The research work reported in this thesis is focused on the chiral analysis, quantification of enantiomeric composition, assignment of absolute configuration of molecules with chosen functional groups. The weak intra-molecular hydrogen bonding interactions are detected by exploiting several multinuclear and multi-dimensional techniques. Pulse sequences have been designed to manipulate the spin dynamics to derive specific information from the complex NMR spectra encountered in diverse situations. Broadly, the thesis can be classified in to three sections. The section I containing two chapters reports the introduction of new chiral auxiliaries and protocols developed for enantiomeric discrimination, measurement of enantiomeric contents, assignment of absolute configuration for molecules possessing specific functional groups using chiral solvating and derivatizing agents. The section II, reports NMR experimental evidence for the observation of the rare type of intramolecular hydrogen bonds involving organic fluorine in biologically important organic molecules, that are corroborated by extensive DFT based theoretical calculations. The section II also discusses the H/D exchange mechanism as a tool for quantification of HB strengths in organic building blocks. The section III reports the two different novel NMR methodologies designed for deriving information on the scalar interaction strengths in an orchestrated manner. The designed sequences are able to completely eradicate the axial peaks, prevents the evolution of unwanted couplings and also yields ultrahigh resolution in the direct dimension, permitting the accurate measurement of scalar couplings for a particular spin. The brief summary about each chapter is given below. Chapter 1 provides a general introduction to one and two dimensional NMR spectroscopy. The pedagogical approach has been followed to discuss the conceptual understanding of spin physics and the NMR spectral parameters. The basic introduction to chirality, existing approaches in the literature for discrimination of enantiomers and the assignment of absolute configuration of molecules with chosen functional groups and their limitations are briefly discussed. The brief introduction to hydrogen bond, experimental methods to obtain the qualitative information about the strengths of hydrogen bonds, and the theoretical approaches employed in the thesis to corroborate the NMR experimental findings have been provided. The mechanism of H/D exchange, the utilization of exchange rates to derive strengths of intra-molecular hydrogen bond in small molecules have also been discussed. This chapter builds the bridge for the rest of the chapters. Each of these topics are discussed at length in the corresponding chapters. Part I: NMR Chiral Analysis: Novel Protocols Chapter 2 discusses a simple mix and shake method for testing the enantiopurity of primary, secondary and tertiary chiral amines and their derivatives, amino alcohols. The protocol involves the in-situ formation of chiral ammonium borate salt from a mixture of C2 symmetric chiral BINOL, trialkoxyborane and chiral amines. The proposed concept has been convincingly demonstrated for the visualization of enantiomers of a large number of chiral and pro-chiral amines and amino alcohols. The protocol also permits the precise measurement of enantiomeric composition. The significant advantage of the protocol is that it can be performed directly in the NMR tube, without any physical purification. The structure of the borate complex responsible for the enantiodifferentiation of amines has also been established by employing multinuclear NMR techniques and DFT calculations. From DOSY and 11B NMR experiments it has been ascertained that there are only two possible complexes or entities which are responsible for differentiating enantiomers. From the combined utility of DFT calculations and the 11B NMR chemical shifts, the structure of the borate complex has been determined to be an amine-coordinated complex with the N atom of the amine. Chapter 3 discusses a simple chiral derivatizing protocol involving the coupling of 2-formylphenylboronic acid and an optically pure [1,1-binaphthalene]-2,2-diamine for the rapid and accurate determination of the enantiopurity of hydroxy acids and their derivatives, possessing one or two optically active centers. It is established that this protocol is not only rapid method for discrimination of enantiomers but also highly effective for assigning the absolute configuration of various chiral hydroxy acids and their derivatives. The developed protocol involves the coupling of 2-formylphenylboronic acid with (R)-[1,1-binaphthalene]-2,2-diamine, and 2-formylphenylboronic acid with (S)-[1,1-binaphthalene]-2,2-diamine as chiral derivatizing agents. The absence of aliphatic peaks from the derivatizing agent, large chemical shift separation between the discriminated peaks of diastereomers, and the systematic change in the direction of displacement of peaks for an enantiomer in a particular diastereomeric complex, permitted the unambiguous assignment of absolute configuration. Part II : Rare Type of Intramolecular Hydrogen Bonding In chapter 4 The rare occurrence of intramolecular hydrogen bonds of the type N–H˖˖˖F–C, in the derivatives of imides and hydrazides in a low polarity solvent, is convincingly established by employing multi-dimensional and multinuclear solution state NMR experiments. The observation of 1hJFH, 2hJFN, and 2hJFF of significant strengths, where the spin polarization is transmitted through space among the interacting NMR active nuclei, provided strong and conclusive evidence for the existence of intra-molecular hydrogen bonds. Solvent induced perturbations and the variable temperature NMR experiments unambiguously supported the presence of intramolecular hydrogen bond. The two dimensional HOESY and 15N–1H HSQC experiments reveals the existence of multiple conformers in some of the investigated molecules. The 1H DOSY experimental results discarded any possibility of self or cross-dimerization of the molecules. The results of DFT based calculations, viz., Quantum Theory of Atoms In Molecules (QTAIM) and Non Covalent Interaction (NCI), are in close agreement with the NMR experimental findings. In chapter 5 the rates of hydrogen/deuterium (H/D) exchange determined by 1H NMR spectra have been utilized to derive the strength of hydrogen bonds and to monitor the electronic effects in the site-specific halogen substituted benzamides and anilines. The theoretical fitting of the time dependent variation in the integral areas of 1H NMR resonances to the first order decay function permitted the determination of H/D exchange rate constants (k) and their precise half-lives (t1/2) with high degree of reproducibility. The comparative study also permitted the determination of relative strengths of hydrogen bonds and the contribution from electronic effects on the H/D exchange rates. Part III: Novel NMR Methodologies for the Precise Measurement of 1H-1H Couplings Chapter 6 describes two novel NMR methodologies developed for the precise measurement of 1H-1H couplings. Poor chemical shift dispersion and the pairwise interaction among the entire coupled network of protons results in the severely complex and overcrowded one dimensional 1H NMR spectra, hampering both the resonance assignments and the accurate determination of nJHH. The available two-dimensional selective refocusing (SERF) based experiments suffer from the evolution of magnetization from uncoupled protons as intense uninformative axial peaks. This creates ambiguity in the identification of peaks belonging to the coupled partners of a selectively excited proton, hindering the extraction of their interaction strengths. This challenge has been circumvented by designing two novel experimental technique, cited as “Clean-G-SERF” and “PS-Clean-G-SERF”. The Clean-G-SERF technique completely eradicates the axial peaks and suppresses the evolution of unwanted couplings while retaining only the couplings to the selectively excited proton. The method permits the accurate determination of spin-spin couplings even from a complex proton NMR spectrum in an orchestrated manner. The PS-Clean-G-SERF technique has been designed for the complete elimination of axial peaks and undesired couplings, with a blend of ultra-high resolution achieved by real time broad band homonuclear decoupling has been discussed in this chapter. The spin dynamics involved in both these pulse sequences have been discussed. The diverse applications of both these novel experiments have been demonstrated.

The research work reported in this thesis is focused on the chiral analysis, quantification of enantiomeric composition, assignment of absolute configuration of molecules with chosen functional groups. The weak intra-molecular hydrogen bonding interactions are detected by exploiting several multinuclear and multi-dimensional techniques. Pulse sequences have been designed to manipulate the spin dynamics to derive specific information from the complex NMR spectra encountered in diverse situations. Broadly, the thesis can be classified in to three sections. The section I containing two chapters reports the introduction of new chiral auxiliaries and protocols developed for enantiomeric discrimination, measurement of enantiomeric contents, assignment of absolute configuration for molecules possessing specific functional groups using chiral solvating and derivatizing agents. The section II, reports NMR experimental evidence for the observation of the rare type of intramolecular hydrogen bonds involving organic fluorine in biologically important organic molecules, that are corroborated by extensive DFT based theoretical calculations. The section II also discusses the H/D exchange mechanism as a tool for quantification of HB strengths in organic building blocks. The section III reports the two different novel NMR methodologies designed for deriving information on the scalar interaction strengths in an orchestrated manner. The designed sequences are able to completely eradicate the axial peaks, prevents the evolution of unwanted couplings and also yields ultrahigh resolution in the direct dimension, permitting the accurate measurement of scalar couplings for a particular spin. The brief summary about each chapter is given below. Chapter 1 provides a general introduction to one and two dimensional NMR spectroscopy. The pedagogical approach has been followed to discuss the conceptual understanding of spin physics and the NMR spectral parameters. The basic introduction to chirality, existing approaches in the literature for discrimination of enantiomers and the assignment of absolute configuration of molecules with chosen functional groups and their limitations are briefly discussed. The brief introduction to hydrogen bond, experimental methods to obtain the qualitative information about the strengths of hydrogen bonds, and the theoretical approaches employed in the thesis to corroborate the NMR experimental findings have been provided. The mechanism of H/D exchange, the utilization of exchange rates to derive strengths of intra-molecular hydrogen bond in small molecules have also been discussed. This chapter builds the bridge for the rest of the chapters. Each of these topics are discussed at length in the corresponding chapters. Part I: NMR Chiral Analysis: Novel Protocols Chapter 2 discusses a simple mix and shake method for testing the enantiopurity of primary, secondary and tertiary chiral amines and their derivatives, amino alcohols. The protocol involves the in-situ formation of chiral ammonium borate salt from a mixture of C2 symmetric chiral BINOL, trialkoxyborane and chiral amines. The proposed concept has been convincingly demonstrated for the visualization of enantiomers of a large number of chiral and pro-chiral amines and amino alcohols. The protocol also permits the precise measurement of enantiomeric composition. The significant advantage of the protocol is that it can be performed directly in the NMR tube, without any physical purification. The structure of the borate complex responsible for the enantiodifferentiation of amines has also been established by employing multinuclear NMR techniques and DFT calculations. From DOSY and 11B NMR experiments it has been ascertained that there are only two possible complexes or entities which are responsible for differentiating enantiomers. From the combined utility of DFT calculations and the 11B NMR chemical shifts, the structure of the borate complex has been determined to be an amine-coordinated complex with the N atom of the amine. Chapter 3 discusses a simple chiral derivatizing protocol involving the coupling of 2-formylphenylboronic acid and an optically pure [1,1-binaphthalene]-2,2-diamine for the rapid and accurate determination of the enantiopurity of hydroxy acids and their derivatives, possessing one or two optically active centres. It is established that this protocol is not only rapid method for discrimination of enantiomers but also highly effective for assigning the absolute configuration of various chiral hydroxy acids and their derivatives. The developed protocol involves the coupling of 2-formylphenylboronic acid with (R)-[1,1-binaphthalene]-2,2-diamine, and 2-formylphenylboronic acid with (S)-[1,1-binaphthalene]-2,2-diamine as chiral derivatizing agents. The absence of aliphatic peaks from the derivatizing agent, large chemical shift separation between the discriminated peaks of diastereomers, and the systematic change in the direction of displacement of peaks for an enantiomer in a particular diastereomeric complex, permitted the unambiguous assignment of absolute configuration. Part II : Rare Type of Intramolecular Hydrogen Bonding In chapter 4 The rare occurrence of intramolecular hydrogen bonds of the type N–H˖˖˖F–C, in the derivatives of imides and hydrazides in a low polarity solvent, is convincingly established by employing multi-dimensional and multinuclear solution state NMR experiments. The observation of 1hJFH, 2hJFN, and 2hJFF of significant strengths, where the spin polarization is transmitted through space among the interacting NMR active nuclei, provided strong and conclusive evidence for the existence of intra-molecular hydrogen bonds. Solvent induced perturbations and the variable temperature NMR experiments unambiguously supported the presence of intramolecular hydrogen bond. The two dimensional HOESY and 15N–1H HSQC experiments reveals the existence of multiple conformers in some of the investigated molecules. The 1H DOSY experimental results discarded any possibility of self or cross-dimerization of the molecules. The results of DFT based calculations, viz., Quantum Theory of Atoms In Molecules (QTAIM) and Non Covalent Interaction (NCI), are in close agreement with the NMR experimental findings. In chapter 5 the rates of hydrogen/deuterium (H/D) exchange determined by 1H NMR spectra have been utilized to derive the strength of hydrogen bonds and to monitor the electronic effects in the site-specific halogen substituted Benz amides and anilines. The theoretical fitting of the time dependent variation in the integral areas of 1H NMR resonances to the first order decay function permitted the determination of H/D exchange rate constants (k) and their precise half-lives (t1/2) with high degree of reproducibility. The comparative study also permitted the determination of relative strengths of hydrogen bonds and the contribution from electronic effects on the H/D exchange rates. Part III: Novel NMR Methodologies for the Precise Measurement of 1H-1H Couplings Chapter 6 describes two novel NMR methodologies developed for the precise measurement of 1H-1H couplings. Poor chemical shift dispersion and the pairwise interaction among the entire coupled network of protons results in the severely complex and overcrowded one dimensional 1H NMR spectra, hampering both the resonance assignments and the accurate determination of nJHH. The available two-dimensional selective refocusing (SERF) based experiments suffer from the evolution of magnetization from uncoupled protons as intense uninformative axial peaks. This creates ambiguity in the identification of peaks belonging to the coupled partners of a selectively excited proton, hindering the extraction of their interaction strengths. This challenge has been circumvented by designing two novel experimental technique, cited as “Clean-G-SERF” and “PS-Clean-G-SERF”. The Clean-G-SERF technique completely eradicates the axial peaks and suppresses the evolution of unwanted couplings while retaining only the couplings to the selectively excited proton. The method permits the accurate determination of spin-spin couplings even from a complex proton NMR spectrum in an orchestrated manner. The PS-Clean-G-SERF technique has been designed for the complete elimination of axial peaks and undesired couplings, with a blend of ultra-high resolution achieved by real time broad band mononuclear decoupling has been discussed in this chapter. The spin dynamics involved in both these pulse sequences have been discussed. The diverse applications of both these novel experiments have been demonstrated.

The thesis entitled “Exploring Diverse Facets of Small Molecules by NMR Spectroscopy” consists of six chapters. The main theme of the thesis is to exploit one and two dimensional NMR methodologies for understanding the diverse facets of small organic molecules, such as, weak intra- and inter- molecular interactions, chiral discrimination, quantification of enantiomeric excess and assignment of absolute configuration. Several new pulse sequences have also been designed to solve specific chemical problems, in addition to extensive utility of existing one and two dimensional NMR experiments. The results obtained on different problems, are discussed under six chapters in the thesis. The brief summary of each of these chapters is given below. Chapter 1 begins with the discussion on the importance of small molecules and their various facets, the analytical techniques available in the literature to study them. The role of NMR spectroscopy as powerful analytical technique to understand the diverse facets of organic molecules and their importance is set out in brief. A short introduction to the basic principles of NMR, the interaction parameters, the commonly employed one and two dimensional homo- and herero- nuclear NMR experiments are also given. The basic introduction to product operators essential for understanding the spin dynamics in the developed pulse sequences is given. The application of diffusion ordered spectroscopy (DOSY), the general problems encountered in the analysis of combinatorial mixtures and the matrix assisted method in circumventing such problems are discussed. Chapter 2 focuses on the chiral discrimination and the measurement of enatiomeric excess. The NMR approach to discriminate enantiomers using chiral auxiliaries such as, solvating agents, derivatizing agents, lanthanide shift reagents, the choice of such auxiliaries and the limitations are discussed in detail. The in-depth discussion on the new protocols developed using both the solvating and derivatizing agents for enantiomeric discrimination of chiral amines, hydroxy acids and diacids are discussed. The new three-component protocols that serve as chiral derivatizing agents for the discrimination of primary amines, diacids and hydroxy acids are discussed. Also the role of organic base such as DMAP in the chiral discrimination is explored for discrimination of acids using BINOL as a chiral solvating agent. Accordingly the discussion is classified into two sections. In the first section the protocol developed utilizing an enantiopure mandelic acid, a primary amine substrate and 2-formylphenylboronic acid that is ideally suited for testing the enantiopurity of chiral primary amines is discussed. The broad applicability of the protocols for testing enantiopurity has been demonstrated on number of chiral molecules using 1H and 19F NMR. The second section contains the results on the new concept developed for discrimination of hydroxy acids. The strategy involves the formation of three component protocol using chiral hydroxy acid, R-alphamethylbenzylamine and 2-formylphenylboronic acid for 1H-NMR discrimination of diacids. The section also includes the utility of ternary ion-pair complex for the discrimination of acids. The ternary ion-pair not only permitted the testing of enantiopurity of chiral acids, but is also found useful for the measurement of enantiomeric excess. Chapter 3 discusses the utilization of the developed three-component protocols for the assignment of absolute configurations of molecules of different functionality. The protocols for the assignments of absolute configuration of primary amines using 2-formylphenylboronic acid and mandelic acid yielded the substantial chemical shift differences between diastereomers. The consistent trend in the direction of change of chemical shifts of the discriminated proton(s) gave significant evidence for employing them as parameters for the assignment of spatial configuration of primary amines. Another protocol using 2-formylphenylboronic acid, hydroxy acids and enantiopure alphamethylbenzylamine permitted their configurational assignment. In the second section a novel solvating agent, obtained by the formation of an ion-pair complex among enantiopure BINOL, DMAP and chiral hydroxy acid for the assignment of the spatial configuration of hydroxy acids is discussed. Chapter 4 focuses on the development of novel NMR methodologies, and also the utility of existing two-dimensional experiments for addressing certain challenging problems. This chapter has been divided into three sections. In Section-I the utilization of well-known homonuclear 2D-J-resolved methodology for unravelling the overlapped NMR spectra of enantiomers, an application for chiral discrimination and the measurement of enantiomeric excess is discussed. The utilization of the chiral auxiliaries, such as, chiral derivatizing agents, chiral solvating agents and lanthanide shift reagents permits enantiodiscrimination and the measurement of excess of one form over the other. Nevertheless many a times one encounters severe problems due to small chemical shift difference, overlap of resonances, complex multiplicity pattern because of the presence of number of interacting spins, and enormous line broadening due to paramagnetic nature of the metal complex. This section is focused on combating such problems utilizing 2D-J-1JNH resolved spectroscopy where a 450 tilting of the spectrum in the F2 dimension, yielded the pure shift NMR spectrum. The method circumvents several problems involved in chiral discrimination and allows the accurate measurement of enantiomeric excess. In Section-II, the development of novel NMR experimental methodology cited in the literature as C-HetSERF and its application for the study of symmetric molecules, such as, double bonded cis- and trans- isomers, and extraction of magnitudes and signs of long range homo- and hetero- nuclear scalar couplings among chemically equivalent protons in polycylic aromatic hydrocarbons is discussed. The extensive utility of the new pulse sequence has been demonstrated on number of symmetric molecules, where the conventional one dimensional experiment fails to yield spectral parameters. In section III, yet another novel pulse sequence called RES-TOCSY developed for unravelling of the overlapped NMR spectrum of enantiomers and the measurement of enantiomeric contents, has been utilized for the accurate measurement of magnitudes and signs of 1H-19F couplings in fluorine containing molecules. The method has distinct advantages as the strengths of the couplings and their relative signs could be extracted on diverse situations, such as, couplings smaller than line widths, the spectrum where the coupling fine structures are absent. Chapter 5 covers the study of nature of intra- and inter- molecular hydrogen bond in amide and its derivative. The chapter is accordingly divided into two sections. In the first section the study of acid and amide hydrogen bonding is discussed and the hydrogen bonded interactions are probed by extensive utility of 1H, 13C and 15N-NMR. The temperature perturbation experiments, measurements of the variation in the couplings, monitoring of diffusion coefficients and the association constants, detection of through space correlation have given unambiguous evidence for the hydrogen bond formation. The results were also supported by DFT calculations. Similar interaction in the solid state has also been derived by obtaining the crystal structure of complex phenylacetic acid with benzamide. In the second section of the chapter the hydrogen bond interaction of organic fluorine in trifluoromethyl derivatives of benzanilides has been explored and the involvement of CF3 group in the hydrogen bonding has been detected. The evidence for the participation of CF3 group in hydrogen bond has been confirmed by number of experiments, such as, the detection of through space couplings, viz., 1hJFH, 1hJFN, and 2hJFF , where the spin polarization between the interacting spins is transmitted through hydrogen bond, the temperature and solvent dependent studies, variation in the 1JNH and two dimensional heteronuclear correlation experiments. In an interesting example of a molecule containing two CF3 groups situated on two phenyl rings of benzanilide, the simultaneous participation of fluorines of two CF3 groups in hydrogen bond has been detected. The confirmatory evidence for such an interaction, where hydrogen bond mediated couplings are not reflected in the NMR spectrum, has been derived by 19F−19F NOESY. Significant deviations in the strengths of 1JNH, in addition to variable temperature, and the solvent induced perturbation studies yielded additional evidence. The NMR results are corroborated by both DFT calculations and MD simulations, where the quantitative information on different ways of involvement of fluorine in two and three centered hydrogen bonds, their percentage of occurrences, and geometries have been obtained. The hydrogen bond interaction energies have also been calculated. The study revealed the rare observation and the first example of the C-F…H-N hydrogen bond in solution state in the molecules containing CF3 groups. Chapter 6 focuses on the mixture analysis using the diffusion ordered spectroscopy (DOSY). High Resolution-DOSY works when the NMR spectrum is well resolved and the diffusion coefficients of the combinatorial mixtures are substantially different from each other. DOSY technique fails when the mixture contains the molecules of nearly identical weights and similar hydrodynamic radii. Thus, the positional isomers, enantiomers consequent to their nearly identical rates of diffusion, are not differentiated. Some of these problems can be overcome by Matrix-Assisted Diffusion Order Spectroscopy (MAD-spectroscopy), where an external reagent acts as a matrix and aids in their diffusion edited separation, provided the molecules embedded in it possess differential binding abilities with the matrix. Such different binding properties of the matrix are the basis for resolution of many isomeric species. In the present study three different novel auxiliaries, micelles-reverse micelles, crown ether and cyclodextrin are introduced for the resolution of positional isomers, double bonded isomers, viz., fumaric acid and maleic acid and also enantiomers. Accordingly, the results of each of these studies are discussed in three different sections.