). For the racemic 10CSA solutions, as expected 1 H NMR spectra of P atropisomers remained isochronous ( Figure S2). Due to the chiral information transmitted in equal proportions from both chiral components (10CSA R and S), the observed inner core signals resonate in an identical manner to achiral acids (BSA and MSA) previously reported by us. [20] One singlet is observed for αβαβ-Pꞏ10CSA, two in α4-Pꞏ10CSA, three signals of relative intensity 1:2:1 in α2β2-Pꞏ10CSA, and finally the spectrum for the unsymmetrical α3β-Pꞏ10CSA atropisomer has four differently shifted signals. The enantiopure solutions are described in the main text.  Figures S4 and S5). The magnitude of chemical shifts of split peaks (∆σ) drastically increased over the first ~7 eq., while the later additions resulted in marginal changes to ∆σ ( Figure S6).   The addition of 12 eq. of TFA was to solubilize and protonate 22-P. Highlighted with red dot and star is two different splitting N-H resonance signals. Note, these spectra were recorded with higher concentrations of 22-P expecting to get a better resolution of the inner N-H. From the obtained spectra it appears that the inner core system of interest originates downfield shifted and overlays with the aliphatic CH3 region.

SUPPORTING INFORMATION
8

Influence of Water on the Complexation
We have previously mentioned that one of the best-known pro-CSA (Bz2oxP) suffers serious sensitivity issues due in part to N-alkylation. [21] Additionally, the competitive binding of water molecules significantly contributes to the magnitude of splitting ∆σ. Trace amounts of water in solutions or titrants results in the necessary use of high guest concentrations to obtain well-resolved spectra. The hypersensitivity towards water is a major limitation for functional pro-CSA, since water is ubiquitous, avoiding it is at least tedious if not almost impossible for most of the solvents and reagents, especially for the analytes bearing high polarity. Hence, investigation of water influence as a competitive agent to the enantio-pure 22-Pꞏ10CSA(S) complex in CD3CN was carried out. From the first instance, the gradual addition of water has a considerable effect on ∆σ. By the addition of 240 eq. of water, the ∆σ of N-H has contracted by 0.132 ppm. ( Figure S9). Despite the competitive nature of water, the overall binding strength of 10CSA(S) remains observable considering the ~12.5 fold higher water consistency in the solution. Even at substantially higher (>4000 eq.) quantity of water the signals remained anisochronous highlighting the strong relationship between the host and chiral guest ( Figure S8-S9).

Detection of Enantiomeric Excess
For example, 19 eq. of an unknown enantiomeric mixture of 10CSA was complexed with 22-P ( Figure   S11). The inner core system shows clearly split resonance signals with ∆σ = 0.13 ppm. A small addition of 10CSA(R) decreased ∆σ (0.051 ppm), while a similar amount of 10CSA(S) increased ∆σ (0.224 ppm) revealing the predominant enantiomeric identity (S > R). Next, a linear calibration plot with 7 data points (e.e.10CSA(S) = 0, 48, 58, 69, 79, 90, 100 %) was constructed. It is worth noting that it is possible to generate a calibration curve from a single measured enantiopure point due to the linear dependency with the second point being ∆σ = 0 ppm. where e.e. = 0%. Lastly, the ∆σ can be fitted to the calibration plot and reveal the unknown e.e. (21%) based on the inner N-H chemical shift difference between the split peaks (for more information on this example see figure S12). The same principle can be applied to quickly test the purity of enantiomers, i.e. by equally pre-mixing stereoisomers of interest with opposite chirality, in the event of matching purity, the 22-P signals will remain isochronous. Overall, monitoring changes of this inner core system in a model chiral environment demonstrates a powerful tool for easy and sensitive detection of enantiomeric compositions.  (20 eq. of 10CSA(R) and 19 eq. of 10CSA(S)) were used to highlight the diversity of the enantiomeric excess detection using 22-P. Spectra recorded in CD3CN. Figure S11. The 1 H NMR spectra of the inner core system N-H unknown e.e.% target compound (highlighted in blue), after small 10CSA(R) (red) and 10CSA(S) (green) additions, and the rest of the spectra for the construction of the calibration curve. Spectra recorded in CD3CN. Figure S12. Graph of the ∆σ dependence on the ee% with 19eq. of 10CSA. Measured from the 1 H NMR split N-H signals recorded in CD3CN (at 100 ee% -19 eq. 22-Pꞏ10CSA(S)). On the right side: calculations of the unknown ee from the calibrated curve and using a one-point system. Note, calculations using a one-point system should only be used for quick, approximate determinations of the ee.

C and 15 N Investigations
To further understand the transfer of chirality to the atropisomeric receptor systems P, we have performed 2D NMR analyses with enantiopure 10CSA. The 15 N resonance signals obtained from 1 H-15 N HSQC varied from 125 to 135 ppm and correlated well with the corresponding inner core system protons.

1.73
Deepest hole/e Å -3 -0.99 Porphyrin α2β2-P is represented in black(C) and blue(N), with the reference structure (CuTPP) in red dotted lines. [2] Right top: top view and sideview of α2β2-P-SO4 structure. Non-essential hydrogens, majority of counter anions and solvent molecules omitted for clarity, thermal ellipsoids shown at 50% probability.

Geometry analysis
Based on the crystallographic data we concluded that one porphyrin cation binds two camphorsulfonic acid molecules. Using this ratio and the α2β2-P[SO4 2-][HSO4 − ]4 crystal data the corresponding host-guest complexes were built with subsequent conformation search. According to the Boltzmann distribution, in acetonitrile one major conformer A (90%) and one minor conformer B (up to 10%) being higher in energy by 1.30 kcal/mol are presented (Table S5).
In all host-guest complexes, the porphyrin plane is significantly distorted (the Cβ-Cβ-Cβopp-Cβopp angles varying in the range of 0˚-25˚ and N-Cα-Cα-N angles varying in the range of 37˚-48˚) because of a steric hindrance between the peripheral substituents (for NSD profile check Figure S13). This deformation results in appearance of two cavities on both sides of the porphyrin macrocycle, which differ by the position of NH3 + groups. In one cavity the ammonia groups are placed on the same side of the cavity and in another -on the opposite sides ( Figure S14a), and the distance between two nitrogen atoms increases from 4.899 Å to 8.7174 Å, respectively. The difference in the position of NH3 + groups results in a non-identical mode of the binding of two camphorsulfonic acids ("standing" and "lying"). Both the NBO and AIMALL analysis showed that the guest molecules interact with the porphyrin cation through the formation of four H-bonds, which cause significant elongation of the N-H bonds by 0.08-0.03 Å in the case of NH3 + groups and by 0.03 Å in the case of inner core protons for the major conformer A ( Figure S14b, Tables S6 and S7).   (1) (1) BD (1) (2) BD (1)

NMR calculations and analysis
In addition, the 13 C and 15 N NMR shielding tensors and chemical shifts, as well as ∆σmax between the corresponding chemical shifts of the main dominant conformer A of α2β2-Pꞏ10CSA(R) were calculated (Tables S8-S10). The calculated 13 C and 15 N NMR chemical shifts were found to deviate from the experimentally measured data, while the calculated and experimental ∆σmax values are in good agreement (Table S8), except the ∆σmax between carbons 2 and 13, 3 and 12, and nitrogens 22 and 24, which showed strong deviation (Table S8). The disagreement between these three ∆σmax values can be explained by the presence and influence of solvent molecules situated in a larger cavity. In contrast to the smaller cavity, where only one "lying" guest molecule can be placed, in the larger cavity in addition to the "standing" guest molecule several solvent molecules can be placed as well. However, the continuum solvent model (SND) used represents solvent as a continuous medium and does not count for individual effects of "explicit" molecules, such as H-bonds formation or charge transfer. We suppose that the deviation from the experiment is due to not accounting for the individual solvent molecules' effects in the larger porphyrin cavity. Between other ∆σmax calculated using theoretical and experimental data, a good correlation is observed, proving that the main calculated conformation corresponds to the dominant conformation presented in solution, which shows non-equivalency of the carbon and nitrogen atoms. To improve agreement with experimental measurements the conformer A of α2β2-Pꞏ10CSA(R) with one and two additional acetonitrile molecules was modelled, and its 13 C and 15 N NMR shielding tensors and chemical shifts, as well as ∆σmax between the corresponding chemical shifts were calculated.
Inclusion of explicit solvent molecules in the model system improved agreement with the experimental data, especially in the case of the ∆σmax between carbons 2 and 13, 3 and 12, and nitrogens 22 and 24 (Table S8). However, the model system is sensitive to the presence of solvent molecules, especially in the case of peripheral atoms. Thus, to perform accurate NMR calculations, the whole solvent shell should be modelled which is beyond the scope of this study. Table S8. Calculated shielding tensors and chemical shifts of 13 C of conformer A of α2β2-Pꞏ10CSA(R) with and without solvent molecules.

NBO partial charges calculations and analysis
In order to clarify the source of ∆σ in 13 C and 15 N NMR, the NBO partial charges were calculated for the The ortho-carbons in the porphyrin cations 4+ and 6+ have similar partial charges (differing up to 0.006) ( Figure S15 and Summarizing, based on the six model complexes and their NBO partial charges it was shown that ∆σ values rely on the interactions with the chiral guests of certain size and chirality transfer effect. Figure S15. NBO partial charges in variety of calculated complexes.