TECHNISCHE UNIVERSITÄT MÜNCHEN

DEPARTMENT FÜR CHEMIE

Design and Characterization of Tectones based on

Guanidinium-Oxoanion Interactions for the Assembly

in Water

Laxman H. Malge

Vollständiger Abdruck der von der Fakultät für Chemie der Technischen Universität München

zur Erlangung des akademischen Grades eines Doktors der Naturwissenschaften

genehmigten Dissertation.

Vorsitzender: Univ.-Prof. Dr. M. Schuster

Prüfer der Dissertation: 1. Univ.-Prof. Dr. F. P. Schmidtchen

2. apl. Prof. Dr. P. Härter

Die Dissertation wurde am 17.06.2010 bei der Technischen Universität München eingereicht

und durch die Fakultät für Chemie am 14.07.2010 angenommen.

To my parents

Acknowledgements

Work presented in this thesis was carried out from April 2006 to April 2010 in the Department of

Organic Chemistry and Biochemistry at Technical University of Munich.

I would like to thank Prof. Dr. F. P. Schmidtchen for his continuous encouragement and support. The

work would not have been completed without his patience and understanding. His vast experience in

the field of organic and supramolecular chemistry helped me to overcome every obstacle during the

work. His timely advice during the difficult times helped me a lot to come out.

I would like to thank Prof. Vladimir Kral for fruitful discussions during his short visits to Munich.

My special thanks go to Dr. Tomas Briza, Dr. Robert Kaplanek and Dr. Bohumil Dolensky for the long

discussions held during their stay in the laboratory and outside.

I would like to thank Mrs. Otte, Mr. Kaviani and Mr. Cordes for providing me mass spectra of the

compounds prepared in this work. I would also be thankful to Dr. Bettina Bechlars for making available

the X-ray crystal structure.

I would take the opportunity to thank Dr. Vinod Jadhav and Dr. Wiebke Antonius for their continuous

help throughout these years. I wish to thank Ashish Tiwari for his help during the stay in laboratory.

I thank all my friends Dr. Sriram Kotkar, Dr. Nagendra Kondekar, Dr. Kulbhushan Durugkar, Dr.

Rameshwar Patil, Dr. Namdev Vatmurge, Dr. Amol Kendhale, Mr. Awadut Giri, Mr. Pandurang

Chouthaiwale and Mr. Ganesh Jogdand for their unwavering support extended to me since campus

days. I am indebted to my brother Mr. M. L. Chapale for his belief in my ability to do the Ph. D.

I would be grateful to my parents and family members for their love and constant encouragement

throughout my studies.

Finally I would like to thank my wife Pallavi for her love, affection and unassuming support extended to

me during this work.

TABLE OF CONTENTS

1. Introduction .............................................................. 1

1.1. Intermolecular non-covalent interactions ...................................................1 1.1.1. Hydrogen bonding ............................................................................................................ 2 1.1.2. Hydrophobic effect............................................................................................................ 3 1.1.3. Van der Waals forces ....................................................................................................... 5 1.1.4. π-π interactions ................................................................................................................ 5 1.1.5. Electrostatic effects........................................................................................................... 6

1.2. Survey of artificial receptors for polar solvents...........................................7

2. Aim of this work ..................................................... 20

3. Synthesis ................................................................ 24

3.1 Synthesis of the hydrophilic bicyclic guanidinium host 34 ...........................24

3.2 Synthesis of the hydrophilic bicyclic guanidinium host 51 ...........................25

3.3 Synthesis of the tetra-cyclic guanidinium host 68........................................32

3.4 Attempted synthesis of the lactone guest 36...............................................32

3.5 Synthesis of the aromatic phosphinate guest 37.........................................38

4. Results and discussion of binding studies ........ 47

4.1 ITC titrations in Water .................................................................................48 4.1.1 ITC titration of host 28 with guest 37 in Water ....................................................................... 48 4.1.2 ITC titration of host 28 with guest 119 in Water ..................................................................... 49 4.1.3 ITC titration of host 34 with guest 37 in Water ....................................................................... 50

4.2 ITC titrations in DMSO................................................................................51 4.2.1 ITC titration of host 28 with guest 37 in DMSO...................................................................... 51 4.2.2 ITC titration of host 34 with guest 37 in DMSO...................................................................... 53 4.2.3 ITC titration of host 51 with guest 37 in DMSO...................................................................... 53 4.2.4 ITC titration of host 51 with guest 119 in DMSO.................................................................... 54

4.3 MD simulations ...........................................................................................56 4.3.1 MD simulations in H2O .......................................................................................................... 57 4.3.2 MD simulations in DMSO ...................................................................................................... 59 4.3.3 MD simulations in MeOH....................................................................................................... 61 4.3.4 MD simulations in CHCl3 ....................................................................................................... 63

5. Experimental Part .................................................. 66

5.1 Reagents, Methods and Materials ..............................................................66

5.2 Experimental Procedures............................................................................69

5.3 Experiments in-silico.................................................................................103 5.3.1 Topology file for guanidinium host 51 .................................................................................. 103 5.3.2 Topology file for guanidinium host 34 .................................................................................. 107 5.3.3 Topology file for guanidinium host 28 .................................................................................. 109 5.3.4 Topology file for phosphinate guest 37 ................................................................................ 111

6. Summary............................................................... 116

7. References............................................................ 119

Abbreviations

Ac Acetyl AHP Anilinium hypophosphinate Ala Alanine AMBER Assisted Model Building with Energy Refinement Arg Arginine arom aromatic Bn Benzyl Boc tert-Butoxycarbonyl bp Boiling Point BTSA Bis(trimethylsilyl)acetamide BTSP bis(trimethylsilyl)phosphonite calc calculated CFF Consistent Force Field CHARMM Chemistry at Harvard Molecular Mechanics δ chemical shift d Dublett DABAL Aluminium-1,4-Diazabicyclo[2.2.2]-octane DABCO 1,4-Diazabicyclo[2.2.2]-octane DCM Dichloromethane de diastereomeric excess DMF N,N-Dimethylformamide DMSBT dimethylsilyl-bis (trifluoromethanesulfonate) DMSO Dimethylsulfoxide DNA Desoxyribonucleic acid DPPA Diphenyl phosphoryl azide dppf 1,1'-Bis(diphenylphosphino)ferrocene dppp 1,3-bis(diphenylphosphino)propane DVB Divinylbenzene EDIPA Ethylenediisopropylamine ESI Electrospray Ionization Et Ethyl Fig Figure FT Fouriertransformation GROMACS Groningen Machine for Chemical Simulations HMDS Hexamethyldisilazane HMPA Hexamethylphosphoramide HPLC High Performance Liquid Chromatography HRMS High Resolution Mass Spectrometry IPP Thiamine diphosphate IR Infrared ITC Isothermal Titration Calorimetry

Kass Association Constant LDA Lithiumdi-iso-propylamide Lys Lysine MD Molecular dynamics

Me Methyl MM Molecular Mechanics MOM Methoxymethyl mp Melting Point MS Mass Spectroscopy MTB Methyltributylammonium MTBE Methyl tert.-butyl ether

NAD+ Nicotinamide adenine dinucleotide NAMD Nanoscale Molecular Dynamics NMR Nuclear Magnetic Resonance NOE Nuclear Overhauser Effect Nos Nosyl OPLS Optimized Potential for Liquid Simulations p Quintett ppm parts per million q Quartett QM Quantum Mechanics RCM Ring closing metathesis RMSD Root-Mean-Square-Deviation

Rt Retention Time RT Room Temperature s Singulett SPE Solid Phase Extraction T Temperature t Triplet TBDMS tert.-butyl dimethyl silane TBDPS tert.-butyl diphenyl silane t-Bu tert.-Butyl TEA Triethyl amine Tf Trifluormethansulfonyl TFA Trifluoroacetic acid THF Tetrahydrofuran TLC Thin Layer Chromatography TMA Tetramethylammonium TMS trimethyl silane Ts Tosyl UV Ultraviolett Val Valine VIS Visible AMP adenosine monophosphate GMP guanosine monophosphate UMP uridine monophosphate

1

1. Introduction

Supramolecular chemistry as defined by Nobel Laureate Jean-Marie Lehn “the chemistry beyond the

molecule”, is the designed chemistry of the intermolecular bond. It is a highly interdisciplinary science

which comprises all three traditional fields of science, chemistry (organic, inorganic and physical

chemistry for creating substances and understanding their thermodynamics), physics (for

computational purposes and analysis), and biology (to understand the similar natural processes).

When a substrate binds to an enzyme or a drug to its target, highly selective interactions occur

between the partners that control the processes. Supramolecular chemistry is concerned with the

study of the basic features of these interactions and with their application in biological as well as in

non-natural (artificial) systems[1, 2].

One of the ambitions of a supramolecular chemist is to synthesize receptors with high binding affinity

and a high selectivity for the substrate molecules in water. Most of the receptors have so far been

studied in organic solvents, but the fact is all of the recognition processes in nature take place in

water. For this very reason designing artificial systems that work in water is challenging as well as

interesting, giving the potential for direct manipulation of biosystems at the molecular level.

The design of artificial hosts which can be used in water poses the following main challenges:

I. The host needs to be soluble in water.

II. The special interactions and approaches have to be chosen to overcome the competitive

influence of the water.

Despite these difficulties, recently there has been a steady increase of artificial systems designed to

work in water for example - molecular receptors (cyclodextrins,[3, 4] crown ethers[5] and

azamacrocycles[6]) and self-assemblies (capsules,[7] metal-organic macro-cycles,[8] hydrogels,[9]

polymerosomes[10] and fibers[11]).

1.1. Intermolecular non-covalent interactions

The study of non-covalent interactions is important to understand biological processes for example,

interactions between a substrate and an enzyme. Biological systems are often the inspiration for

supramolecular research. Unlike traditional chemistry which focuses on the covalent bond,

supramolecular chemistry explores the weaker and reversible non-covalent interactions between

molecules. These forces include hydrogen bonding, hydrophobic forces, van der Waals forces, π- π

interactions and electrostatic effects.

2

1.1.1. Hydrogen bonding

Hydrogen bonds are omnipresent and are described as ‘master key interactions in supramolecular

chemistry’.[12] They arise from the interaction of a hydrogen atom attached to an electronegative atom

(donor, D) and a proximate molecule or functional group (acceptor, A) (fig. 1.1.1-a).

Fig.1.1.1-a: Schematic representation of hydrogen bonding

These interactions are directional and their strength ranges from 4 to 120 kJ mol-1 e.g. amines and

guanidines (-N···H) are hydrogen bond donors whereas carboxylates, phosphates or phosphinates act

as hydrogen bond acceptor molecules.

For example, the double-helix of DNA in biological systems represents the real-life example of

hydrogen bonding. In DNA, base pairs are held together by several hydrogen bond donors and

acceptors (Figure 1.1.1-b).

Fig.1.1.1-b: Hydrogen bonding between Cytosine and Guanine

Of late, there are some reports suggesting the hydrogen bonding between a hydrogen atom attached

to carbon, instead to an electronegative atom such as N and O (electronegativities: C: 2.55, H: 2.20,

N: 3.04, O: 3.44). Usually these interactions are weak, but can be significant depending upon the

acidity of C – H proton[13] (figure 1.1.1-c).

3

Fig.1.1.1-c: (a) X-ray crystal structure showing C—H···N (2.21 Å) and C—H···O (2.41 Å, average)

hydrogen bonding in a complex of with nitromethane crown ether (b)

1.1.2. Hydrophobic effect

Hydrophobic or solvophobic effects are the causes for exclusions of insoluble molecules from the

polar solvents, particularly water. For example, oil in water or n-hexane in water. Organics or the

simple hydrocarbons like alkanes have little attraction for each other. However, water molecules

attract one another very strongly. The hydrophobic effect drives the hydrocarbons together in water

resulting in agglomeration of these non-polar organic molecules. It plays the important role in

biological molecular recognition and it also contributes in protein folding, membrane formation and

many times, small molecule binding by receptors in water.

Hydrophobic effects can be observed from two different expressions. One is the immiscibility of

hydrocarbons in water, which is studied by considering ∆G° for the transfer of organic molecules from

the gas phase or hydrocarbon solution to water. The other expression is the tendency of organics to

associate or aggregate in water, which is studied by measuring ∆G° of association or binding

constant. Immiscibility is the characteristic feature of hydrophobic effect, besides other thermodynamic

effects like entropy and large heat capacity. To a very good approximation, ∆G° of transfer relates with

surface area of the hydrocarbon that is exposed to water on dissolution.

The hydrophobicity of organic groups can also be measured by the partitioning of organic molecules

between n-octanol and water. It was found that small organic substituents make constant and additive

contributions to the hydrophobicity of a molecule. This supports the view that hydrophobicity arises

simply from the surface area of the group, and is not much affected by the environment. Once they are

4

in water, hydrocarbons try to minimise the exposure to water in two ways: by changing their shape or

through aggregation e.g. change from anti-conformation of n-butane in water to the more compact

gauche-conformation is an example of shape change, while the formation of micelles of stearic acid in

water is an example of aggregation.[14]

Hydrophobic effects are crucial for the binding of organic compounds in polar solvents or water and

can be divided into two energetic components: enthalpic and entropic. The enthalpic component of the

hydrophobic effect can be described as follows. It is a well known fact that water has a high cohesive

energy and equally high surface tension. For dissolution of a hydrocarbon solute in water, it has to

create a cavity in water which involves a significant penalty for breaking water-water interactions.

Second, hydrocarbons are not soluble in water therefore very little binding occurs between the solute

and the solvent which otherwise, would have compensated for the lost interactions between solvent

molecules. Moreover, hydrocarbons are polarisable and water is much less so, water would prefer to

interact with water while hydrocarbons would rather bind hydrocarbons on account of their lower

cohesive energy and low surface tension. All these factors constitute the enthalpic component of

hydrophobic effect but the observation of thermodynamic signature of the hydrophobic effect suggests

there is an entropic component too.

The entropic component of the hydrophobic effect can be explained by considering the following

model (figure 1.1.2). Assume that two hydrocarbon molecules (a host shown in green and a guest

shown as blue) are put into water. Water molecules surrounding the hydrocarbon surface area lose

the water-water contacts because of the cavity created by hydrocarbon molecules.

Fig. 1.1.2: Schematic illustration of hydrophobic effects in water

To compensate for the loss of water-water contacts, these molecules strengthen the remaining water-

water contacts by increasing the strength and number of individual water hydrogen bonds around the

solute. During the process, the enthalpy of water before and after dissolution of solute remains nearly

the same but certainly the entropy becomes more negative which leads to the low solubility of the

5

organic structure. This is an example of enthalpy-entropy compensation, where decreased enthalpy

leads to decreased entropy, too.

On the other hand, upon dimerization or binding of the host to the guest, the surface area exposed to

the water decreases which leads to either an unfavourable enthalpy or a close-to-zero enthalpy

change. But there is a definite increase in the disorder of the water due to the release of water

molecules from the hydrophobic area to the bulk solvent. Hence the association is entropically

favourable. The net effect is that the T∆S° term outweighs the ∆H° term, producing a favourable ∆G°.

Therefore, the hydrophobic association is entropy driven.

1.1.3. Van der Waals forces

Van der Waals or London dispersion forces are weak electrostatic attractions or repulsions (figure

1.1.3) occurring from the polarization of the electron cloud of one molecule when it comes in the

vicinity of the other molecule’s nucleus. The most important feature of these interactions is that the

energies of these interactions are distance dependent e. g. induced-dipole–induced-dipole or van der

Waals interactions (r-6). They are non-directional and are found in ‘inclusion’ complexes in

supramolecular chemistry.

Fig. 1.1.3: Van der Waals interactions

1.1.4. π-π interactions

These are the weak interactions between two aromatic rings among others. As depicted in the figure

1.1.4-a, π-π interactions can be of two types, face-to-face, where centre of one aromatic ring interacts

with corner of the other ring and edge-to-face, where hydrogen atom from one ring interacts in a

perpendicular orientation with respect to the centre of the other ring. The π- π interactions arise from

the attraction between the negatively charged π-electron cloud of one conjugated system and the

positively charged sigma framework of a neighbouring molecule.[15]

6

Fig.1.1.4-a: Different geometries of aromatic π-π interactions

Graphite is an example of π-π stacking interactions as shown in figure 1.1.4-b. The layered structure

of graphite is held together by weak, face-to-face π-π interactions.

Fig.1.1.4-b: Face-to-face π-π interactions in graphite, top view (a) and side view (b)

1.1.5. Electrostatic effects

Electrostatic interactions are a very important class of noncovalent interactions which are ubiquitous in

biological systems. Moreover, these are often the major interactions between the substrate and an

enzyme. They can be classified further as ion-ion interactions, ion-dipole interactions and dipole-dipole

(r-3) interactions (figure 1.1.5). Ion-ion interactions are non-directional and the strongest of the class

whereas ion-dipole and dipole-dipole interactions are directional and their strength depends upon the

orientation of the molecules involved.

7

Fig. 1.1.5: Examples of electrostatic interactions

1.2. Survey of artificial receptors for polar solve nts

Supramolecular chemistry in polar solvents or water is a constantly growing area of research because

noncovalent interactions in aqueous media are necessary for a better understanding of biological

systems.[16] Polar solvents or water present a challenge, as individual solvent molecules interact

intermolecularly by forming a dynamic network of hydrogen bonds. On the other hand, polar molecules

experience strong hydration by water and participate in the hydrogen bonding network, which

dramatically influences the properties of the solvated species. These properties of water provide two

main challenges for supramolecular chemistry in aqueous media. One is how to achieve high water

solubility and second, how to avoid, minimize or exploit the strong involvement of water in noncovalent

processes.

In order for binding to occur between a host and a guest, the energy barrier of host-solvent or guest-

solvent has to be overcome. This desolvation process has both enthalpic and entropic consequences.

Enthalpically, energy must be given to break the solvent-host or solvent-guest bonds. The release of

solvent molecules from host and guest to the bulk leads to increase in the entropy and also the

formation of solvent-solvent bonds.

Polar solvents particularly inhibit binding of charged species, as the solvent dipole can interact

strongly with charged centers thus making the solvent-host or solvent-guest interactions harder to

break. Also some solvents can disturb the binding through electron-pair or hydrogen bond donation or

acceptance. For example, DMSO can act as both electron-pair donor and hydrogen bond acceptor

due to the presence of oxygen and sulphur lone pairs.

In spite of these challenges there has been continuous improvement in the host-guest chemistry of

artificial receptors designed for polar solvents. The synthetic receptors described herein, are

categorized based on the type of functional group responsible for anion binding, particularly in

aqueous media. The analysis of the energetic signature of the interaction between artificial host and

8

guest by means of, for example, microcalorimetry, was highlighted as it would give an important

information on the enthalpic and entropic components of the complex formation. A description of

recent developments in the field of supramolecular chemistry in polar solvents, with a special

emphasis on water as a solvent, is presented through the following section.

Molecular Tweezers and Clips

Tweezers[17, 18] and clips[18-21] are two-armed acyclic receptors with flexible hydrophobic cavities which

can wrap around guests or clip them between two rigid molecular planes, respectively. Molecular clips

1-2 and tweezers 3-4, features a rigid torus-shaped unpolar cavity decorated with two rotatable

peripheral anionic phosphonate groups. The molecules were designed with a suitably pre-organized

cleft having high electron density on its inner surface, which should facilitate the complete desolvation

of tetrahedral alkylammonium ions.

1 2

3 4

R

R

R = OPMeO2Li R = OPMeO2Li

R

R

R = OPMeO2Li R = OPMeO2Li

R

R

R

R

Klärner and co-workers have recently reported on the water-soluble benzene-spaced molecular clip

1[17] and the benzene-naphthalene-spaced molecular tweezer 4.[19, 21] These receptors bind biologically

important molecules such as cofactors of enzymes and peptides in aqueous solution with remarkable

efficiency and selectivity.

9

Clip 1 strongly binds to enzyme cofactors such as N-methylnicotineamide iodide, nicotinamide

adenine dinucleotide (NAD+) and thiamine diphosphate (TPP) in D2O with Ka = 8.3 x 104 M-1, 9.1 x 103

M-1 and 1.4 x 104 M-1, respectively. Molecular tweezer 4 is an excellent receptor for lysine and

arginine. Tweezer 4 not only selectively recognizes simple protected peptides in water (Ka values for

Ac-Lys-OMe and Ts-Arg-OEt in D2O are 2.3 x 104 M-1 and 7.8 x 103 M-1, respectively), but is also able

to bind lysine or arginine incorporated into a peptidic framework (for example, KKLVFF, the lysine-

containing self-complementary central part of the Alzheimer peptide, is bound with a Ka = 3.8 x 104 M-1

in 25 mM NaH2PO4 buffer in D2O/CD3OD 1:1).[17]

Traditionally, hydrophobic interactions are predicted to increase steadily and incrementally as the

surface area of the solute is enlarged with respect to the well-ordered water molecules in liquid phase.

The release of water molecules into the solvent bulk during the aggregation of the solute leads to an

increase in entropy (classical hydrophobic effect, entropy-driven).[22, 23] Recent investigations of

recognition processes, for example, of host-guest complex formations, enzyme substrate binding, or

DNA intercalation by arenes, however, show that in these cases an enthalpic gain is the origin of

hydrophobic interactions (non-classical hydrophobic effect).[22, 24-26] The molecular clip 2 and tweezer 4

are the nice examples of the non-classical hydrophobic effect as evidenced from their thermodynamic

parameters. By enlarging the respective π-faces of clip 1 and tweezer 3, the authors sought to

improve the recognition abilities of the synthetic receptors 2 and 4 through the increased hydrophobic

interactions. Subsequently, the water-soluble phosphonate substituted benzene-anthracene-spaced

clip 2 and naphthalene-spaced tweezer 4 were synthesized. The molecular clip 2 and tweezer 4

(R=OPMeO2ֿLi+) form highly stable dimers in water [Ka values in D2O at 25°C are 2.28 x 106 M-1 (∆G°

= -36.4 kJ mol-1, ∆H° = -87.4 kJ mol -1, T∆S° = -51.0 kJ mol -1) and 1.6 x 105 M-1 (∆G° = -29.7 kJ mol -1,

∆H° = -57.7 kJ mol -1, T∆S° = -28.0 kJ mol -1), respectively].[18] However, the dimers were dissociated

upon addition of N-methylnicotinamide as a guest to the receptor 2 or 4.

Another interesting example of a molecular tweezer based on the glycouril units, was reported by Lyle

Isaacs and co-workers.[27] The complexation of this type of receptors with dialkylammonium ions, for

example, arises from the hydrophobic interactions between the central polymethylene part of the guest

and the tweezer interior, as well as hydrogen bonding and ion-dipole interactions between the urea

oxygen atoms and the ammonium ends of the guest. The glycouril tweezer 5 is an excellent example

of self-assembly.

10

The glycouril tweezer 5 dimerizes isostructurally in a wide range of solvents from C6D6 to D2O, and

keeps the same association motif both in nonpolar aprotic media, such as chloroform, and in polar

protic competitive solvents,[27] such as methanol and water (dimerization of 5 in D2O Ka = 3.6 x 104

M-1). The behavior is maintained by compensative cooperative hydrogen-bonding and π–π

interactions. Hydrogen bonds provide the main self-association force in nonpolar media, while π–π

interactions dominate in water.[27]

Cyclophanes

The cyclophanes are the macromolecules that contain a bridged aromatic ring forming a cavity which

can bind guests. A variety of groups such as pyridinium,[28, 29] ammonium,[30] carboxylic, phosphonic

moieties and saccharides have been used to solubilize cyclophane scaffolds in water.[31]

Schneider and co-workers have reported on the phenanthridinium-containing cyclophane[29] 6 which

binds AMP (Ka = 6.3 x 105 M-1) selectively over GMP or UMP which have very negligible change in

fluorescence upon binding with 6.

11

Receptor 7 efficiently recognizes benzenetricarboxylates (BTC) in water (for trimesate Ka values are

between 1.5 x 104 M-1 and 3.9 x 106 M-1, depending on the degree of protonation of 7).[30] The

molecular recognition of 7 with different tricarboxylic acids (1,3,5-BTC, 1,2,4-BTC, 1,2,3-BTC and

1,3,5-BTA: benzenetriacetic acid) containing aromatic subunits constitutes a very nice example of

Emil-Fischer lock and key principle.[32, 33] The guest 1,3,5-BTC, fits very well electronically and

stereochemically within the cavity of relatively rigid host species 7. It gives one of the largest

selectivities reported in water [∆ (log K) ca. 2.0 for 1,3,5-BTC-7 over 1,2,3-BTC-7 and for 1,3,5- BTC-7

over 1,3,5-BTA-7 at pH 4].[30]

Another important class of cyclophanes which binds catecholamines selectively and efficiently was

reported by Schrader et al.[34] Adrenaline is the lead compound of a whole class of catecholamine

neurotransmitters and mediates signal transduction across cell membranes.[35] It is a small, highly

polar molecule, which is bound very shortly and efficiently by its natural receptor. This recognition

eventually leads to a conformational change within the transmembrane helices of the receptor and in

turn triggers the activation of the G protein on the cytosolic side of the membrane.[36] A deep binding

pocket is needed to provide a sufficiently hydrophobic environment for complete desolvation of the

charged hormone. Cyclophanes 8 and 9 which contain exo- and endocyclic phosphonate groups are

good receptors for catecholamines in aqueous media.[34, 37-40]

8 9

NH

O O

HN

O2N NO2

O OP P

OO

OO

P

O

OCH3

O

O O

OO OO

P

P

O

OCH3

O

H3CO

O

O

PH3CO

O

O

Li

LiLi

Li

Li

Li

12

The receptor 8 binds adrenaline, noradrenaline and dopamine to form 1:1 complexes in

methanol/water (1:1)[37] where Ka value ranges between 1.5–2.5 x 102 M-1. Two guest molecules can

be bound by cyclophane 9 in water. Although the binding is non-cooperative, its affinity for the guest is

higher than that of receptor 8 toward catecholamine and related structures such as β-blockers with

extended aromatic π- surfaces (Ka values up to 7 x 103 M-1 for each single complexation step or 5 x

107 M-2 for both steps).[34]

Anion recognition is ubiquitous in biological systems which guides the development of efficient artificial

receptors for anions in the modern day supramolecular chemistry.[41] Artificial receptors in which the

substrate is bound by hydrogen bonds as in natural systems are, however, often only active in

nonpolar solvents.[42] For complexation in water, stronger interactions, such as electrostatic or

coordinative interactions, are normally necessary because of the high solvation energy of many

anions.[41] Stefan Kubik and co-workers have developed the cyclic hexapeptide receptor 10.

Cyclopeptide receptor 10 binds sulphate or iodide in aqueous solution to form a 2:1 complex in which

two C3-symmetric receptors 10 provide six hydrogen bonds to a single desolvated anion within a

sandwich-like complex.[43-46] It shows a strong cooperativity for sulphate, K1 ≈ 3.6 x 102 M-1 and K2 ≈

8.8 x 103 M-1 (in D2O/CD3OD 1:1), where K2 is 98 times higher than its statistical value. Such a high

increase in binding strength is due to receptor-receptor interactions[46] which become significant even

in a mixture of water and methanol because of perfect alignment of receptors around the sandwiched

anion.

10

NNH

OO

HN

O N

O

N

O

NH ON

N N

Inspired from the sandwich-like complex-structure of 10 with anion, the authors have extended the

study by incorporating a linker to form an oyster like hexavalent cyclopeptide 11a, in which two

cyclohexapeptide subunits containing L-proline and 6-aminopicolinic acid subunits in an alternating

sequence are connected via an adipinic acid spacer. The receptor 11a effectively increases the

molarity for complexation with the second cyclopeptide.[44] The receptor 11a binds sulphate and iodide

13

in 50% D2O/CD3OD at 25°C with log Ka value 4.55, ∆G° = -26.0 kJ mol -1, ∆H° = -15.0 kJ mol -1, T∆S° =

11.0 kJ mol-1 for sulphate and log Ka = 3.79, ∆G° = -21.6. kJ mol -1, ∆H° = -13.2 kJ mol -1, T∆S° = 8.4 kJ

mol-1 for iodide respectively. Microcalorimetric investigations in this case showed that anion binding is

enthalpically as well as entropically driven.[44]

Moreover, dynamic combinatorial studies to optimize the size and length of the linker gives

cyclopeptides 11b and 11c, which the authors claim are the strongest receptors for inorganic ions in

aqueous media to date. For example, iodide and sulphate binding by 11b and 11c in acetonitrile/water

(2:1) is shown in table1.2.1.[45]

Table 1.2.1: Association constants, Gibbs energies,

enthalpies and entropies of binding of KI and K2SO4 to

receptors 11b and 11ca.

Guest Receptor Ka ∆G° ∆H° T∆S°

11b 2.9 x 104 -25.5 -20.7 4.8 KI

11c 5.6 x 104 -27.1 -13.4 13.7

11b 5.4 x 106 -38.4 1.8 40.1 K2SO4

11c 6.7 x 106 -39.0 3.7 42.7 arecorded in 2:1 (v/v) acetonitrile/water at 298K; binding constants in M-1 and energies in kJ mol-1.

14

Cavitands

Cavitands are synthetic organic compounds with enforced cavities large enough to complex

complementary organic compounds or ions.[47, 48] Cavitands consist of multiple arene rings covalently

linked in a highly constrictive manner to give a well-formed hydrophobic cavity. The parent cavitands

12 (X = -CH2, R1 = alkyl, R2 = H, alkyl) reported by Reinhoudt and co-workers are insoluble in water.

Charged groups,[49-51] saccharide functions[52] and dendritic oxo substituents[50] have been introduced

to the upper and bottom rims to make them soluble in aqueous media.

Cavitand[52] 12 (X = -CH2, R1 = Me, R2 = -CH2NHCSNH-glucose or R2 = -CH2NHCSNH-galactose

moiety) binds acetate in water/acetonitrile (1:1) where the complexation has an unfavourable enthalpy

and is entropically driven (Ka = 2.15 x 103 M-1, ∆G° = -19.0 kJ mol -1, ∆H° = 2.9 kJ mol -1, ∆S° = 73.4

JK-1 mol-1). However, in anhydrous acetonitrile, the acetate complexation is enthalpically driven.

Rebek and co-workers have studied the molecular recognition in aqueous media of a variety of broad

deep cavity cavitands, such as 13,[53] which were made water-soluble by the attachment of

carboxylate, ammonium, or amino groups. For example, cavitand 13 (R = -CH2COOֿ), which has a

solubility of 5 mM in water, forms complexes with a variety of guests such as S-nicotinium,

quinuclidinium (Ka> 104 M-1), tetraalkylammonium bromides (3.8 x 103–1.2 x 104 M-1), L-carnitine (1.5 x

102 M-1), choline chloride (2.6 x 104 M-1), acetylcholine chloride (1.5 x 104 M-1).

15

13

14 15

O

O

N

NH

R

O

O

N

HNR

O

O

N

NHR

O

O

N

NHR

Et Et

Et Et

S PO

OO

O

O

O

N

Na

Na

R = -CH2COO , -p-CH2C6H4COO ; X = Na

R = -CH2NH3 ; X = Br

4X

Cavitand 13 (R = -CH2COOֿ) also forms complexes with the surfactants 14 and 15 so that the long

alkyl chains of the guests spontaneously form a helix upon encapsulation.[54, 55] In this case, the alkyl

chain guests adopt a coiled conformation in order to better fill the hydrophobic cavity and maximize

the CH–π interactions with the aromatic surface of the host. Also it was proved subsequently that the

roles of host and guest are reversed above the critical micellar concentration (cmc) of sodium dodecyl

sulphate (SDS) 14: the cavitand 13 is bound within micelles of the surfactant.[55]

Guanidinocarbonylpyrroles, -pyridines, and -pyrazol es and bicyclic guanidinium cations

The design of artificial receptors for the selective molecular recognition of a given substrate is still a

challenge, especially in polar solvents.[56] A large number of artificial receptors for hydrophobic

solvents such as chloroform have been described.[57] As the polarity of the surrounding solvent

increases, the strength of hydrogen bonds and electrostatic interactions mainly used for molecular

recognition, decreases rapidly, due to the competitive solvation of donor and acceptor sites by the

solvent. Schmuck and co-workers introduced a novel class of receptor molecules, known as

guanidinocarbonylpyrroles, for the binding of carboxylates in aqueous media. The idea was to improve

the binding affinity of guanidinium cations, well known for the complexation of oxoanions in organic

solvents such as CHCl3 or CH3CN,[41] by adding additional binding sites. To achieve the enhanced

binding, they selected substituted 2-(guanidiniocarbonyl)-1H-pyrroles as the core binding motif. The

16

pyrrole NH as well as suitable donor sites in the side chain should be able to bind the carboxylate

through hydrogen bonds in addition to ion pairing with the guanidinium unit.[58]

Schmuck et al. have studied a range of guanidinocarbonylpyrroles[59] 16 and 17 and found that the

combination of additional hydrogen bonding along with electrostatic interactions allows the effective

binding of amino acids and peptides in aqueous solution.

Addition of NH or charged substituents to either the pyrrole or the guanidinium moiety of 16

significantly increases the affinity toward carboxylates. Compound 16 (R1=R2=H) binds Ac-L-Ala-Oֿ

with Ka = 130 M-1 (in water/DMSO 2:3).[60] Attachment of a peptide at the guanidinium moiety leads to

receptor 16 (R1 = H, R2 = CH2CH2CO-Val) which strongly binds carboxylates or amino acids with Ka ≥

103 M-1 in an aqueous buffer solution.[61] Functionalization of the pyrrole moiety gives receptor 16 (R1 =

C(O)NHEt, R2 = H), which binds acetate with Ka ≈ 3 x 103 M-1 and N-acetylated amino acids with Ka =

360–1700 M-1 (water/DMSO 2:3). Extra ionic interactions introduced by the imidazolium moiety in de

novo designed receptor 17 led to the efficient binding of dipeptides in water,[62] with binding constants

up to 5.43 x 104 M-1, which is almost 10 times higher than the binding affinity toward simple amino

acids.

Artificial receptors based on guanidinium and amidinium groups are synthesized to mimic the anion

binding properties of the side chain of arginine. The advantage of guanidinium and amidinium groups

lies in their ability to combine Coulomb attraction in the binding of oxoanions such as carboxylates or

phosphates with the formation of two strong parallel hydrogen bonds. Additionally, both groups remain

protonated over a wide pH range because of their strong basicity (pKa values typically ranging

between 11 and 13).[63]

Kilburn and co-workers synthesized closely related guanidinium based receptors[64] 18 and 19 for

carboxylate binding. ITC data for these receptors were shown in table 1.2.2.

17

Table 1.2.2: Binding data for 18 and 19 as PF6- salts with tetrabutylammonium acetate

Guanidinium salt Solvent Ka (1:1) /M-1 ∆G°

(kJ mol-1)

∆H°

(kJ mol-1)

T∆S°

(kJ mol-1)

18 DMSO 5300a -21.2 -19.4 1.8

19 DMSO 22000a -24.8 -8.0 16.8

19 10% H2O-DMSO 3900b -20.5 -7.4 13.1

19 30% H2O-DMSO 480b -15.3 -2.0 13.3 aITC data were fitted using two site model, giving reported 1:1 binding constants and much smaller 1:2 binding constants (Ka (1:2) < 150 M-1 in each case). bITC data was fitted using one site (1:1) binding model.

Comparison of the data reveals that guanidinium 19 featuring a pyridine moiety provides better binding

results than its analogue 18 where the benzene ring replaces the pyridine moiety. Also it is clear from

the ITC data that the complexation involving 19 was entropically driven whereas, in case of 18 the

complexation was driven mainly by enthalpy.

Schmidtchen and co-workers reported a ditopic water soluble receptor 20 (R = H) based on chiral

bicyclic guanidinium anchor group.[65]

The dichloride salt of receptor 20 forms reasonably stable host-guest complexes with biologically

important phosphates in water (e.g. for HPO42-, Ka = 970 M-1). The compound 20 (R = SiPh2t-Bu) was

18

reported earlier and found to have very good binding with the phosphates in a polar solvent like

methanol (Ka ≈ 1.8 – 3.8 x 104 M-1).

Schmidtchen et al. studied guanidinium-carboxylate interactions by isothermal titration calorimetry

(ITC).[66] The binding constant calculated from ITC titrations for host 21 as the bromide salt and

tetraethylammonium acetate in acetonitrile was found to be Ka = 2.0 x 105 M-1 and the energetic

signature (∆G° = -30.7 kJ mol -1, ∆H° = -15.5 kJ mol -1, ∆S° = 50.2 JK -1 mol-1). In DMSO, the

corresponding binding constant was found to be 30 times lower (Ka = 6.5 x 103 M-1, ∆G° = -22.1 kJ

mol-1, ∆H° = -14.2 kJ mol -1, ∆S° = 26.1 JK -1 mol-1). The results indicate that the binding process was

both entropically and enthalpically favourable for 1:1 complex formation. Thermodynamic parameters

could be derived from both CH3CN and DMSO but in MeOH the heat output was too low to quantify.

This result shows that the stabilization of guanidinium-carboxylate is not only due to electrostatic

interactions (∆H°) but also due to favourable release of solvent molecules (∆S°), which strongly

emphasizes the importance of solvation in host-guest interactions, a factor often neglected in receptor

design.

Recently Schmidtchen and Jadhav reported on the complexation between 22 and a variety of

phosphates of different sizes. The complex formation was investigated by both 1H NMR and ITC

techniques in acetonitrile.[67] For 22, 1H NMR gave a satisfactory fit to a 1:1 stoichiometry model

whereas ITC predicted a 1:2 host-guest binding model and the complexation was driven mainly by

entropy. Hence ITC can prevent the misleading predictions by NMR when rapidly interconverting

species are in equilibrium. It also cautions against the incorporation of several H-bond donors in a

scaffold which could act opposite to the enthalpic stabilization of a complex.

Anslyn and co-workers developed a tripodal guanidinium based receptor 23 which shows very high

selectivity for citrate 24 in water (Ka = 6.9 x 103 M-1, 1H NMR titrations).[68] This host was able to

complex citrate even from a crude orange juice which indicates its high selectivity towards citrate in

presence of carboxylates. It is clear from the results that positioning of several hydrogen bond donors

in a suitable platform can overcome the solvent effect also.

19

Over the last decade, arginine based peptides were used for transport of non-permeable molecules

into the cytoplasm which opens the new field for guanidine-based synthetic receptors as potential drug

carriers. De Mendoza and co-workers recently reported on a non-peptidic oligoguanidinium salt 25

which readily internalizes into human cancer cells and hence could be a potential drug transporter.[69]

20

2. Aim of this work

Biological systems are the key motivation behind the artificial supramolecular host or guest designs. In

biological systems such as enzymes and antibodies, the guanidinium groups present in the side chain

of the amino acid arginine, form strong noncovalent interactions with anionic groups like carboxylates,

phosphates, sulphates and nitrates through hydrogen bonding and electrostatic interactions.[41]

Guanidiniums are also responsible for stabilization of protein tertiary structures via internal salt bridges

mainly with carboxylates.[70] Owing to the importance of the guanidinium functionality in biological

systems, it is obvious to find them in many drug substances as well as a binding motif in molecular

recognition studies involving artificial receptors.[71, 72]

Bicyclic guanidines are different from their acyclic counterparts in physical, chemical and electronic

properties.[73] The rigidity of the structure is responsible for their unique properties. Here the hydrogen

bond donors are properly aligned to interact with an oxygen atom from the carboxylates or phosphates

and the guanidinium cation complements through electrostatic interactions with the anion (fig. 2.1).

Figure 2.1: Sketch of the bicyclic guanidinium motif interacting with an oxoanion

Besides hydrogen bonding and charge-charge interactions, solvent also has a role to play and

influences the energetic outcome of the host-guest system. In polar solvents such as water, the

strength of hydrogen bonds and electrostatic interactions decreases rapidly as the polarity of the

surrounding solvent increases.[74-76] In the preceding work in our laboratory, a trend analysis was

performed involving closely related guanidinium hosts (26-30) and phosphinic acid guests (31-33) (fig.

2.2) with increasing congestion around the binding site to explore the structure-activity relationship in

acetonitrile, a relatively polar solvent, keeping the ambiguous interactions at minimum.[77] In the series

of hosts and guests, the most congested phosphinate 33 binds tightly with the least congested host 26

corroborating the hypothesis that assembly occurs with the shielding of respective binding sites. The

thermodynamic data obtained from ITC, indicates maximum negative enthalpy for the ion-pair (26·33)

confirming the dominance of the classical ionic-hydrogen bonding interactions (fig. 2.1) over the total

21

interactions. The increase in steric bulk around the binding site deteriorates the proper arrangement of

binding partners, thereby raising the association enthalpy to more positive values. The ordinary

geometric fit allows more flexibility between host and guest which was reflected in more positive

entropies of association. It was proved that the positive entropy changes during the complexation are

not only due to the release of solvent molecules to the bulk (desolvation) but also the possible

changes in the configuration resulting from thermal population of a variety of distinct binding modes

(potential minima) that includes the participation of solvent molecules.[77]

Figure 2.2: Ensemble of guanidinium hosts 26-30 and phosphinate guests 31-33 prepared earlier by

Schmidtchen and Haj-Zaroubi.[77]

NH

N

NH

O OSi SitBu tBu N

H

N

NH

NH

N

NH

H3C

H3C

CH3

CH3

NH

N

NH N

H

N

NH

PO O

PO O

P

O O

O

TBA TBATBA

26 27 28

29 30

31 32 33

Moreover, the replacement of phosphinate 33 with the more relaxed ‘bay phosphinate’ 32 or the

parent phosphinate 31 does not lead to the increased affinity with the series of hosts 26-30 indicating

the existence of a broad variety of complex configurations rather than a single binding mode to

represent the associated host-guest pair. The detailed analysis of energetic signatures of both the

phosphinates 32 and 31 with the series of hosts (26-30) deduced that the positive change in entropies

22

of associations was not only due to the solvent reorganisation but also because of the direct host-

guest binding. The change in association entropy can be related to the tightness of the mutual fit of the

host-guest partners, which approaches a minimum limit and is interpreted as the unique lock-and-key

binding mode. The obvious extension of the work would be the study of the complexation

phenomenon in water involving the artificial receptors to appreciate the mechanism of natural

processes such as an enzyme binding to a substrate. This would definitely be a step closer to the

understanding of natural or biological systems.

In the present work, the main goal is to investigate pair-wise interaction of the guanidinium hosts with

phosphinate guests in water and polar solvents like dimethyl sulfoxide and methanol using ITC

techniques. In order to achieve this goal, the design and synthesis of a host-guest pair was planned

which can meet the following criteria:

1. The synthetic hosts and guests should be soluble in polar solvents like water, DMSO and

methanol.

2. The ion-pair should complex in such solvents to produce a measurable energetic response

enabling us to predict distinct binding modes of the ion-pair.

Unlike the guanidinium cation 29 (fig. 2.2), which was prepared by guanidinylation of its corresponding

tetraphenyl substituted trisamine salt with thiocarbonyldiimidazole;[78, 79] guanidinium host 35 (fig. 2.3)

required a new synthetic route due to the lack of its analogous tetra substituted 4-hydroxyphenyl

compound. It was premeditated to synthesize α,α,α’,α’-tetrasubstituted guanidinium cation 35 with 4-

hydroxy phenyl moiety as the substituent. The flanking 4-hydroxy phenyl moieties would protect the

guanidinium core from solvation as well as facilitate the solubility of the host in polar solvents like

water and methanol. Similarly, another tetra-substituted host 34 based on four hydroxypropyl chains

around the guanidinium core was planned to be synthesized according to the literature.[80] The desired

purpose of protecting the solvation shell would be served with the less sterically demanding propyl

chains and the terminal hydroxyl functionalities would help in dissolving the molecule in polar

solvents.[80, 81] Allyl guanidinium host 28[80] as its bromide salt would be included in the study which

completes the ensemble of guanidinium cations with varied flexibility and steric congestion nearby the

binding site.

Phosphinate guests 36, 37 and 38 (fig. 2.3), based on the similar concept of protecting the binding

sites from the solvation with adjoining hydrophobic groups were designed. The synthetic procedures

used for the preparation of phosphinates 32 and 33 (fig. 2.2) can not be used for the synthesis of the

phosphinates 37 and 38 as the related starting materials are not available commercially. Therefore,

both the aliphatic and aromatic phosphinates (36-38) required a novel synthetic method in view of their

expected roles in the present study.

23

Aliphatic phosphinate 36 was conceptualized expecting the rigid spiro-lactone rings would form a

hydrophobic shell in the vicinity of the phosphinate anion and the sulphone-lactone functionalities

would help in solubility of the entire compound in polar solvents.

The aromatic phosphinates 37 and 38 differing in the bite angle around the phosphinate anion were

appended on both sides with phenyl acetylene groups adjacent to the solvation shell of phosphinate

anion anticipating that these groups will minimize the solvation of the anion. Furthermore, polyhydroxy

functionalities were constructed far remote from the phosphinate anion which would enhance the

overall solubility of the guest molecules yet would not interfere in the complexation process.

Figure 2.3: Proposed synthesis of guanidinium hosts (34-35) and phosphinate guests (36-38) in the

present work

On successful preparation of the ensemble of host and guest molecules (fig. 2.3), the ITC and NMR

titrations will be carried out to study the structure-activity relationships of the ion-pairs.

24

3. Synthesis

3.1 Synthesis of the hydrophilic bicyclic guanidini um host 34

The synthesis of guanidinium host 34 was started off with the hydroboration-oxidation of allyl

guanidine 28a.[82] The hydroboration–oxidation reaction is a two-step reaction that converts an alkene

into an alcohol by the net addition of water across the double bond. The hydrogen and hydroxyl group

are added in a syn addition leading to cis stereochemistry. It is an anti-Markovnikov reaction, with the

hydroxyl group attaching to the less-substituted carbon. The tandem procedure for the synthesis of

guanidinium host 34 is depicted in scheme 3.1.[80, 81] The first step consisted of a simple addition

reaction between the 2-methyl-2-butene 39 and borane-THF complex 40, both commercially available,

to give the disiamyl borane 41.[83] Followed by the addition of tetra-allyl guanidine 28a to the disiamyl

borane 41 prepared in situ afforded an intermediate tetra-substituted guanidine-borane-complex. The

disiamyl borane reagent 41 was chosen for the hydroboration of allyl guanidine 28a due to its well

known ability to give exclusively the anti-Markonikov’s product on account of its steric bulk. Hydrolysis

of the intermediate borane-complex using sodium hydroxide and hydrogen peroxide 42 gave

guanidinium host 34. The guanidinium host 34 was precipitated as its tetra-phenyl borate salt in 51%

yield by treating it with aqueous sodium tetraphenyl borate 43. Allyl guanidine 28a[66, 82, 84] was

obtained by de-protonation of tetra-allyl guanidinium iodide 28 with sodium hydroxide in CCl4 at room

temperature.

Scheme 3.1: Synthetic strategy for the guanidinium host 34

25

An attempt to prepare host 34 as its bromide salt directly by the addition of 47% HBr solution

immediately after basic hydrolysis had failed. The host 34 could have been degraded because of

harsh basic conditions followed by drastic acidic conditions. Ultimately, the bromide and perchlorate

salts of the guanidinium cation 34 were obtained by ion exchange using BioRad AG 4-X4 resin

acrylic matrix ion exchanger in the corresponding bromide or perchlorate form in aqueous methanol,

followed by crystallization from acetonitrile.

3.2 Synthesis of the hydrophilic bicyclic guanidini um host 51

The retro-synthetic analysis for the preparation of the hydrophilic bicyclic guanidinium host 51 is

depicted in scheme 3.2.1.

Scheme 3.2.1: Retro-synthetic route for the guanidinium host 51

HO

HO

OH

OH

NH

N

NH

BrN

OMe

OMeMeO

OMe

NH2NH2

N

MeO

MeO

OMe

OMeN NC C

OO

N

MeO

MeO

OMe

OMe

CONH2 H2NOC

MeO

CONH2

OMe

NI I

51

CN

CNCN

CN

MeO

COOEt

OMe MeO

CCl3

OMe

4445

47

46

4948

50

26

It was conceived that the preparation for the host 51 would start off with the conversion of

commercially available 1,1,1-trichloro-2,2-bis(p-anisyl)ethane or methoxychlor 44 to the corresponding

ester derivative 45 by a known procedure.[85] The ester compound 45 would be transformed into its

amide derivative 46 which in turn, would give an intermediate precursor 48 upon alkylation with bis-

iodoethylcyanamide 47. The Hofmann rearrangement[86] of the obtained amide precursor 48 would

yield the bis-bisisocyanate compound 49 which on hydrolysis would afford the corresponding bisamine

compound 50. The bisamine 50 would be transformed into bicyclic guanidinium host 51 utilizing the

cyano-carbon for guanidylation in ethanol/HCl (Pinner reaction)[87-90] (cf. scheme 3.2.1).

The synthesis was started with the preparation of ethyl 2,2-bis(4-methoxyphenyl)acetate 45 from the

commercially available methoxychlor 44 following a reported procedure[85]. Methoxychlor 44 was

refluxed with silver nitrate in absolute ethanol to obtain the corresponding ester 45. The yield of

compound 45 was disappointing (15%), rather the major product was found to be the corresponding

ketone, bis(4-methoxyphenyl) methanone 52 (85% yield) [(M+H)+ = 243] (scheme 3.2.2).

Scheme 3.2.2: Synthesis of ethyl 2,2-bis(4-methoxyphenyl)acetate 45

An attempted conversion of the ester compound 45 to its amide derivative using a variety of reagents

viz. aq. NH4OH, NH3 in ethanol and NH4Cl with catalytic AlMe3 failed to give the desired 2,2-bis(4-

methoxyphenyl)acetamide 46. Moreover, an attempt to hydrolyse the ester to its corresponding acid

by refluxing in aq. NaOH also failed (scheme 3.2.3).

Scheme 3.2.3: Attempted synthesis of 2,2-bis(4-methoxyphenyl)acetamide 46

27

Alternatively, the alkylation of the ester derivative 45 with bis-iodoethylcyanamide 47 was probed

using LDA/THF, but failed to achieve the conversion. On account of the modest yield of the required

ester 45 and subsequent failure to get the corresponding amide 46 or acid, it was essential to

formulate a new approach for the synthesis of host 51.

The modified synthetic plan for preparation of the hydrophilic bicyclic guanidinium host 51 was based

on the Curtius rearrangement of bisacid 57 to give the related bisamine in place of the Hofmann

degradation of the analogous bisamide (scheme 3.2.4). The desired precursor 57 for the Curtius

reaction would be synthesized by treating the monoacid 55 with bis-iodoethyltosylamide 56. The

monoacid 55 would be easily prepared from glyoxalic acid 53 and anisole 54.

Scheme 3.2.4: The modified retro-synthetic path for the intermediate 57

N

S

MeO

MeO

OMe

OMe

COOH HOOC

OO

MeO

COOH

OMe

OMe

OHCCOOH H2O

N

S OO

I I

5755 56

54 53

The monoacid 55[91] was produced in about 80% yield following a known procedure where glyoxalic

acid 53 was condensed with anisole 54 in glacial acetic acid in presence of catalytic sulphuric acid at

0°C (scheme 3.2.5). Alkylation of compound 55 was carried out with bis-iodoethylcyanamide 47 (cf:

scheme 3.2.1) and LDA/THF to get the related bisacid analogue of 57. The resulting bisacid was

treated with oxalyl chloride or DPPA to prepare the corresponding bisacid chloride or azide but in both

cases an unwanted reaction occurred at the CN-group (-CN becomes -CONH2). The cyano-group in

the bisacid was found to be very sensitive toward moisture and led to its hydrolysis product urea. The

purpose of using –CN protection (for the Pinner reaction) of the amine was not served in this case;

therefore it was mandatory to replace it with a more stable protecting group such as a p-

toluenesulfonamide group. Subsequently, the compound 47 was replaced by the analogous bis-

iodoethyltosylamide 56 in the alkylation step.

28

Scheme 3.2.5: Synthesis of the intermediate bisacid 57

Alkylation of the compound 55 was effected with compound 56 and in situ prepared LDA in absolute

THF at -10°C to obtain 57 in 72% yield. The compound 61 (scheme 3.2.6) was planned to be formed

by Curtius reaction of the bisacid 57. The acyl azide 59 required for the Curtius rearrangement would

be prepared by reacting bisacid compound 57 with diphenylphosphoryl azide (DPPA) 58.[92]

Successive thermal decomposition of 58 would yield the corresponding bis-isocyanate 60 which on

hydrolysis would render the desired bisamine 61.

The bisacid compound 57 was first converted into the corresponding bisacid azide by treating it with

DPPA 58 in dry dichloromethane at room temperature. Curtius rearrangement of the resulting acyl

azide 59 was carried out by refluxing in absolute toluene to yield the corresponding bisisocyanate 60.

After completion of the reaction, toluene was evaporated and the residue was dissolved in

tetrahydrofuran. The THF solution was acidified with dilute HCl and heated to reflux till the hydrolysis

was complete. The bisamine 61 was isolated from the crude, but the yield was very low. The low yield

of bisamine 61 was probably due to the polymerization during Curtius rearrangement under normal

heating of acyl azide in toluene. To overcome the polymerisation, it was decided to trap the isocyanate

in the form of its more stable urethane form before converting it to the corresponding bisamine 61

under milder conditions. As an alternative, the microwave irradiation[93, 94] was opted for its mild and

quick nature of heating.

During the course of reaction of the acyl azide 59 and allyl alcohol under microwave irradiation to

prepare the corresponding urethane or carbamate, to our surprise, the clean bis-isocyanate 60 (a

strong band in IR at 2250 cm-1, characteristic of –C=N bond) was obtained instead of the urethane. All

the three reactions were carried out in tandem starting from acyl azide till bisamine stage to avoid

unnecessary time and exposure for the intermediates to degrade. A decent 45% overall yield for the

three successive steps was achieved (scheme 3.2.6).

29

Scheme 3.2.6: Synthetic strategy for the trisamine 62

Deprotection of tosyl group in compound 61 was achieved by employing a well known method of

dissolving sodium metal in liquid ammonia.[95] The conversion afforded about 78% trisamine 62 as

pale yellow liquid under basic workup conditions. The compound 62 was immediately used for further

cyclization reaction. The guanidylation of 62 was carried out using thiocarbonyldiimidazole 63 and

methyl iodide 64 in acetonitrile to give the bicyclic guanidinium compound 65 (scheme 3.2.7).

Demethylation of compound 65 was achieved by using 1M solution of BBr3 66[96] in dichloromethane at

-78°C which afforded bicyclic guanidinium host 51 in 80% yield. The guanidinium host 51 as the

bromide salt was found to be water soluble at a modest one millimolar concentration. The X-ray crystal

structure of the final host 51 is shown in fig. 3.2.1.

30

Scheme 3.2.7: Synthetic strategy for the hydrophilic host 51

Figure 3.2.1: X-ray crystal structure of the host 51 as its bromide salt

31

Table 3.1: Data from crystal structure analysis of guanidinium host 51 as its

bromide salt

Compound Name Guanidinium host 51 Formula C31H30BrN3O4 ⋅ 1.75 NC2H3 ⋅ 0.75H2O M (g/mol) 673.85 Crystal description Colorless fragment Crystal dimensions (mm3) 0.23x0.18x0.08 Temperature (K) 173(2) crystal system, space group

Orthorhombic, Cmc21

a (Å) 30.8939(11) b (Å) 21.3922(7) c (Å) 19.6064(9) α (°) 90 β (°) 90

γ (°) 90 V (Å3) 12957.6(9) Z 16 dcalc (g/cm3) 1.382 F000 5600

µ (mm-1) 1.317

Index ranges (±h, ±k, ±l) -35/37, -25/23, -23/23,

θ ranges (°) 1.16-25.39 Collected refelections 51728 Unique reflections [all data]

11720

Rint/Rσ 0.0293/0.0350

Unique reflections [I0>2 σ(I0)] 10297 Data/Restraints/Parameter 11720/1/844 GoF (on F2) 1.027 R1/wR2 [I0>2 σ(I0)]

0.0436/0.1227

R1/wR2 [all data]

0.0500/0.1185

Max./Min. residual electron density 1.973/-0.763

Since it was not possible to describe a disorder of some solvent molecules correctly, the electron

density for these molecules (96 electrons) was removed by SQUEEZE, which then allowed further

refinement. The data, however, is not of good enough quality to reliably discuss the extracted bond

distances and angles since they contain significant errors. Nevertheless, the identity of the

guanidinium host 51 is proved unambiguously.

32

3.3 Synthesis of the tetra-cyclic guanidinium host 68

Scheme 3.3.1: Synthetic strategy of the tetra-cyclic guanidinium compound 68

Synthesis of 68 (scheme 3.3.1) was carried out by using the well known procedure for ring closing

metathesis (RCM)[97]. The allyl guanidinium compound 28 does not have good solubility in acetonitrile

nor in dichloromethane. Hoveyda-Grubb’s second generation catalyst 67 was used for its efficacy and

solubility in methanol along with substrate 28. The RCM was completed in about 3h at room

temperature using 10 mol% of the catalyst 67. Further modifications at the double bonds on both ends

of the tetra-cyclic compound 68 were envisaged to minimize the solvation at the core guanidinium

functionality and maximize the overall solubility in polar solvents, but the reaction was not carried out.

3.4 Attempted synthesis of the lactone guest 36

As shown in scheme 3.4.3 (p. 35), depicted the strategy towards guest 36, the synthesis of the key

intermediate spiro-lactone 74 was started off using Michael-type addition reaction. The synthesis for

the compound 74 is shown in scheme 3.4.1. The Michael-type addition reaction between enolate of

ethyl bromopyruvate 69 and tetrahydrothiopyran-4-one 72 will lead to an intermediate lactone 73

which upon usual oxidation reaction gives the desired spiro-lactone 74. The enolate of ethyl

bromopyruvate 71 was prepared by treating ethyl bromopyruvate 69 with lithiumdiisopropylamide

(LDA) in absolute tetrahydrofuran at -78°C. The successive reaction of the enolate 71 with thiopyran-

4-one 72 did not work even at room temperature. The disappointing results were probably due to the

low stability of the lithium enolate and/or the less reactivity of the carbonyl group of thiopyranone at the

working temperatures. Attempts were made to maximize the reactivity and stability of the substrates

by silylation[98] of ethyl pyruvate as well as its enolate, but failed to synthesize the compound 74 using

the Michael-type addition reaction.

33

Scheme 3.4.1: Synthetic route to intermediate 74

Br

COOEt

O

N Li

THF

-78oC

Br

COOEtOLi

S

O

O OH

LiBr

EtOH

S

O

S

O

O OH

O O

oxidation

74

73

72

71

70

69

Br

Br

As an alternative, dimethylsilylbis(trifluoromethanesulfonate)[99] [DMSBT, Me2Si(OTf)2] was prepared

from the reaction of dichlorodimethylsilane and trifluoromethane sulphonic acid at 60-70°C for 12 h.

DMSBT is a strong silylating agent which should help in stabilisation of the enolate and simultaneously

activate the carbonyl carbon of thiopyranone. Moreover, the reaction between thiopyran-4-one 72 and

DMSBT in presence of the EDIPA as a base did not take place.[100]

An attempt was made to prepare a similar guest 82 inspired by the known synthesis of 4-

chlorobenzene lactone 79[101] as depicted in the scheme 3.4.2. A simple base catalyzed condensation

between 4-chlorobenzaldehyde 75 and pyruvic acid 76 afforded the unsaturated acid 77 in about 80%

yield. Molecular bromine was added to the unsaturated acid 77 in chloroform at 0°C to yield a white

crystalline dibromide 78 in 70-80% yield. On heating in water at 65°C, compound 78 cyclised to the

corresponding lactone 79.

To synthesize the phosphinic acid guest 82 from the intermediate 4-chlorobenzene lactone 79, an

Arbuzov reaction[102-104] was employed. Ammonium phosphinate and hexamethyldisilazane were

heated together at 110°C under nitrogen atmosphere to afford the reactive intermediate bis-

(trimethylsilyl)-phosphonite (BTSP)[103] 81 which was diluted with anhydrous dichloromethane at room

temperature. The BTSP solution thus prepared was made to react with lactone 79 at ambient

temperature to generate the desired phosphinic acid 82.

Although the model reactions of BTSP with benzyl bromide and ethyl acrylate produced the

corresponding phosphinic acids, compound 79 did not yield the phosphinic acid 82 at the given

conditions.

34

Scheme 3.4.2: Synthetic route for 4-chlorobenzene lactone based guest 82

An attempt was made to increase the electrophilicity at the β-carbon of α, β-unsaturated lactone 79 by

protecting the hydroxyl group by TMS, TBDMS or TBDPS group. Several attempts to either activate or

react compound 79 with a BTSP reagent prepared in situ or as a solution failed to give the desired

compound 82.

The synthetic strategy shown in scheme 3.4.1 did not work, however, it was obvious that a very strong

and stable nucleophile was required to attack the relatively unreactive carbonyl group of the

thiopyranone. Moreover, it was desired to have a clean and neat BTSP[105] reagent for the synthesis of

the phosphinic acid compounds.

A literature[106] for the synthesis of pyruvic acid dianion equivalent used to create a similar spiro-

lactone starting from cyclohexanone was followed for the preparation of spiro-lactone 74. The

dimethylhydrazone of pyruvic acid 85 from stoichiometric amounts of dimethyl hydrazine and pyruvic

acid 76 was synthesized. The corresponding dianion 86 was generated by treating this substance with

MeLi in THF / HMPA as a yellow solution (scheme 3.4.3). This was found to be very reactive even with

the oxidised thiopyranone-4-one 84. The spiro-lactone 74 was prepared in 65% yield using the dianion

86. The compound 84 in turn, was obtained in 75% yield by oxidizing tetrahydrothiopyran-4-one 72

with Oxone monosulphate 83 in acetonitrile at room temperature.[107]

Moreover, the BTSP reagent 81 was prepared by heating together at 100°C ammonium

hypophosphite and HMDS. When ammonia evolution stopped, the mixture was vacuum distilled at 51-

54°C and 11 mbar pressure under inert atmosphere to yield the pure reagent 81.[105]

35

Scheme 3.4.3: Synthetic strategy for aliphatic phosphinate guest 36

The BTSP reagent 81 was carefully stored under argon and handled with caution during reactions

since it is extremely pyrophoric and can explode when it comes in contact with moisture. The reactions

between the reagent BTSP 81 and various analogues of spiro-lactone 74 under different conditions

are listed in table 3.4.1.

Table 3.4.1: Reactions between BTSP 81 and spiro-lactones 74 and 87-91

Spiro-

lactone

BTSP

Mol% Solvent

Temperature /

time Reagents Product

74 200 DCM r.t. TMSCl / TEA No reaction

87 200 DCM r.t. / O/N* TMSCl / TEA No reaction

88 200 Benzene r.t. TMSCl / TEA No reaction

89 100 Benzene 0°C – reflux / 1 h TMSCl / TEA No reaction

90 200 THF r.t. TMSCl / TEA No reaction

Benzyl

bromide 50 THF r.t. – reflux / 1 h TMSCl / TEA

dibenzylphosphinic

acid

* O/N: overnight

All endeavour to trigger some reaction between different spiro-lactones and BTSP resulted only in

failure. To our knowledge there was no such reaction reported on vinylic carbon and nucleophilic

phosphorus to form a C-P bond under these conditions. There was no doubt left about the quality and

36

reactivity of BTSP as it was evident from its reaction with benzyl bromide. The non-reactivity of the

spiro-lactone could be due to its unique structure (figure 3.4.1). The electrophilicity of β-carbon varies

with the type of group attached to the α-carbon. In case of the spiro-lactone 87, both electron-donating

(-OMe, 88) as well as electron-withdrawing (-OTBDPS, 90) groups coupled with bromine as better

leaving group on β-carbon were attempted to react with the BTSP reagent 81, but no reaction

occurred.

Figure 3.4.1: Chem3D model of spiro-lactone 87

Alternatively, we sought a different approach based on the phosphate-phosphonate rearrangement as

portrayed in scheme 3.4.4.[108]

Scheme 3.4.4: Schematic portray of phosphate-phosphonate type rearrangement of 74

The reactions of spiro-lactone 74 with methyl dichlorophosphate 92 under different conditions were

arranged in table 3.4.2. Most of the reactions show conversion of starting lactone 74, but the product

was too elusive to be isolated.

37

Table 3.4.2: Reactions of 74 with methyl dichlorophosphate 92

Substrate Base Solvent Temp. Result

74 EDIPA DCM 0°C degraded

74 DABCO DCM 0°C DABCO-Cl

74 DABCO DCM r.t. DABCO-Cl

74 HMPA DCM 0°C no reaction

74 piperidinomethyl on

polystyrene resin 2% DVB DCM r.t. S.M.*

74 aq. NaOH / cat. Triton B DCM r.t. no reaction

74 LDA THF -10°C no reaction

90 pyridine benzene 80°C no reaction

91 imidazole DMF 50°C no reaction

*S.M.: starting material, consumed but no desired product obtained

Reaction with EDIPA as a base yielded a product, but this was very unstable and degraded in the

NMR tube at room temperature before analysis was complete. However, there was a positive

indication that the lactone reacted in few cases to obtain a new product even though it was unstable.

Moreover, a number of palladium-catalyzed C-P bond formation reactions were carried out on various

analogues of 74 using anilinium hypophosphinate (AHP)[109] under different conditions, but without

success. Montchamp[110-113] and co-workers reported palladium catalyzed cross-coupling reaction of

AHP with a variety of electrophiles such as Ar-X[114] and alkenyl halides.[115] We explored some

structurally related reactions from these references to suit our substrate and the results are collected

in table 3.4.3.

On the contrary, cross-coupling reactions of compounds containing two phosphorus-hydrogen bonds

are much less common, because of the possibility for competing transfer hydrogenation with these

substrates.[116] In fact, the preparatively useful transfer hydrogenation of alkenes, alkynes, aldehydes,

ketones and aryl halides is well precedented to take place with hypophosphorus acid or its sodium and

amine salts, under the influence of almost all transition-metals[117]. The reaction is believed to occur via

insertion of the metal into P-H bond with subsequent formation of metal hydride which is the

catalytically active reducing agent. In order to achieve the desired C-P bond formation, the metal must

instead insert into a C-X bond.

38

Table 3.4.3: Catalytic C-P bond formation reactions on 88* and 80§

Lactone Reagent$ Catalyst Solvent T (°C) Result

88 AHP Pd(OAc)2/dppp/

(BuO)4Si CH3CN

r.t. -

70 no reaction

88 AHP Pd(OAc)2/dppp THF reflux no reaction

80 AHP Pd(OAc)2/dppp THF r.t. degradation

80 AHP Pd(OAc)2/dppp THF reflux degradation

80 AHP Pd(OAc)2/Johnphos/D

ABCO CH3CN r.t.

deprotection of

TBDPS

80 AHP Pd(OAc)2/dppf/

(EtO)3Si(CH2)3NH2 CH3CN reflux elimination of Br

80 AHP Pd(OAc)2 /

tBu-X-

Phos/TEA CH3CN 60

Monosubstituted

product

1-bromo-

naphthalene AHP Pd(PPh)4/TEA DMF 85

monosubstituted

product

*Refer to scheme 3.4.3; § Refer to scheme 3.4.2; $AHP: anilinium hypophosphinate

Consequently, the plan to prepare the aliphatic phosphinate 36 was aborted on account of the failure

of both non-catalytic and catalytic synthetic approaches.

3.5 Synthesis of the aromatic phosphinate guest 37

The retro-synthetic analysis for the aromatic phosphinate guest 37 is described in scheme 3.5.1. The

desired phosphinate functional group would be easily incorporated in the initial stages as ethylester by

treating commercially available 3,5-dimethoxy phenol 94 with the ethyl dichlorophosphate 95. The

phosphate-phosphonate rearrangement[108] on the resulting compound 96 would yield the phosphinate

97. The corresponding bistriflate 99 obtained from bisphenol 97 would be cyclized into a tricyclic

compound 102 by using Suzuki coupling[118, 119] reaction. The successive deprotection-alkylation-

deprotection reactions would lead to the compound 113, which on triflation followed by the

Sonogashira coupling[120-122] with phenyl acetylene would give the penultimate phosphinate. This

compound on hydrolysis would afford the hydrophilic aromatic phosphinate 37.

39

Scheme 3.5.1: The retro-synthetic strategy for the aromatic phosphinate guest 37

P

HO O

O O

OHHO

HO OH

OHHO37

P

HO O

O O

OO

O OO

O114

OF

FF

S

O

O

OF

FF

S

O

O

P

HO O OHHO

O O

OO

O OO

O

P

O O OO

O O

OO

O OO

O

109

P

O O OO

HO OH

104

113

P

O O OO

O O

P

O

99

O

O O

F

F

F

S

O

O

O

O

O

OF

FF

S

O

O

OH

MeO OMe

OMe

O

MeO

PO

OMeMeO

O

OEt

OH

MeO

P

O

OEt

OMe

MeO

OH

OMe

949697

102

Synthesis of the aromatic phosphate 96 started off with a relatively straightforward reaction between

3,5-dimethoxy phenol 94 and ethyl dichlorophosphate 95 in a heterogeneous solution of aqueous

sodium hydroxide and dichloromethane in presence of a phase transfer catalyst Triton B (scheme

3.5.2). The compound 96 was isolated in a decent yield of 85–90% as a high boiling colourless liquid.

The compound 96 rearranged to the corresponding phosphinate 97 when treated with LDA in

anhydrous tetrahydrofuran at -78°C in about 80% yield.[108]

40

Scheme 3.5.2: Synthetic route to tetramethoxy compound 102

The conversion of 97 into corresponding bistriflate 99 (scheme 3.5.3) was attempted using standard

reaction conditions, Tf2O in pyridine at 0°C, but instead the compound 100 (m/z = 741) was obtained.

Scheme 3.5.3: Synthesis of the compound 99

A rather expensive reagent N-Phenyl bistrifluoromethanesulphonamide (PhNTf2) 98 was opted for the

desired triflation of compound 97. The desired bistriflate 99 was obtained in 72% yield by treating

compound 97 with PhNTf2 98 and sodium hydride in absolute tetrahydrofuran at 0°C. Afterwards

several attempts were made to optimize the intramolecular Suzuki coupling[118, 119] on bistriflate 99 to

arrive at the tricyclic phosphinate 102 using various catalysts and solvents. All the results obtained are

collected in table 3.5.1.

41

Table 3.5.1: Optimization of intramolecular Suzuki coupling on the bistriflate 99

Sr.

No.

Bis-

triflate

99

(eq.)

Boronic

Ester

101

(eq.)

K3PO4

(eq.) Solvent

Ligand

(mol%)

Palladium

Catalyst

(mol%)

T (°C) Time

(h) Result

1 1 1.5 2.5 DMF TPP

(10)

Pd(PPh3)2Cl2

(10) 100 24

not

completed

2 1 0.5 2 toluene X-Phos

(40) Pd2(dba)3 (20) 100 24

no

reaction

3 1 1 2 t-BuOH X-Phos

(40) Pd(OAc)2 (20) 100 24

no

reaction

4 1 1 3 DMF X-Phos

(40) Pd(OAc)2 (20) 100 24 (1 : 5)*

5 1 1.2 2.5 THF TPP

(15)

Pd(PPh3)2Cl2

(10) reflux 12

no

reaction

6 1 1.4 2.5 DMA dppf

(10)

Pd(dppf)2Cl2

(10) 100 72

no

reaction

7 1 1.2 2.5 dioxane TPP

(15)

Pd(PPh3)2Cl2

(10)

80-

100 12 completed

8 1 1.2 5 DMF TPP

(15)

Pd(PPh3)2Cl2

(10) 100 4 completed

* ratio of the products 102 and 117 respectively

Suzuki coupling of bistriflate 99 in presence of bis-(triphenylphosphine) palladium(II) dichloride,

bis(neopentylglycolato)diboron 101 and triphenyl phosphine as ligand in anhydrous

dimethylformamide at 100°C gave 65% yield of the tricyclic compound 102. However, addition of a

molar equivalent of water to the bistriflate 99 and sodium hydride as a base, bistriflate 99 rendered

another desired compound 117 [(M+H)+ = 381.2] (scheme 3.5.4) which could be transformed

similarly to get a tweezer-like phosphinate 38.

42

Scheme 3.5.4: Synthesis of the compound 117

Several experiments were carried out with compound 102 to prepare exclusively its tetrahydroxy

analogue using boron tribromide in dichloromethane. The temperature of the reaction was varied from

room temperature to reflux and the rate of addition of BBr3 was changed from few minutes to several

hours. Moreover, different dilutions were tried, but the reaction always afforded a mixture of mono-,

di-, tri- and tetra- hydroxy compounds 103, 104, 105 and 106 respectively (scheme 3.5.5). This was

due to the increasing insolubility of the resulting product after each subsequent deprotection of methyl

groups. There was a severe restriction on the use of different solvents owing to their incompatibility

with boron tribromide or the solubility of the substrate 102.

Scheme 3.5.5: Demethylation of the tetramethoxy compound 102

Subsequently, with the progress of the reaction second [(M+H)+ = 337], third [(M+H)+ = 323] and the

fourth methyl group [(M+H)+ = 309] were deprotected, but after the third methyl deprotection the

reaction became very slow. There was a peak at the front in the HPLC chromatogram (with least

retention time), the mass of which corresponds to the completely deprotected compound i.e. the free

phosphinic acid [(M+H)+ = 281]. However, the ultimate output was a mixture of compounds, instead of

any single compound.

43

When the reaction was caried out at lower temperatures (-60°C to -40°C), it was possible to isolate the

bismethoxy compound [(M+H)+ = 337] as a major product. The purified product was subjected to

further analysis by NMR spectroscopy. The 1H NMR spectrum indicated that the isolated compound

was symmetric: the chemical shifts for the aromatic protons have only two peaks (in case of

unsymmetric compound there should be four peaks corresponding to each proton) and there was only

one peak representing the –OMe protons (singlet of 6H). Therefore, the compound must correspond

to either the ortho- (C1 and C4) or the para-methoxy (C2 and C3) groups deprotection (cf. the

structure for 102 above). This can easily be identified from the 2D-NMR spectroscopy (COSY and

NOESY) experiments. If the para-methoxy groups are cleaved, there will be a spatial interaction

between the protons of ortho-methyl group and the protons of ethoxy group on phosphorus in the

NOESY experiment; otherwise there will be no cross peaks.

Meticulous analysis of the NOESY spectrum showed the cross peaks between the methoxy and

ethoxy protons which confirmed the identity of the compound as having methyl groups on C1 and C4

intact. On the other hand, the methyl groups on C2 and C3 were cleaved selectively. The isolated

bismethoxy compound was confirmed to be the compound 104 (scheme 3.5.5).

Consequently the conditions for demethylation of tetramethoxy compound 102 were optimized. The

compound 102 was converted smoothly to 104 in dichloromethane at -40°C in about 60% yield using

neat boron tribromide under strictly anhydrous conditions.

The attempted alkylation of isolated tetrahydroxy compound 106 (scheme 3.5.6) using 4-

(bromomethyl)-2,6,7-trioxabicyclo[2.2.2]octane 107 under different conditions is illustrated in table

3.5.2.

Scheme 3.5.6: Alkylation of the tetrahydroxy compound 106

44

Table 3.5.2: Alkylation of compound 106 using 107

Sr. No.

106 mmol

107 mmol Base T (°C) Time Solvent Result

1 0.16 0.36 K2CO3 reflux O/N§ acetone no reaction 2 0.16 0.36 K+ -OtBu reflux 8 h acetone no reaction 3 0.16 0.36 NaH 100 2 h DMF no reaction 4 0.032 0.07 K2CO3 150 O/N DMF degradation 5 0.08 0.18 K2CO3 155 O/N cyclohexanone no reaction 6 0.08 0.36 K2CO3 155 O/N cyclohexanone no reaction 7 0.08 0.18 K2CO3 r.t. - 100 O/N cyclohexanone no reaction 8 0.032 0.07 P1Base r.t. - 60 O/N acetonitrile no reaction 9 0.032 0.07 K2CO3 100 - 250 24 h sulpholane no reaction

10 0.032 0.08 NaH r.t. - 180 12 h DMSO degradation 11 0.10* 0.10 K2CO3 reflux O/N acetone no reaction

12 0.10* 0.20 K2CO3 155 4.5 h cyclohexanone mono alkyl. (M-H)ֿ= 355.2

*Substrate was Bisphenol A; §O/N: overnight

During workup of the reaction between the compound 106 and the compound 107, the bicyclic ortho-

ester rings were found to be very unstable toward acid leading to the opening-up of the ortho-ester

rings in presence of moisture. Therefore, the more stable analogue 4-(bromomethyl)-1-methyl-2,6,7-

trioxabicyclo[2.2.2]octane 108 was probed for the alkylation reaction. The compound 109 was

synthesized using the compound 108 and Cs2CO3 as a base in DMF in a moderate 65% yield

(scheme 3.5.7). A direct Suzuki coupling between the compound 109 and 2-phenyl-1-ethynylboronic

acid pinacol ester 123 using a Nickel catalyst was tried unsuccessfully to prepare the compound

110.[123]

Scheme 3.5.7: Synthetic strategy for the compound 110

45

In continuation of the envisioned synthetic strategy (cf. scheme 3.5.1) for the aromatic phosphinate 37,

the next step involved the deprotection of remaining methyl groups at ortho-position of the compound

109.

Scheme 3.5.8: Synthetic strategy for the hydrophilic aromatic phosphinate 37

P

O O OHHO

O O

OO

O OO

O

113

PhNTf298

heptyl thiol /NaH

53%

pH = 7-8

Na

112

P

O O

O O

OHHO

HO OH

OHHO

115

116

P

O O

O O

OHO

HO OH

OHO37

OO

P

O O OTfTfO

O O

OO

O OO

O

114

1) 4N NaOH

2) TFA

pH = 5- 6

2) TFA

NaNa

Na

109

1)

The boron tribromide reagent 66 could not be used for demethylation due to its high Lewis acidity

which would catalyze the opening-up of the ortho-ester rings of the compound 109 in presence of a

trace of the moisture. Therefore, a basic reagent was required for the demethylation of the compound

109. Sodium heptyl thiolate 112[124] was chosen due to its strong nucleophilicity and its ability to

catalyze the cleavage of aromatic methyl groups. The compound 109 and the sodium heptyl thiolate

112 in DMF were heated to 120°C to yield the corresponding demethylated product, but in the process

the ethyl group on phosphorus was also cleaved and afforded the corresponding phosphinic acid 113

(scheme 3.5.8). To avoid the opening-up of the ortho-ester rings, the product was isolated at pH~6-7

as the sodium phosphinate 113 in about 53% yield.

A tandem reaction sequence was followed to achieve the final phosphinate 37 starting from the

compound 114. The triflation of the compound 113 was carried out using PhNTf2 98 and sodium

46

hydride as a base in absolute DMF at 0°C. The DMF was removed by Kugelrohr distillation and the

product was used in situ for further modifications. Sonogashira coupling[120-122] of the resulting product

114 was carried out in anhydrous acetonitrile with phenyl acetylene 115, (PPh3)4Pd, TPP and triethyl

amine at reflux temperature. After completion of the reaction, the final phosphinate 37 was isolated

after acid-base-acid hydrolysis of the Sonogashira product 116 formed in situ. The output was a

modest 25% over three steps as a sodium salt of the desired aromatic phosphinic acid 37.

47

4. Results and discussion of binding studies

The experimental observations from ITC titrations made by using the specially designed bicyclic

guanidinium hosts and phosphinate guests for the polar solvents are discussed in this section. The

complexation of the host-guest ion pairs could be better understood if the free energy ∆G° of

association is dissected into the corresponding enthalpy ∆H° and entropy ∆S° of binding. As

demonstrated earlier, the enthalpy and the entropy of the binding compensate each other[125, 126],

therefore it is essential to measure the magnitude of each component individually.

Calorimetric measurements were conducted using an isothermal titration calorimeter (MCS-ITC,

Microcal, USA), and employed carefully degassed and filtered (0.2 mm Teflon syringe filter) solutions

of the host and the guest compounds in absolute DMSO (Fluka, water ≤ 0.005%) and deionised water.

All titrations were corrected by corresponding blind titrations to eliminate systematic errors (heats of

dilution, calibration errors etc.). Data analysis relied on Origin 5.0 software supplied by the instrument

manufacturer. The choice of the binding model and the subsequent fit procedure used various

(commonly 15–20) different starting parameter sets to raise the probability of finding the global error

minimum. Usually convergence was obtained after less than 50 iteration steps.

Table 4.1: Summary of ITC titrations of various host-guest ion pairs at 303 K

Sr.

No. Host Guest

Ka

[M-1]

-∆H°

[kcal mol-1]

∆S°

[cal K-1 mol-1]

T∆S°

[kcal M-1]

-∆G°

[kcal M-1]

Water

1 28 37 69150 ± 2700 3.24 ± 0.01 +11.5 +3.5 6.7

2 28 119 --- --- --- --- ---

3 34 37 --- --- --- --- ---

DMSO

4 28 37 6184 ± 103 7.18 ± 0.03 -6.2 -1.9 5.3

5 34 37 1950 ± 69 8.64 ± 0.19 -13.4 -0.6 8.1

6 51 37 542 ± 17 7.97± 0.71 -13.8 -4.2 3.8

7 51 119 2953 ± 169 3.02 ± 0.08 +5.9 +1.8 4.8

A typical ITC plot (cf. figure 4.1.1) from an ITC titration can be graphed into two panels: an upper

panel corresponding to the differential heat pulses (absorbed or released) over time and the lower

panel indicating the binding isotherm generated by integration of each peak plotted versus the molar

48

ratio. The titration curve gives the molar enthalpy ∆H° as the step height and the free energy ∆G° from

the slope at the inflection point. The molar entropy ∆S° could be easily calculated from the Gibb’s-

Helmholtz equation and the stoichiometry n is derived from the curve-fit as an independent parameter.

The titrations of hosts 28, 34 and 51 with the phosphinate guests 37 and 119 were carried out in water

and DMSO at 30°C (303 K) as represented in table 4.1.

4.1 ITC titrations in Water

4.1.1 ITC titration of host 28 with guest 37 in Wat er

The one-site binding model curve-fitting shows a 1:1 stoichiometry of host-guest association. The

calculation of the slope at the inflection point gives a very high association constant (Ka ≈ 70,000). The

huge binding constant obtained was a result of the higher positive entropy as well as the favourable

enthalpy. Ostensibly, the higher value of the entropy could be attributed to the desolvation process

occurring at the guanidinium core and the phosphinate “bay” during the binding.

In a high dielectric (ε ≈ 80 at 20°C) solvent like water it was expected to obtain a higher positive value

for the entropy (favourable) of association between the host 28 and the guest 37 due to the release of

solvent molecules from the respective hydrophobic solvation shells of the host and the guest

molecule. The host 28 and the phosphinate guest 37 were tailor-made to suit the polar solvents like

water with respect to the solubility and the hydrophobicity or solvophobicity. The resulting

complexation between the host and the guest was anticipated to maximize the favourble enthalpy (<

0) and the favourable entropy (> 0) of association.

The ITC titration (fig. 4.1.1) clearly indicates the expected results of negative enthalpy and positive

entropy of association with a huge binding constant in water. The solvent reorganization may not be

responsible solely for the large positive entropy of association observed in the experiment, however

there are enough reasons to believe that it must be arising due to the direct host-guest bindding.[77]

The binding studies involving the series of hosts and guests including the allyl guanidinium host 28

and the analogous phosphinate guest 32 (cf. section 2) in a relatively polar but aprotic solvent

acetonitrile (ε ≈ 36) revealed that there exists a broad variety of complex configurations rather than a

single binding mode to represent the associated host-guest pair.[77]

49

Figure 4.1.1: ITC traces of titration of host 28 (34.3 mM) into guest 37 (2.48 mM) in water at 303 K

28

NH

N

NH

Br

-40

-20

0

-10 0 10 20 30 40 50 60 70 80 90 100110120

Time (min)

µca

l/sec

0,0 0,5 1,0 1,5 2,0 2,5 3,0 3,5-4

-2

0

Data: V137A_NDHModel: OneSitesChi^2 = 430,496N 0.9257 ±0.001535K 6.915 E4 ±2689∆H -3238 ±8.554∆S 11,47

Molar Ratio

kcal

/mol

e of

inje

ctan

t

P

O O

O O

OHHO

HO OH

OHHO37

Na

Moreover, the corresponding MD simulation (fig. 4.3.1 I & II) data indicated that there were many

plausible modes of binding between the host 28 and the guest 37. In addition to the anticipated

hydrogen bonding between the guanidinium N–H’s and the phosphinate O-atoms, the aromatic π-π

interactions and guanidinium cation-π interactions were responsible for the stability of the complex

and prevented the likely complete dissociation. All these possible modes of interactions contribute to

the positive value of the association entropy.

The observed small binding enthalpy and the larger positive entropy of association suggest that the

enhanced affinity of the host-to-phosphinate results from the weakened structuring.

4.1.2 ITC titration of host 28 with guest 119 in Wa ter

The titration of the allyl guanidinium host 28 and the sodium diphenylphosphinate guest 119 in water

indicates the endothermic process, however no affinity for the guest was observed. Unlike the bay

phosphinate 37, the solvation shell of the guest 119 was not protected by the hydrophobic groups from

the potential solvation in water. As a result, massive solvation of the exposed phosphinate anion of

119 in water leads to the nil affinity for the allyl guanidinium host 28 (fig. 4.1.2).

50

Figure 4.1.2: ITC traces of titration of the host 28 (34.3 mM) into the guest 119 (5.2 mM) in water at

303 K

28

NH

N

NH

Br

0

1

2-10 0 10 20 30 40 50 60 70 80

Time (min)

µca

l/sec

0,0 0,5 1,0 1,5

-0,05

0,00

0,05

0,10

0,15

0,20

v138bl.itc

v138a.itc

Molar Ratio

kcal

/mol

e of

inje

ctan

t

PO O

Na

119

4.1.3 ITC titration of host 34 with guest 37 in Wat er

The titration of the host 34 into the guest 37 in water does not indicate any complex formation though

the heat pulses were exothermic. The observed negative results can be explained in many ways like

there must be a formation of micelle like structures which prevents the contact of host-guest ion pairs.

The host 34 is flanked by flexible and longer hydrophilic chains which must be involved in solvation

rather than preventing the solvation of the guanidinium core in water. Moreover, there is a probability

of non-complementarity of the host size and shape with the bay-like structure of the guest 37. The

heat pulses obtained must be due to the aggregation of host and/or guest during titration and not from

any specific complexation between the host 34 and the guest 37.

51

Figure 4.1.3: ITC traces of titration of host 34 (50.6 mM) into guest 37 (2.48 mM) in water at 303 K

P

O O

O O

OHHO

HO OH

OHHO37

Na

NH

N

NH

OH

OH

HO

HO

Br

34

-4

-2

0

2-10 0 10 20 30 40 50 60 70 80 90 100110120

Time (min)

µca

l/sec

-0,5 0,0 0,5 1,0 1,5 2,0 2,5 3,0 3,5 4,0 4,5 5,0

-0,22-0,20

-0,18

-0,16-0,14

-0,12

-0,10-0,08

-0,06-0,04

-0,02

0,000,02

v135bl.itc

v135a.itc

Molar Ratio

kcal

/mol

e of

inje

ctan

t

The trend analysis of the different host-guest ion pairs (28·37, 28·119 and 34·37) suggested that the

protection of the solvation shells around the binding motifs of the respective host as well as the guest

by constructing the adjacent hydrophobic walls was essential to produce the effective association

between the ion-pairs in highly polar solvent such as water.

4.2 ITC titrations in DMSO

4.2.1 ITC titration of host 28 with guest 37 in DMS O

Polar solvents are able to interact with host or guest molecules via electrostatic interactions. Such

solvents are particularly able to inhibit binding of charged species, as the solvent dipole can interact

strongly with a charged centre, thus making the solvent-host or solvent-guest interactions harder to

break. DMSO acts as both an electron-pair donor and hydrogen bond acceptor by virtue of oxygen

and sulphur lone pairs.[15] Because of these and other such unique properties, DMSO has been called

“water’s alter ego.”[127] The binding reaction between the allyl guanidinium host 28 and the bay

52

phosphinate 37 was strongly exothermic giving a good quality fit-curve using a one-site binding model

but with an unusual stoichiometry of 2:3 of the host 28 to the guest 37.

Figure 4.2.1: ITC traces of titration of allyl guanidinium host 28 (31.6 mM) into guest 37 (2.11 mM) in

DMSO at 303 K

28

NH

N

NH

Br

P

O O

O O

OHHO

HO OH

OHHO37

Na

-60

-40

-20

0

0 20 40 60 80 100 120

Time (min)

µca

l/sec

-0,5 0,0 0,5 1,0 1,5 2,0 2,5 3,0 3,5 4,0

-6

-4

-2

0

Data: V133A_NDHModel: OneSitesChi^2 = 549,620N 0.6607 ±0.001980K 6184 ±103.6∆H -7147 ±28.64∆S -6,218

Molar Ratio

kcal

/mol

e of

inje

ctan

t

The binding constant for the ion-pair was moderate (Ka ≈ 6184 M-1) in DMSO. The thermodynamic

data obtained from the ITC titration indicated the association was dominated by the favourable

enthalpy. It was clear from the ITC data that the entropy change is unfavourable for the complex

formation; therefore the binding was enthalpically driven. The strong negative enthalpy and the

unfavourable entropy of association point towards a lock-and-key binding relation. The view was also

supported by the MD simulation data of the respective host-guest ion-pair which suggested a stable

complex formation over about 7 ns simulation period (fig.4.3.2, blue trace).

53

4.2.2 ITC titration of host 34 with guest 37 in DMS O

The association of the tetra-hydroxypropyl guanidinium host 34 and the phosphinate guest 37 in

DMSO shows a strong exothermicity which is an indication of strong hydrogen bonding between the

ion-pair. The stoichiometry of the host to guest binding was found to be 1:2. The large negative

entropy (∆S° = -13.43 e.u.) represents the restriction on the movement of the complexed species

which could be interpreted as a better fit between the host and guest.

Figure 4.2.2: ITC traces of titration of host 34 (55.5 mM) into guest 37 (2.11 mM) in DMSO at 303 K

P

O O

O O

OHHO

HO OH

OHHO37

Na

NH

N

NH

OH

OH

HO

HO

Br

34

-100

-50

0

0 20 40 60 80 100 120

Data: V131A_NDHModel: OneSitesChi^2 = 1161,57N 0.5683 ±0.010K 1950 ±69.32∆H -8640 ±190.1∆S -13,43

Time (min)

µca

l/sec

0 1 2 3 4 5 6

-6

-4

-2

0

Molar Ratio

kcal

/mol

e of

inje

ctan

t

4.2.3 ITC titration of host 51 with guest 37 in DMS O

The tetra-(4-hydroxyphenyl) guanidinium host 51 was titrated into the solution of the aromatic

phosphinate guest 37 in anhydrous dimethyl sulfoxide at 303 K. The titration curve obtained by using a

one-site binding model was shown in fig. 4.2.3. The titration results indicate that the complexation was

exothermic though the curve was not an ideal sigmoidal one but the curve fits well using a one site

54

binding model. The stoichiometry of 1:4 was found to be too low (n = 0.25) which must be due to some

aggregation or the higher order complexation in the system.

Figure 4.2.3: ITC traces of titration of host 51 (19.5 mM) into guest 37 (2.11 mM) in DMSO at 303 K

P

O O

O O

OHHO

HO OH

OHHO37

Na

-15

-10

-5

0

0 20 40 60 80 100 120

Time (min)

µca

l/sec

0,0 0,5 1,0 1,5 2,0 2,5-2

0

Data: V129A_NDHModel: OneSitesChi^2 = 37,7968N 0.2521 ±0.02039K 542.1 ±16.99∆H -7969 ±710.2∆S -13,77

Molar Ratio

kcal

/mol

e of

inje

ctan

t

HO

HO

OH

OH

NH

N

NH

Br

51

4.2.4 ITC titration of host 51 with guest 119 in DM SO

The binding reaction of the diphenyl phosphinate guest 119 with the host 51 was exothermic and the

resulting complexation was entropy-driven. The stoichiometry of the host to the guest was found to be

1:2 with a modest association constant of 2953 M-1 derived by applying the one-site binding model (fig.

4.2.4.). The large positive entropy of association observed must be arising due to the desolvation of

the more exposed phosphinate anion 119.

55

Figure 4.2.4: ITC traces of titration of host 51 (19.5 mM) into guest 119 (2 mM) in DMSO at 303 K

PO O

Na

119

HO

HO

OH

OH

NH

N

NH

Br

51

-20

-10

0

0 10 20 30 40 50

Time (min)

µca

l/sec

0,0 0,5 1,0 1,5 2,0

-2

0

Data: V130A_NDHModel: OneSitesChi^2 = 339,192N 0.4828 ±0.01000K 2953 ±169.0∆H -3020 ±83.34∆S 5,922

Molar Ratio

kcal

/mol

e of

inje

ctan

t

A definite trend was observed from the titration data between the closely related but with increasing

steric bulk around the guanidinium cations 28, 34 and 51 and the phosphinate guests 37 and 119. The

increase in negative value of the change in association entropy going from the least congested host

28 to the most congested one 51 in association with the guest 37 was consistent with the increased

surface area exposed to the solvent molecules. The release of solvent molecules to the bulk is

proportional to the corresponding exposed surface area of the host or the guest compound.[128] All the

complexes with the guest 37 of the respective hosts were enthalpy-driven except for the ion-pair

51·119, where the complexation was entropically driven.

The trend could be explained as with the increase in steric hindrance around the binding motifs led to

the increase in secondary interactions like π-π stacking of aromatic residues and the dipole

interactions of the hydroxyl groups which results in the weak structuring of the complexes. The

association constant of the ion-pair 28·37 was about three-fold higher than the ion-pair 34·37 which in

turn, was about four times higher than the corresponding 51.37 ion-pair.

56

4.3 MD simulations

The MD simulations were performed using GROMOS’96[129] software and Pentium PC’s. The

GROMOS software was initially developed for proteins and widely used for small molecules having

peptide residues like arginine. Owing to the similarities between the acyclic guanidine in the side chain

of the amino acid arginine and the bicyclic guanidines it is easier to employ this software for the

present calculations. The GROMOS’96 program not only offers the force field 45a4 “united-atom” but

also a software package for analysis of the resulting data.

Simulations were carried out in different solvents and at 300 K (for the host 51 at 400 K as well) to

study their effect on the complexation as presented in table 4.3.1. The polar solvents like water,

DMSO, methanol and the non-polar solvent chloroform were used for simulations at 300 K.

Table 4.3.1: All simulations in this table were carried out with the hosts 28,

34, 51 and the guest 37

Host Solvent Temperature

[K]

Box size

[Å]

No. of the solvent

molecules

H2O 300 40.7 1053

DMSO 300 42.1 316

MeOH 300 41.8 553 28

CHCl3 300 42.3 275

H2O 300 42.3 1177

DMSO 300 42.2 316

MeOH 300 41.0 525 34

CHCl3 300 46.4 367

300 44.05 1373 H2O

400 44.05 1373

300 44.05 348 DMSO

400 44.05 348

300 43.2 616 MeOH

400 43.2 616

300 44.1 314

51

CHCl3 400 44.1 314

57

The MD simulations are successfully run by GROMOS’96 program using two different set of files– the

topology files and the coordinate files. The topology files for host and guest were constructed

according to the rules described in the GROMOS manual.[129] The partial charges for phosphorous

atom and bond parameters for the C–P bond in the guest molecule 37 were adapted from the

literature.[130] The combined coordinate file was prepared by first drawing the structures of the

respective host and guest molecule in the ChemSketch (ACD/Labs Release: 12.0) program (drawn

close enough but separate structures as the initial conformation of the plausible complex) and then the

combined structure was saved as 3D coordinate file (*.mol). Using the OpenBabel (version 2.2.1)

program the 3D coordinate file (*.mol) was converted into the GROMOS file (*.gr96). Then the

respective host and guest topology files were combined along with the 3D coordinates followed by the

energy minimization in vacuum using the GROMOS’96 program. The MD simulations can be run in

vacuum or in a solvent. The boundary conditions (box size and shape) and the solvent were decided

prior to the energy minimization.The truncated octahedron box was chosen for all the MD simulation

studies as it offers the spherical surrounding which bear a resemblance to the natural systems.

The energy minimized structures were placed into the pre-equilibrated box filled with the explicit

solvent molecules. The MD simulations were performed using different solvents like H2O, DMSO,

MeOH and CHCl3 at 300 K temperature (for the host 51 at 400 K as well) for a period of 10 ns as

shown in the table 4.3.1.

The results obtained from the MD simulations were analysed by using the provided software. The

resulting non-bonded interactions or energies and H-bond analysis from the MD simulations between

the tetra-allyl guanidinium host 28, tetra-hydroxypropyl guanidinium host 34, tetra-(4-hydroxyphenyl)

guanidinium host 51 and the aromatic tricyclic phosphinate guest 37 in the respective solvents i.e.

H2O, DMSO, MeOH and CHCl3 are discussed through the following sub-sections.

4.3.1 MD simulations in H 2O

Water is the most polar solvent used in the present study. As discussed throughout the previous

sections, the complexation studies involving artificial receptors and oxoanions in water is one of the

ambitions of a supramolecular chemist. Water is a unique solvent with a very high dielectric constant

(ε ≈ 80 at 20°C). It has a very high affinity for polar compounds and binds through hydrogen bonding.

Due to this feature of the water, polar host and guest molecules have to overcome the solvation by

water molecules to achieve effective association.

58

Figure 4.3.1: I. Time series of non-bonded energies between hosts 28, 34, 51 and guest 37 at 300 K

in H2O.

-450

-400

-350

-300

-250

-200

-150

-100

-50

0

50

0 1000 2000 3000 4000 5000 6000 7000 8000 9000 10000

Time [ps ]

Ene

rgy

[kJ/

mol

]

28

34

51

a

b

c

de

II. The conformations at the indicated intervals in the corresponding time series.

a b c

d e

59

The molecular dynamics or MD studies offer a first hand impression about the proposed molecules

and their association properties. These studies can also form the additional supporting data for the

experimental results.

In the present study the hydrophilic hosts 28, 34, 51 and the hydrophilic aromatic guest 37 were

synthesized and the corresponding ITC measurements were performed to establish the binding

properties of the host-guest ion pairs (cf. section 4.2).

The MD simulations run in water by using the GROMOS’96 program indicate that initially the allyl

guanidinium host 28 sits very well into the “bay” formed by the phenyl acetylene residues adjacent to

the phosphinate anion of the guest 37 (fig. 4.3.1 II a). After about 200 ps of the simulation time, the

host 28 dissociates shortly (fig. 4.3.1 II b) from the phosphinate bay and then again binds the guest

through the cation-π interactions (fig. 4.3.1 II c). It is clearly seen from these conformations (a, b and

c) that the allyl guanidinium host 28 binds to the phosphinate guest 37 in water mainly through the

hydrophobic interactions between the aromatic part of the guest and the guanidinium cation rather

than the corresponding H-bonding interactions between N–H···O–P bonds. The complexation between

the tetra-propylhydroxy guanidinium host 34 and the phosphinate guest 37 was stabilized by hydrogen

bonding between the O-atom from the phosphinate anion and the H-atoms from the propyl chains

instead of N–H···O–P bonds (fig. 4.3.1 II d). The guanidinium host was not dissociated completely

probably because of these secondary interactions only. The guanidinium motif was seen far-flung from

the phosphinate anion throughout the simulation time period. However, the tetra-phenyl guanidinium

host 51 formed a relatively stable complex with the phosphinate anion 37 in water and was stable over

the 10 ns simulation time (fig. 4.3.1 I, green trace and II. e). At a time only one oxygen atom from the

phosphinate 37 was participating in the H-bonding with the guanidinium N–H’s. Apparently the “bay”

formed by the tweezer-like phenyl acetylene arms of the phosphinate guest 37 was not wide enough

to accommodate the entire host 51, therefore one arm of the anion was seen to be interacting with the

two flanking phenyl groups from the same side of the host. The remaining two phenyl groups from the

other side of the host were interacting with the tricyclic aromatic part of the guest whereas the second

phenyl acetylene arm was seen out of the plane.

4.3.2 MD simulations in DMSO

Dimethyl sulphoxide is an aprotic and relatively less polar solvent (dielectric constant, ε ≈ 47 at 20°C)

than the water. DMSO forms very strong hydrogen bonds with H-bond donors such as guanidinium N–

H’s. Therefore, the polar host and the guest molecules will be massively solvated by the solvent.

60

Figure 4.3.2: I. Time series of non-bonded energies between hosts 28, 34, 51 and guest 37 at 300 K

in DMSO.

-450

-400

-350

-300

-250

-200

-150

-100

-50

0

50

0 1000 2000 3000 4000 5000 6000 7000 8000 9000 10000

Time [ps]

Ene

rgy

[kJ/

mol

]

28

34

51

a

b

c

d

II. The conformations at the indicated intervals in the corresponding time series.

61

The binding partners have to overcome the solvation barrier by breaking the solvent-host or solvent-

guest bonds.The MD simulations carried on the allyl guanidinium host 28 and the phosphinate guest

37 indicated the complex between the ion-pair was stable over 65% of the simulation time but after

about 7.5 ns it was completely dissociated. The partially dissociated structure of the complex 28·37 is

represented by fig. 4.3.2 II-d at 7555 ps. Initially, the host molecule 28 fits well into the bay of

phosphinate 37 (fig.4.3.2 II-c) and stabilized by hydrogen bonding (N–H···O–P) as well as cation-π

interactions between flanking aromatic residues from phenyl acetylene walls and guanidinium cation.

On the contrary, the host 34 does not fit into the bay of the phosphinate 37 rather the complete

dissociation was prevented by the secondary hydrogen bonding interactions between propyl -OH

groups and the phosphinate O-atoms (fig.4.3.2 II-a and b). The host 34 probably fails to approach the

guest’s well protected binding site due to the relatively longer alkyl chains and the substantial solvation

of the polar host compound by the solvent molecules. However, the guarded guanidinium motif in the

host 51 and the phosphinate guest 37 forms a stable complex in DMSO at 300 K as depicted in (fig.

4.3.2 I. green trace). The stability of the ion-pair 51·37 (structure III) in dimethy sulphoxide must be a

consequence of the protection of the binding motifs with hydrophobic groups.

III: A conformation of the complex between host 51 and guest 37 in DMSO.

4.3.3 MD simulations in MeOH

The MD simulations in another polar protic solvent methanol (dielectric constant, ε ≈ 33 at 20°C) were

performed using the GROMOS’96 program; the corresponding non-bonded energies plotted versus

time are depicted in the fig. 4.3.3. The ion-pair 28·37 was dissociated after around 5 ns of the

simulation time. The host 28 was not interacting with the phosphinate 37 probably due to the higher

solvation of both the partners in methanol. The simulation between the hydroxypropyl host 34 and the

phosphinate guest 37 indicated that the short-lived resulting complex was rather due to the secondary

62

H-bonds between propyl –OH and the phosphinate anion. The polar host 34 was massively bound by

the solvent molecules and stayed in the box throughout the simulation time whereas the more

hydrophobic phosphinate guest 37 was flying-out of the box. The aromatic host 51 formed a relatively

stable complex with 37, however the secondary interactions were seen to be dominating (fig.4.3.3 II).

The complex was not dissociating completely due to these secondary interactions only, otherwise,

there were no N–H···O–P interactions observed during entire simulation time to stabilize the complex.

Figure 4.3.3: I Time series of non-bonded energies between hosts 28, 34, 51 and guest 37 at 300 K

in MeOH

-450

-400

-350

-300

-250

-200

-150

-100

-50

0

50

0 1000 2000 3000 4000 5000 6000 7000 8000 9000 10000

Time [ps]

Ene

rgy

[kJ/

mol

]

28

34

51

II. A conformation of the complex between host 51 and guest 37 in MeOH.

63

4.3.4 MD simulations in CHCl 3

The non polar solvent chloroform (dielectric constant, ε ≈ 4.8 at 20°C) was also used for the simulation

of hydrophilic host-guest ion-pairs. The ensemble of hosts 28, 34, 51 and the phosphinic acid guest 37

were probed in chloroform for in-silico complexations using the GROMOS’96 program. All the

receptors indicated a stable comlex formation with the guest 37 (fig. 4.3.4 I). The host 28 was sitting

very well in the phosphinate bay and the ion-pair was stabilized by the strong hydrogen bonding (N–

H···O–P) interactions. Interestingly, the hydroxypropyl chains in the host 34 folded in such a way to

accommodate the entire host molecule in the “bay” of the phosphinate guest 37 (fig. 4.3.4 II). Two of

the propyl-hydroxy groups, one from each side of the guanidinium core, were involved in the hydrogen

bonding with the phosphinate anion in addition to the primary N–H···O–P bonds. The remaining two

propyl chains form intramolecular H-bond with one another. The aromatic host 51 also formed a stable

complex with the guest 37 in chloroform and was additionally stabilized by π-π interactions between

aromatic residues of the host and guest. The complexation of the ensemble of hosts with the

phosphinate guest 37 clearly indicated the lack of solvation of the binding partners in chloroform

results in straightforward binding between the corresponding hydrophilic ion-pairs.

The binding energies for the ion-pairs 28·37 and 34·37 were found to be higher and keep fluctuating (0

to -350 kJ mol-1) in solvents like water, DMSO and methanol, however it was much lower for the ion-

pair 51·37 (approx. -350 kJ mol-1). In chloroform, the binding energies were in the range of -350 to -

450 kJ mol-1 and remained constant for the corresponding ion-pair throughout the simulation period.

The observation was consistent with the concomitant drop in exothermicity due to the solvation of the

binding partners in more polar solvents.

64

Figure 4.3.4: I. Time series of non-bonded energies between hosts 28, 34, 51 and guest 37 at 300 K

in CHCl3

-600

-550

-500

-450

-400

-350

-300

-250

-200

-150

0 1000 2000 3000 4000 5000 6000 7000 8000 9000 10000

Time [ps]

Ene

rgy

[kJ/

mol

]

28

34

51

II. The conformation at 5000 ps of the complex between host 34 and guest 37.

65

Figure 4.3.5: Number of H-bonds participating in the association of the host 51 and the guest 37 at

300 K in H2O

0

1

2

3

4

5

6

7

8

0 1000 2000 3000 4000 5000 6000 7000 8000 9000 10000

Time [ps]

Num

ber

of H

-Bon

ds

Furthermore, the hydrogen bond analysis of the related MD simulations showed that the number of H-

bonds between the binding partners decrease with the increase in polarity of the solvent e.g. in CHCl3

six to seven H-bonds participated in the complex stabilization (fig.4.3.6) whereas in water ony three H-

bonds were involved (fig. 4.3.5). The H-bonding analysis includes the intermolecular as well as the

intramolecular H-bonds besides the guanidinium NH’s. The hydrogen bonding was unaffected by the

increase in temperature ca. from 300 K to 400 K.

Figure 4.3.6 : Conformation showing six H-bonds participating in 51·37 complex in CHCl3

66

5. Experimental Part

5.1 Reagents, Methods and Materials

All reactions were carried out in oven-dried glassware under an inert nitrogen / argon atmosphere. All

chemicals were purchased from Sigma-Aldrich, Fluka, ABCR, Strem Chemicals or Fluorochem and

used as received for the synthesis. Anhydrous solvents like dichloromethane, acetonitrile and dimethyl

formamide were purchased from Sigma-Aldrich stored on molecular sieves and used without further

purification. Tetrahydrofuran, toluene, 1,4-dioxane and ethanol were dried and purified as described

below.

Tetrahydrofuran (THF), Toluene and 1,4-dioxane

Technical grade solvents were first distilled under nitrogen atmosphere. Then sodium wire and

benzophenone (about 15 grams/litre as an indicator reagent) were added. The flask was stoppered

and kept in the dark overnight. A light blue-green to deep purple colour change during reflux indicates

the absence of water and free oxygen in the solvent. The solvent was refluxed under nitrogen for an

hour and the required quantity of the solvent was distilled off every time.

Absolute ethanol

6 g sodium metal per litre was dissolved in freshly distilled ethanol. To the solution 20 g of diethyl

phthalate was added and the ethanol was refluxed for 1 h under inert nitrogen atmosphere. Then the

ethanol was distilled over a vigreux column and stored on 4Å molecular sieves.

All other solvents were distilled before use (first 10-15% fractions were discarded). Triethyl amine and

EDIPA were stored on potassium hydroxide before distillation from calcium hydride.

Almost all reactions were monitored by RP-HPLC. HPLC instrumentation included Merck-Hitachi L-

6200A Intelligent Pump, Knauer UV-VIS Filter-Photometer and Sedex 55 LSD detector.

Chromatogram was recorded by Kipp and Zonen BD112 two channel recorder. Following are the

reverse phase HPLC columns used:

• Macherey-Nagel EC 250/4.6 Nucleodur C18 Pyramid, 3µ

• Macherey-Nagel EC 250/4 Nucleodur 100-5 C18 EC

• Macherey-Nagel EC 250/4 Nucleodur 100-5 CN

67

• Phenomenex Aqua 125/4 C18, 5µ

HPLC grade methanol and acetonitrile were purchased from VWR International, Germany. All eluents

were prepared by mixing methanol / acetonitrile and water with 0.1 % TFA as buffer.

Silica gel TLC and Aluminium oxide TLC-PET foils from Fluka were also used occasionally for reaction

progress monitoring; visualization was effected with UV and/or by developing in iodine. Uniplate Silica

gel GF Prep-TLC’s from Analtech Inc., USA and commercially available solid phase extraction (SPE)

cartridges from Alltech (High Capacity C18) were used for small scale purifications. Silica gel 100

(0.063-0.200 mm) from Merck was used for medium to large scale purifications.

Moreover, medium pressure reverse phase Flash-Chromatography was performed using C8 or CN-

modified silica gel in Michael-Miller columns connected to HPLC Pump 64, UV detector and a recorder

(all from Knauer).

Microwave reaction was carried out in a house hold microwave oven, Model MW 20 Digital W / MW 20

Digital S from TechnoStar.

Kugelrohr distillation was performed using a Büchi Glass oven B-585 connected to a high vacuum

pump.

NMR spectra were recorded on a Brucker AV-250 / 360 / 500 MHz spectrometer at 250.13 / 360.13 /

500.13 (1H), 62.89 / 90.55 (13C) MHz and 121 MHz (31P) at 298 K. Chemical shifts are reported in δ

[ppm] relative to the solvent residual peak CDCl3 / DMSO-d6 / MeOD / CD3CN as internal standards

(1H and 13C)[131] and H3PO4 as external standard for 31P. Data for 1H are reported as follows: chemical

shift, multiplicity (s = singlet, d = doublet, t = triplet, bs = broad signal, m = multiplet), coupling

constants (J) in Hz and integration.

Mass spectra were recorded on LCQ Classic Electrospray-Ionisation (ESI, HPLC-MS). High-resolution

mass spectra were recorded under ESI+ / HRMS using a MicroTOF-Q 77.

IR-Spectra were recorded on JASCO IR-4100 spectrometer (as direct substance or CCl4 solution).

Melting points were measured in open capillary tubes using Fisher-Jones apparatus and are

uncorrected.

Elemental analysis was done by in-house micro analytical laboratory of TU Munich.

68

Calorimetric titrations were performed on the Isothermal Titration Calorimeter MCS-ITC from Microcal

Inc., USA.

MD simulations were done by using GROMOS’96 software[129] and desktop PC’s.

69

5.2 Experimental Procedures

2,2,8,8-Tetrakis-(3-hydroxy-propyl)-1,3,4,6,7,9-hex ahydro-2 H-pyrimido[1,2- a]pyrimidinium

tetraphenylborate 34 [80, 81]

A 100 ml two-necked round-bottomed flask, equipped with a septum and a magnetic stirring bar was

charged with borane-THF solution (10 ml, 10.02 mmol, 1 M in THF) via a syringe under nitrogen

atmosphere. The flask was cooled to -10°C in an ice-salt bath. Then 2-methyl-2-butene solution (2.123

ml, 20.04 mmol in 5 ml of THF) was added to the flask at -10°C and the reaction mixture was stirred

for 1h maintaining the temperature between -10°C and -5°C. A solution of allyl guanidinium iodide

(500 mg, 1.67 mmol) in 7.5 ml of THF was added to the reaction mixture over a 40-minute interval

using a syringe pump. The resulting solution was stirred for 1h maintaining the temperature below

0°C. Stirring was continued for an additional 1h keeping the temperature below 10°C. After completion

of the reaction, it was quenched slowly with water (750 µl). Then a 4N NaOH solution (3 ml) was

added to the reaction mixture followed by the addition of 30% H2O2 solution (9 ml) below 0°C. Stirring

was continued further for 1h keeping the temperature between 0°C and room temperature. The

reaction mass was reduced to half by blowing down with nitrogen stream in a hood. Then an aqueous

solution of sodium tetraphenyl borate (3 mmol) was added to the reaction mixture and the content was

stirred further for about 2h at room temperature. The solid product was filtered and the mother liquor

was worked up further to collect the second crop of the product.

34: C19H38N3O4

+ −BPh4 (MW = 691.75 g/mol) Yield: 51% m.p.: 145°C

MS-ESI: m/z = 372.4 [(M) +, 100%] HRMS: Calculated = 372.2857, Observed = 372.2884

HPLC: Rt = 5 min. EC 250/4.6 Nucleodur C18 Pyramid, 3µ, UV254, Gradient from 30% to 90% in 10

minutes and subsequently from 90% to 90% in 5 minutes, CH3OH/H2O, 0.1% TFA

70

1H NMR (360 MHz, CD3CN) : δ [ppm] = 7.36 – 7.26 (m, 8H, ArH), 7.03 (t, J = 7.4 Hz, 8H, ArH), 6.88 (t,

J = 7.2 Hz, 4H, ArH), 6.05 (s, 2H, -NH), 3.60 – 3.42 (m, 8H, -CH2OH), 3.26 (t, J = 6.2 Hz, 4H, -NCH2),

2.82 (t, J = 4.9 Hz, 4H, -CH2OH), 1.80 (t, J = 6.2 Hz, 4H, -NCH2CH2), 1.64 – 1.41 (m, 16H, -

CH2CH2CH2OH).

13C NMR (91 MHz, CD3CN) : δ [ppm] = 164.30, (ArC), 149.33 (Guanidinium, C), 135.43 (d, J = 1.2 Hz,

ArCH), 125.27 (dd, J = 5.5, 2.7 Hz, ArCH), 121.47 (ArCH), 61.14 (-CH2OH), 53.53 (quart., C), 43.20 (-

NCH2CH2), 33.83 (-NCH2CH2), 28.66 (-CH2CH2CH2), 25.63 (-CH2CH2CH2).

13C NMR-DEPT (91 MHz, CD3CN): δ [ppm] = 135.43 (ArCH), 125.27 (ArCH), 121.47 (ArCH), 61.14 (-

CH2OH), 43.20 (-NCH2CH2), 33.83 (-NCH2CH2), 28.66 (-CH2CH2CH2), 25.63 (-CH2CH2CH2).

3',4',6',7',9',9'a-hexahydro-1'H-dispiro[cyclopenta ne-1,2'-[1,3]diazino[1,2-a]pyrimidine-8',1''-cyclopentane]-3,3''-dien-9'a-ylium iodide 68

An oven dried screw-cap vial equipped with a magnetic stirring bar, was charged with allyl

guanidinium iodide 28 (50 mg) and Hoveyda-Grubbs 2nd generation catalyst 67[97, 132, 133] (7.5 mg, 10

mol% by wt.). 2.5 ml of anhydrous methanol was injected to the reaction mixture and the resulting

solution was stirred at room temperature. After 3h, the reaction was complete although a small peak of

the mono-cyclized product was observed in the HPLC chromatogram. The solvent was evaporated by

a nitrogen stream and the residue was washed twice with ethyl acetate (5 ml) to remove the catalyst.

The solid residue was then purified by preparative HPLC (C8 column, 30 to 50% methanol in water as

an eluent) to afford the pure product.

68: C15H22IN3 (M = 371.26 g/mol) Yield: 65%

MS-ESI: m/z = 244.4 [M+, 100%]; 272.4 [(M) +, 60%, Monocyclized]

71

HPLC: Rt = 5.5 min. EC 250/4 Nucleodur 100-5 C18 EC, UV254, Gradient from 10% to 50% in 10

minutes and subsequently from 50% to 90% in 10 minutes, CH3OH/H2O, 0.1% TFA 1H NMR (250 MHz, CDCl3): δ [ppm] = 8.01 (s, 2H, -NH), 5.60 (s, 4H, -CH), 3.43 (t, J = 5.9 Hz, 4H, -

NCH2), 2.63 (d, J = 15.6 Hz, 4H, -CH2CH), 2.39 (d, J = 15.4 Hz, 4H, -CH2CH), 1.94 (t, J = 5.9 Hz, 4H, -

CH2CH2).

13C NMR (63 MHz, CDCl3): δ [ppm] = 149.82 (C, Guanidinium), 128.04 (-CH), 59.03 (C, quart.), 46.10

(-NCH2CH2), 44.83 (-NCH2), 31.72 (-CH2CH).

13C NMR-DEPT (63 MHz, CDCl3): δ [ppm] = 128.04 (-CH), 46.10 (-NCH2CH2), 44.83 (-NCH2), 31.72 (-

CH2CH).

4-(4-Chloro-phenyl)-2-oxo-but-3-enoic acid 77 [134-136]

9.025 g of 4-chlorobenzaldehyde 75 (64.2 mmol) was added to 37 ml of a 25% solution of potassium

hydroxide in methanol while stirring and to this mixture 5.8 g of pyruvic acid 76 (65.86 mmol) was

added rapidly to keep the aldehyde in molten condition. The yellow granular salt obtained was washed

with alcohol and then with ether and was dissolved in water cooled solution was acidified to yield the

unsaturated acid 77.

77: C10H7ClO3 (M = 210.61 g/mol) Yield: 80% m.p.: 130°C

MS-ESI: m/z = 209.1 [(M-H)-, 100%], 165.1 [(M-H-CO2)-, 65%]; 441.1 [(2M-2H+Na)-, 10%]

HPLC: Rt = 14 min. EC 250/4 Nucleodur 100-5 C8 EC, UV254, Gradient from 10% to 50% in 10

minutes and subsequently from 50% to 90% in 10 minutes, CH3OH/H2O, 0.1% TFA

72

3,4-Dibromo-4-(4-chloro-phenyl)-2-oxo-butyric acid 78[134]

The dried acid 77 was suspended in anhydrous chloroform and bromine was added to the well cooled

mixture. After one equivalent of bromine was added and the red-brown mixture was shaken until the

solution becomes clear and nearly colourless. The excess of bromine and the solvent were removed

under the nitrogen stream. The crystalline dibromide 78 was washed with a few millilitres of warm

benzene and crystallized from a small volume of boiling benzene.

78: C10H7Br2ClO3 (M = 370.42 g/mol) Yield: 80% m.p.: 135°C

MS-ESI: m/z = 598.8 [(2M+Na)-, 90%] corresponding lactone

HPLC: Rt = 17 min. EC 250/4 Nucleodur 100-5 C8 EC, UV254, Gradient from 10% to 50% in 10

minutes and subsequently from 50% to 90% in 10 minutes, CH3OH/H2O, 0.1% TFA.

4-Bromo-5-(4-chloro-phenyl)-3-hydroxy-5 H-furan-2-one 79 [134-137]

Cl

O Br

OHO

79

A 100 ml two-necked round-bottomed flask equipped with a magnetic stirring bar and septum was

charged with the dibromide 78 (1 g, 2.7 mmol) and distilled water (30 ml). The reaction mixture was

heated to 65-70°C in an oil bath while stirring the suspension rapidly. After stirring the reaction mass

for 30-40 minutes at 65-70°C, it was cooled slowly and left overnight to increase the conversion of the

less stable keto-acid 78 to the lactone. It was transferred to ether solution and the mixture was

73

extracted with pH 7.2 buffer (0.1 M KH2PO4-Na2HPO4) to remove the keto-acid impurity. The ether

solution was dried over MgSO4, evaporated and dried azeotropically with toluene on a rotating vacuum

evaporator. The crude product was then re-crystallized from benzene-petroleum ether to obtain the

colourless crystals.

79: C10H6BrClO3 (M = 289.51 g/mol) Yield: 65% m.p.: 140°C

MS-ESI: m/z = 286.9 [(M-H)-, 50%], 598.8 [(2M+Na)-, 100%]

HPLC: Rt = 16.5 min. EC 250/4 Nucleodur 100-5 C18 EC, UV254, Gradient from 10% to 50% in 10

minutes and subsequently from 50% to 90% in 10 minutes, CH3OH/H2O, 0.1% TFA.

1H NMR (360 MHz, CDCl3): δ [ppm] = 7.42, 7.39 (d, 2H, ArH), 7.25, 7.23 (d, 2H, ArH), 5.79 (s, 1H,

bridge-CH)

13C NMR (90.55 MHz, CDCl3): δ [ppm] = 167.02 (-C=O), 140.75 (-COH), 136.26 (-ArC), 131.51

(ArCCl), 129.36, 128.93 (ArCH), 112.39 (-C-Br), 82.03 (bridge –CH)

13C NMR-DEPT (90.55 MHz, CDC13): δ [ppm] = 129.35, 128.93 (ArCH), 82.03 (bridge –CH)

1,1-Dioxo-tetrahydro-1 λλλλ6-thiopyran-4-one 84 [107]

To the tetrahydrothiopyran-4-one 72 (0.4 g, 3.4 mmol) solution in acetonitrile (4.5 ml) at room

temperature was added an aqueous Na2·EDTA solution (3 ml, 4x10-4 M). To this mixture was added in

portions a mixture of Oxone® 83 (6.3 gm 10.3 mmol) and sodium bicarbonate (2.7 g, 32 mmol) over

20-minute period and then the reaction mass was left stirring overnight. After overnight stirring at room

temperature, the mixture was diluted with 80 ml dichloromethane, dried over anhydrous MgSO4 and

74

filtered through a pad of celite. The filtrate was concentrated to dryness under reduced pressure to

yield the product 84 as a white solid.

84: C5H8O3S (M = 148.18 g/mol) Yield: 75% m.p.: 170°C

HPLC: Rt = 4 min. (only LSD-peak) EC 250/4 Nucleodur 100-5 C18 EC, UV220, Gradient from 10% to

50% in 10 minutes and subsequently from 50% to 90% in 10 minutes, CH3OH/H2O, 0.1% TFA.

1H NMR (360 MHz, CDCl3): δ [ppm] = 3.38 (t, J= 10.2 Hz, 4H, -CH2S), 2.98 (t, J= 10.2 Hz, 4H, -CH2)

13C NMR (90.55 MHz, CDCl3): δ [ppm] = 202.02 (-C=O), 49.59 (-SCH2), 38.17(-CH2)

13C NMR-DEPT (90.55 MHz, CDCl3): δ [ppm] = 49.59 (-SCH2), 38.17 (-CH2)

The NMR data and melting point was found to be in accordance with the literature.[107]

2-(Dimethyl-hydrazono)-propionic acid 85 [106]

N

OH

O

NMe2

85

A solution of pyruvic acid 76 (1 g, 11.36 mmol) in anhydrous diethyl ether (5 ml) was taken in a 25 ml

round-bottomed flask equipped with a stirring bar and a rubber septum. 1,1-dimethylhydrazine (860 µl,

11.36 mmol) was injected to the reaction mass slowly at room temperature over 10-minute interval.

Gradually solid was started to precipitate in the reaction flask. The reaction mass was stirred for 2h at

room temperature. Then the solid was filtered and washed with diethyl ether and re-crystallized from

n-hexane: chloroform (1:1).

85: C5H10N2O2 (M = 130.15 g/mol) Yield: 62%

MS-ESI: m/z = 131.0 [(M+H)+, 100%], 155.0 [(M+2H+Na)+, 85%]

75

HPLC: Rt = 5 min. EC 250/4 Nucleodur 100-5 C18 EC, UV220, Gradient from 10% to 50% in 10

minutes and subsequently from 50% to 90% in 10 minutes, CH3OH/H2O, 0.1% TFA.

1H-NMR (250 MHz; CDCl 3): δ [ppm] = 2.99 (s, 6H, -NCH3), 2.14 (s, 3H, -CH3)

3-Hydroxy-8, 8-dioxo-1-oxa-8 λλλλ6-thia-spiro [4.5] dec-3-en-2-one 74 [106]

S

O O

O

O OH

74

A dry Schlenk tube was charged with compound 85 (500 mg), evacuated for 5 minutes and then filled

with dry nitrogen. Absolute tetrahydrofuran (10 ml) was injected to the reaction flask via a syringe

followed by freshly distilled HMPA (3.4 ml, 5 eq.). The Schlenk tube was cooled to -20°C in a dry ice-

acetone bath. Methyl lithium (7.2 ml, 3eq.) was slowly injected to the reaction mixture via a syringe at -

20°C. After 15 minutes of stirring at -20°C, compound 84 (626 mg, 1.1eq.) was added quickly and re-

placed the septum. The temperature of the reaction flask was allowed to rise to room temperature by

removing the cooling bath. The reaction mixture was stirred for overnight at room temperature. It was

quenched by pouring on ice slowly. The aqueous solution was acidified with hydrochloric acid (pH~2-

3) and stirring continued further for 8h at room temperature. The reaction mixture was extracted with

ethyl acetate (3 x 20 ml) and the combined organic fractions were washed with water (1 x 10 ml). The

organic fraction was dried over MgSO4 and the solvent was evaporated in vacuo. The thick yellow oil

obtained was dissolved in anhydrous diethyl ether, and the insoluble white product (desired lactone

74) was filtered.

74: C8H10O5S (M = 218.23 g/mol) Yield: 65% m.p.: 132°C

MS-ESI: m/z = 217.1 [(M-, 45%], 435.1 [(2M+H)-, 30%]

HPLC: Rt = 5.5 min. EC 250/4 Nucleodur 100-5 C18 EC, UV220, Gradient from 10% to 50% in 10

minutes and subsequently from 50% to 90% in 10 minutes, CH3OH/H2O, 0.1% TFA.

1H NMR (250 MHz, CD3CN): δ [ppm] = 6.34 (s, 1H, -CH), 3.32 (t, 2H, -SCH2), 3.04 (d, 2H, -SCH2),

2.46 (t, 2H, -CH2), 2.08 (d, 2H, -CH2).

76

13C NMR (62.89 MHz; CD3CN): δ [ppm] = 168.75 (-C=O), 143.91 (-COH), 122.49 (-CH), 80.35 (spiro,

C), 48.28 (-SCH2), 36.99 (-CH2)

13C NMR-DEPT (62.89 MHz; CD3CN): δ [ppm] = 122.49 (-CH), 48.28 (-SCH2), 36.99 (-CH2)

3-Methoxy-8,8-dioxo-1-oxa-8 λλλλ6-thia-spiro [4.5] dec-3-en-2-one 89

S

O O

O

O OMe

89

Compound 74 (100 mg, 0.46 mmol) was suspended in 10 ml anhydrous dichloromethane. The

reaction mixture was cooled to 0°C using an ice-salt bath. A solution of diazomethane (5.4 ml, 0.92

mmol) in ether (0.17M) was added to the cooled reaction mass slowly. The reaction was completed in

about one hour. Dichloromethane was then evaporated in a hood under nitrogen stream to afford the

desired product.

89: C9H12O5S (M = 232.25 g/mol) Yield: 70% m.p.: 275°C

HPLC: Rt = 8.5 min. EC 250/4 Nucleodur 100-5 C18 EC, UV220, Gradient from 10% to 50% in 10

minutes and subsequently from 50% to 90% in 10 minutes, CH3OH/H2O, 0.1% TFA.

1H NMR (360 MHz, DMSO-d6): δ [ppm] = 6.88 (s, 1H, -CH), 3.71 (s, 3H, -OCH3), 3.24 (t, 4H, -SCH2),

2.35 (dd, 2H, -CH2), 2.18 (dd, 2H, -CH2).

13C NMR (360 MHz, DMSO-d6): δ [ppm] = 165.66 (-C=O), 145.85 (-COCH3), 121.40 (-CH), 80.03

(spiro, C), 57.98 (-OCH3), 47.09 (-SCH2), 33.69 (-CH2)

13C NMR-DEPT (360 MHz, DMSO-d6): δ [ppm] = 121.40 (-CH), 80.03 (spiro, C), 57.98 (-OCH3), 47.09

(-SCH2), 33.69 (-CH2)

77

4-Bromo-3-hydroxy-8, 8-dioxo-1-oxa-8 λλλλ6-thia-spiro [4.5] dec-3-en-2-one 87

S

O O

O

O OH

Br

87

A solution of sulphone-lactone 74 (218 mg, 1 mmol) in 10 ml of anhydrous acetonitrile, was slightly

warmed with a heat gun to dissolve the compound completely. A solution of bromine (100 µl, 2 mmol)

in 1 ml of acetonitrile was added to the reaction flask under nitrogen atmosphere. After 20 minutes

stirring at room temperature, the solid was separated from the reaction mass. The reaction mixture

was stirred further for additional 30 minutes at room temperature. The solvent was evaporated in

vacuo and the residue was washed with ether to yield the desired product 87.

87: C8H9BrO5S (M = 297.12 g/mol) Yield: 80% m.p.: 270°C

MS-ESI: m/z = 295.0 [(M-H)-, 20%], 614.9 [(M-H+Na)-, 100%]

HPLC: Rt = 12 min. EC 250/4 Nucleodur 100-5 C18 EC, UV254, Gradient from 10% to 50% in 10

minutes and subsequently from 50% to 90% in 10 minutes, CH3OH/H2O, 0.1% TFA.

1H NMR (500 MHz, CD3CN): δ [ppm] = 3.37 (t, 2H, -SCH2), 3.05 (d, 2H, -SCH2), 2.61 (t, 2H, -CH2),

2.04 (d, 2H, -CH2).

4-Bromo-3-methoxy-8, 8-dioxo-1-oxa-8 λλλλ6-thia-spiro [4.5] dec-3-en-2-one 88

S

O O

O

O OMe

Br

88

78

To the solution of bromo-lactone 87 (30 mg, 0.1 mmol) in 5 ml anhydrous dichloromethane, a solution

of diazomethane (1.2 ml, 0.2 mmol, 0.17M in ether) was added over a 10-minute period at 0°C under

nitrogen atmosphere. Then the reaction mixture was allowed to warm to room temperature. After

completion of the reaction, the solvent was evaporated under nitrogen jet in a hood to obtain a white

solid product.

88: C9H11BrO5S (M = 311.15 g/mol) Yield: 70% m.p.: 205°C

HPLC: Rt = 15 min. EC 250/4 Nucleodur 100-5 C18 EC, UV254, Gradient from 10% to 50% in 10

minutes and subsequently from 50% to 90% in 10 minutes, CH3OH/H2O, 0.1% TFA

1H NMR (500 MHz; CDCl 3): δ [ppm] = 4.13 (s 3H, -OCH3), 3.46 (t, 2H, -SCH2), 3.07 (d, 2H, -SCH2),

2.87 (t, 2H, -CH2), 1.95 (d, 2H, -CH2).

13C NMR (62.89 MHz, CDCl3): δ [ppm] = 163.56 (-C=O), 142.43 (-COCH3), 120.38 (-CBr), 80.55

(spiro, C), 58.83 (-OCH3), 46.82 (-SCH2), 33.27 (-CH2)

13C NMR-DEPT (62.89 MHz, CDCl3): δ [ppm] = 58.83(-OCH3),46.82 (-SCH2), 33.27 (-CH2)

4-Bromo-3-( tert-butyl-diphenyl-silanyloxy)-8, 8-dioxo-1-oxa-8 λλλλ6-thia-spiro [4.5] dec-3-en-2-one 90

S

O O

O

O

Br

O Si

90

A 25 ml round-bottomed flask was charged with sulphone lactone 87 (100 mg, 0.3367 mmol) and

DMAP (4 mg, 10 mol %). The reaction mass was kept under nitrogen atmosphere. Anhydrous

dichloromethane (7 ml) and triethyl amine (120 µl, 1.01 mmol) were added to the reaction mass via a

syringe at room temperature. A solution of tert.-butyldiphenylsilyl chloride (140 mg, 0.505 mmol) in 5

ml dichloromethane was added to the reaction mixture. The reaction was monitored by HPLC. After

completion of the reaction, it was diluted with 50 ml dichloromethane. The reaction mixture was

79

extracted with brine (1 x 10 ml), dried over MgSO4 and the solvent was evaporated on a rotatory

evaporator. The crude product obtained was purified by silica gel chromatography.

90: C24H27BrO5SSi (M = 535.52 g/mol) Yield: 70% m.p.: 122°C

HPLC: Rt = 27 min. EC 250/4 Nucleodur 100-5 C18 EC, UV254, Gradient from 10% to 50% in 10

minutes and subsequently from 50% to 90% in 10 minutes and 90% to 90% in 5 minutes CH3OH/H2O,

0.1% TFA

Bis-(trimethylsilyl)phosphonite (BTSP) 81 [103, 105]

TMSOP

TMSO

H

81

To 8.3 g water free ammonium hypophosphite (0.1 mol) was added 22.9 ml of HMDS (0.1 mol). The

reaction mixture was heated to 100°C in an oil bath for 8h till the ammonia gas evolution ceased. Then

the resulting solution was vacuum distilled (11 mbar) at 51 to 54°C to yield the pure product. After

distillation the product was stored very carefully under argon balloon since it was an extremely

pyrophoric reagent and could also explode upon contact with a trace of air.

81: C6H19O2PSi2 (M = 210.36 g/mol) Yield: >95% b.p.: 51-54°C at 11 mbar

Bis-(4-methoxy-phenyl)-acetic acid 55 [91]

Glacial acetic acid (55 ml), glyoxalic acid 53 (5 g) and anisole 54 (12.35 g) were placed in a 250 ml

round-bottomed flask fitted with a septum and a magnetic stirring bar. The mixture was cooled to 0°C

in an ice-salt bath while gently stirring. Conc. H2SO4 (2 ml) was added drop-wise to the reaction

80

mixture at 0°C. The reaction mixture was allowed to warm to room temperature and then heated at

50°C in an oil bath for overnight. After completion of the reaction, it was poured into ice and extracted

with dichloromethane (3 x 100 ml). The combined dichloromethane fractions were washed with water

(1 x 100 ml) followed by extraction with aq. 4N NaOH solution (3 x 100 ml). Aqueous basic fraction

was then acidified with hydrochloric acid slowly to pH~1-2 and was extracted with dichloromethane (3

x 50 ml), washed with water (1 x 30 ml), dried over MgSO4 and the solvent was evaporated in vacuo to

afford the product.

55: C16H16O4 (M = 272.29 g/mol) Yield: 80% m.p.: 105°C

HPLC: Rt = 11 min. EC 250/4 Nucleodur 100-5 C18 EC, UV254, Gradient from 10% to 50% in 10

minutes and subsequently from 50% to 90% in 10 minutes, CH3OH/H2O, 0.1% TFA

1H NMR (500 MHz, CDCl3): δ [ppm] = 7.27 (d, J = 8.6 Hz, 4H, ArH), 6.89 (d, J = 8.7 Hz, 4H, ArH),

4.98 (s, 1H, -CH), 3.81 (s, 6H, -CH3).

13C NMR (63 MHz; CDCl 3): δ [ppm] = 55.06 (-OCH3), 55.25(-CH), 158.64, 130.46, 129.52, 113.88 (-

ArC), 178.37 (-COOH)

4-[[3-Carboxy-3,3-bis-(4-methoxy-phenyl)-propyl]-(t oluene-4-sulfonyl)-amino]-2,2-bis-(4-methoxy-phenyl)-butyric acid 57

An oven dried 250 ml-round-bottomed flask equipped with a nitrogen bubbler, a septum and a

magnetic stirring bar, was charged with absolute tetrahydrofuran (50 ml). The reaction flask was

cooled to -10°C in an ice-salt bath. To the cold THF 5.7 ml of diisopropylamine (freshly distilled and

dried over molecular sieves) was injected, followed by the addition of butyl lithium (1.6 M solution in n-

hexane, 33 ml). After stirring for 30 minutes at -10°C, a solution of monoacid 55 (1 g, in 15 ml absolute

THF dried over molecular sieves) was added to the reaction mixture over a 10-minute interval. The

81

reaction mass was stirred further for 15 minutes at -10°C. Then a solution of bisiodo-tosyl compound

56 (4.4 g in 15 ml absolute THF, dried over molecular sieves) was added to it over a 10-minute period.

The reaction mixture was stirred for 1h at -10°C and then allowed to warm to room temperature. The

reaction mixture was stirred at room temperature till completion of the reaction (110 h). The solvent

was evaporated in vacuo after completion of the reaction. The resulting solid was washed with n-

hexane, sonicated for 10 minutes to crush the lumps and then filtered under suction. The powder was

dried and dissolved in distilled water (100 ml, pH≈10 of the solution). It was extracted with diethyl ether

(3 x 50 ml) to remove the impurities. Then the aqueous basic solution was acidified with 6N HCl

solution to pH~1-2 and extracted again with diethyl ether (3 x 50 ml). The combined ether fractions

were washed with water (1 x 50 ml), brine (1 x 50 ml), dried over MgSO4 and the solvent was

evaporated to yield the crude product. Subsequently the crude was purified by silica gel column

chromatography and eluted with 20-40% ethyl acetate in petroleum ether to obtain the pure bisacid

57.

57: C43H45NO10S (M = 767.88 g/mol) Yield: 72% m.p.: 170°C

MS-ESI: m/z = 768 [(M+H)+, 92%]; 790.2 [(M+Na)+, 100%]

HRMS: Calculated = 767.2764, Observed = 767.2679

HPLC: Rt = 17 min. EC 250/4 Nucleodur 100-5 C18 EC, UV254, Gradient from 10% to 50% in 10

minutes and subsequently from 50% to 90% in 10 minutes, CH3OH/H2O, 0.1% TFA

1H NMR (360 MHz, DMSO): δ [ppm] = 7.23 (dd, J = 22.0, 7.9 Hz, 4H, ArH), 7.08 (d, J = 8.6 Hz, 8H,

ArH), 6.82 (s, 8H, ArH), 3.72 (s, 12H, -OCH3), 2.34 (s, 8H, -CH2), 2.08 (s, 3H, -CH3).

13C NMR (91 MHz, DMSO): δ [ppm] = 176.25 (-C=O), 157.45, 142.92, 129.54, 126.67, 112.91, (ArC),

54.98 (-OCH3), 52.28 (quart., C), 45.68 (-NCH2), 37.24 (-CH2), 20.93 (-CH3).

13C NMR-DEPT (91 MHz, DMSO): δ [ppm] = 129.55, 126.68, 112.91 (ArCH), 54.98 (-OCH3), 45.68 (-

NCH2), 37.24 (-CH2), 20.93 (-CH3).

82

4-[[3-Azidocarbonyl-3,3-bis-(4-methoxy-phenyl)-prop yl]-(toluene-4-sulfonyl)-amino]-2,2-bis-(4-methoxy-phenyl)-butyryl azide 59 [92]

To the solution of bisacid 57 (11.06 g, 14.4 mmol) in anhydrous dichloromethane (25 ml) was added

triethyl amine (6 ml, 43.2 mmol) slowly at 0°C. The reaction mass was stirred for 15 minutes at 0°C.

Diphenylphosphoryl azide (DPPA) (6.5 ml, 30.2 mmol) was added to the reaction mass via a syringe

at 0°C. The reaction mixture was allowed to warm to room temperature over 1h. The reaction was

completed in about three hours. It was diluted with dichloromethane (50 ml) and washed with 5 % aq.

NaHCO3 solution (2 x 50 ml) followed by the extractions with 10% methanol in water (4 x 50 ml), brine

(1 x 50 ml), dried over MgSO4 and then the solvent was evaporated in vacuo to yield the crude

product. The crude product was used as such for further modifications.

59: C43H43N7O8S (M = 817.91 g/mol)

HPLC: Rt = 19 min. (mono-azide), 22.5 min. (Bis-azide) EC 250/4 Nucleodur 100-5 C18 EC, UV254,

Gradient from 10% to 50% in 10 minutes and subsequently from 50% to 90% in 10 minutes,

CH3OH/H2O, 0.1% TFA

N,N-Bis-[3-isocyanato-3,3-bis-(4-methoxy-phenyl)-propy l]-4-methyl-benzenesulfonamide 60 [93,

94]

83

The crude bisacid azide 59 was dissolved in absolute toluene (20 ml) and the solution was irradiated

with microwave (power: 30%) for 7 minutes at one-minute intervals (at each interval nitrogen formed

was evacuated using a syringe). The HPLC chromatogram after 7 minutes irradiation shows complete

conversion of bisazide 59 to the corresponding bis-isocyanate 60. The solvent was evaporated in

vacuo to yield the crude product. The crude product obtained was used for subsequent reaction

without further purification

60: C43H43N3O8S (M = 761.88 g/mol) IR: 2250 cm-1 characteristic band for –C=N=O bond

HPLC: Rt = 24 min. EC 250/4 Nucleodur 100-5 C18 EC, UV254, Gradient from 10% to 50% in 10

minutes and subsequently from 50% to 90% in 10 minutes, CH3OH/H2O, 0.1% TFA

N,N-Bis-[3-amino-3,3-bis-(4-methoxy-phenyl)-propyl]-4- methyl-benzenesulfonamide 61

The crude bisisocyanate 60 was dissolved in tetrahydrofuran (5 ml). Conc. HCl (1 ml) was added to

the solution slowly and the reaction mass was then heated to reflux in an oil bath. The reaction

progress was followed by HPLC. After completion of the reaction, the solvent was removed on a

rotatory evaporator to dryness. The crude bisamine salt 61 obtained was purified by re-crystallization

from acetonitrile.

61: C41H49Cl2N3O6S (M = 782.81 g/mol) Yield: 45% (over three steps) m.p.: 195°C

MS-ESI: m/z = 709.9 [(M+H) +, 55%], 1419.0 [(2M+H) +, 35%]

84

HPLC: Rt = 13 min. EC 250/4 Nucleodur 100-5 C18 EC, UV254, Gradient from 10% to 50% in 10

minutes and subsequently from 50% to 90% in 10 minutes, CH3OH/H2O, 0.1% TFA

1H NMR (250 MHz, CDCl3): δ [ppm] = 7.56 (dd, 2H, -NH), 7.25 (d, 4H, ArH), 7.13 (dd, 2H, -NH), 6.92

(m, 8H, ArH), 6.80 (m, 8H, ArH), 3.78 (s, 12H, -OCH3), 2.35 (s, 8H, -CH2), 2.39 (s, 3H, -CH3).

N3-[3-Amino-3,3-bis-(4-methoxy-phenyl)-propyl]-1,1-bi s-(4-methoxy-phenyl)-propane-1,3-diamine 62 [95]

The suspension of bisamine salt 61 (900 mg) in 10 ml of tetrahydrofuran was added to approx. 10 ml

of liq. NH3 at -78°C. Sodium-metal pieces (~500 mg) were added to the reaction mixture (solution

became deep purple in colour). The reaction mixture was stirred further for additional 15 minutes and

then quenched with solid NH4Cl till it became colourless. Liq. NH3 was then evaporated under nitrogen

stream and the reaction mass was diluted with distilled water (pH>10). It was extracted with ethyl

acetate (3 x 25 ml), the combined ethyl acetate fraction was washed with water, dried over MgSO4 and

the solvent was evaporated in vacuo to yield the crude trisamine free base 62 as a pale yellow liquid.

62: C34H41N3O4 (M = 555.71 g/mol) Yield: 78%

HPLC: Rt = 9 min. EC 250/4 Nucleodur 100-5 C18 EC, UV254, Gradient from 10% to 50% in 10

minutes and subsequently from 50% to 90% in 10 minutes, CH3OH/H2O, 0.1% TFA

1H NMR (360 MHz, DMSO): δ [ppm] = 7.21 (dd, J = 6.9, 5.0 Hz, 8H, ArH), 6.79 (d, J = 8.9 Hz, 8H,

ArH), 3.69 (s, 12H, -OCH3), 2.30 (d, J = 7.5 Hz, 4H, -NCH2), 2.19 (d, J = 7.5 Hz, 4H, -CH2).

13C NMR (91 MHz, DMSO): δ [ppm] = 157.19, 141.74, 127.33, 113.03 (ArC), 59.24 (quart. C), 54.97 (-

OCH3), 45.50 (-NCH2), 41.41 (-CH2).

85

13C NMR-DEPT (91 MHz, DMSO): δ [ppm] = 127, 113.03 (ArC), 54.97 (-OCH3), 45.50 (-NCH2),

41.41 (-CH2).

2,2,8,8-Tetrakis-(4-methoxy-phenyl)-1,3,4,6,7,9-hex ahydro-2 H-pyrimido[1,2- a]pyrimidinium iodide 65

A 25 ml single neck round-bottomed flask equipped with a septum and a magnetic stirring bar, was

charged with trisamine 62 (190 mg, 0.34 mmol) in 5 ml anhydrous acetonitrile.

Thiocarbonyldiimidazole (73 mg, 0.41 mmol) was added to the reaction solution at 15-20°C and the

stirring was continued for 30 minutes. After 30 minutes, the HPLC chromatogram indicated the

conversion of starting material to an intermediate. The reaction mass was stirred further for additional

30 minutes. Glacial acetic acid (50 µl, 0.85 mmol) was added to the reaction mixture and stirring was

continued further for 30 minutes followed by the addition of methyl iodide (65 µl, 1.03 mmol). The

reaction was stirred for 1 hour and then EDIPA (175 µl, 1.03 mmol) was added to the mixture. The

reaction mass was refluxed for about 5h till the complete conversion of the intermediate to the desired

product. It was cooled to room temperature and the solvent was evaporated. The residue was diluted

with dichloromethane (10 ml), washed with water, brine and dried the organic phase was dried over

MgSO4. The mixture was filtered and the solvent was evaporated to afford the crude product. The

crude product was re-crystallized from acetonitrile.

65: C35H38N3O4+ (M = 564.69 g/mol) Yield: 70% m.p.: 270°C

MS-ESI: m/z = 564.3 [M+, 100%]; 565.3 [(M+H)+, 38%]

HPLC: Rt = 21.5 min. EC 250/4 Nucleodur 100-5 C18 EC, UV254, Gradient from 10% to 50% in 10

minutes and subsequently from 50% to 90% in 10 minutes, CH3CN/H2O, 0.1% TFA

86

1H NMR (250 MHz, CDCl3): δ [ppm] = 9.19 (s, 2H, -NH), 7.42 – 7.31 (m, 8H, ArH), 6.93 – 6.84 (m, 8H,

ArH), 3.77 (s, 12H, -OCH3), 3.13 (t, J = 6.0 Hz, 4H, -NCH2), 2.62 (t, J = 5.9 Hz, 4H, -CH2).

13C NMR (63 MHz, CDCl3): δ [ppm] = 159.03 (ArC), 150.87(Guanidinium, C), 134.96, 127.63, 114.25

(ArC), 60.86 (quart. C), 55.29 (-OCH3), 44.73 (-NCH2), 32.62 (-CH2).

13C NMR-DEPT (63 MHz, CDCl3): δ [ppm] = 127.63, 114.25 (ArCH), 55.30(-OCH3), 44.73 (-NCH2),

32.62 (-CH2).

2,2,8,8-Tetrakis-(4-hydroxy-phenyl)-1,3,4,6,7,9-hex ahydro-2 H-pyrimido[1,2- a]pyrimidinium

bromide 51

An oven dried screw-cap vial equipped with a triangular magnetic stirring bar, was charged with tetra-

methoxy guanidinium compound 65 (50 mg). It was filled with argon and evacuated twice. Anhydrous

dichloromethane (2 ml) was injected to the vial and the mixture was stirred at room temperature till the

solid dissolved completely to became a clear solution. The vial was cooled to -78°C in a dry ice-

acetone bath. 1M solution of BBr3 in dichloromethane (1.1 ml) was injected to the reaction mixture

over a 5-minute period. It was stirred at -78°C for 30 minutes and the cooling bath was taken out

allowing the reaction vial to warm to room temperature. HPLC analysis of the reaction mixture after 2

hours at room temperature was shown complete conversion of the starting material. The reaction was

quenched by injecting 1 ml methanol: water (1:1) into the vial at 0°C. The dichloromethane phase was

separated from the aqueous methanolic phase containing the desired product using a syringe. The

solvent was evaporated in vacuo to yield the crude product which was then re-crystallized from

acetonitrile.

51: C31H30N3O4+ (M = 508.22 g/mol) Yield: 80% m.p.: 210°C

MS-ESI: m/z = 508.1 [M+, 100%]; 1097.0 [(2M+HBr)+, 22%]

87

HRMS: Calculated = 508.2231 Observed = 508.2268

HPLC: Rt = 19.5 min. EC 250/4 Nucleodur 100-5 C18 Pyramid, 3µ, EC, UV254, Gradient from 10% to

50% in 10 minutes and subsequently from 50% to 90% in 10 minutes, CH3OH/H2O, 0.1% TFA

1H NMR (360 MHz, DMSO): δ [ppm] = 9.54 (s, 4H, -OH), 8.18 (s, 2H, -NH), 7.10 (d, J = 8.6 Hz, 8H,

ArH), 6.78 (d, J = 8.6 Hz, 8H, ArH), 3.08 (t, J = 5.5 Hz, 4H, -NCH2), 2.59 (t, J = 5.3 Hz, 4H, -CH2).

13C NMR (91 MHz, DMSO): δ [ppm] = 156.66 (ArC), 149.71 (Guanidinium, C), 134.32, 127.14, 115.33

(ArC), 59.79 (quart. C), 44.05 (-NCH2), 31.60 (-CH2).

13C NMR-DEPT (91 MHz, DMSO): δ [ppm] = 127.14, 115.34 (ArCH), 44.07 (-NCH2), 31.60 (-CH2).

Bis-(3,5-dimethoxy-phenyl)phosphoric acid ethyl est er 96

3,5-dimethoxy phenol 94 (10 g, 64.86 mmol) was dissolved in aq. NaOH solution (2.6 g NaOH in 40 ml

water, 65 mmol) under stirring till the clear solution was obtained. Triton B (740 mg, 3.24 mmol) was

added to the solution at room temperature. A solution of ethyl dichlorophosphate 95 (3.66 ml, 30.84

mmol) in 20 ml dichloromethane was added dropwise to the reaction mixture maintaining the

temperature below 20°C over a 20-minute interval. The reaction was monitored by TLC. It was stirred

for overnight in a water bath at room temperature. When the reaction was completed, two phases

were separated and the aqueous layer was extracted with dichloromethane (1 x 100 ml). The

combined organic fractions were washed successively with 1N NaOH solution (1 x 50 ml), distilled

water (1 x 50 ml) and brine (1 x 50 ml). The dichloromethane fraction was dried over MgSO4 and the

solvent was evaporated in vacuo to afford the crude product as a pale yellow liquid. The crude product

was subsequently distilled in Kugelrohr apparatus at 300°C / 4 x 10-2 mbar to yield the pure product as

a colourless liquid.

96: C18H23O8P (M = 398.34 g/mol) Yield: 85% b.p.: 300°C / 4x10-2 mbar

88

MS-ESI: m/z = 399.2 [(M+H)+, 100%]

HPLC: Rt = 21 min. EC 250/4.6 Nucleodur C18 Pyramid, 3µ, UV254, Gradient from 10% to 50% in 10

minutes and subsequently from 50% to 90% in 10 minutes, CH3OH/H2O, 0.1% TFA

1H NMR (250 MHz, CDCl3): δ [ppm] = 6.41 (dd, J = 2.2, 1.1 Hz, 4H, ArH), 6.29 (dd, J = 2.9, 1.4 Hz,

2H, ArH), 4.32 (q, J = 8.4, 7.1 Hz, 2H, -OCH2CH3), 3.75 (s, 12H, -OCH3), 1.38 (t, J = 7.1, 1.1 Hz, 3H, -

OCH2CH3).

13C NMR (63 MHz, CDCl3): δ [ppm] = 161.26, 151.99 (d), 98.77 (d), 97.68 (ArC), 65.59 (d) (-

OCH2CH3), 55.47 (-OCH3), 16.11 (d) (-OCH2CH3).

13C NMR-DEPT (63 MHz, CDCl3): δ [ppm] = 98.77 (d), 97.68 (ArCH), 65.59 (d) (-OCH2CH3), 55.47 (-

OCH3), 16.11, (d, J = 6.7 Hz, -OCH2CH3).

Bis-(2-hydroxy-4,6-dimethoxy-phenyl)-phosphinic aci d ethyl ester 97

7.5 g of phosphate ester 96 (18.83 mmol) was dissolved in 190 ml of absolute tetrahydrofuran. The

solution was cooled to -78°C in a dry ice-acetone bath. 1.8 M solution of lithium di-isopropylamine (in

THF / heptane / ethylbenzene) (32 ml, 58 mmol) was slowly added to the reaction mass over a 20-

minute period. It was stirred further for 20-30 minutes at -78°C. After about 30 minutes, the reaction

showed complete conversion of the starting material and the intermediates to the desired product. The

reaction was stirred further for additional 30 minutes. It was then poured slowly in an ice-cold dilute

hydrochloric acid (20 ml of 4N HCl in 100 ml ice-cold water) solution followed by the extractions with

dichloromethane (2 x 50 ml). The organic fractions were washed twice with brine (25 ml), dried over

MgSO4 and the solvent was evaporated in vacuo to yield the crude product. The crude product was

then purified by re-crystallization from diethyl ether at -8°C.

97: C18H23O8P (M = 398.34 g/mol) Yield: 80% m.p.: 118°C

89

MS-ESI: m/z = 399.1 [(M+H)+, 100%], 819.0 [(2M+Na)+, 30%]

HPLC: Rt = 9 min. EC 250/4.6 Nucleodur C18 Pyramid, 3µ, UV254, Gradient from 70% to 90% in 5

minutes and subsequently from 90% to 90% in 5 minutes, CH3OH/H2O, 0.1% TFA

1H NMR (360 MHz, DMSO): δ [ppm] = 10.56 (s, 2H, -OH), 6.07 – 6.01 (m, 4H, ArH), 4.06 (q, J = 14.2,

7.1 Hz, 2H, -OCH2CH3), 3.75 (s, 6H, -OCH3), 3.58 (s, 6H, -OCH3), 1.27 (t, J = 7.0 Hz, 3H, -OCH2CH3).

13C NMR (91 MHz, DMSO): δ [ppm] = 164.90 (d), 96.72 (d), 94.01 (d), 90.68 (d) (ArC), 61.36 (d) (-

OCH2CH3), 55.93 (d) (-OCH3), 16.08 (d) (-OCH2CH3).

13C NMR-DEPT (91 MHz, DMSO): δ [ppm] = 94.01 (d, J = 9.8 Hz, ArCH), 90.68 (d, J = 7.6 Hz, ArCH),

61.36 (d, J = 5.8 Hz, -OCH2CH3), 55.93 (d, J = 5.8 Hz, -OCH3), 16.05 (d, J = 6.2 Hz, -OCH2CH3)

Bis-(2-trifluoromethanesulfonyloxy-4,6-dimethoxy-ph enyl)-phosphinic acid ethyl ester 99

Sodium hydride (231 mg, 5.77 mmol, 60% dispersion in mineral oil) and 20 ml absolute

tetrahydrofuran were placed in a 100 ml oven dried round-bottomed flask. The suspension was cooled

to 0°C in an ice-salt bath. A solution of bisphenol phosphinate 97 (1 g, 2.51 mmol) in 20 ml of absolute

THF was added to the reaction mixture via a syringe pump over a 20-minute interval. After 30 minutes

of stirring at 0°C, a solution of PhNTf2 98 (1.97 g, 5.52 mmol) in 30 ml THF was added to the reaction

mass over a 30-minute period. The reaction mixture was allowed to warm to room temperature and

stirring continued for overnight. After completion of the reaction, it was poured into a 1:1 solution of

ethyl acetate: water (100 ml). The organic phase was separated and the aqueous phase was

extracted with ethyl acetate (1x 50 ml). The combined ethyl acetate fraction was washed successively

with half-saturated brine (1 x 50 ml) and saturated brine (1 x 50 ml), dried over MgSO4 and the solvent

was evaporated in vacuo to yield the crude product. The crude product was purified by silica gel

chromatography using 40% ethyl acetate in petroleum ether as an eluent.

90

99: C20H21F6O12PS2 (M = 662.47 g/mol) Yield: 72% m.p.: 125°C

MS-ESI: m/z = 663.0 [(M+H)+, 70%], 685.0 [(M+Na)+, 100%]

HPLC: Rt = 13.5 min.(bistriflate), 13 min. (monotriflate) EC 250/4 Nucleodur 100-5 C18 Pyramid, 3µ,

UV254, Gradient from 70% to 90% in 5 minutes and subsequently from 90% to 90% in 5 minutes,

CH3OH/H2O, 0.1% TFA

1H NMR (250 MHz, CDCl3): δ [ppm] = 6.42 (dd, J = 3.9, 1.7 Hz, 2H, ArH), 6.33 (dd, J = 4.0, 2.0 Hz,

2H, ArH), 4.23 (q, J = 9.8, 7.1 Hz, 2H, -OCH2CH3), 3.81 (s, 6H, -OCH3), 3.60 (s, 6H, -OCH3), 1.28 (t, J

= 7.1 Hz, 3H, -OCH2CH3).

13C NMR (63 MHz, CDCl3): δ [ppm] = 163.90 (d), 163.62, 152.69 (ArC), 116.35(-CF3), 109.78 (ArC),

100.33 (d, J = 7.5 Hz, ArCH), 98.06 (d, J = 7.1 Hz) (ArCH), 61.87 (d) (-OCH2CH3), 55.86 (-OCH3),

16.99 (d) (-OCH2CH3).

13C NMR-DEPT (63 MHz, CDCl3): δ [ppm] = 100.33 (d), 98.06 (d) (ArCH), 61.87 (d, J = 6.0 Hz, -

OCH2CH3), 55.86 (-OCH3), 16.99, (d) (-OCH2CH3).

5-Ethoxy-2,4,6,8-tetramethoxy-dibenzophosphole-5-ox ide 102

A 25 ml oven dried single-neck round-bottomed flask equipped with a septum and a stirring bar, was

charged with potassium phosphate (800 mg, 3.77 mmol), bis-(neopentylglycolato)diboron 101 (410

mg, 1.81 mmol), bis-(triphenylphosphine)palladium(II)dichloride (106 mg, 10 mol%) and triphenyl

phosphine (60 mg, 15 mol%). Anhydrous DMF (7 ml) was added to the reaction mixture and nitrogen

was bubbled through for 10 minutes followed by the addition of a solution of bistriflate 99 (1 g, 1.51

mmol, dried over molecular sieves). The reaction mixture was heated to 100°C in an oil bath and was

monitored by HPLC. After 4 hours at 100°C, the reaction was completed. It was worked up first by

centrifuging the reaction mass to remove the insoluble impurities and then the supernatant was diluted

91

with dichloromethane (50 ml) and washed successively with water (3 x 25 ml), dilute HCl (20 % 6N

HCl in water, 1 x 25 ml) and brine (1 x 25 ml). The dichloromethane fraction was dried over MgSO4

and the solvent was evaporated in vacuo to yield the crude oily product. The crude was then

precipitated with diethyl ether and filtered. The crude product was then re-crystallized from MTBE/n-

hexane or toluene.

102: C18H21O6P (M = 364.33 g/mol) Yield: 65% m.p.: 140°C

MS-ESI: m/z = 365.1 [(M+H)+, 100%]

HPLC: Rt = 16 min. EC 250/4 Nucleodur 100-5 C18 Pyramid, 3µm, UV254, Gradient from 50% to 90%

in 10 minutes and subsequently from 90% to 90% in 10 minutes, CH3OH/H2O, 0.1% TFA

1H NMR (360 MHz, CDCl3): δ [ppm] = 6.78 (t, J = 1.9 Hz, 2H, ArH), 6.37 (dd, J = 4.9, 1.6 Hz, 2H,

ArH), 4.23 (q, J = 8.8, 7.1 Hz, 2H, -OCH2CH3), 3.90 (d, J = 2.3 Hz, 12H, -OCH3), 1.31 (t, J = 7.1 Hz,

3H, -OCH2CH3).

13C NMR (91 MHz, CDCl3): δ [ppm] = 161.81, 143.30, 110.27, 99.28 (d, J = 13.5 Hz), 98.66 (d, J = 7.6

Hz) (ArC), 62.56 (d, J = 6.2 Hz, -OCH2CH3), 55.88 (d, J = 4.8 Hz, -OCH3), 16.55 (-OCH2CH3)

13C NMR-DEPT (91 MHz, CDCl3): δ [ppm] = 99.28 (d), 98.66 (d) (ArCH), 62.56 (-OCH2CH3), 55.88 (-

OCH3), 16.55 (-OCH2CH3)

10-Ethoxy-1,3,7,9-tetramethoxy-phenoxaphosphine-10- oxide 117

A screw-cap vial was charged with bis-triflate compound 99 (100 mg, 0.151 mmol), sodium hydride (8

mg, 0.2 mmol, 60% dispersion in mineral oil) and anhydrous dimethyl formamide (1 ml). Distilled water

(approx. 3 µl, 0.167 mmol) was added to the reaction mixture and was stirred for 15 minutes at room

temperature. The reaction mixture was then heated to 100°C in an oil bath and the reaction progress

92

was followed by HPLC. After completion of the reaction, the reaction mixture was diluted with 10 ml

dichloromethane: water (1:1) and the organic phase was separated from the aqueous one. It was

washed successively with water (3 x 2 ml), brine (1 x 2 ml) and dried over MgSO4. The solvent was

evaporated and the residue was dried in high vacuum. The crude product obtained was purified by re-

crystallization from toluene.

117: C18H21O7P (M = 380.33 g/mol) Yield: 52% m.p.: 125°C

MS-ESI: m/z = 381.2 [(M+H)+, 100%]; 403.2 [(M+Na)+, 38%]

HPLC: Rt = 14 min. EC 250/4 Nucleodur 100-5 C18 EC, UV254, Gradient from 10% to 50% in 10

minutes and subsequently from 50% to 90% in 10 minutes, CH3OH/H2O, 0.1% TFA.

1H NMR (250 MHz, CDCl3): δ [ppm] = 6.25 (dd, J = 5.7, 2.0 Hz, 2H, ArH), 6.19 (dd, J = 2.9, 2.2 Hz,

2H, ArH), 4.29 – 4.13 (m, 2H, -OCH2CH3), 3.85 (s, 6H, -OCH3), 3.79 (s, 6H, -OCH3), 1.27 (t, J = 7.1

Hz, 3H, -OCH2CH3).

13C NMR (63 MHz, CDCl3): δ [ppm] = 164.28, 162.67, 158.67, 94.34, 93.88, 77.67 (ArC), 62.72 (-

OCH2CH3), 55.61 (-OCH3), 16.29 (-OCH2CH3).

13C NMR-DEPT (63 MHz, CDCl3): δ [ppm] = 94.26 (d, J = 6.2 Hz, ArCH), 93.79 (d, J = 7.4 Hz, ArCH),

62.64 (d, J = 6.2 Hz, -OCH2CH3), 55.61 (-OCH3), 16.29 (d, J = 7.3 Hz, -OCH2CH3).

5-Oxo-5H-5λλλλ5-dibenzophosphole-2,4,5,6,8-pentaol 120

An oven dried 25 ml two-necked round-bottomed flask equipped with a septum, a reflux condenser

and a stirring bar kept under nitrogen atmosphere, was charged with 1M BBr3 solution (4 ml, 4.116

mmol, 30 eq.). A solution of tetra-methoxy phosphinate 102 (50 mg, 0.1372 mmol, 1 eq.) in 5 ml of

anhydrous dichloromethane was added slowly to the refluxing reaction mixture over a 5-hour interval

using a syringe pump at the rate of 1 ml / h. The reaction mixture was stirred further overnight. After

93

overnight stirring, dichloromethane was removed by decantation and the precipitated solid was

quenched slowly with methanol (25 ml) at 0°C. The methanol fraction was centrifuged to remove

insoluble impurities and the solvent was evaporated in vacuo to yield the crude product. The crude

product was then washed with ether (5 x 10 ml), and with dichloromethane (2 x 10 ml) to remove the

impurities and the residue obtained as an oily product 120.

120: C35H38N3O4+ (M = 564.69 g/mol) Yield: 25%

MS-ESI: m/z = 309.1 [(M+H)+, 100%], 639.6 [(2M+Na)+, 30%]

HPLC: Rt = 21 min. EC 250/4 Nucleodur 100-5 C18 EC, UV254, Gradient from 10% to 50% in 10

minutes and subsequently from 50% to 90% in 10 minutes, CH3CN/H2O, 0.1% TFA.

1H NMR (250 MHz, DMSO): δ [ppm] = 7.67 (dd, J = 5.8, 3.2 Hz, 2H, ArH), 7.58 (dd, J = 5.8, 3.2 Hz,

2H, ArH).

13C NMR (63 MHz, DMSO): δ [ppm] = 169.10, 133.24, 131.21, 128.77 (ArC)

13C NMR-DEPT(63 MHz, DMSO): δ [ppm] = 131.21, 128.77 (ArCH).

5-Ethoxy-4,6-dimethoxy-5-oxo-5 H-5λλλλ5-dibenzophosphole-2,8-diol 104

To a solution of tetra-methoxy compound 102 (500 mg, 1.37 mmol) in 5 ml anhydrous

dichloromethane, cooled to -40°C using dry ice-acetone, neat boron tri-bromide (400 µl, 4.12 mmol)

was added. The reaction mixture was stirred at -40°C for about 2 hours. The reaction was followed by

HPLC. After about 2h stirring at -40°C, the HPLC chromatogram was indicated the desired product as

a major peak. The reaction was then quenched with methanol. The solvent was evaporated and the

crude (containing 103, 105 and 106) was purified by re-crystallization from acetonitrile (or column

chromatography – Prep. column: Silica C8, Eluents: 30% Methanol in water).

94

104: C16H17O6P (M = 336.28 g/mol) Yield: 60% m.p.: 276°C

MS-ESI: m/z = 337.1 [(M+H)+, 100%], 694.9 [(2M+Na)+, 60%]

HRMS: Calculated = 335.0690 Observed = 335.0678

HPLC: Rt = 12 min. EC 250/4 Nucleodur 100-5 C18 Pyramid, 3µ, UV254, Gradient from 50% to 90% in

10 minutes and subsequently from 90% to 90% in 5 minutes, CH3OH/H2O, 0.1% TFA.

1H NMR (500 MHz, DMSO): δ [ppm] = 10.32 (s, 2H, -OH), 6.74 (s, 2H, ArH), 6.44 – 6.32 (m, 2H, ArH),

3.87 (q, J = 14.2, 7.0 Hz, 2H, -OCH2CH3), 3.79 (s, 6H, -OCH3), 1.13 (t, J = 7.0 Hz, 3H, -OCH2CH3).

13C NMR (91 MHz, DMSO): δ [ppm] = 164.01 (d, J = 1.8 Hz), 161.26 (d, J = 3.4 Hz), 142.41, 107.38,

101.11 (d, J = 12.8 Hz), 99.44 (d, J = 7.5 Hz) (ArC), 61.26 (d, J = 5.8 Hz, -OCH2CH3), 55.42 (-OCH3),

16.12 (d, J = 6.3 Hz, -OCH2CH3).

13C NMR (91 MHz, DMSO): δ [ppm] = 101.11 (d, J = 12.8 Hz), 99.44 (d, J = 7.5 Hz) (ArCH), 61.26 (d,

J = 5.8 Hz, -OCH2CH3), 55.42 (-OCH3), 16.12 (d, J = 6.3 Hz, -OCH2CH3).

5-Ethoxy-6-methoxy-5-oxo-5 H-5λλλλ5-dibenzophosphole-2,4,8-triol 105

105: C15H15O6P (M = 322.25 g/mol)

HPLC: Rt = 15 min. EC 250/4 Nucleodur 100-5 CN EC, UV254, Gradient from 10% to 50% in 10

minutes and subsequently from 50% to 90% in 10 minutes, CH3OH/H2O, 0.1% TFA.

1H NMR (360 MHz, DMSO): δ [ppm] = 10.18 (s, 3H, -OH), 6.73 (s, 1H, ArH), 6.62 (s, 1H, ArH), 6.38

(d, J = 5.0 Hz, 1H, ArH), 6.28 (d, J = 4.9 Hz, 1H, ArH), 3.89 (dd, J = 9.0, 7.1 Hz, 2H, -OCH2CH3), 3.78

(d, J = 2.8 Hz, 3H, -OCH3), 1.14 (t, J = 7.0 Hz, 3H, -OCH2CH3).

95

5-Ethoxy-5-oxo-5 H-5λλλλ5-dibenzophosphole-2,4,6,8-tetraol 106

1H NMR (360 MHz, DMSO): δ [ppm] = 10.12 (s, 4H, -OH), 6.61 (s, 2H, ArH), 6.27 (d, J = 2.9 Hz, 2H,

ArH), 3.98 – 3.83 (m, 2H, -OCH2CH3), 1.15 (t, J = 6.7 Hz, 3H, -OCH2CH3).

HPLC: Rt = 12 min. EC 250/4 Nucleodur 100-5 CN EC, UV254, Gradient from 10% to 50% in 10

minutes and subsequently from 50% to 90% in 10 minutes, CH3OH/H2O, 0.1% TFA.

3-Bromo-2,2-bis-hydroxymethyl-propan-1-ol 121

In a 2-L, three-necked flask equipped with a reflux condenser and an addition funnel were placed 200

g (1.47 mol) of pentaerythritol and 16 ml of 48% hydrobromic acid in about 500 ml of glacial acetic

acid. The mixture was refluxed for ~0.5h until all pentaerythritol was dissolved. Then 170 ml of 48%

hydrobromic acid was added, followed by 310 ml of acetic anhydride, and the mixture was refluxed for

3h. Then 94 ml of 48% hydrobromic acid was added followed by 150 ml of acetic anhydride and the

mixture was refluxed for additional 3h. The acetic acid was then removed as completely as possible

on a rotary evaporator. Ethanol (95%, 750 ml) and 17 ml of 48% hydrobromic acid were then added to

the residue. The flask was equipped with a 1-m vigreux column and the ethyl acetate / ethanol

azeotrope was slowly distilled. When 500 ml of distillate had been collected, an additional 750 ml of 95

% ethanol was added and the distillation was continued until about 400 ml remained. After cooling, the

precipitated pentaerythritol was filtered. The solvent was evaporated and the residue was dissolved in

water. The aqueous phase was extracted twice with carbon tetrachloride and twice with ether. The

water was evaporated on the rotary evaporator and the last traces of water were removed by

96

azeotropic distillation with toluene using a Dean-Stark trap. The solid was re-crystallized from

chloroform containing 10% acetonitrile.

121: C5H11BrO3 (M = 199.04 g/mol) Yield: 65% m.p.: 74°C

HPLC: Rt = 21 min. EC 250/4 Nucleodur 100-5 C18 EC, UV254, Gradient from 10% to 50% in 10

minutes and subsequently from 50% to 90% in 10 minutes, CH3CN/H2O, 0.1% TFA

1H NMR (360 MHz, DMSO): δ [ppm] = 4.38 (s, 3H, -OH), 3.47 (s, 2H, -CH2Br), 3.36 (s, 6H, -CH2OH).

13C NMR (91 MHz, DMSO): δ [ppm] = 60.35 (-CH2OH), 45.22 (quart., C), 37.22 (-CH2Br)

13C NMR-DEPT (91 MHz, DMSO): δ [ppm] = 60.35 (-CH2OH), 37.22 (-CH2Br)

4-(Bromomethyl)-2,6,7-trioxabicyclo-[2.2.2]-octane 107 and 4-(Bromomethyl)-1-methyl-2,6,7-

trioxabicyclo-[2.2.2]-octane 108

A mixture of monobromopentaerythritol (15.1 g, 0.025 mmol), the corresponding triethyl ester (4.54 g,

0.028 mmol, triethyl formic ester), p-toluene sulfonic acid (190 mg, 0.001 mmol), and toluene (10 ml)

were placed in a 100 ml round-bottomed flask. The mixture was heated at 90°C for 7 h. The ethanol

by-product formed during the reaction was azeotropically removed. After cooling and concentration of

the reaction mixture, sublimation gave a white powder of the corresponding product 107 or 108.

107: C6H9BrO3 (M = 209.04 g/mol) ESI-MS: m/z = 209.0 [(M+H) +, 52%]

1H NMR (360 MHz, DMSO): δ [ppm] = 5.54 (s, 1H, -CH), 4.01 (s, 6H, -OCH2), 3.12 (s, 2H, -CH2Br).

13C NMR (91 MHz, DMSO): δ [ppm] = 102.00 (-CH), 69.32 (-OCH2), 34.37 (quart., C), 29.54 (-CH2Br). 13C NMR-DEPT (91 MHz, DMSO): δ [ppm] = 102.00 (-CH), 69.32 (-OCH2), 29.54(-CH2Br).

97

108: C7H11BrO3 (M = 223.06 g/mol) Yield: 65% m.p.: 70°C

MS-ESI: m/z = 223.0 [(M+H) +, 67%], 709.0 [(3M+H2O+Na) +, 100%]

1H NMR (360 MHz, DMSO): δ [ppm] = 3.92 (s, 6H, -CH2O), 3.44 (s, 2H, -CH2Br), 1.32 (s, 3H, -CH3).

13C NMR (91 MHz, DMSO): δ [ppm] = 107.92 (quart., C), 68.97 (-CH2O), 33.40 (quart., C) 31.14 (-

CH2Br), 23.15 (-CH3)

13C NMR-DEPT (91 MHz, DMSO): δ [ppm] = 68.97 (-CH2O), 31.14 (-CH2Br), 23.16 (-CH3)

4,6-Dimethoxy-2,8-bis-(1-methyl-2,6,7-trioxa-bicycl o-[2.2.2]-oct-4-ylmethoxy)-5-Ethoxy-5-oxo-5H-5λλλλ5-dibenzophosphole 109

To a solution of bis-methoxy compound 104 (600 mg, 1.7842 mmol) in 20 ml dimethyl formamide,

caesium carbonate (1.75 g, 5.3526 mmol) and bromomethyl bicyclo-compound 108 (1.28 g, 5.7094

mmol) were added at room temperature. Then the reaction mass was heated at 60-80°C till the

completion of the reaction (4-6 h). After completion of the reaction, it was diluted with dichloromethane

(50 ml), washed successively with water (3 x 50 ml), brine (1 x 50 ml), dried over MgSO4 and the

solvent was evaporated. The crude was washed with n-hexane (50 ml) and the residue was then

precipitated from methanol as a white solid.

109: C30H37O12P (M = 620.58 g/mol) Yield: 65% m.p.: 225°C

MS-ESI: m/z = 621.5 [(M+H) +, 100%]; 639.5 [(M+H) +, 83%] mass corresponds to one ortho-ester ring

opened-up product.

HRMS: Calculated = 643.1915, Observed = 643.1879 as sodium adduct

98

HPLC: Rt = 12.25 min. EC 250/4 Nucleodur 100-5 C18 Pyramid, 3µ, UV254, Gradient from 50% to 90%

in 10 minutes and subsequently from 90% to 90% in 5 minutes, CH3OH/H2O, 0.1% TFA

1H NMR (250 MHz, DMSO): δ [ppm] = 7.17 (s, 2H, ArH), 6.59 (dd, J = 4.9, 1.5 Hz, 2H, ArH), 4.06 (s,

12H, -CH2O), 4.01 (s, 4H, -OCH2), 3.94 – 3.86 (m, 2H, -OCH2CH3), 3.84 (s, 6H, -OCH3), 1.35 (s, 6H, -

CH3), 1.11 (t, J = 7.0 Hz, 3H, -OCH2CH3).

4,6-Dimethoxy-2,8-bis-(2,6,7-trioxa-bicyclo-[2.2.2] -oct-4-ylmethoxy)-5-Ethoxy-5-oxo-5 H-5λλλλ5-dibenzophosphole 122

Same procedure as 109.

1H NMR (250 MHz, DMSO): δ [ppm] = 7.18 (d, J = 1.8 Hz, 2H, ArH), 6.60 (dd, J = 4.9, 1.5 Hz, 2H,

ArH), 5.63 (s, 2H, -CH), 4.07 (s, 12H, -CH2O,ring), 3.92 (s, 6H, -OCH2), 3.85 (s, 6H, -OCH3), 1.11 (t, J

= 7.0 Hz, 3H, -OCH2CH3).

2,8-Bis-(1-methyl-2,6,7-trioxa-bicyclo-[2.2.2]-oct- 4-ylmethoxy)-5-oxo-5,9b-dihydro-4a H-5λλλλ5-dibenzophosphole-4,5,6-triol 113

A solution of bis-methoxy-alkylated compound 109 (200 mg, 0.3222 mmol), sodium hydride (65 mg,

1.611 mmol as 60% oil dispersion) and heptyl thiol (162 µl, 1.031 mmol) in anhydrous DMF (10 ml)

was heated to 120°C in an oil bath. The reaction was completed in about 2-3h. After completion, the

reaction was quenched with water (10 ml) and pH was adjusted to 6-7 with 0.1 M HCl solution. The

99

mixture was extracted with diethyl ether (3 x 20 ml) to remove heptyl thiol. The aqueous DMF phase

was evaporated to dryness and the residue was re-crystallized from ethanol.

113 and 113a: C26H33O14P (M = 600.51 g/mol) Yield: 53% m.p.: 295°C (as Na-

Salt)

MS-ESI: m/z = 565.2 [(M+H) +, 72%], 583.2 [(M+H2O) +, 100%], 601.2 [(M+H) +, 100%, ring opened]

HPLC: Rt = 5 min. EC 250/4 Nucleodur 100-5 C18 Pyramid, 3µ, UV254, Gradient from 50% to 90% in

10 minutes and subsequently from 90% to 90% in 5 minutes, CH3OH/H2O, 0.1% TFA

1H NMR (250 MHz, MeOD): δ [ppm] = 6.98 (s, 2H, ArH), 6.37 (d, J = 4.7 Hz, 2H, ArH), 4.20 (s, 4H, -

OCH2), 4.06 (s, 4H, -CH2OCO), 3.70 (s, 8H, -CH2OH), 2.05 (s, 6H, -COCH3).

5-hydroxy-2,8-bis-(1-methyl-2,6,7-trioxa-bicyclo-[2 .2.2]-oct-4-ylmethoxy)-5-oxo-6-

trifluoromethanesulfonyloxy-5a,9a-dihydro-5 H-5λλλλ5-dibenzophosphol-4-olate sodium salt 114

To a solution of the compound 113 (25 mg, 0.0443 mmol) in anhydrous dimethyl formamide (4 ml)

sodium hydride (3 mg, 0.1329 mmol) was added followed by the addition of PhNTf2 (48 mg, 0.1329

mmol) at 0°C. The reaction was allowed to warm to room temperature. Then it was heated to 60°C for

about 4h till the complete conversion of compound 113. After completion, the solvent was removed

and the residue was used for the subsequent reaction without further purification.

114: C28H26F6NaO16PS2 (M = 850.58 g/mol)

MS-ESI: m/z = 865.3 [(M+H) +, 100%] (mass corresponds to opened-up ortho-ester under acidic

conditions)

100

HPLC: Rt = 15.5 min. EC 250/4 Nucleodur 100-5 C18 Pyramid, 3µ, UV254, Gradient from 50% to 90%

in 10 minutes and subsequently from 90% to 90% in 5 minutes, CH3OH/H2O, 0.1% TFA

1H NMR (360 MHz, MeOD): δ [ppm] = 7.63 (s, 2H, ArH), 6.91 (s, 2H, ArH), 4.19 (d, J = 26.1 Hz, 8H, -

OCH2), 3.72 (s, 8H, -CH2OH), 2.05 (s, 6H, -COCH3).

13C NMR (91 MHz, MeOD): δ [ppm] = 172.87 (-COCH3), 164.94, 150.87 (ArC), 116.12 (-CF3), 109.21,

108.48 (ArCH), 68.67, 63.93, 61.32 (-CH2), 46.24 (quart., C), 20.72 (-COCH3)

13C NMR-DEPT (91 MHz, MeOD): δ [ppm] = 109.22, 108.49 (ArCH), 68.66, 63.92, 61.31 (-CH2), 20.72

(-COCH3)

2,8-Bis-(3-hydroxy-2,2-bis-hydroxymethyl-propoxy)-5 -oxo-4,6-bis-phenylethynyl-5,9b-dihydro-4aH-5λλλλ5-dibenzophosphol-5-olate sodium salt 37

An oven-dried screw-cap vial equipped with a triangular magnetic stirring bar, was charged with bis-

hydroxy compound 113 (100 mg, 0.1705 mmol) and sodium hydride (20 mg, 0.5115 mmol, 60%

dispersion in mineral oil). Anhydrous dimethyl formamide (2 ml) was injected to the reaction mixture

kept under nitrogen atmosphere followed by the addition of a solution of PhNTf2 (183 mg, 0.5115

mmol) in 2 ml of DMF. The reaction mixture was then stirred at room temperature till completion of the

reaction. After completion of the reaction, DMF was removed in Kugelrohr apparatus under high

vacuum to dryness. The residue was washed with diethyl ether (10 ml), suspended in anhydrous

acetonitrile (5 ml) and was filtered through a pad of MgSO4. The clear solution obtained (bistriflate

114) was used as such for the subsequent Sonogashira coupling.

To the acetonitrile solution of 114 was added (Ph3P)4Pd (20 mg, 10 mol %), triphenyl phosphine (6

mg, 15 mol%), triethyl amine (60 µl, 0.8525 mmol) and phenyl acetylene (40 µl, 0.682 mmol). The

reaction mixture was then heated to reflux in an oil bath. After completion of the reaction, the solvent

101

was evaporated and the residue obtained was washed with diethyl ether (10 ml). Then the crude

product was dissolved in methanol:water (1:1) and was acidified with TFA (pH ~ 2-3). The solution

was stirred further for 30 minutes at room temperature. After 30 minutes, it was basified with aqueous

4N NaOH solution to pH~10 and stirring was continued for additional 30 minutes. The reaction was

followed by HPLC till the complete hydrolysis of an intermediate ester to the corresponding alcohol

(ortho-ester ring opening). Then the reaction mass was neutralized with TFA (pH~6). It was extracted

with ether to remove hydrophobic impurities from the catalyst and the ligand. The aqueous methanolic

fraction was then purified using an SPE cartridge to yield the phosphinate 37. It was re-crystallized

from methanol:toluene (1:2) solvent mixture.

37: C38H37O10P (MW = 684.67 g/mol) Yield: 30 mg , 25% m.p.: > 300°C

MS-ESI: m/z = 685.2 [(M+H)+, 100%], 1369.1 [(2M+H)+, 30%]

HRMS: Calculated = 685.2203, Observed = 685.2214

HPLC: Rt = 10.5 min (for bis-triflate product) and Rt = 11.5 min (for bis-Sonogashira product) EC 250/4

Nucleodur 100-5 CN, UV254, Gradient from 30% to 90% in 10 minutes and subsequently from 90% to

90% in 5 minutes, MeOH/H2O, 0.1% TFA

1H NMR (250 MHz, DMSO): δ [ppm] = 7.65 (d, J = 7.8 Hz, 4H, ArH), 7.42 (d, J = 10.0 Hz, 8H, ArH),

6.91 (s, 2H, ArH), 4.59 (s, 6H, -CH2OH), 3.92 (s, 4H, -OCH2), 3.48 (d, J = 4.4 Hz, 12H, -CH2OH).

13C NMR (91 MHz, DMSO): δ [ppm] = 160.30, 139.48, 135.62, 134.18 (ArC), 131.71, 128.44 (ArCH),

123.28, 118.99 (ArC), 117.05, 107.92 (fused ArCH), 91.08 (acetylenic, C), 67.0(-CH2OH), 59.94 (-

OCH2), 45.85 (quart., C).

13C NMR-DEPT (91 MHz, DMSO): δ [ppm] = 131.72, 128.45 (ArCH), 117.10 (d, J = 9.8 Hz, fused

ArCH), 107.96 (d, J = 11.2 Hz, fused ArCH), 67.04 (-CH2OH), 59.94 (-OCH2).

102

2,2,8,8-Tetraallyl-octahydro-pyrimido-[1,2- a]-pyrimidinium bromide 28 [80]

500 mg of allyl guanidinium iodide was dissolved in 70% methanol in water. This solution was loaded

on anion-exchange resin AG4- X4 bromide form (10 ml on wet basis), was slowly eluted with 50%

methanol in water. The solvent was evaporated on the rotatory evaporator and the residue obtained

was crystallized from acetonitrile / diethyl ether.

28: C19H30BrN3 (MW = 380.37 g/mol) Yield: 95% m.p.: 84°C

MS-ESI: m/z = 300.3 [(M)+, 100%]

HPLC: Rt = 5 min. EC 250/4 Nucleodur 100-5 C18 Pyramid,3µ, UV254, Gradient from 30% to 90% in

10 minutes and subsequently from 90% to 90% in 5 minutes, MeOH/H2O, 0.1% TFA

1H NMR (360 MHz, DMSO): δ [ppm] = 7.15 (s, 2H, -NH), 5.87 – 5.70 (m, 4H, allylic –CH), 5.19 (dd, J

= 10.0, 7.9 Hz, 8H, allylic –CH2), 3.35 (dd, J = 14.4, 8.0 Hz, 4H, -NCH2), 2.26 (d, J = 7.3 Hz, 4H, -

NCH2CH2), 1.79 (t, J = 6.2 Hz, 8H, -CH2CH).

13C NMR (91 MHz, DMSO): δ [ppm] = 149.70 (Guanidinium, -C), 132.13 (allylic, -CH), 119.98 (allylic, -

CH2), 53.17 (quart., C), 42.90 (-NCH2), 42.04 (-NCH2CH2), 28.17.( -CH2CH)

13C NMR-DEPT (91 MHz, DMSO): δ [ppm] = 132.13 (allylic, -CH), 119.98 (allylic, -CH2), 42.90 (-

NCH2), 42.04 (-NCH2CH2), 28.17 (-CH2CH)

103

5.3 Experiments in-silico All the MD simulations were carried out using desktop computers (Pentium PC’s) and GROMOS’96

(an acronym of the GROningen MOlecular Simulation) software provided generously by Prof. W.F.

van Gunsteren, ETH-Zürich.[129]

All topology files were prepared following the instructions given in GROMOS manual[129]. Co-ordinate

files were made by first drawing the molecule structure in ChemSketch (ACD/Labs version 12.0) and

saved these *.sk files as *.mol files. Second, with the help of Open Babel program (version 2.2.1)

converted the *.mol files into *.gr96 co-ordinate files. The numbering of the structures is not according

to IUPAC system.

The partial charges for phosphorous atom and bond parameters for the C–P bond in the guest

molecule 37 were adapted from the literature.[130] The individual host-guest species were deliberately

placed in a solvent box (CHCl3, DMSO, Methanol and Water) and the system energetically relaxed.

The molecular dynamic runs were conducted for 10 ns. Frames were taken every 5 picoseconds. The

structural analysis was carried out using the GROMOS’96 software package.[138]

5.3.1 Topology file for guanidinium host 51

2,2,8,8-Tetrakis-(4-hydroxy-phenyl)-1,3,4,6,7,9-hex ahydro-2 H-pyrimido[1,2- a]pyrimidinium

(GUA1)

N34

C+

16

33

N14

32

13

36

19

35

N17

8

20

49

37

5754

59

52

50

4745

4238 403028

25

21

23

9

11

6

34

O2

O26

O43

O55

H15

H18

H58

H60

H53

H51

H48

H46

H39

H41H

31

H29

H22

H24

H10

H12

H7

H5

H1

H27

H44

H56

MTBUILDBLSOLUTE # building block (residue, nucleotide, etc.) # RNME GUA1 # number of atoms, number of preceding exclusions # NMAT, NLIN 60 0 # atoms #ATOM ANM IACM MASS CGM ICGM MAE MSAE 1 HO1 18 1 0.398 0 2 2 3

104

2 O1 3 16 -0.548 0 7 3 4 5 6 9 11 12 3 C1 11 12 0.150 1 9 4 5 6 7 8 9 10 11 12 4 C2 11 12 -0.100 0 7 5 6 7 8 9 11 12 5 HC2 17 1 0.100 1 4 6 7 8 11 6 C3 11 12 -0.100 0 6 7 8 9 10 11 13 7 HC3 17 1 0.100 1 3 8 9 13 8 C4 11 12 0.000 1 13 9 10 11 12 13 14 15 16 20 21 30 32 33 9 C5 11 12 -0.100 0 4 10 11 12 13 10 HC5 17 1 0.100 1 3 11 12 13 11 C6 11 12 -0.100 0 2 12 13 12 HC6 17 1 0.100 1 0 13 C7 45 12 0.200 0 14 14 15 16 17 20 21 22 23 28 30 31 32 33 34 14 N1 10 14 -0.110 0 6 15 16 17 20 32 34 15 HN1 18 1 0.240 0 1 16 16 C8 11 12 0.340 0 6 17 18 19 33 34 35 17 N2 10 14 -0.110 0 6 18 19 34 36 37 49 18 HN2 18 1 0.240 0 1 19 19 C9 45 12 0.200 1 17 34 35 36 37 38 39 40 45 47 48 49 50 51 52 57 59 60 20 C10 11 12 0.000 1 9 21 22 23 24 25 28 29 30 31 21 C11 11 12 -0.100 0 8 22 23 24 25 26 28 30 31 22 HC11 17 1 0.100 1 4 23 24 25 30 23 C12 11 12 -0.100 0 7 24 25 26 27 28 29 30 24 HC12 17 1 0.100 1 3 25 26 28 25 C13 11 12 0.150 0 6 26 27 28 29 30 31 26 O2 3 16 -0.548 0 4 27 28 29 30 27 HO2 18 1 0.398 1 0 28 C14 11 12 -0.100 0 3 29 30 31 29 HC14 17 1 0.100 1 2 30 31 30 C15 11 12 -0.100 0 1 31 31 HC15 17 1 0.100 1 0 32 C16 44 4 0.000 1 2 33 34 33 C17 44 4 0.055 0 2 34 35 34 N3 10 14 -0.110 0 2 35 36 35 C18 44 4 0.055 1 1 36 36 C19 44 4 0.000 1 2 37 49 37 C20 11 12 0.000 1 9 38 39 40 41 42 45 46 47 48 38 C21 11 12 -0.100 0 8 39 40 41 42 43 45 47 48 39 HC21 17 1 0.100 1 4 40 41 42 47 40 C22 11 12 -0.100 0 7 41 42 43 44 45 46 47 41 HC22 17 1 0.100 1 3 42 43 45 42 C23 11 12 0.150 0 6 43 44 45 46 47 48 43 O3 3 16 -0.548 0 4 44 45 46 47 44 HO3 18 1 0.398 1 0 45 C24 11 12 -0.100 0 3 46 47 48 46 HC24 17 1 0.100 1 2 47 48 47 C25 11 12 -0.100 0 1 48 48 HC25 17 1 0.100 1 0 49 C26 11 12 0.000 1 9 50 51 52 53 54 57 58 59 60 50 C27 11 12 -0.100 0 6 51 52 53 54 55 57 51 HC27 17 1 0.100 1 4 52 53 54 59 52 C28 11 12 -0.100 0 7 53 54 55 56 57 58 59 53 HC28 17 1 0.100 1 3 54 55 57 54 C29 11 12 0.150 0 6 55 56 57 58 59 60 55 O4 3 16 -0.548 0 4 56 57 58 59 56 HO4 18 1 0.398 1 0 57 C30 11 12 -0.100 0 3 58 59 60 58 HC30 17 1 0.100 1 2 59 60

105

59 C31 11 12 -0.100 0 1 60 60 HC31 17 1 0.100 1 0 # bonds # NB 65 # IB JB MCB IB JB MCB IB JB MCB 1 2 1 2 3 12 3 4 15 3 11 15 4 5 3 4 6 15 6 7 3 6 8 15 8 9 15 8 13 26 9 10 3 9 11 15 11 12 3 13 14 10 13 20 26 13 32 26 14 15 2 14 16 10 16 17 10 16 34 10 17 18 2 17 19 10

19 36 26 19 37 26 19 49 26 20 21 15 20 30 15 21 22 3 21 23 15 23 24 3 23 25 15 25 26 12 25 28 15 26 27 1 28 29 3 28 30 15 30 31 3 32 33 26 33 34 20 34 35 20 35 36 26 37 38 15 37 47 15 38 39 3

38 40 15 40 41 3 40 42 15 42 43 12 42 45 15 43 44 1 45 46 3 45 47 15 47 48 3 49 50 15 49 59 15 50 51 3 50 52 15 52 53 3 52 54 15 54 55 12 54 57 15 55 56 1 57 58 3 57 59 15 59 60 3

# bond angles # NBA 104 # IB JB KB MCB IB JB KB MCB 1 2 3 11 2 3 4 26 2 3 11 26 4 3 11 26 3 4 5 24 5 4 6 24 3 4 6 26 4 6 7 24 4 6 8 26 7 6 8 24 6 8 9 26 6 8 13 26 9 8 13 26 8 9 10 24 8 9 11 26 10 9 11 24 9 11 12 24 9 11 3 26 12 11 3 24 8 13 14 12 14 13 32 12 32 13 20 12 8 13 20 12 8 13 32 12 14 13 20 12 13 14 15 22 13 14 16 26

15 14 16 22 14 16 17 27 14 16 34 27 17 16 34 27 16 17 18 22 16 17 19 26 18 17 19 22 17 19 49 12 17 19 36 12 17 19 37 12 36 19 49 12 37 19 49 12 36 19 37 12 13 20 30 26 13 20 21 26 30 20 21 26 20 21 23 26 22 21 23 24 20 21 22 24 21 23 25 26 21 23 24 24 24 23 25 24 23 25 26 26 23 25 28 26 26 25 28 26 25 26 27 11 25 28 29 24

106

25 28 30 26 29 28 30 24 28 30 31 24 28 30 20 26 31 30 20 24 13 32 33 12 32 33 34 12 33 34 35 26 33 34 16 26 16 34 35 26 34 35 36 12 35 36 19 12 19 37 38 26 19 37 47 26 38 37 47 26 37 38 39 24 37 38 40 26 39 38 40 24 38 40 41 24 38 40 42 26 41 40 42 24 40 42 43 26 40 42 45 26 43 42 45 26 42 43 44 11

42 45 46 24 42 45 47 26 46 45 47 24 45 47 48 24 45 47 37 26 48 47 37 24 19 49 50 26 19 49 59 26 50 49 59 26 49 50 52 26 49 50 51 24 51 50 52 24 50 52 53 24 50 52 54 26 53 52 54 24 52 54 55 26 52 54 57 26 55 54 57 26 54 55 56 11 54 57 58 24 54 57 59 26 58 57 59 24 57 59 60 24 57 59 49 26 60 59 49 24

# improper dihedrals # NIDA 56 # IB JB KB LB MCB IB JB KB LB MCB 3 2 4 11 1 4 3 5 6 1 6 4 7 8 1 8 6 9 13 1 9 8 10 11 1 11 9 12 3 1 3 4 6 8 1 4 6 8 9 1 6 8 9 11 1 8 9 11 3 1 9 11 3 4 1 11 3 4 6 1 20 13 30 21 1 21 20 22 23 1 23 21 24 25 1 25 23 26 28 1 28 25 29 30 1 30 28 31 20 1 20 21 23 25 1 21 23 25 28 1 23 25 28 30 1 25 28 30 20 1 28 30 20 21 1 30 20 21 23 1 37 19 47 38 1 38 37 39 40 1 40 38 41 42 1 42 40 43 45 1

45 42 46 47 1 47 45 48 37 1 37 38 40 42 1 38 40 42 45 1 40 42 45 47 1 42 45 47 37 1 45 47 37 38 1 47 37 38 40 1 49 19 50 59 1 50 49 51 52 1 52 50 53 54 1 54 52 55 57 1 57 54 58 59 1 59 57 60 49 1 49 50 52 54 1 50 52 54 57 1 52 54 57 59 1 54 57 59 49 1 57 59 49 50 1 59 49 50 52 1 14 13 15 16 1 16 14 17 34 1 17 16 18 19 1 34 33 16 35 1 14 16 34 33 1 17 16 34 35 1 19 17 16 34 1 13 14 16 34 1

# dihedrals # NDA

107

16 # IB JB KB LB MCB 1 2 3 4 2 27 26 25 23 2 44 43 42 40 2 56 55 54 52 2 32 13 8 9 5 14 13 20 30 5 17 19 37 38 5 36 19 49 59 5

16 14 13 32 4 14 13 32 33 20 13 32 33 34 17 32 33 34 16 19 33 34 35 36 19 34 35 36 19 17 35 36 19 17 20 36 19 17 16 4

END 5.3.2 Topology file for guanidinium host 34

2,2,8,8-Tetrakis-(3-hydroxy-propyl)-1,3,4,6,7,9-hex ahydro-2 H-pyrimido[1,2- a]pyrimidinium

tetraphenyl borate (GUA2)

N20

C+

9

19

N7

18

6

22

12

21

N10 28

29

30O31

H32

23

24

25

O26

H27

5

4

3O2

H1

13

14

15

O16

H17

H8

H11

MTBUILDBLSOLUTE # building block (residue, nucleotide, etc.) # RNME GUA2 # number of atoms, number of preceding exclusions # NMAT,NLIN 32 0 # atoms #ATOM ANM IACM MASS CGM ICGM MAE MSAE 1 HO1 18 1 0.398 0 2 2 3 2 O1 3 16 -0.548 0 2 3 4 3 C1 13 12 0.150 1 2 4 5 4 C2 13 12 0.000 0 2 5 6 5 C3 13 12 0.000 1 4 6 7 13 18 6 C4 45 12 0.200 0 7 7 8 9 13 14 18 19 7 N1 10 14 -0.110 0 6 8 9 10 13 18 20 8 HN1 18 1 0.240 0 1 9 9 C5 11 12 0.340 0 6 10 11 12 19 20 21 10 N2 10 14 -0.110 0 6 11 12 20 22 23 28 11 HN2 18 1 0.240 0 1 12 12 C6 45 12 0.200 1 6 21 22 23 24 28 29 13 C7 13 12 0.000 0 3 14 15 18 14 C8 13 12 0.000 1 2 15 16 15 C9 13 12 0.150 0 2 16 17 16 O2 3 16 -0.548 0 1 17 17 HO2 18 1 0.398 1 0 18 C10 44 4 0.000 1 2 19 20 19 C11 44 4 0.055 0 2 20 21

108

20 N3 10 14 -0.110 0 2 21 22 21 C12 44 4 0.055 1 1 22 22 C13 44 4 0.000 1 2 23 28 23 C14 13 12 0.000 0 3 24 25 28 24 C15 13 12 0.000 1 2 25 26 25 C16 13 12 0.150 0 2 26 27 26 O3 3 16 -0.548 0 1 27 27 HO3 18 1 0.398 1 0 28 C17 13 12 0.000 0 2 29 30 29 C18 13 12 0.000 1 2 30 31 30 C19 13 12 0.150 0 2 31 32 31 O4 3 16 -0.548 0 1 32 32 HO4 18 1 0.398 1 0 # bonds # NB 33 # IB JB MCB 1 2 1 2 3 17 3 4 26 4 5 26 5 6 26 6 7 10 6 13 26 6 18 26 7 8 2 7 9 10 9 10 10 9 20 10 10 11 2 10 12 10 12 22 26 12 23 26 12 28 26

13 14 26 14 15 26 15 16 17 16 17 1 18 19 26 19 20 20 20 21 20 21 22 26 23 24 26 24 25 26 25 26 17 26 27 1 28 29 26 29 30 26 30 31 17 31 32 1

# bond angles # NBA 44 # IB JB KB MCB 1 2 3 11 2 3 4 12 3 4 5 12 4 5 6 12 5 6 7 12 5 6 13 12 5 6 18 12 7 6 18 12 7 6 13 12 13 6 18 12 6 7 8 22 6 7 9 26 8 7 9 22 7 9 10 27 7 9 20 27 10 9 20 27 9 10 11 22 9 10 12 26 11 10 12 22

10 12 22 12 10 12 23 12 10 12 28 12 22 12 23 12 23 12 28 12 22 12 28 12 6 13 14 12 13 14 15 12 14 15 16 12 15 16 17 11 6 18 19 12 18 19 20 12 19 20 9 26 19 20 21 26 9 20 21 26 20 21 22 12 21 22 12 12 12 23 24 12 23 24 25 12

109

24 25 26 12 25 26 27 11 12 28 29 12

28 29 30 12 29 30 31 12 30 31 32 11

# improper dihedrals # NIDA 8 # IB JB KB LB MCB 7 6 8 9 1 9 7 10 20 1 10 9 11 12 1 20 9 19 21 1

6 7 9 20 1 7 9 20 19 1 10 9 20 21 1 12 10 9 20 1

# dihedrals # NDA 24 # IB JB KB LB MCB 1 2 3 4 12 2 3 4 5 17 3 4 5 6 17 4 5 6 7 20 17 16 15 14 12 16 15 14 13 17 15 14 13 6 17 14 13 6 18 20 27 26 25 24 12 26 25 24 23 17 25 24 23 12 17 24 23 12 10 20

32 31 30 29 12 31 30 29 28 17 30 29 28 12 17 29 28 12 22 20 9 7 6 18 4 7 6 18 19 20 6 18 19 20 17 18 19 20 21 19 19 20 21 22 19 20 21 22 12 17 21 22 12 10 20 22 12 10 9 4

END

5.3.3 Topology file for guanidinium host 28

2,2,8,8-Tetraallyl-octahydro-pyrimido-[1,2- a]-pyrimidine (GUA3)

N16

C+

10

15

N8

14

7

18

13

17

N11

223

21

4

5

6

2324

19

20

21

H9

H12

MTBUILDBLSOLUTE # building block (residue, nucleotide, etc.) # RNME GUA3 # number of atoms, number of preceding exclusions # NMAT,NLIN 24 0 # atoms #ATOM ANM IACM MASS CGM ICGM MAE MSAE 1 C1 16 4 0.000 0 2 2 3 2 C2 16 3 0.000 0 2 3 7 3 C3 13 4 0.000 1 4 4 7 8 14

110

4 C4 13 4 0.000 0 5 5 6 7 8 14 5 C5 16 3 0.000 0 2 6 7 6 C6 16 4 0.000 1 0 7 C7 11 12 0.200 0 5 8 9 10 14 15 8 N1 10 14 -0.110 0 5 9 10 11 14 16 9 HN1 18 1 0.240 0 1 10 10 C8 11 12 0.340 0 6 11 12 13 15 16 17 11 N2 10 14 -0.110 0 6 12 13 16 18 19 22 12 HN2 18 1 0.240 0 1 13 13 C9 11 12 0.200 1 6 17 18 19 20 22 23 14 C10 44 4 0.000 1 2 15 16 15 C11 44 4 0.055 0 2 16 17 16 N3 10 14 -0.110 0 2 17 18 17 C12 44 4 0.055 1 1 18 18 C13 44 4 0.000 1 2 19 22 19 C14 13 4 0.000 0 3 20 21 22 20 C15 16 3 0.000 0 1 21 21 C16 16 4 0.000 1 0 22 C17 13 4 0.000 0 2 23 24 23 C18 16 3 0.000 0 1 24 24 C19 16 4 0.000 1 0 # bonds # NB 25 # IB JB MCB 1 2 9 2 3 26 3 7 26 4 5 26 4 7 26 5 6 9 7 8 10 7 14 26 8 9 2 8 10 10 10 11 10 10 16 10 11 12 2

11 13 10 13 18 26 13 19 26 13 22 26 14 15 26 15 16 20 16 17 20 17 18 26 19 20 26 20 21 9 22 23 26 23 24 9

# bond angles # NBA 36 # IB JB KB MCB 1 2 3 26 2 3 7 12 7 4 5 12 4 5 6 26 3 7 8 12 8 7 14 12 4 7 14 12 3 7 4 12 3 7 14 12 4 7 8 12 7 8 9 22 7 8 10 26 9 8 10 22 8 10 11 27 8 10 16 27

11 10 16 27 10 11 12 22 10 11 13 26 12 11 13 22 11 13 22 12 19 13 22 12 18 13 22 12 11 13 18 12 11 13 19 12 18 13 19 12 7 14 15 12 14 15 16 12 10 16 15 26 10 16 17 26 15 16 17 26

111

16 17 18 12 17 18 13 12 13 19 20 12

19 20 21 26 13 22 23 12 22 23 24 26

# improper dihedrals # NIDA 8 # IB JB KB LB MCB 8 7 9 10 1 10 8 11 16 1 11 10 12 13 1 16 15 10 17 1

7 8 10 16 1 13 11 10 16 1 11 10 16 17 1 8 10 16 15 1

# dihedrals # NDA 16 # IB JB KB LB MCB 1 2 3 7 20 6 5 4 7 20 13 19 20 21 20 13 22 23 24 20 2 3 7 8 20 5 4 7 14 20 11 13 22 23 20 18 13 19 20 20

10 8 7 14 4 10 11 13 18 4 8 7 14 15 20 11 13 18 17 20 7 14 15 16 17 16 17 18 13 17 14 15 16 17 19 15 16 17 18 19

END

5.3.4 Topology file for phosphinate guest 37

2,8-Bis-(3-hydroxy-2,2-bis-hydroxymethyl-propoxy)-5 -oxo-4,6-bis-phenylethynyl-5,9b-

dihydro-4a H-5λλλλ5-dibenzophosphol-5-ol (GST1)

53 P50

16

49

17

35 33

1820

54

14

13

O51

O-

5255 36

3756

38

4739

4541

43

57

66

58

6460

62

O12

11

O21

22

234

24

O25

30

O31

3

O2

8

O9

5O6

27O28

H34

H19

H

H15

H48

H40

H46

H42

H44

H67

H59

H65

H61

H63

H26

H32

H1

H10

H7 H

29

MTBUILDBLSOLUTE # building block (residue, nucleotide, etc.) # RNME GST1

112

# number of atoms, number of preceding exclusions # NMAT,NLIN 69 0 # atoms #ATOM ANM IACM MASS CGM ICGM MAE MSAE 1 HO1 18 1 0.398 0 2 2 3 2 O1 3 16 -0.548 0 2 3 4 3 C1 13 4 0.150 1 4 4 5 8 11 4 C2 11 12 0.000 1 6 5 6 8 9 11 12 5 C3 13 4 0.150 0 4 6 7 8 11 6 O2 3 16 -0.548 0 1 7 7 HO2 18 1 0.398 1 0 8 C4 13 4 0.150 0 3 9 10 11 9 O3 3 16 -0.548 0 1 10 10 HO3 18 1 0.398 1 0 11 C5 13 4 0.274 0 2 12 13 12 O4 3 16 -0.548 0 7 13 14 15 16 54 68 69 13 C6 11 12 0.274 1 9 14 15 16 17 53 54 55 68 69 14 C7 11 12 -0.100 0 10 15 16 17 18 49 50 53 54 68 69 15 HC7 17 1 0.100 1 4 16 17 53 68 16 C8 11 12 0.000 0 13 17 18 19 20 35 49 50 51 52 53 54 55 68 17 C9 11 12 0.000 1 13 18 19 20 21 33 35 36 49 50 51 52 53 54 18 C10 11 12 -0.100 0 10 19 20 21 22 33 34 35 49 50 53 19 HC10 17 1 0.100 1 4 20 21 33 49 20 C11 11 12 0.274 0 8 21 22 23 33 34 35 36 49 21 O5 3 16 -0.548 0 8 22 23 24 27 30 33 34 35 22 C12 13 4 0.274 1 4 23 24 27 30 23 C13 11 12 0.000 1 6 24 25 27 28 30 31 24 C14 13 4 0.150 0 4 25 26 27 30 25 O6 3 16 -0.548 0 1 26 26 HO6 18 1 0.398 1 0 27 C15 13 4 0.150 0 3 28 29 30 28 O7 3 16 -0.548 0 1 29 29 HO7 18 1 0.398 1 0 30 C16 13 4 0.150 0 2 31 32 31 O8 3 16 -0.548 0 1 32 32 HO8 18 1 0.398 1 0 33 C17 11 12 -0.100 0 6 34 35 36 37 49 50 34 HC17 17 1 0.100 1 3 35 36 49 35 C18 11 12 0.000 1 8 36 37 38 49 50 51 52 53 36 C19 11 12 0.000 0 6 37 38 39 47 49 50 37 C20 11 12 0.000 1 8 38 39 40 41 45 47 48 49 38 C21 11 12 0.000 1 9 39 40 41 42 43 45 46 47 48 39 C22 11 12 -0.100 0 8 40 41 42 43 44 45 47 48 40 HC22 17 1 0.100 1 4 41 42 43 47 41 C23 11 12 -0.100 0 6 42 43 44 45 46 47 42 HC23 17 1 0.100 1 3 43 44 45 43 C24 11 12 -0.100 0 5 44 45 46 47 48 44 HC24 17 1 0.100 1 3 45 46 47 45 C25 11 12 -0.100 0 3 46 47 48 46 HC25 17 1 0.100 1 2 47 48 47 C26 11 12 -0.100 0 1 48 48 HC26 17 1 0.100 1 0 49 C27 11 12 -0.300 0 5 50 51 52 53 54 50 P1 27 31 1.000 0 6 51 52 53 54 55 68 51 O9 2 16 -0.700 0 3 52 53 54 52 O10 2 16 -0.700 0 2 53 54 53 C28 11 12 -0.300 1 5 54 55 56 68 69 54 C29 11 12 0.000 1 5 55 56 57 68 69

113

55 C30 11 12 0.000 0 6 56 57 58 66 68 69 56 C31 11 12 0.000 1 8 57 58 59 60 64 66 67 68 57 C32 11 12 0.000 1 9 58 59 60 61 62 64 65 66 67 58 C33 11 12 -0.100 0 8 59 60 61 62 63 64 66 67 59 HC33 17 1 0.100 1 4 60 61 62 66 60 C34 11 12 -0.100 0 6 61 62 63 64 65 66 61 HC34 17 1 0.100 1 3 62 63 64 62 C35 11 12 -0.100 0 5 63 64 65 66 67 63 HC35 17 1 0.100 1 3 64 65 66 64 C36 11 12 -0.100 0 3 65 66 67 65 HC36 17 1 0.100 1 2 66 67 66 C37 11 12 -0.100 0 1 67 67 HC37 17 1 0.100 1 0 68 C38 11 12 -0.100 0 1 69 69 HC38 17 1 0.100 1 0 # bonds # NB 73 # IB JB MCB IB JB MCB IB JB MCB 1 2 1 2 3 17 3 4 26 4 5 26 4 8 26 4 11 26 5 6 17 6 7 1 8 9 17 9 10 1 11 12 17 12 13 12 13 14 15 13 68 15 14 15 3 14 16 15 16 17 15 16 53 15 17 18 15 17 49 15 18 19 3 18 20 15 20 21 12 20 33 15 21 22 17

22 23 26 23 24 26 23 27 26 23 30 26 24 25 17 25 26 1 27 28 17 28 29 1 30 31 17 31 32 1 33 34 3 33 35 15 35 36 15 35 49 15 36 37 50 37 38 15 38 39 15 38 47 15 39 40 3 39 41 15 41 42 3 41 43 15 43 44 3 43 45 15 45 46 3

45 47 15 47 48 3 49 50 51 50 51 23 50 52 23 50 53 51 53 54 15 54 55 15 54 68 15 55 56 50 56 57 15 57 58 15 57 66 15 58 59 3 58 60 15 60 61 3 60 62 15 62 63 3 62 64 15 64 65 3 64 66 15 66 67 3 68 69 3

# bond angles # NBA 110 # IB JB KB MCB IB JB KB MCB 1 2 3 11 2 3 4 12 3 4 8 12 3 4 5 12 3 4 11 12 8 4 11 12 5 4 11 12 5 4 8 12 4 5 6 12 5 6 7 11 4 8 9 12

8 9 10 11 4 11 12 12 11 12 13 11 12 13 14 26 12 13 68 26 14 13 68 26 13 14 15 24 13 14 16 26 15 14 16 24 14 16 17 38 14 16 53 26

114

17 16 53 6 16 17 18 38 16 17 49 6 18 17 49 26 17 18 19 24 17 18 20 26 19 18 20 24 18 20 21 26 18 20 33 26 21 20 33 26 20 21 22 11 21 22 23 12 22 23 24 12 24 23 27 12 27 23 30 12 22 23 30 12 24 23 30 12 22 23 27 12 23 24 25 12 24 25 26 11 23 27 28 12 27 28 29 11 23 30 31 12 30 31 32 11 20 33 34 24 20 33 35 26 34 33 35 24 33 35 36 26 33 35 49 26 36 35 49 26 35 36 37 48 36 37 38 48 37 38 39 26 37 38 47 26 39 38 47 26 38 39 40 24 38 39 41 26 40 39 41 24 39 41 42 24 39 41 43 26 42 41 43 24 41 43 44 24 41 43 45 26 44 43 45 24

43 45 46 24 43 45 47 26 46 45 47 24 45 47 48 24 45 47 38 26 38 47 48 24 35 49 50 53 35 49 17 26 17 49 50 52 49 50 51 50 51 50 52 51 52 50 53 50 49 50 53 49 49 50 52 50 51 50 53 50 50 53 54 53 50 53 16 52 16 53 54 26 53 54 55 26 53 54 68 26 55 54 68 26 54 55 56 48 55 56 57 48 56 57 58 26 56 57 66 26 58 57 66 26 57 58 59 24 57 58 60 26 59 58 60 24 58 60 61 24 58 60 62 26 61 60 62 24 60 62 63 24 60 62 64 26 63 62 64 24 62 64 65 24 62 64 66 26 65 64 66 24 64 66 57 26 64 66 67 24 57 66 67 24 54 68 69 24 54 68 13 26 13 68 69 24

# improper dihedrals # NIDA 53 # IB JB KB LB MCB IB JB KB LB MCB 13 12 14 68 1 14 13 15 16 1 16 14 17 53 1 53 16 50 54 1 54 53 55 68 1 68 54 69 13 1 13 14 16 53 1 14 16 53 54 1 16 53 54 68 1 53 54 68 13 1 54 68 13 14 1

68 13 14 16 1 17 16 18 49 1 18 17 19 20 1 20 18 21 33 1 33 20 34 35 1 35 33 36 49 1 49 35 50 17 1 17 18 20 33 1 18 20 33 35 1 20 33 35 49 1 33 35 49 17 1

115

35 49 17 18 1 49 17 18 20 1 16 17 49 50 1 17 49 50 53 1 49 50 53 16 1 50 53 16 17 1 53 16 17 49 1 38 37 39 47 1 39 38 40 41 1 41 39 42 43 1 43 41 44 45 1 45 43 46 47 1 47 45 48 38 1 38 39 41 43 1 39 41 43 45 1 41 43 45 47 1

43 45 47 38 1 45 47 38 39 1 47 38 39 41 1 57 56 58 66 1 58 57 59 60 1 60 58 61 62 1 62 60 63 64 1 64 62 65 66 1 66 64 67 57 1 57 58 60 62 1 58 60 62 64 1 60 62 64 66 1 62 64 66 57 1 64 66 57 58 1 66 57 58 60 1

# dihedrals 20 # NDA # IB JB KB LB MCB

IB JB KB LB MCB

1 2 3 4 12 2 3 4 11 20 4 5 6 7 12 8 4 5 6 20 4 8 9 10 12 5 4 8 9 20 3 4 11 12 20 4 11 12 13 12 11 12 13 14 2 18 20 21 22 2

20 21 22 23 12 22 23 24 25 20 21 22 23 24 20 23 24 25 26 12 22 23 27 28 20 23 27 28 29 12 22 23 30 31 20 23 30 31 32 12 51 50 53 16 42 51 50 49 17 42

END

116

6. Summary

The work presented here describes the synthesis and binding studies of bicyclic guanidinium

receptors specially designed to act in polar solvents like water, DMSO and methanol. The

receptor 51 was designed to contain the bicyclic guanidinium group as the main binding motif

flanked by hydrophobic substituents on both sides to guard the anchor group from potential

solvation in polar solvents. It was constructed with water solubilising hydroxyl groups in the para-

position of the phenyl rings which will orient away from the guanidinium motif and can not

interfere in the binding process.

The host 34 was prepared with similar substitution pattern, but with less steric hindrance at the

guanidinium anchor group in the form of flexible hydroxypropyl chains. Another water soluble

receptor 28 was also studied on account of its established binding affinity for the oxoanions in

polar aprotic solvents like acetonitrile.

The water soluble aromatic hydrophilic phosphinate guest 37 was also designed specially for

this study. The phosphinate guest 37 was constructed based on the similar concept of protecting

the binding motif (phosphinate anion) from the potential solvation in polar solvents using

hydrophobic groups without compromising the affinity. It was incorporated with wall-like

structures (phenyl acetylene groups) adjacent to the binding site to create a hydrophobic shell

around the phosphinate anion and polyhydroxy groups to enhance the overall solubility. The

more exposed guest 119 was chosen for the trend analysis because of its similarity with the

phosphinate 37.

117

The complexation between hosts 51, 34, 28 and phosphinate guests 37 and 119 was probed

using isothermal titration calorimetry (ITC) in polar protic solvent water and polar aprotic solvent

DMSO. The ITC is a very precise method, which in a single experiment gives the complete

assessment of the energetic parameters like enthalpy, entropy and free energy of association.

The titration plot also furnishes the stoichiometry of the complex.

The host 51 and the guest 37 were precipitated during the titration at the experimental

concentrations in water. However, the host-guest ion pair 34·37 does not show any

complexation in water. Interestingly, the allyl guanidinium host 28 binds very strongly with the

aromatic phosphinate 37 in water with a huge association constant of about 70000 M-1. The

binding was strongly driven by entropy and the stoichiometry was found to be 1:1 for the ion-pair

28·37 (fig. 6.1).

Figure 6.1: A conformation of the complex 28·37 in water

However, the same host 28 does not bind at all to the control phosphinate 119 in water.

118

In DMSO, the host 51 binds to the guest 37 with a modest association constant of 542 M-1 and

the complexation was exothermic. The stoichiometry was rather low which indicates some

aggregates might be forming during the binding process. Surprisingly, host 51 complexes guest

119 with a five-fold higher binding constant (Ka =2953 M-1) indicating the easy access to the

highly hindered guanidinium core cavity. The host 34 binds to the phosphinate 37 with a lower

binding constant (Ka = 1950 M-1) than with the host 28 and gave large unfavourable entropy

value. The host 28 with least steric hindrance at the binding site was found to be the better

complementary structure to the guest 37 as it gives the highest binding constant (Ka = 6184 M-1)

in dimethyl sulphoxide. The asociations between the hosts (28, 34 and 51) with the phosphinate

guest 37 in DMSO were enthalpically driven except with the guest 119 where it was entropically

driven.

The results from the titrations inferred the protection of the solvation shell around both the host

and the guest successfully improved the binding of the phosphinate in polar solvents like water

and dimethyl sulphoxide. The host 51 needs to be incorporated with more hydroxyl groups to

further enhance its water solubility.

MD simulation studies were performed on the ensemble of hosts 28, 34 and 51 with the

phosphinate 37 in various solvents like H2O, DMSO, MeOH and CHCl3 at 300 K (also at 400 K in

case of the host 51) using the GROMOS’96 software. The non-bonded energy analysis of the

simulations indicated the weakening of the electrostatic interactions due to the solvation of the

binding partners in polar solvents which also led to the structural fuzziness in the complexation.

The H-bond analysis showed the increase in polarity of the solvent leads to the decrease in

number of H-bonds participating in the complexation process.

119

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Important Compounds