Structural and Dynamic Properties of Amorphous Solid Dispersions: The Role of Solid-State Nuclear Magnetic Resonance Spectroscopy and Relaxometry

Amorphous solid dispersions (ASDs) are one of the frontier strategies to improve solubility and dissolution rate of poorly soluble drugs and hence tackling the growing challenges in oral bioavailability. Pharmaceutical performance, physicochemical stability, and downstream processability of ASD largely rely on the physical structure of the product. This necessitates in-depth characterization of ASD microstructure. Solid-state nuclear magnetic resonance (SS-NMR) techniques bear the ultimate analytical capabilities to provide the molecular level information on the dynamics and phase compositions of amorphous dispersions. SS-NMR spectroscopy/relaxometry, as a single and nondestructive technique, can reveal diverse and critical structural information of complex ASD formulations that are barely amenable from any other existing technique. The purpose of the current article is to review the recent most important studies on various sophisticated and information-rich one-dimensional and two-dimensional SS-NMR spectroscopy/relaxometry for the analysis of molecular mobility, miscibility, drug–carrier interactions, crystallinity, and crystallization in ASD. Some specific examples on microstructural elucidations of challenging ASD using multidimensional and multinuclear SS-NMR are presented. Additionally, some relevant examples on the utility of solution-NMR and NMR-imaging techniques for the investigation of the dissolution behavior of ASD are gathered. © 2014 Wiley Periodicals, Inc. and the American Pharmacists Association J Pharm Sci 103:2635–2662, 2014 Amorphous solid dispersions (ASDs) are one of the frontier strategies to improve solubility and dissolution rate of poorly soluble drugs and hence tackling the growing challenges in oral bioavailability. Pharmaceutical performance, physicochemical stability, and downstream processability of ASD largely rely on the physical structure of the product. This necessitates in-depth characterization of ASD microstructure. Solid-state nuclear magnetic resonance (SS-NMR) techniques bear the ultimate analytical capabilities to provide the molecular level information on the dynamics and phase compositions of amorphous dispersions. SS-NMR spectroscopy/relaxometry, as a single and nondestructive technique, can reveal diverse and critical structural information of complex ASD formulations that are barely amenable from any other existing technique. The purpose of the current article is to review the recent most important studies on various sophisticated and information-rich one-dimensional and two-dimensional SS-NMR spectroscopy/relaxometry for the analysis of molecular mobility, miscibility, drug–carrier interactions, crystallinity, and crystallization in ASD. Some specific examples on microstructural elucidations of challenging ASD using multidimensional and multinuclear SS-NMR are presented. Additionally, some relevant examples on the utility of solution-NMR and NMR-imaging techniques for the investigation of the dissolution behavior of ASD are gathered. © 2014 Wiley Periodicals, Inc. and the American Pharmacists Association J Pharm Sci 103:2635–2662, 2014 The use of the amorphous state of poorly water-soluble active pharmaceutical ingredients (APIs) can provide remarkably higher aqueous solubility/dissolution rate compared with the crystalline counterpart.1.Hancock B.C. Zografi G. Characteristics and significance of the amorphous state in pharmaceutical systems.J Pharm Sci. 1997; 86: 1-12Abstract Full Text PDF PubMed Google Scholar Owing to the higher molecular mobility and energy state, this thermodynamic benefit of solubility is often negated by the inherent propensity of metastable amorphous form to undergo nucleation and crystal growth through solid-state or solution-mediated routes.2.Sun D.D. Lee P.I. Evolution of supersaturation of amorphous pharmaceuticals: The effect of rate of supersaturation generation.Mol Pharm. 2013; 10: 4330-4346Crossref PubMed Scopus (0) Google Scholar, 3.Murdande S.B. Pikal M.J. Shanker R.M. Bogner R.H. Solubility advantage of amorphous pharmaceuticals: I. A thermodynamic analysis.J Pharm Sci. 2010; 99: 1254-1264Abstract Full Text Full Text PDF PubMed Scopus (159) Google Scholar Therefore, the amorphous form of the pure API is seldom used in the drug product. Amorphous composite systems wherein amorphous API is dispersed, to the best, at molecular level in the hydrophilic polymer matrices constitutes amorphous solid dispersion (ASD).4.Huang Y. Dai W.-G. Fundamental aspects of solid dispersion technology for poorly soluble drugs.Acta Pharm Sin B. 2014; 4: 18-25Crossref PubMed Google Scholar Polymers used are generally amorphous and with high glass transition temperature (Tg). This leads to significant reduction in molecular mobility of API and hence stabilizes against crystallization. In addition, specific intermolecular interactions between API and polymer in ASD aid to inhibit nucleation and crystal growth. These benefits of ASD have been exploited as one of the key approaches of oral bioavailability improvement of poorly water soluble drugs.5.Gurunath S. Pradeep Kumar S. Basavaraj N.K. Patil P.A. Amorphous solid dispersion method for improving oral bioavailability of poorly water-soluble drugs.J Pharm Res. 2013; 6: 476-480Abstract Full Text Full Text PDF Scopus (0) Google Scholar, 6.Van den Mooter G. The use of amorphous solid dispersions: A formulation strategy to overcome poor solubility and dissolution rate.Drug Discov Today. 2012; 9: e79-e85Crossref Scopus (111) Google Scholar Despite distinct advantages, very few commercial formulations based on ASD have reached the market through more than four decades of academic and industrial research in this field.6.Van den Mooter G. The use of amorphous solid dispersions: A formulation strategy to overcome poor solubility and dissolution rate.Drug Discov Today. 2012; 9: e79-e85Crossref Scopus (111) Google Scholar The key hurdles associated with the development of ASD are poor understanding of the physical structures and the relationship of the latter with the pharmaceutical performance and physical stability of the product. ASD manufactured by industrial processes such as spray drying (SD), hot-melt extrusion (HME), milling, and so on generally consists of API dispersed in the polymer at the concentration level higher than solid-state solubility of API in the particular polymer.7.Janssens S. Zeure A. Paudel A. Humbeeck J. Rombaut P. Mooter G. Influence of preparation methods on solid state supersaturation of amorphous solid dispersions: A case study with itraconazole and eudragit E100.Pharm Res. 2010; 27: 775-785Crossref PubMed Scopus (0) Google Scholar, 8.Paudel A. Nies E. Van den Mooter G. Relating hydrogen-bonding interactions with the phase behavior of naproxen/PVP K 25 solid dispersions: Evaluation of solution-cast and quench-cooled films.Mol Pharm. 2012; 9: 3301-3317Crossref PubMed Scopus (0) Google Scholar As shown in Figure 1, the supersaturated ASD systems exhibit (partial) amorphous–amorphous demixing and/or nucleation and crystal growth of API upon exposure to elevated heat, humidity, or mechanical stress through downstream processing, in vitro or in vivo environment, or storage.9.Paudel A. Van Humbeeck J. Van den Mooter G. Theoretical and experimental investigation on the solid solubility and miscibility of naproxen in poly(vinylpyrrolidone).Mol Pharm. 2010; 7: 1133-1148Crossref PubMed Scopus (62) Google Scholar, 10.Rumondor A.C.F. Marsac P.J. Stanford L.A. Taylor L.S. Phase behavior of poly(vinylpyrrolidone) containing amorphous solid dispersions in the presence of moisture.Mol Pharm. 2009; 6: 1492-1505Crossref PubMed Scopus (0) Google Scholar, 11.Marsac P.J. Rumondor A.C.F. Nivens D.E. Kestur U.S. Stanciu L. Taylor L.S. Effect of temperature and moisture on the miscibility of amorphous dispersions of felodipine and poly(vinyl pyrrolidone).J Pharm Sci. 2010; 99: 169-185Abstract Full Text Full Text PDF PubMed Scopus (98) Google Scholar Generally, these stresses induce thermodynamic (binodal/spinodal) immiscibility, weakening of stabilizing interactions between API and carrier (e.g., ionic, H-bonding), enhanced molecular mobility (α, β processes), or combinations.12.Ayenew Z. Paudel A. Van den Mooter G. Can compression induce demixing in amorphous solid dispersions? A case study of naproxen-PVP K25.Euro J Pharm Biopharm. 2012; 81: 207-213Crossref PubMed Scopus (0) Google Scholar This eventually result into the loss of the claimed solubility advantages of these enabling formulations. It has been demonstrated through several researches that the physical stability of ASD can be markedly improved by generating molecularly mixed amorphous dispersions wherein API and polymer molecules are at the closest proximity and involved in stronger molecular interactions such as ionic, H-bonding, dipolar interactions, or electrostatic interactions.13.Marsac P. Shamblin S. Taylor L. Theoretical and practical approaches for prediction of drug-polymer miscibility and solubility.Pharm Res. 2006; 23: 2417-2426Crossref PubMed Scopus (0) Google Scholar, 14.Janssens S. Van den Mooter G. Review: Physical chemistry of solid dispersions.J Pharm Pharmacol. 2009; 61: 1571-1586Crossref PubMed Google Scholar There are very few analytical techniques that offer spatial capability to probe the miscibility at molecular level in complex ASD.5.Gurunath S. Pradeep Kumar S. Basavaraj N.K. Patil P.A. Amorphous solid dispersion method for improving oral bioavailability of poorly water-soluble drugs.J Pharm Res. 2013; 6: 476-480Abstract Full Text Full Text PDF Scopus (0) Google Scholar, 15.Guo Y. Shalaev E. Smith S. Solid-state analysis and amorphous dispersions in assessing the physical stability of pharmaceutical formulations.TrAC Trend Anal Chem. 2013; 49: 137-144Crossref Scopus (0) Google Scholar This is one of the key hurdles in identifying, at nanoscopic level, whether API molecules are dispersed at molecular level (glass solution), present as amorphous clusters (amorphous solid suspension) in carrier matrices or present spatial heterogeneity of composition.16.Dahlberg C. Dvinskikh S.V. Schuleit M. Furó I. Polymer swelling, drug mobilization and drug recrystallization in hydrating solid dispersion tablets studied by multinuclear NMR microimaging and spectroscopy.Mol Pharm. 2011; 8: 1247-1256Crossref PubMed Scopus (0) Google Scholar For example, differential scanning calorimetry (DSC), the most commonly used technique for miscibility studies, can demonstrate immiscibility through multiple glass transitions only if the minimum size of phase-separated domains is approximately 30 nm or higher.17.Newman A. Engers D. Bates S. Ivanisevic I. Kelly R.C. Zografi G. Characterization of amorphous API:polymer mixtures using X-ray powder diffraction.J Pharm Sci. 2008; 97: 4840-4856Abstract Full Text Full Text PDF PubMed Scopus (0) Google Scholar, 18.Bikiaris D. Papageorgiou G.Z. Stergiou A. Pavlidou E. Karavas E. Kanaze F. Georgarakis M. Physicochemical studies on solid dispersions of poorly water-soluble drugs: Evaluation of capabilities and limitations of thermal analysis techniques.Thermochim Acta. 2005; 439: 58-67Crossref Scopus (0) Google Scholar, 19.Qian F. Huang J. Zhu Q. Haddadin R. Gawel J. Garmise R. Hussain M. Is a distinctive single Tg a reliable indicator for the homogeneity of amorphous solid dispersion?.Int J Pharm. 2010; 395: 232-235Crossref PubMed Scopus (0) Google Scholar, 20.Bates S. Zografi G. Engers D. Morris K. Crowley K. Newman A. Analysis of amorphous and nanocrystalline solids from their X-ray diffraction patterns.Pharm Res. 2006; 23: 2333-2349Crossref PubMed Scopus (0) Google Scholar Likewise, often used vibrational spectroscopy can only provide qualitative information on the molecular interactions for most of the ASD.21.Vogt F.G. Clawson J.S. Strohmeier M. Edwards A.J. Pham T.N. Watson S.A. Solid-state NMR analysis of organic cocrystals and complexes.Cryst Growth Des. 2008; 9: 921-937Crossref Google Scholar, 22.Rumondor A.F. Wikström H. Van Eerdenbrugh B. Taylor L.S. Understanding the tendency of amorphous solid dispersions to undergo amorphous-amorphous phase separation in the presence of absorbed moisture.AAPS PharmSciTech. 2011; 12: 1209-1219Crossref PubMed Scopus (0) Google Scholar Otherwise, it cannot provide direct quantitative information on the intermolecular proximity, molecular conformations, H-bonding geometries, and so on for most of the complex multicomponent systems. Furthermore, generally used techniques for molecular mobility studies such as dielectric spectroscopy (DES) lack the ability of speciation of the dynamic heterogeneity and probing the exact molecular origin of the primary and different secondary molecular relaxation processes.23.Kaatze U. Measuring the dielectric properties of materials. Ninety-year development from low-frequency techniques to broadband spectroscopy and high-frequency imaging.Meas Sci Technol. 2013; 24012005Crossref Scopus (0) Google Scholar Apart from confirming amorphicity, no direct and model-free information on the structural features can be extracted from the diffuse X-ray halo patterns conventionally obtained for amorphous powders.17.Newman A. Engers D. Bates S. Ivanisevic I. Kelly R.C. Zografi G. Characterization of amorphous API:polymer mixtures using X-ray powder diffraction.J Pharm Sci. 2008; 97: 4840-4856Abstract Full Text Full Text PDF PubMed Scopus (0) Google Scholar, 20.Bates S. Zografi G. Engers D. Morris K. Crowley K. Newman A. Analysis of amorphous and nanocrystalline solids from their X-ray diffraction patterns.Pharm Res. 2006; 23: 2333-2349Crossref PubMed Scopus (0) Google Scholar Analytical limits of the existing tools often hinder the assessment on the level of disorder and heterogeneity at the nanoscopic or mesoscopic structure in partially crystalline systems. Moreover, subtle alterations on these structural attributes of ASD originated from different manufacturing processes, process parameters, or other formulation variables, for the same system can prove deleterious to the physical stability and other quality attributes of the final product.24.Paudel A. Loyson Y. Van den Mooter G. An investigation into the effect of spray drying temperature and atomizing conditions on miscibility, physical stability, and performance of naproxen–PVP K 25 solid dispersions.J Pharm Sci. 2013; 102: 1249-1267Abstract Full Text Full Text PDF PubMed Scopus (0) Google Scholar Probing such delicate structural variation at molecular level needs strong analytical techniques.4.Huang Y. Dai W.-G. Fundamental aspects of solid dispersion technology for poorly soluble drugs.Acta Pharm Sin B. 2014; 4: 18-25Crossref PubMed Google Scholar Solid-state nuclear magnetic resonance (SS-NMR) techniques can be broadly divided into three categories, namely, relaxometry, spectroscopy, and imaging. In this review, the use of the term “spectroscopy” will be used to the measurements intended to yield the spectral properties (experimental observables that can be determined from spectra), whereas “relaxometry” will be referred to all the techniques concerning the measurement of nuclear relaxation times. These techniques have been proven as one of the supreme tools for elucidating the physical structure of ASD with the maximum details among the existing techniques.25.Vogt F.G. Solid-state NMR in drug discovery and development.in: Garrido L. Beckmann N. New applications of NMR in drug discovery and development. Cambridge, UK, The Royal Society of Chemistry2013: 43-100Crossref Google Scholar, 26.Paradowska K. Wawer I. Solid-state NMR in the analysis of drugs and naturally occurring materials.J Pharm Biomed Anal. [Epub ahead of print]. 2013; Google Scholar Expeditious application of SS-NMR techniques for the identification, characterization, and quantification of various solid forms of drug candidates (polymorphs, hydrate, solvate, salt, cocrystal, amorphous, mesomorphous, etc.) during drug development is eminent.21.Vogt F.G. Clawson J.S. Strohmeier M. Edwards A.J. Pham T.N. Watson S.A. 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Probing intermolecular interactions and nitrogen protonation in pharmaceuticals by novel 15N-edited and 2D14N-1H solid-state NMR.CrystEngComm. 2012; 14: 2654-2659Crossref Scopus (0) Google Scholar Beyond the applicability as “a solid-state-meter,” SS-NMR techniques can yield comprehensive structural information of multicomponent amorphous systems ranging from molecular dynamics, associations, intra- and intermolecular interactions, molecular miscibility, crystallinity, and so on.31.Kruk D. Privalov A. Medycki W. Uniszkiewicz C. Masierak W. Jakubas R. NMR studies of solid-state dynamics.in: Graham A.W. Annual reports on NMR spectroscopy. Academic Press, Amsterdam, The Netherlands2012: 67-138Crossref Scopus (0) Google Scholar, 32.Tatton A.S. Development of solid-state NMR techniques for the characterisation of pharmaceutical compounds. University of Warwick, PhD thesis2012: 195Google Scholar This analytical superiority has recently increased the utilization of SS-NMR in mechanistically understanding the structural features of a variety of complex drug delivery systems including ASD and in relating the same to the physical stability of the products.33.Vogt F.G. Clawson J.S. Strohmeier M. Pham T.N. Watson S.A. Edwards A.J. Gad S.C. New approaches to the characterization of drug candidates by solid-state NMR. In Pharmaceutical sciences encyclopedia. John Wiley & Sons, Inc., New Jersy, USA2011Crossref Google Scholar, 34.Vogt F.G. Evolution of solid-state NMR in pharmaceutical analysis.Future Med Chem. 2010; 2: 915-921Crossref PubMed Scopus (28) Google Scholar Very recently, Skorupska et al.35.Skorupska E. Jeziorna A. Kazmierski S. Potrzebowski M.J. Recent progress in solid-state NMR studies of drugs confined within drug delivery systems.Solid State Nucl Mag Reson. 2014; 57–58: 2-16Crossref PubMed Scopus (0) Google Scholar published a compilation of case studies reflecting the application of SS-NMR for the characterization of a wide diversity of enabling drug delivery systems, namely, mesoporous silica-based systems, polymeric dispersions, cyclodextrin complexes, and so on. This review covers different SS-NMR methodologies with several case studies that provide readers with general insights on the utility of these techniques for the in-depth elucidation of the phase structures and dynamics of ASD. Nuclear magnetic resonance spectroscopy is a top-tier characterization tool in chemistry for structural elucidation, chemical identification, quantification of concentration/composition, and study of diverse molecular dynamics (molecular rotations, diffusion, etc.) of a wide variety of materials including liquids, gels, and solids.36.Courtier-Murias D. Farooq H. Masoom H. Botana A. Soong R. Longstaffe J.G. Simpson M.J. Maas W.E. Fey M. Andrew B. Struppe J. Hutchins H. Krishnamurthy S. Kumar R. Monette M. Stronks H.J. Hume A. Simpson A.J. Comprehensive multiphase NMR spectroscopy: Basic experimental approaches to differentiate phases in heterogeneous samples.J Magn Reson. 2012; 217: 61-76Crossref PubMed Scopus (0) Google Scholar With the chemical and spatial specificity of the measuring principle and versatility of operations, NMR can analyze every type of very subtle alteration in the chemical environment of small to large molecules in all phases of materials and thus deliver supreme quantitative molecular information that is inaccessible by other techniques. The fundamental bases of NMR and, in particular, SS-NMR, are available in many books,37.Duer M.J. Introduction to solid-State NMR. Wiley-Blackwell, Oxford, UK2004Google Scholar, 38.Schmidt-Rohr K. Spiess H.W. Principles of NMR of organic solids. In Multidimensional solid-state NMR and polymers. Academic Press, San Diego, CA1994: 13-68Google Scholar, 39.Levitt M.H. Spin dynamics: Basics of nuclear magnetic resonance.2nd ed. Wiley, Chichester, UK2008Google Scholar, 40.Apperley D.C. Harris R.K. Hodgkinson P. Solid state NMR: Basic principles & practice. Momentum Press, New York, USA2012Crossref Google Scholar and in the following only a very brief and certainly incomplete summary of them is reported. The NMR phenomenon involves the Zeeman interaction between a static magnetizing field (B0) and the nuclear spin. A first, obvious consequence is that only nuclei having a non-null spin are NMR active. This interaction results in the precession of the spin about B0 (commonly defined as z-axis) at an angular frequency ω0 (the Larmor frequency), which is the product of B0 and the gyromagnetic ratio (γI), the latter having a characteristic value for each nuclear species I. The sum of all the nuclear spins of the same type is defined as magnetization. At equilibrium, under the sole action of B0, the magnetization assumes a small magnitude and it is directed along B0. The equilibrium can be perturbed by applying an rf-pulse (with an associated magnetic field B1) with a frequency of about ω0 along a direction perpendicular to B0. This causes the precession of the magnetization on the plane perpendicular to this direction. After the pulse is applied for a suitable time, all the magnetization or part of it can be found on the x–y plane, perpendicular to B0. If the sample is placed within a suitable conductive coil, when the magnetization evolves by precessing around B0 and tending to the equilibrium, it generates a small electric current within the coil, whose evolution with time represents the NMR signal (free induction decay, FID). When subjected to Fourier transformation, the FID gives rise to the classical NMR spectrum (intensity vs. frequency). In the real case, the nucleus is not “naked” and the currents of electrons revolving around it generate a small local three-dimensional magnetic field slightly altering B0 at the nucleus. The local field experienced by the nucleus (Bloc) is therefore the product of B0 and (1 − σζζ), where σζζ is an element of the shielding tensor, determined by the surrounding electrons. In general, σζζ also depends on the molecular orientation with respect to B0. However, in solution-state NMR, because of the fast and isotropic Brownian molecular tumbling experienced by the molecules, σζζ = σ, known as shielding factor, no longer depending on the molecular orientation. Therefore, nuclei of the same species can experience diverse values of σ because of the different local chemical environments. Consequently, these spins precess with different frequencies ω = γIB0(1-σ). In the NMR spectrum, they are revealed as well-separated peaks. The intensity of a peak at a particular frequency in the NMR spectrum of a specific nucleus of a sample is quantitatively proportional to the population of that nucleus in a given chemical environment, typically related to a chemical group. The collection of diverse nuclear resonances reflects molecular/supramolecular environments and it is depicted in the resulting NMR spectrum. The choice of a nucleus for NMR is based on the natural abundance of spin-active nuclei and on the intention of the study. Although proton NMR (1H-NMR) and 13C-NMR are universal NMR methodologies for most of the organic materials, X-NMR experiments (X = 15N, 19F, 17O, 31P, 2H, etc.) are sometimes preferred for their selective and richer information on the molecular structure. For each nucleus, besides the frequency scale, the x-axis of NMR spectrum is very commonly expressed in terms of chemical shift (δ) with the unit of parts per million (ppm). This scale is useful as the values of δ, linked to the extent of shielding experienced by the nucleus of interest, are independent of the intensity of the external magnetic field B0. Moreover, this scale is not absolute, but referred to a given reference substance, typically showing a single peak whose chemical shift is conventionally taken equal to zero. For example, some reference substances are tetramethylsilane (13C-, 1H-, and 29Si-NMR), CFCl3 (19F-NMR), and H2O (17O-NMR).41.Pham T.N. Watson S.A. Edwards A.J. Chavda M. Clawson J.S. Strohmeier M. Vogt F.G. Analysis of amorphous solid dispersions using 2D solid-state NMR and 1H T1 relaxation measurements.Mol Pharm. 2010; 7: 1667-1691Crossref PubMed Scopus (90) Google Scholar Coming back to solids, we remind that in this case the resonance frequency depends on σζζ, which also depends on molecular orientation, as in this case molecular tumbling is absent. This means that if we have a “powder” sample (where all the molecular orientations can be populated with the same probability), the same nucleus in different molecules gives rise to different resonance frequencies, resulting in a severe line broadening and consequent loss of resolution. This phenomenon is even more dramatic if we think that it also involves all the other “internal” interactions experienced by the nuclei (dipolar, scalar, quadrupolar, etc.), all of which have a strong dependence on molecular orientations, causing themselves further line broadening and loss of resolution. With several nuclear interactions experienced by the same nucleus, a broadly distributed resonance frequency produces a complex superposition pattern. This eventually results in very weak and broad NMR peaks with overlapping signals: this poor spectral resolution of SS-NMR spectra would make their interpretation very difficult or, more often, impossible. The advent of magic angle spinning (MAS) helped in acquiring improved SS-NMR spectra. It involves spinning the sample during measurement at high speed about an axis forming an angle of 54.74° with respect to B0: the spatial dependence of the NMR interactions (i.e., their anisotropic component) is fully averaged out provided the sample is spun at infinite speed. The second-order quadrupolar interaction represents an exception to this rule, but it will not be treated in the present review. Unfortunately, in practice, MAS spinning frequency is limited, although it can be very high in modern apparatuses (up to 70 kHz when using rotors with very small diameters of about 1.2 mm). This is sufficient for MAS to eliminate most broadening effects arising from chemical shielding, but not from dipolar coupling. For this reason, MAS must be combined to suitable heteronuclear and homonuclear decoupling schemes to record high-resolution spectra of rare (e.g., 13C) and abundant (e.g., 1H) spins, respectively. Contrary to solution-state NMR, decoupling in SS-NMR requires high-power amplifiers (up to 1 kW) because of the noticeable strength of the dipolar coupling. When the MAS frequency is smaller than chemical shielding anisotropy, MAS is not capable to completely average the shielding tensor. This leads to the appearance in the spectrum, in addition to the isotropic peaks, of spinning sidebands, separated from the corresponding isotropic peaks by integer multiples of the spinning frequency. Spinning sidebands can be easily discriminated from the isotropic signal as they shift by changing the MAS speed. Nonetheless, sidebands can be removed by the application of special pulse sequences, such as “total suppression of sidebands” (TOSS). However, often MAS alone is not sufficient to remove the severe line-broadening effects produced by homonuclear and heteronuclear dipolar coupling, and therefore several types of decoupling schemes, consisting of carefully selected high-power rf-pulses, have been developed, capable of removing such effect and further enhancing spectral quality. The application of homo-decoupling sequences in MAS mode is commonly known as combined rotation and multiple pulse sequences (CRAMPS).37.Duer M.J. Introduction to solid-State NMR. Wiley-Blackwell, Oxford, UK2004Google Scholar Different types of CRAMPS are used for recording one-dimensional SS-NMR high-resolution spectra of abundant nuclei. In order to record one-dimensional SS-NMR high-resolution spectra of rare nuclei simpler heteronuclear decoupling techniques (continuous wave, CW, TPPM, SPINAL, XiX, etc.) can be successfully applied in combination with MAS. Beside MAS and high-power decoupling schemes, cross polarization (CP) must be mentioned: this is a pulse sequence used to greatly increase the sensitivity of rare nuclei (actually without increasing spectral resolution). CP exploits the magnetization transfer from abundant (“a,” typically 1H or 19F) to rare, low-γ nuclei (“r,” e.g., 15N, 13C) taking place by simultaneously irradiating both nuclei under the particular Hartmann–Hahn conditions γaB1a = γrB1r. The enhancement of sensitivity is achieved because both the magnetization transferred in a single scan is larger than that obtained by direct excitation, and the time to wait between consecutive scans is determined by the T1 of the abundant rather than the rare nuclei, the former being generally up to two to three orders of magnitude shorter. In pharmaceutical analysis, including ASD,