Trapped ion magnetic resonance: concepts and designs

A novel spectroscopy of trapped ions is proposed which will bring single-ion detection sensitivity to the observation of magnetic resonance spectra. The approaches developed here are aimed at resolving one of the fundamental problems of molecular spectroscopy, the apparent incompatibility in existing techniques between high information content (and therefore good species discrimination) and high sensitivity. Methods for studying both electron spin resonance (ESR) and nuclear magnetic resonance (NMR) are designed. They assume established methods for trapping ions in high magnetic field and observing the trapping frequencies with high resolution (<1 Hz) and sensitivity (single ion) by electrical means. The introduction of a magnetic bottle field gradient couples the spin and spatial motions together and leads to a small spin-dependent force on the ion, which has been exploited by Dehmelt to observe directly the perturbation of the ground-state electron's axial frequency by its spin magnetic moment.

A series of fundamental innovations is described m order to extend magnetic resonance to the higher masses of molecular ions (100 amu = 2x 10^5 electron masses) and smaller magnetic moments (nuclear moments = 10^(-3) of the electron moment). First, it is demonstrated how time-domain trapping frequency observations before and after magnetic resonance can be used to make cooling of the particle to its ground state unnecessary. Second, adiabatic cycling of the magnetic bottle off between detection periods is shown to be practical and to allow high-resolution magnetic resonance to be encoded pointwise as the presence or absence of trapping frequency shifts. Third, methods of inducing spindependent work on the ion orbits with magnetic field gradients and Larmor frequency irradiation are proposed which greatly amplify the attainable shifts in trapping frequency.

The dissertation explores the basic concepts behind ion trapping, adopting a variety of classical, semiclassical, numerical, and quantum mechanical approaches to derive spin-dependent effects, design experimental sequences, and corroborate results from one approach with those from another. The first proposal presented builds on Dehmelt's experiment by combining a "before and after" detection sequence with novel signal processing to reveal ESR spectra. A more powerful technique for ESR is then designed which uses axially synchronized spin transitions to perform spin-dependent work in the presence of a magnetic bottle, which also converts axial amplitude changes into cyclotron frequency shifts. A third use of the magnetic bottle is to selectively trap ions with small initial kinetic energy. A dechirping algorithm corrects for undesired frequency shifts associated with damping by the measurement process.

The most general approach presented is spin-locked internally resonant ion cyclotron excitation, a true continuous Stern-Gerlach effect. A magnetic field gradient modulated at both the Larmor and cyclotron frequencies is devised which leads to cyclotron acceleration proportional to the transverse magnetic moment of a coherent state of the particle and radiation field. A preferred method of using this to observe NMR as an axial frequency shift is described in detail. In the course of this derivation, a new quantum mechanical description of ion cyclotron resonance is presented which is easily combined with spin degrees of freedom to provide a full description of the proposals.

Practical, technical, and experimental issues surrounding the feasibility of the proposals are addressed throughout the dissertation. Numerical ion trajectory simulations and analytical models are used to predict the effectiveness of the new designs as well as their sensitivity and resolution. These checks on the methods proposed provide convincing evidence of their promise in extending the wealth of magnetic resonance information to the study of collisionless ions via single-ion spectroscopy.

Applications of nuclear magnetic resonance spectroscopy to the study of medium-sized rings. I. Conformational properties of cycloheptane. II. Conformational properties of cycloöctane

Conformational equilibrium in medium-sized rings has been investigated by the temperature variation of the fluorine-19 n.m.r. spectra of 1, 1-difluorocycloalkanes and various substituted derivatives of them. Inversion has been found to be fast on the n.m.r. time scale at -180˚ for 1, 1-difluorocycloheptane, but slow for 1, 1-difluoro-4, 4-dimethylcycloheptane at -150˚. At low temperature, the latter compound affords a single AB pattern with a chemical-shift difference of 841 cps. which has been interpreted in terms of the twist-chair conformation with the methyl groups on the axis position and the fluorine atoms in the 4-position. At room temperature, the n.m.r. spectrum of 1, 1-difluoro-4-t-butylcycloheptane affords an AB pattern with a chemical-shift difference of 185 cps. The presence of distinct trans and gauche couplings from the adjacent hydrogens has been interpreted to suggest the existence of a single predominant form, the twist chair with the fluorine atoms on the axis position.

Investigation of 1, 1-difluorocycloöctane and 1, 1, 4, 4-tetrafluorocycloöctane has led to the detection of two kinetic processes both having activation energies of 8-10 kcal./mole but quite different A values. In light of these results eleven different conformations of cycloöctane along with a detailed description of the ways in which they may be interconverted are discussed. An interpretation involving the twist-boat conformation rapidly equilibrating through the saddle and the parallel-boat forms at room temperature is compatible with the results.

I. Nuclear magnetic resonance analysis of ferrocenylcarbonium ion. II. Ferrocene catalyzed photochemistry

I. Nuclear magnetic resonance spectra of appropriately substituted ferrocenylcarbonium ions reveal the α-protons of the substituted ring to be more shielded than β-protons. The observation is discussed in terms of various models proposed for the ferrocenylcarbonium ion and is found to support a model in which the iron is bonded to all six carbona of the substituted ring.

II. Ferrocene catalyzes the photoisomerization of the piperylenes and the photodimerization of isoprene. Our results suggest a mechanism in which a complex of ferrocene and diene is excited to its second singlet state which dissociates to a triplet-state ferrocene molecule and a triplet-state diene molecule. The triplet-state diene, then, proceeds to isomerize or attack ground-state diene to form dimers.

I. The crystal structure of potassium bis(tricyanovinyl)amine. II. Nuclear magnetic resonance studies of alkali halide solutions. III. Elucidation of the intramolecular hydrogen bond in acetylacetone by the deuterium isotope effect on the chemical shift of the bridge hydrogen

Part I

Potassium bis-(tricyanovinyl) amine, K⁺N[C(CN)=C(CN)₂]₂^-, crystallizes in the monoclinic system with the space group Cc and lattice constants, a = 13.346 ± 0.003 Å, c = 8.992 ± 0.003 Å, B = 114.42 ± 0.02°, and Z = 4. Three dimensional intensity data were collected by layers perpendicular to b* and c* axes. The crystal structure was refined by the least squares method with anisotropic temperature factor to an R value of 0.064.

The average carbon-carbon and carbon-nitrogen bond distances in –C-CΞN are 1.441 ± 0.016 Å and 1.146 ± 0.014 Å respectively. The bis-(tricyanovinyl) amine anion is approximately planar. The coordination number of the potassium ion is eight with bond distances from 2.890 Å to 3.408 Å. The bond angle C-N-C of the amine nitrogen is 132.4 ± 1.9°. Among six cyano groups in the molecule, two of them are bent by what appear to be significant amounts (5.0° and 7.2°). The remaining four are linear within the experimental error. The bending can probably be explained by molecular packing forces in the crystals.

Part II

The nuclear magnetic resonance of ⁸¹Br and ¹²⁷I in aqueous solutions were studied. The cation-halide ion interactions were studied by studying the effect of the Li⁺, Na⁺, K⁺, Mg⁺⁺, Cs⁺ upon the line width of the halide ions. The solvent-halide ion interactions were studied by studying the effects of methanol, acetonitrile, and acetone upon the line width of ⁸¹Br and ¹²⁷I in the aqueous solutions. It was found that the viscosity plays a very important role upon the halide ions line width. There is no specific cation-halide ion interaction for those ions such as Mg⁺⁺, Di⁺, Na⁺, and K⁺, whereas the Cs⁺ - halide ion interaction is strong. The effect of organic solvents upon the halide ion line width in aqueous solutions is in the order acetone ˃ acetonitrile ˃ methanol. It is suggested that halide ions do form some stable complex with the solvent molecules and the reason Cs⁺ can replace one of the ligands in the solvent-halide ion complex.

Part III

An unusually large isotope effect on the bridge hydrogen chemical shift of the enol form of pentanedione-2, 4(acetylacetone) and 3-methylpentanedione-2, 4 has been observed. An attempt has been made to interpret this effect. It is suggested from the deuterium isotope effect studies, temperature dependence of the bridge hydrogen chemical shift studies, IR studies in the OH, OD, and C=O stretch regions, and the HMO calculations, that there may probably be two structures for the enol form of acetylacetone. The difference between these two structures arises mainly from the electronic structure of the π-system. The relative population of these two structures at various temperatures for normal acetylacetone and at room temperature for the deuterated acetylacetone were calculated.

I. Efforts toward the synthesis of aliphatic iodonium salts. II. Flourine-19 nuclear magnetic resonance spectroscopy of cyclic and bicyclic systems

The synthesis of iodonium salts of the general formula [C₆H₅IR]⁺X^-, where R is an alkyl group and x- is a stabilizing anion, was attempted. For the choice of R three groups were selected, whose derivatives are known to be sluggish in S_N1 and S_N2 substitutions: cyclopropyl, 7, 7 -dimethyl-1-norbornyl, and 9 -triptycyl. The synthetic routes followed along classical lines which have been exploited in recent years by Beringer and students. Ultimately, the object of the present study was to study the reactions of the above salts with nucleophiles. In none of the three cases, however, was it possible to isolate a stable salt. A thermodynamic argument suggests that this must be due to kinetic instability rather than thermodynamic instability. Only iodocyclopropane and 1-iodoapocamphane formed isolable iododichlorides.

Several methylated 2, 2-difluoronorbornanes were prepared with the intent of correlating fluorine -19 chemical shifts with geometric features in a rigid system. The effect of a methyl group on the shielding of a β -fluorine is dependent upon the dihedral angle; the maximum effect (an upfield shift of the resonance) occurs at 0° and 180°, whereas almost no effect is felt at a dihedral angle of 120°. The effect of a methyl group on a γ -fluorine is to strongly shift the resonance downfield when fluorine and methyl group are in a 1, 3 - diaxial-like relationship. Molecular orbital calculations of fluorine shielding in a variety of molecules were carried out using the formalism developed by Pople; the results are, at best, in modest agreement with experiment.

Nuclear magnetic resonance studies of protein conformation: I. Ribonuclease S. II. Hemoglobin

Fluorine nuclear magnetic resonance techniques have been used to study conformational processes in two proteins labeled specifically in strategic regions with covalently attached fluorinated molecules. In ribonuclease S, the ϵ-amino groups of lysines 1 and 7 were trifluoroacetylated without diminishing enzymatic activity. As inhibitors bound to the enzyme, changes in orientation of the peptide segment containing the trifluoroacetyl groups were detected in the nuclear magnetic resonance spectrum. pH Titration of one of the histidines in the active site produced a reversal of the conformational process.

Hemoglobin was trifluoroacetonylated at the reactive cysteine 93 of each β chain. The nuclear magnetic resonance spectrum of the fluorine moiety reflected changes in the equilibrium position of the β chain carboxy terminus upon binding of heme ligands and allosteric effectors. The chemical shift positions observed in deoxy- and methemoglobin were pH dependent, undergoing an abnormally steep apparent titration which was not observed in hemoglobin from which histidine β 146 had been removed enzymatically. The abnormal sharpness of these pH dependent processes is probably due to interactions between several ionizing groups.

The carbon monoxide binding process was studied by concurrent observation of the visible and nuclear magnetic resonance spectra of trifluoroacetonylated hemoglobin at fractional ligand saturations throughout the range 0-1.0. Comparison of the ligand binding process observed in these two ways yields evidence for a specific order of ligand binding. The sequence of events is sensitive to the pH and organic phosphate concentration of the medium, demonstrating the delicately balanced control system produced by interactions between the hemoglobin subunits and the effectors.

The binding and catalytic properties of lysozyme

The binding and catalytic properties of hen's egg white lysozyme have been studied by a variety of techniques. These studies show that the enzyme has three contiguous binding subsites, A, B, and C. The application of nuclear magnetic resonance (NMR) spectroscopy to probe the binding environment of several saccharides to lysozyme has demonstrated that the reducing end sugar rings of chitotriose, chitobiose and the β-form of N-acetylglucosamine all bind in subsite C. The central sugar ring of chitotriose and the sugar ring at the nonreducing end of chitobiose were found to bind in subsite B, while the nonreducing end sugar residue of chitotriose occupied subsite A. The dynamics of the binding process has also been investigated by NMR. The formation rate constant of chitobiose--and chitotriose-enzyme complexes were found to be about 4 X 10^-6 M^-1 sec^-1 with small activation energies.

The stereochemical path of the lysozyme catalyzed hydrolysis of glycosidic bonds has been shown to proceed with at least 99.7% retention of configuration at C-1 of the sugar. The lysozyme catalyzed hydrolysis of glucosidic bonds has been shown to be largely carbonium ion in character by virtue of the α-deuterium kinetic isotope effect (k_H/k_D = 1.11) observed for the reaction. It is probable that the mechanism of action of the enzyme involves a carbonium ion intermediate which is stereospecifically quenched by solvent. However, acetamido group participation cannot be ruled out for natural substrates.

Spin dynamics of pulsed nuclear magnetic double resonance in solids

In the first part of this thesis, experiments utilizing an NMR phase interferometric concept are presented. The spinor character of two-level systems is explicitly demonstrated by using this concept. Following this is the presentation of an experiment which uses this same idea to measure relaxation times of off-diagonal density matrix elements corresponding to magnetic-dipole-forbidden transitions in a ^(13)C-^1H, AX spin system. The theoretical background for these experiments and the spin dynamics of the interferometry are discussed also.

The second part of this thesis deals with NMR dipolar modulated chemical shift spectroscopy, with which internuclear bond lengths and bond angles with respect to the chemical shift principal axis frame are determined from polycrystalline samples. Experiments using benzene and calcium formate verify the validity of the technique in heteronuclear (^(13)C-^1H) systems. Similar experiments on powdered trichloroacetic acid confirm the validity in homonuclear (^1H- ^1H) systems. The theory and spin dynamics are explored in detail, and the effects of a number of multiple pulse sequences are discussed.

The last part deals with an experiment measuring the ^(13)C chemical shift tensor in K_2Pt(CN)_4Br_(0.3) • 3H_2O, a one-dimensional conductor. The ^(13)C spectra are strongly affected by ^(14)N quadrupolar interactions via the ^(13)C - ^(14)N dipolar interaction. Single crystal rotation spectra are shown.

An appendix discussing the design, construction, and performance of a single-coil double resonance NMR sample probe is included.

I. Nuclear spin-internal-rotation-coupling. II. Fluorine spin-rotation interaction and magnetic shielding in fluorobenzene

Part I.

The interaction of a nuclear magnetic moment situated on an internal top with the magnetic fields produced by the internal as well as overall molecular rotation has been derived following the method of Van Vleck for the spin-rotation interaction in rigid molecules. It is shown that the Hamiltonian for this problem may be written

H_SR = Ῑ · M · Ĵ + Ῑ · M” · Ĵ”

Where the first term is the ordinary spin-rotation interaction and the second term arises from the spin-internal-rotation coupling.

The F¹⁹ nuclear spin-lattice relaxation time (T₁) of benzotrifluoride and several chemically substituted benzotrifluorides, have been measured both neat and in solution, at room temperature by pulsed nuclear magnetic resonance. From these experimental results it is concluded that in benzotrifluoride the internal rotation is crucial to the spin relaxation of the fluorines and that the dominant relaxation mechanism is the fluctuating spin-internal-rotation interaction.

Part II.

The radiofrequency spectrum corresponding to the reorientation of the F¹⁹ nuclear moment in flurobenzene has been studied by the molecular beam magnetic resonance method. A molecular beam apparatus with an electron bombardment detector was used in the experiments. The F¹⁹ resonance is a composite spectrum with contributions from many rotational states and is not resolved. A detailed analysis of the resonance line shape and width by the method of moments led to the following diagonal components of the fluorine spin-rotational tensor in the principal inertial axis system of the molecule:

F/Caa = -1.0 ± 0.5 kHz

F/Cbb = -2.7 ± 0.2 kHz

F/Ccc = -1.9 ± 0.1 kHz

From these interaction constants, the paramagnetic contribution to the F¹⁹ nuclear shielding in C₆H₅F was determined to be -284 ± ppm. It was further concluded that the F¹⁹ nucleus in this molecule is more shielded when the applied magnetic field is directed along the C-F bond axis. The anisotropy of the magnetic shielding tensor, σ_” - σ_⊥, is +160 ± 30 ppm.

Carbon-13 magnetic resonance spectroscopy

High-resolution, natural-abundance ¹³C spectra have been obtained from a wide variety of organic compounds; ¹³C chemical shifts and coupling constants have been correlated with other molecular properties.

Geminal and vicinal, carbon-proton couplings in benzene and the five- and six-membered aromatic heterocycles have been related to the corresponding proton-proton couplings in substituted ethylenes. The carbon-proton coupling constants in benzene are J_CCH = + 1.0, J_CCCH = +7.4 and J_CCCH = -1.1 Hz. Extended Hückel wavefunctions are uniformly poor in explaining the long-range, carbon-proton couplings in aromatic systems.

Couplings between carbon and elements other than hydrogen have been observed in proton decoupled ¹³C spectra. All of the carbons in fluorobenzene and 1-fluoronaphthalene, but only six of the carbons in 2-fluoronaphthalene are coupled to the fluorine. One-bond, carbon-phosphorus coupling in trialkylphosphines is negative, while one-bond, carbon-phosphorus coupling in tetra-alkylphosphonium ions is positive. Atoms which do not use hybrid orbitals to form bonds to carbon (F, P(III), Se, Te) may have negative, one-bond coupling constants because of the failure of the average energy approximation. One-bond couplings between carbon and carbon, silicon, tin, lead and mercury appear to be explainable in terms of an effective nuclear charge and the s-bond order of the metal. Couplings between carbon and nitrogen and phosphorus (IV) have significant negative contributions to the Fermi contact coupling expression, though, within one series, correlations with s-bond order may be valid. Carbon-carbon coupling in cyclopropane derivatives (10-15 Hz) is consistent with a high degree of p character in the interior orbitals. Some two- and three-bond carbon-carbon coupling constants have also been observed.

Substituent effects of hydroxyl groups on the ¹³C chemical shifts of continuous-chain alkanes depend both on steric and electronic factors. The hydroxyl substituent effects in the long-chain, primary alcohols are α = -48.3, β = -10.2, and γ = +6.0 ppm. The upfield γ effect is attributed to steric crowding in the gauche conformations. Additivity of the hydroxyl and carbonyl and alkyl substituent effects in alkyl-substituted cyclohexanols and cyclohexanones has been demonstrated.

Proton magnetic resonance studies of ribonucleic acid complexes. I. Complexes of biological bases and oligonucleotides with RNA. II. Template recognition and the degeneracy of the genetic code

Part I. Complexes of Biological Bases and Oligonucleotides with RNA

The physical nature of complexes of several biological bases and oligonucleotides with single-stranded ribonucleic acids have been studied by high resolution proton magnetic resonance spectroscopy. The importance of various forces in the stabilization of these complexes is also discussed.

Previous work has shown that purine forms an intercalated complex with single-stranded nucleic acids. This complex formation led to severe and stereospecific broadening of the purine resonances. From the field dependence of the linewidths, T₁ measurements of the purine protons and nuclear Overhauser enhancement experiments, the mechanism for the line broadening was ascertained to be dipole-dipole interactions between the purine protons and the ribose protons of the nucleic acid.

The interactions of ethidium bromide (EB) with several RNA residues have been studied. EB forms vertically stacked aggregates with itself as well as with uridine, 3'-uridine monophosphate and 5'-uridine monophosphate and forms an intercalated complex with uridylyl (3' → 5') uridine and polyuridylic acid (poly U). The geometry of EB in the intercalated complex has also been determined.

The effect of chain length of oligo-A-nucleotides on their mode of interaction with poly U in D₂0 at neutral pD have also been studied. Below room temperatures, ApA and ApApA form a rigid triple-stranded complex involving a stoichiometry of one adenine to two uracil bases, presumably via specific adenine-uracil base pairing and cooperative base stacking of the adenine bases. While no evidence was obtained for the interaction of ApA with poly U above room temperature, ApApA exhibited complex formation of a 1:1 nature with poly U by forming Watson-Crick base pairs. The thermodynamics of these systems are discussed.

Part II. Template Recognition and the Degeneracy of the Genetic Code

The interaction of ApApG and poly U was studied as a model system for the codon-anticodon interaction of tRNA and mRNA in vivo. ApApG was shown to interact with poly U below ~20°C. The interaction was of a 1:1 nature which exhibited the Hoogsteen bonding scheme. The three bases of ApApG are in an anti conformation and the guanosine base appears to be in the lactim tautomeric form in the complex.

Due to the inadequacies of previous models for the degeneracy of the genetic code in explaining the observed interactions of ApApG with poly U, the "tautomeric doublet" model is proposed as a possible explanation of the degenerate interactions of tRNA with mRNA during protein synthesis in vivo.

Studying conscious and unconscious vision with functional Magnetic Resonance Imaging : the BOLD promise

Waking up from a dreamless sleep, I open my eyes, recognize my wife’s face and am filled with joy. In this thesis, I used functional Magnetic Resonance Imaging (fMRI) to gain insights into the mechanisms involved in this seemingly simple daily occurrence, which poses at least three great challenges to neuroscience: how does conscious experience arise from the activity of the brain? How does the brain process visual input to the point of recognizing individual faces? How does the brain store semantic knowledge about people that we know? To start tackling the first question, I studied the neural correlates of unconscious processing of invisible faces. I was unable to image significant activations related to the processing of completely invisible faces, despite existing reports in the literature. I thus moved on to the next question and studied how recognition of a familiar person was achieved in the brain; I focused on finding invariant representations of person identity – representations that would be activated any time we think of a familiar person, read their name, see their picture, hear them talk, etc. There again, I could not find significant evidence for such representations with fMRI, even in regions where they had previously been found with single unit recordings in human patients (the Jennifer Aniston neurons). Faced with these null outcomes, the scope of my investigations eventually turned back towards the technique that I had been using, fMRI, and the recently praised analytical tools that I had been trusting, Multivariate Pattern Analysis. After a mostly disappointing attempt at replicating a strong single unit finding of a categorical response to animals in the right human amygdala with fMRI, I put fMRI decoding to an ultimate test with a unique dataset acquired in the macaque monkey. There I showed a dissociation between the ability of fMRI to pick up face viewpoint information and its inability to pick up face identity information, which I mostly traced back to the poor clustering of identity selective units. Though fMRI decoding is a powerful new analytical tool, it does not rid fMRI of its inherent limitations as a hemodynamics-based measure.

I. The effect of N,N,N',N' -tetramethylethylenediamine on the Schlenk Equilibrium. II. The nature of the Di-Grignard reagent

I. The influence of N,N,N’,N’-tetramethylethylenediamine on the Schlenk equilibrium

The equilibrium between ethylmagnesium bromide, diethylmagnesium, and magnesium bromide has been studied by nuclear magnetic resonance spectroscopy. The interconversion of the species is very fast on the nmr time scale, and only an averaged spectrum is observed for the ethyl species. When N,N,N’,N’-tetramethylethylenediamine is added to solutions of these reagents in tetrahydrofuran, the rate of interconversion is reduced. At temperatures near -50°, two ethylmagnesium species have been observed. These are attributed to the different ethyl groups in ethylmagnesium bromide and diethylmagnesium, two of the species involved in the Schlenk equilibrium of Grignard reagents.

II. The nature of di-Grignard reagents

Di-Grignard reagents have been examined by nuclear magnetic resonance spectroscopy in an attempt to prove that dialkylmagnesium reagents are in equilibrium with alkylmagnesium halides. The di-Grignard reagents of compounds such as 1,4-dibromobutane have been investigated. The dialkylmagnesium form of this di-Grignard reagent can exist as an intramolecular cyclic species, tetramethylene-magnesium. This cyclic form would give an nmr spectrum different from that of the classical alkylmagnesium halide di-Grignard reagent. In dimethyl ether-tetrahydrofuran solutions of di-Grignard reagents containing N N,N,N’,N’-Tetramethylethylenediamine, evidence has been found for the existence of an intramolecular dialkylmagnesium species. This species is rapidly equilibrating with other forms, but at low temperatures, the rates of interconversion are reduced. Two species can be seen in the nmr spectrum at -50°. One is the cyclic species; the other is an open form.

Inversion of the carbon at the carbon-magnesium bond in di-Grignard reagents has also been studied. This process is much faster than in corresponding monofunctional Grignard reagents.

Studies of dinucleoside monophosphates and monomer-polynucleotide interactions by proton magnetic resonance

The nature of the intra- and intermolecular base-stacking interactions involving several dinucleoside monophosphates in aqueous solution have been investigated by proton magnetic resonance spectrosocopy, and this method has been applied to a study of the interaction of polyuridylic acid with purine and adenosine monomers.

The pmr spectra of adenylyl (3' → 5') cytidine (ApC) and cytidylyl (3' → 5') adenosine (CpA) have been studied as a function of concentration and temperature. The results of these studies indicate that the intramolecular base-stacking interactions between the adenine and cytosine bases of these dinucleoside monophosphates are rather strong, and that the stacking tendencies are comparable for the two sequence isomers. The chemical shifts of the cytosine H₅ and adenine H₂ protons, and their variations with temperature, were shown to be consistent with stacked conformations in which both bases of the dinucleoside monophosphates are preferentially oriented in the anti conformation as in similar dApdC, and dCpdA (dA = deoxyadenosine; dC = deoxycytidine) segments in double helical DNA. The intramolecular stacking interaction was found to have a pronounced effect on the conformations of the ribose moieties, and these conformational changes are discussed. The concentration studies indicate extensive self-association of these dinucleoside monophosphates, and analysis of the concentration data facilitated determination of the dimerization constant for the association process as well as the nature of the intermolecular complexes.

The dependence of the ribose conformation upon the extent of intramolecular base-stacking was used to demonstrate that the base-base interaction in cytidylyl (3' → 5') cytidine (CpC) is rather strong, while there appears to be little interaction between the two uracil bases of uridylyl (3' → 5') uridine (UpU).

Studies of the binding of purine to several ribose and deoxyribose dinucleoside monophosphates show that the mode of interaction is base-stacking, and evidence for the formation of a purine-dinucleoside monophosphate intercalated complex is presented. The purine proton resonances are markedly broadened in this complex, and estimates of the purine linewidths in the complex and the equilibrium constant for purine intercalation are obtained.

A study of the interaction of unsubstitued purine with polyuridylic acid at 29°C by pmr indicated that purine binds to the uracil bases of the polymer by base-stacking. The severe broadening of the purine proton resonances observed provides strong evidence for the intercalation of purine between adjacent uracil bases of poly U. This interaction does not result in a more rigid or ordered structure for the polymer.

Investigation of the interaction between adenosine and polyuridylic acid revealed two modes of interaction between the monomer and the polymer, depending on the temperature. At temperatures above 26°C or so, monomeric adenosine binds to poly U by noncooperative A-U base stacking. Below this temperature, a rigid triple-stranded 1A:2U complex is formed, presumably via cooperative hydrogen-bonding as has previously been reported.

These results clearly illustrate the importance of base-stacking in non-specific interactions between bases, nucleosides and nucleotides, and also reveal the important role of the base-stacking interactions in cooperatively for med structures involving specific base-pairing where both types of interaction are possible.

I. Configurational stability and redistribution equilibria in organomagnesium compounds. II. Redistribution equilibria in organocadmium compounds

I. CONFIGURATIONAL STABILITY AND REDISTRIBUTION EQUILIBRIA IN ORGANOMAGNESIUM COMPOUNDS

The dependence of the rate of inversion of a dialkylmagnesium compound on the solvent has been studied.

Examination of the temperature dependence of the nuclear magnetic resonance spectrum of 1-phenyl-2-propylmagnesium bromide in diethyl ether solution indicates that inversion of configuration at the methylene group of this Grignard reagent occurs with an approximate rate of 2 sec^-1 at room temperature. This is the first example of a rapid inversion rate in a secondary Grignard reagent.

The rates of exchange of alkyl groups between dineopentylmagnesium and di-s-butylmagnesium, bis-(2-methylbutyl)-magnesium and bis-(4, 4-dimethyl-2-pentyl)-magnesium respectively in diethyl ether solution were found to be fast on the nmr time scale. However, the alkyl group exchange rate was found to be slow in a diethyl ether solution of dineopentylmagnesium and bis-(2-methylbutyl)-magnesium containing N, N, N', N'-tetramethylethylenediamine. The unsymmetrical species neopentyl-2-methylbutyl-magnesium was observed at room temperature in the nmr spectrum of the solution containing the diamine.

II. REDISTRIBUTION EQUILIBRIA IN ORGANOCADMIUM COMPOUNDS

The exchange of methyl groups in dimethylcadmium has been studied by nuclear magnetic resonance spectroscopy. Activation parameters for the methyl group exchange have been measured for a neat sample and for a solution in tetrahydrofuran. The exchange is faster in the basic solvent tetrahydrofuran relative to the neat sample and in tetrahydrofuran solution is retarded by the solvating agent N, N, N’, N’-tetramethylethylenediamine and greatly increased by cadmium bromide. The addition of methanol to a solution of dimethylcadmium in tetrahydrofuran appears to have very little effect on the rate of exchange. The exchange was found to proceed with retention of configuration. The rate-limiting step for the exchange of methyl groups in a basic solvent appears to be the dissociation of coordinating solvent from dimethylcadmium.

The equilibrium between methylcadmium bromide, dimethylcadmium and cadmium bromide in tetrahydrofuran solution has also been studied. At room temperature the interconversion of the species is very fast on the nmr time scale but at -100° distinct absorptions for methylcadmium bromide and imethylcadmium are observed.

The species ethylmethylcadmium has been observed in the nmr spectrum.

The rate of exchange of vinyl groups in a solution of divinylcadmium in tetrahydrofuran has been found to be fast on the nmr time scale.