Dynamic aspects of phosphorus in four- and five-coordinated compounds : DNA as an example

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Dynamic aspects of phosphorus in four- and five-coordinated

compounds : DNA as an example

Citation for published version (APA):

van Lier, J. J. C. (1983). Dynamic aspects of phosphorus in four- and five-coordinated compounds : DNA as an example. Technische Hogeschool Eindhoven. https://doi.org/10.6100/IR38658

DOI:

10.6100/IR38658

Document status and date: Published: 01/01/1983 Document Version:

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DYNAMIC ASPECTSOF PHOSPHORUS IN

FOOI·"AND FIVE-COORDINATED COMPOUNDS

DNA

as an

example

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DYNAMIC ASPECfS OF PHOSPHORUS IN

FOUR- AND FIVE-COORDINATED COMPOUNDS

DNA as an example

PROEFSCHRIFT

TER VERKRIJGING VAN DE GRAAD VAN DOCrOR IN DE TECHNISCHE WETENSCHAPPEN AAN DE TECHNISCHE HOGESCHOOL EINDHOVEN, OP GEZAG VAN DE RECfOR MAGNIFICUS, PROF. DR. S. T. M. ACI<El{MANS, VOOR EEN COMMISSIE AANGEWEZEN DOOR HET COLLEGE VAN, DEKANEN IN HET OPENBAAR TE VERDEDIGEN OP

DINSDAG 25 OKTOBER 1983 TE 16.00 UUR DOOR

JOHANNES JACOBUS CORNELIA VAN LIER

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DIT PROEFSCHRIFT IS GOEDGEKEURD DOOR

DE PROMOTOREN

PROF. DR. H.M. BUCK

EN

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"Wijs is hij/zij~ die van aZZen ?Jeet te tel"en"

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-Chapter I

Chapter 11

s-.z

transition in aZternating d(A-T)n potymers

V.3 MNDO caleuZations

V. 4 Computational data and stereo model studies

V. 5 Discussion

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Chapter VI

Appendix

SuiD.ID.ary

SaiD.envatting

Tetraoxaspirophosphoranes with a six- 116 -membered ring as model compounds for_ the hydrolysis of ribonucleic acids by RNase A

VI.1 Introduation

VI.2 NMR speatrosaopia study of the formation of tetrao~aspirophos

phoranes aontaining a si~

-membered ring and a P-H bond VI.3 Base-aataLyzed ring aLosure as

a modeL foP the aation of RNase A VI.4 Disaussion

VI.S E~perimentaL

Referenaes and Notes

143

147

150

CurriculuiD. vitae 153

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CHAPTER I

General introduetion

I.1 Reaent developments in organophosphorus chemistry.

The research i~ phosphorus chemistry has developed rapidly inthelast decade1_, _{A number of systems reveal the}

unique possibilities of phospporus for coordination in diffe-rent valenee states2

• Recent progress has stimulated research

o~ the structural and dynamic stereochemistry of organophos-phorus compounds, particularly in the field of biomolecules.

In this respect, the discovery of stabie penta-coordinated (P(V)) organophosphorus derivatives3 _{has been of paramount}

importance. Westheimer's~-6 _{studies on the hydrolysis of} five--membered cyclic phosph(on)ates have greatly advanced the understanding of the mechanistic aspects of phosphorylation reactions (i.e. substitution at phosphorus). The enormous rate enhancement of the phosphorylation, as observed.for these compounds, was elegantly explained by assuming P(V) trigonal bipyramidal (TBP) intermediates with the capability of ligand reorganizations (pseudorotation, vide infra). Interestingly, group transfer reactions (i.e. substitution at a phosphorus ligand) may also proceed via P(V) TBP inter-mediates. Voncken, Castelijns, van Aken and Buck1_-9 _clearly established that, in this case, the selective step inheres a nucleÓphilic attack on one of the pseudo-equatorial posi-tions in the TBP configuration.

P(V) TBP intermediate structures certainly play an important role in bicmolecules containing phosphorus. A well-known example of a biochemica! process in which this P(V) intermediacy has been accepted, is the hydrolysis of ribonucleic acids, catalyzed by bovine pancreatie ribonuclease

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-WHis 119 ROH =

~··

0 ' \

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(RNase A). This hydrolysis occurs via anchimeric assistance of the 2'-0H group of the ribose ring10

- 12, which is in

close proximity to histidine 12 and the 3'-phosphate group. In the first transphosphorylation step, histidine 12 abstracts the 2'-0H proton thus facilitating apical attack on the

phosphate group and formation·of a P(V) TBP intermediate (Fig. 1,1), This intermediate is stabilized by hydrogen bonding between protonated lysine 41 and the equatorial anionic oxygen ligands. Activation of the leaving 5'-nucleo-tide by protonated histidine 119 generates the 2',3'-cyclic phosphate13

'14, which, aft~r a similarly catalyzed hydrolysis,

gives rise to the products (see Fig. 1.1).

Like RNase A, staphyZococcal nuclease hydrolyzes nucleic acids but: in contrast, operates on both RNA and DNA sub-strates15. Of the many known nucleases this enzyme is perhaps. the best characterized. The crystal structure has been re-solved for the native enzyme16 _{as wellas for the}

nuclease--thymidine-3',5'-diphosphate (pdTp)-calcium ion complex17

(resolution, 1.5 ~). The X-ray data16_•17 _{and additional NMR}

studies18 • 19 implicate two tyrosines (85 and 113) at the active site in direct interaction with the nucleotide in-hibitor, pdTp 17 • Moreover, in this complex 17 the Ca2+ ion is coordinated by asparagine 21, asparagine 40 and glutamine 43 and located at 4.7 ± 0.2 ~ distance from the phosphorus atom. The X-ray data17 _{suggest that an intervening water or}

hydroxyl ligand can be accommodated as a ligand of the ca 2+

ion. Subsequent nucleophilic attack of this ligand at phos-phorus generates a P(V) TBP intermediate20

•21 (Fig. !.2).

This intermediate structure is stabilized by hydrogen-bonding from the protonated arginines 35 and 87 to the anionic oxygen ligands. Proton transfer from arginine 87 to the 0(5') atom leads tp apical departure of the 5'-nucleotide. Interestingly, hydrolysis of the model substrate deoxythymidine-3'-phosphate--5'-p-nitrophenylphosphate, catalyzed by this enzyme, gave exclusive formation of p-nitrophenyl phosphate, whereas nonenzymatic base-catalyzed hydrolysis of the same substrate causes displacement of nitrophenoxide, a much better leaving group than the 5'-oxyanion of deoxythymidine22_• _{This is a}

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0 I " I , _,b

-go

-;)>o. ___

r,~,

Tyr' 113 5 --- • :.. - - -H + Arg --~

C

_)p~3~

o

'"

s1

₀

+

-#\

0 Arg H ----0 b 35 0-H _, Asp;-1 -- - - 1 ... *'+ ... '·Ca' / ' - '-GI u~-

3

----r Asp' 40

Fig. I.2 Proposed cataZytic mechanism for the action of staphyZoaoccaZ nucZease on DNA substrates.

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strong argument in favour of the proposed mechanism.

!.2 GeneraZ properties of penta-aoordinated phosphorus aompounds.

An important factor governing the stereochemistry of P(V) compounds, in contrast to P(IV) compounds, is that the distribution of the ligands around the central phosphorus atom cannot be spherically symmetrical, i.e., the ligands in P(V) compounds are not equivalent.23 _{As shown by X-ray}

analysis24

- 26, usually two types of structures of very similar

energy23 _{are considered, the TBP and the square pyramid}

(SP). The TBP and SP geometries are characterized by non-equivalent bonding. In the TBP there are three equatorial and two apical substituents, whereas in the SP one apical and four basal ligands are encountered (Fig. I.3). In the

a a

apical ligand e,

_I

_b~.

_{I ..}

_b a:

'p-e

_{b_....:p~b} b: bas al ligand

e"J

e: equator i al ligand

a

TBP SP

Fig. I.J Different geometrie a of P(V) aompounds. TBP configu;ation, the apical honds are langer and weaker

than the equatorial bonds27

• Apical sites are preferred by

electron-withdrawing ligands, whereas electron-donating substituents !end to occupy equatoral positions28_• _This

'polarity rule is basedon experimental data29_•30 _{and is}

supported by semi-empirica! calculations31

' 32• The polarity

rule can be explained on the basis of a hybridization of the Pz and dz2 orbitals in the TBP to account for the apical honds, combined with three sp 2 orbitals in the equatorial plane33

• Although this classica! description of the phosphorus

hybridization in a TBP structure offers a good explanation for the observed selectivity, recent ESR studies of phos-phoranyl radicals performed by Hamerlinck~ Schipper and

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honds. This means that the discriminatien between the equato-rial and apical ligands is determined by a smal! overbalance of s-character in the equatorial position, thus still

suppor-ting the physical organic properties of TBP phosphoranes. Furthermore, it has been found that smal! rings are easily accommodated if they.span an apical and an equatorial posi-tion. This strain rute6 _{is a result of the 90° angle}

be-tween apical and equatorial honds in the TBP and is demon-strated by the fact that in most of the stabie phosphoranes the phosphorus atom is part of a cyclic system. The diffuse dzz orbital accounts for the greater apical bond length. Apical sites are preferentially occupied by electron-with-drawing substituents, whereas the equatorial sp2 orbitals demand for electrens from the corresponding ligands. In addition, equatorial ligands are more capable to form d~-pn

bonds to phosphorus (backdonation)21 •

One of the consequences of the differences in bond strength in a TBP is that leaving groups preferentially depart from an apical position6

- 31 • Conversely, as required

by the principle of microscopie reversibility6

, nucleophilic

attack on tetra~coordinated phosphorus forms TBP interme-diates and/or transition states in which the extra ligand occupies an apical position. Interestingly, the TBP confi-guration is stereochemically non-rigid23

, This was established

first for PF 5 by 19F-NMR showing one fluorine resonance37 ,

while other investigations38 _{indicate that the fluorines}

exchange their positions fast on the NMR time scale. A

mechanism which accounts for this permutational isomerization is the Berry pseudorotation (BPR)39

• In this process two

equatorial and both apical ligands change place via an intermediate SP configuration, the remaining ligand being the pivot (Fig. 1.4). The energy harrier for BPR is conside-rably increased upon introduetion of ligands with different electron-withdrawing character or in case of small rings31_•

Therefore, an alternative process which accounts for the same permutation is the Turnstite rotation (TR) which may

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TBP SP TBP Fig. I.4 The BePry pseudorotation proaess.

be favoured in (bi)cyclic phosphoranes40

' 41 (Fig. I.S).

From X-ray diffraction studies it has been established that in the solid state the structure of P(V) compounds is distor-ted more or less from anideal TBP toward an SP geometry42

,

and that these distortions closely follow the local

c

_2v een-straint of the Berry intramolecular exchange coordinate. These data suggest that opening of an equatorial angle in the TBP is associated with an approximately equal degree of closing of the axial angle43

•

top pair

bottam

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1.3 Different configurations of DNA.

The detailed knowledge with respect to the structural and dynamic stereochemistry of organophosphorus compounds (vide supra) can also be applied to the phosphate groups in the helix backbene of the DNA molecule. It is well known that the genetic information is linearly encrypted in the sequence of bases of nucleic acids which are the genetic material of all living organisms. The means by which the

information is contained and transmitted in these molecules is a subject of extensive research which has been greatly advanced by the application of recombinant DNA techniques44

•

Another major souree of information is the elucidation of the three-dimensional structures of nucleic acids by means of high-resalution single crystal X-ray diffraction studies45

•

The main structural elements of nucleic acids are i) a five-membered sugar ring which is a ribose for RNA

and a 2'-deoxyribose for DNA.

ii) the heterocyclic bases adenine (A). guanine (G).

cytosine (C), thymine (T) and. in case of RNA, uracil (U) replacing T. The bases are bound to C(l') of the

sugar ring in the ~-configuration.

iii) the 3'-5'-phosphodiester linkage joining the individual nucZeosides (i.e. the combination of sugar and base). Double-stranded nucleic acids are formed by Watson-Crick46

hydrogen bonding between the complementary G and C or A and T(U) bases in both strands of the structure. The complemen-tary strands are anti-paraZZeZ, i.e. have opposite S'-.3' direction. The numbering scheme47 _{for the various structural}

units which bas been used in this work is given in Fig. !.6. DNA usually crystallizes as a right-handed double helix with anti conformation of the base residues (B-DNA)48_• _Other

right-handed helices have been classified and designated as A-DNA49

' 50, C-DNA51 and D-DNA52' 53•

Double-stranded DNA eligomers and polymers containing a strictly alternating CG sequence display unusual confor-mational properties. Using circulair dichroism (CD) studies, PohZ and Jovin54 _{demonstrated that poly d(G-C) undergoes a}

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thymine

cytosine

Fig. I.6 Structure of a DNA fragment containing the four common bases in DNA~ tagether with the numbering echemes ueed for the pyrimidine (C~ T) and purine

(A~ G) bases and the 2'-deo~yriboses. In the cor-reeponding RNA oligomer# the 2'-deo~yriboee wiZZ be replaced by ribose and T wiZZ be substituted by

u.

salt-induced conformational change which is characterized by a speetral inversion in high-salt solution. This transi-tion appeared to be reversible, Interestingly, 1H- and

31 P-NMR studies revealed that the d(G-C)_{8 duplex in high-salt} solution contained two different types of nucleotide and phosphate group conformations including glycosidic dihedral angles which are different from B-DNA 5 5

' 56• Recently, Wang

et al, 51 _{reported a novel} _{Zeft-handed Z-DNA} _{helix with the}

sequence d(C-G) 3 . The dG residues in the structure have an C(3')-endo (3E) pucker and a syn conformation in contrast

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to the C(2')-endo (2E) pucker and anti conformation which is observed for B-DNA (Fig. I.7). A similar left-handed duplex structure bas been observed in d(G-C) 358 _and

d(G-m5c)

359 single crystals. In solution, the Raman spectra

\

0 0

J

o-o~P~

\

0 NHz

er

Z-DNA Fig. I.? Geometrie differenaes between B-DNA and Z-DNA for

a 5'-dpG fragment.

of the high- and low-salt forms of poly d(G-C) differ from each other60

• Recent Raman studies show that the spectrum

of crystalline d(G-C) 3 is essentially the same as that of poly d(G-C) in high-salt solution61

• Thus the conformations

are identical. This means that the salt-induced conversion as observed for the poly d(G-C) duplex can be interpreted as a transition from a right-handed helix into its left--handed isomer (B-.z transition). Furthermore, X-ray diffrac-tion data on Z-DNA single crystals57

- 59 revealed the unusual

y- and yt conformations around the C(4')-C(S') bond. A yt conformation bas also been established for model

organophosphorus substrates in the active site of RNase A10_- 12

and of staphylococcal nuclease17

• Interestingly, for RNase A,

intermediate P(V) TBP structures are known to be involved in the hydrolysis of ribonucleic acids, whereas for staphylo-coccal nuclease similar intermediates have been proposed

(c.f. Chapter I.l). These results seem to parallel the

calculated decreased preferenee of the y+ conformer in favour of the y- and yt conformers upon P(IV)-P(V) TBP activatien of the phosphate groups in the helix backbone of the DNA structure (c.f. Chapter III).

Characteristic differences between right-handed and left-handed helices are given in Table I.l. Additional stu-dies revealed that the B-.z transition is generally observed

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Table I.1 St~ucturaZ pa~amete~s in DNA.

Helix sense Right-handed Left-handed

Designation A B

c

D

z

Sugar pucker 3Ea ZE ZE ZE ZE (dC); 3E (dG)

Glycosyl angle anti anti anti anti anti (dC); syn (dG)

Twist per base 32.7° 36° 38.6° 45° -60°/2b

pair

Bases per turn 11 10 9.33 8 12

Rise per base 2.6 3.4 3.3 3.0 3.7

pair, Ä

Base tilt0 19° -60 -80 -16° -70

a 3E, C(3')-endo; 2E, C(2')-endo.

bAs a result of the alternating sugar pucker the repetitive unit is a dinucleotide.

0

Deviation from a plane perpendicular to the helix axis. in DNA structures with an alternating purine-pyrimidine sequence such as poly d(A-C). poly d(G-T)62 _and

poly d(G-m 5

c).

poly d(G-m 5C)63

• Moreover, Z-DNA

struc-tures have been detected in vivo6_~'65_• _{Very recently,}

Singleton et aZ. 66 _{showed that a Z-DNA is induced by}

super-coiling (i.e. higher order structure of DNA) under physio-logical conditions. Therefore, the mechanistic aspects of the B-.Z transition deserve attention.

I.4 Biologiaal p~ope~tiea of DNA.

DNA has two major and discrete functions. One is to carry the genetic information that brings about the specific phenotype of the cel!. The other function of DNA is its own replication67_• _{In DNA} _~epliaation,_{a comple,mentary copy}

of each strand is first made. Since each new strand remains bound to one of the old strands the net result is two double helices, each identical with the one originally present. Thus, for duplicating the genotype of the cell, DNA serves as a template for converting one chromosome into two identi-cal chromosomes. This 'semi-conservative1 _{mode of replicatien}

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of DNA was originally predicted by Watson and Crick~6_•

DNA, whicJl resides in the nucleus of the cell, does not determine the amino acid sequence of a protein directly. Instead, the base sequence of DNA serves as a template for the synthesis of a single strand of messenger RNA (mRNA). This transcription process produces mRNA with a base

sequence complementary to the transcribed DNA strand, with uracil replacing thymine. Very recently, Wang et al.68

repor-ted the detailed three-dimensional structure of a short DNA-RNA hybrid helix joined to double helical DNA and showed that this fragment adopts a helix close to 11-fold DNA (A-DNA). Once mRNA has been sythesized, it passes out of the nucleus into the cytoplasm to the ribesomes where translation of the base sequence into an amino acid sequence of a protein is accomplished.

I.S Scope of this thesis.

As evidence in favour of the important dynamic role of phosphorus in the DNA molecule is increasing10

, it is useful

to apply the knowledge of phosphorane intermediates to the phosphate groups in the helix backbone of DNA in order to obtain a better understanding of the salt-induced B-.z transi-tion. Chapter II describes a study of the interaction of model organophosph(on)ates with lithium salts in solution using multi-nuclear NMR. The 35

c1-

and 81Br NMR results can be explained by assuming fast equilibria between lithium balides in the complexed (with phosph(on)ates) and the free form. The 7Li NMR chemica! shift and line-broadening data reveal complexation of the phosphoryl oxygen atom by the Li+ ion, preferentially in 1:1 mol ratio salt/phosphate. Furthermore, kinetic studies on the combined phosphorylation and group transfer (de-alkylation) in the salt/phosphate/ methanol system reveal that the phosphorylation reaction is retarded in the presence of lithium halides. The magnitude of the effect is related to the ionic radius of the halide anion as well as to the solvation properties of this anion.

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The data therefore provide evidence for the proximity of the halide anion to the phosphorus atom. Low-temperature 31P NMR measurements combined with CND0-2 calculations reveal that the close-ion pair structure is the NMR observed configuration, whereas short-lived P(V) structures are encountered as inter-mediates in the reactions. Based on these data and on

three--dimensional structural features of B and Z forms of DNA, a detailed model description of the salt-induced B-.z transition

in DNA with alternating purine-pyrimidine sequences is given in Chapter III. The performed CND0-2 and MNDO quantum-cnemical calculations support the suggested selective role of P(V) TBP intermediates within 5'-dpC structural units in DNA as an initial inducer of the B-.Z transition. Moreover, the calcu-lations reveal a significant difference in selectivity between purine and pyrimidine bases in the structure which offers a good rationalization for the observed features in the process.

Chapter IV provides evidence for the considerable impact of specific methylation of C.G base pairs on the relative stability of B and Z isomers of DNA based on calculations using methylated tetrahydrofuryl model systems. The obtained data are related to known experimental details. The calcula-tions reveal that an important stabilization of the Z confor-mer is obtained upon specific methylation. The possible biologica! significanee of methylated cytosine residues in

the genomic (supercoiled) DNA is discussed in relation to the obtained computational results and known experimental data.

The molecular aspects of methylated adenine in DNA, specifically focused on an altered anti-syn equilibrium due to selective methylation, are discussed in Chapter V, based on calculations on tetrahydrofuryl model structures and experimental data.

In Chapter VI, model compounds derived from tetraoxa-spirophosphoranes are used to study the base-catalyzed ring opening and ring ciosure process. This behaviour mimics the possible role of the enzymatic sites in the active cleft of RNase A during hydrolysis of ribonucleic acids.

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HefePenoes and Notes.

1. For up-to-date reviews on the subject, see the series "Organophosphorus Chemistry" (Specialist Periodical Reports), S. Trippett, ed., The Chemica! Society, London. 2. The following abbreviations are used: P(IV)

tetra--coordinated phosphorus; P(V) pentatetra--coordinated phosphorus; TBP trigonal bipyramid(al); SP square pyramid(al);

BPR Berry pseudorotation; TR Turnstile rotation. 3. F. Ramirez, R.B. Mitra and N.B. Dessai, J. Am. Chem.

Soc., 1960, 82, 2651.

4. J. Kumamoto, J.R. Cox and F.H. Westheimer, J. Am. Chem. Soc., 1956, ?8, 4858,

5. A. Eberhard and F.H. Westheimer, J. Am. Chem. Soc., 1965, 8?, 253.

6. F.H. Westheimer, Acc. Chem. Res., 1968, 1, 70.

7. W.G. Voncken, Ph. D. Thesis, Eindhoven University of Technology, 1976.

8. A.M.C.F. Castelijns, Ph. D. Thesis, Eindhoven University of Technology, 1979.

9. D. van Aken, Ph. D. Thesis, Eindhoven University of Technology, 1981.

10. R.R. Holmes, "Penta-coordinated Phosphorus" (ACS Monograph 176), Vol.II, American Chemica! Society, Washington D.C., 1980, 180.

11. F.M. Richards and H.W. Wyckoff, "The Enzymes", Vol.IV, P.D. Boyer, ed., Academie Press, New York, 1971.

12. D.G. Gorenstein, A.M. Wyrwycz and J, Bode, J. Am. Chem. Soc., 1976, 98, 2308.

13. R.R. Holmes, J.A. Deiters and J.C. Galluci, J. Am. Chem. Soc., 1978, 100, 7393.

14. C.A. Deakyne and L.C. Allen, J. Am. Chem. Soc., 1979,

101, 3951.

15. C.B. Anfinsen, P. Cuatrecasas and H. Taniuchi, "The Enzymes", P.D. Boyer, Ed., 3rd ed., Vol. IV, Academie Press, New York, 1971, 177.

16, F.A. Cotton and E.E. Hazen, Jr., "The Enzymesn, P.D. Boyer, Ed., 3rd ed., Vol. IV, Academie Press, New York,

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1971, 153.

17. F.A. Cotton, E.E. Hazen, Jr. and M.J. Legg, Proc. Natl, Acad. Sci. u.s.A., 1979, 76, 2551 and raferences cited therein.

18. J.L. Markley and 0. Jardetzky, J. Mol. Biol., 1970, 50,

223.

19. G.C.K. Roberts and 0. Jardetzky, Adv. Prot. Chem., 1970,

24, 44 7.

20. A.S. Mildvan, Annu. Rev. Biochem., 1974, 43, 357.

21. A.S. Mildvan and C.M. Grisham, Struct. Bonding(Berlin),

1974, 20, 1.

22. B.M. Dunn, C. DiBello and C.B. Anfinsen, J. Biol. Chem.,

1973, 248, 4769.

23. R. Luckenbach, "Dynamic Stereochemistry of Penta-coordi-nated Phosphorus and Related Elements", G. Thieme, Stuttgart, 1973.

24. E.L. Muetterties and R.A. Schunn, Quart. Rev. Chem. Soc.,

1966, 20, 245.

25. T.E. Clark, R.O, Day and R.R. Holmes, Inorg. Chem., 1979,

18, 1653.

26, T.E. Clark, R.O. Day and R.R. Holmes, Inorg. Chem., 1979,

18, 1668.

27. F. Ramirez and I. Ugi, "Advances in Physical Organic Chemistry", Vol. 9, V. Gold, ed., Academie Press, London,

19 71 •

28. P. Gillespie, P. Hoffmann, H. Klusacek, D. Marguarding, S. Pfohl, F. Ramirez, E.A. Tsolis and I. Ugi, Angew. Chem.,

1971,83,691.

29. E.L. Muetterties, W, Mahler and R. Schmutzler, Inorg. Chem., 1963, 2, 613,

30. E.L. Muetterties, K.J. Packer and R. Schmutzler, Inorg. Chem., 1964, 3, 1298.

31. D. Marguarding, F. Ramirez, I. Ugi and P. Gillespie, Angew. Chem., 1973, 85, 99.

32. F. Keil and W. Kutzelnigg, J. Am. Chem. Soc., 1975, 97,

3623.

33. R.F. Hudson and M. Green, Angew. Chem., 1963, 75, 47.

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of Technology, 1982.

35. J.H.H. Hamerlinck, P. Schipper and H.M. Buck, J. Org. ehem., 1983, 48, 306.

I

36. J.H.H. Hamerlinck, P. Schipper and H.M. Buck, J. Am. ehem. Soc., 1983, 105, 385.

37. H.S. Gutowski, D.M. Meeall and e.P. Slichter, J. Chem. Phys., 1953, 21, 279.

38. H.S. Gutowski and A.D. Liehr, J. ehem. Phys., 1953, 20,

1652.

39. R.S. Berry, J. ehem. Phys., 1960, 32, 933.

40. F. Ramirez, S. Pfohl, E.A. Tsolis, J.F. Pilot, e.P. Smith, I. Ugi, D. Marguarding, P. Gillespie and P. Hoffmann, Phosphorus, 1971, 1, 1.

41. I. Ugi, D, Marguarding, H. Klusacek, P. Gillespie and F. Ramirez, Acc. ehem. Res., 1971, 4, 288.

42. R.R. Holmes, Acc. ehem. Res. 1979, 12, 257.

43. H.-B. Bürgi and J.D. Dunitz, Acc. ehem. Res., 1983, 16,

153.

44. M. Singer, "Genetic Engineering", A. Hollaender and J. Setlow, Eds., Vol. I, Plenum Press, New York, 1979. 45. N.e. Seeman, "Nucleic Acid Geometry and Dynamics", R.H.

Sarma, ed., Pergamon Press, New York, 1980, 47.

46. J.D. Watson and F.H.e. erick, Nature(Lond.), 1953, 171,

737.

47. Abbreviations and symbols follow IUPAC-IUB Recommenda-tions, see Eur. J. Biochem., 1983, 131, 9.

48. H.R. Drew, R,M, Wing, T. Takano, C. Broka,

s.

Tanaka, K. Itakura and R.E. Dickerson, Proc. Natl. Acad. Sci.

u.s.A.,

1981, 78, 2179.

49. B.N. Conner, T. Takano, S. Tanaka, K. Itakura and R.E. Dickerson, Nature(Lond.), 1982, 295, 294.

50. A.H.-J. Wang, S. Fujii, J.H. van Boom and A. Rich, Proc. Natl. Acad. Sci. U.S.A., 1982, 79, 3968,

51. s. Arnott, R. Chandrasekaran, D.W.J. Hukins, R.s.c. Smith and L. Watts, J. Mol. Biol., 1974, 88, 523,

52. S. Arnott and E. Selsing, J. Mol. Biol., 1975, 98, 265. 53. A. Mahendrasingam, N.J. Rhodes, D.C. Goodwin, e. Nave,

(28)

Nature(Lond.), 1983, SOl, 535,

54. F.M. Pohl and T.M. Jovin, J. Mol. Biol. 1972, 67, 375. 55. D.J. Patel, L.L. Canuel and F.M. Pohl, Proc. Natl. Acad.

Sci. u.s.A., 1979, 76, 2508.

56. D.J. Patel, "Stereodynamics of Molecular Systems", R.H. Sarma, ed., Pergamon Press, New York, 1979, 397.

57. A.H.-J. Wang, G.J. Quigley, F.J. Kolpak, J.L. Crawford, J.H. van Boom, G. van derMareland A. Rich, Nature(Lond,), 1979, 282, 680.

58. H. Drew, T. Takano, S. Tanaka, K. Itakura and R. Dickerson, Nature(Lond.), 1980, 286, 567.

59. A.H.-J. Wang, S. Fujii, J.H. van Boom and A. Rich, Cold Spring Harhor Symp. Quant. Biol., 1982, 47, 5.

60. F.M. Pohl, A. Renade and M. Stockburger, Biochim. Biophys. Acta, 1973, SS6, 85.

61, T.J. Thamann, R.C. Lord, A.H.-J. Wang and A. Rich, Nucleic Acids Res., 1981, 9, 5443.

62,

s.

Arnott, R. Chandrasekaran, D.L. Birdsall, A.G.W. Leslie and R.L. Ratliff, Nature(Lond.), 1980, 28S, 743.

63. J. Nickol, M. Behe and G. Felsenfeld, Proc. Natl. Acad. Sci. u.s.A., 1982, ?9, 1771.

64, A. Nordheim, M.L. Pardue, E. M. Lafer, A. Möller, B.D. Stollar and A. Rich, Nature(Lond.), 1981, 294, 417.

65. H. Hamada and T. Kakunaga, Nature(Lond.), 1982, 298, 396. 66. C.K. Singleton, J. Klysik, S.M. ~tirdivant and R.D.

Wells, Nature(Lond.), 1982, 299, 312.

67. A. Kornberg, "DNA Replication", A.C. Bartlett and P. Brewer, eds., W.H. Preeman and Company, San Francisco, 1980.

68. A.H.-J. Wang, s. Fujii, J.H. van Boom, G.A. van der Marel, S.A.A. van Boeckel and A. Rich, Nature(Lond.), 1982, 299, 601.

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CHAPTER 11

LlthiuiD halide and lithluiD perchlorate binding to phosphates. A IDulti -nuclear IDagnetic resonance spectroscopie study

11.1 Introduation.

Growing recognition of the importance of biologically relevant metal ion interactions with nucleic acids and nucleotides has stimulated research focused on the chemistry of the complexes formed1

-4• lnteractions between

alkaline earth metal species and nucleosides (without phosphate group) are weak and can be explained by means of only a few specified binding criteria5

, 6• In nucleic acids,

the phosphate-to-metal bonding dominates. Recent ab initia

d . . . f L.+ N + B 2+ dM 2+ . h

stu 1es on 1nteract1ons o 1 , a , e an g Wlt H2Po4- reveal significant electron transfer for all

complexes, except those invalving Na+7 • This implies that these interactions are not purely ionic. Ion pairs in organic solvents have been studied extensively by combinations of multi-nuclear magnetic resonance techniques8_•9_•

Recently a thorough study has been performed by Caatelijns10' 11 on the reactivity, the stereochemistry and

the kinetics of de-alkylation reactions of phosphates and phosphonates with LiCl and LiBr in solvents of various polarities. The NMR- and kinetic data provided strong evidence for the involvement of a penta-coordinated (P(V)) intermediate with only a moderate charge separation in the rate-determining step of the de-alkylation reaction (Fig.

11.1). Although no conclusion could be drawn concerning the

amount of charge separation in the P(V) intermediate, it was clearly demonstrated that its reactivity obeyed the stability rules of pentavalent trigonal bipyramidal (TBP)

(30)

0 ₀ _. 11 • R, /P, /R' LiX

"o·;t

'o' .

~ R"

o-R-o, 1 ' + 'p -o-: .... u R-0

... _ , e

I'

Li - R'-X+ ;p~ R'/ 0 X :F,Cl,Br

~!

0 0

'\

..

Fig. II.1 P(V) intermediate in the de-alkylation reaation. phosphorus compounds. These results are in contrast with previous work which argues in favour of an "Arbusov"-type mechanism in which reactions proceed via a nucleophilic attack on a saturated carbon, linked through oxygen with the tetravalent phosphonium ion (A)12

- 1

'*.

Even in reactions

in which the initial formation of a P(V) compound was established, the de-alkylation was supposed to occur via an intramolecular SNZ displacement from the ion-pair,

although a a 2s + crza- thermal pericyclic disproportionation reaction from the covalent P(V) compound (B) was not ruled

A OR RO .. ,,

I

C?u~~-OR

I

x

R • B . out15_-20

• With this in mind, it was tempting to investigate

the nature of the interaction of lithium balides and lithium perchlorate with model phosphates and a model phosphonate in salution by means of multi~nuclear NMR. Therefore 1H NMR spectra of 2-isopropoxy-2-oxo-5-methyl--1,2-oxaphosphol-4-ene (compound 1, Fig. II.2) were

recorded in a number of organic solvents. Also 7Li-, 35

cr-and 81Br NMR spectra of the salt/phosphate complexes in acetone were recorded to yield chemica! shift data for the 7Li nuclei and line-width data for the 35

c1-

and 81 Br nuclei. Moreover, the solvent- and salt-dependent nonequivalence of the methylene protons in 1 was ex~ined. Besides the study of aggregates by physical means, it also appeared

(31)

worthwhile to investigate the effect of lithium halides on the rate of phosphorylation of 2-methoxy-2-oxo-4,5-dimethyl--1,3,2-dioxaphosphol-4-ene (compound 2, Fig. 11.2). Bromide anions cause a remarkable decrease in the rate of

phosphorylation toward methanol in different cyclic phosph(on)ates11_• _{In order to establish whether this}

retardation is a more general property of halide anions in organic solvents, detailed kinetic data of the system LiX(X=F, Cl, Br)/2/methanol in deuteriochloroform were determined. Finally, low-temperature 31P spectra for the combination 2/LiF in tetrahydrofuran were investigated to examine the possibility of the formation of a covalent P-F bond. Some of the conclusions are corroborated by the results of CND0-2 quantum-chemical calculations.

II.2 The Peaation of a five-membePed ayatic P(IV) compound with alcohol in the pPeaence of lithium halidea. Addition of LiX (X=F, Cl, Br, Cl0 4) to solutions of compounds 1-4 (Fig. II.Z) in acetone reveals deshielding

0'\,

YO

.

CH 3 0'\_p/OJCH3 CH3 /p

I

/ \

~0

, H H3C-O 0 CH3 ~-~~·'7

',

CHJ H H 1 ₂ 0 11

rQ1

,.-P..\:'o~

if:

~

4

Fig. II.2 Model aompounda 1-4.

of the proton resonances in the 1H NMR most likely as a consequence of the complexation of the phosphoryl oxygen atom with the lithium cation. This results also in a

(32)

deshielding of the phosphorus atom. These results are in accord with former workon salt/phosphate aggregates11

•21- 24•

The above-mentioned adducts all disproportionate on prolonged standing to the corresponding de-alkylated products in case of LiF, LiCl and LiBr. The weakly nucleophilic CI0₄ ion is not capable of inducing the de-alkylation process.

Addition of one equivalent of methanol to a 0.5 M salution of 2 in tetrahydrofuran (THF) results in a fast

phosphorylation reaction. However, in the presence of one equivalent of LiBr, almost complete de-alkylation is

observed11_• _{In order to elucidate the observed decreasein}

the rate of phosphorylation, the model system LiX(X=F, Cl, Br)/2/methanol was studied in CDC1 3 in equimolar ratio of the components (0.3 M). To dissect the reaction rates of the combined phosphorylation (Path II, Fig. II.3) and de-alkylation (Path I) in ~he system, kinetic measurements in CDC1 3 were performed on 2/methanol (equimolar ratio, 0.3 M) and 2/LiBr (equimolar ratio, 0.3 M). In view of the poor solubility of LiBr in CDCI₃at 298 K (8.10- 3 mol/L), the effective LiBr concentratien in salution during the de-alkylation process can be regarded as constant and the reaction rate becomes pseudo-first order in phosphate with -d!Pl/dt = _{k2.!PJ and k2}= _{k 1.!BJ in which !Pl is the}

concentratien of phosphate (mol/L) a~d (BJ is the concentratien LiBr (mol/1). This gives ln(IPJt/[PI₀) = -k 1.!Bl.t with an average k1 = 3.41 10-3 L/mol s.

[BI was estimated at 10-Z mol/1 of LiBr. Earlier kinetic measurements of the de-alkylation (Path I) in the more polar acetone-d 6 revealed that the reaction rate was first order in phosphate and first order in LiBr10

• In case of

phosphorylation (Path II), kinetic data on the system

2/methanol in deuteriochloroform revealed that the reaction was first order in phosphate and of zero order in methanol

-3 -1

with an average rate constant of 2.15 10 s • As the phosphorylation proceeds approximately 24 times faster than the de-alkylation, it is reasanabie to neglect the latter path in case of the phosphate/salt/alcohol system. Therefore,

(33)

Me • 0 OJMe

o

0 1

~

Me Li • • '\. 1 1 _ L j - - -

w/,,,..

r

I p . _ . ', P-O L'B _~~B~ _o/ \ _o _Me _M _L'o~~I ~ e - , 0 0 Me I / ~Br

Me-O:>~OlMe

MeOH Me Me ... MeOH

~ 0~

Me

~

-0

H-Q~,~.

I

l,,·p-o -Me-0,.-

I

11 H Me , .. P'-..

y

Me Me-o''l

o'

Y

Me-0

°

0 '-...Me 0 _~IOJMe P

I

+MeBr / \ Li-0 0 Me

~

MeOH 0 11 H Me

··P XyM

"'''' -... . e Me-0\"/ 0 Li-0 O

Fig. II.3 Reaation path~aya of the ayatem LiBP/2/methanoZ in deutePioahZoPofoPm.

(34)

this system can be approximated by a first order reaction in phosphate. The kinetic first order plots of the phospho-rylation (Fig. !!.4) reveal that the reaction rate decreases

0 In [ Pl t

I

5.0 6.0 0 Fig. II.4 T=298 K Li Br LiF sa I t-f ree 600 1200 1800 2400 3000 3600 - - - - t Is I

Least squares plot of Zn[P]t vs. t for the system LiX(X=F$ CZ$ Br)/2/methanoZ in CDCZ 3•

The initiaZ amounts of the different compounds are equimoZar (O,S M).

in the order salt-free>fluoride>chloride>bromide (Table II.1). This result is particularly striking because complexation of the phosphoryl oxygen by the lithium cation increases the phosphorylating ability of this compound toward

alcohols. The measured rate constant for the phosphorylation is somewhat lower than the real value because a slight amount of methanol is located in the inner solvation shell of the lithium cation. As a consequence, the amount of free methanol is slightly reduced.

The results show a relation between the reaction rate of the phosphorylation and the ionic radius of the different anions studied. The observed relation is not linear because of the different solvent reorientations induced by the halide ions involved9_• _{This creates unique solvation spheres for the}

various anions. As a result of their ordered solvation spheres, the halide ions, which appear as close-ion pairs9

(35)

Table. II.1 Kinetia data of the system LiX(X=F~ CZ~ Br)/2/ methanol in CDCZ₃at T=298 K.a salt t . b

1 ,

m1n. r,a Ä 10 3k, s -1 d krel e

-

5.57

-

2.15 1.00 LiF 11 • 38 1.33 1.02 0.48 LiCl 13.17 1.81 0.88 0.40 LiBr 31 • 81 1.96 0.36 0.17

aThe initia! concentrations of the various compounds are equimolar (0.3 M).

bobtained via the equation t!=ln2/60k. aAnionic radius without solvation spheres.

'

dDetermined via the slope of the straight line in Fig. 11.4.

e -3

krel=k/(2,15 10 ).

in combination with the metal ion, are capable of shielding the phosphorus atom against a nucleophilic attack of methanol (Fig. II.5). Consequently, the phosphorylation of methanol is retarded in comparison with the salt-free experiment. Most likely, attack of methanol proceeds by a displacement of the halogen atom followed by a fast intramolecular nucleophilic attack at the phosphorus atom. These kinetic data therefore provide evidence for the proximity of the halide ion to the phosphorus atom.

0

.

; ' ir---·uu--~-0 0 .. •' i

J~~---~----.--e

Fig. II.5 Stereosaopia ORTEP drawing of the LiBr/2 aggre-gate. The alosest P-Br distanae in the aompZe~

was aaZaulated to be 4.0

i.

The different radii used for saaZing do not repreaent aotvation.

(36)

11,3 Solvent-and salt-induced diffe~ential shielding effects in a five-membe~ed cyalic phosphonate.

In order to improve the knowledge about the location of the Li+ ion in the salt/phosphate complexes studied here, the paramagnatie Eu3+ ion was used in combination with compound 1, Addition of Eu(fod)₃25 _{to a solution of}₁ _in

CDC1 3 reveals deshielding for all the proton resonances in the NMR spectrum due to phosphoryl oxygen complexation by the Eu 3+ ion (Fig. II.6). The shift of the tertiairy isopro-poxy proton (4) shows a high concentratien dependence. Obviously, the Eu 3+ ion is located in the proximity of this proton. This confirms the previously observed complexation of the phosphoryl oxygen atom by the Li+ ion. Moreover, the asymmetrie ring in the structure causes, via preferred orientation of the Eu(fod) 3 complex, increased shift dif-ferences of the isopropoxy methyl gróups at higher Eu(fod) 3

~ppm

l

0 0 Me 14

; >çr

~0 H 4 12 _Me _: 5 1 Me H. H ! 3' 3 10 8 oL---~o.~2---~o.~4----~o~.a~----~o~.8~----~1~~~-- eq Eu lfodl3

Fig. II.6 60 MHz 1H NMR data of compound 1 in CDCZ 3 ~ith

diffe~ent concent~ations _{of Eu(fodJ 3 at P=298 K.}

ö in ppm vs. TMS,

concentrations (viz. two resonances 1). The asymmetrie loca-tion of the Eu 3+ ion with respect to the ring methylene

(37)

protons (3 and 31₎ _{causes a different deshielding for both}

protons. From molecular model studies on compound 1 one would expect intrinsically nonequivalent resonances of the methylene protons. Indeed, the 250 MHz 1H NMR spectrum of

1 in acetone-d₆revealed nonequivalent resonances (Ao=25 Hz; 0.10 ppm; Fig. 11.7). This value appeared to be solvent

dependent and should therefore be ascribed to solvent-induced differential shielding effects (viz. Table !1.2). From

earlierresearch by CasteZijns10_•11 _{it was established that}

the shift difference between the methylene protons was depen-dent on the anion, the cation and also on the bulkiness of the exocyclic alkoxy ligand. Comparison with literature data on solutions of alkali halides in organic solvents9 _revealed

that a large Aö is observed in these cases where the salt-ions form close pairs and in which one or both ions are surrounded by partially oriented solvent molecules. Therefore, the

observed Ao in case of close-ion pairs can best be explained by reorientation effects of the solvent structure near the close-ion pair and concomitant electric field effects. From Table 11.2 it is evident that there is no salt-induced Aê of the methylene protons in case of fully complexated or solvated cations and anions. Hence, only possible solvent--induced differential shielding can be measured (Aê solvent, Table II.2). In case of

so

_{2 liq we have fully solvated} halide_anions which prevent salt-induced Aö of the protons.

o~Pçro

I

t.;e

H4 /

.\-a \

~

Me· Me H

H

I I 3 3' - I J ppm

Fig. II,? 250.13 MHa 1H NMR spect~um of compound 1 in acetone-d at T=298 K.

(38)

Table II.2 Solventand salt effeats on the ahemiaal shift differenae äö of the methylene protons of 1.4

clóse;-ion salt solvent solventa saltd

. b pa1r M ppm llO ppm

-

(CD 3) 2SO 0

--

-

co

₂_{c1 2} 0.048

--

-.

_{CDC1 3} 0.100

-yes _LiC104 CD₃

No

₂ 0 0.048 yes LiCl(satd) _{CD 3}

No

₂ 0 0.060 yes LiCl(sàtd) _{(CD 3) 2}

co

0.01 0.050

yes LiBr(1 equiv) _{(CD 3) 2}

Co

0.01 0.286

yes LiBr(satd) _C6D6 0. 1768 _0.504

no LiCl0₄ SOz liq 0 0

no LiBr(1 equiv)

so

₂liq 0 0

no LiBr _{(CD 3) 2}

so

0 0

no A1Cl₃ _{(CD 3) 2CO} 0.010 0

no LiBr/kryptofix _{(CD 3) 2CO} 0.010 0

(1: 1)f

a250.13 MHz 1H NMR (T=298 K). bAccording to Popov6 _and

Weingärtner9

• aSolvent-induced differentlal shielding.

d~o(salt): measured total effect, containing contributions from both solvent and ion pairs. eThis large effect can be explained by aromatic-solvent-induced• shift (ASIS) which is known for phenyl fragments. fKryptofix 221/LiBr in 1:1 mol ratio.

In case of A1C1 3 we have a Lewis aaid ions (X=F, Cl, Br) in the presence of 7Li NMR measurements of a 1:2 mixture

which.forms A1C1 3

x-halide anions11_,

26

of kryptofix 221 and LiBr in acetone revealed two resonances of equal inten-sity at o( 7Li) 2.43 ppm and o( 7Li) -0.11 ppm which can be attributed to the LiBr in salution and the fully complexed kryptofix 221.Li+, respectively. In a 1:1 mixture there is no free LiBr in solution. As a result there is no

(39)

Formation of a covalent bond between the Li+ ion and the phosphoryl oxygen atom would lead to a more positively charged phosphorus atom. In order to investigate a possible change in net atomie charge on the phosphorus atom upon addition of lithium salt, the accurate values of the 2JHH geminal coupling constants27 _{had to be determined via}

computer simulation of the 250 MHz 1H NMR spectra of 1

in acetone-d 6, in the presence of öne equivalent of LiBr in acetone-d6 and in benzene-de (Fig. II.8-II.10). The results are summarized in Table 11.3. From this table it is evident, that the difference in 2JHH between acetone-de and acetone-d 6 with one equivalent of LiBr is 0.34 Hz. This implies a negligible change in the net atomie charge on the phosphorus atom. Moreover, this conclusion is consistent with the observed small (approximately 1 ppm) 31 P chemical

shift differences which occur upon addition of salt.

al ~~--l-~~~~--'---'--- scale, 10Hz

Fig. II.B RecoPded (a) and computeP-eimulated (bJ 2S0.1J

MHs 1

n

NMR epectPum of the methylene pPotone of compound 1 in acetone-d6• Notice the Pesemblance of the fine structuPe in the lo~-field domain

(40)

Ao _ _ - . .

··~~···~-~· scale 10 Hz

o.so _ ~ppm '

Fig. II.9 Reaorded (a) and aomputer-simulated (b) 250.13 MHs 1H NMR speatrum of the methylene protons of aompound 1 in the presenae of one equivalent LiBr in aaetone-d 6,

-~ppm

Fig. II.10 Reaorded (a) and aomputer-simulated (b) 250,13 MHz 1H NMR speatrum of the methylene protons of aompound 1 in bensene-d₆•

(41)

Tab~e II.3 Indireat aoup~ing aonstants {J) and chemiaal shift values (W) via computer-simulated 260.13 MHz 1H NMR spectra of 1. 0 0 Me

~p))'

s Me / '

.\-o '.

~

Me' H H H 2 J W a

'

ppm J, Hz Acetone-d 6, Rms error=0.042 W(1) _J₁₂ 12.64 3 24 2.75 W(2) 0.538 3₁₃ 15.74 _{3 25} 2.55 W(3) 0.548 _{3 14} 33.61 3 34 3.08 W(4) 3.201 _{3 15} 0.80 _{3 35} 2.28 W(S) 0 3₂₃ -18.67 _{3 45} 1.44 Benzene-d6 , Rms error=0.116 W(1) _{3 12} 13.46 _{3 24} 2. 72 W(Z) 0,681 _{3 13} 15.24 _{3 25} 2.49 W(3) 0.503 _J14 33.54 3₃₄ 3.03 W(4) 2.917 _{3 15} 0,85 _{3 35} 2.30 W(S) 0 _{3 23 -18.41} ₃₄₅ 1. 45

Acetone-d0/LiBr 1:1 mol ratio, Rms error=0.167

W(1) _J12 14.28 3₂₄ 2.69

W(Z) 0.551 _{3 13} 15.03 _{3 25} 2.48

W(3) 0.656 _{3 14} 34.58 3₃₄ 2.74

W(4) 3.297 _J₁₅ 0.74 _{3 35} 2.36

W(5) 0 3₂₃ -19.01 ₃₄₅ 1.43

(42)

31 . . t. f.

11.4 Lo~-temperature P NMR ~nvest~ga ~on on a

~ve-membered ayctic phosphate in the presenae of lithium fluoride,

Earlier work concerning the kinetics of de-alkylation reactions in organophosphorus compounds strongly points to the involvement of penta-coordinated intermediates10

'11 •

In this. respect it is interesting to investigate possible intermediate structures by means of low-temperature 31 P NMR techniques. Recentworkof Granoth et at.28 _{showed structures}

intermediate between halophosphoranes and phosphonium halides with covalent phosphorus-halogen honds, which have a very large degree of ionic character. 31 P chemica! shifts were measured near ö(31 P) +40 ppm which points to phosphonium structures. Granoth took recourse to electric field induced contributions in order to explain the deshielding of one of the aromatic protons H

0 (Fig. !1.11), Over such small

dis-,_-'fr~

~~-

x

b-Ho

Fig. II.11

tances relatively large uncertainties exist in the use of the formalism, both in order of magnitude and direction29_,

Moreover, an ion pair suitably oriented with respect to the C-H₀ bond, could very well result in the same effect, i.e. a covalent or ionic P-X bond (X = halogen) is not necessary in order to rationalize deshielding of the H₀ proton. The orientation of the halide ion with respect to the phosphorus atom is prQbably determined largely by steric factors.

For the salt/phosphate aggregates studied here it is interesting to determine whether a stabie penta-coordinated structure can be trapped at low temperature. In case of the bromide anion, compound 2 revealed the highest rate of

de-alkylation and thus a concomitant low activatien energy (Ba::::: 12.5 kJ/mol) to form the intermedia te state11_• _Therefore,

(43)

recorded. The low dielectric constant of THF suppresses the ion pair dissociation (e=7.4, T=298 K) thus supporting the generation of a penta-coordinated intermediate with moderate charge separation. Covalent character of the phosphorus--fluoride interaction is accurately measurable due to the large 1JPF z1000 Hz30_, _{Addition of one equivalent LiF at'}

213 K to a salution of 2 (0,3 M) in _{THF-d 8} resulted in a line broadening of 1.75 Hz and an upfield shift of 0.40 ppm. No doublet in the 31 P resonance could be resolved in the temperature region 213-298 K. A similar result was obtained in case of NaF.

Recently Richman et a~. 31 publisbed the 31P- and 19F NMR results of cyclenfluorophosphoranes and demonstrated. a clear distinction between covalent and ionic structures.

In the ionic structures, no 1JPF was observed whereas in the penta-coordinated cornpounds couplings of 800-900 Hz were rneasured. Based on these observations and our 31 P NMR data it can therefore be concluded that for the salt

aggregates studied here, NMR techniques reveal a change in solvation but no penta-caordinated intermediate structure with a life-time short on the NMR time scale32

• In addition,

different interrnediate geometries of compound 2 with one equivalent LiBr were calculated using the CND0-2 rnethod33_•

(Por the theory of the CND0-2 method, see Appendix Chapter III). Optimization of the different structures a, b and a toward

lowest energy (Fig. II.12) revealed a decreasing stability in the order a > b > c. The kinetic data on de-alkylation reactions point to penta-coordinate interrnediate structures10_•

Me Me-0,_

~~Me

'P-O Li-O,...j Br c Fig. II.12 DilfePent geometPiee of the LiBP/2 comple~.

a) c~oee-ion paiP stPuatuPe.

b) phosphonium etPuctuPe. c) penta-cooPdinate etPuctuPe,

(44)

Combined with the CND0-2 calculations this implies the for-mation of a short-lived structure of type a as a result of an interaction of the d-orbitals of the phosphorus atom with the unpaired electron pairs of the bromide anion. Structure

a seems the most stable configuration observable using NMR

spectroscopy.

II.S 7Li-~ 35

ct-

and 81Br NMR investigations on salt/phosphate aggregates in aaetone.

The behaviour of the Li+ ions in the salt/phosphate aggregates in acetone was studied by means of 7Li NMR. With respect to the behaviour of the anion in the salt/phosphate complexes, 35c1- and 81Br NMR techniques were applied. The data of the 7Li, 35c1 and 81Br studies are shown in Fig. II.13 - II.16 and in Tables II.4- II.S.

The properties of 7Li nuclei are quite favourable for NMR studies. The resonance lines of the Li+ ion in solutions are exceptionally narrow cw

₁

~z Hz)3_~ _{and chemica! shifts}

(vs. _{4.0 M aqueous LiCl0 4 solution) can be measured with} considerable accuracy. Literature data available for chlorine-and bromine NMR in non-aqueous solvents are limited. Halide ion quadrupale relaxation rates have·been reported for methanol9

'35' 36, dimethylsulfoxide9' 35, nitromethane37,

formic acid9_, _{N-methylformamide}9_, _{dimethylformamide}9_,

acetonitrile9_, _acetone9 _{and for mixtures of acetonitrjle}38_,

methanol36_' 39 _{and acetone}39 _{with water.}

From Table II.4 it is evident that the 7Li chemica! shifts of lithium bromide are concentratien dependent. This concentratien dependenee can be attributed to the formation of contact-ion pairs, i.e. to cases where the anion directly replaces a solvent molecule or molecules in the inner solvation shell of the cation8

, It has been

previously observed40

'41 that the contact-ion pair

equili-brium strongly depends on the donor ability of the solvent molecule as well as on the bulk dielectric constant e of

(45)

Tabte II.4 97.21 MHs Lithium-? ahemiaat shifts (ppm) of 1:1 mol ~atio LiB~/modet compound agg~egates

in acetone at T=298 K,a

LiBr/1 LiBr/3 LiBr/4 LiBr

Mb _ö(7Li)0

M ó(7Li) M o{7Li) M o(7Li)

0.11 2.23 0. 13 2.41 0.11 2.15 0.09 2. 50

0.31 2.04 0,25 2.32 0.26 2.03 0.24 2.48

0.47 1.98 0.50 2.17 0.49 1.89 0.53 2.47

0,94 1. 81 1. 00 1. 95 0.98 1. 78 0.94 2.36

a 7Li chemica! shifts against 4.0 M aqueous LiCio₄ solution. bconcentration of model compound (mol/L).

0

Estimated accuracy of ö{7Li): 2 10- 2 ppm.

the medium. Acetone has a dielectric constant of 20,7 (298 K),

its donor ability is reasonable high and on Gutmann's scale~2

its donor number is 17.0. Contact-ion pair formation in acetone occurs likewise in case of lithium chloride (Table 11.7). From the 7Li chemica! shift data it can be concluded that the chemica! shifts are clearly dèpendent on the

counter ion (Cl-, Br-, CI0_{4-) and on the concentration. The}

7Li chemica! shift values agree reasonably well with those reported by Maciel et al.43 _{and by}_Akitt_and_Downs44

• From

Table 11.4 we see that addition of one equivalent of the phosphoryl compounds 1, 3 or 4 to the lithium bromide solution results inshielding of the 7Li nucleus. This shielding occurs as a result of replacement of at least one of the four carbonyl groups in the inner solvation shell of the lithium ion by a phosphoryl group45

• Addition

of small amounts of water results in an analogous replacement by water. Limiting 7Li chemica! shift values for infinite dilution obtained via extrapolation from data in Table 11.4 are listed in Table 11.5. From the CND0-2 geometry optimi-zations of the model compounds 1 and 4 there seems to exist a correlation between the dipole moment of the phosph(on)ate (8.23 and 8.83 D for 1 and 4, respectively) and the value of

(46)

Tabte II.5 Limiting vatues at T=298 K foF infinite ditution of tithium-7 chemiaat shifts (ppm) and bFomine-81

line widths (kHz) of 1:1 mol Fatio comple~es

of model aompound/salt in aoetone. limiting va lues compound o(7Li)a W (81Br)b

!

LiBr sec 2.53 6.3 1/LiBr 2.36 8.5 3/LiBr 2.53 6.6 4/LiBr 2.26 7.2

a _{Estimated accuracy: 2 10}-2 _{ppm; chemical shift in ppm} _vs. 4.0 M aqueous LiC104 solution. bEstimated accuracy: 0.5 kHz.

the limiting 7Li chemical shift. A larger dipale moment seems to result in a larger shielding of the 7Li nucleus. Compound 3 reveals a different behaviour (dipole moment, 13.05 D). The results of the 81 Br line width measurements of the 1:1 LiBrimodel compound complexes are plotted in Fig. II.13. There is considerable broadening of the 81 Br resonance with increasing concentration of the complexes in acetone. The observed concentration-dependent broadening of the 81 Br resonance is indicative of contact-ion pair formation because of asymmetrical soivation of the bromide anion, caused by a contribution of the (Li-0-P) fragment, thus enhancing the electrical anisotropy around the 81 Br nucleus. This results into an increase in line width with increasing salt concentration. The contribution of the (Li-0-P) fragment to the electrical anisotropy of the

ellipsoidal 81 Br nucleus can also account for the difference in limiting values of the 81 Br line widths in the 1:1

complexes salt/phosphate and in LiBr (viz. Table II.S). The limiting 81 Br line widths were obtained via extrapolation from data in Fig. 11.13 to infinite dilution. Table II.4 and Fig. II.13 reveal, that no distinction can be made between the cyclic and the acyclic model compounds. The different behaviour of 3 (Fig. 11.13) might be explained