Removal of the phosphate group in mechanism-based inhibitors of inositol monophosphatase leads to unusual inhibitory activity.

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Removal of the phosphate group in mechanism-based inhibitors of

inositol monophosphatase leads to unusual inhibitory activity

David J. Miller, M. Bashir-Uddin Surfraz, Mahmoud Akhtar, David Gani and

Rudolf K. Allemann*

School of Chemistry, The University of Birmingham, Edgbaston, Birmingham, UK B15 2TT.

E-mail: r.k.allemann@bham.ac.uk; Fax: ϩ44 121 414 4446

Received 15th October 2003, Accepted 13th January 2004

First published as an Advance Article on the web 5th February 2004

Inositol monophosphatase is widely held to be the therapeutic target for inhibition by lithium ion in the treatment of

bipolar disorder. In a continued eﬀort to improve the bioavailability of alternative inhibitors, we have designed and

tested two new series of compounds; phosphonates and product-like mimics. Phosphonate substrate mimics were

competitive inhibitors of reduced potency as compared to phosphate based inhibitors. Product mimics however,

showed various inhibitory modes of action. The 6-butylamino derivative 6p was an uncompetitive inhibitor when

acting alone (K_i= 0.3 mM) but displayed non-competitive inhibition in the presence of inorganic phosphate.

This compound represents a new lead in the search for a viable replacement for lithium ion therapy.

Introduction

The chronic and debilitating condition of manic depression

(bipolar disorder) aﬀects up to 340 million people worldwide.¹

It remains a severe problem as there is no cure and the current

treatments with lithium salts have serious limitations due to the

high toxicity of these compounds.²Inositol monophosphatase

(IMPase, E.C. 3.1.3.25) is widely believed to be the therapeutic

target of lithium ion and as such has been the focus of much

eﬀort to discover a more attractive therapy for the condition.^3–9

IMPase is a key enzyme in a brain secondary messen-

ger system.¹⁰Its purpose is to provide free myo-inositol for

Fig. 1 Schematic representation of the hydrolysis of -Ins-1-P by

the biosynthesis of the second messenger precursor phospha-

IMPase. Key interactions of the substrate with the active site are

shown.

tidylinositol-4,5-bisphosphate.^10,11In response to extra-cellular

stimuli, phosphatidylinositol-4,5-bisphosphate is hydrolysed to

1,2-diacylglycerol and inositol-1,4,5-trisphosphate. These so

knowledge. Substrate analogues of cyclitol phosphate 1 with

deleted 3-OH and 5-OH groups bind to the enzyme more

tightly than the natural substrates, and if the 6-OH group is

called second messengers give rise to protein kinase C activ-

ation and intracellular calcium release respectively.^10–12It is

thought that over-activity of this cellular response mechanism

either deleted or alkylated, the resulting molecule is a tight

binding competitive inhibitor of IMPase.^9,16,17For example

leads to the violent mood swings characteristic of manic-

depressive illness.¹³Once inositol-1,4,5-trisphosphate has been

compound 2 is such an inhibitor and displays an IC₅₀value of

3 µM. Similarly, compound 3 is the most potent known inhibi-

released it is rapidly dephosphorylated by a cascade of enzymes

to give ultimately, myo-inositol.¹⁰IMPase catalyses the ﬁnal

tor of IMPase (IC₅₀= 40 nM).⁹Modeling studies have sug-

step of this dephosphorylation sequence and is responsible

gested that the long lipophilic side chain of inhibitors such as 3

for the hydrolysis of monophosphates of -myo-inositol in

enhance binding to the enzyme by interaction with a lipophilic

the 1, 3, 4 or 6 positions.¹⁴Suppression of IMPase activity

reduces the pool of myo-inositol available in the brain and

therefore slows down the resynthesis of the second messenger

precursors.¹³

pocket near the active site bounded by Val40 and Leu42.^5cIt is

now relatively easy to prepare such compounds and we have

used epoxide 4 as a common precursor to all such inhibitors.¹⁸

Previous work has lead to a detailed understanding of the

structure and mode of action of IMPase. The enzyme is a

58 kDa homodimer with a requirement of 2 Mg^2ϩions per

active site.^3–8Kinetic data indicated that one metal ion (Mg^2ϩ1)

binds before binding of the substrate and the second (Mg^2ϩ2)

binds after (Fig. 1).^3–8A water molecule associated with Mg^2ϩ

1

deep within the active site acts as a nucleophile and attacks

the phosphate group of the substrate -inositol-1-phosphate

(-Ins-1-P) 1 and a second water molecule, coordinated to

Mg^2ϩ2 and hydrogen bonded to the 6-OH group of 1 acts as a

general acid, protonating the inositolate group as it leaves.¹⁵

Deletion of this 6-OH group removes any substrate activity

from related compounds and it is known that the 4-OH and

2-OH groups along with the 1-O atom provide strong inter-

actions for the binding of the substrate to the active site.¹⁶

Many mechanism-based inhibitors of IMPase have been syn-

thesized and evaluated during the accumulation of the above

These materials have proved invaluable as probes for elucid-

ation of the mechanism of action of IMPase but they are of no

use in vivo. In common with all known organic phosphate

T h i s j o u r n a l i s © T h e R o y a l S o c i e t y o f C h e m i s t r y 2 0 0 4

O r g . B i o m o l . C h e m . , 2 0 0 4 , 2, 6 7 1 – 6 8 8

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inhibitors of IMPase there are bioavailability limitations that

preclude their use as drugs.⁹These inhibitors possess a double

negative charge at physiological pH which renders them too

hydrophilic to cross the lipophilic blood–brain barrier in order

to have a therapeutic eﬀect.¹⁹The aim of the work described

here was to design and prepare ‘reduced charge’ mechanism

based inhibitors of IMPase. Guided by data available for the

importance of binding interactions it was reasoned that we

could either modify or delete the phosphate group of IMPase

inhibitors in known materials in order to retain good inhibitory

activity against the enzyme.

uncompetitive inhibitor and acts synergistically with inorganic

phosphate (P_i).³It replaces Mg^2ϩ2 at the active site of IMPase

in an enzyme–product complex and prevents the release of

P_i, thereby preventing access by substrate molecules. It was

proposed to prepare compounds that would mimic the

myo-inositol product of the reaction catalysed by the enzyme.

Such molecules would be inositol analogues with similar struc-

tural features to previously prepared inhibitors, except, with the

phosphate group absent. The presence of a good lipophilic side

chain at the 6-position of the inositol ring was expected to

compensate for the binding energy loss that would inevitably

arise from removal of the phosphate group from such com-

pounds. It was anticipated that, like Li^ϩ, these compounds

might act synergistically with the product P_isince they would

not need to displace it from the active site in order to bind. It is

known that P_iis a product inhibitor of the enzyme with K_i=

0.3 mM (at pH 8.0³) and that brain phosphate levels are

∼3 mM. Therefore the enzyme is largely P_i-bound under physio-

logical conditions in the cell, which would aid these compounds

in vivo should they transport eﬀectively to the brain. At the

outset of this work myo-inositol itself was the only known

product inhibitor of IMPase and as one would expect, it has a

very large K_iof 400 mM.³Since no good leads were available for

this approach, it was not expected to devise a tight binding

inhibitor straight away but rather to produce compounds

displaying suﬃcient potency as inhibitors of IMPase, and

the required cooperative eﬀects with P_ito validate the design

strategy. The product-like inhibitors envisaged are illustrated

as general structure 6 in Fig. 3.

Results and discussion

Two approaches towards overcoming the problems of poor

bioavailability were contemplated. Firstly, modeling studies

derived from X-ray data have indicated that one of the oxygen

atoms of the phosphate group of -Ins-1-P does not form a

direct interaction with the active site.^5cMoreover, it is evident

that the P–O bond points directly towards the lipophilic pocket

bounded by Val40 and Leu42 (illustrated in Fig. 2).^5cIt seemed

that the preparation of various phosphonate derivatives with

alkyl groups attached to both the phosphorus atom and to the

6-position of the inositol ring would give inhibitors with a

single negative charge at physiological pH. In addition, if the

6-alkoxy group could be replaced with an alkylamino group at

this position then such compounds would be neutral overall.

These compounds are illustrated as general structure 5 in Fig. 3.

The second approach was entirely diﬀerent and uses know-

ledge of the mechanism of action of Li^ϩas an inhibitor of

IMPase. At its therapeutic concentrations (∼1 mM) Li^ϩis an

Synthesis of phosphonate inhibitors

As with our previous work, the enantiomerically pure epoxide 4

was used as the common intermediate for the preparation of all

the compounds in this work. It is available in multigram quanti-

ties using an 11 step synthesis from (Ϫ)-quinic acid.¹⁸Scheme 1

shows how the phosphonate compounds were prepared.

The simplest of these compounds to prepare were the phos-

phonates bearing a 6-hydoxy 5a or 6-propyloxy group 5b and

5c. These compounds diﬀer in both the identity of the alkoxy

group at the C-6 and the alkyl group attached to phosphorus.

Firstly the epoxide 4 was stereoselectively ring opened with

benzyl alcohol using ytterbium() triﬂate as a Lewis acid

catalyst as reported previously.^18,20The resulting alcohol 7 was

then transformed into phosphonate 8a by treatment with

methyl phosphonic dichloride. The intermediate phosphonyl

chloride was hydrolysed in a mixture of t-butanol–water (1 : 1;

v/v) at 50 ЊC to give the acid.²¹The phosphonic acid 5a was

isolated in 86% yield after removal of the benzyl groups using

sodium in liquid ammonia, and conversion of the 1,2,4,6 tetrol

phosphonic acid into its cyclohexylammonium salt.

The 6-propyloxy alcohol 9b was obtained by ring opening of

epoxide 4 with 1-propanol as described above. The phosphon-

ates 10b and 10c were obtained (52% and 23% respectively) in a

similar manner to above using respectively methyl and ethyl

phosphonic dichlorides as phosphonylating agents. Debenzyl-

ation of these phosphonic acids, was however achieved using

catalytic hydrogenolysis using 10% palladium on activated

charcoal, and the required 6-alkoxy phosphonates 5b and

5c were isolated after transformation into their cyclohexyl-

ammonium salts in 41% and 98% yield respectively.

Fig. 2 Model of Ins-1-P bound at the active site of IMPase. The

binding interactions between the 4-OH and 2-OH groups of Ins-1-P

with Glu213 and Asp93 of the enzyme can be clearly seen. The

hydrophobic loop containing Val40 and Leu42 is also illustrated and

one can also clearly see that both the 6-OH group and one P–O bond of

Ins-1-P point directly towards this exploitable area of the active site.

The preparation of the phosphonate compounds possessing

a 6-alkylamino side chain 5d–f was slightly more complicated as

the amino functionality needed protection prior to the phos-

phonylation step. The ring opening of epoxide 4 proved slightly

more problematic than had previously been encountered due to

the bulkiness of some of the amines used in this step. The reac-

tion with methylamine was carried out as had been previously

communicated²⁰and involved heating a mixture of 4, methyl-

amine in water and ytterbium() triﬂate in a sealed vessel.

Fig. 3 Proposed reduced charge inhibitors of IMPase.

O r g . B i o m o l . C h e m . , 2 0 0 4 , 2, 6 7 1 – 6 8 8

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Scheme 1 Synthesis of phosphonate inhibitors of IMPase. Reagents and conditions: (i) BnOH, Yb(OTf )₃, 1,2-dichloroethane, reﬂux; (ii) R₂POCl₂,

TEA, DMAP, DCM, then t-BuOH/H₂O, 50 ЊC; (iii) Na or Ca in NH₃(liq.), then MeOH; (iv) R₁OH, Yb(OTf )₃, 1,2-dichloroethane, reﬂux. v. H₂,

Pd–C, MeOH; (vi) R₃NH₂, Yb(OTf )₃, various solvents; (vii) BnOCOCl, TEA, DCM.

After protection of the resulting secondary amine with benzyl

chloroformate, alcohol 11d was isolated in 78% yield. For the

preparation of alcohol 11e hexylamine was used to open the

epoxide 4, but under the above conditions or using a solution of

hexylamine in 1,2-dichloroethane none of the required amine

was isolated, and under forcing conditions the amine reacted

with 1,2-dichloroethane. However, treatment of epoxide 4 with

hexylamine and ytterbium() triﬂate in a reﬂuxing mixture of

toluene–THF (3 : 1) gave the desired secondary amine, which

was Cbz protected to give alcohol 11e in 68% yield. For the

synthesis of alcohol 11f, the epoxide 4 was ring opened with

2-phenylethylamine and ytterbium() triﬂate in reﬂuxing 1,2-

dichloroethane, and the resulting secondary amine Cbz pro-

tected to give alcohol 11f in 70% yield. These three protected

amines were then phosphonylated using methylphosphonic

dichloride under the same conditions as before. Phosphonate

12d was carried forward to the next step without isolation,

whereas 12e and 12f were isolated in 84% and 90% yield respect-

ively. The resulting phosphonic acids 12d–f were deprotected

using dissolving metal reduction since catalytic hydrogenolysis

proved slow and unreliable for amine bearing compounds of

this type. The 6-methylamino phosphonate 5d was isolated

in 36% yield after reaction of the intermediate phosphonate

12d with sodium in liquid ammonia to facilitate global

deprotection. Similarly, the 6-hexylamino phosphonate 5e was

isolated in 44% yield from phosphate 12f. However, in the case

of phosphonate 12f, in order to avoid possible Birch reduction

Scheme 2 Synthesis of product-like inhibitors of IMPase. Reagents

and conditions: (i) R₁OH, Yb(OTf )₃, 1,2-dichloroethane, reﬂux; (ii) H₂,

Pd–C, MeOH; (iii) NaH, n-hexyl iodide, DMF, 0 ЊC; (iv) TBAF, THF.

v. TFA(cat.), MeOH; (vi) 4-bromo-1-butene, acetone, reﬂux; (vii) BH₃ؒ

THF, Et₂O, then NaOH/H₂O₂; (viii) R₃NH₂, Yb(OTf )₃, various

solvents; (ix) TMSBr, CHCl₃, 50 ЊC.

of the phenyl group in the 2-phenylethyl side chain, calcium

in liquid ammonia was used for the deprotection step.^22,23

6-(2-Phenylethyl)amino phosphonate 5f was isolated in 47%

yield.

Synthesis of product-like inhibitors

The synthesis of the product mimics is shown in Scheme 2. The

preparation and initial evaluation of some these compounds as

inhibitors of IMPase was the subject of an earlier communi-

cation.²⁴

Two series of compounds were prepared; one had a 6-alkyl-

oxy side chain and the other a 6 amino or 6-alkylamino side

chain. We have previously demonstrated that 6-alkylamino side

chains are tolerated at the active site of IMPase; 6-amino- and

6-methylamino-phosphate derivatives having been shown to be

substrates of IMPase.^15,20Compound 6i is merely the potent

inhibitor 3 without a phosphate group attached at C-1. It was

hoped that since the phosphate group had been removed then

the pendant arm of 3, above all others, would compensate for

the binding energy that would be lost through this change.

Compounds 6l and 6m were designed with an amino group at

the terminus of the 6-alkyloxy group. It was hoped that this

group, which would be positively charged at physiological pH,

would replace the Mg^2ϩ2 ion at the active site of IMPase and

O r g . B i o m o l . C h e m . , 2 0 0 4 , 2, 6 7 1 – 6 8 8

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Table 1 Inhibitory constants and modes of action of substrate-like inhibitors of IMPase

Compound

Phosphonate (R₁)

Side chain on C-6 (XR₂)

Mode of action

K_ior IC₅₀(mM)

myo-Inositol 1-P

5

0.1 (K_M)

Hydroxyl

Methyl

Ethyl

Methyl

OH

O(CH₂)₂CH₃

NHCH₃

NH(CH₂)₅CH₃

NH(CH₂)₂Ph

Substrate

—

Competitive

—

0.025^16c(K_M)

260 16

5a

5b

5c

5d

5e

5f

0.04 0.004 (K_i)

0.16 0.016 (K_i)

>8

1.3 0.2

>54

Scheme 3 Synthesis of product-like inhibitor of IMPase bearing a diamino functionality. Reagents and conditions: (i) BnOCOCl, TEA, DMAP,

1,4-dioxane–water (1 : 1; v/v), 0 ЊC to RT; (ii) TsCl, DMAP, TEA, DCM; (iii) 4-phenylbutylamine, 50 ЊC, closed system, then K₂CO₃, CHCl₃;

(iv) 4, Yb(OTf )₃, toluene–THF (3 : 1; v/v), reﬂux. v. TMSBr, CHCl₃, 50 ЊC.

that this would mimic the action of Li^ϩ. The 6-alkylamino

derivatives 6e–f and 6n–r were designed with varying alkyl

chain lengths in order to gain a structure activity relationship

should these compounds be successful as inhibitors of IMPase.

The synthesis of the 6-alkyloxy derivatives involved simple

ring opening of epoxide 4, under standard conditions, with

n-propanol, alcohol 15 (prepared from catechol as shown in

Scheme 2) and Cbz-protected ethanolamine and propanol-

amine to give compounds 9g,i–k in 65%,¹⁸72%, 71% and

52% yield, respectively. Deprotection was achieved using

catalytic hydrogenolyisis over palladium–charcoal in excellent

yields (84–95%).

In an eﬀort to simplify the synthetic route to these 6-alkyloxy

derivatives we decided to investigate the direct alkylation of the

4-silyl ether 16; an intermediate in the synthesis of the epoxide

4.¹⁸Treatment of 4-silyl ether 16 with n-hexyl iodide in the

presence of sodium hydride gave the fully protected tetrol 17h in

64% yield. Sequential removal of the silyl and cyclohexylidene

ketal protecting groups with TBAF in THF and TFA in

MeOH respectively, gave the 6-hexyloxy 1,2,4-triol 6h in 63%

yield. This route, which is considerably shorter and higher

yielding oﬀers an eﬃcient way of synthesizing these and further

6-alkyloxy derivatives.

lipophilic pocket at that site we decided to synthesize the

diaminotriol 25, Scheme 3.

Ethanolamine 20 was Cbz protected in 80% yield using

benzyl chloroformate in the presence of DMAP. The free

alcohol 21 was converted into the corresponding tosylate in

88% using tosyl chloride in the presence of DMAP and

TEA. Nucleophilic substitution of the tosyl group with

4-phenylbutylamine gave the secondary amine 23 in 45% yield.

Although we were concerned that ring opening of epoxide 4

would be diﬃcult, due the added steric bulk of the secondary

amine, the reaction proceeded eﬃciently under the standard

conditions of epoxide ring opening to give the diamino alcohol

24 in 58% yield. Global deprotection using either hydro-

genolysis or dissolving metal reduction proved unfruitful.

However when the diamino alcohol was treated with TMS-

bromide the resulting diamino triol 25 was isolated in 38%

yield.

Biological evaluation

Phosphonates. For the testing of the phosphonate com-

pounds 5a–f as inhibitors of IMPase, the colorimetric assay of

Itaya et al. was used.^3,25This employs a malachite green/

molybdic acid colorimetric reagent that is sensitive to the

release of P_ithat occurs upon turnover of the substrate. Using

this assay 5a (260 µM) was shown to be a competitive inhibitor

of IMPase but with reduced potency relative to a phosphate

derivative (K_M= 25 µM^16c), Table 1. It was also noted that this

compound may be a substrate for the enzyme. Incubation of 5a

with the enzyme under standard assay conditions and following

The 6-amino and 6-alkylamino derivatives, of general struc-

ture 19, were prepared in the same manner as that outlined

above for the 6-alkylaminophosphate derivatives (5d–f ) starting

from epoxide 4. Removal of the benzyl groups was however

achieved by heating the amino alcohols 19 at 50 ЊC with tri-

methylsilyl bromide in chloroform, and yields ranged from 54%

to 88%.

1

In order to investigate whether the introduction of a second-

ary alkyl group at C-6 would increase binding due to the

the progress of the reaction by H and ³¹P NMR spectroscopy

showed (due to no change in the NMR spectrum) that the com-

O r g . B i o m o l . C h e m . , 2 0 0 4 , 2, 6 7 1 – 6 8 8

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Table 2 Inhibitory constants and modes of action of product-like inhibitors of IMPase

Compound

Side chain (XR₂)

Mode of action

K_ior IC₅₀(mM)

myo-Inositol

—

Non-competitive

—

Competitive

—

400 (K_i)³

∼150 (IC₅₀)

∼10 (IC₅₀)

6g

6h

6i

O(CH₂)₂CH₃

O(CH₂)₅CH₃

O(CH₂)₄O-[(2-OH)C₆H₄]

O(CH₂)₂NH₂

O(CH₂)₃NH₂

NH₂

NHCH₂CH₃

NH(CH₂)₃CH₃

NH(CH₂)₅CH₃

NH(CH₂)₇CH₃

NH(CH₂)₂Ph

NH(CH₂)₄Ph

H₂N(CH₂)₂N(CH₂)₄Ph

4

5

0.6 (K_i)

1 (IC₅₀)

6l

6m

6n

6o

6p

6e

6q

6f

10 2 (IC₅₀)

>50 (IC₅₀)

0.3 0.03 (K_i)

0.3 0.04 (K_i)

—

Uncompetitive (in absence of P_i)

Uncompetitive

—

Non-competitive

—

4

6

9

0.4 (IC₅₀)

0.6 (IC₅₀)

0.9 (K_i)

6r

25

0.9 (IC₅₀)

pound displayed no substrate activity. The methyl and ethyl

phosphates 5b (K_i= 40 µM) and 5c (K_i= 160 µM) were also

shown to be competitive inhibitors of IMPase but again with

reduced potency relative to a phosphate derivative with a

6-propyloxy side chain (K_i= 0.85 µM^17,26). It seemed therefore

that increasing the length of the alkyl chain directly attached to

the phosphorus in these compounds had a detrimental eﬀect on

their inhibitory activity. Attempts to make further homologues

of this type were complicated due to problems encountered in

separating the ﬁnal products from contaminating phosphonic

acids. The crude materials, when tested, did not show any sig-

niﬁcant inhibitory activity. The phosphonates 5d–f possessing

the 6-alkylamino functionality and hence no net charge were

even more disappointing as inhibitors of IMPase. Neither the

6-methylamino derivative 5d nor the 6-(2-phenylethyl)amino

derivative 5f gave any measurable inhibition of the enzyme.

Indeed the latter did not retard the turnover of substrate even at

concentrations up to 50 mM. Of these, only the 6-hexylamino

derivative 5e was active against IMPase and this had a relatively

poor IC₅₀of 1.3 mM. The results are summarized in Table 1.

We therefore concluded that reduction of the charge on IMPase

by modiﬁcation of the phosphate group to a phosphonate was

not likely to lead to an eﬀective inhibitor of IMPase.

diﬀered in its mode of action. myo-Inositol is known to be a

non-competitive inhibitor of IMPase but the double reciprocal

plots for 6i clearly intersected on the y-axis showing this com-

pound to be a competitive inhibitor of IMPase,²⁷Fig. 4. This

result demonstrates that this compound blocks the binding site

from access by the substrate.

Fig. 4 Double reciprocal plots of initial rate versus [Ins-1-P] at 6i

concentrations of 0 (᭹), 1 mM (red ᭿), 2 mM (green ꢀ) and 3 mM

(blue ᭜). Experiments were performed at 25 ЊC and at pH 8.0.

Intersection of the lines on the y-axis indicates that 6i is a competitive

inhibitor of IMPase.

Product mimics. The product mimics prepared in this work

proved much more interesting as inhibitors of IMPase. Because

it was anticipated that any inhibitor showing promise would be

tested as an inhibitor in the presence of P_i(in order to assess

whether the two would act cooperatively) it was necessary to

use a diﬀerent assay. The colorimetric assay is sensitive to a

maximum concentration of 100 µM P_i²⁵and our needs required

assay conditions containing concentrations 10–30 times higher

than this. The assay used was the radiochemical assay of Gee

et al.^3,14employing tritium labeled -inositol-1-phosphate as the

substrate. The concentration of -Ins-1-P was 0.2 mM. For

each compound in this series an IC₅₀was calculated and for

promising compounds a full K_idetermination was performed to

assess the mode of action of the inhibitor. The results are sum-

marized in Table 2.

All inhibitors tested showed a marked improvement in aﬃn-

ity for the enzyme as compared to the natural product myo-

inositol. The 6-alkyloxy derivatives were the least promising of

these compounds, yet bound to the enzyme approximately 2

orders of magnitude more tightly than myo-inositol itself.

Interestingly, the double reciprocal plots (1/v versus 1/[S]) for 6i

possessing the catechol derived side chain of 3 showed that it

The two 6-alkyloxy compounds containing an amino group

in the pendant arm 6l and 6m showed a lower aﬃnity for the

enzyme than 6i. Two further 6-alkyloxy derivatives containing

simple alkyl chains, 6-propyloxy 6g and 6-hexyloxy 6h, were

also prepared and tested but were found to be of lower aﬃnity

(IC₅₀∼150 mM and ∼10 mM respectively). In addition the

relatively low water solubility of these compounds precluded

accurate IC₅₀measurements.²⁴Interestingly, 6l showed a 30-fold

increase in potency as compared to 6g, indicating that replace-

ment of the terminal CH₃by an NH₂group enhances binding

almost certainly through its interaction with the Mg^2ϩ2 binding

site.

The 6-alkylamino derivatives showed more potency as inhibi-

tors of IMPase as well as having a diﬀerent mode of action.

The 6-(4-phenylbutyl) derivative 6r when tested as an inhibitor

gave double reciprocal plots that intersect on the x-axis, Fig. 5.

This is clearly indicative of non-competitive inhibition and is

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Fig. 5 Double reciprocal plots of initial rate versus [Ins-1-P] at 6r

concentrations of 0 (᭹), 5 mM (red ᭿), 10 mM (green ꢀ) and 15 mM

(blue ᭜). The plots intersect on the x-axis indicating non-competitive

inhibition. Experiments were performed at 25 ЊC and at pH 8.0.

Fig. 6 Double reciprocal plots of initial rate versus [Ins-1-P] at 6p

concentrations of 0 (᭺), 0.25 mM (ᮀ), 0.5 mM (᭝) and 0.75 mM (◊).

The parallel line pattern is clearly indicative of uncompetitive

inhibition. Also shown are the same measurements {6p concentrations

of 0 (᭺), 0.25 mM (ᮀ), 0.5 mM (᭝) and 0.75 mM (◊)} in the presence

of 1 mM Pi. Under these conditions the plots now intersect on the x-

axis indicating a change of behaviour to non-competitive inhibition.

Experiments were performed at 25 ЊC and at pH 8.0.

the same mode of action as shown by the natural product of

IMPase. This indicates that 6r does not compete with Ins-1-P

for the active site of IMPase yet it does lower the k_catof the

reaction. Presumably this compound is acting as a substrate

for the enzyme in the reverse direction and hence inhibits in a

non-competitive fashion.

The diamino derivative 25 had an IC₅₀value (9 mM) margin-

ally lower than the 6-(4-phenylbutyl) derivative 6r (13 mM).

Presumably the 4-phenylbutyl group binds with the Val40–

Leu42 lipophilic pocket and the primary amino group in the

Mg^2ϩsite. This could be investigated further using other

analogues. It would also be interesting to see whether replace-

ment of oxygen in the 2-aminopropyloxy compound 6l by

nitrogen would lead to a better inhibitor than either 6l or the

diamino derivative 25.

The most interesting of all of these product-like inhibitors

proved to be the 6-butylamine and 6-hexylamine derivatives 6p

and 6e, with sub-millimolar IC₅₀values. Moreover upon meas-

urement of their K_ivalues it became apparent that they again

had a diﬀerent mode of action as inhibitors of IMPase. The

double reciprocal plots at diﬀerent inhibitor concentrations

showed a parallel line pattern indicating that these compounds

were uncompetitive inhibitors of IMPase—both with K_i=

0.3 mM. This suggests that these compounds have eﬀectively no

aﬃnity at all for the free enzyme and only bind to the enzyme

substrate complex, not aﬀecting k_cat/K_m.²⁷The butylamine

derivative 6p was then assessed as an inhibitor in the presence

of P_iand showed a change in mode of action; the double

reciprocal plots now showed a non-competitive inhibition

pattern, see Fig. 6. Presumably this change in behaviour is due

to the secondary amine functionality of 6p, which will be posi-

tively charged under the assay conditions (pH 8.0) and there-

fore have no aﬃnity for an active site designed for the anionic

inositol monophosphates. Only when the requirement for an

anion to be bound is fulﬁlled does it bind to the enzyme active

site. Under these circumstances it is presumably acting as a

substrate for the enzyme in the reverse direction and so, like

inositol itself will act as a non-competitive inhibitor. The K_ifor

6p was then measured at concentrations of 1, 2 and 3 mM P_ito

see if the two were cooperative in their inhibition of IMPase.

The results are shown in Fig. 7 and are inconclusive. Certainly

the presence of P_iaﬀects the magnitude of K_ifor 6p but it seems

less eﬀective rather than more eﬀective. Moreover, from 1–2

mM P_ithe K_iof 6p gets worse but then improves again as the

concentration of P_iincreases to 3 mM.

Fig. 7 K_ifor 6p at P_iconcentrations of 0, 1, 2 and 3 mM. Note that at

[P_i] = 0 6p displays uncompetitive inhibition and at 1–3 mM it is a non-

competitive inhibitor.

amino derivatives 6n and 6o (of general structure 6) were pre-

pared but these were completely ineﬀective as inhibitors of

IMPase with IC₅₀values > 50 mM. This indicates that at least a

4-carbon chain length seems to be necessary on the 6-amino

group for eﬀective inhibition of the enzyme.

Conclusion

Phosphonate inhibitors of IMPase are far less eﬀective than

their phosphate counterparts, although at K_i= 40 µM, 5b is

amongst the most potent mono-anionic inhibitors of IMPase

yet produced.⁹Introduction of a 6-amino or 6-alkylamino

group on these materials further reduces their potency as

inhibitors of IMPase.

The product-like inhibitor series were not tight binding

inhibitors of IMPase but were 2–3 orders of magnitude more

eﬀective as product inhibitors than myo-inositol. No conclusive

evidence that these compounds might cooperatively inhibit

IMPase with P_iwas found but the K_iof 6p did seem to vary

somewhat with the P_iconcentration indicating that this might

be possible. The change in the mode of action of this inhibitor

in the presence of P_iis certainly encouraging. These compounds

therefore represent good lead structures from which more

potent inhibitors of this type might be derived. Given that they

It seems from the data shown in Table 2 that as the 6-alkyl-

amino side chain is reduced in length then the more potent the

inhibitor (albeit 6p and 6e are of the same potency within

experimental error). Consequently the 6-amino and 6-ethyl-

O r g . B i o m o l . C h e m . , 2 0 0 4 , 2, 6 7 1 – 6 8 8

676

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have a very diﬀerent nature to the phosphate inhibitors pre-

pared in our previous work, it is certainly possible that they

might have very diﬀerent pharmacological properties and

future work will aim at investigating their transport properties

in vivo.

(؉)-(1S,2R,4R,6R)-2,4-Bis(benzyloxy)-1,6-epoxycyclohexane 4

This compound was prepared as previously described.¹⁸

(؊)-(1S,2R,4S,6R)-2,4,6-Tris(benzyloxy)-cyclohexanol 7

To a stirred solution of the epoxide (ϩ)-4 (300 mg, 0.968 mmol)

and benzyl alcohol (2.00 g, 19.4 mmol) in 1,2-dichloroethane

(30 cm³) was added Yb()(OTf )₃(120 mg, 0.194 mmol) and the

solution was heated under reﬂux for 16 h. After cooling, the

solvent was removed under reduced pressure and the excess

benzyl alcohol was removed by distillation in vacuo using a

Kugelrohr apparatus. The residue was then puriﬁed by ﬂash

silica column chromatography (petroleum ether–ethyl acetate;

1 : 1) to give the alcohol 7 as a colourless oil (359 mg, 89%);

(HRMS: found: [M ϩ Na]^ϩ, 441.2031. C₂₇H₃₀O₄Na requires

441.2042); [α]_DϪ33.0 (c 0.98 in MeOH); ν_max(thin ﬁlm)/cm^Ϫ1

3562, 3460, 2930, 2867, 1952, 1876, 1810, 1605, 1586, 1496 and

1454; δ_H(300 MHz; C²HCl₃) 1.37–1.49 (2 H, m, 3-H and 5-H),

2.34–2.42 (1 H, m, 3-H), 2.50–2.55 (1 H, m, 5-H), 2.58 (1 H, d,

J_HH4.5, OH), 3.60–3.82 (3 H, m, 1 H, 4-H and 6-H), 3.97–4.00

(1 H, m, 2-H), 4.47–4.74 (6 H, m, PhCH₂) and 7.28–7.39 (15 H,

m, Ar-H); δ_C(75 MHz; C²HCl₃) 33.91 and 35.34 (3-C and 5-C),

70.60, 71.64 and 71.93 (PhCH₂), 71.59 (1-C), 75.54, 76.22 and

76.89 (2-C, 4-C and 6-C), 127.59, 127.62, 127.72 and 128.39

(Ar-CH) and 138.40 and 138.50 (Ar-C quaternary); m/z (TOF

ES^ϩ) 441 (100%, [M ϩ Na]^ϩ).

Experimental

Elemental microanalyses were performed in the departmental

micro-analytical laboratory using a Carlo Erba EA1110 CHNS

elemental analyzer. NMR spectra were recorded on a Bruker

AC-300 spectrometer (¹H, 300 MHz; ³¹P, 121.5 MHz), Bruker

AV-300 spectrometer (¹³C, 75.4 MHz), Bruker AMX-400

spectrometer (¹H, 400 MHz; ¹³C, 100 MHz) and a Bruker

DRX-500 spectrometer (¹H, 500 MHz; ¹³C, 125 MHz).

Chemical shifts are described in parts per million downﬁeld

shift from SiMe₄and are reported consecutively as position

(δ_Hor δ_C), relative integral, multiplicity (s = singlet, d = doublet,

t = triplet, q = quartet, dd = double of doublets, sep = septet,

m = multiplet, and br = broad), coupling constant (J/Hz) and

assignment (numbering according to the IUPAC nomenclature

for the compound). ¹H-NMR were referenced internally on

CH₃OH (δ 3.35), HOH (δ 4.68) or CHCl₃(δ 7.27). ¹³C-NMR

2

were referenced on CH₃OH (δ 49.0), C²HCl₃or (δ 77.0). ³¹P

NMR spectra were referenced to an external capillary contain-

ing 1 mol dm^Ϫ3solution of H₃PO₄in deuterated water (δ 0.00).

Infra-red spectra were recorded on a Nicolet Avatar 360 FT-IR

spectrometer. The samples were prepared as KBr discs, Nujol

mulls, solutions in chloroform or thin ﬁlms between sodium

chloride discs. The frequencies (ν) as absorption maxima are

given in wavenumbers (cm^Ϫ1) relative to a polystyrene standard.

Mass spectra and accurate mass (HRMS) measurements were

recorded on a VG ProSpec or VG ZabSpec spectrometer (EI

and CI), Micromass LCT TOF spectrometer (ES) and VG

ZabSpec spectrometer [LSIMS (modern version of FAB)].

LSIMS spectra were recorded using m-nitrobenzyl alcohol as a

matrix. Chemical ionization (CI) spectra were recorded using

ammonia as a reagent gas. Electrospray (ES) spectra were

recorded using MeOH or acetonitrile as the mobile phase.

Major fragments were given as percentages of the base peak

intensity (100%). Melting points were taken on an Electro-

thermal Gallenkamp melting point apparatus and are uncor-

rected. Optical rotations were measured at room temperature

(25 ЊC) on Optical Activity AA-2001 polarimeter using a 5, 10 or

20 cm path length cell and are given in 10^Ϫ1deg cm²g^Ϫ1. Flash

chromatography was performed according to the method of Still

et al.²⁸using Fluka silica gel 60 (3570 µm mesh). Analytical thin

layer chromatography (TLC) was carried out on 0.25 mm pre-

coated silica gel glass plates (Whatman K6F) and compounds

were visualised using UV ﬂuorescence, ethanolic phosphomolyb-

dic acid, ninhydrin or iodine. Preparative cation exchange column

chromatography was performed using Amberlite X 131 WET

(H^ϩion-exchange resin, 120 × 25 mm), eluting with distilled

water at a rate of 1 drop every 3 seconds. RP HPLC was carried

out using a Dionex Summit System. Analytical RP HPLC was

performed on a Phenomenex Luna C18 (2) 10 µm column (250 ×

4.6 mm). Preparative RP HPLC was performed on a Pheno-

menex Luna C18 (2) 10 µm column (250 × 21.2 mm).

(؊)-(1R,2R,4R,6R)-2,4,6-Trihydroxycyclohexyl cyclohexyl-

ammonium 1-methylphosphonate [cyclohexylammonium salt

of 5a]

To a stirred solution of alcohol (Ϫ)-7 (142 mg, 0.340 mmol) in

dry dichloromethane (10 cm³) under argon was added dry TEA

(237 mm³, 1.70 mmol) and DMAP (42 mg, 0.340 mmol) and

the solution was cooled to 5 ЊC. Methyl phosphonic dichloride

(226 mg, 1.70 mmol) was added and the solution was stirred at

room temperature for 16 h. The solvent was removed under

reduced pressure and the residue was redissolved in t-butanol–

water, 1 : 1 (10 cm³) and the solution was heated at 50 ЊC for 2 h.

After cooling, the solvent was removed under reduced pressure

and the residue was partitioned between 10% aqueous sodium

bicarbonate (10 cm³) and diethyl ether (10 cm³). The separated

aqueous layer was extracted with diethyl ether (10 cm³) then

acidiﬁed to pH 1 by addition of concentrated HCl and then the

resulting white precipitate was extracted with ethyl acetate

(3 × 20 cm³). The pooled organic extracts were dried (MgSO₄),

ﬁltered and evaporated to give the intermediate protected

phosphonate 8a as a colourless oil. This was then dissolved in

dry THF (2 cm³).

Anhydrous ammonia was condensed at Ϫ78 ЊC into a dry

three-necked ﬂask ﬁtted with a cold-ﬁnger condenser until

approximately 10 cm³had condensed. To this stirred liquid

ammonia was added freshly cut sodium (75 mg, 3.28 mmol)

and the solution turned a deep blue colour. The solution of the

protected phosphonate 8a in dry THF was then added via a

cannula (plus washings) and the mixture stirred at Ϫ78 ЊC for

30 min. MeOH (2 cm³) was then added and the resulting colour-

less solution was allowed to stand at room temperature for 16 h

to allow the volatile solvents to evaporate. The solid white resi-

due was dissolved in water (2 cm³) and passed down a column

of Amberlite IR 118H cation exchange resin (pre-loaded with

H^ϩions, eluting with water). To the pooled, acidic column frac-

tions was added a 1% solution of cyclohexylamine in water

until the solution reached pH 6 and the solution was lyophil-

ised. The resulting white powder was crystallised from MeOH–

diethyl ether to give the phosphonate 5a as a white crystalline

solid (95 mg, 86%), mp >200 ЊC (decomp.) (HRMS: found:

[M ϩ Na]^ϩ, 249.0499. C₇H₁₅O₆NaP requires 249.0504); [α]_D

Ϫ25.6 (c 0.5 in MeOH); ν_max(KBr disc)/cm^Ϫ13405 br, 2938,

2860, 1646, 1540 and 1456; δ_H(300 MHz; ²H₂O) 0.96–1.06 (1 H,

myo-Inositol-phosphatase was supplied by Sigma and was

used according to the manufacturers instructions. (2-³H)-myo-

inositol-1-phosphate was supplied by Tocris-Cookson Ltd.

Amberlite X 131 WET ion exchange resin and phosphoryl-

ating reagents were obtained from Aldrich Chemical Co. Ltd.

(Gillingham, Dorset, UK). The solvents were used either

distilled or of Analar quality and petroleum ether refers to that

portion boiling between 40 and 60 ЊC. Solvents were dried

according to literature procedures.²⁹MeOH was dried using

magnesium turnings, dichloromethane and TEA were distilled

over calcium hydride and THF was distilled over sodium–

benzophenone.

O r g . B i o m o l . C h e m . , 2 0 0 4 , 2, 6 7 1 – 6 8 8

677

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m, 1 × 4-H of Cha), 1.09–1.31 (4 H, m, 2 × 2-H of Cha

and 2 × 3-H of Cha), 1.36–1.50 (3 H, m, 1 × 4-H of Cha, 3-H

and 5-H), 1.61–1.64 and 1.77–1.83 (4 H, m, 2 × 2-H of Cha and

2 × 3-H of Cha), 1.93–2.01 (1 H, m, 3-H), 2.07–2.13 (1 H, m,

5-H), 2.90–3.05 (1 H, m, 1-H of Cha), 3.67–3.79 (2 H, m, 1-H

and 6-H), 3.88 (1 H, tt, J_HH4.5 and 11.5, 4-H) and 4.04–4.10

J_HH12, PhCH₂), 4.62 (2 H, ABq, J_HH12, PhCH₂) and 7.26–

7.38 (10 H, m, Ar-H); δ_C(75 MHz; C²HCl₃) 6.42 (d, J_CP7,

PCH₂CH₃), 10.43 (OCH₂CH₂CH₃), 19.33 (d, J_CP144, PCH₂-

CH₃), 23.08 (OCH₂CH₂CH₃), 34.30 and 35.65 (5-C and 3-C),

70.52 (PhCH₂O), 71.25 (4-C), 72.08 (OCH₂CH₂CH₃), 72.75

(PhCH₂O), 75.06 (d, J_CP5, 6-C), 76.09 (2-C), 79.02 (d, J_CP4,

1-C), 127.49, 127.55, 127.56, 128.25, 128.34 (Ar-CH) and 138.4

and 138.7 (Ar-C quaternary); δ_P(121 MHz; C²HCl₃) 36.19;

m/z (ES^ϩTOF) 485 (100%, [M ϩ Na]^ϩ).

2

(1 H, m, 2-H); δ_C(75 MHz; H₂O) 14.55 (d, J_CP137, PCH₃),

26.26 (3-C of Cha), 26.75 (4-C of Cha), 32.79 (2-C of Cha),

40.46 (5-C), 42.35 (3-C), 52.78 (1-C of Cha), 66.17 (4-H), 69.14

(d, J_CP5, 6-C), 70.91 (2-C) and 81.55 (d, J_CP6, 1-C); δ_P(121

MHz; ²H₂O) 27.23; m/z (TOF ES^ϩ) 303 (25%, [M Ϫ H ϩ 2K]^ϩ),

287 (10, [M Ϫ H ϩ Na ϩ K]^ϩ), 271 (100, [M Ϫ H ϩ 2Na]^ϩ) and

249 (40%, [M ϩ Na]^ϩ).

(؊)-(1R,2R,4R,6R)-6-Propyloxy-2,4-dihydroxycyclohexyl

cyclohexylammonium 1-methylphosphonate [cyclohexyl-

ammonium salt 5b]

To a stirred solution of protected phosphonate 10b (88 mg,

0.197 mmol) in ethanol (10 cm³) was added a drop of acetic

acid followed by 10% palladium on activated charcoal (20 mg)

and the mixture was stirred under an atmosphere of hydrogen

for 72 h. The solution was ﬁltered through a thin pad of

Celite to remove the catalyst and then the solvent was removed

under reduced pressure. The resulting residue was dissolved in

water (1 cm³) and the solution passed through a column of

Amberlite IR 118H cation exchange resin (H^ϩform, eluting

with water). The acidic fractions were pooled and a 1% solution

of freshly distilled cyclohexylamine in water was added until the

solution reached pH 7. The solvent was then lyophilised to

give phosphonate 5b as a white powder (29.6 mg, 41%); mp

>200 ЊC (decomp.) (HRMS: found: [M ϩ Na]^ϩ, 291.0983.

C₁₀H₂₁O₆NaP requires 291.0973); [α]_DϪ33.6 (c 0.23 in MeOH);

ν_max(KBr disc)/cm^Ϫ13379 br, 2937, 2860, 1633, 1535 and 1448;

δ_H(300 MHz; ²H₂O) 0.71 (3 H, t, J_HH7, OCH₂CH₂CH₃)

0.97–1.49 (13 H, m, 3-H, 5-H, OCH₂CH₂CH₃, PCH₃and

6 × Cha-H), 1.56–1.66 and 1.76–1.84 (4 H, m, 4 × Cha-H),

1.91–1.98 and 2.18–2.23 (2 × 1 H, 2 × m, 3-H and 5-H), 2.96

(1 H, m, 1-H of Cha), 3.37–3.56 (3 H, m, OCH₂CH₂CH₃and

6-H), 3.79–3.89 (2 H, m, 1-H and 4-H) and 4.02–4.10 (1 H, m,

2-H); δ_C(75 MHz; ²H₂O), 12.24 (OCH₂CH₂CH₃), 14.13 (d,

J_CP138, PCH₃), 24.92 (OCH₂CH₂CH₃), 26.26 (3-C of

Cha), 26.74 (4-C of Cha), 32.78 (2-C of Cha), 40.31 (3-C),

40.43 (5-C), 52.81 (1-C of Cha), 66.16 (4-C), 70.75 (2-C),

75.05 (OCH₂CH₂CH₃), 77.22 (d, J_CP5.5, 6-C) and 81.15

(؊)-(1S,2R,4S,6R)-(O¹-Methylphosphonyl)-2,4-bis(benzyloxy)-

6-propyloxycyclohexanol 10b

To a stirred solution of (Ϫ)-(1S,2R,4S,6R)-2,4-bis(benzyloxy)-

6-propyloxycyclo-hexanol 9b¹⁸(200 mg, 0.541 mmol) in dry

dichloromethane (10 cm³) under argon was added dry TEA

(150 mm³, 1.08 mmol) and DMAP (66 mg, 0.541 mmol) and the

solution was cooled to 5 ЊC (ice–water bath). A solution of

methyl phosphonic dichloride (359 mg, 2.70 mmol) in dry di-

chloromethane (10 cm³) was then added via a cannula. After

stirring for 16 h at room temperature the solvent was removed

under reduced pressure and the residue was redissolved in

1 : 1 water : tert-butanol (10 cm³). This solution was then heated

at 60 ЊC for 2 h. After cooling the solution was diluted with

10% sodium hydrogen carbonate solution (10 cm³) and then

extracted with diethyl ether (2 × 20 cm³). The separated aque-

ous layer was acidiﬁed to pH 1 by dropwise addition of concen-

trated HCl and the resulting white precipitate was extracted

with ethyl acetate (3 × 20 cm³). The pooled organic extracts

were dried (MgSO₄), ﬁltered and concentrated under reduced

pressure to give the protected phosphonate 10b as a white solid

(125 mg, 52%); mp 76–78 ЊC (HRMS: found: [M ϩ Na]^ϩ,

471.1905. C₂₄H₃₃O₆NaP requires 471.1912); [α]_DϪ35 (c 0.16

in CH₂Cl₂); ν_max(KBr disc)/cm^Ϫ13437 br, 2960, 2858, 1737,

1495, 1455, 1368 and 1368; δ_H(300 MHz; C²HCl₃) 0.84 (3 H, t,

J_HH7, OCH₂CH₂CH₃), 1.27–1.59 (4 H, m, 1 × 3-H, 1 × 5-H,

OCH₂CH₂CH₃), 1.47 (3 H, d, J_HP18, PCH₃), 2.12–2.23 (1 H, m,

1 × 3-H), 2.34–2.44 (1 H, m, 1 × 5-H), 3.43 (2 H, ddt, J_HH7,

9.5 and 22, OCH₂CH₂CH₃), 3.58–3.77 (2 H, m, 6-H and

4-H), 3.97–4.02 (1 H, m, 2-H), 4.19 (1 H, dt, J_HH3 and 8,

1-H), 4.43 (2 H, ABq, J_HH11, 4-C OCH₂Ph), 4.58 (2 H,

ABq, J_HH12, 2-C OCH₂Ph) and 7.19–7.32 (10 H, m, Ar-H);

δ_C(125 MHz; C²HCl₃), 10.51 (OCH₂CH₂CH₃), 12.07 (d,

J_CP146, PCH₃), 23.23 (OCH₂CH₂CH₃), 34.56 (3-C), 35.66

(5-C), 70.58 (4-C OCH₂Ph), 71.36 (4-C), 71.79 (OCH₂-

CH₂CH₃), 72.88 (2-C OCH₂Ph), 74.93 (d, J_CP3, 6-C), 76.06

(2-C), 79.55 (d, J_CP4, 1-C), 127.5, 127.6, 128.3 and 128.4

(Ar-CH) and 138.6 and 138.8 (2 × Ar-C quaternary);

δ_P(121 MHz; C²HCl₃) 33.90; m/z (ES^ϩ) 471 (100%, [M ϩ Na]^ϩ)

and 449 (15, [M ϩ H]^ϩ).

2

(1-C); δ_P(121 MHz; H₂O) 30.07; m/z (ES TOF) 291 (100%,

[M ϩ Na]^ϩ) and 269 (10, [M ϩ H]^ϩ).

(؊)-(1R,2R,4R,6R)-6-Propyloxy-2,4-dihydroxycyclohexyl

cyclohexylammonium 1-ethylphosphonate [cyclohexylammonium

salt 5c]

This compound was prepared in a manner identical with that

described for the phosphonate 5b using the protected phos-

phonate 10c (46 mg, 98.7 µmol) to give a crude oil which was

crystallised from MeOH–diethyl ether to aﬀord phosphonate 5c

as a white powder (34 mg, 98%); mp >200 ЊC (decomp.)

(HRMS: found: [M ϩ Na]^ϩ, 305.1137. C₁₁H₂₃O₆NaP requires

305.1130); ν_max(KBr disc)/cm^Ϫ13435 br, 2942, 2865, 1679, 1643,

1529 and 1448; [α]_DϪ23.3 (c 0.18 in MeOH); δ_H(500 MHz;

²H₂O), 0.84 (3 H, t, J_HH7.5, OCH₂CH₂CH₃), 0.94–1.12 (4 H, m,

PCH₂CH₃and 1 × 4-H of Cha), 1.17–1.30 (3 H, m, 5-H and

2 × 3-H of Cha), 1.43–1.59 (5 H, m, PCH₂CH₃, OCH₂CH₂CH₃

and 1 × 4-H of Cha), 1.64–1.74 (2 H, m, 2 × 3-H of Cha), 1.86–

1.92 (2 H, m, 4 × 2-H of Cha), 1.98–2.05 (1 H, m, 3-H), 2.24–

2.30 (1 H, m, 5-H), 2.96–3.09 (1 H, m, 1-H of Cha), 3.55 (2 H, t,

J_HH6.5, OCH₂CH₂CH₃), 3.59 (1 H, dt, J_HH10 and 4, 6-H),

3.85–3.95 (2 H, m, 1-H and 4-H) and 4.15 (1 H, br s, 2-H);

(؊)-(1S,2R,4S,6R)-(O¹-Ethylphosphonyl)-2,4-bis(benzyloxy)-

6-propyloxycyclohexanol 10c

This compound was prepared in a manner identical with that

described for the phosphonate 10b using ethyl phosphonic

dichloride (397 mg, 2.71 mmol) as the phosphonylating agent

to give phosphonate 10c as a colourless oil which solidiﬁed

upon standing (58 mg, 23%); mp 53–56 ЊC (HRMS: found:

[M ϩ Na]^ϩ, 485.2053. C₂₅H₃₅O₆NaP requires 485.2069); [α]_D

Ϫ21.8 (c 0.17 in CH₂Cl₂); ν_max(KBr disc)/cm^Ϫ13431 br, 2919,

1649, 1495, 1459 and 1362; δ_H(300 MHz; C²HCl₃) 0.90 (3 H, t,

J_HH7.5, OCH₂CH₂CH₃), 1.11–1.88 (9 H, m, PCH₂CH₃,

OCH₂CH₂CH₃, 1 × 3-H and 1 × 5-H), 2.49–2.29 and 2.43–

2.47 (2 × 1 H, 2 × m, 1 × 3-H and 1 × 5-H), 3.53 (2 H, m,

OCH₂CH₂CH₃), 3.69–3.82 (2 H, m, 6-H and 4-H), 4.09 (1 H,

m, 2-H), 4.35 (1H, d t, J 2.5 and 9, 1-H), 4.50 (2 H, ABq,

2

δ_C(125 MHz; H₂O) 9.28 (d, J_CP6, PCH₂CH₃), 10.77 (OCH₂-

CH₂CH₃), 22.51 (d, J_CP164, PCH₂CH₃), 25.01 (OCH₂CH₂-

CH₃), 26.31 (3-C of Cha), 26.81 (4-C of Cha), 32.86 (2-C of

Cha), 40.31 (3-C), 40.45 (5-C), 52.92 (1-C of Cha), 66.42 (4-C),

71.16 (2-C), 75.05 (OCH₂CH₂CH₃), 77.56 (d, J_CP3, 6-C) and

80.53 (d, J_CP6, 1-C); δ_P(121 MHz; ²H₂O) 33.60; m/z (ES TOF)

305 (100%, [M ϩ Na]^ϩ) and 283 (10, [M ϩ H]^ϩ).

O r g . B i o m o l . C h e m . , 2 0 0 4 , 2, 6 7 1 – 6 8 8

678

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(؊)-(1S,2R,4S,6R)-2,4-Bis(benzyloxy)-(N-benzyloxycarbonyl)-

(N-methyl)-6-aminocyclohexanol 11d

125.62, 125.77, 125.81, 126.43 and 126.47 (Ar-CH), 134.89,

136.16 and 136.46 (Ar-C, quaternary) and 155.10 (CO); m/z

(ES^ϩTOF) 568.3 (100%, [M ϩ Na]^ϩ).

A mixture of epoxide (ϩ)-4 (583 mg, 1.88 mmol) and Yb()-

(OTf )₃(42 mg, 0.67 mmol) was suspended in a solution of

methylamine in water (20 cm³, 40% w/v) and the resulting solu-

tion was carefully heated at 60 ЊC in a sealed ﬂask for 12 h. The

reaction mixture was concentrated under reduced pressure and

then dissolved in a 1 M hydrochloric acid solution (20 cm³) and

extracted with diethyl ether (2 × 30 cm³). To the resulting aque-

ous layer was added, dropwise, a 4 M NaOH solution until the

pH was 14. The solution was then extracted with ethyl acetate

(3 × 50 cm³) and the pooled organic fractions were washed with

brine (100 cm³), dried (MgSO₄), ﬁltered and then concentrated

under reduced pressure to give the intermediate amine as a yel-

low oil (569 mg, 89%). A portion of this material (318 mg, 1.32

mmol) was dissolved in dry dichloromethane (40 cm³) and to

this stirred solution was added DMAP (22 mg, 0.180 mmol)

and TEA (202 mm³, 1.45 mmol). After cooling to 5 ЊC benzyl

chloroformate (581 mm³, 1.45 mmol) was added and stirring

continued at room temperature for 16 h. The solution was con-

centrated under reduced pressure to give an oil which was dis-

solved in ethyl acetate (30 cm³) and successively washed with

1 M HCl (20 cm³), water (20 cm³) and brine (20 cm³). The result-

ing organic layer was dried (MgSO₄), ﬁltered and concentrated

to give an oil that was puriﬁed by ﬂash silica column chromato-

graphy (petroleum ether–ethyl acetate; 4 : 1) to give the fully

protected N-methylamino alcohol 11d as a white solid (470 mg,

78%); mp 102–103 ЊC (Found: H, 7.05; C, 73.3; N, 2.95.

C₂₉H₃₃NO₅requires H, 7.0; C, 73.25; N, 2.95%); (HRMS:

found: [M ϩ Na]^ϩ, 498.2258. C₂₉H₃₃NO₅Na requires 498.2256);

[α]_DϪ29.1 (c 0.12 in CHCl₃); ν_max(CH₂Cl₂)/cm^Ϫ13500 br, 3165,

2921, 2850, 1689, 1494, 1400, 1344, 1149, 1088, 1026, 733 and

692; δ_H(300 MHz; C²HCl₃) 1.30–1.36 (1 H, m, 5-H), 1.50–1.58

(1 H, m, 3-H), 1.65 (1 H, br s, OH), 1.98–2.06 (1 H, m, 5-H),

2.31–2.46 (2 H, m, 3-H and 6-H), 2.87 (3 H, s, N-CH₃), 3.56

(1 H, m, 1-H), 3.84–3.90 (1 H, m, 4-H), 3.95–4.00 (1 H, m, 2-H),

4.42–4.64 (4 H, m, 2 × OCH₂Ph), 5.10–5.26 (2 H, m, CO-

OCH₂Ph) and 7.27–7.34 (15 H, m, Ar-H); δ_C(75.4 MHz;

C²HCl₃) 28.33 (N-CH₃), 34.00 and 34.16 (3-C and 5-C), 54.02

(6-C), 71.30 (OCH₂Ph), 71.70 and 76.66 (2 × OCH₂Ph), 71.30

(4-C), 71.88 (2-C), 76.87 (1-C), 127.69, 127.78 and 128.52

(Ar-CH), 136.88, 138.11 and 138.43 (3 × Ar-C quaternary) and

157.4 (CO-OCH₂Ph); m/z (ES^ϩTOF) 514.2 (75%, [M ϩ K]^ϩ)

and 498.2 (100, [M ϩ Na]^ϩ).

(؊)-(1S,2R,4S,6R)-2,4-Bis(benzyloxy)-(N-benzyloxycarbonyl)-

(N-2-phenylethyl)-6-aminocyclohexano1 11f

This compound was prepared in a manner identical with that

described for the N-methylamino alcohol 11d using epoxide

(ϩ)-4 (222 mg, 0.536 mmol) and a mixture of 2-phenyl-

ethylamine in 1,2-dichloroethane (40 cm³, 1 : 1, v/v) to give

initially the unprotected hexylamino alcohol [see compound 19f

for full experimental details and data] and then the N-2-phenyl-

ethylamino alcohol 11f as a colourless oil (53% overall yield);

(Found: H, 7.0; C, 76.5; N, 2.5. C₃₆H₃₉NO₅requires H, 6.95; C,

76.45; N, 2.5%); (HRMS: found: [M ϩ Na]^ϩ, 588.2729.

C₃₆H₃₉NO₅Na requires 588.2726); [α]_DϪ78.6 (c 0.12 in CHCl₃);

ν_max(CH₂Cl₂)/cm^Ϫ13440 br, 3090, 3050, 3020, 2938, 2856, 1691

br, 1500, 1413, 1276, 1102, 1075, 1032, 738 and 699; δ_H(300

MHz; C²HCl₃) 1.27–1.71 (2 H, m, 3-H and 5-H), 2.16 (1 H, br s,

OH), 2.36–2.51 (2 H, m, 3-H and 5-H), 2.76–3.13 (3 H, m, 1 of

N-CH₂CH₂Ph and N-CH₂CH₂Ph), 3.27–3.38 (4 H, m, 1 of

N-CH₂CH₂Ph, 6-H, 1-H and 4-H), 3.39–4.02 (1 H, m, 2-H),

4.27–4.71 (4 H, m, 2 × OCH₂Ph), 5.01–5.37 (2 H, m, CO-

OCH₂Ph) and 7.04–7.52 (20 H, m, Ar-H); δ_C(75.4 MHz;

C²HCl₃) 34.71 (N-CH₂CH₂Ph), 35.11 and 35.30 (3-C and 5-C),

36.71 (N-CH₂CH₂Ph), 55.42 (6-C), 67.44 (CO-OCH₂Ph), 70.87

and 71.81 (2 × OCH₂Ph), 72.06 (4-C), 72.11 (2-C), 76.83 (1-C),

127.78, 127.96, and 128.98 (Ar-CH), 136.86, 138.27, 138.62,

139.43 (Ar-C quaternary) and 157.31 (CO-OCH₂Ph); m/z

(ES^ϩTOF) 588.3 (100%, [M ϩ Na]^ϩ).

(؊)-(1S,2R,4S,6R)-6-Methylamino-2,4-dihydroxycyclohexyl

cyclohexylammonium 1-methylphosphonate [cyclohexyl-

ammonium salt 5d]

To a stirred solution of the 6-methylamino compound 11d (200

mg, 0.421 mmol) in dry dichloromethane (10 cm³) under argon

was added dry TEA (293 mm³, 2.12 mmol) and DMAP (51 mg,

0.421 mmol) and the solution was cooled to 5 ЊC. Methyl phos-

phonic dichloride (280 mg, 2.105 mmol) was added and the

solution was stirred at room temperature for 16 h. The solvent

was removed under reduced pressure and the residue was redis-

solved in t-butanol–water (10 cm³, 1 : 1, v/v) and the resulting

solution was heated at 50 ЊC for 2 h. After cooling, the solvent

was removed under reduced pressure and the residue par-

titioned between 10% sodium hydrogen carbonate solution

(15 cm³) and diethyl ether (15 cm³). The separated aqueous

layer was further extracted with diethyl ether (15 cm³) then acid-

iﬁed to pH 1 by addition of concentrated hydrochloric acid.

The resulting white precipitate was extracted into ethyl acetate

(3 × 20 cm³) and the pooled organic extracts were dried

(MgSO₄), ﬁltered and concentrated under reduced pressure.

The intermediate protected phosphonic acid 12d was isolated as

a white foam and dissolved in dry THF (5 cm³). Anhydrous

ammonia was condensed at Ϫ78 ЊC (dry ice–acetone bath) into

a three necked ﬂask ﬁtted with a cold-ﬁnger condenser until

approximately 10 cm³had collected. Freshly cut sodium metal

(82 mg, 3.58 mmol) was added to the stirred liquid ammonia

and the solution turned a deep blue colour. The solution of 12d

in THF was then added via a cannula and the resulting solution

was stirred under argon at Ϫ78 ЊC for 30 min. MeOH (2 cm³)

was added (causing the solution to become colourless) and the

solution allowed to stand at room temperature for 16 h. The

volatile solvents were then removed under reduced pressure to

give a white powder which was dissolved in water (2 cm³) and

was passed through a column of Amberlite IR 118H cation

exchange resin (H^ϩform, eluting with water). The acidic frac-

tions were pooled and pH adjusted to 6 by dropwise addition of

a 1% solution of freshly distilled cyclohexylamine in water. The

solution was then lyophilised and the resulting white solid was

(؊)-(1S,2R,4S,6R)-2,4-Bis(benzyloxy)-(N-benzyloxycarbonyl)-

(N-hexyl)-6-aminocyclohexano1 11e

This compound was prepared in a manner identical with that

described for the N-methylamino alcohol 11d using epoxide

(ϩ)-4 (260 mg, 0.83 mmol) and n-hexylamine (111 mm³,

0.83 mmol) to give initially the unprotected hexylamino alcohol

[see compound 19e for full experimental details and data] and

then the N-Cbz protected N-hexylamino alcohol 11e as a pale

yellow oil (68% overall yield) (Found: H, 7.95; C, 74.9; N, 2.6.

C₃₄H₄₃NO₅requires H, 7.95; C, 74.85; N, 2.55%); (HRMS:

found: [M ϩ Na]^ϩ, 568.3045. C₃₄H₄₃NO₅Na requires 568.3039);

[α]_DϪ27.2 (c 0.11 in CHCl₃); ν_max(CH₂Cl₂)/cm^Ϫ13325 br, 3091,

3029, 2926, 2865, 1689, 1452, 1416, 1350, 1262, 1041, 1114, 739

and 693; δ_H(300 MHz; C²HCl₃) 0.88 (3 H, s, CH₃) 1.05–1.39

(8 H, m, 6 of CH₂(CH₂)₄CH₃, 3-H and 5-H), 1.40–1.67 [3 H, m,

2 of CH₂(CH₂)₄CH₃and OH], 2.10–2.46 [3 H, m, 1 of CH₂-

(CH₂)₄CH₃, 3-H and 5-H], 2.98–3.29 [2 H, m, 1 of CH₂-

(CH₂)₄CH₃and 6-H], 3.51–3.84 (2 H, m, 1-H and 4-H), 3.90

(1 H, s, 2-H), 4.47–4.70 (4 H, m, 2 × OCH₂Ph), 5.02–5.27 (2 H,

m, CO-OCH₂Ph) and 7.15–7.42 (15 H, m, Ar-H); δ_C(75.4 MHz;

C²HCl₃) 12.01, 20.57, 24.76, 28.12 and 29.45 [CH₂(CH₂)₄CH₃],

31.93 (3-C), 33.5 (5-C), 41.20–42.70 (br, 6-C), 43.20–45.10

[CH₂(CH₂)₄CH₃], 67.34 (CO-OCH₂Ph), 68.76 and 69.72

(2 × OCH₂Ph), 69.89 and 70.03 (2-C and 4-C), 74.92 (1-C),

O r g . B i o m o l . C h e m . , 2 0 0 4 , 2, 6 7 1 – 6 8 8

679

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crystallised from MeOH–diethyl ether to give the N-methyl

methylphoshonate 5d as a white crystalline solid (51 mg, 36%);

mp >200 ЊC (decomp.) (HRMS: found: [M ϩ H]^ϩ, 240.1008.

C₈H₁₉NO₅P requires 240.1001); [α]_DϪ27.3 (c 1.0 in MeOH);

ν_max(KBr disc)/cm^Ϫ13415 br, 2937, 1634, 1540 and 1456; δ_H(300

(Ar-CH), 137.37, 139.23, 139.42, 139.65 (Ar-C quaternary) and

159.31 (CO-OCH₂Ph); δ_P(121 MHz; C²HCl₃) 33.85; m/z (ES^ϩ

TOF) 704.5 (20%, [M Ϫ H ϩ Na ϩ K]^ϩ), 688.5 (25, [M Ϫ H ϩ

2Na]^ϩ), 682.4 (50, [M ϩ K]^ϩ), 666.5 (100, [M ϩ Na]^ϩ), 496 (25,

[M Ϫ CH₄PO₃Ϫ C₇H₇ϩ K]^ϩ) and 480.4 (40, [M Ϫ CH₄PO₃

Ϫ C₇H₇ϩ Na]^ϩ).

2

MHz; H₂O) 0.98–1.24 (8 H, m, H-4 of Cha, 2 × 2-H of Cha,

2 × 3-H of Cha and PCH₃), 1.33–1.51 (3 H, m, 4-H of Cha, 3-H

and 5-H), 1.57–1.72 (2 H, m, 2 × 3-H of Cha), 1.76–1.88 (2 H,

m, 2 × 2-H of Cha), 1.98–2.08 (1 H, m, 3-H), 2.23–2.09 (1 H, m,

5-H), 2.60 (3 H, s, NCH₃), 2.92–3.04 (1 H, m, 1-H of Cha),

3.32–3.41 (1 H, m, 6-H), 3.91–4.00 (1 H, m, 4-H), 4.05–4.12

(1 H, m, 1-H) and 4.16–4.22 (1 H, m, 2-H); δ_C(75 MHz; ²H₂O)

14.86 (d, J_PC136.5, PCH₃), 26.27, 26.76 and 32.79 (3 × CH₂of

Cha), 32.43 (NHCH₃), 35.68 (3-C), 40.12 (5-C), 57.91 (d, J_CP5,

6-C), 65.63 (4-C), 69.97 (2-C) and 76.33 (d, J_CP5.5, 1-C); δ_P(121

MHz; ²H₂O) 27.24; m/z (TOF ES^ϩ) 284 (15%, [M Ϫ H ϩ

2Na]^ϩ), 262 (70, [M ϩ Na]^ϩ) and 240 (100, [M ϩ H]^ϩ).

(؊)-(1S,2R,4S,6R)-6-Hexylamino-2,4-dihydroxycyclohexyl

1-methylphosphonate 5e

To a stirred blue solution of freshly cut sodium (84 mg,

3.65 mmol) in liquid ammonia (15 cm³) was added dropwise a

solution of the fully protected N-hexyl methylphoshonate 12e

(252 mg, 0.405 mmol) in dry THF. After stirring for 1 h, 2 drops

of MeOH were added and the mixture was left to evaporate.

The resulting yellow solid was washed thoroughly with MeOH

and the ﬁltrate concentrated under reduced pressure to give a

yellow solid which was dissolved in water and loaded onto a

cation exchange chromatography column (Amberlyst 131 WET

resin, H^ϩform, 4.5 g). The column was eluted with water and

the acidic fractions were combined and concentrated under

reduced pressure to give a pale yellow solution that was lyophil-

ised. The resulting crude white solid was recrystallised from

hexane–MeOH (8 : 1) to give triol 5e as a white solid (55 mg,

44%); mp >166 ЊC (decomp.); (Found: H, 9.15; C, 50.55; N,

4.55. C₁₃H₂₈NO₄P requires H, 9.1; C, 50.5; N, 4.55%); (HRMS:

found: [M ϩ H]^ϩ, 293.1756. C₁₃H₂₉NO₄P requires 310.1783);

[α]_DϪ33.6 (c 0.12 in CHCl₃); ν_max(CH₂Cl₂)/cm^Ϫ13445 br, 3092,

3058, 3028, 2956, 2867, 1690, 1349, 1265, 1026, 1133 and 1042;

δ_H(300 MHz; C²H₃O²H) 0.89 (3 H, s, CH₃), 1.15–1.89 [11 H, m,

6 of CH₂(CH₂)₄CH₃, P-CH₃, 3-H and 5-H], 1.92–2.64 [4 H, m,

2 of CH₂(CH₂)₄CH₃, 3-H and 5-H], 2.82–2.43 [3 H, m,

CH₂(CH₂)₄CH₃and 6-H], 3.42–3.66 (1 H, m, 4-H), 3.71–3.79

(1 H, m, 2-H) and 3.92–4.06 (1 H, m, 1-H); δ_C(75.4 MHz;

(؊)-(1S,2R,4S,6R)-(O¹-Methylphosphonyl)-2,4-bis(benzyloxy)-

N-benzyloxycarbonyl-6-hexylamino-cyclohexanol 12e

This compound was prepared in a manner identical with that

described for the phosphonate 10b using n-hexylamino alcohol

11e (340 mg, 0.624 mmol) and methyl phosphonic dichloride as

the phosphorylating agent to give the fully protected N-hexyl

methylphoshonate 12e as a colourless oil (327 mg, 84%);

(Found: H, 7.45; C, 67.5; N, 2.25. C₃₅H₄₆NO₇P requires H, 7.45;

C, 67.4; N, 2.25%); (HRMS: found: [M ϩ Na]^ϩ, 646.2908.

C₃₅H₄₆NO₇PNa requires 646.2910); [α]_DϪ26.7 (c 0.12 in

CHCl₃); ν_max(CH₂Cl₂)/cm^Ϫ13447 br, 3096, 3061, 3031, 2922,

2856, 2617 br, 1756, 1691, 1500, 1419, 1353, 1195, 1095, 1065,

754 and 694; δ_H(300 MHz; C²HCl₃) 0.81–0.94 [3 H, m,

CH₂(CH₂)₄CH₃], 1.11–1.78 [13 H, m, CH₂(CH₂)₄CH₃, P-CH₃,

3-H and 5-H], 2.19–2.30 (2 H, m, 3-H, 5-H), 2.72–3.68 [3 H, m

CH₂(CH₂)₄CH₃and 6-H], 3.68–3.75 (1 H, m, 4-H), 4.07–4.19

(1 H, m, 2-H), 4.43–4.82 (5 H, m, 2 × OCH₂Ph and 1-H), 4.99–

5.36 (2 H, m, CO-OCH₂Ph), 7.23–7.32 (15 H, m, Ar-H) and

10.46 (I H, br, OH); δ_C(75.4 MHz; C²HCl₃) 16.48, 25.0, 28.95,

31.20 and 32.41 [CH₂(CH₂)₄CH₃], 34.00 (P-CH₃), 35.26 (3-C),

37.46 [CH₂(CH₂)₄CH₃], 37.78 (5-C), 54.13–61.12 (br, 6-C),

69.41 (CO-OCH₂Ph), 73.06 and 74.26 (2 × OCH₂Ph), 75.60

(4-C), 77.08 (2-C), 77.45 (1-C), 130.03, 130.35, 130.42, 130.57,

130.78 and 130.88 (Ar-CH), 139.18, 141.03 and 141.68 (Ar-C

quaternary) and 158.11 (CO-OCH₂Ph); δ_P(121 MHz; C²HCl₃)

33.17; m/z (ES^ϩTOF) 646.2 (100%, [M ϩ Na]^ϩ) and 528.2 (45,

[M Ϫ CH₄PO₃]^ϩ).

1

C²H₃O²H) 14.24 [CH₂(CH₂)₄CH₃], 17.56 (d, J_P-C145.0,

P-CH₃), 23.36, 27.18, 27.28 and 32.31 [CH₂(CH₂)₄CH₃], 35.65

(3-C), 39.40 (5-C), 46.17 [CH₂(CH₂)₄CH₃], 58.23 (6-C), 64.35

and 64.45 (2-C and 4-C) and 71.34 (1-C); δ_P(121 MHz; C²HCl₃)

28.10; m/z (ES^ϩTOF) 310.2 (100%, [M Ϫ H]^ϩ) and 214.2 (50,

[M Ϫ CH₃PO₃H]^ϩ).

(؊)-(1S,2R,4S,6R)-6-(2-Phenylethyl)amino-2,4-dihydroxy-

cyclohexyl 1-methylphosphonate 5f

To a stirred blue solution of calcium turnings (94 mg,

2.35 mmol) in liquid ammonia (15 cm³) was added dropwise a

solution of the fully protected N-(2-phenylethyl) methyl-

phoshonate 12f (150 mg, 0.235 mmol) in dry THF. After 1 h,

2 drops of MeOH were added and the reaction mixture left to

evaporate. The resulting yellowish solid was then washed thor-

oughly with MeOH and the ﬁltrate concentrated under reduced

pressure to give a yellow solid which was dissolved in water

(2 cm³) and then loaded on to a cation exchange chromato-

graphy column (Amberlyst 131 WET resin, 4.5 g). The column

was eluted with water and the acidic fraction concentrated

under reduced pressure to give a pale yellow solution that was

lyophilised. The crude white solid was recrystallised from hex-

ane–MeOH (8 : 1) to give the triol 5f as a white solid (36 mg,

47%); mp >145 ЊC (decomp.); (Found: H, 7.4; C, 54.8; N, 4.25.

C₁₅H₂₄NO₅P requires H, 7.35; C, 54.7; N, 4.25%); (HRMS:

found: [M ϩ Na]^ϩ, 352.1288. C₁₅H₂₄NO₅NaP requires 352.1290);

[α]_DϪ24.5 (c 0.13 in CHCl₃); ν_max(CH₂Cl₂)/cm^Ϫ13447 br, 3096,

3063, 3031, 2922, 2856, 2617 br, 1756, 1691, 1500, 1419, 1353,

1195, 1095, 1065, 754 and 694; δ_H(300 MHz; C²H₃O²H) 1.33

(؊)-(1S,2R,4S,6R)-(O¹-Methylphosphonyl)-2,4-bis(benzyloxy)-

N-benzyloxycarbonyl-6-(2-phenylethyl)amino-cyclohexanol 12f

This compound was prepared in a manner identical with that

described for the phosphonate 10b using the 2-phenylethyl-

amino alcohol 11f (305 mg, 0.540 mmol) and methyl phos-

phonic dichloride as the phosponylating agent to give the fully

protected N-(2-phenylethyl) methylphoshonate 12f as a yellow

oil (312 mg, 90%); (Found: H, 6.6; C, 69.1; N, 2.2. C₃₇H₄₂NO₇P

requires H, 6.6; C, 69.05; N, 2.2%); (HRMS: found: [M ϩ Na]^ϩ,

666.2612. C₃₇H₄₂NO₇PNa requires 666.2597); [α]_DϪ31.8

(c 0.14 in CHCl₃); ν_max(CH₂Cl₂)/cm^Ϫ13346–3548 br, 3096, 3063,

2922, 2856, 2617 br, 1756, 1691, 1500, 1419, 1353, 1195, 1095,

1065, 754 and 694; δ_H(300 MHz; C²HCl₃) 1.08–1.48 (5 H, m,

P-CH₃, 3-H and 5-H), 1.98–2.07 (2 H, m, N-CH₂CH₂Ph),

2.21–2.33 (2 H, m, 3-H and 5-H), 2.77–2.87 (2 H, m N-CH₂-

CH₂Ph), 3.50–3.56 (1 H, m, 6-H), 3.69–3.74 (1 H, m, 4-H),

3.76–3.83 (1 H, m, 4-H), 4.07–4.11 (1 H, m, 2-H), 4.17–4.77

(5 H, m, 2 × OCH₂Ph and 1-H), 5.13–5.41 (2 H, m, CO-

OCH₂Ph), 6.67 (1 H, br s, OH) and 7.22–7.35 (20 H, m, Ar-H);

δ_C(75.4 MHz; C²HCl₃) 31.73 (P-CH₃), 33.22 and 34.84 (3-C

and 5-C), 35.67 (N-CH₂CH₂Ph), 45.39 (N-CH₂CH₂Ph), 54.63

(6-C), 67.59 (CO-OCH₂Ph), 71.37 and 73.62 (2 × OCH₂Ph),

72.89 (4-C), 73.00 (2-C), 76.45 (1-C), 127.27, 128.40, and 129.55

2

(3 H, d, J 16.7, P-CH₃), 1.44–162 (2 H, m, 3-H and 5-H),

H-H

2.93–3.10 (2 H, m CH₂CH₂Ph), 3.24–3.37 (1 H, m, 6-H), 3.41–

3.58 (2 H, m CH₂CH₂Ph), 3.95–4.08 (1 H, m, 4-H), 4.18–4.24

(1 H, m, 2-H), 4.24–4.32 (1 H, m, 1-H) and 7.31–7.39 (20 H, m,

1

Ar-H); δ_C(75.4 MHz; C²H₃O²H) 14.82 (d, J_P-C138.0, P-CH),

34.39 and 36.41 (3-C and 5-C), 40.23 (CH₂CH₂Ph), 48.79

(N-CH₂CH₂Ph), 56.62 (6-C), 65.79 (4-C), 70.29 (2-C), 76.32

O r g . B i o m o l . C h e m . , 2 0 0 4 , 2, 6 7 1 – 6 8 8

680

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(1-C), 130.06, 131.49 and 131.75 (Ar-CH) and 139.03 (Ar-C

quaternary); δ_P(121 MHz; C²HCl₃) 27.93; m/z (ES^ϩTOF) 703.3

(10%, [(2 × M) Ϫ H ϩ 2 Na]^ϩ), 681.3 (100, [(2 × M) ϩ Na]^ϩ),

352.1 (30, [M ϩ Na]^ϩ), 330.2 (25, [M ϩ H]^ϩ) and 257.1 (15,

[M Ϫ CH₄PO₃ϩ Na]^ϩ).

dried (MgSO₄) and concentrated under reduced pressure. The

resulting oil (alcohol) 18h was dissolved in MeOH (75 cm³)

and TFA (0.15 cm³) added. The solution was stirred for 6 days

and then the solvent removed under reduced pressure to give a

dirty white solid which was recrystallised from petroleum

ether–ethyl acetate (5 : 1) to give the 6-hexyloxy triol 6h

as a white solid (163 mg, 63%), mp 84–85 ЊC; (Found: H,

10.5; C, 72.1. C₁₂H₂₄O₄requires H, 10.4; C, 72.05%); (HRMS:

found: [M ϩ Na]^ϩ, 255.1517. C₁₂H₂₄O₄Na requires 255.1572);

[α]_DϪ27.8 (c 0.14 in CHCl₃); ν_max(CHCl₃)/cm^Ϫ13396 br, 2921,

2867–2921, 1456, 1374, 1070, 1170 and 1018; δ_H(300 MHz;

(؊)-(1S,2R,4R,6R)-6-Propyloxycyclohexane-1,2,4-triol 6g

To a stirred solution of (Ϫ)-(1S,2R,4S,6R)-2,4-bis(benzyloxy)-

6-propyloxycyclohexanol 9b¹⁸(636 mg, 1.72 mmol) in MeOH

(20 cm³) was added 5% palladium on activated carbon (182 mg,

0.086 mmol) followed by a small drop of acetic acid. The mix-

ture was stirred under an atmosphere of hydrogen for 16 h then

the catalyst was removed by ﬁltration through a small plug of

Celite. The solvent was removed under reduced pressure to give

the triol 6g as an oﬀ-white solid (320 mg, 98%); mp 86–89 ЊC

(HRMS: found: [M ϩ Na]^ϩ, 213.1107. C₉H₁₈O₄Na requires

213.1103); [α]_DϪ72.1 (c 1.38 in MeOH); ν_max(KBr disc)/cm^Ϫ1

3432 br, 2930, 1470 and 1451; δ_H(300 MHz, ²H₂O) 0.81 (3 H, t,

J_HH7.5, CH₃), 1.17 (1 H, q, J_HH11, 3-H), 1.39–1.55 (3 H, m,

5-H and CH₃CH₂), 1.99–2.04 (1 H, m, 5-H), 2.31–2.36 (1 H, m,

3-H), 3.39–3.59 (4 H, m, 1-H, 6-H and OCH₂), 3.90 (1 H,

ddd, J_HH4, 11 and 15.5, 4-H) and 4.02–4.03 (1 H, m, 2-H);

δ_C(75 MHz, ²H₂O) 10.87 (CH₃), 23.56 (CH₃CH₂), 38.72

and 39.44 (3-C and 5-C), 65.05, 69.95, 75.19 and 77.01 (1-C,

2-C, 4-C and 6-C), 72.67 (OCH₂); m/z (ES^ϩTOF) 213 (100%,

[M ϩ Na]^ϩ).

3

C²HCl₃) 0.94 (3 H, m, J_H-H6.4, n-hexyl), 1.21–1.43 (6 H, m,

n-hexyl) 1.48–1.72 (4 H, m, 2 of n-hexyl, 5-H and OH), 2.01

(1 H, br, OH), 2.10–2.23 (1 H, m, 3-H), 2.25–2.40 (2 H, m, 3-H

and 5-H), 2.71 (1 H, br, OH), 3.32–3.39 (1 H, m, 1 of n-hexyl),

3.55–3.69 (3 H, m, 1 of n-hexyl, 1-H and 6-H) and 4.00–4.15

(1 H, m, 4-H) and 4.16–4.20 (1 H, m, 2-H); δ_C(75.4 MHz;

C²HCl₃) 12.48, 21.05, 24.27, 28.46 and 30.09 (n-hexyl), 34.32

and 36.68 (3-C and 5-C), 62.55 (n-hexyl), 63.95 (4-C), 65.87

(6-C), 72.18 (2-C) and 73.38 (1-C); m/z (FAB^ϩ) 233.2 (100%,

[M ϩ H]^ϩ) and 197.2 (15, [M Ϫ C₃H₈– OH ϩ Na]^ϩ).

2-(3-Butenyloxy)-phenol 14³⁰

To a stirred mixture of catechol 13 (6.00 g, 54.5 mmol) and

potassium carbonate (7.53 g, 54.5 mmol) in acetone (150 cm³)

was added 4-bromo-1-butene (1.85 cm³, 18.2 mmol) and

potassium iodide (50 mg). This mixture was stirred and heated

under reﬂux for 2 days. After cooling the solvent was carefully

removed under reduced pressure and the residue was par-

titioned between diethyl ether (200 cm³) and 1 M HCl

(200 cm³). The separated organic layer was washed with water

(200 cm³) and brine (200 cm³) and then dried (MgSO₄), ﬁltered

and concentrated under reduced pressure to give a yellow

solid that was puriﬁed by ﬂash silica column chromatography

(petroleum ether–ethyl acetate; 4 : 1) to give alcohol 14 as a

colourless oil (1.09 g, 37%); bp 96–98 ЊC/4 mmHg; ν_max(thin

ﬁlm)/cm^Ϫ13528 br, 3081, 2934, 1643 and 1597; δ_H(300 MHz,

(؊)-(1R,2R,4S,6R)-1,2-Cyclohexyldienedioxy-4-(tert-butyl-

diphenylsilyloxy)-6-n-hexyloxy)cyclohexane 17h

A stirred solution of (Ϫ)-(1S,2R,4S,6R)-1,2-cyclohexylidene-

dioxy-4-(tert-butyl-diphenylsilyloxy)cyclohexan-6-ol 16¹⁸(1.12

g, 2.37 mmol) in dry DMF (20 cm³) was cooled to 0 ЊC in an

ice–water bath. NaH (270 mg, 6.77 mmol) was added in small

portions under argon followed by dropwise addition of n-hexyl

iodide (3.56 cm³, 23.7 mmol). The reaction mixture was grad-

ually brought to room temperature and stirred for 12 h. The

resulting solution was diluted with water (40 cm³) and then

extracted with ethyl acetate (3 × 50 cm³). The organic layers

were pooled and washed with brine (40 cm³), dried (MgSO₄),

ﬁltered and then concentrated under reduced pressure. The

resulting yellow oil was puriﬁed by ﬂash silica column chrom-

atography (petroleum ether–ethyl acetate; 4 : 1) to give the

6-hexyloxycyclohexane 17h as a colourless oil (834 mg, 64%);

(Found: H, 9.2; C, 74.2. C₃₄H₅₀O₄Si requires H, 9.15; C,

74.15%); (HRMS: found: [M ϩ Na]^ϩ, 573.3378. C₃₄H₅₀O₄Si Na

requires 573.3376); [α]_DϪ33.7 (c 0.14 in CHCl₃); ν_max(Nujol)/

cm^Ϫ13085, 3063, 3028, 2932, 2857, 1462, 1363, 1112, 1058, 940,

823, 739 and 702; δ_H(300 MHz; C²HCl₃) 0.88 [3 H, t, ³J_H-H7.2,

CH₂(CH₂)₄CH₃], 1.05 [6 H, s, 6 of C(CH₃)₃], 1.07 [3 H, s, 3 of

C(CH₃)₃], 1.24–2.30 [22 H, m, cyclohexylidene, CH₂(CH₂)₄CH₃,

2 × 3-H and 2 × 5-H], 3.05–4.35 [6 H, m, CH₂(CH₂)₄CH₃, 1-H,

2-H, 4-H and 6-H], 7.32–7.45 (6 H, m, Ar-H) and 7.64–7.77

(4 H, m, Ar-H); δ_C(75.4 MHz; C²HCl₃) 16.65 [CH₂(CH₂)₄-

CH₃], 21.10 [SiC(CH₃)₃], 25.21, 26.51, 27.69, 28.30, 29.53,

32.47, 34.20, 35.80, 37.69, 37.80, 38.75 and 40.55 (C × 5, cyclo-

hexylidene, CH₂(CH₂)₄CH₃, C(CH₃)₃, 3-C and 5-C), 68.86

(4-C), 71.17 [CH₂(CH₂)₄CH₃], 74.00 (2-C), 75.87 (C-6), 81.69

(1-C), 119.39 (C quaternary, cyclohexylidene), 129.95, 130.16

and 132.19 (Ar-CH) and 138.36 and 138.75 (Ar-C quaternary);

m/z (ES^ϩTOF) 573.4 (100%, [M ϩ Na]^ϩ).

C²HCl ) 2.54–2.61 (2 H, m, CH CH CH᎐CH ), 4.11 (2 H, t,

᎐

3

2

J_HH7, OCH ), 5.13–5.24 (2 H, m, CH ᎐CH), 5.66 (1 H, s,

᎐

2

ArCOH ), 5.83–5.96 (1 H, m, CH ᎐CH ), 6.83–6.96 (4 H, m,

᎐

2

Ar-CH); δ_C(75 MHz; C²HCl₃) 34.50 (OCH₂CH₂), 68.93

(OCH₂), 112.98, 115.46, 120.93, and 122.54 (Ar-CH), 118.36

(CH᎐CH ), 135.01 (CH ᎐CH), 146.84 and 146.86 (Ar-C

᎐

2

quaternary); m/z (EI^ϩ) 164 (34%, M^ϩ), 110 (73, [C₆H₄(OH)₂]^ϩ)

and 55 (100%, CH CH CH᎐CH ^ϩ).

᎐

2

2-(4-Hydroxybutyloxy)-phenol 15

To a stirred solution of alcohol 14 (227 mg, 1.38 mmol) in dry

THF (10 cm³), under argon, was added borane–THF complex

(1 M solution in THF, 0.914 cm³) and the mixture stirred for

1 h. Sodium hydroxide solution (1 M, 2.74 cm³) was then added

and when eﬀervescence had ceased hydrogen peroxide solution

(1 M, 2.74 cm³) was added and the solution stirred for 3 h. 1 M

HCl (10 cm³) was then added and the separated aqueous layer

was extracted with diethyl ether (2 × 20 cm³). The pooled ether-

eal extracts were dried (MgSO₄), ﬁltered and concentrated

under reduced pressure to a yellow oil that was puriﬁed by ﬂash

silica column chromatography (hexane–ethyl acetate; 1 : 1) to

give diol 15 as a white crystalline compound (169 mg, 67%); mp

82–84 ЊC (HRMS: found: [M ϩ Na]^ϩ, 205.0387, C₁₀H₁₄O₃Na

requires 205.0841); ν_max(KBr disc)/cm^Ϫ13461, 3066, 2963, 2865

and 1597; δ_H(300 MHz; C²HCl₃) 1.63 (1 H, br s, CH₂OH ),

1.73–1.82 and 1.90–1.99 (4 H, m, OCH₂CH₂CH₂), 3.76 (2 H, t,

J_HH6, OCH₂), 4.10 (2 H, t, J_HH6, OCH₂), 5.91 (1 H, br s,

ArCOH ) and 6.80–6.96 (Ar-H); δ_C(75 MHz; C²H₃O²H) 28.75

and 32.02 (OCH₂CH₂CH₂), 64.53 and 71.69 [OCH₂(CH₂)₂-

CH₂OH], 116.06, 118.26, 122.74 and 124.11 (Ar-CH) and

149.71 and 150.13 (Ar-C quaternary); m/z (EI^ϩ) 182 (12%, M^ϩ)

and 110 (100, [C₆H₄(OH)₂]^ϩ).

(؊)-(1S,2R,4S,6R)-6-Hexyloxycyclohexane-1,2,4-triol 6h

To a stirred solution of the 6-hexyloxy silyl ether 17h (802 mg,

1.46 mmol) in DCM (30 cm³) was added, dropwise, TBAF

(1.6 cm³, 1.60 mmol). After 5 h, the resulting solution was

diluted with DCM (30 cm³) and water (40 cm³). The aqueous

layer was then back extracted with DCM (3 × 40 cm³). The

organic layers were pooled and washed with brine (50 cm³),

O r g . B i o m o l . C h e m . , 2 0 0 4 , 2, 6 7 1 – 6 8 8

681

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(1S,2R,4S,6R)-2,4-Bis(benzyloxy)-6-[4-(2-hydroxyphenyloxy)-

butyloxy]cyclohexanol 9i

(N-Benzyloxycarbonyl)-3-aminopropanol

To a stirred solution of 3-aminopropanol (7.65 cm³, 100 mol) in

1,4-dioxane–water (1 : 1, v/v, 200 cm³) was added DMAP

(50 mg, 4 mmol), benzyl chloroformate (16 cm³, 110 mmol) and

TEA (13.9 cm³, 100 mmol) and the reaction mixture stirred for

12 h. The solvents were removed under reduced pressure to give

a thick solution which was diluted with DCM (150 cm³) and

water (50 cm³). The aqueous layer was then back-extracted with

DCM (3 × 50 cm³). The pooled organic fractions were then

successively washed with 0.1 M solution of HCl (100 cm³),

water (100 cm³), and brine (100 cm³), dried (MgSO₄) and ﬁl-

tered. Removal of the solvents under reduced pressure gave a

white solid which was recrystallised from petroleum ether–ethyl

acetate (3 : 1) to give the Cbz protected 3-aminopropanol as

a white solid (19.2 g, 92%); mp 51–52 ЊC (lit.,³²51–52 ЊC);

(Found: H, 7.3; C, 63.2; N, 6.7. Calc. for C₁₁H₁₅O₃: H, 7.25; C,

63.15; N, 6.7%); (HRMS: found: [M ϩ Na]^ϩ, 232.0956. Calc.

for C₁₁H₁₅O₃Na: 232.0950); ν_max(CH₂Cl₂)/cm^Ϫ13325 br, 3057,

3043, 3029, 2955, 2873, 1682, 1536, 1453, 1327, 1265, 1143,

1024, 750, 725 and 697; δ_H(300 MHz; C²HCl₃) 1.69 (2 H, qt,

³J_H-H5.9, NCH₂CH₂CH₂OH), 2.62 (1 H, br, OH), 3.35 (2 H, q,

To a stirred solution of diol 15 (134 mg, 0.738 mmol) in

1,2-dichloroethane (5 cm³) was added the epoxide (ϩ)-4

(76 mg, 0.246 mmol) as a solution in 1,2-dichloroethane

(5 cm³). Yb()(OTf )₃(31 mg, 0.049 µmol) was then added

and the solution was heated under reﬂux for 1 h. After cool-

ing the solvent was removed under reduced pressure and the

residue was dissolved in diethyl ether (20 cm³) and washed

with water (2 × 20 cm³) and brine (20 cm³). The ethereal

solution was dried (MgSO₄), ﬁltered and concentrated under

reduced pressure to give a brown oil that was puriﬁed by

ﬂash silica column chromatography (petroleum ether–ethyl

acetate; 1 : 1) to give the alcohol 9i as a colourless oil (87 mg,

72%); (HRMS: found: [M ϩ Na]^ϩ, 515.2401, C₃₀H₃₆O₆Na

requires 515.2410); [α]_DϪ7.66 (c 1.11 in MeOH); ν_max(thin

ﬁlm)/cm^Ϫ13538 br, 3425 br, 3086, 3060, 3024, 2943, 2880,

1740, 1602 and 1509; δ_H(300 MHz; C²HCl₃) 1.27–1.44 (2 H,

m, 3-H and 5-H), 1.75–1.96 [4 H, m, OCH₂(CH₂)₂], 2.42–

2.50 (2 H, m, 3-H and 5-H), 3.52–3.74 (5 H, 1-H, 2-H, 6-H and

OCH₂CH₂), 3.95–3.96 (1 H, m, 4-H), 4.08 (2 H. t. J_HH6,

OCH₂CH₂), 4.51 (2 H, ABq, J_HH12, ArCH₂), 4.60 (2 H, s,

ArCH₂), 5.91 (1 H, br s, ArCOH ) and 6.62–6.95 and 7.26–7.39

(14 H, m, Ar-H); δ_C(75 MHz; C²HCl₃) 25.27 and 25.87

[OCH₂(CH₂)₂], 33.06 and 34.51 (3-C and 5-C), 67.88 and

68.09 [OCH₂(CH₂)₂CH₂O], 69.70 and 71.10 (PhCH₂O), 70.69,

74.58, 75.41 and 76.18 (1-C, 2-C, 4-C and 6-C), 111.08, 113.85,

119.10, 120.56, 126.70, 126.79 and 127.50 (Ar-CH), 138.50,

139.00 and 143.40 (Ar-C quaternary); m/z (ES^ϩTOF) 515

(100%, [M ϩ Na]^ϩ).

³J_H-H6.3, NCH₂CH₂CH₂OH), 3.66 (2 H, t, J_H-H6.3, NCH₂-

3

CH₂CH₂OH), 5.09 (3 H, s, OCH₂Ph and NH) and 7.25–7.36

(5 H, m, Ar-H); δ_C(75.4 MHz; C²HCl₃) 31.82 (NCH₂CH₂-

CH₂OH), 37.01 (NCH₂CH₂CH₂OH), 58.85 (NCH₂CH₂-

CH₂OH), 66.15 (CO-OCH₂Ph), 127.38, 127.45 and 127.82 (Ar-

CH), 135.72 (Ar-C quaternary) and 156.12 (CO); m/z (ES^ϩ

TOF) 232.1 (100%, [M ϩ Na]^ϩ).

(؊)-(1S,2R,4S,6R)-2,4-Bis(benzyloxy)-6-(N-benzyloxy-

carbonyl-3-aminopropyl)cyclohexanol 9k

(؊)-(1S,2R,4S,6R)-2,4-Bis(benzyloxy)-6-(N-benzyloxy-

This compound was prepared in a manner identical with that

described for (2-aminoethyl) alcohol 9j using (N-benzyloxy-

carbonyl)-3-aminopropanol (284 mg, 1.36 mmol) to give alco-

hol 9k as a colourless oil (279 mg, 52%); (Found: H, 7.2; C,

71.7; N, 2.7. C₃₁H₃₇NO₆requires H, 7.2; C, 71.65; N, 2.7%);

(HRMS: found: [M ϩ Na]^ϩ, 542.2533. C₃₁H₃₇NO₆Na requires

542.2519); [α]_DϪ22.1 (c 0.12 in CHCl₃); ν_max(CH₂Cl₂)/cm^Ϫ1

3410 br, 3086, 3053, 3030, 2932, 2861, 1711, 1562, 1454, 1362,

1258, 1217, 1093, 1028, 752 and 697; δ_H(300 MHz; C²HCl₃)

1.25–1.42 (2 H, m, 3-H and 5-H), 1.74 (2 H, t, ³J_H-H5.7, OCH₂-

CH₂CH₂NH), 2.31–2.43 (2 H, m, 3-H and 5-H), 2.73 (1 H, d,

³J_H-H5.9, OH), 3.24–3.55 (5 H, m, OCH₂CH₂CH₂NH and 1-H),

3.67–3.75 (2 H, m, 4-H and 6-H), 3.90–3.96 (1 H, m, 2-H), 4.45

(1 H, A of ﬁrst AB system, ²J_H-H11.8, 1 of OCH₂Ph), 4.51 (1 H,

B of ﬁrst AB system, ²J_H-H11.8, 1 of OCH₂Ph), 4.53 (1 H, A of

carbonyl-2-aminoethyl)cyclohexanol 9j

To a stirred solution of epoxide (ϩ)-4 (261 mg, 0.842 mmol) in

1,2-dichloroethane (30 cm³) was added Yb()(OTf )₃(25 mg,

0.32 mmol) followed by N-benzyloxycarbonyl-ethanolamine³¹

(378 mg, 1.94 mmol) and the resulting mixture heated under

reﬂux for 20 h. The reaction mixture was concentrated under

reduced pressure and then dissolved in 1 M HCl solution

(20 cm³). To this solution was added, dropwise, a 4 M sodium

hydroxide solution until the pH was 14. The solution was then

extracted with ethyl acetate (3 × 50 cm³) and the pooled organic

fractions were washed with brine (50 cm³), dried (MgSO₄),

ﬁltered and then concentrated under reduced pressure to give

a light pale yellow oil which was puriﬁed by ﬂash silica

column chromatography (ethyl acetate–MeOH; 9 : 1 with addi-

tion of 0.5 cm³of TEA per 10 cm³of solvent) to give alcohol

9j as a colourless oil (302 mg, 71%); (Found: H, 7.0; C,

71.3; N, 2.8. C₃₀H₃₅O₆requires H, 6.7; C, 71.25; N, 2.75%);

(HRMS: found: [M ϩ Na]^ϩ, 528.2359. C₃₀H₃₅O₆Na requires

528.2362); [α]_DϪ20.7 (c 0.12 in CHCl₃); ν_max(CH₂Cl₂)/cm^Ϫ1

3410 br, 3086, 3056, 3030, 2932, 2861, 1711, 1562, 1454, 1362,

1258, 1217, 1093, 1028, 752 and 697; δ_H(300 MHz; C²HCl₃)

1.22–1.46 (2 H, m, 3-H and 5-H), 2.29–2.44 (2 H, m, 3-H and

5-H), 2.56 (1 H, br, OH), 3.32–3.41 (2 H, m, OCH₂CH₂NH),

3.44–3.63 (3 H, m, 1-H, 2-H and 6-H), 3.72–3.79 (2 H, m,

OCH₂CH₂NH), 3.91–3.97 (1 H, m, 2-H), 4.47 (1 H, A of

ﬁrst AB system, ²J_H-H11.8, 1 of OCH₂Ph), 4.51–4.55 (2 H, B of

ﬁrst AB system and A of second AB system, 2 × 1 of OCH₂Ph),

2

second AB system, J_H-H11.7, 1 of OCH₂Ph), 4.57 (1 H, B of

2

second AB system, J_H-H11.7, 1 of OCH₂Ph), 5.07 (2 H, s,

CO-OCH₂Ph), 5.47 (1 H, br, NH) and 7.25–7.33 (15 H, m,

Ar-H); δ_C(75.5 MHz; C²HCl₃) 31.98 and 35.73 (3-C and 5-C),

37.23 (OCH₂CH₂CH₂NH), 41.03 (OCH₂CH₂CH₂NH), 68.58

(OCH₂CH₂CH₂NH), 69.71 (CO-OCH₂Ph), 72.73 (OCH₂Ph),

73.64 (4-C), 73.98 (OCH₂Ph), 77.25, 78.49 and 79.40 (1-C, 2-C

and 6-C), 129.75, 129.86, 130.07 and 130.56 (Ar-CH), 137.16,

139.16 and 139.91 (Ar-C quaternary) and 157.15 (CO);

m/z (ES^ϩTOF) 542.3 (100%, [M ϩ Na]^ϩ).

(؊)-(1S,2R,4S,6R)-6-[4-(2-Hydroxyphenyloxy)butyloxy]cyclo-

hexane-1,2,4-triol 6i

2

4.57 (1 H, B of second AB system, J_H-H12.0, 1 of OCH₂Ph),

5.08 (2 H, s, CO-OCH₂Ph), 5.46 (1 H, br, NH) and 7.24–

7.41 (15 H, m, Ar-H); δ_C(75.5 MHz; C²HCl₃) 32.71 and 34.46

(3-C and 5-C), 40.56 (OCH₂CH₂NH), 65.90 (OCH₂CH₂NH),

67.82 (CO-OCH₂Ph), 69.88 (OCH₂Ph), 70.65 (4-C), 71.03

(OCH₂Ph), 74.31 (6-C), 75.54 (2-C), 76.88 (1-C), 126.87,

127.00, 127.09, 127.28, 127.69 and 127.73 (Ar-CH), 135.97,

137.11 and 137.48 (Ar-C quaternary) and 156.16 (CO);

m/z (ES^ϩTOF) 528.2 (100%, [M ϩ Na]^ϩ) and 327.3 (15,

[M Ϫ C₁₀H₁₂NO₂]^ϩ).

To a stirred solution of alcohol 9i (372 mg, 0.755 mmol) in

MeOH (20 cm³) was added 5% palladium on activated charcoal

(318 mg) and a drop of acetic acid. The mixture was stirred

vigorously under an atmosphere of hydrogen gas for 16 h and

then ﬁltered through a pad of Celite. The pad was further

washed with MeOH (10 cm³) and water (20 cm³), and the

pooled ﬁltrate concentrated under reduced pressure. The resi-

due was redissolved in a small amount of water (∼5 cm³) and

lyophilised to give the 6-[4-(2-hydroxyphenyloxy)butyloxy] triol

O r g . B i o m o l . C h e m . , 2 0 0 4 , 2, 6 7 1 – 6 8 8

682

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6i as a white powder (222 mg, 84%); mp 92–97 ЊC (decomp.);

(HRMS: found: [M ϩ Na]^ϩ335.1468, C₁₆H₂₄O₆Na requires

335.1471); [α]_DϪ48.5 (c 0.6 in MeOH); ν_max(KBr disc)/cm^Ϫ1

3412 br, 2937, 2867, 1614, 1599 and 1498; δ_H(300 MHz; ²H₂O)

1.12–1.20 and 1.40–1.55 (2-H, m, 3-H and 5-H), 1.65–1.85 (4 H,

m, OCH₂(CH₂)₂CH₂O), 2.00–2.12 (1 H, m, 5-H), 2.30–2.40

(1 H, m, 3-H), 3.40–4.12 (8-H, m, OCH₂(CH₂)₂CH₂O, 1-H,

2-H, 4-H and 6-H), 6.80–7.05 (4-H, m, Ar-H); δ_C(75 MHz;

²H₂O) 23.19, 23.80, 35.66 and 36.39 (3-C, 5-C and OCH₂CH₂-

CH₂CH₂O), 61.96, 66.87, 72.14 and 74.08 (1-C, 2-C, 4-C and 6-

C), 66.80 and 67.29 (OCH₂CH₂CH₂CH₂O), 111.91, 113.64,

118.84 and 119.65 (Ar-CH) and 143.21 and 144.48 (Ar-C

quaternary); m/z (ES^ϩTOF) 357 (9%, [M ϩ 2Na Ϫ H]^ϩ) and

335 (100, M ϩ Na]^ϩ).

hydroxide solution until the pH was 14. The solution was then

extracted with ethyl acetate (3 × 50 cm³) and the pooled organic

fractions were washed with brine (50 cm³), dried (MgSO₄), ﬁl-

tered and then concentrated under reduced pressure to give a

light pale yellow oil which was puriﬁed by ﬂash silica column

chromatography (ethyl acetate–MeOH; 9 : 1, with addition

of 0.5 cm³of TEA per 10 cm³of solvent mixture) to give the

6-hexylamino alcohol 19e as a colourless oil (310 mg, 90%);

(Found: H, 9.1; C, 75.95; N, 3.4. C₂₆H₃₇NO₃requires H, 9.05;

C, 75.85; N, 3.4%); (HRMS: found: [M ϩ H]^ϩ, 412.2855.

C₂₆H₃₈NO₃requires 412.2852); [α]_DϪ26.1 (c 0.13 in CHCl₃);

ν_max(CH₂Cl₂)/cm^Ϫ13380 br, 3091, 3060, 3020, 2957, 2828, 1493,

1359, 1096, 1027, 734 and 665; δ_H(300 MHz; C²HCl₃) 0.83 [3 H,

3

t, J_H-H6.8, CH₂(CH₂)₄CH₃], 1.09–1.73 [11 H, m, CH₂(CH₂)₄-

CH₃, 3-H, 5-H and NH], 2.35–2.50 (2 H, m, 3-H and 5-H),

2.60–2.75 [2 H, m, CH₂(CH₂)₄CH₃], 2.68–2.79 (3 H, m, 1 of

1-H, 6-H and OH), 3.72–3.78 (1 H, m, 4-H), 3.90–3.94 (1 H, m,

2-H), 4.46–3.69 (4 H, m, 2 × OCH₂Ph) and 7.19–7.32 (10 H, m,

Ar-H); δ_C(75.4 MHz; C²HCl₃) 14.26 [CH₂(CH₂)₄CH₃], 22.81,

26.71, 27.24 and 31.96 [CH₂(CH₂)₄CH₃], 34.09 and 35.71 (3-C

and 5-C), 42.12 [CH₂(CH₂)₄CH₃], 56.54 (6-C), 70.78 and 71.97

(2 × OCH₂Ph), 72.48 (4-C), 74.86 (2-C), 76.55 (1-C), 127.80

and 128.60 (Ar-CH) and 138.68 and 138.88 (Ar-C quaternary);

m/z (ES^ϩTOF) 412.3 (100%, [M ϩ H]^ϩ).

(؊)-(1S,2R,4S,6R)-6-(2-Aminoethyloxy)cyclohexane-1,2,4-triol

6l

To a stirred solution of alcohol 7j (172 mg, 0.340 mmol) in

ethanol (20 cm³) was added 10% palladium on activated char-

coal (180 mg) and hydrogen gas was bubbled through the mix-

ture for 5 min followed by vigorous stirring for 18 h under a

constant hydrogen pressure. The reaction mixture was then ﬁl-

tered through Celite and the resulting solution concentrated

under reduced pressure to give the 6-(2-aminoethyloxy) triol 6l

as a colourless oil (63 mg, 97%); (Found: H, 9.0; C, 50.3; N,

7.35. C₈H₁₇NO₄requires H, 8.95; C, 50.25; N, 7.3%); (HRMS:

found: [M ϩ H]^ϩ, 192.1227. C₈H₁₈NO₄requires 192.1236); [α]_D

Ϫ17.3 (c 0.10 in MeOH); ν_max(thin ﬁlm)/cm^Ϫ13382 br, 2931,

2859, 1457, 1343, 1215, 1074 and 1022; δ_H(300 MHz; C²H₃O²H)

1.19–1.35 (1 H, m, 5-H), 1.39–1.48 (1 H, m, 3-H), 2.04–2.13 (1

H, m, 3-H), 2.28–2.37 (1 H, m 5-H), 3.05–3.23 (2 H, m, OCH₂-

CH₂NH), 3.47 (1 H, dd, ³J_H-H9.1, 2.8, 1-H), 3.53–3.61 (1 H, m,

6-H), 3.64–3.71 (1 H, m, 4-H), 3.85–3.99 (2 H, m, OCH₂-

CH₂NH) and 4.01–4.07 (1 H, m, 2-H); δ_C(75.5 MHz; C²H₃O²H)

28.59 and 39.91 (3-C and 5-C), 40.30 (OCH₂CH₂NH), 64.43

(4-C), 65.37 (OCH₂CH₂NH), 69.42 (6-C), 75.41 (C-2) and

77.45 (1-C); m/z (ES^ϩTOF) 192.1 (100%, [M ϩ H]^ϩ).

(؊)-(1S,2R,4S,6R)-2,4-Bis(benzyloxy)-6-(2-phenylethyl)-

aminocyclohexanol 19f

To a solution of epoxide (ϩ)-4 (222 mg, 0.536 mmol) in a mix-

ture of 2-phenylethylamine and 1,2-dichloroethane (40 cm³,

1 : 1, v/v) was added Yb()(OTf )₃(111 mg, 0.179 mmol) and

the resulting mixture was heated under reﬂux for 20 h. The

reaction mixture was concentrated under reduced pressure and

then dissolved in 1 M HCl (20 cm³). The solution was washed

with diethyl ether (2 × 30 cm³) and the separated aqueous layer

was adjusted to pH 14 by dropwise addition of 4 M NaOH

solution. The solution was then extracted with ethyl acetate

(3 × 50 cm³) and the pooled organic fractions were washed with

brine (50 cm³), dried (MgSO₄), ﬁltered and then concentrated

under reduced pressure to give a light yellow oil. Traces of

2-phenylethylamine were removed by Kugelrohr distillation to

give the 6-(2-phenylethyl)amino alcohol 19f as a pale yellow oil

(231 mg, 75%); (Found: H, 7.75; C, 78.0; N, 3.25. C₂₈H₃₃NO₃

requires H, 7.7; C, 77.95; N, 3.25%); (HRMS: found: [M ϩ H]^ϩ,

432.2537. C₂₈H₃₄NO₃requires 432.2539); [α]_DϪ25.7 (c 0.12 in

CHCl₃); ν_max(CH₂Cl₂)/cm^Ϫ13303–3602 br, 3085, 3055, 2933,

2807, 1500, 1359, 1135, 1032, 749 and 699; δ_H(300 MHz;

C²HCl₃) 1.09–1.45 (3 H, m, NH and 3-H and 5-H), 2.31–2.49

(3 H, m, 1 of N-CH₂CH₂Ph, 3-H and 5-H), 2.74–2.87 (4 H, m,

1 of N-CH₂CH₂Ph, OH and 2 × N-CH₂CH₂Ph), 3.03–3.09

(1 H, m, 6-H), 3.36–3.41 (1 H, dd, ³J_H-H9.1 and 3.0, 1-H), 3.67–

3.78 (1 H, m, 4-H), 3.89–3.93 (1 H, m, 2-H), 4.43 (1 H, A of an

AB system, ³J_H-H11.5, 1 of OCH₂Ph), 4.52 (43 (1 H, B of an AB

(؊)-(1S,2R,4S,6R)-6-(3-Aminopropyloxy)cyclohexane-1,2,4-triol

6m

This compound was prepared in a manner identical with that

described for the 6-(2-aminoethyloxy) triol 6l using alcohol 7k

(173 mg, 0.333 mmol) to give 6-(3-aminopropyloxy) triol 6m as

a colourless oil (65 mg, 95%); (Found: H, 9.35; C, 52.7; N, 6.85.

C₉H₁₉NO₄requires H, 9.35; C, 52.65; N, 6.8%); (HRMS: found:

[M ϩ H]^ϩ, 206.1388. C₉H₂₀NO requires 206.1392); [α]_DϪ16.5

(c 0.11 in MeOH); ν_max(thin ﬁlm)/cm^Ϫ13396 br, 2944, 2854,

1454, 1343, 1216, 1072 and 1022; δ_H(300 MHz; C²H₃O²H) 1.13–

1.48 (2 H, m, 3-H and 5-H), 1.54–1.82 (2 H, m, OCH₂CH₂-

CH₂NH₂), 2.03–2.15 (1 H, m, 3-H), 2.26–2.35 (1 H, m 5-H),

2.65–3.04 (2 H, m, OCH₂CH₂CH₂NH₂), 3.26–3.73 (3 H, m,

OCH₂CH₂CH₂NH₂and 6-H), 3.78–3.84 (1 H, m, 1-H) and

3.89–4.04 (2 H, m, 2-H and 4-H); δ_C(75.5 MHz; C²H₃O²H)

31.12 (OCH₂CH₂CH₂NH₂), 39.04 and 40.04 (3-C and 5-C),

40.42 (OCH₂CH₂CH₂NH₂), 65.02 (4-C), 68.92 (OCH₂CH₂-

CH₂NH₂), 70.05 (6-C), 76.07 (C-2) and 77.80 (1-C); m/z (ES^ϩ

TOF) 206.1 (100%, [M ϩ H]^ϩ).

3

system, J_H-H11.5, 1 of OCH₂Ph), 4.55 (1 H, m, A of an AB

3

system, J_H-H12.2, 1 of OCH₂Ph), 4.64 (1 H, B of an AB sys-

tem, ³J_H-H12.2, 1 of OCH₂Ph), and 7.19–7.38 (15 H, m, Ar-H);

δ_C(75.4 MHz; C²HCl₃) 29.48 and 30.90 (3-C and 5-C), 48.11

(N-CH₂CH₂Ph), 56.17 (6-C), 60.52 (N-CH₂CH₂Ph), 70.53 and

71.75 (2 × OCH₂Ph), 72.15 (4-C), 74.54 (2-C), 76.25 (1-C),

126.63, 127.65 and 128.87 (Ar-CH) and 138.41, 138.59 and

139.87 (Ar-C quaternary); m/z (FAB) 432 (38%, [M ϩ H]^ϩ), 416

(44, [M Ϫ OH ϩ 2H]^ϩ), 295 (65, [M Ϫ OH Ϫ PhCH₂CH₂NH ϩ

2H]^ϩ), 181 (100, [2 × C₇H₇Ϫ H]^ϩ) and 105 (25, [PhCH₂CH₂]^ϩ).

(؊)-(1S,2R,4S,6R)-2,4-Bis(benzyloxy)-6-hexylaminocyclo-

hexanol 19e

To a solution of epoxide (ϩ)-4 (260 mg, 0.839 mmol) in tolu-

ene–THF mixture (3 : 1, 3 cm³) was added Yb()(OTf )₃

(25 mg, 0.040 mmol) followed by hexylamine (111 mm³,

0.84 mmol) and the resulting mixture was heated under reﬂux

for 18 h. The solvents were removed under reduced pressure

and the residue dissolved in a 1 M hydrochloric acid solution

(20 cm³) and extracted with diethyl ether (2 × 30 cm³). To the

resulting aqueous layer was added, dropwise, a 4 M sodium

(؊)-(1S,2R,4S,6R)-2,4-Bis(benzyloxy)-6-aminocyclohexanol

19n

To a stirred mixture of epoxide (ϩ)-4 (500 mg, 1.6 mmol) and

ammonia solution (50 cm³), containing a few drops of ethanol

to aid solvation, was added Yb()(OTf )₃(300 mg, 0.48 mmol).

The ﬂask was sealed and heated to 70 ЊC for 12 h. After cooling,

O r g . B i o m o l . C h e m . , 2 0 0 4 , 2, 6 7 1 – 6 8 8

683

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the ﬂask was opened and the reaction mixture was extracted

with ethyl acetate (2 × 50 cm³). The pooled organic extracts

were dried (MgSO₄), ﬁltered and concentrated under reduced

pressure to an oﬀ-white solid which was puriﬁed by ﬂash silica

column chromatography (petroleum ether–ethyl acetate; 10 : 1)

to give the 6-amino alcohol 19n as white crystals (459 mg, 87%);

mp 81–83 ЊC; (HRMS: found: [M ϩ H]^ϩ, 328.1906. C₂₀H₂₆NO₃

requires 328.1913); [α]_DϪ10.6 (c 0.67 in EtOAc); ν_max(KBr

disc)/cm^Ϫ13853, 3748, 3304 br, 2929, 2867, 2362, 1648, 1543,

1459, 1364, 1106 and 695; δ_H(300 MHz; C²HCl₃) 1.13–1.24 and

1.30–1.40 (2 × 1 H, 2 × m, 3-H and 5-H), 1.75–1.85 (3 H, br s,

OH and NH₂), 2.17–2.25 and 2.29–2.37 (2 × 1 H, 2 × m, 3-H

and 5-H), 2.81–2.90 (1 H, m, 6-H), 3.14 (1 H, dd, J_HH3 and 9,

1-H), 3.62–3.72 (1 H, m, 4-H), 3.82–3.85 (1 H, m, 2-H), 4.41

(2 H, Abq, J_HH12, OCH₂Ph), 4.52 (2 H, ABq, J_HH11.5, OCH₂-

Ph) and 7.19–7.31 (10 H, m, Ar-H); δ_C(75 MHz; C²HCl₃) 33.61

and 38.26 (3-C and 5-C), 49.82 (6-C), 70.36 and 71.32 (2 ×

OCH₂Ph), 71.85, 76.57 and 76.94 (1-C, 2-C and 4-C), 127.32,

127.36, 127.52, 128.32 and 128.38 (Ar-CH) and 138.28

and 138.58 (Ar-C quaternary); m/z (ES^ϩTOF) 350 (100%,

[M ϩ Na]^ϩ) and 328 (85%, [M ϩ H]^ϩ).

ν_max(CH₂Cl₂)/cm^Ϫ13450 br, 3093, 3061, 3028, 2952, 2802, 1506,

1454, 1361, 1136, 1029, 754 and 697; δ_H(300 MHz; C²HCl₃) 0.91

(3 H, m, J_H-H7.2, NCH₂CH₂CH₂CH₃), 1.14–1.28 (1 H, m,

3

5-H), 1.30–1.52 (6 H, m, NCH₂CH₂CH₂CH₃, 3-H and NH),

2.34–2.55 (4 H, m, 3-H, 5-H, OH and 1 of NCH₂CH₂CH₂CH₃),

2.72–2.74 (2 H, m, 1 of NCH₂CH₂CH₂CH₃and 6-H), 3.39

(1 H, dd, ³J_H-H9,6 and 3.1, 1-H), 3.70–3.77 (1 H, m, 4-H), 3.90–

3.95 (1 H, m, 2-H), 4.46 (1 H, A of an AB system, ³J_H-H11.8, 1

3

of OCH₂Ph), 4.53 (1 H, A of an AB system, J_H-H11.8, 1 of

3

OCH₂Ph), 4.54 (1 H, m, B of an AB system, J_H-H11.7, 1

3

of OCH₂Ph), 4.61 (1 H, B of an AB system, J_H-H11.7, 1 of

OCH₂Ph) and 7.25–7.33 (10 H, m, Ar-H); δ_C(75.4 MHz;

C²HCl₃) 13.85 (NCH₂CH₂CH₂CH₃), 20.30 and 31.54 (NCH₂-

CH₂CH₂CH₃), 33.71 and 34.54 (3-C and 5-C), 46.29 (NCH₂-

CH₂CH₂CH₃), 56.54 (6-C), 70.66 and 71.83 (2 × OCH₂Ph),

72.04 (4-C), 73.89 (2-C), 76.11 (1-C), 127.65, 127.78 and 128.46

(Ar-CH) and 138.62 and 138.89 (Ar-C quaternary); m/z

(ES^ϩTOF) 406.2 (5%, [M ϩ Na]^ϩ) and 384.2 (100, [M ϩ H]^ϩ).

(؊)-(1S,2R,4S,6R)-2,4-Bis(benzyloxy)-6-octylaminocyclo-

hexanol 19q

This compound was prepared and puriﬁed in a manner identi-

cal with that described for the 6-butylamino alcohol 19p using

(ϩ)-4 (257 mg, 0.831 mmol) and n-octylamine (138 mm³, 0.83

mmol) to give the 6-octylamino alcohol 19q as a colourless oil

(310 mg, 85%); (Found: H, 9.45; C, 76.6; N, 3.2. C₂₈H₄₁NO₃

requires H, 9.4; C, 76.5; N, 3.2%); (HRMS: found: [M ϩ H]^ϩ,

440.3170. C₂₈H₄₂NO₃requires 440.3165); [α]_DϪ31.7 (c 0.12 in

CHCl₃); ν_max(CH₂Cl₂)/cm^Ϫ13398 br, 3089, 3060, 3030, 2926,

2854, 1479, 1357, 1095 br, 1026, 738, 696 and 664; δ_H(300 MHz;

C²HCl₃) 0.84 (3 H, m, ³J_H-H6.8, n-octyl), 1.06–1.45 (15 H, m, 12

of n-octyl, 3-H, 5-H, and NH), 1.58 (1 H, s, OH), 2.31–2.46

[3 H, m, 1 of N-CH₂(CH₂)₆CH₃, 3-H and 5-H], 2.65–2.75 [2 H,

m, 1 of N-CH₂(CH₂)₆CH₃and 6-H], 3.31 (1 H, dd, ³J_H-H9,6 and

3.1, 1-H), 3.67–3.71 (1 H, m, 4-H), 3.85–3.89 (1 H, m, 2-H),

4.43 (1 H, A of ﬁrst AB system, ³J_H-H11.8, 1 of OCH₂Ph), 4.49

(1 H, B of ﬁrst AB system, ³J_H-H11.8, 1 of OCH₂Ph), 4.50 (1 H,

(؊)-(1S,2R,4S,6R)-2,4-Bis(benzyloxy)-6-ethylaminocyclo-

hexanol 19o

To a stirred mixture of epoxide (ϩ)-4 (520 mg, 1.68 mmol) and

ethylamine (70% solution in water, 70 cm³) was added Yb()-

(OTf )₃(200 mg, 0.33 mmol). The ﬂask was sealed and heated to

65 ЊC for 48 h. After cooling, the ﬂask was opened and the

excess ethylamine gas was allowed to escape by bubbling with

nitrogen gas for several hours. The aqueous layer was then

extracted with ethyl acetate (3 × 50 cm³). The pooled organic

extracts were dried (MgSO₄), ﬁltered and concentrated under

reduced pressure to give the ethylamino alcohol 19o as an oil

(620 mg, 95%); (HRMS: found: M^ϩ, 355.2129. C₂₂H₂₉NO₃

requires 355.2147); [α]_DϪ23.6 (c 0.19 in EtOAc); ν_max(thin ﬁlm)/

cm^Ϫ13302 br, 3088, 3063, 3030, 2903, 2867, 1737 and 1657;

δ_H(300 MHz; C²HCl₃) 1.05 (3 H, t, J_HH7, NHCH₂CH₃), 1.07–

1.38 (2 H, m, 3-H and 5-H), 1.97–2.53 (5 H, m, 6-H, 3-H, 5-H,

OH and NH), 2.68–2.77 (2 H, m, NHCH₂CH₃), 3.31 (1 H, dd,

J_HH3.5 and 10, 1-H), 3.67 (1 H, tt, J_HH4 and 11, 4-H), 3.84–

3.87 (1 H, m, 2-H), 4.44 (2 H, Abq, J_HH12, OCH₂Ph), 4.51

(2 H, ABq, J_HH11.5, OCH₂Ph) and 7.19–7.31 (10 H, m, Ar-H);

δ_C(75 MHz; C²HCl₃) 15.51 (NHCH₂CH₃), 33.86 and 35.33 (3-C

and 5-C), 41.12 (NHCH₂CH₃), 56.28 (6-C), 70.60 and 71.78

(2 × OCH₂Ph), 72.19, 74.53 and 76.44 (1-C, 2-C and 4-C),

127.64, 127.74, 127.94, 128.46 and 128.91 (Ar-CH) and 138.44

and 138.65 (Ar-C quaternary); m/z (EI^ϩ) 356 (3%, [M ϩ H]^ϩ),

264 (65, [M Ϫ C₇H₇]^ϩ), 248 (10, [M Ϫ C₇H₇O]^ϩ) and 91 (100,

C₇H₇^ϩ).

3

m, A of second AB system, J_H-H11.7, 1 of OCH₂Ph), 4.57

3

(1 H, B of second AB system, J_H-H11.7, 1 of OCH₂Ph) and

7.24–7.33 (10 H, m, Ar-H); δ_C(75.4 MHz; C²HCl₃) 14.16, 22.70,

27.42, 29.31, 29.55, 30.55 and 31.88 [N-CH₂(CH₂)₆CH₃], 33.90

and 35.55 (3-C and 5-C), 46.97 [N-CH₂(CH₂)₆CH₃], 56.37

(6-C), 70.60 and 71.78 (2 × OCH₂Ph), 72.29 (4-C), 74.70 (2-C),

76.38 (1-C), 127.65, 127.72 and 128.46 (Ar-CH) and 138.50

and 138.69 (Ar-C quaternary); m/z (ES^ϩTOF) 440.2 (100%,

[M ϩ H]^ϩ).

(؊)-(1S,2R,4S,6R)-2,4-Bis(benzyloxy)-6-(4-phenylbutyl)-

aminocyclohexanol 19r

This compound was prepared and puriﬁed in a manner identi-

cal with that described for the 6-butylamino alcohol 19p using

(ϩ)-4 (260 mg, 0.838 mmol) and 4-phenylbutylamine (130 mm³,

0.84 mmol) to give the 6-(4-phenylbutyl)amino alcohol 19r as a

colourless oil (369 mg, 96%); (Found: H, 8.15; C, 78.5; N, 3.05.

C₃₀H₃₇NO₃requires H, 8.1; C, 78.4; N, 3.05%); (HRMS: found:

[M ϩ H]^ϩ, 460.2855. C₃₀H₃₈NO₃requires 460.2852); [α]_DϪ36.1

(c 0.10 in CHCl₃); ν_max(CH₂Cl₂)/cm^Ϫ13399 br, 3085, 3062, 3027,

2933, 2855, 1497, 1361, 1092, 1026, 754 and 664; δ_H(300 MHz;

C²HCl₃) 1.08–1.28 (1 H, m, 5-H), 1.35–1.43 (1 H, m, 3-H),

1.46–1.70 [6 H, m, NHCH₂(CH₂)₂CH₂Ph and OH], 2.35–2.44

(2 H, m, 3-H and 5-H), 2.45–2.53 [1 H, m, 1 of NHCH₂-

(؊)-(1S,2R,4S,6R)-2,4-Bis(benzyloxy)-6-butyl-

aminocyclohexanol 19p

To a solution of epoxide (ϩ)-4 (300 mg, 0.968 mmol) in 1,2-

dichloroethane was added Yb()(OTf )₃(25 mg, 0.040 mmol)

followed by butylamine (3 cm³, 50 mmol) and the resulting mix-

ture was heated under reﬂux for 20 h. The reaction mixture was

concentrated under reduced pressure and then dissolved in a

1 M hydrochloric acid solution (20 cm³) and extracted with

diethyl ether (2 × 30 cm³). To the resulting aqueous layer was

added, dropwise, a 4 M sodium hydroxide solution until the pH

was 14. The solution was then extracted with ethyl acetate

(3 × 50 cm³) and the pooled organic fractions were washed with

brine (50 cm³), dried (MgSO₄), ﬁltered and then concentrated

under reduced pressure to give a light yellow oil. Traces of

butylamine were removed by Kugelrohr distillation to give the

6-butylamino alcohol 19p as a yellow oil (266 mg, 72%);

(Found: H, 8.7; C, 75.2; N, 3.65. C₂₄H₃₃NO₃requires H, 8.65;

C, 75.15; N, 3.65%); (HRMS: found: [M ϩ H]^ϩ, 384.2541.

C₂₄H₃₄NO₃requires 384.2539); [α]_DϪ34.7 (c 0.14 in CHCl₃);

3

(CH₂)₂CH₂Ph], 2.58–2.63 [2 H, t, J_H-H7.6, NHCH₂(CH₂)₂-

CH₂Ph], 2.68–2.79 [1 H, m, 1 of NHCH₂(CH₂)₂CH₂Ph and

6-H], 3.33 (1 H, dd, ³J_H-H9.7 and 3.1, 1-H), 3.68–3.75 (1 H, m,

4-H), 3.90–3.95 (1 H, m, 2-H), 4.46 (1 H, A of ﬁrst AB system,

²J_H-H11.8, 1 of OCH₂Ph), 4.52 (1 H, B of ﬁrst AB system, ²J_H-H

11.8, 1 of OCH₂Ph), 4.53 (1 H, A of second AB system,

²J_H-H11.7, 1 of OCH₂Ph), 4.62 (1 H, B of second AB system,

²J_H-H11.8, 1 of OCH₂Ph), 7.15–7.17 (3 H, m, Ar-H) and 7.24–

O r g . B i o m o l . C h e m . , 2 0 0 4 , 2, 6 7 1 – 6 8 8

684

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7.36 (12 H, m, Ar-H); δ_C(75.4 MHz; C²HCl₃) 29.23 and 30.03

[NCH₂(CH₂)₂CH₂Ph], 33.88 and 35.40 (3-C and 5-C), 35.85

[NCH₂(CH₂)₂CH₂Ph], 46.73 [NCH₂(CH₂)₂CH₂Ph], 56.40

(6-C), 70.63 and 71.80 (2 × OCH₂Ph), 72.25 (4-C), 74.56 (2-C),

76.36 (1-C), 125.77, 127.68, 127.76, 128.35 and 128.48 (Ar-CH)

and 138.49, 138.68 and 142.45 (Ar-C quaternary); m/z (ES^ϩ)

482.4 (5%, [M ϩ Na]^ϩ) and 460.4 (100, [M ϩ H]^ϩ).

19n (85 mg, 0.260 mmol). The 6-amino triol 6p was isolated as

oﬀ white sticky solid (23 mg, 60%); (HRMS: found: [M ϩ H]^ϩ,

148.0975. C₆H₁₄NO₃requires 148.0974); [α]_DϪ64.7 (c 0.38 in

CH₃OH); ν_max(KBr disc)/cm^Ϫ13373 br, 3049, 2942, 1622 and

2

1499; δ_H(300 MHz; H₂O) 1.38–1.55 (2 H, m, 3-H and 5-H),

2.09 (1 H, dq, J_HH4 and 7, 5-H), 2.24 (1 H, ddd, J_HH4, 7 and 12,

3-H), 3.34 (1 H, dt, J_HH4 and 10, 6-H), 3.59 (1 H, dd, J_HH3 and

10, 1-H), 3.99 (1 H, tt, J_HH4.5 and 11.5, 4-H) and 4.06 (1 H,

2

dd, J_HH3 and 6.5, 2-H); δ_C(75.4 MHz; H₂O) 35.93 and 38.05

(؊)-(1S,2R,4S,6R)-6-Hexylaminocyclohexane-1,2,4-triol 6e

(3-C and 5-C), 48.93 (6-C), 63.16, 68.33 and 71.40 (1-C, 2-C

and 4-C); m/z (EI^ϩ) 148 (40%, M^ϩ).

To a solution of 6-hexylamino alcohol 19e (168 mg, 0.409 mmol)

in chloroform (20 cm³) was added trimethylsilyl bromide

(270 mm³, 2.05 mmol), dropwise, under argon. The resulting

solution was stirred and heated at 50 ЊC under an argon atmos-

phere for 36 h. After cooling, the solution was concentrated

under reduced pressure and the residue was dissolved in 0.2 M

HCl. This solution was washed with ethyl acetate (2 × 30 cm³)

and then adjusted to pH 14 by dropwise addition of 4 M NaOH.

The solution was then extracted with ethyl acetate (3 × 50 cm³)

and the pooled organic extracts were washed with brine (70 cm³),

dried (MgSO₄), ﬁltered and concentrated under reduced pressure

to give the 6-hexylamino triol 6e as a colourless oil (68.2 mg,

72%); mp 90–91 ЊC; (Found: H, 10.9; C, 62.4; N, 6.05. C₁₂H₂₅NO₃

requires H, 10.9; C, 62.3; N, 6.05%); (HRMS: found: [M ϩ H]^ϩ,

232.1905. C₁₂H₂₆NO₃requires 232.1913); [α]_DϪ18.6 (c 0.16 in

CH₃OH); ν_max(CHCl₃)/cm^Ϫ13385 br, 2993, 2859, 1457, 1344,

1222, 1076 and 1017; δ_H(300 MHz; C²H₃O²H) 0.77 [3 H, m, ³J_H-H

6.4, NCH₂(CH₂)₄CH₃], 1.02–1.14 (1 H, m, 5-H), 1.18–1.34 [6 H,

m, 6 of NCH₂(CH₂)₄CH₃], 1.36–1.53 [3 H, m, 2 of NCH₂-

(CH₂)₄CH₃and 3-H], 2.01–2.12 (1 H, m, 5-H), 2.17–2.24 (1 H, m,

3-H), 2.32–2.43 [1 H, m, 1 of NCH₂(CH₂)₄CH₃], 2.53–2.66 [1 H,

m, 1 of NCH₂(CH₂)₄CH₃], 2.67–2.74 (2 H, m, 6-H), 3.36 (1 H, dd,

³J_H-H9.3 and 2.1, 1-H), 3.88–3.96 (1 H, m, 4-H) and 4.01–4.07

(1 H, m, and 2-H); δ_C(75.4 MHz; C²H₃O²H) 11.72 (NCH₂-

(CH₂)₄CH₃), 20.71, 25.01, 26.89 and 29.72 (NCH₂(CH₂)₄CH₃),

34.46 and 37.32 (3-C and 5-C), 44.10 (NCH₂(CH₂)₄CH₃), 52.95

(6-C), 62.53 (4-C), 68.08 (2-C) and 71.33 (1-C); m/z (ES^ϩTOF)

232.0 (100%, [M ϩ H]^ϩ).

(؊)-(1S,2R,4S,6R)-6-Ethylaminocyclohexane-1,2,4-triol 6o

This compound was prepared in a manner identical with that

described for the 6-hexylamino triol 6e using 6-ethylamino

alcohol 19o (100 mg, 0.282 mmol). The 6-ethylamino triol 6o

was isolated as an oﬀ white sticky solid (39 mg, 78%); (HRMS:

found: [M ϩ H]^ϩ, 176.1280. C₈H₁₈NO₃requires 176.1287); [α]_D

Ϫ36.3 (c 0.20 in CH₃OH); ν_max(KBr disc)/cm^Ϫ13302 br, 3055,

2952, 2980, 2814, 1720 and 1638; δ_H(300 MHz; ²H₂O) 1.23 (3 H,

t, J_HH7.5, NCH₂CH₃), 1.34–1.55 (2 H, m, 3-H and 5-H), 2.06–

2.12 (1 H, m, 5-H), 2.31–2.36 (1 H, m, 3-H), 3.10 (2 H, ddt, J_HH

7.5 20 and 41, NCH₂CH₃), 3.31 (1 H, m, 6-H), 3.65 (1 H, dd,

J_HH3 and 10.5, 1-H), 3.99 (1 H, tt, J_HH4.5 and 11.5, 4-H) and

2

4.06 (1 H, m, 2-H); δ_C(75.4 MHz; H₂O) 15.79 (NCH₂CH₃),

38.94 (NCH₂CH₃), 43.17 and 45.17 (3-C and 5-C), 59.86 (6-C),

68.63, 73.76 and 76.07 (1-C, 2-C and 4-C); m/z (ES TOF^ϩ) 198

(10%, [M ϩ Na]^ϩ) and 176 (100, [M ϩ H]^ϩ).

(؊)-(1S,2R,4S,6R)-6-Butylaminocyclohexane-1,2,4-triol 6p

This compound was prepared in a manner identical with that

described for the 6-hexylamino triol 6e using 6-butylamino

alcohol 19p (228 mg, 0.592 mmol). The 6-butylamino triol 6p

was isolated as a pale yellow solid (92 mg, 76%); mp 87–89 ЊC;

(Found: H, 10.45; C, 59.15; N, 6.9. C₁₀H₂₁NO₃requires H, 10.4;

C. 59.1; N, 6.9%); (HRMS: found: [M ϩ H]^ϩ, 204.1594.

C₁₀H₂₂NO₃requires 204.1600); [α]_DϪ21.4 (c 0.13 in CH₃OH);

ν_max(CHCl₃)/cm^Ϫ13382 br, 2931, 2859, 1457, 1343, 1215, 1074

(؊)-(1S,2R,4S,6R)-6-(2-Phenylethyl)aminocyclohexane-1,2,4-

triol 6f

3

and 1022; δ_H(300 MHz; C²H₃O²H) 0.96 (3 H, m, J_H-H6.4,

NCH₂CH₂CH₂CH₃), 1.33–1.62 (4 H, m, NCH₂CH₂CH₂CH₃,

3-H and 5-H), 1.67–1.86 (2 H, m, NCH₂CH₂CH₂CH₃), 2.09–

2.21 (1 H, m, 5-H), 2.31–2.43 (1 H, m, 3-H), 3.01–3.25 (2 H, m,

NCH₂CH₂CH₂CH₃), 3.32–3.47 (2 H, m, 6-H), 3.64–3.73 (1 H,

m, 1-H) and 4.00–4.15 (2 H, m, 4-H and 2-H); δ_C(75.4 MHz;

C²H₃O²H) 15.19 (NCH₂CH₂CH₂CH₃), 22.07 and 30.44 (NCH₂-

CH₂CH₂CH₃), 37.00 and 41.41 (3-C and 5-C), 46.93 (NCH₂-

CH₂CH₂CH₃), 57.96 (6-C), 66.00 (4-C), 71.20 (2-C) and 73.69

(1-C); m/z (ES^ϩTOF) 406.2 (5%, [M ϩ Na]^ϩ) and 384.2 (100%,

[M ϩ H]^ϩ).

To a solution of 6-(2-phenylethyl)amino alcohol 19f (164 mg,

0.380 mmol) in MeOH (20 cm³) was added 10% palladium on

activated charcoal (180 mg) and hydrogen gas was bubbled

through the mixture for 5 min followed by vigorous stirring for

10 h under a constant hydrogen pressure. The reaction mixture

was then ﬁltered through Celite and the resulting solution con-

centrated under reduced pressure to give a pale yellow oil. This

whole procedure was repeated twice for the reaction to go to

completion. The 6-(2-phenylethyl)amino triol 6f was isolated as

a colourless oil (62 mg, 65%); (Found: H, 8.5; C, 67.0; N, 5.6.

C₁₄H₂₁NO₃requires H, 8.4; C, 66.9; N, 5.55%); (HRMS: found:

[M ϩ Na]^ϩ, 274.1425. C₁₄H₂₁NO₃Na requires 274.1419); [α]_D

Ϫ54.9 (c 0.17 in CH₃OH); ν_max(thin ﬁlm)/cm^Ϫ13365 br, 3091,

3058, 3023, 2939, 2850, 1576, 1451, 1315, 1080, 1127, 756 and

704; δ_H(300 MHz; C²H₃O²H) 1.09–1.22 (2 H, dt, ³J_H-H11.5 and

7.2, 3-H), 1.39–1.48 (1 H, dt, ³J_H-H11.2 and 2.6, 5-H), 2.05–2.13

(1 H, m, 5-H), 2.17–2.24 (1 H, m, 3-H), 2.71–2.91 (4 H, m,

N-CH₂CH₂Ph), 2.95–3.02 (1 H, m, 6-H), 3.29–3.32 (1 H,

m, 1-H), 3.91–4.01 (1 H, m, 4-H and 2-H) and 7.15–7.30 (5 H,

m, Ar-H); δ_C(75.4 MHz; C²H₃O²H) 37.37 and 39.41 (3-C and

5-C), 41.59 (N-CH₂CH₂Ph), 49.64 (N-CH₂CH₂Ph), 56.79

(6-C), 66.13 (4-C), 71.22 (2-C), 75.92 (1-C), 128.13, 130.34 and

130.48 (Ar-CH) and 141.43 (Ar-C quaternary); m/z (ES^ϩTOF)

274.1 (15%, [M ϩ Na]^ϩ) and 252.1 (100, [M ϩ H]^ϩ).

(؊)-(1S,2R,4S,6R)-6-Octylaminocyclohexane-1,2,4-triol 6q

This compound was prepared in a manner identical with that

described for the 6-hexylamino triol 6e using the 6-octylamino

alcohol 19q (174 mg, 0.396 mmol) to give the 6-octylamino triol

6q as a thick pale yellow oil (55.5 mg, 54%); (Found: H, 11.3; C,

64.9; N, 5.45. C₁₄H₂₉NO₃requires H, 11.25; C, 64.85; N, 5.4%);

(HRMS: found: [M ϩ H]^ϩ, 260.2229. C₁₄H₃₀NO₃requires

260.2226); [α]_DϪ16.7 (c 0.23 in CHCl₃); ν_max(thin ﬁlm)/cm^Ϫ1

3393 br, 2930, 2867, 1454, 1343, 1214, 1076 and 1021; δ_H(300

3

MHz; C²HCl₃) 0.86 (3 H, m, J_H-H6.3, n-octyl), 1.16–1.36

(11 H, m, 10 of n-octyl and 5-H), 1.38–1.58 (3 H, m, 2 of

n-octyl and 3-H), 1.96–2.36 (3 H, m, 3-H, 5-H and NH), 2.41–

2.54 [1 H, m, 1 of N-CH₂(CH₂)₆CH₃], 2.67–2.89 [2 H, m, 1 of

N-CH₂(CH₂)₆CH₃and 6-H], 3.14–3.70 (4 H, m, 3 × OH and

4-H) and 3.94–4.22 (2 H, m, 1-H and 2-H); δ_H(75.4 MHz;

C²HCl₃) 13.94 [N-CH₂(CH₂)₆CH₃], 22.49 and 27.15 [2 of N-CH₂-

(CH₂)₆CH₃], 28.10 (5-C), 29.14 and 29.35 [2 of N-CH₂-

(CH₂)₆CH₃], 29.91 (3-C), 31.67 [1 of N-CH₂(CH₂)₆CH₃], 36.32

(؊)-(1S,2R,4S,6R)-6-Aminocyclohexane-1,2,4-triol 6n

This compound was prepared in a manner identical with that

described for the 6-hexylamino triol 6e using 6-amino alcohol

O r g . B i o m o l . C h e m . , 2 0 0 4 , 2, 6 7 1 – 6 8 8

685

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[N-CH₂(CH₂)₆CH₃, due to folding of octyl chain], 38.66 [1 of

N-CH₂(CH₂)₆CH₃], 46.43 and 53.45 [N-CH₂(CH₂)₆CH₃], 55.68,

55.91 and 59.82 (6-C), 65.23 and 65.56 (4-C), 67.50 and 67.71

(2-C) and 70.27 and 73.44 (1-C); m/z (ES^ϩTOF) 274.2 (40%,

[M Ϫ C₂H₄Ϫ OH ϩ 2Na]^ϩ) and 260.2 (100, [M ϩ H]^ϩ).

was dissolved in ethyl acetate (50 cm³) and washed with water

(50 cm³). The aqueous layer was then back extracted with ethyl

acetate (3 × 50 cm³). The combined organic fractions were

washed with brine (50 cm³), dried (MgSO₄), ﬁltered and con-

centrated under reduced pressure. The resulting oil was puriﬁed

by ﬂash silica column chromatography (petroleum ether–ethyl

acetate; gradient from 1 : 2 to 2 : 1) to give the tosylate 22 as a

white solid (5.97 g, 88%); mp 51–52 ЊC (lit.,³³51–52 ЊC);

(Found: H, 5.55; C, 58.5; N, 4.05. C₁₇H₁₉NO₅S requires H, 5.5;

C, 58.45; N, 4.0%); (HRMS: found: [M ϩ Na]^ϩ, 372.0893.

C₁₇H₁₉NO₅NaS requires 372.0882); ν_max(CH₂Cl₂)/cm^Ϫ13390 br,

3091, 3060, 3030, 2950, 2850, 1716, 1597, 1455, 1358, 1259,

1189, 1176, 1096, 1010, 915, 754, 698 and 663; δ_H(300 MHz;

(؊)-(1S,2R,4S,6R)-6-(4-Phenylbutyl)aminocyclohexane-1,2,4-

triol 6r

This compound was prepared in a manner identical with that

described for the 6-hexylamino triol 6e using the 6-(4-phenyl-

butyl)amino alcohol 19r (161 mg, 0.351 mmol) to give the

6-(4-phenylbutyl)amino triol 6r as a white amorphous solid

(86 mg, 88%), mp 91–92 ЊC; (Found: H, 9.05; C, 68.9; N, 5.0.

C₁₆H₂₅NO₃requires H, 9.0; C, 68.8; N, 5.0%); (HRMS: found:

[M ϩ H]^ϩ, 280.1924. C₁₆H₂₆NO₃requires 280.1913); [α]_DϪ50.4

(c 0.06 in CHCl₃); ν_max(CHCl₃)/cm^Ϫ13315 br, 3086, 3056, 3026,

2930, 2856, 1602, 1453, 1092, 1026, 747 and 699; δ_H(300 MHz;

C²HCl₃) 1.13–1.24 (1 H, m, 5-H), 1.41–1.54 [3 H, m, 2 of

N-CH₂(CH₂)₂CH₂Ph and 3-H], 1.96–2.18 (6 H, m, 3-H, 5-H,

NH and 3 × OH), 2.46–2.54 [1 H, m, 1 of N-CH₂(CH₂)₂-

CH₂Ph], 2.60 [2 H, t, ³J_H-H6.0, N-CH₂(CH₂)₂CH₂Ph], 2.76–2.84

[2 H, m, 1 of N-CH₂(CH₂)₂CH₂Ph and 6-H], 3.29–3.33 (1 H, m,

1-H), 4.01–4.10 (1 H, m, 4-H), 4.14–4.17 (1 H, m, 2-H), 7.15–

7.19 (3 H, m, Ar-H) and 7.25–7.30 (2 H, m, Ar-H); δ_C(75.4

MHz; C²HCl₃) 29.62 and 30.60 [N-CH₂(CH₂)₂CH₂Ph], 36.33

[N-CH₂(CH₂)₂CH₂Ph], 39.06 and 40.07 (3-C and 5-C), 47.03

[N-CH₂(CH₂)₂CH₂Ph], 56.51 (6-C), 66.52 (4-C), 68.08 (2-C),

74.43 (1-C), 125.35 and 128.91 (Ar-CH) and 142.84 (Ar-C

quaternary); m/z (ES^ϩTOF) 302.1 (10%, [M ϩ Na]^ϩ), 280.2

(100, [M ϩ H]^ϩ) and 262.2 (5, [M Ϫ OH]^ϩ).

3

C²HCl₃) 2.42 (3 H, br, tosyl), 3.45 (2 H, q, J_H-H5.4, NCH₂-

3

CH₂O), 4.08 (2 H, t, J_H-H5.0, NCH₂CH₂O), 5.05 (3 H, s,

OCH₂Ph and NH), 7.25–7.38 (7 H, m, Ar-H) and 7.78 (2 H, d,

³J_H-H8.2, Ar-H); δ_C(75.4 MHz; C²HCl₃) 22.58 (CH₃), 41.20

(NCH₂CH₂O), 67.89 (NCH₂CH₂O), 70.04 (OCH₂Ph), 128.87,

128.97, 129.15, 129.50 and 130.91 (Ar-CH), 135.50, 138.12 and

148.12 (Ar-C quaternary) and 156.17 (CO); m/z (ES^ϩTOF)

372.2 (100%, [M ϩ Na] ^ϩ).

N-(4-Phenylbutyl)-N-(benzyloxycarbonyl)-ethane-1,2-diamine

23

A sealed round bottom ﬂask containing a stirred solution of the

tosylate 22 (5.02 g, 14.4 mmol) in phenebutylamine (23.2 cm³,

144 mmol) was heated at 50 ЊC for 12 h. Chloroform (150 cm³)

and K₂CO₃(2.20 g, 15.9 mmol) were then added and the result-

ing solution was stirred for a further 3 h. The solution was

diluted with chloroform (30 cm³) and washed with water

(150 cm³), saturated solution of NH₄Cl (120 cm³), water

(120 cm³) and brine (120 cm³). The resulting organic layer was

dried (MgSO₄), ﬁltered and then concentrated under reduced

pressure to give a light yellow which was puriﬁed by distillation

in vacuo at 80 ЊC using a Kugelrohr apparatus to give the

diamine 23 as a colourless oil (2.11 g, 45%); (Found: H, 8.1; C,

73.65; N, 8.6. C₂₀H₂₆N₂O₂requires H, 8.05; C, 73.6; N, 8.6%);

(HRMS: found: [M ϩ H]^ϩ, 327.2054. C₂₀H₂₇N₂O₂requires

327.2073); ν_max(CH₂Cl₂)/cm^Ϫ13326 br, 2091, 3062, 3033, 2961,

2849, 1706, 1537, 1454, 1258, 1181, 1122, 1030, 1006, 754 and

697; δ_H(300 MHz; C²HCl₃) 1.42–1.71 [4 H, m, CH₂(CH₂)₂-

CH₂Ph], 2.48–2.69 [6 H, m, NH, CONHCH₂CH₂and CH₂-

(N-Benzyloxycarbonyl)-2-aminoethanol 21

To a solution of ethanolamine (6 cm³, 100 mmol) in 1,4-

dioxane–water (1 : 1, 200 cm³) was added DMAP (50 mg,

4 mmol), benzyl chloroformate (16 cm³, 110 mmol) and TEA

(13.9 cm³, 110 mmol) and the reaction mixture stirred for 12 h.

The solvents were removed under reduced pressure to give a

thick solution which was diluted with DCM (150 cm³) and

water (50 cm³). After separation, the aqueous layer was back-

extracted with DCM (3 × 50 cm³). The pooled organic layers

were then successively washed with 0.1 M solution of HCl

(100 cm³), water (100 cm³), and brine (100 cm³), dried (MgSO₄)

and ﬁltered. The solvent was removed under reduced pressure

and the resulting white crystalline solid was recrystallised from

petroleum ether–ethyl acetate (3 : 1) to give N-Cbz 2-amino-

ethanol 21 as white crystals (15.6 g, 80%); mp 61–62 ЊC (lit.,³¹

61–62 ЊC); (Found: H, 6.8; C, 61.8; N, 7.2. Calc. for C₁₀H₁₃NO₃:

H, 6.7; C, 61.55; N, 7.15%) (HRMS: found: [M ϩ Na]^ϩ,

218.0802. Calc. for C₁₀H₁₃NO₃Na: 218.0793); ν_max(CH₂Cl₂)/

cm^Ϫ13370 br, 3250, 3140, 3030, 2950, 2850, 1692, 1546, 1453,

1276, 1150, 747 and 696; δ_H(300 MHz; C²HCl₃) 2.85 (1 H, br

3

(CH₂)₂CH₂Ph], 3.22 (2 H, t, J_H-H6.2, CONHCH₂CH₂), 5.08

(2 H, s, OCH₂Ph) and 7.12–7.38 (10 H, m, Ar-H); δ_C(75.4 MHz;

C²HCl₃) 28.18 and 28.56 [CH₂(CH₂)₂CH₂Ph], 34.80 [CH₂-

(CH₂)₂CH₂Ph], 39.26 [CH₂(CH₂)₂CH₂Ph], 48.01 (CONHCH₂-

CH₂), 48.51 (CONHCH₂CH₂), 65.47 (OCH₂Ph), 124.46,

127.10, 127.18, 127.51, 128.69 and 128.73 (Ar-CH), 136.13 and

141.44 (Ar-C quaternary) and 156.91 (CO); m/z (ES^ϩTOF)

327.2 (100%, [M ϩ H]^ϩ) and 300.2 (40, [M Ϫ C₂H₄ϩ 2H]^ϩ).

(؊)-(1S,2R,4S,6R)-2,4-Bis(benzyloxy)-6-{N-(4-phenylbutyl)-N-

[(NЈ-benzyloxycarbon-yl)-2-aminoethyl]amino}cyclohexanol 24

3

OH), 3.30 (2 H, q, J_H-H5.2, NCH₂CH₂OH), 3.65 (2 H, m,

NCH₂CH₂OH), 5.07 (2 H, s, OCH₂Ph), 5.39 (1 H, br, NH) and

7.25–7.32 (5 H, m, Ar-H); δ_C(75.4 MHz; C²HCl₃) 43.74

(NCH₂CH₂OH), 62.37 (NCH₂CH₂OH), 67.16 (OCH₂Ph),

128.35, 128.43 and 128.79 (Ar-CH), 136.64 (Ar-C quaternary)

and 157.41 (CO); m/z (FAB^ϩ) 218 (50%, [M ϩ Na]^ϩ), 199 (15,

[M Ϫ H₂O Ϫ H ϩ Na]^ϩ), 176 (100, M Ϫ H₂O Ϫ H]_ϩ), 105 (30,

[M Ϫ C₇H₇ϩ H]^ϩ) and 92 (25, C₇H₈O^ϩ)

This compound was prepared in a manner identical with that

described for amino alcohol 19e using diamine 23 (1.28 g,

3.93 mmol). The resulting oil was puriﬁed by ﬂash silica column

chromatography (petroleum ether–ethyl acetate; 6 : 1) to give

the alcohol 24 as a colourless oil (290 mg, 58%); (Found: H, 7.7,

C; 73.6, N; 4.4. C₄₀H₄₈N₂O₅requires H, 7.6; C, 75.45; N, 4.4%);

(HRMS: found: [M ϩ H]^ϩ, 637.3645. C₄₀H₄₉N₂O₅requires

637.3641); [α]_DϪ26.8 (c 0.12 in CHCl₃); ν_max(CHCl₃)/cm^Ϫ13395

br, 3089, 3052, 3024, 2954, 2851, 1724, 1526, 1454, 1242, 1094,

742 and 697; δ_H(300 MHz; C²HCl₃) 1.15–1.75 [6 H, m, 4 of

CH₂(CH₂)₂CH₂Ph, 3-H and 5-H], 2.11–2.20 (1 H, m, 5-H),

2.26–2.34 (1 H, m, 3-H), 2.44–2.74 [7 H, m, CH₂(CH₂)₂CH₂Ph,

NCH₂CH₂NH and OH], 2.88–2.95 (1 H, m, 6-H), 3.17–3.23

(2 H, m, NCH₂CH₂NH), 3.36–3.40 (1 H, m, 1-H), 3.70–3.76

(1 H, m, 4-H), 3.93–4.01 (1 H, m, 2-H), 4.45 (1 H, A of ﬁrst

(N-Benzyloxycarbonyl)-2-aminoethyl p-toluenesulfonate 22

A solution of alcohol 21 (3.79 g, 19.4 mmol) and DMAP

(50 mg, 4 mmol) in dry DCM (50 cm³) was cooled to 0 ЊC in

an ice–water bath and then tosyl chloride (4.08 g, 21.4 mmol)

was added, dropwise, under argon followed by TEA (2.98 cm³,

21.4 mmol). The reaction mixture was gradually brought to

room temperature and stirred for 5 h. The solvents were

removed under reduced pressure to give a white residue which

2

AB system, J_H-H11.9, OCH₂Ph), 4.50 (1 H, B of AB system,

O r g . B i o m o l . C h e m . , 2 0 0 4 , 2, 6 7 1 – 6 8 8

686

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²J_H-H11.9, OCH₂Ph), 4.60 (1 H, A of second AB system,

²J_H-H12.1, OCH₂Ph), 4.73 (1 H, B of second AB system,

²J_H-H12.1, OCH₂Ph), 5.09 (2 H, s, CO-OCH₂Ph), 5.28 (1 H,

br, NH) and 7.13–7.37 (20 H, m, Ar-H); δ_C(75.4 MHz;

C²HCl₃) 29.58, 29.91, 30.32 [CH₂(CH₂)₂CH₂Ph], 35.74

and 36.61 (3-C and 5-C), 40.91 [CH₂(CH₂)₂CH₂Ph], 50.15

(NCH₂CH₂NH), 51.02 (NCH₂CH₂NH), 58.87 (6-C), 67.43

(CO-OCH₂Ph), 71.51 (OCH₂Ph), 72.73 (4-C), 73.48 (OCH₂-

Ph), 73.82 (2-C), 76.66 (1-C), 126.58, 128.33, 128.44, 128.89 and

129.18 (Ar-CH), 136.17, 138.42, 138.89 and 142.31 (Ar-C

quaternary) and 156.55 (CO-OCH₂Ph); m/z (ES^ϩTOF) 659.4

(100%, [M ϩ Na]^ϩ), 637.4 (30, [M ϩ H]^ϩ).

at 25 ЊC and the reaction was quenched by the addition of

colorimetric reagent (800 mm³) at the required time followed 1

minute later by 34% trisodium citrate solution (100 mm³).

Absorbance at 660 nm was measured in 10 mm pathlength

cuvette. Phosphate concentrations were determined by com-

parison of absorbance value to a preconstructed standard curve

prepared using known phosphate concentrations.

Radiolabelled assay. Assays (50 mm³), which were initiated by

addition of enzyme (5 mm³) contained 50 mM KCl, 100 mM

Tris.HCl, 3 mM MgCl₂, substrate {( )-[2-³H]-Ins-1-P, 500 dpm

nmol^Ϫ1} at varying concentrations in the assay buﬀer, inhibitor

at varying concentrations in the assay buﬀer and K₂HPO₄(at 0,

1, 2 or 3 mM in the assay buﬀer) were incubated at 25 ЊC and

the reaction was quenched after 20 min by addition of 1 M

NaOH (5 mm³). The solution was then diluted with water

(100 mm³) and applied to a pre-equilibrated DOWEX-1X2-400

anion exchange column (OH^Ϫform, 200 mg). The column was

eluted by micro-centrifugation and the eluates (plus washings)

were emulsiﬁed with scintillation ﬂuid. The radioactivity in

each sample was determined by liquid scintillation counting

using a Packard Tri-Carb 2500 TR scintillation counter.

(؊)-(1S,2R,4S,6R)-6-[N-(4-phenylbutyl)-N-(2-aminoethyl)-

amino]cyclohexane-1,2,4-triol 25 [hydrochloride salt]

This compound was prepared in a manner identical with

that described for the 6-hexylamino triol 6e using alcohol 24

(285 mg, 0.45 mmol) to give a crude oil which dissolved in

concentrated HCl (10 cm³) and stirred for 1 h. Removal of the

solvent under reduced pressure gave a yellow oil which was

dissolved in MeOH (1 cm³). The solution was added dropwise

to diethyl ether (250 cm³), under stirring, and the resulting

white solid was ﬁltered, washed with diethyl ether (100 cm³) and

dried under high vacuum to give the diamine triol hydro-

chloride 25 as a white solid (55 mg, 38%); mp 93–94 ЊC (Found:

H 8.75; C, 60.3; N, 7.85. C₁₈H₃₁ClN₂O₃requires H, 8.7; C,

60.25; N, 7.8%); (HRMS: found: [M Ϫ Cl]^ϩ, 323.2324.

C₁₈H₃₁N₂O₃requires 323.2335); [α]_DϪ25.9 (c 0.10 in MeOH);

ν_max(KBr disc)/cm^Ϫ13396 br, 3089, 3056, 2967, 2854, 1506,

1454, 1246, 1094, 742 and 698; δ_H(300 MHz; C²H₃O²H) 1.48–

1.91 [6 H, m, 4 of CH₂(CH₂)₂CH₂Ph, 3-H and 5-H], 2.11–2.19

(1 H, m, 5-H), 2.29–2.34 (1 H, m, 3-H), 2.68–2.73 [2 H, m,

CH₂(CH₂) ₂CH₂Ph], 2.98–3.14 [1 H, m, 1 of CH₂(CH₂)₂-

CH₂Ph], 3.32–4.18 [9 H, m, 1 of CH₂(CH₂)₂CH₂Ph, NCH₂-

CH₂NH, 6-H, 1-H, 4-H and 2-H], and 7.19–7.37 (5 H, m,

Ar-H); δ_C(75.4 MHz; C²H₃O²H) 25.38 and 29.41 [CH₂(CH₂)₂-

CH₂Ph], 32.89 and 35.80 (3-C and 5-C), 36.09 [CH₂(CH₂)₂-

CH₂Ph], 40.27 (NCH₂CH₂NH₃), 49.00 [CH₂(CH₂)₂CH₂Ph],

53.72 (NCH₂CH₂NH), 61.80 (6-C), 64.98 (4-C), 70.12 (2-C),

70.38 (1-C), 127.04 and 129.48 (Ar-CH) and 141.18 (Ar-C

quaternary); m/z (ES^ϩTOF) 323.3 (100%, [M Ϫ Cl]^ϩ), 306.3

(70, [M Ϫ Cl Ϫ NH₃]^ϩ) and 280.3 (30, [M Ϫ Cl Ϫ C₂H₇N ϩ

2H]^ϩ).

Acknowledgements

The authors would like to thank the BBSRC and EPSRC for

grants 6/B13490 and GR/K89788, the University of Birming-

ham for some ﬁnancial support, and Peter R. Ashton and

Neil Spencer for help with mass spectrometry and NMR

spectroscopy.

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Article Doi

Removal of the phosphate group in mechanism-based inhibitors of inositol monophosphatase leads to unusual inhibitory activity.

DOI: 10.1039/b312808c

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Article abstract of DOI:10.1039/b312808c

Full text of DOI:10.1039/b312808c

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