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Sep 25, 2017 - We present a new method for differentiating racemic crystals from enantiopure crystals. Recently, developments in optical filters have ...
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Characterization of Crystal Chirality in Amino Acids Using Low-Frequency Raman Spectroscopy Hagit Aviv, Irena Nemtsov, Yitzhak Mastai, and Yaakov R. Tischler J. Phys. Chem. A, Just Accepted Manuscript • DOI: 10.1021/acs.jpca.7b07033 • Publication Date (Web): 25 Sep 2017 Downloaded from http://pubs.acs.org on October 3, 2017

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The Journal of Physical Chemistry

Characterization of Crystal Chirality in Amino Acids Using Low-Frequency Raman Spectroscopy

Hagit Aviv†, Irena Nemtsov†, Yitzhak Mastai*, Yaakov R. Tischler*

Department of Chemistry and Bar-Ilan Institute for Nanotechnology and Advanced Materials (BINA), Bar Ilan University, Ramat Gan, 5290002, Israel

† Equally contributing authors * Corresponding authors

[email protected] , [email protected]

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Abstract We present a new method for differentiating racemic from enantiopure crystals. Recently, developments in optical filters have enabled the facile use of Raman spectroscopy to detect low-frequency vibrational (LFV) modes. Here, for the first time, we use Raman spectroscopy to characterize the LFV modes for crystalline organic materials composed of chiral molecules. The LF-Raman spectra of racemic and enantiopure crystals exhibit a significant variation, which we attribute to different hydrogen-bond networks in the chiral crystal structures. Across a representative set of amino acids, we observed that when comparing racemic versus enantiopure crystals, the available LFV modes and their relative scattering intensity are strong functions of side chain polarity. Thus, LF-Raman can be used as a complementary method to the currently used methods for characterizing crystal chirality due to simpler, faster, and more sensitive measurements, along with the small sample size required, which is limited by the laserbeam diameter in the focus.

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Introduction Most living systems are composed of chiral molecules, amongst which are amino acids, sugars, proteins, enzymes, and nucleic acids. The left- and right-handed enantiomers of a chiral compound possess different biological activities and physiological effects in the human body, due to different interactions with enzymes and proteins.1,2 It is therefore extremely important to achieve chiral separation and produce enantiomerically pure products, e.g. by recognizing racemic structures or eliminating an unwanted enantiomer. Molecular separation of enantiomers is most readily accomplished by means of chiral chromatography.3,4 Some of the chiral chromatography techniques include: gas chromatography,5 high performance liquid chromatography,6 supercritical fluid chromatography,7 and simulated moving bed technology.8 Moreover, it is important to identify chiral molecules since their solutions and mixtures are very common. The basic measurements used for this purpose in solution are still based on well-established methods that have been used for decades, such as optical rotation and circular dichroism. More modern methods are vibrational circular dichroism9 and second-harmonic generation circular dichroism.10,11 Chirality measurements in the solid state are considered difficult; the common methods used today are X-ray diffraction (XRD) and differential scanning calorimetry.12,13 Several groups have reported on chirality measurements using low-frequency vibrations by terahertz (THz) absorption,14–16 and inelastic neutron scattering.17 In this study, we present a new method for characterizing chirality of crystals based on the use of low-frequency (LF) Raman spectroscopy. In a crystalline structure composed of molecules, there are vibrational modes associated with chemical bonds within each molecule, intermolecular interactions, and with overall oscillations of the

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crystal lattice, i.e. phonons. For a given vibration at frequency ω, the oscillations can interact with light producing signals that are detectable by different methods of vibrational spectroscopy.18 Until recently, low-frequency vibrational (LFV) modes were detected mostly via far-infrared and THz absorption;19–23 however, the development in recent years of volume holographic gratings (VHG) has enabled more facile use of Raman spectroscopy for this purpose.24–26 LF-Raman possesses a number of advantages over other methods: first, the high spectral bandwidth, which enables recording the vibrational frequencies from 5 cm-1 to 4000 cm-1, in a single measurement, in contrast to THz and Far-IR spectroscopies.27 Second, LF-Raman is insensitive to the presence of water, which in other methods adds a significant background signal. Third, both Stokes and anti-Stokes shifts are observed, thereby improving signal discrimination. Finally, the crystallite sample can be as small as the laser-beam diameter in the focus.18,27 LFV assignment for a specific mode is usually obtained using several theoretical calculations.23,28,29 Up to now, relatively little data is available on the assignment of LFRaman shifts. LF-Raman can be used as an improved method for crystal chirality investigation, particularly for distinguishing between racemic and enantiopure organic crystals. Chirality plays a key role in the crystallization of chiral molecules that are normally divided into two main crystallization forms. For pure enantiomers, the resulting crystal is composed of only one enantiomer. This crystal is defined as being enantiopure, i.e. an enantiomerically pure crystal. For racemic compounds, the final crystal contains equal amounts of each enantiomer, and is called a racemate. Most racemic mixtures crystallize as racemic crystals; however, in some cases (5 to 10% in nature), racemic compounds

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crystallize in a conglomerate form that is a mixture of homochiral crystals.30 According to Wallach’s rule, racemic crystals are denser than the corresponding enantiopure crystals;31,32 this is generally explained by different hydrogen-bond networking. In the enantiopure case, crystallization is dictated by the chirality and thus the hydrogen-bond network is limited, while for racemic crystals, many more modes of hydrogen bonding are available and eventually define the structure.33,34 These significant differences in the crystalline structures and intermolecular interactions of racemic and enantiopure crystals lead to distinct vibrational modes that are detectable by LF-Raman. In particular, we show that LF-Raman provides completely different spectra for racemic and enantiopure crystals. Moreover, LF-Raman offers faster and more sensitive chiral characterization in crystals than currently used methods, enabling facile measurements for micro-crystals and detection of defects in chiral crystals. Materials and Methods Materials. The following analytical-grade chemicals were purchased from AldrichSigma and measured after recrystallization: L-phenylalanine (>99%), D-phenylalanine (>98%), DL-phenylalanine (99%), L-leucine (>98%), D-leucine (99%), DL-leucine (>99%), L-alanine (>99.5%), D-alanine (>98%), DL-alanine (>99%), L-valine (>99.5%), D-valine (>98%), DL-valine (>99%), L-aspartic acid (>99%), D-aspartic acid (99%), DL-aspartic acid (>99%), L-arginine (>98%), D-arginine (>98%), DL-arginine (>97%), L-threonine

(>99.5%),

D-threonine

(>98%),

and

DL-threonine

(>95%).

For

recrystallization, double distilled water was used, obtained by purifying water through a Barnstead EASY Pure II osmosis system (Thermo Fisher Scientific Inc.).

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Recrystallization

of

phenylalanine.

Supersaturated

stock

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solutions

of

D/L-

phenylalanine (Phe) and DL-Phe were prepared by dissolving 30 mg/mL in water, warmed up to 60 ºC. The solutions were stirred for 30 min and then left to cool and crystallize at room temperature for 24 hours. The crystals were smashed and examined by XRD, SEM, Raman, and LFV-Raman. XRD data plots are presented in the supplementary information, Fig. S1. Recrystallization of leucine. Supersaturated stock solutions of D/L-leucine (Leu) and DL-Leu were prepared by dissolving 80 mg/mL in water, warmed up to 60 ºC. The solutions were treated, as described above, for solutions of Phe. XRD data plots are presented in the supplementary information, Fig. S2. Recrystallization of Alanine. Supersaturated stock solutions of D/L-Alanine (Ala) and DL-Ala were prepared by dissolving 250 mg/mL in warm water, as described above. XRD data plots are presented in the supplementary information, Fig. S3. Recrystallization of Valine. Supersaturated stock solutions of D/L-Valine (Val) and DLVal were prepared by dissolving 115 mg/mL in warm water, as described above. XRD data plots are presented in supplementary Fig. S4. Recrystallization of Aspartic acid. Supersaturated stock solutions of D/L-Aspartic acid (Asp) and DL-Asp were prepared by dissolving 13.5 mg/mL in warm water, as described above. XRD data plots are presented in supplementary Fig. S5. Recrystallization of Arginine. Supersaturated stock solutions of D/L-Arginine (Arg) and DL-Arg were prepared by dissolving 250 mg/mL in warm water, as described above. XRD data plots are presented in supplementary Fig. S6.

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Recrystallization of Threonine. Supersaturated stock solutions of D/L-Threonine (Thr) and DL-Thr were prepared by dissolving 300 mg/mL in warm water, as described above. XRD data plots are presented in supplementary Fig. S7. Structure drawings. Drawings of the amino acid structures were performed by the Mercury program using CIF numbers from the ConQuest program. The CIF of the Leu structures are: LEUCIN02 and DLLEUC02, and those of the Asp structures are: LASPRT03 and DLASPA12. XRD measurements. The phases of the crystals were studied by powder XRD using a Bruker AXS D8 Advance diffractometer with Cu Ka (λ = 1.5418 Å) operating at 40 kV/40 mA, and collecting from 2θ =10° to 80°. SEM measurements. The crystal shape was studied using the Environmental Scanning Electron Microscope Quanta FEG 250, FEI. All crystal images are presented in Fig. S8. Raman of the chemical fingerprint range. Regular Raman scattering in the chemical fingerprint range were taken using a micro-Raman instrument (HORIBA Scientific LabRAM HR) in air at room temperature. The crystals were excited by a laser with an excitation wavelength of λex = 532 nm with 30 mW of optical power with an acquisition time of 10 seconds and a grating groove density of 600 g/mm. These Raman spectra are presented in supplementary Figs. S9-S15. The spectral baselines were shifted for presentation purposes. LF-Raman. LF-Raman measurements were taken using an integrated laser and VHG filter system (ONDAX, XLF-MICRO 532nm) with 50 mW of optical power at an excitation wavelength of λex = 532 nm. The laser output was routed into a lab-built

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microscope, and the Raman signal was fiber-coupled into an imaging spectrometer (Princeton Instruments, SP-2500i) with EM-CCD camera (Princeton Instruments, ProEM 16002). Acquisition times of 100 seconds were used and the grating groove density of 1800 g/mm was selected. In order to avoid edge effects and LF-Raman signals from air, single crystals were measured by first setting the focus to the top crystal surface and then slightly lowering the focal plane into the depth of the crystal. At least five different crystals were measured for every material, and for each crystal, 3 different focal areas were measured. The spectral baselines were shifted for presentation purposes. Results and Discussion In order to investigate the LFV modes of racemic and enantiopure crystals in amino acids, we chose representative amino acids for each group: Phe and Leu for hydrophobic, Ala and Val for small aliphatic, Asp and Arg for hydrophilic, and Thr for a conglomerate system. XRD, SEM, and Raman measurements were performed on D, L, and DL crystals in order to verify purity and crystallinity prior to LF-Raman measurements. Regular Raman spectra were measured in order to compare the spectra of the chemical fingerprints of the racemic and enantiopure crystals. XRD data plots, SEM images, and regular Raman spectra are included in the supplementary information. D and L crystals of the same amino acid showed similar XRD data plots as expected;35 the similarity between the D and L crystal structures produces similar LF-Raman spectra for recrystallized materials that avoids contaminations, and when the focal area measures the core of the crystal. In our work, the L crystal form of each amino acid was used to represent the XRD data plots, Raman spectra, and LF-Raman spectra of both D and L crystals.

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In molecular lattices, LFV modes represent the molecular degrees of freedom, translational and rotational motions, accompanied by intermolecular interactions and phonons associated with lattice vibrations.27,36,37 For layered materials, the LFV spectrum exhibit layer breathing and shear modes.38–40 Overall, LF-Raman provides information on hydrogen bonding, hydrophobic interactions, and degrees of freedom in crystalline structures. Previous publications have stated that vibrational modes in the region of 0-100 cm-1 are generated by global fluctuations (e.g., molecular degrees of freedom and shear modes) and phonons, while vibrational modes in the region of 100-200 cm-1 are generated by more localized intermolecular interactions.41,42 In enantiopure crystals, crystallization is dictated by the chirality, and the hydrogen-bond network is limited, while in racemic crystals, many more modes of hydrogen bonds are available and eventually define the structure.33,34 It is well established, namely via THz absorption, that hydrogen bonds show LFV modes. Within a crystal, hydrogen-bond stretching modes exist and can be found in the range of 100-200 cm-1.43–45 All crystal structures of hydrophobic amino acids have the same basic organization: the side chains form hydrophobic layers and the polar head groups form hydrophilic layers, and a hydrogen bond network is formed between each pair of adjacent hydrophilic layers.46 The strong network of the racemic crystal is energetically preferred, and thus leads to a weakening of the hydrophobic interactions among the side chains. This enables the dominance of shear modes between neighboring bilayers and rotation modes of the side chains; these modes have been reported in the range of 0-100 cm-1.36,38– 40

In the case of hydrophilic amino acids, an additional hydrogen-bond network is created

which, similar to the main network, is significantly stronger for the racemate. This

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network populates more stretching modes of hydrogen bonds but also limits other modes in the crystal motion. These vibrational dynamics in turn affect the phonon dispersion relation of the amino acid crystal lattice.

a

b

c

d

Fig. 1: A scheme presenting the structures along with hydrogen bonds of (a) L-Leu, (b) DL-Leu, (c) L-Asp, and (d) DL-Asp. The structures represent 2*2*2 unit cells. Red balls represent oxygen atoms, purple represents nitrogen atoms, red dashed lines represent oxygen-hydrogen bonds, and turquoise dashed lines represent nitrogen-hydrogen bonds.

Figure 1 presents different crystal structures of two representative amino acids, Leu for hydrophobic and Asp for hydrophilic residues. The L-Leu structure (Figure 1a) presents the hydrogen-bond network along with the hydrophobic layers. For DL-Leu, the

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stronger hydrogen-bond network leads to the weakening of the van der Waals bonds as observed in Figure 1b. Figure 1c shows that L-Asp forms an additional hydrogen-bond network between the side chains, and DL-Asp, which is not limited by the chirality, forms stronger networks in a more densely packed crystal structure (Figure 1d).

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Fig. 2: LF-Raman spectra of (a) L-Phe, DL-Phe crystals and (b) L-Leu, DL-Leu crystals.

Figure 2 presents LF-Raman spectra of Phe (Figure 2a) and Leu (Figure 2b) in both enantiopure and racemic forms. For each amino acid, the disparity between the enantiopure and racemic spectra reflects different crystal structures. The LFV modes observed for L-Phe are at: 11, 25, 41, 48, 67, 102, 147, and 209 cm-1. For DL-Phe, the observed modes are at: 17, 29, 58, 77, 107, 125, 144, 182, and 214 cm-1. The LFV modes observed for L-Leu are at: 28, 42, 66, 89, 109, and 170 cm-1. For DL-Leu, the observed modes are at: 38, 55, 80, 88, 124, 139, 167, and 215 cm-1. In some cases, two close shifts appear as one broad shift or as a shift with a shoulder. A comparison between the LFRaman spectra of racemic and enantiopure crystals shows that in both cases more modes exist in the hydrogen-bond stretching region (100-200 cm-1) for the racemic crystal. However, the lower energy modes have higher intensity, probably due to stronger shear modes and side-chain rotation modes. The scattering spectrum shows higher intensity in

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the lower energy modes rather than the higher energy modes when compared to the enantiopure crystals. Phe demonstrated more significant differences than Leu, probably because Phe is a more complex molecule.

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L-Val DL-Val

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Fig. 3: LF-Raman spectra of (a) L-Ala, DL-Ala crystals and (b) L-Val, DL-Val crystals.

This trend applies to small aliphatic amino acids, Ala and Val, as well, as shown in the LF-Raman spectra of Figure 3. The LFV modes observed for L-Ala (Figure 3a) are at: 39, 48, 74, 114, 140, 160, and 194 cm-1. For DL-Ala (Figure 3b), the observed modes are at: 40, 74, 91, 101, 119, 151, 162, 187, and 230 cm-1. The LFV modes observed for LVal are at: 36, 48, 56, 73, 93, 135, 160, 181, and 208 cm-1. For DL-Val, the observed modes are at: 47, 60, 68, 106, 117, 134, 176, and 215 cm-1. The racemic spectra of both amino acids are comprised of more Raman modes at lower frequencies, which can be assigned to shear modes between hydrophobic layers and to rotation modes of the side chains that are populated only in the racemic crystal. The scattering intensity of the racemic crystals demonstrates higher population in the lower energy modes. Also, for Ala, the most populated mode is probably shared for both racemic and enantiopure structures. The peak demonstrates a blue shift from 114 cm-1 in the enantiopure crystal to 119 cm-1 in the racemic counterpart that can be explained by the higher spring constant

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for this mode in the racemic crystal; this can further support the assumption that this peak is assigned to hydrogen-bond stretching.

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Fig. 4: LF-Raman spectra of (a) L-Asp, DL-Asp crystals and (b) L-Arg, DL-Arg crystals.

Figure 4 presents the LF-Raman spectra of Asp (Figure 4a) and Arg (Figure 4b). The LFV modes observed for L-Asp are at: 45 and 88 cm-1. For DL-Asp, the observed modes are at: 64, 83, 101, 115, 129, 165, 182, and 215 cm-1. The LFV modes observed for L-Arg are at: 32, 60, 90, 119, 151, and 195 cm-1. For DL-Arg, the observed modes are at: 42, 51, 66, 100, 122, 144, 165, 192, and 256 cm-1. These spectra show the opposite trend in comparison to Phe, Leu, Ala, and Val, i.e. more highly populated modes are observed in the higher region of LFV, at 100-200 cm-1. This can be explained by the additional hydrogen-bond network that is created by the polar substituent of the amino acid side chain. This network populates more stretching modes of hydrogen bonds and also limits other modes in the crystal motion, such as shear modes of neighboring hydrophobic layers and rotation modes of the side chains. Therefore, amino acids with hydrophilic side chains show a different distribution in scattering intensity relative to hydrophobic amino acids when comparing racemic and enantiopure crystals. We believe that the shifts at 88 cm-1 for L-Asp and 83 cm-1 for DL-Asp represent the same LFV

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mode; the red shift in the racemic crystal indicates a lower spring constant that supports our assumption that this mode is assigned to one of the limited crystal motions. For Arg (Figure 4b), the differences between the spectra of the racemic and enantiopure crystals are smaller. Possible reasons could be the longer side chain that decreases the limitations in the crystal motion, and the weaker hydrogen bonds formed by this molecule.

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Fig. 5: LF-Raman spectra of L-Thr and DL-Thr crystals.

Threonine was measured as representative of a conglomerate crystal; the LFRaman spectra of L-Thr and DL-Thr (Figure 5) show the same shifts at: 49, 73, 94, 104, 130, 157, 196, and 215 cm-1. A conglomerate crystal is composed of an equimolar mixture of two crystalline enantiomers, and is therefore expected to have the same modes for the enantiopure and racemic crystals as confirmed by the data. Conclusions In this study, we presented a new method for characterizing the chirality of crystals composed of chiral molecules using LF-Raman spectroscopy. We showed that with LF-Raman, it is possible to distinguish between racemic and enantiopure crystals of amino acids. LF-Raman spectra of racemic and enantiopure crystals were significantly different, attributable to different hydrogen-bond networks in the chiral crystal structures.

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This method represents an advance in identification and recognition of crystal chirality, a field that, in solids, suffers from very limited available techniques for chirality measurements. LF-Raman spectroscopy offers fast and sensitive chiral characterization compared to currently used methods. Moreover, LF-Raman enables measurement of single micro-crystals and defect detection. We believe this method can be applied to many other crystalline structures, and can open a new frontier in the study of chiral crystals and surfaces. Finally, the possibility to measure the LF-Raman spectra of chiral surfaces enables interesting research, such as investigating the correlation between crystal phonons and structures. Supporting Information The Supporting Information is available free of charge on the ACS Publications website. The PDF file includes XRD data plots, SEM images of crystals, and regular Raman spectra of the investigated amino acids. Acknowledgements We gratefully acknowledge the technical support given by ONDAX. This research was financially supported by the Israeli National Nanotechnology Initiative (INNI) Focal Technology Area Project, FTA (grant number 458004).

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References (1)

Mohan, S. J.; Mohan, E. C.; Yamsani, M. R. Chirality and Its Importance in Pharmaceutical Field- An Overview. Int. J. Pharm. Sci. Nanotechnol. 2009, 1, 309–316.

(2)

Davies, N. M.; Wei Teng, X. Importance of Chirality in Drug Therapy and Pharmacy Practice: Implications for Psychiatry. Adv. Pharm. 2003, 1, 242–252.

(3)

Davankov, V. a. Analytical Chiral Separation Methods. Pure App. Chem. 1997, 69, 1469–1474.

(4)

Pirkle, W. H.; Pochapsky, T. C. Considerations of Chiral Recognition Relevant to the Liquid Chromatography Separation of Enantiomers. Chem. Rev. 1989, 89, 347–362.

(5)

Schurig, V. Separation of Enantiomers by Gas Chromatography. J. Chromatogr. A 2001, 906, 275–299.

(6)

Gasparrini, F.; Misiti, D.; Villani, C. High-Performance Liquid Chromatography Chiral Stationary Phases Based on Low-Molecular-Mass Selectors. J. Chromatogr. A 2001, 906, 35–50.

(7)

Petersson, P.; Markides, K. E. Chiral Separations Performed by Supercritical Fluid Chromatography. J. Chromatogr. A 1994, 666, 381–394.

(8)

Schulte, M.; Strube, J. Preparative Enantioseparation by Simulated Moving Bed Chromatography. J. Chromatogr. A 2001, 906, 399–416.

(9)

Freedman, T. B.; Cao, X.; Dukor, R. K.; Nafie, L. A. Absolute Configuration Determination of Chiral Molecules in the Solution State Using Vibrational Circular Dichroism. Chirality 2003, 15, 743–758.

(10)

Foerier, S.; Kolmychek, I. A.; Aktsipetrov, O. A.; Verbiest, T.; Valev, V. K. Optical Second Harmonic Generation Chiral Spectroscopy. ChemPhysChem 2009, 10, 1431–1434.

(11)

Verbiest, T.; Kauranen, M.; Persoons, A.; Ikonen, M.; Kurkela, J.; Lemmetyinen, H.; Krauss, T.; Brus, L. Second-Order Nonlinear Optical Properties of Chiral Thin Films. J. Mater. Chem. 1999, 9, 2005–2012.

(12)

Medina, D. D.; Goldshtein, J.; Margel, S.; Mastai, Y. Enantioselective Crystallization on Chiral Polymeric Microspheres. Adv. Funct. Mater. 2007, 17, 944–950.

(13)

Nie, Q.; Gong, J. B.; Wang, J. K.; Wang, S. Formation of Mixed Crystals in Crystallization of 11alpha -Hydroxy-16alpha ,17alpha -Epoxyprogesterone and 16alpha ,17alpha -Epoxyprogesterone. Ind. Eng. Chem. Res. 2006, 45, 432–437.

(14)

King, M. D.; Buchanan, W. D.; korter, T. M. Understanding the Terahertz Spectra

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of Crystalline Pharmaceuticals: Terahertz Spectroscopy and Solid-State Density Functional Theory Study of (S)-(+)-Ibuprofen and (RS)-Ibuprofen. J. Pharm. Sci. 2011, 100, 1116–1129. (15)

Yamaguchi, M.; Miyamaru, F.; Yamamoto, K.; Tani, M.; Hangyo, M. Terahertz Absorption Spectra of L-, D-, and DL-Alanine and Their Application to Determination of Enantiometric Composition. Appl. Phys. Lett. 2005, 86, 53903– 53905.

(16)

Nishikiori, R.; Yamaguchi, M.; Takano, K.; Enatsu, T.; Tani, M.; de Silva, U. C.; Kawashita, N.; Takagi, T.; Morimoto, S.; Hangyo, M.; et al. Application of Partial Least Square on Quantitative Analysis of L-, D-, and DL-Tartaric Acid by Terahertz Absorption Spectra. Chem. Pharm. Bull. (Tokyo). 2008, 56, 305—307.

(17)

Bordallo, H. N.; Kolesov, B. A.; Boldyreva, E. V.; Juranyi, F. Different Dynamics of Chiral and Racemic (L- and DL-) Serine Crystals: Evidenced by Incoherent Inelastic Neutron and Raman Scattering. J. Am. Chem. Soc. 2007, 129, 10984– 10985.

(18)

Adibi, A.; Asakura, T.; Hänsch, H.; Kamiya, T.; Krausz, F.; Monemar, B. A.; Venghaus, H.; Weber, H.; Weinfurter, H. Theory of Crystal Phonons. In Terahertz Spectroscopy and Imaging; 2013; pp 155–177.

(19)

Mittleman, D. M. Frontiers in Terahertz Sources and Plasmonics. Nat. Publ. Gr. 2013, 7, 666–669.

(20)

El Haddad, J.; Bousquet, B.; Canioni, L.; Mounaix, P. Review in Terahertz Spectral Analysis. Trends Anal. Chem. 2013, 44, 98–105.

(21)

Hielscher, R.; Friedrich, T.; Hellwig, P. Far-and Mid-Infrared Spectroscopic Analysis of the Substrate-Induced Structural Dynamics of Respiratory Complex I. ChemPhysChem 2011, 12, 217–224.

(22)

Duncan, W. D.; Williams, G. P. Infrared Synchrotron Radiation from Electron Storage Rings. Appl. Opt. 1983, 22, 2914–2923.

(23)

Hoshina, H.; Ishii, S.; Yamamoto, S.; Morisawa, Y.; Sato, H.; Uchiyama, T.; Ozaki, Y.; Otani, C. Terahertz Spectroscopy in Polymer Research: Assignment of Intermolecular Vibrational Modes and Structural Characterization of poly(3Hydroxybutyrate). IEEE Trans. Terahertz Sci. Technol. 2013, 3, 248–258.

(24)

Ali, M. M.; Lim, K. S.; Becir, A.; Lai, M. H.; Ahmad, H. Optical Gaussian Notch Filter Based on Periodic Microbent Fiber Bragg Grating. IEEE Photonics J. 2014, 6, 1–8.

(25)

Raymond, S. G.; Wagner, P.; Panczyk, M.; Williams, G. V. M.; Stevens, K. J.; Monfils, I.; Hirst, D.; Whaanga, J.; Kutuvantavida, Y.; Bhuiyan, M. D. H.; et al. Development of Fibre Bragg Grating Based Strain/temperature Sensing System. Org. Photonic Mater. Devices XIV 2012, 8258, 1–8.

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(26)

Glebov, A. L.; Mokhun, O.; Rapaport, A.; Vergnole, S.; Smirnov, V.; Glebov, L. B. Volume Bragg Gratings as Ultra-Narrow and Multiband Optical Filters. Proc. SPIE 2012, 8428, 84280C:1–11.

(27)

Parrott, E. P. J.; Zeitler, J. A. Terahertz Time-Domain and Low-Frequency Raman Spectroscopy of Organic Materials. Appl. Spectrosc. 2015, 69, 1–25.

(28)

Jepsen, P. U.; Clark, S. J. Precise Ab-Initio Prediction of Terahertz Vibrational Modes in Crystalline Systems. Chem. Phys. Lett. 2007, 442, 275–280.

(29)

Ogawa, Y.; Hayashi, ichiro; Oikawa, M.; Otani, C.; Kawase, K.; Fukui, S.; Feizi, T.; Galustian, C.; Lawson, A. M.; Chai, W. Interference Terahertz Label-Free Imaging for Protein Detection on a Membrane. Sci. Curr. Opin. Chem. Biol. Angew. Chem. Int. Ed. Opt. EXPRESS 1995, 270, 467–470.

(30)

Addadi, L.; Berkovitch-Yellin, Z.; Domb, N.; Gati, E.; Lahav, M.; Leiserowitz, L. Resolution of Conglomerates by Stereoselective Habit Modifications. Nature 1982, 296, 21–26.

(31)

Brock, C. P.; Schweizer, W. B.; Dunitz, J. D. On the Validity of Wallach Rule - on the Density and Stability of Racemic Crystals Compared with Their Chiral Counterparts. J. Am. Chem. Soc. 1991, 113, 9811–9820.

(32)

Seibel, J.; Parschau, M.; Ernst, K. H. From Homochiral Clusters to Racemate Crystals: Viable Nuclei in 2D Chiral Crystallization. J. Am. Chem. Soc. 2015, 137, 7970–7973.

(33)

Kennedy, A. R.; Morrison, C. A.; Briggs, N. E. B.; Arbuckle, W. Density and Stability Differences between Enantiopure and Racemic Salts: Construction and Structural Analysis of a Systematic Series of Crystalline Salt Forms of Methylephedrine. Cryst. Growth Des. 2011, 11, 1821–1834.

(34)

Dunitz, J. D.; Gavezzotti, A. Proteogenic Amino Acids: Chiral and Racemic Crystal Packings and Stabilities. J. Phys. Chem. B 2012, 116, 6740–6750.

(35)

Lidor-Shalev, O.; Pliatsikas, N.; Cramiel, Y.; Patsalas, P.; Mastai, Y. Chiral MetalOxide Nanofilms by Cellulose Template Using Atomic Layer Deposition Process. ACS Nano 2017, 11, 4753–4759.

(36)

Williams, M. R. C.; Aschaffenburg, D. J.; Ofori-Okai, B. K.; Schmuttenmaer, C. A. Intermolecular Vibrations in Hydrophobic Amino Acid Crystals: Experiments and Calculations. J. Phys. Chem. B 2013, 117, 10444–10461.

(37)

Niehues, G.; Heyden, M.; Schmidt, D. A.; Havenith, M.; Berzofsky, J. A.; DeLisi, C.; Haller, E. E.; Havenith, M. Exploring Hydrophobicity by THz Absorption Spectroscopy of Solvated Amino Acids. Faraday Discuss. 2011, 150, 193–207.

(38)

He, R.; Baren, J. Van; Yan, J.; Xi, X.; Ye, Z.; Ye, G.; Lu, I.; Leong, S. M.; Lui, C. H. Interlayer Breathing and Shear Modes in NbSe2 Atomic Layers. 2D Mater. 2016, 2, 1–16.

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(39)

Zhao, Y.; Luo, X.; Li, H.; Zhang, J.; Araujo, P. T.; Gan, C. K.; Wu, J.; Zhang, H.; Quek, S. Y.; Dresselhaus, M. S.; et al. Interlayer Breathing and Shear Modes in Few-Trilayer MoS2 and WSe2. Nano Lett. 2013, 13, 1007–1015.

(40)

Cong, C.; Yu, T. Enhanced Ultra-Low-Frequency Interlayer Shear Modes in Folded Graphene Layers. Nat. Commun. 2014, 5, 4709–4715.

(41)

Kalanoor, B. S.; Ronen, M.; Oren, Z.; Gerber, D.; Tischler, Y. R. New Method to Study the Vibrational Modes of Biomolecules in the Terahertz Range Based on a Single-Stage Raman Spectrometer. ACS Omega 2017, 2, 1232–1240.

(42)

Woods, K. N.; Pfeffer, J.; Dutta, A.; Klein-Seetharaman, J.; Haak, J. R. Vibrational Resonance, Allostery, and Activation in Rhodopsin-like G Protein-Coupled Receptors. Sci. Rep. 2016, 6, 37290–37306.

(43)

Takahashi, M. Terahertz Vibrations and Hydrogen-Bonded Networks in Crystals. Crystals 2014, 4, 74–103.

(44)

Saito, S.; Inerbaev, T. M.; Mizuseki, H.; Igarashi, N.; Note, R.; Kawazoe, Y. Terahertz Phonon Modes of an Intermolecular Network of Hydrogen Bonds in an Anhydrous β-D-Glucopyranose Crystal. Chem. Phys. Lett. 2006, 423, 439–444.

(45)

Allis, D. G.; Fedor, A. M.; Korter, T. M.; Bjarnason, J. E.; Brown, E. R. Assignment of the Lowest-Lying THz Absorption Signatures in Biotin and Lactose Monohydrate by Solid-State Density Functional Theory. Chem. Phys. Lett. 2007, 440, 203–209.

(46)

Görbitz, C. H.; Leirvåg, A. B.; Jacobsen, Ø. Hydrogen Bond Architecture in Crystal Structures of N-Alkylated Hydrophobic Amino Acids. CrystEngComm 2014, 16, 9631–9637.

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