The synthesis of "apo-proteinic" multidentate, N2SS* and N2S, ligands to model type I trigonal and trigonal-pyramidal copper protein sites

FULL PAPER

The Synthesis of ‘‘Apo-Proteinic’’ Multidentate, N₂SS* and N₂S, Ligands to

Model Type I Trigonal and Trigonal-Pyramidal Copper Protein Sites

Timothy C. Higgs*^[a]and C. J. Carrano^[b]

Keywords: Protein models / Biomimetic synthesis / Type I copper protein / N,S ligands / Steric hindrance

The syntheses of the novel sterically highly hindered asym-

tocol which is highly effective despite the high involatility

metric tripodal tridentate N₂S and ‘‘distended’’ tripodal tetra- of the N₂O precursors/N₂S products. This process has been

dentate N₂SS* (S: thiolate; S*: thioether donor) ligands, illustrated crystallographically for the sequence of conver-

2-[bis(3Ј-isopropylpyrazol-1Ј-yl)methyl]-6-tert-butyl-4- sions of 2-[bis(pyrazol-1Ј-yl)methyl]-6-tert-butyl-4-methyl-

methylbenzenethiol (11) and 3-{2Ј-[bis(3ЈЈ-isopropylpyrazol-

phenol (3) to O-{2-[bis(pyrazol-1Ј-yl)methyl]-6-tert-butyl-4-

1ЈЈ-yl)methyl]phenylsulfanyl}-1,1-diphenylpropane-1-thiol methylphenyl}-N,N-dimethylthiocarbamate (4) to S-{2-[bis-

(28) are reported. These molecules possess the atom donor

arrays of ‘‘trigonal’’ and ‘‘trigonal-pyramidal’’ type I ‘‘blue’’

copper proteins respectively. The success of the synthesis has

transpired from the development of an effective flexible

route to achieving the functional group interconversion of the

phenolic group of the N₂O heteroscorpionate ligands based

upon the already known 2-[bis(pyrazol-1Ј-yl)methyl]phenol

framework to their benzenethiol analogs. This has been ef-

fected using the Newman−Kwart rearrangement of the thi-

(pyrazol-1Ј-yl)methyl]-6-tert-butyl-4-methylphenyl}-N,N-

dimethylthiocarbamate (5). These ‘‘all-in-one’’ ligands rep-

resent a new strategy to modeling ‘‘blue’’ copper proteins and

should be capable of complementing recent significant de-

velopments in the field since they are sterically somewhat

less bulky than pre-existing ligands whilst maintaining vi-

able coordination vectors and thus allow for the adoption of

more ‘‘open’’ metal coordination environments.

ocarbamic acid O-ester of the N₂O-phenol by a ‘‘Direct Inser- ( Wiley-VCH Verlag GmbH, 69451 Weinheim, Germany,

tion Vapour Phase Flash Vacuum Pyrolysis’’ (DIVPFVP) pro- 2002)

Introduction

Mixed [N_xS_yO_z] coordination arrays are almost ubiquit-

ous in metalloproteins systems, notably with zinc^[1Ϫ6]and

copper.^[7Ϫ16]Recently, considerable effort has been ex-

pended to synthesise unsymmetrical ligands possessing

Figure 1. The molecular structure of the N₂S donor ligand

2-[bis(3Ј,5Ј-dimethylpyrazol-1Ј-yl)methyl]-6-tert-butyl-4-methyl-

mixed N and/or S and/or O donor sets. Vahrenkamp et

al.^[17Ϫ21]have reported a number of tripodal N₃O and N₃S

ligands that incorporate carboxylate, phenolate, or thiolate

functionalities. Parkin et al.^[22Ϫ24]have successfully synthe-

sized a series of facially coordinating N₂O and N₂S heteros-

corpionate ligands possessing thioether, thioimidazolyl, or

carboxylate moieties attached to the BϪH group. Rior-

dan^[25,26]and Fenton^[27]have also pursued this goal with

significant success. A recent noteworthy achievement in this

field has been accomplished by Hammes, Carrano et

al.^[28Ϫ31]who have used the N₂S donor ligand, L2SH (Fig-

benzenethiol (L2SH)^[28Ϫ31]

ure 1), to probe mechanisms of reactivity of Zn^IIligated by

N₂S₁S*₁donor sets (S*: exogenous thiolate ligand) specific-

ally in relation to alkylation of the exogenous RS* ligand.

These studies are especially significant in light of the recent

advances in recognizing the role of zinc metalloproteins in

alkyl group transfer reactions.^[32,33]

The simplicity of its method of synthesis and its consider-

able potential for future derivatization is a particularly at-

tractive feature of this ligand design. To date though a de-

finitive description of the synthetic protocols and experi-

mental techniques required to produce ligands similar to

L2SH has not been reported. Herein we finally report a

definitive experimental method to produce these N₂S li-

gands, based upon a 2-[bis(pyrazol-1Ј-yl)methyl]benzene-

thiol skeleton, and our attempts to further derivatise these

ligands to achieve a total synthesis of the ‘‘apo-proteinic’’

[a]

Laboratory N2.16, The Department of Chemistry, The

University of Edinburgh, King’s Buildings,

West Mains Road, Edinburgh, EH9 3JJ, Scotland, UK

Fax: (internat.) ϩ 44-131/650-6452

E-mail: Tim.Higgs@ed.ac.uk

[b]

The Department of Chemistry, Southwest Texas State

University,

San Marcos, Texas, 78666, USA

E-mail: CC05@swt.edu

3632

 2002 WILEY-VCH Verlag GmbH & Co. KGaA, 69451 Weinheim 1434Ϫ193X/02/1121Ϫ3632 $ 20.00ϩ.50/0 Eur. J. Org. Chem. 2002, 3632Ϫ3645

Ligands to Model Type I Trigonal and Trigonal-Pyramidal Copper Protein Sites

FULL PAPER

coordination sites of trigonal and trigonal-pyramidal type other potential ligands.^[45,46]This rack-induced displace-

I copper proteins, recently described by Tolman and Spen- ment of the apical methionine ligand often observed in this

cer^[34]as the ‘‘holy grail’’ of synthetic bioinorganic chem- class of metalloprotein, relative to the Cu ion has been pos-

istry.

tulated as modulating the rather high redox potential of

the Cu^I/Cu^IIcouple observed in these systems.^[45,48,60]More

evidence though is required to support such speculation

Type I ‘‘Blue’’ Copper Proteins (Cupredoxins)

Type I copper proteins, are found in bacterial and plant and hence the need for viable model systems with accurate

systems, having a role in biological electron transfer N_xS_ydonor sets, this latter requirement necessitating novel

processes.^{[7Ϫ10,35Ϫ43]}For this task they are ideally suited approaches to asymmetric ligand design.

possessing a trigonal, N(his)₂S(cys)-ligated Cu^I/IIredox

center in an ‘‘entatic state’’ with a coordination number/

geometry intermediate between that preferred by the ϩ

and ϩ Cu oxidation states. This ‘‘entatic’’ state is possibly

enforced by the rack-induced bonding mechanism, first

proposed by Malmström in the 1960s,^[44]whereby the fold-

ing of the protein leading to a minimum overall energy con-

formation can stabilise a local energy maximum in the vi-

cinity of the metal site (e.g. to support an unconventional

arrangement of ligands). Considerable effort has been ex-

pended to support or challenge the rack hypothesis. Gray,

Malmström et al.^[8,45Ϫ48]have produced experimental evid-

ence that supports it whereas Ryde et al.^[49,50]have under-

taken quantum chemical calculations that challenge it for

the oxidized [Cu^II] protein.

Synthetic Bioinorganic Chemistry: Type I Copper Protein

Models

Synthesising a Cu^IIcomplex that can simultaneously

model the extraordinarily anomalous structural, spectro-

scopic, and electrochemical properties of type I copper pro-

teins, is a considerable challenge and one that has preoccu-

pied synthetic bioinorganic chemists for over 25 years. The

situation is further complicated by the notorious extremely

facile reduction of mercaptoϪCu^IIcomplexes, postulated to

proceed by a two-step free-radical coupling mechan-

ism.^[61,62]

The initial homolytic cleavage (1) of the Cu^IIϪSR bond

forming Cu^Iand a thiolate radical is reversible being in dy-

namic equilibrium with the corresponding bond formation

but (1) is driven completely to the right by subsequent irre-

versible 2 ϫ RS^·radical coupling (2) which forms stable dis-

ulfide.

Type I copper proteins exhibit extraordinarily anomalous

spectroscopic properties with an intense absorption band at

ca. 620 nm attributed to S(cys) Ǟ Cu^IICT.^{[10,38,41,42,51]}The

EPR spectroscopic properties are similarly intriguing with

the observance of both rhombic and axial type spectra,^[52,53]

all possessing unusually small A_//hyperfine coupling con-

stants of Յ 70ϫ10^Ϫ4cm^Ϫ1compared to 180Ϫ200ϫ10^Ϫ4

cm^Ϫ1for ‘‘normal’’ tetragonal Cu^IIcomplexes. Intensive

Cu^IIϪSR v Cu^Iϩ RS^·(1)

2 RS^·Ǟ RSSR (2)

Early attempts at synthesising effective type I models,

studies using sulfur K-edge X-ray absorption spectroscopy such as those of Bosnich et al.,^[61][62]were thwarted by the

have indicated this is due to extensive delocalization of the facile mercapto-mediated reduction of Cu^II. Brader, Borch-

type I Cu^IIsite^[10]with an unusually short (ca. 2.1 A), cova- ardt, and Dunn^[63]elegantly avoided many of the synthetic

˚

lent Cu^IIϪS(cys) bond which effectively delocalises the un- difficulties associated with the required ligand systems by

paired electron away from the Cu^IInucleus thus reducing using an insulin-stabilized monomeric Cu^IIchromophore as

the spin-coupling interaction of the former with the latter. a framework onto which were introduced a series of exogen-

Numerous members of this class of proteins possess subtle ous thiolate-based ligands (C₆F₅S^Ϫ, C₆HF₄S^Ϫ). The result-

structural additions to the fundamental, commonal ant adducts effectively mimicked several of the spectro-

N(his)₂S(cys) Cu ligation, and the effect these variations scopic properties (EPR, electronic spectroscopy) of type I

have on the spectrochemical properties of these proteins are sites. Unfortunately, these adducts still possessed redox in-

currently undergoing rigorous scrutiny. These variants in- stability over a time-scale of hours and X-ray characteriza-

clude the ‘‘pure trigonal’’ site (fungal laccase,^[54]ceruloplas- tion of these systems was not possible. Only with the sem-

min^[55,56]), the ‘‘trigonal-pyramidal’’ site wherein a weakly inal work of Kitajima et al.^[64Ϫ67]in the early 1990s was the

˚

coordinated (ca. 2.6 A) fourth ligand, typically a methion- redox instability overcome to some extent by the use of the

ine-S thioether, occupies an axial site (plastocyanin^[57]), and sterically highly hindered tridentate N₃-donor ligand hy-

the ‘‘trigonal-bipyramidal’’ site wherein two ligands occupy drotris(3,5-ddiisopropylpyrazol-1-yl)borate and similarly

˚

axial positions at long distances, Ͼ 2.8 A, with one again sterically hindered thiol ligands, including tBuSH, Ph₃CSH,

being methionine and the other a backbone carbonyl oxy- and C₆F₅SH. These ligands were used to synthesise ‘‘spec-

gen atom (azurin^[58,59]).

troscopically accurate’’ four-coordinate Cu^IIcomplexes

The quantum chemical calculations of Ryde et al.^[49,50]with N₃S ligation (S: thiolate), the sterically hindered nature

have led to a refinement of the rack-model for type I copper of the ligands conferring at least low-temperature stability

proteins. This revised model states that whilst protein fold- onto the complexes so that in a couple of cases, structural

ing has a minimal effect on the Cu^IIcoordination geometry characterizations could be made. These model systems have

it has a significant effect on the Cu^Isite of the reduced several structural inaccuracies in that the N₃S coordination

protein, forcing the elongation of the axial methionine li- sphere is formerly incorrect, having one too many N-donors

gand in addition to shielding the metal from water and and no equivalent to the S(met) donor and the complexes

Eur. J. Org. Chem. 2002, 3632Ϫ3645

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T. C. Higgs, C. J. Carrano

FULL PAPER

exhibit flattened tetrahedral, not trigonal or trigonal-pyr-

amidal, geometries. Tolman, Holland et al.^[68Ϫ71]have re-

cently reported a stunning breakthrough in this field, hav-

ing successfully synthesized tractable, accurate structural

model complexes of both three- (N₂S) and four-coordinate

(N₂SS*) ‘‘blue’’ copper sites. This again achieved by the use

of sterically highly crowded ligands, the (1Ϫ)-bidentate N₂-

donor ligand 2,4-bis(2,6-ddiisopropylphenylimido)pentane,

the (1Ϫ)-monodentate ligand triphenylmethylthiolate (for

the N₂S model) and the (1Ϫ)-bidentate thiolatoϪthioether

donor 2-(methylsulfanyl)-1,1-diphenylethanethiolate (for

the N₂SS* model). The N₂S model closely mimics the spec-

troscopic properties of ‘‘trigonal’’ type I sites with a similar

S(cys)ǞCu^IILMCT band position and intensity and an

‘‘axial’’ type EPR spectrum with small A_//values. The

N₂SS* model is the first to formerly model the ligation of

the classic type I ‘‘blue’’ Cu^IIsite, and possesses many of

the essential spectroscopic properties of these sites. This

complex though does not accurately model the structure

of ‘‘trigonal-pyramidal’’ sites, the Cu^IIN₂SS* moiety being

somewhat ‘‘squashed’’ by the extensive steric hindrance re-

quired to ‘‘redox-stabilise’’ the Cu^IIϪS_thiolatebond, con-

sequently forming a flattened tetrahedral geometry which

more resembles the Cu sites of nitrite reductases and certain

azurin mutants (His, or Glu replacing the Met ligand).^[69]

Therefore, there is still a requirement for ‘‘blue’’ copper li-

gands which allow for more ‘‘open’’ Cu^IIcoordination with

a long ‘‘axial’’ S(met) interaction whilst maintaining the

fundamental short S(cys)ϪCu^IIbond.

Scheme 1

This involved the synthesis of the N₂O phenol analog

(based upon a 2-[bis(pyrazol-1Ј-yl)methyl]phenol frame-

work) of the desired benzenethiol ligand with the required

alkyl-substitution pattern, on the pyrazole rings and/or the

phenol (using an appropriately substituted salicylaldehyde,

Results and Discussion

Total Synthesis of Flexible ‘‘Apo-proteinic’’ Type I Copper

Protein Binding Sites

The development of ligands based upon a 2-[bis(pyrazol- typically

3-tert-butyl-2-hydroxy-5-methylbenzaldehyde).

1Ј-yl)methyl]phenol unit by the direct solventless reaction Synthesis of a thiocarbamic acid O-ester from this phenol,

of various bis(pyrazolyl) ketones and salicylaldehyde,^[72Ϫ76]then its pyrolytic conversion by NewmanϪKwart

and subsequent development of a thiol analog, 2-[bis(pyra- rearrangement^[78Ϫ80]into the isomeric thiocarbamic acid S-

zol-1Ј-yl)methyl]benzenethiol, by substituting 2-thiocyana- ester and lastly cleavage of this by either base hydrolysis

tobenzaldehyde^[77]for the salicylaldehyde led to the consid- (KOH) or reduction (LiAlH₄) yielded the benzenethiol li-

eration of the latter’s feasibility as a Cu^II-coordinating gand. This sequence of synthetic conversions is illustrated

agent capable of forming complexes that model type I cop- by crystallographic plots of molecules 3, 4, and

5

per properties. Facile reduction of the coordinated Cu^IIion (Scheme 1) in Figure 2.

by the benzenethiolate group of the 2-[bis(pyrazol-1Ј-yl)me-

The NewmanϪKwart rearrangement is normally per-

thyl]benzenethiol ligand was the most pressing considera- formed (i) in high-boiling solvents (diphenyl ether, benzal-

tion. The precedents set by Kitajama et al.^[64Ϫ67]and Tol- dehyde, anisole, etc.), (ii) as a ‘‘melt’’ in the absence of solv-

man, Holland et al.^[68Ϫ71]indicated that it would be neces- ent, (iii) in the vapour phase at atmospheric pressure by a

sary to introduce a high degree of steric hindrance onto flash pyrolysis process with the thiocarbamic acid O/S-ester

this ligand system. Reactions of 2-thiocyanatobenzaldehyde being transported through the furnace by an inert carrier

with sterically hindered bis(pyrazolyl) ketones were unsuc- gas.^[80]Neither of these methods worked satisfactorily for

cessful (yields of Յ 10% were typical after long workups). sterically substituted 2-[bis(pyrazol-1Ј-yl)methyl]phenol-

In addition, synthetic protocols for hindered analogs of 2- based thiocarbamic acid O-esters, in order: (i) primarily due

thiocyanatobenzaldehyde (e.g. with 3-tert-butyl substitu- to the limitations on the amount of material that could con-

tion) are not available. This necessitated the development of verted in a realistic (i.e. small) amount of solvent, (ii) due

a new, more flexible, approach to synthesizing these N₂S to rapid thermal decomposition of the melt, (iii) due to the

heteroscorpionate ligands, detailed in Schemes 1 and 2.

extremely low volatility of these compounds at atmospheric

3634

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Ligands to Model Type I Trigonal and Trigonal-Pyramidal Copper Protein Sites

FULL PAPER

Figure 2. Crystallographic plots (ORTEP, 30% thermal ellipsoids)

of molecules 3 (a), 4 (b), and 5 (c), illustrating the conversion of 2-

[bis(pyrazol-1Ј-yl)methyl]-6-tert-butyl-4-methylphenol to its ben-

zenethiol analog by NewmanϪKwart rearrangement

acid O-ester is volatilized at high temperature in vacuo and

passed through a quartz tube inserted into a furnace at very

high temperature (typically Ͼ 500°C). Conventional vapour

phase pyrolysis protocols often involve pre-vapourization of

the starting material before its transfer into the pyrolysis

vessel, this proved impractical with our thiocarbamic acid

O-esters, again due to their very low volatility, even under

vacuum, at ‘‘easily accessible’’ elevated temperatures (i.e. Յ

200 °C). Therefore, a modification to these conventional

procedures was developed whereby the starting thiocar-

bamic acid O-ester (typically 1.0Ϫ2.0 g) was packed into a

5-g weighing vial and packed down with glass wool. This

sample loading vial was inserted into the end of the quartz-

Scheme 2

pressure (thermal decomposition occurring long before va- vacuum tube which was then securely stoppered. The ap-

pourization). Thus, we have had to develop a variant of the paratus was then assembled at room temp. and placed un-

above vapour pyrolysis method wherein the thiocarbamic der vacuum (0.10 Torr) with the end of the quartz tube

Eur. J. Org. Chem. 2002, 3632Ϫ3645

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T. C. Higgs, C. J. Carrano

FULL PAPER

containing the loading vial outside of the tube furnace. The inhibit oligomerization of its Cu^IIcomplex through µ₂-S

tube furnace was then activated and heated to the required bridging, and more critically dimerization of RϪS^·radicals,

temperature (dependent on the steric bulk on the ligand) resulting from reductive homolysis of the Cu^IIϪSR bond,

and once the apparatus had thermally equilibrated the por- thereby increasing the redox solution stability of the com-

tion of the quartz tube containing the loading vial was in- plex. Furthermore the complementary 3-iPr substitution on

serted directly into the furnace (CAUTION! Care required!). the pyrazolyl rings will assist in constraining the coordina-

Under these low-pressure and high-temperature conditions tion number of the Cu^IIcomplex such as preventing ‘‘sand-

the O-ester rapidly volatilised, underwent the pyrolytic re- wich’’ complex formation^[72,73]and reducing exogenous

arrangement reaction and the product [and side product(s)] ligation (e.g. by solvent) therefore promoting the formation

condensed on the far side of the furnace (see Exp. Sect.). of low-coordination number complexes relevant to the

By careful control of these ‘‘Direct Insertion Vapour Phase redox-active centers in ‘‘blue’’ copper systems.

Flash Vacuum Pyrolysis’’ (DIVPFVP) conditions complete

An exogenous monodentate thioether ligand would be

reaction of the thiocarbamic acid O-ester can be achieved unlikely to insert into the coordination sphere of the Cu^II

with only two significant products, the desired thiocarbamic atoms coordinated to 6 or 11 N₂S-donor ligands due to

acid S-ester and 1H-pyrazole resulting from thermal de- the poor Cu^II-ligating ability of thioether-S.^[69]Therefore,

composition of the O/S ester (usually 20Ϫ40% depending to utilise these N₂S ligands for modeling N₂SS* ‘‘trigonal-

on temperature, higher temperatures leading to more de- pyramidal’’ type I ligation spheres, an ‘‘all-in-one’’ adaptive

gradation). The efficacy of this batch technique is a trade- strategy was required. The critical thioether-S* ligand was

off between the furnace temperature and the amount of incorporated into the pre-synthesized N₂S ligand to form

starting material converted per run. Greater steric bulk in an N₂SS* donor tetradentate asymmetric ‘‘distended’’ tri-

the starting material requires higher furnace temperatures pod ligand. This was achieved relatively simply as shown in

for complete rearrangement, but temperatures above 600 °C Scheme 2 by treating an appropriate 2-[bis(pyrazol-1Ј-yl)-

tended to lead to excessive thermal decomposition of the methyl]benzenethiol-based ligand (with 3,5-dimethyl- or 3-

thiocarbamic acid O-ester (predominantly in the loading isopropylpyrazole substitution and no thiophenol substitu-

vial). A solution to this development was to use smaller tion) with a vast excess of 1,2-dibromoethane thus con-

amounts of material at lower temperatures (Յ 1.0 g for verting the thiophenol to a thioether-S*, and subsequent

sterically very bulky O-esters such as 8). Using this method substitution of the 2-bromo group of the ethyl chain for a

this group has managed to synthesize numerous 2-[bis(pyra- lithiated 2-[(diphenylmethyl)thio]tetrahydropyran group.

zol-1Ј-yl)methyl]benzenethiol-based ligands with various Deprotection of the tetrahydropyran yielded a 3-(1,1-di-

permutations of alkyl substitition (Me, iPr, tBu) on the pyr- phenyl-1-mercaptopropane) chain tethered to the sulfanyl

1

azoles and/or the thiophenol. The ¹H NMR spectrum group of the tripod phenyl ring. The H NMR spectrum of

(CDCl₃) for the most hindered ligand synthesized to date the sterically most hindered ligand of this type synthesized

using

this

procedure,

2-[bis(3Ј-isopropylpyrazol-1Ј- to date, 28, is shown in Figure 4.

yl)methyl]-6-tert-butyl-4-methylbenzenethiol (11), is shown

in Figure 3.

These ‘‘all-in-one’’ N₂SS* ligands retain all the advant-

ages of the ligands employed for the Katjima et al.^[64Ϫ67]

Ligands 6 and 11, each of which possesses tert-butyl sub- and Holland, Tolman et al.^[68Ϫ71]type I copper protein

stitution on the carbon atom adjacent to the thiophenol model systems in that they possess a high degree of steric

group, both have N₂S donor capability, i.e. that observed hindrance (especially in the case of the 28 ligand), thus re-

in ‘‘trigonal’’ type I copper proteins. The tert-butyl group straining the adoption of higher coordination numbers (i.e.

‘‘covering’’ the ϪSH group of the ligand (Figure 1) should Ͼ 4). The sterically bulky 1,1-diphenyl substitution of the

1

Figure 3. 400 MHz H NMR spectrum of 11 in CDCl₃

3636

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Ligands to Model Type I Trigonal and Trigonal-Pyramidal Copper Protein Sites

FULL PAPER

1

Figure 4. 400 MHz H NMR spectrum of 28 in CDCl₃

ϪC(Ph)₂SH group should confer at least low-temperature ‘‘trigonal-pyramidal’’ copper proteins. We anticipate that

solution redox metastability, by inhibiting inter-radical such ligand systems or ones based upon them could

RϪS^·close contacts (these would favor disulfide formation complement the seminal work of Kitajima et al.^[64Ϫ67]and

concomitant with irreversible reduction of Cu^IIǞ Cu^I), Holland, Tolman et al.^[68Ϫ71]in elucidating structural-spec-

onto any Cu^IIcomplex of this N₂SS* ligand. The steric troscopic-redox relationships in this class of metalloprotein.

bulk, whilst still encouraging distorted coordination geo-

metries, is not as nearly great as that employed on the N-

donor ligands of the Holland, Tolman et al.^[68Ϫ71]systems

(2,6-diisopropylphenyl groups) and so should allow more

Experimental Section

‘‘open’’ tetraligated Cu^IIcoordination modes (i.e. less

General: All reactions were run under a blanket of argon, using

‘‘squashed’’ or ‘‘flattened’’). Furthermore, Cu^IIligation by

standard anaerobic protocols, unless stated otherwise. Anhydrous,

this thioether-S* ligand will be strongly encouraged due to

degassed THF was freshly distilled from sodium/benzophenone,

its internalized position within the N₂SS* ligand at the

‘‘trichelating pivot’’ of the ligand, able to form three simul-

taneous chelate rings when the the ligand is fully coordin-

whilst all other solvents were purchased from Aldrich Chemical

1

Co. and used as received. H NMR spectra were acquired using a

400 MHz Varian NMR spectrometer. All reagents were purchased

ated, two seven-membered rings with the pyrazole-N from Aldrich Chemical Company unless otherwise stated. The

400 MHz ¹H NMR spectroscopic characterizations are tabulated

in Tables 1Ϫ4, each one summarising a particular synthetic se-

quence. The analytical integrity of ligands synthesized by this flash

donors and one six-membered ring with the thiolate-S

donor. The incorporation of the thiolate ligand into a six-

membered chelate ring (with the thioether-S* donor via the

vacuum pyrolysis strategy was confirmed by Hammes, Carrano et

linking alkylpropyl chain) upon coordination should assist

al.^[28Ϫ31]through the Zn^IIcomplexes of their similarly synthesized

in enhancing the stability of its Cu^IIcomplex. Although the

L2SH ligand.

homolytic cleavage/formation equilibrium of the Cu^IIϪSR

bond will be unaffected, the resultant thiolate radical will

be effectively tethered to its ‘‘parent’’ Cu^Icomplex making

Ligand Syntheses

the long-range dissociation of RϪS^·from its ‘‘parent’’ Cu^I

(1) 2-[Bis(pyrazol-1Ј-yl)methyl]-6-tert-butyl-4-methylbenzenethiol

and subsequent radical disulfide coupling less facile.

(a) 3-tert-Butyl-2-hydroxy-5-methylbenzaldehyde (1): This was pre-

pared according to a previously published method.^[81]

Conclusion

(b) Bis(pyrazol-1-yl) Ketone (2): This synthesis has been previously

reported by this group.^[72]

Herein we have reported the hitherto unspecified syn-

thetic protocol used by Hammes, Carrano et al.^[28Ϫ31]to

produce ligands of the type L2SH which they have em-

ployed successfully in the elucidation of possible exogenous

thiolate alkylation reaction mechanisms of Zn^IIcomplexes.

Further to this we have described the extension of these

procedures to encompass sterically highly hindered tripodal

heteroscorpionate N₂S (11) and asymmetric ‘‘distended’’

tripod N₂SS* (28) ligands which possess identical ligand

(c) 2-[Bis(pyrazol-1Ј-yl)methyl]-6-tert-butyl-4-methylphenol (3): 1

(6.93 g, 0.036 mol), 2 (3.90 g, 0.024 mol), and a catalytic amount of

CoCl₂(anhydrous, 0.05 g) were placed together in a 100-mL round-

bottom flask fitted with a reflux condenser. The reaction mixture

was then steadily brought to a temperature of 90Ϫ120 °C during

which time a deep blue ‘‘melt’’ formed which effervesced due to

evolution of CO₂(g). The heating was continued for 90 min before

the reaction mixture was allowed to cool to room temp. The result-

donor arrays to those observed in type I ‘‘trigonal’’ and ant thick, viscous dark blue oil was dissolved in CH₂Cl₂(70 mL)

Eur. J. Org. Chem. 2002, 3632Ϫ3645 3637

T. C. Higgs, C. J. Carrano

Table 2. Proposed H NMR assignments for 8, 9, 10, 11; chemical

FULL PAPER

Table 1. Proposed ¹H NMR assignments for 3, 4, 5, 6; chemical

1

shifts δ in ppm; ambiguous assignments are represented by merging shifts δ in ppm; ambiguous assignments are represented by merging

the cells of the possible contributors; for overlapping peaks the

individual components are deconvoluted in the table (dd: doublet

of doublets; td: triplet of doublets; bs: broad singlet; qd: quadruplet

of doublets; cm: complex multiplet)

the cells of the possible contributors; for overlapping peaks the

individual components are deconvoluted in the table (dd: doublet

of doublets; td: triplet of doublets; bs: broad singlet; qd: quadruplet

of doublets; cm: complex multiplet)

[a]

3 phenolic, ϪOH, a singlet at δ ϭ 11.52 ppm. ^[b]6 thiophenolic,

^[a]8 phenolic, ϪOH, a singlet at δ ϭ 12.33 ppm. ^[b]11 thiophenolic,

[c]

ϪSH, a singlet at δ ϭ 2.68 ppm.

stereotopic protons.

Carbon atom possesses dia-

[c]

ϪSH, a singlet at δ ϭ 2.78 ppm.

Carbon atom possesses dia-

stereotopic protons.

and this was extracted with distilled water (2 ϫ 200 mL). The

CH₂Cl₂layer was separated and dried with anhydrous MgSO₄, and

the solution subsequently concentrated to dryness in a rotary evap-

orator yielding a yellow oil which was distilled under a reduced

pressure removing approximately 3.2 g of 1 (which was recycled for

use in later syntheses). The brown residual oil in the distillation

flask was dissolved in CH₂Cl₂(20 mL) and n-hexane (30 mL) ad-

ded, this solution was then transferred to a crystallising dish and

the product was allowed to crystallise as large colorless crystals (X-

ray quality). These were collected by filtration, washed with n-hex-

IR (KBr disk): ν˜ ϭ 3050 s, 2947 s, 2721 s, 2618 s, 1778 w, 1727 w,

1064 w, 1515 m, 1478 s, 1435 s, 1384 s, 1356 w, 1304 s, 1252 s, 1210

s, 1149 m, 1087 s, 1050 s, 975 m, 914 w, 834 m, 806 s, 754 s, 707 s,

651 w, 614 w, 561 w, 528 w, 467 w cm^Ϫ1

.

(d)

O-{2-[Bis(pyrazol-1Ј-yl)methyl]-6-tert-butyl-4-methylphenyl}-

N,N-dimethylthiocarbamate (4): 3 (3.60 g, 11.6ϫ10^Ϫ3mol), pyridine

(0.94 mL, 11.6ϫ10^Ϫ3mol), and dimethylthiocarbamoyl chloride

(1.51 g, 0.122 mol) were dissolved in o-xylene (75 mL) in a 250-

mL round-bottom flask fitted for reflux. The reaction mixture was

brought to reflux for 24 h before being allowed to cool to room

temp. and the solvents were evaporated to dryness under a reduced

pressure. The resultant residual green oil was dissolved in CH₂Cl₂

(50 mL) and this solution washed with distilled water (3 ϫ 300

mL). The organic layer was separated and dried with anhydrous

MgSO₄and then concentrated to dryness in a rotary evaporator

yielding a yellow oil which was triturated with n-hexane (30 mL).

This material recrystallized from CH₂Cl₂/n-hexane to yield yellow

blocks of the product (of crystallographic quality). Yield: 3.3 g

1

ane (10 mL) and dried in vacuo. Yield: 4.1 g (42%), subsequent H

NMR spectroscopy and crystal structure analysis of this material

indicated a significant component of it was (pz)₂CH[3-tBu-5-Me-

PhOC(ϭO)pz] (3e), an ester of the desired phenol ligand. 3e

(4.00 g, 9.90ϫ10^Ϫ3mol) and KOH (3.00 g, 0.053 mol) were re-

fluxed in MeOH (80 mL) for 1 h. The volume of the reaction mix-

ture was then reduced to about 15 mL under a reduced pressure

and distilled water (80 mL) then added. The solution was then

acidified to pH ϭ 6 with 6  HCl(aq) which caused a dense white

microcrystalline solid to deposit. This water/MeOH/3 suspension

was extracted with CH₂Cl₂(40 mL) and the organic layer was sep-

arated and washed with distilled water (2 ϫ 100 mL). The CH₂Cl₂

layer was then separated and dried with anhydrous MgSO₄, n-hex-

ane (100 mL) was added and the volume of the solution was re-

duced to about 20 mL in a rotary evaporator. The solution was

transferred to a crystallising dish and overnight large colorless

1

(71%). H NMR (400 MHz, CDCl₃): Table 1. IR (KBr disk): ν˜ ϭ

3396 w, 3123 w, 2965 m, 2860 m, 1632 s, 1432 s, 1390 s, 1286 s,

1207 s, 1170 s, 1133 s, 1086 s, 1044 s, 970 m, 913 w, 865 w, 807 m,

744 s, 708 m, 655 m, 613 m, 503 w, 445 w cm^Ϫ1

.

(e)

S-{2-[Bis(pyrazol-1Ј-yl)methyl]-6-tert-butyl-4-methylphenyl}-

N,N-dimethylthiocarbamate (5): 4 (0.20 g, 5.04ϫ10^Ϫ4mol) was con-

block crystals developed (of crystallographic quality), these were verted into 5 by ‘‘direct insertion vapour phase flash vacuum pyro-

collected by filtration, washed with n-hexane (5 mL) and dried in lysis’’ (DIVPFVP) at a furnace temperature of 550 °C (0.10 Torr).

vacuo. Yield: 2.4 g (78%). ¹H NMR (400 MHz, CDCl₃): Table 1. The product condensed on the far side of the furnace as a yellow/

3638

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Ligands to Model Type I Trigonal and Trigonal-Pyramidal Copper Protein Sites

FULL PAPER

1

Table 3. Proposed H NMR assignments for 14, 15, Li[16]·THF, 17, 20, 21; all spectra were run in CDCl₃except for Li[16]·THF which

was run in CD₃OD; chemical shifts δ in ppm; ambiguous assignments are represented by merging the cells of the possible contributors;

for overlapping peaks the individual components are deconvoluted in the table (dd: doublet of doublets; td: triplet of doublets; bs: broad

singlet; qd: quadruplet of doublets; cm: complex multiplet)

[a]

[b]

THF solvate: δ ϭ 1.89 (m, ca. 3 H, 2,3-CH₂of THF), 3.75 (m, ca. 3 H, 1,4-CH₂of THF) ppm. Carbon atom possesses diastereo-

topic protons.

orange glass (further along the quartz tube, dendritic clumps of

white crystals of pyrazole condensed). The condensate was re-

moved from the quartz glass tube by solvation with CH₂Cl₂(100

mL), this solution was then concentrated to dryness in a rotary

evaporator. The ¹H NMR (400 MHz, CDCl₃) of the crude product

indicated the only significant contaminant was free pyrazole, re-

sulting from thermal decomposition. After five more pyrolysis runs

of 4, the batches of crude product were then combined and dis-

solved in CH₂Cl₂(30 mL) and this was extracted with distilled

(f)

2-[Bis(pyrazol-1Ј-yl)methyl]-6-tert-butyl-4-methylbenzenethiol

(6): 5 (0.50 g, 1.26ϫ10^Ϫ3mol) and KOH (0.71 g, 25.2ϫ10^Ϫ3mol)

were dissolved together in MeOH (30 mL), in a 100-mL round-

bottom flask fitted for reflux, forming a deep yellow solution. The

apparatus was purge-filled with Ar(g) three times and then the reac-

tion mixture brought to reflux for 18 h. The reaction mixture was

then allowed to cool to room temp. and was halved in volume using

a rotary evaporator. Distilled water (80 mL) was then added to

the remaining methanolic solution, and this mixture was carefully

water (3 ϫ 75 mL) to remove the pyrazole impurity. The organic acidified to pH ϭ 6 using 6  HCl(aq) causing the solution color

layer was separated and dried with anhydrous MgSO₄, the dried

CH₂Cl₂solution was then concentrated to dryness in a rotary evap-

orator yielding a yellow glass which was recrystallized from a

CH₂Cl₂/n-hexane solution (1:3) to yield colorless block crystals of

the 5 (of crystallographic quality); these were collected, washed

to fade from deep yellow to colorless and for a white solid to pre-

cipitate. The resulting suspension was extracted into CH₂Cl₂(30

mL) and this layer was washed with distilled water (100 mL). The

organic layer was separated and dried with anhydrous MgSO₄and

then this CH₂Cl₂solution concentrated to dryness in a rotary evap-

with n-hexane (5 mL) and dried in vacuo. Yield: 0.12Ϫ0.14 g orator yielding a pale yellow, slightly oily ‘‘foam’’. This was tritur-

(60Ϫ70%) per batch. ¹H NMR (400 MHz, CDCl₃): Table 1. IR

ated with n-hexane (10Ϫ15 mL) to yield a very pale yellow micro-

(KBr disk): ν˜ ϭ 3434 w, 3103 w, 2960 m, 2926 m, 1680v s, 1587 w, crystalline solid which was collected by filtration, washed with n-

1

1515 w, 1449 m, 1432 m, 1393 s, 1355 s, 1305 m, 1259 s, 1167 w, hexane (5 mL) and dried in vacuo. Yield: 0.30 g (73%). H NMR

1085 s, 1040 s, 980 w, 958 w, 919 m, 831 w, 809 s, 798 w, 765 s, 743 (400 MHz, CDCl₃): Table 1. IR (KBr disk): ν˜ ϭ 3406 w, 3108 m,

s, 704 m, 685 m, 657 m, 616 w, 553 w cm^Ϫ1

.

2959 s, 2919 s, 2870 s, 2562 w, 2235 w, 1770 w, 1716 w, 1602 m,

3639

Eur. J. Org. Chem. 2002, 3632Ϫ3645

T. C. Higgs, C. J. Carrano

FULL PAPER

1

Table 4. Proposed H NMR assignments for 23, 24, 25, 26, 27, 28; chemical shifts δ in ppm; all spectra were run in CDCl₃; ambiguous

assignments are represented by merging the cells of the possible contributors; for overlapping peaks the individual components are

deconvoluted in the table (sp: septuplet; dd: doublet of doublets; td: triplet of doublets; bs: broad singlet; qd: quadruplet of doublets;

cm: complex multiplet)

[a]

[b]

25 thiol, ϪSH, singlet at δ ϭ 3.195 ppm. Carbon atom possesses diastereotopic protons.

1512 s, 1477 m, 1428 s, 1388 s, 1363 m, 1320 s, 1254 s, 1214 m,

1195 m, 1165 m, 1088 s, 1046 s, 971 m, 917 s, 823 s, 807 s, 748 s,

dryness in a rotary evaporator yielding a viscous yellow oil. This

oil was dissolved in n-hexane (100 mL) and this too was then con-

centrated to dryness under a reduced pressure (to remove off any

residual CH₂Cl₂azeotropically). The resultant yellow oil was dis-

solved in pentane (100Ϫ150 mL) and this placed in a freezer for

several days at Ϫ30 °C during which time a microcrystalline very

pale yellow solid precipitated. This solid was collected on a cold

sinter and washed with cold (Ϫ30 °C) pentane (2 ϫ 20 mL) and

then dried in vacuo. Further batches of material could be obtained

by sequentially reducing the volume of the pentane solution and

cooling it to Ϫ30 °C to effect crystallization of the product. Yield:

698 s, 654 s, 614 m cm^Ϫ1

.

(2) 2-[Bis(3Ј-isopropylpyrazol-1Ј-yl)methyl]-6-tert-butyl-4-methyl-

benzenethiol

(a) Bis(3-isopropylpyrazol-1-yl) Ketone (7): This synthesis has been

previously described by this group.^[72]

(b)

2-[Bis(3Ј-isopropylpyrazol-1Ј-yl)methyl]-6-tert-butyl-4-methyl-

phenol (8): 1 (24.0 g, 0.125 mol), 7 (30.5 g, 0.125 mol), and CoCl₂

(anhydrous, ca. 0.50 g) were mixed together in a 150-mL round-

bottom flask fitted with a reflux condenser and for magnetic stir-

1

24.0 g (49%). H NMR (400 MHz, CDCl₃): Table 2.

ring. This reaction mixture was brought to a temperature of 120 (c) O-{2-[Bis(3Ј-isopropylpyrazol-1Ј-yl)methyl]-6-tert-butyl-4-meth-

°C for 96 h [during which time a dark green-blue ‘‘melt’’ formed ylphenyl}-N,N-dimethylthiocarbamate (9): NaH (0.66 g, 0.027 mol)

from which very slow evolution of CO₂(g) was observed]. The was placed in a 3-necked 500-mL round-bottom flask, under Ar(g),

course of the reaction was monitored by taking small aliquots (ca.

0.10 g) of the reaction mixture out at 24-h intervals and elucidating

which was fitted with both a 250-mL pressure-equalising addition

funnel and a reflux condenser. In a separate 250-mL round-bottom

its contents by ¹H NMR (CDCl₃). After 96 h, the reaction was flask, 8 (5.42 g, 13.7ϫ10^Ϫ3mol) was dissolved in THF (150 mL)

considered to have proceeded to a satisfactory point (90ϩ% conver-

sion), and the mixture was allowed to cool to room temp. The

‘‘melt’’ was then dissolved in CH₂Cl₂(200 mL) and this solution

extracted with distilled water (2 ϫ 300 mL). The CH₂Cl₂layer was

separated, dried with anhydrous MgSO₄, and then concentrated to

under Ar(g). The 8_THFsolution was then cannulated into the addi-

tion funnel and then added dropwise to the NaH [Caution: H₂(g)

evolution as Na[20] formed]. Separately, in a 150-mL round-bottom

flask, dimethylthiocarbamoyl chloride (1.87 g, 15.1ϫ10^Ϫ3mol) was

dissolved in THF (40 mL). This solution was cannulated into the

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Eur. J. Org. Chem. 2002, 3632Ϫ3645

Ligands to Model Type I Trigonal and Trigonal-Pyramidal Copper Protein Sites

FULL PAPER

addition funnel of the above apparatus. The dimethylthiocarba-

with THF (30 mL), then water (2 ϫ 30 mL). These filtrates were

moyl chloride/THF solution was then added dropwise to the combined and concentrated to a volume of about 60 mL thus re-

Na[8]_THFsolution over a period of 15 min, the reaction mixture

was then refluxed for 3 h, during which time the mixture turned

green and an NaCl precipitate formed. The reaction mixture was

allowed to cool to room temp. before the NaCl/excess NaH col-

lected by filtration in air and washed with THF (20 mL). The yel-

low filtrate was concentrated to dryness in a rotary evaporator

yielding a yellow oil which was dissolved in CH₂Cl₂(65 mL). This

CH₂Cl₂solution was extracted with two portions of saturated brine

(2 ϫ 150 mL) and the organic layer then separated and dried with

anhydrous MgSO₄. The CH₂Cl₂solution was then concentrated in

a rotary evaporator leaving a thick yellow oil which was dried in

vacuo. This oil was dissolved in a minimum volume of n-hexane

(10 mL) and purified by dry flash column chromatography [TLC-

grade silica gel w/o binder, CH₂Cl₂/EtOAc (2% increments of

EtOAc), fractions monitored by ¹H NMR]. The pure fractions were

combined and the solvents evaporated to dryness yielding a color-

moving most of the THF and causing a yellow oil to deposit. The

solution was then acidified to pH ϭ 5.5 by dropwise addition of

6  HCl(aq) before being extracted with CH₂Cl₂(2 ϫ 50 mL). The

CH₂Cl₂extracts were combined and dried with anhydrous MgSO₄.

The CH₂Cl₂solution was then concentrated to dryness in a rotary

evaporator yielding a yellow oil which was dissolved in a minimum

volume of n-hexane (5 mL) and then purified by dry flash column

chromatography [TLC-grade silica gel w/o binder, CH₂Cl₂/EtOAc

(2% increments of EtOAc), fractions monitored by ¹H NMR]. The

fractions containing pure 11 were combined and dried in vacuo.

Appearance: very viscous pale yellow oil. Yield: 1.4 g (82%). ¹H

NMR (400 MHz, CDCl₃): Table 2.

(3) 3-{2Ј-[Bis(3ЈЈ,5ЈЈ-dimethylpyrazol-1ЈЈ-yl)methyl]phenylsulfanyl}-

1,1-diphenylpropane-1-thiol

(a) Bis(3,5-dimethylpyrazol-1-yl) Ketone (12): This synthesis has

1

been previously described by this group.^[72]

less viscous oil. Yield: 5.1 g (77%). H NMR (400 MHz, CDCl₃):

Table 2.

(b) 2-[Bis(3Ј,5Ј-dimethylpyrazol-1Ј-yl)methyl)]phenol (13): The pro-

(d) S-{2-[Bis(3Ј-isopropylpyrazol-1Ј-yl)methyl]-6-tert-butyl-4-meth-

cedure for this synthesis has been described by this group in a previ-

ylphenyl}-N,N-dimethylthiocarbamate (10): 9 (Յ 1.00 g, 2.09ϫ10^Ϫ3ous publication.^[72]

mol) was converted into 10 by DIVPFVP at a furnace temperature

(c)

O-{2-[Bis(3Ј,5Ј-dimethylpyrazol-1Ј-yl)methyl]phenyl}-N,N-di-

of 570 °C (0.10 Torr), smaller batches of material than usual

(2.00 g) were allowed to react to limit thermal decomposition. The

product condensed on the far side of the furnace as a black tar

[further along the quartz tube, a yellow oil condensed which was

predominantly 3-isopropylpyrazole (3-iPrpz)]. The condensate was

removed from the quartz glass tube by solvation with CH₂Cl₂(100

mL), this solution was then concentrated to dryness in a rotary

evaporator. After ten pyrolysis runs of 9, the batches of crude prod-

uct were then combined and most of the 3-iPrpz contamination

distilled off in vacuo (0.1 Torr) using a kugelrohr over a temper-

ature range of 100Ϫ120 °C. The residual black tar was dissolved

in MeOH (150 mL) and this solution decolorized using activated

charcoal. The solution was then concentrated to dryness in a rotary

evaporator yielding a brown oil, this was then dried in vacuo. This

material was then dissolved in a minimum volume of n-hexane (10

mL) and purified by dry flash column chromatography [TLC-grade

silica gel w/o binder, CH₂Cl₂/EtOAc (2% increments of EtOAc),

fractions monitored by ¹H NMR], the fractions containing pure 10

were combined and dried in vacuo yielding a pale yellow oil. Yields:

0.40Ϫ0.60 g (40Ϫ60%) per batch. ¹H NMR (400 MHz, CDCl₃):

Table 2.

methylthiocarbamate (14): The same procedure as used to synthes-

ise 9 was applied using 13 (20.0 g, 0.0676 mol), NaH (1.95 g,

81.1ϫ10^Ϫ3mol), dimethylthiocarbamoyl chloride (8.77 g,

70.9ϫ10^Ϫ3mol). The final product was triturated with Et₂O to

yield a pale yellow solid. Yield: 18.6 g (72%). ¹H NMR (400 MHz,

CDCl₃): Table 3.

(d)

S-{2-[Bis(3Ј,5Ј-dimethylpyrazol-1Ј-yl)methyl]phenyl}-N,N-di-

methylthiocarbamate (15): 14 (2.00 g, 5.22ϫ10^Ϫ3mol) was con-

verted into 15 by DIVPFVP at 520 °C (0.1 Torr). Ten batches of 5

were subjected to DIVPFVP, the batches of crude 15 were com-

bined and distilled under vacuum (0.10 Torr) using a kugelrohr, to

remove the 3,5-dimethylpyrazole contamination, over a temper-

ature range of 60Ϫ120 °C, the 3,5-dimethylpyrazole distillate con-

densing as dendritic clumps of white crystals. The dark brown oily

residue from the distillation was dissolved in MeOH (150 mL) and

decolorized with activated charcoal. After removal of the charcoal,

the solution was concentrated to dryness in a rotary evaporator

yielding a light yellow-orange oil was dried in vacuo. This material

was used in the next step of the synthesis without further purifica-

tion. Yield: 1.2Ϫ1.4 g (60Ϫ70%) per batch. ¹H NMR (400 MHz,

CDCl₃): Table 3.

(e)

2-[Bis(3Ј-isopropylpyrazol-1Ј-yl)methyl]-6-tert-butyl-4-methyl-

benzenethiol (11): 10 (2.00 g, 4.17ϫ10^Ϫ3mol) was dissolved in THF (e) Lithium 2-[Bis(3Ј,5Ј-dimethylpyrazol-1Ј-yl)methyl]benzenethio-

(50 mL) in a 150-mL round-bottom flask. Separately, 1.0  LiAlH₄

(7.00 mL, 7.00ϫ10^Ϫ3mol) was transferred via gastight syringe into

a 500-mL 3-necked round-bottom flask, fitted with both a water

condenser and a 150-mL pressure-equalising addition funnel. The

late؊THF Solvate {Li[16]·THF}: 15 (12.2 g, 31.9ϫ10^Ϫ3mol) was

placed into a 250-mL round-bottom flask under Ar(g). THF (100

mL) was added, and the flask was shaken until 15 had dissolved.

Separately, 1.0  LiAlH₄(32.0 mL, 31.9ϫ10^Ϫ3mol) was transferred

10_THFsolution was then cannulated into the addition funnel and (using a gas-tight 50-mL syringe) into a 1-L 3-necked round-bot-

its flask washed out with a further portion of THF (50 mL) into

the addition funnel. The 10_THFsolution was then added dropwise

to the 1.0  LiAlH₄solution over a period of 20 min during which

time the reaction mixture turned to a deep yellow color. Following

the addition, the reaction mixture was refluxed for 3 h before being

tom flask fitted with both a water condenser and a 250-mL pres-

sure-equalising addition funnel. The 6_THFsolution was cannulated

into the addition funnel and the flask was washed out with a fur-

ther portion of THF (50 mL) into the addition funnel. The 15_THF

solution was then added dropwise to the 1.0  LiAlH₄solution

allowed to cool to room temp. The reaction mixture at this point over a period of 20 min. As the addition of 15_THFproceeded the

contained a large excess of LiAlH₄, this was destroyed by careful

dropwise addition of first a 10% NaOH(aq) solution (1 mL), then

distilled water (1 mL; Caution: foaming due to H₂evolution!). The

reaction mixture turned deep orange in color and started refluxing

due to the highly exothermic reduction reaction. After several mi-

nutes, a white microcrystalline solid started to precipitate from the

{Al(OH)₃}_nthat precipitated from the reaction mixture after reaction mixture (Li[16]·THF). After completion of the addition,

quenching was collected by filtration through Celite and washed

the reaction mixture was refluxed for a further 90 min before being

3641

Eur. J. Org. Chem. 2002, 3632Ϫ3645

T. C. Higgs, C. J. Carrano

FULL PAPER

allowed to cool to room temp. The white solid was collected by

filtration under Ar(g), using standard Schlenk line techniques,

washed with THF (2 ϫ 30 mL), and dried in vacuo. The THF

(j) 3-{2Ј-[Bis(3ЈЈ,5ЈЈ-dimethylpyrazol-1ЈЈ-yl)methyl]phenylsulfanyl}-

1,1-diphenylpropane-1-thiol (21): 20 (2.75 g, 4.41ϫ10^Ϫ3mol) was

dissolved in MeOH (40 mL). Separately, AgNO₃(1.65 g,

filtrate containing the excess LiAlH₄was then destroyed by careful 9.70ϫ10^Ϫ3mol) was dissolved in MeOH (100 mL)/pyridine (10 mL).

dropwise addition of H₂O (Caution: foaming due to H₂evolution!).

Yield: 7.3 g (59%). H NMR (400 MHz, CD₃OD): Table 3.

The flask containing the 20_MeOHsolution was wrapped with Al

foil to completely eliminate all light and the AgNO_{3(MeOH/pyridine)}

solution was added. The reaction mixture was stirred for 24 h at

room temp. in the dark. After 24 h, a white microcrystalline solid

had precipitated from the solution, it was collected by filtration,

washed with MeOH (2 ϫ 10 mL) and air-dried on the sinter to

give approximately 3.0 g of material. This solid was dissolved in

pyridine (5 mL), then diluted with CH₂Cl₂(100 mL) to give a

brown solution, and H₂S [generated by dropwise addition of 6 

HCl(aq) to pellets of K₂S] bubbled through this solution for 20 min

causing the dense precipitation of black [Ag₂S]_n. The [Ag₂S]_nwas

then removed by filtration, and the filtrate concentrated to dryness

using a kugelrohr. This yielded a viscous yellow oil which was dis-

solved in CH₂Cl₂(40 mL), then extracted with distilled water (3 ϫ

100 mL). The CH₂Cl₂layer was separated, dried with anhydrous

MgSO₄and then concentrated in a rotary evaporator to yield a

pale yellow oil which was triturated to a white microcrystalline

solid with n-hexane/diethyl ether. Yield: 1.7 g (73%). ¹H NMR

(400 MHz, CDCl₃): Table 3.

1

(f) 1-{2Ј-[Bis(3ЈЈ,5ЈЈ-dimethylpyrazol-1ЈЈ-yl)methyl]phenylsulfanyl}-

2-bromoethane (17): Li[16]·THF (0.50 g, 1.28ϫ10^Ϫ3mol) was dis-

solved in 1,2-dibromoethane (21.0 g, 0.11 mol)/MeOH (10 mL) in

a 100-mL round-bottom flask fitted with a reflux condenser, this

mixture was brought to reflux for 15 min. After cooling to room

temp., the MeOH component of the reaction solvents was evapor-

ated under a reduced pressure, this resulted in the precipitation of

LiBr as a white microcrystalline solid. CH₂Cl₂(20 mL) was added

and the mixture extracted with distilled water (2 ϫ 100 mL) to

remove the LiBr. The organic layer was separated and dried with

anhydrous MgSO₄. The solvents (CH₂Cl₂, BrCH₂CH₂Br) were

then distilled off under vacuum yielding a pale yellow solid in the

distillation flask. This solid was recrystallised from CH₂Cl₂/n-hex-

ane, crystallising as small pale yellow blocks. Yield: 0.47 g (87%).

¹H NMR (400 MHz, CDCl₃): Table 3.

(g) Diphenylmethanethiol (18). This synthesis of this compound was

achieved by the published procedure of Archer et al.^[82]

(4) 3-{2Ј-[Bis(3ЈЈ-isopropylpyrazol-1ЈЈ-yl)methyl]phenylsulfanyl}-1,1-

diphenylpropane-1-thiol

(h) 2-[(Diphenylmethyl)thio]tetrahydropyran (19): The method of

Berg and Holm was utilized to effect synthesis of this com-

pound.^[83]

(a) 2-[Bis(3Ј-isopropylpyrazol-1Ј-yl)methyl]phenol (22): The proced-

ure for this synthesis has been described by this group in a previ-

ous publication.^[72]

(i)

2-(3Ј-{2ЈЈ-[Bis(3ЈЈЈ,5ЈЈЈ-dimethylpyrazol-1ЈЈЈ-yl)methyl]phenyl-

(b) O-{2-[Bis(3Ј-isopropylpyrazol-1Ј-yl)methyl]phenyl}-N,N-dimeth-

ylthiocarbamate (23): The same procedure as used to synthesise 9

was applied using 22 (20.0 g, 61.7ϫ10^Ϫ3mol), NaH (1.93 g, 0.080

mol), dimethylthiocarbamoyl chloride (8.77 g, 0.0709 mol). The fi-

nal product was triturated with n-hexane to yield a slightly ‘‘sticky’’

yellow solid. Yield: 19.3 g (76%). ¹H NMR (400 MHz, CDCl₃):

Table 4.

sulfanyl}-1Ј,1Ј-diphenylpropyl-1Ј-sulfanyl)tetrahydropyran (20): 19

(2.00 g, 7.04ϫ10^Ϫ3mol) was dissolved in THF (100 mL) in a 1-L

2-necked round-bottom flask fitted with a 250-mL pressure-

equalising addition funnel. The solution was then cooled to Ϫ78

°C using a dry-ice/acetone bath and 1.6  nBuLi in n-hexane (4.40

mL, 7.04ϫ10^Ϫ3mol) was added using a Hamilton 5.0-mL gas-tight

syringe. Upon addition of the nBuLi the color of the solution

changed from colorless to deep orange. The solution was stirred

at Ϫ78 °C for 30 min during which time a microcrystalline solid

precipitated from the solution (presumably Li[19]). The mixture

was then allowed to warm to room temp. over a period of about

1 h, as the mixture neared room temp. the solid re-dissolved once

again forming a deep orange solution. Separately, 17 (1.48 g,

3.52ϫ10^Ϫ3mol) was dissolved in THF (50 mL) in a 100-mL round-

bottom flask and this solution was cannulated into the addition

funnel of the above apparatus. This 8_THFsolution was then added

dropwise to the Li[19]_THFsolution (at room temp.) over a period

of 1 h. During the addition the color of the reaction mixture

darkened to a brown color. After completion of the addition, the

reaction mixture was stirred at room temp. for a further 30 min

before being quenched with distilled water (5 mL) which caused

the color of the solution to change to yellow. The solution was then

concentrated to dryness in a rotary evaporator yielding a yellow

oil, this was dissolved in CH₂Cl₂(60 mL) and extracted with dis-

tilled water (2 ϫ 150 mL). The CH₂Cl₂layer was separated and

dried with anhydrous MgSO₄, then concentrated to dryness. The

residual yellow oil was dissolved in a minimum amount of n-hexane

(20Ϫ30 mL) and then purified by dry flash column chromato-

graphy [TLC-grade silica gel w/o binder, CH₂Cl₂/EtOAc (2% incre-

ments of EtOAc), fractions monitored by ¹H NMR]. The fractions

containing the desired product were combined and the solvents

evaporated to dryness yielding a ‘‘foam’’ which was broken up to

give a white solid. Yield: 2.0 g (92%). ¹H NMR (400 MHz,

CDCl₃): Table 3.

(c) S-{2-[Bis(3Ј-isopropylpyrazol-1Ј-yl)methyl]phenyl}-N,N-dimeth-

ylthiocarbamate (24): 23 (2.00 g, 4.86ϫ10^Ϫ3mol) was converted

into 24 by DIVPFVP at a furnace temperature of 580 °C (0.10

Torr). The product condensed on the far side of the furnace as a

very dark brown oil (further along the quartz tube, a yellow oil

condensed which was predominantly 3-iPrpz). The condensate was

removed from the quartz glass tube by solvating it with CH₂Cl₂

(150 mL), and this solution was concentrated to dryness in a rotary

evaporator. Ten batches of 23 were subjected to DIVPFVP. The

batches of crude product were then combined and distilled under

vacuum (0.10 Torr) using a kugelrohr, to remove most of the 3-

iPrpz contamination, over a temperature range of 100Ϫ120 °C, the

3-iPrpz distillate condensing as a yellow oil. The residual dark

brown oil from the distillation was dissolved in MeOH (150 mL)

and decolorized with activated charcoal. The solution was then

concentrated yielding a pale brown oil which was then dried in

vacuo. This material was then purified further by dry flash column

chromatography [TLC-grade silica gel w/o binder, CH₂Cl₂/EtOAc

(2% increments of EtOAc), fractions monitored by ¹H NMR]. The

pure fractions were combined, and concentrated yielding a viscous

yellow oil. Yield: 1.0Ϫ1.4 g (50Ϫ70%) per batch. ¹H NMR

(400 MHz, CDCl₃): Table 4.

(d) 2-[Bis(3Ј-isopropylpyrazol-1Ј-yl)methyl]benzenethiol (25): The

same LiAlH₄reduction procedure as used to synthesise 10 was ap-

plied using 24 (12.5 g, 0.030 mol) and LiAlH₄(1.0  in THF, 30.4

mL, 0.026 mol). The crude product, a yellow oil, was purified by

3642

Eur. J. Org. Chem. 2002, 3632Ϫ3645

Ligands to Model Type I Trigonal and Trigonal-Pyramidal Copper Protein Sites

dry flash column chromatography [TLC-grade silica gel w/o binder, thesis of 27 (from 26 and 19) was exactly the same as that of 20

FULL PAPER

CH₂Cl₂/EtOAc (2% increments of EtOAc), fractions monitored by (from 17 and 19), substituting 26 for 17 and using the following

¹H NMR]. The fractions containing pure 25 were combined and

reactant ratios: 19 (3.43 g, 0.0121 mol), 1.6  nBuLi in n-hexane

dried in vacuo. Appearance: viscous pale yellow oil. Yield: 8.3 g

(7.55 mL, 0.0121 mol), and 26 (2.70 g, 6.04ϫ10^Ϫ3mol). Yield: 1.7 g

1

(80%). H NMR (400 MHz, CDCl₃): Table 4.

(43%). H NMR (400 MHz, CDCl₃): Table 4.

(e) 1-{2Ј-[Bis(3ЈЈ-isopropylpyrazol-1ЈЈ-yl)methyl]phenylsulfanyl}-2-

bromoethane (26): NaH (1.27 g, 0.053 mol) was placed in a two-

necked 500-mL round-bottom flask fitted with a 150-mL pressure-

equalising addition funnel. Separately, 25 (12.0 g, 0.035 mol) was

dissolved in THF (100 mL) in a 250-mL round-bottom flask, and

this was cannulated into the addition funnel. The 25_THFwas added

(g) 3-{2Ј-[Bis(3ЈЈ-isopropylpyrazol-1ЈЈ-yl)methyl]phenylsulfanyl}-1,1-

diphenylpropane-1-thiol (28): The same procedure as used to synth-

esise 21 was applied using 27 (1.80 g, 2.77ϫ10^Ϫ3mol) and AgNO₃

(1.04 g, 6.09ϫ10^Ϫ3mol) in MeOH (60 mL)/pyridine (10 mL). The

resultant Ag^Ithiolate complex was again cleaved using H₂S(g)

1

[from K₂S ϩ 6  HCl(aq)]. The H NMR spectrum of the crude

dropwise to the NaH [Caution: effervescence due to H₂(g) genera- product indicated a 9:1 ratio of 28/27. This material was purified by

tion]. Upon completion of the addition, the mixture was stirred for

a further 20 min at room temp. before the excess NaH was collected

by filtration. The deep yellow filtrate was concentrated to dryness

under a reduced pressure, then MeOH (5 mL)/dibromoethane

(48.45 g, 0.258 mol) was added to the resulting yellow ‘‘foam’’. The

yellow material quickly dissolved, and this reaction mixture was

then refluxed for 25 min. After cooling to room temp., the MeOH

was removed in a rotary evaporator, causing the precipitation of

LiBr, CH₂Cl₂(50 mL) was then added and the mixture extracted

with distilled water (2ϫ150 mL) to remove the LiBr. The organic

layer was separated and dried with anhydrous MgSO₄before being

concentrated to dryness under vacuum to yield the crude product,

a yellow oil. This material was purified by dry flash column chro-

matography [TLC-grade silica gel w/o binder, CH₂Cl₂/EtOAc (2%

increments of EtOAc), fractions monitored by ¹H NMR]. The pure

fractions of 26 from the dry flash chromatography were combined,

dried in vacuo and then recrystallized from n-hexane, pure 26 was

isolated as a pale yellow solid. Yield: 7.2 g (46%). ¹H NMR

(400 MHz, CDCl₃): Table 4.

dry flash column chromatography [TLC-grade silica gel w/o binder,

CH₂Cl₂/EtOAc (2% increments of EtOAc), 28 eluted first, fractions

monitored by ¹H NMR]. The pure fractions were combined and

the solvents evaporated to dryness yielding a very viscous pale yel-

low oil [Note: air-sensitive material, stored under Ar(g)]. Yield:

1

1.1 g (71%). H NMR (400 MHz, CDCl₃): Table 4.

Crystallography: Crystals of 3, 4, and 5 were all grown by solvent

evaporation of CH₂Cl₂/n-hexane solutions of the respective com-

pounds (see Exp. Sect., vide supra). For all three compounds a

crystals was glued to a glass fiber with epoxy resin and mounted

onto a goniometer head and this loaded into a Siemens P4 dif-

fractometer, with a sealed-tube Mo-K_αX-ray source (λ ϭ 0.71073

˚

A), under PC control with an installed Siemens XSCANS 2.1 soft-

ware package. Automatic searching, centering, indexing, and least-

squares routines were carried out on 3, 4, and 5 for at least 30

reflections in the range 24° Յ 2θ Յ 25° and these subsequently

used to determine unit cell parameters. During the data collection,

the intensities of three representative reflections were monitored

every 97 reflections for each of the three crystals, no significant

decay was observed for any of the crystals. The three data sets were

corrected for Lorentz and polarization effects. No semi-empirical

(f) 2-(3Ј-{2ЈЈ-[Bis(3ЈЈЈ-isopropylpyrazol-1ЈЈЈ-yl)methyl]phenylsulfan-

yl}-1Ј,1Ј-diphenylpropyl-1Ј-sulfanyl)tetrahydropyran (27): The syn-

Table 5. Crystallographic data and data collection parameters for 3, 4, and 5

Compound^[a]

3

4

5

Empirical formula

Bravais lattice

Space group

C₁₈H₂₂N₄O

monoclinic

P2₁/c

15.2701(17)

9.9914(12)

12.1924(12)

C₂₁H₂₇N₅OS

monoclinic

P2₁/c

12.6279(6)

9.6906(8)

18.4360(17)

C₂₁H₂₇N₅OS

triclinic

¯

P1

˚

a [A]

9.3154(12)

9.8197(17)

12.5959(12)

96.050(11)

100.216(11)

106.366(13)

1073.2(3)

1.230

˚

b [A]

˚

c [A]

α [°]

β [°]

γ [°]

110.786(6)

104.494(6)

3

˚

V [A ]

1739.1(3)

1.185

664

2184.2(3)

1.209

848

ρ [g·cm^Ϫ3

F(000)

]

424

Crystal size [mm]

0.7 ϫ 1.2 ϫ 1.0

Colorless, block

0.076

0.7 ϫ 0.7 ϫ 0.7

Yellow, cube

0.169

0.2 ϫ 0.2 ϫ 0.3

Colorless, block

0.172

Crystal color, habit

µ [mm^Ϫ1

]

2θ collection range [°]

No. unique data

3.5Ϫ45.0

2256

3.5Ϫ45.0

2846

3.5Ϫ45.0

2771

No. observed data, [|F| Ͼ 4σ(|F|)]

Data/restraints/parameters

R₁[|F| Ͼ 4σ(|F|)]

1683

2418

1679

2256/0/213

2846/0/282

2771/0/254

4.19^[b]

4.49^[c]

5.85^[d]

wR₂

12.66

ϩ0.184

Ϫ0.159

13.16

ϩ0.289

Ϫ0.239

14.24

ϩ0.226

Ϫ0.233

Ϫ3

˚

Largest difference peak [e A

]

Ϫ3

Largest difference hole [e A

]

2

^[a]Temp. 298 K; radiation Mo-K_α; scan type θ-2θ. ^[b]w ϭ 1/[σ²(F_o) ϩ (0.0673P)²ϩ 0.3881P] where P ϭ (F_oϩ 2F_c)/3. ^[c]w ϭ 1/[σ²(F_o)

ϩ (0.0683P)²ϩ 1.5934P] where P ϭ (F_oϩ 2F_c)/3. w ϭ 1/[σ²(F_o) ϩ (0.0457P)²ϩ 0.8206P] where P ϭ (F_oϩ 2F_c)/3.

2

[d]

2

Eur. J. Org. Chem. 2002, 3632Ϫ3645

3643

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FULL PAPER

[13]

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absorption corrections were applied since examination of the ψ

scan data collected for each crystal indicated little anisotropic radi-

ation absorption. The SHELXTL version 5^[84]software package

was used to solve all three structures by direct methods and sub-

sequently refine them. A summary of cell parameters, data collec-

tion conditions, transmission factors, and refinement results can be

found in Table 5. Details pertinent to the individual refinements

are outlined below. The structure of 3 was solved by direct methods,

the asymmetric unit containing a single complete molecule of 3. All

non-hydrogen atoms were refined isotropically and subsequently

anisotropically. All hydrogen atoms were included in calculated po-

sitions except for H1B, which is attached to O1 and hydrogen-bon-

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˚

ded to N2 [O1···N2; 2.64(1) A], which was located on the E-map

by difference Fourier synthesis. The structure of 4 was also solved

by direct methods, with once again a single complete molecule

within the asymmetric unit. The non-hydrogen atoms were first re-

fined isotropically whereupon disorder over two positions was ob-

served in the tert-butyl group (C15, C16, C17, C18) of the C7ϪC12

phenyl ring. The tert-butyl methyl groups (C16, C17, and C18) had

their occupancies tied to their disordered counterparts (C16Ј, C17Ј,

and C18Ј) to give a total of 1.0 for each pair, then these were refined

to give respective occupancy values of 0.68 and 0.32. All the non-

hydrogen atoms were then refined anisotropically. Hydrogen atoms

were included in calculated positions (including the disordered tert-

butyl group, the H occupancies corresponding to those of the C

atom to which they were bonded) using a riding model and fixed

isotropic thermal parameters. The structure of 5 was solved by dir-

ect methods with one complete molecule being contained within

the asymmetric unit. All non-hydrogen atoms were refined with

anisotropic thermal parameters and the hydrogen atoms sub-

sequently included in the refinement in calculated positions using

a riding model with fixed isotropic thermal parameters. CCDC-

187103 (3), -187104 (4) and -187105 (5) contain the supplementary

crystallographic data for this paper. These data can be obtained

free of charge at www.ccdc.cam.ac.uk/conts/retrieving.html or from

the Cambridge Crystallographic Data Centre, 12 Union Road,

Cambridge CB2 1EZ, UK [Fax: (internat.) ϩ 44-1223/336-033;

E-mail: deposit@ccdc.cam.ac.uk].

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Acknowledgments

This work was supported by Grants AI-1157 from the Robert A.

Welch Foundation, the ACF/PRF, and Grant SF-93Ϫ12 from the

Dreyfus foundation. NSF Program Grants USE-9151286 and

CHE-9601574 are gratefully acknowledged for partial support of

the X-ray diffraction and NMR facilities, respectively, at Southwest

Texas State University. We would also like to thank Dr. Hamish

McNabb for a valuable discussion on the presentation of this work.

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Received June 12, 2002

[O02314]

Eur. J. Org. Chem. 2002, 3632Ϫ3645

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Article Doi

The synthesis of "apo-proteinic" multidentate, N2SS* and N2S, ligands to model type I trigonal and trigonal-pyramidal copper protein sites

DOI: 10.1002/1099-0690(200211)2002:21<3632::AID-EJOC3632>3.0.CO;2-2

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Article abstract of DOI:10.1002/1099-0690(200211)2002:21<3632::AID-EJOC3632>3.0.CO;2-2

Full text of DOI:10.1002/1099-0690(200211)2002:21<3632::AID-EJOC3632>3.0.CO;2-2

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