Proton transfer and hydrogen bonding in the organic solid state: a combined XRD/XPS/ssNMR study of 17 organic acid–base complexes †

The properties of nitrogen centres acting either as hydrogen-bond or Brønsted acceptors in solid molecular acid–base complexes have been probed by N 1s X-ray photoelectron spectroscopy (XPS) as well as 15 N solid-state nuclear magnetic resonance (ssNMR) spectroscopy and are interpreted with reference to local crystallographic structure information provided by X-ray diffraction (XRD). We have previously shown that the strong chemical shift of the N 1s binding energy associated with the protonation of nitrogen centres unequivocally distinguishes protonated (salt) from hydrogen-bonded (co-crystal) nitrogen species. This result is further supported by significant ssNMR shifts to low frequency, which occur with proton transfer from the acid to the base component. Generally, only minor chemical shifts occur upon co-crystal formation, unless a strong hydrogen bond is formed. CASTEP density functional theory (DFT) calculations of 15 N ssNMR isotropic chemical shifts correlate well with the experimental data, confirming that computational predictions of H-bond strengths and associated ssNMR chemical shifts allow the identification of salt and co-crystal structures (NMR crystallography). The excellent agreement between the conclusions drawn by XPS and the combined CASTEP/ssNMR investigations opens up a reliable avenue for local structure characterization in molecular systems even in the absence of crystal structure information, for example for non-crystalline or amorphous matter. The range of 17 different systems investigated in this study demonstrates the generic nature of this approach, which will be applicable to many other molecular materials in organic, physical, and materials chemistry.


Introduction
The crystallization of solid state acid-base donor-acceptor systems with more than one component is a commonly used route to tailor the physical and chemical properties of solid organic products and their formulations. For example, during the development of active pharmaceutical ingredients (APIs), the physicochemical properties of the pure API may be unsuitable for a chosen drug delivery route. Combining with a Brønsted donor or acceptor can result in intermolecular bonding, either by forming an ionic (salt) or hydrogen-bonded (co-crystal 1-3 ) complex with superior properties, 1-4 which may include solubility, 5 bioavailability, 5,6 chemical and physical stability, 7,8 hygroscopicity, 9 and mechanical properties. 10,11 Single crystal X-ray diffraction (XRD) is most commonly used for determining whether proton transfer or hydrogen bonding takes place between acid and base components, 3,5,[12][13][14][15][16][17][18] often in conjunction with an analysis of structural indicators such as bond angles and bond lengths. 3,12,16,19 However, the unequivocal determination of hydrogen positions is not always straightforward, particularly with systems exhibiting proton disorder, temperaturedependent migration, or other unusual behaviour. 18 Analysis by XRD is also constrained by a requirement for suitable single crystals, which may not be available when solid-state preparation techniques such as milling are used to form materials with more than one component. 20,21 Where single crystal XRD fails other commonly available laboratory techniques such as vibrational spectroscopies can sometimes provide the required information. 12,13 Often, however, the determination of hydrogen and proton positions has to rely on more advanced techniques such as neutron diffraction, 12,18,19 as demonstrated for, e.g., urea/phosphoric acid, 17,22,23 4,4 0 -bipyridyl/ benzene-1,2,4,5-tetracarboxylic acid, 24 4-methylpyridine/pentachlorophenol, 25 and benzoic acid. 26,27 Over the last decade, solid-state NMR (ssNMR) methods combined with computational chemical shift analysis 5,28,29 have been added to the inventory of techniques for the characterisation of hydrogen bonding and Brønsted interactions in the organic solid state. Studies have shown that H-bonding and proton transfer in two-component systems can be distinguished using 15 N ssNMR, 5,21,[29][30][31][32] as for example shown for co-crystals and salts of a cancer-treatment API 5 and for theophylline (1,3dimethyl-7H-purine-2,6-dione) systems. 21,28,30 Density functional theory (DFT) can be applied to correlate 15 N ssNMR parameters with crystal structures. For example, calculations for the co-crystal form of a pharmaceutical API correctly predicted chemical shifts (to low frequency) relative to the free base on formation of strong H-bonds, in agreement with experimental data. 5 Similarly, comparison of predicted 15 N chemical shift values for complexes of dabco (1,4-diazabicyclo[2.2.2]-octane) and dicarboxylic acids with experimental data confirmed Brønsted and H-bonding interactions in these systems. 29 Recently, we reported that X-ray photoelectron spectroscopy (XPS) also reliably detects proton transfer: a strong positive N 1s binding energy shift occurs due to protonation and identifies the formation of a salt. 21,28,30,33 The information of this chemical shift analysis is conceptually very similar to that used in the ssNMR approach, 28 opening up an avenue to mutual validation of results obtained with either technique. The principle of a large N 1s shift with protonation was applied to identify the nature of meta-aminobenzoic acid polymorphs (non-ionic vs. zwitterionic), 34 and an O 1s shift to higher binding energy was also observed for an oxygen proton acceptor with formation of a H-bond on moving from the gas to condensed phase. 35 Very importantly, long-range order is not a pre-requisite for the application of XPS and ssNMR; both techniques are also applicable to systems for which single crystal XRD would be unfeasible, for example when a suitable single crystal cannot be obtained, or when non-crystalline or amorphous samples are analysed. 36 The feasibility of a combined ssNMR/XPS approach to local structure characterization in the organic solid state was introduced for theophylline co-crystal/salt systems, 28 but to explore it more systematically we report here the examination of the nature of the intermolecular interactions between components of 17 acid-base complexes (Table 1 and Fig. 1), comparing XPS results with experimental and calculated 15 N ssNMR data. As in our previous XPS study, 33 we incorporate several development API substances alongside a range of non-development acid-base complexes including theophylline, aminobenzoic, and isonicotinamide base components with different acid co-formers and thus differing acid strength.
These systems cover a wide range of pK a differences (DpK a ), from À3.9 to +17.7, between the acceptor and donor functional groups, which will allow us to examine correlations between DpK a values and the chemical shifts observable by XPS and ssNMR in a systematic manner. Of the 17 complexes, 6 (4 salts, 2 co-crystals) are within the range 0 o DpK a o 3 (Table 1), for which it is generally unpredictable whether H-bonding or proton transfer takes place. 12,37 Experimental

Preparation of salts and co-crystals
The non-development co-crystals and salts were prepared by solid-state-grinding (milling) or solution crystallization. Milling was performed with a Retsch MM200 mixer mill at a rate of 30 Hz, using two 5 mL stainless steel jars, each containing one 7 mm-diameter stainless steel ball. The complexes were prepared as follows: (i) 1 : 1 Theophyllinium salicylic-5-sulfonate dihydrate salt was prepared by both milling and solution crystallization as described previously. 28 (ii) 1 : 1 Theophyllinium salicylic-5-sulfonate monohydrate salt was prepared by both milling and solution crystallization as described previously. 30 (iii) 2 : 1 Theophylline/oxalic acid co-crystal was prepared by both milling and solution crystallization as described previously. 30 (iv) 1 : 1 Theophylline/maleic acid co-crystal. Anhydrous theophylline (2.023 g, 11.23 mmol) and maleic acid (1.303 g, 1 equiv.) were dissolved in 35 mL of a 6 : 1 chloroform : methanol solvent mixture under reflux (53 1C). The solution was removed from heat and allowed to cool to ambient temperature while stirring. The resulting precipitate was removed by filtration under vacuum to give co-crystals with an excess of anhydrous theophylline. The filtrate was then allowed to stand overnight, Table 1 DpK a values for the seventeen complexes (base/acid), where DpK a = pK a (base) ÀpK a (acid) based on pK a values. 4,12,[38][39][40] For complexes involving two acids, the pK a of the more basic compound (with more basic substituent) is taken as pK a (base) affording the physically pure 1 : 1 co-crystal, which was isolated by filtration under vacuum. The 1 : 1 co-crystal was also formed by milling theophylline (109.99 mg, 0.61 mmol) and maleic acid (70.80 mg, 1 eq.) for 20 minutes.
(vi) 1 : 1 Theophylline/citric acid co-crystal was prepared by milling as described previously. 21 (vii) 1 : 1 Theophylline/glutaric acid co-crystal. Milling anhydrous theophylline (93.7 mg, 0.52 mmol) and glutaric acid (68.7 mg, 1 eq.) for 60 minutes formed the 1 : 1 co-crystal, which was physically pure as determined by X-ray powder diffraction. Theophylline (0.4972 g, 2.76 mmol) and glutaric acid (0.3646 g, 1 equiv.) were also dissolved in 35 mL chloroform under reflux (59 1C). The solution was removed from heat and seeded with B35 mg of co-crystal formed by milling. The solution was allowed to evaporate until B20 mL volume remained, and the precipitate (present at the surface) was filtered under vacuum to give the 1 : 1 co-crystal.

X-ray diffraction (XRD)
Powder XRD patterns were collected using a Rigaku Miniflex instrument utilizing Cu K a radiation (l = 1.5406 Å), operating over 5-401 2y at 1.51 min À1 with a 0.031 step, 30 kV voltage, and 15 mA current. Typically 5 mg of the sample was placed on a small sample attachment and smoothed to achieve a level surface.
Single crystal XRD data were collected at 100 K using an Oxford Diffraction X-Calibur 2 diffractometer utilizing Mo K a radiation (l = 0.71073 Å) and an Oxford CryoSystems Cryostream Controller 700. Data reduction, cell refinement, and multi-scan absorption corrections were carried out using the program CrysAlis RED (Oxford Diffraction Ltd., version 1.171. 32.24, 2008). The isonicotinamide/4-hydroxybenzoic acid co-crystal structure was solved with SHELXS-97 and refined on F 2 against all reflections with SHELXL-97. 45 The isonicotinamide/2,4,6-trihydroxybenzoic acid salt structure was solved with SIR92 46 and refined on F 2 against all reflections using SHELXL-97. 45 All non-hydrogen atoms were refined by direct methods anisotropically; hydrogen atoms were located in difference Fourier maps and refined isotropically, or placed in geometric positions and refined as riding atoms. CCDC 960081 and 960082. † X-ray photoelectron spectroscopy (XPS) XP spectra were recorded using a Kratos Axis Ultra instrument employing a monochromatic Al K a source (1486.69 eV), a hemispherical analyser with a hybrid (electrostatic and magnetic) lens system, charge neutralization by filament-generated, magnetically channeled low-energy electrons, and a delay line detector (DLD). Samples were fixed using double-sided tape. Experiments were performed while operating the X-ray source with a power of 180 W (15 kV and 12 mA), with the pressure below 10 À8 mbar during analysis. The instrument was operated in CAE (constant analysis energy) mode, with a pass energy of 20 eV for high resolution scans of the photoemission from individual core levels, with a calibrated intensity/energy response and transmission function. 36 High resolution spectra were measured within the spectral range of interest (ca. AE20 eV around the core level emission peaks) with 0.1 eV steps and 300-500 ms dwell time per data point. Repeats were carried out to check for radiation damage.
Analysis of the data was carried out using Casa XPS software. 47 A linear background was used in all curve-fitting (minimizing w 2 ) 47,48 along with a GL(30) lineshape (70% Gaussian, 30% Lorentzian using the Gaussian/Lorentzian product form). 47 For the remainder of this paper, when referring to spectral contributions of specific atoms in the molecular structure in the text, the atom of interest will be indicated by being underlined. Samples naturally containing aliphatic À C-C environments were referenced to adventitious hydrocarbon contamination at 285 eV. 48 Samples without C-C environments were referenced to the lowest E B photoemission; those containing citric acid were referenced to À C-COOH (b-COOH) at 285.2 eV 49 due to the absence of a clear hydrocarbon shoulder; samples containing 5-sulfosalicylic acid were referenced to À CQC at 284.8 eV. 50 Individual chemical environments/functional groups often exhibit similar ranges of binding energy values where the bonding or electronegativity is similar, thus where peaks arising from photoemission from different chemical environments occurred at the same position or within AE0.1 eV, a single peak was used to represent both environments for clarity and ease of discussion. Repeatability of the peak positions was within 0.1 eV.

Solid-state nuclear magnetic resonance (ssNMR)
Solid-state 15 N cross-polarisation magic angle spinning (CP-MAS) data were acquired using a Bruker DRX500 spectrometer operating at 50.69 MHz, equipped with a standard bore magnet and a 4 mm CP-MAS probe. The magic angle adjustment was optimized using KBr and field homogeneity optimized using adamantane. Data were collected using a spectral width of 30.3 kHz, 3072 complex data points, and an acquisition time of 50.8 ms. The relaxation delay and contact time were optimized for each individual sample. Sample rotation rates of 5.0-8.0 kHz were employed and typically greater than 2048 transients acquired. 1 H decoupling was achieved using a TPPM-15 sequence (86.2 kHz). Chemical shifts (d N ) were referenced indirectly with respect to nitromethane, a standard recommended by IUPAC, 51,52 via the high frequency signal of a traceable standard of ammonium nitrate (d N = À5.1 ppm). Chemical shifts for the theophylline and API 1 salts and co-crystals are compared with the values for the free base forms by the change in chemical shift (Dd = d complex À d free base ).
Isotropic chemical shift differences 53 were calculated using the CASTEP density functional theory (DFT) code, 54-56 which employs the gauge including projector augmented wave (GIPAW) algorithm. The generalised gradient approximation (GGA) PBE functional was used with core-valence interactions described by ultrasoft pseudo-potentials generated on-the-fly. Integrals over the Brillouin zone were completed using a Monkhorst-Pack grid with a reciprocal space k-point spacing of 0.04 Å À1 . A cut-off energy of 610 eV was defined for kinetic energy of planewaves used to provide a basis for wavefunctions. For each crystal structure, full geometry optimization was performed using an energy cut-off of 300 eV and a k-point spacing of 0.05 Å À1 , with fixed lattice parameters. Calculations were performed using Materials Studio version 4.4, as provided by Accelrys. 57 CASTEP DFT chemical shielding values (s) were obtained for the nitrogen atoms of the theophylline, 4-aminobenzoic acid, 3,5-diaminobenzoic acid, and isonicotinamide complexes using the previously reported 7,21,28,[41][42][43] and new isonicotinamide co-crystal/salt crystal structures as input. Shielding values for the co-crystal and salts are compared with chemical shifts by plotting calculated chemical shieldings (s) versus experimental chemical shifts (d N ). A plot of calculated chemical shieldings (s) versus experimental chemical shifts (d N ) for all of the CASTEP data gives a linear correlation with y = À0.9525x À 153.61 (R 2 = 0.9893). Predicted chemical shift differences use the free base as a reference [Dd calc = À(s complex À s free base )].

XPS
The four nitrogen environments (Fig. 1) in the theophylline complexes result in two X-ray photoemission peak components. 21,28,30 The peak arising from the C-À N/ À N-CQO nitrogens occurs at 401.0 AE 0.1 eV for both the co-crystal and salt forms (Fig. 2, Table 2). In the five theophylline co-crystals, the CQ À N photoemission occurs at 399.6 AE 0.1 eV. However, for the two theophylline salts, this second photoemission peak is at 401.9 eV (Fig. 2b, Table 2), with protonation (CQ À NH + ) shifting the CQN peak by +2.3 eV. 28,30 The 4-aminobenzoic and 3,5-diaminobenzoic complexes all have at least one primary amine nitrogen C-À NH 2 on the base component and a nitro group À NO 2 on the acid component ( Fig. 1 and 3). For the 4-aminobenzoic acid/3,5-dinitrobenzoic acid and 4-aminobenzoic acid/4-hydroxy-3-nitrobenzoic acid co-crystals (heterodimer bonding) this gives rise to two peaks at 399.5 AE 0.1 eV and 406.3 eV, from C-À NH 2 and À NO 2 respectively ( Fig. 3a and b, Table 2). 40 When a salt is formed, as in the case of the 3,5-diaminobenzoic acid/3,5-dinitrobenzoic acid complex, one of the two amine nitrogens of 3,5-diaminobenzoic acid is protonated (C-À NH 3 + ), shifting its peak position to a higher binding energy of 401.9 eV, a shift of +2.4 eV (Fig. 3c, Table 2). 40 There are three different types of nitrogen atom in isonicotinamide/4-hydroxy-3-nitrobenzoic acid -pyridine CQ À N and amide À N-CQO of isonicotinamide and nitro À NO 2 of the acid molecule (Fig. 1). The nitrogen 1s XPS (Fig. 4a) shows two maxima, with the high binding energy peak at 406.3 eV arising from the À NO 2 photoemsission, while the lower energy peak can be fit with two components at 399.6 and 400.1 eV representing the CQ À N and amide À N-CQO nitrogen functionalities respectively. With 2,4,6trihydroxybenzoic acid, there are no nitrogens in the acid molecule (no À NO 2 photoemission), leaving just the CQ À N and À N-CQO from isonicotinamide. Aside from the isonicotinamide À N-CQO peak at 400.1 eV, the CQ À N peak is shifted by 2.0 eV compared to that of isonicotinamide/4-hydroxy-3-nitrobenzoic acid and is observed at 401.6 eV, representing the change in chemical state from pyridine CQ À N to the pyridinium nitrogen CQ À NH + and evidence of salt formation (Fig. 4b, Table 2). Subsequent acquisition of the single crystal structure confirms the XPS-assignment of isonicotinamide/4-hydroxy-3-nitrobenzoic  28 showing the ratios for types of nitrogen and the corresponding intermolecular H-bonding/protonation from the single crystal XRD structures 7,28 with the nitrogen sensitive to H-bonding/protonation marked. acid as a co-crystal and isonicotinamide/2,4,6-trihydroxybenzoic acid as a salt (Fig. 4).
The API 1 provides a variety of nitrogen environments (Fig. 1), which can be grouped into pyridine derivatives (CQ À N Â 3), amines (C-À N Â 2), and amides ( À N-CQO Â 2). For the free base form (API 1), the photoemission by the C-À N and CQ À N nitrogens appears around the same binding energy position at 399.1 AE 0.1 eV, while that of À N-CQO ranges from 400.3 to 400.6 eV (Fig. 5, Table 2). Differences with respect to the free base are clearly evident for the salt 1s, with the presence of a pyridinium CQ À NH + peak component at 401.2 eV shifted by +2.1 eV compared to unprotonated, pyridine nitrogen (Fig. 5, Table 2). The relative intensity of this peak is in accord with the protonation of two of the pyridine nitrogens in the di-HCl salt 1s.
There is also an observable difference for the co-crystal 1c, with a decrease in the intensity of the C-À N/CQN peak relative to the free base and a broadening between the two free base peak components. This is accounted for by a third peak at 399.8 eV, of a relative intensity indicative of one of the C-À N/CQ À N nitrogens (Fig. 5, Table 2). A shift of +0.6 eV from the C-À N/ CQN photoemissions is too small a magnitude for complete proton transfer, so it appears likely that formation of a new hydrogen bond results in a high binding energy shift for the nitrogen acceptor atom.
The difumarate salt 2 has three nitrogen environments (Fig. 1), comprising two heterocyclic amines (C-À N Â 2) and one sulfonamide ( À N-SO 2 Â 1). The sulfonamide nitrogen gives rise to the peak at 399.5 eV, while protonation of the amine nitrogens results in a peak at 401.7 eV (Fig. 6a, Table 2), although this API is particularly sensitive to X-ray induced reduction of NH 3 + to NH 2 (occurring at coinciding binding energy with À N-SO 2 ), leading to a departure from the expected 2 : 1 ratio. 58 The HCl salts 3 and 4 have two and three nitrogens respectively   ( Fig. 1), with 3 including an amine (C-À N Â 1) and sulfonamide ( À N-SO 2 Â 1), while 4 has a heterocyclic amine (C-À N Â 1) and two amides ( À N-CQO Â 2). The sulfonamide peak of 3 and amide peak of 4 occur at 399.6 and 399.9 eV, respectively, while the protonated amine C-À NH + nitrogen photoemissions occur at higher binding energy, 401.7 and 401.5 eV, respectively ( Fig. 6b and c, Table 2).

N ssNMR
The spectra of the theophylline complexes, 21,28,30 1s, and 1c, were compared with the free base form of each in order to investigate the changes in chemical shifts occurring with formation of the complexes. 50-100 ppm shifts relative to the starting components (chemical shift difference Dd) are typical for protonation of aromatic, heterocyclic nitrogen, 5,28-31 and shifts of 47-118 ppm to low frequency confirm salt formation (protonated nitrogen, CQNH + ) for the two theophylline/5-sulfosalicylic acid salts 28,30 (Table 3), the isonicotinamide/2,4,6-trihydroxybenzoic acid salt (Fig. 7, Table 5) and the di-HCl salt 1s ( Table 4). The theophylline salts are protonated at CQ À N (N9, Fig. 1, Table 3), with ca. À50 ppm shifts (Dd), 28,30 and the isonicotinamide salt has a À68.76 ppm shift on protonation (Fig. 7, Table 5), while di-HCl salt 1s is protonated in two pyridine positions, N5 and N7, with shifts of À108.0 and À117.8 ppm respectively (Table 4). In contrast, the fumaric acid co-crystal 1c (Table 4) and the theophylline co-crystal resonances 21,30 (Table 3) show only minor changes in 15 N chemical shifts compared to the base component (Dd), with the exception of the pyridine N5 resonance in 1c. A shift of 32.6 ppm to low frequency occurs for N5 of 1c (Table 4), indicating formation of a strong hydrogen bond that is not present in the free base form. A component of this change in chemical shift may be ascribed to conformational/crystallographic packing differences induced by the hydrogen bonding, which is observed on comparing different polymorphic forms of the free base.
Aliphatic nitrogen often exhibits much smaller shifts with protonation than its heterocyclic counterparts (e.g. amines vs. pyridines). 59 The aliphatic nature of the protonated nitrogen (C-À NH 3 + ) in the 3,5-diaminobenzoic acid/3,5-dinitrobenzoic acid salt results in a smaller magnitude shift of À14.11 ppm relative to the unprotonated amine, but this is still sufficient to distinguish formation of a salt from a co-crystal. There are no significant shifts indicating protonation of the amine in the 4-aminobenzoic acid co-crystals (Table 5), confirming they are not salts.  Density functional theory (DFT) calculations (chemical shielding values, s) were performed for the nitrogens of the theophylline, aminobenzoic, and isonicotinamide complexes in order to compare with experimental data (chemical shifts, d). The plot of s versus d (Fig. 8) shows a linear correlation with y = À0.9525x À 153.61 (R 2 = 0.9893). This demonstrates the robustness of using CASTEP for relating 15 N ssNMR chemical shifts to molecular structure and, therefore, providing NMR based evidence for proof of co-crystal or salt formation.
The predicted chemical shift differences can be related to the H-bond length, giving a measure of H-bond strength, and providing an example of how NMR crystallography can be used in a complementary way with respect to X-ray crystallography. 60 For example, the predicted values as a function of distance between N of the base and H of the acid component show a clear separation between unprotonated (co-crystals, d Z 1.53 Å) and protonated nitrogen (salts, d r 1.11 Å) for the theophylline complexes ( Fig. 9). Additional multi-dimensional solid-state NMR studies are in progress to further characterize the nature of the hydrogen bonding in these co-crystals and salts.

DpK a correlations
Plotting the nitrogen XPS and ssNMR values for the co-crystals and salts against DpK a (Fig. 10) shows the dependence of the extent of proton transfer on the strength of the acidic component. Of the 17 complexes, the co-crystals have DpK a r 0.3, and the salts have DpK a Z 1.8, with DpK a ranging from À3.9 to 17.7, and four salts and two co-crystals have been assigned that are within the range for DpK a typically viewed as the transition region (0-3) for which formation of a salt vs. co-crystal is not predictable. 12 Recently, pK a matching has been used as a tool for predicting H-bond strengths 61 (including the proposed pK a   62 In line with this work, we find a clear separation between N 1s XPS chemical shifts of the co-crystals around 399.6 eV and salts around 401.7 eV (Fig. 10). Measurement of the N 1s binding energy unambiguously determines whether protonation has occurred, with a mean N 1s binding energy difference of +2.1 eV for N -NH + . As the 15 N ssNMR chemical shift d varies significantly with each base moiety and thus the absolute value is not a unique identifier for protonation state, the chemical shift difference (Dd, Fig. 10) is used to compare against the DpK a for the complexes (no ssNMR data for API 2, 3, and 4). There is a clustering of Dd values for the co-crystals, although there is an overlap between co-crystals and salts. The smallest chemical shift observed for protonation of nitrogen (À14.11 ppm) and salt formation occurs for 3,5-diaminobenzoic acid/3,5-dinitrobenzoic acid and is a result of the reduced magnitude of the shift for aliphatic rather than heterocyclic aromatic nitrogen 59 as mentioned previously, and the theophylline and isonicotinamide salts exhibit Dd of À50 to 70 ppm while the di-salt API 1s shows larger magnitude shifts of around 110 ppm. The largest (most negative) shift for nitrogen within the co-crystals occurs for the development drug co-crystal 1c, causing it to be greater than that of the signal from the aliphatic salt -formation of a strong hydrogen bond that is not present in the free base form (API 1) is intimated for co-crystal 1c, influencing the chemical shift, as well as a possible contribution from conformational/crystallographic packing differences. Comparing shifts between compounds sharing a component (such as salts and co-crystals which share the same free base) rather than the whole series clearly identifies each form correctly (Fig. 7, Tables 3-5).
XPS core level chemical shifts reflect primarily the influence of the most immediate atomic neighbours on the electronic state of the photoexcited atom, 33,48,50,63 with the effect of strong local intermolecular interactions such as ionic and H-bonding dominating over the comparatively weaker van-der-Waals and dipole interactions. It is for this reason that the core level binding energies reported in Table 2 and Fig. 10 are so universally sensitive to local structure and thus protonation state. In contrast, NMR is far more dependent on the extended environment and interactions, which leads to greater variance in Dd as an indicator of protonation, although it does provide spatial resolution (allowing structurally inequivalent atoms of the same type of functional group to be distinguished). The combination of XPS and ssNMR to identify when hydrogen transfer has occurred from acid to base component is therefore especially strong, making assignment of salt or co-crystal unambiguous.

Conclusions
Nitrogen XPS and ssNMR chemical shifts clearly separate protonated from unprotonated nitrogen for a wide range of different solid-state donor-acceptor systems, including five pharmaceutical drug substances. The nature of the intermolecular interaction between the acid-base components was correctly identified for the eight salts and nine co-crystals. Brønsted proton transfer to nitrogen of the base component (salt formation) leads to chemical shifts towards higher core level binding energies in XPS and to lower frequency in ssNMR, while the hydrogenbonded co-crystals exhibited only minor shifts compared to the free base form, with the exception of one of the pharmaceutical co-crystals (1c). The salt and co-crystal of the same API (1) Fig. 10 Correlations between N 1s XPS and 15 N ssNMR chemical shifts with DpK a , illustrating the clear separation between protonated (salt) and unprotonated nitrogen (co-crystal) with XPS and the overlap for new H-bond formation in a co-crystal (1c) and protonation of aliphatic nitrogen in a salt (3,5-diamino/3,5-dinitro) with ssNMR. allowed comparison between the magnitude of the shift for formation of a strong hydrogen bond (at a position where there was none in the free base) compared to that for complete hydrogen transfer (protonation) at the same position in the salt. DFT-calculated 15 N ssNMR results show good correlation with experimental data, allowing prediction of chemical shifts and assignment of salt or co-crystal, and the strong influence of H-bond strength on the chemical shift.
Our data demonstrate both the versatility and unambiguity of such a combined XPS/ssNMR approach, which has, to our knowledge, never been systematically pursued before across such a large body of samples. The results will inform a more critical assessment of the possibilities and limitations associated with XPS and ssNMR, and provide a reference database for future investigations of hydrogen-bonding and protonation in other systems, for example the study of solvation effects in the nucleation and growth of organic crystals. 64