Bis(spirolactam) 1,3-double-armed calix[4]arene compounds and their application as extractants for the determination of heavy metal ions

Article abstract of DOI:10.1007/s10847-015-0574-z

A number of double-armed calix[4]arene compounds, with proline-derived spirolactam ligating groups on the lower rim, have been synthesised and investigated as extractants of toxic heavy metal ions from aqueous solution. Pedersen's extraction technique was

Full text of DOI:10.1007/s10847-015-0574-z

J Incl Phenom Macrocycl Chem
DOI 10.1007/s10847-015-0574-z
ORIGINAL ARTICLE
Bis(spirolactam) 1,3-double-armed calix[4]arene compounds
and their application as extractants for the determination of heavy
metal ions
James Ward1 Li Li2 Fiona Regan2 Mary Deasy1 Fintan Kelleher1,3
•
•
•
•
Received: 10 August 2015 / Accepted: 13 October 2015
Ó Springer Science+Business Media Dordrecht 2015
Abstract
A number of double-armed calix[4]arene
compounds, with proline-derived spirolactam ligating
groups on the lower rim, have been synthesised and
investigated as extractants of toxic heavy metal ions from
aqueous solution. Pedersen’s extraction technique was
employed to determine the capability of these new 1,3-
distal derived calix[4]arene spiroalactams to extract
O
OR'
O
OR'
selected heavy metal ions (e.g. Pb2?, Cd2?, Zn2?, Cu2?
,
O
O
N
N
O
O
Ni2?, Co2?) from an aqueous phase into an organic phase.
The percentage extraction was calculated using UV–Vis
spectroscopy. All of the compounds synthesised demon-
strated good selectivity for the heavy metals selected over
the alkali metal and alkaline earth metal ions, in particular
for Pb2?. The level of selectivity observed was dependent
on the N-substituent of the spirolactam moiety attached to
alternate phenolic oxygens of the narrow calix[4]arene rim.
Interestingly, higher sensitivity and lower selectivity was
found when the two remaining –OH groups were replaced
by –OCH3 groups.
N
N
R
R
R = Benzyl, allyl, -CH2COOCH3
R' = H, CH3
Keywords Spirocyclic double-armed calix[4]arenes Á
Spirolactam Á Heavy metal ion determination Á Pb2?
determination Á Optical chemical sensors Á Ionophore
Graphical Abstract
Introduction
Selective monitoring of hazardous heavy metal and tran-
sition metal ions and their determination at low concen-
trations is a longstanding goal of many research and
commercial groups, not least because of the need for high
quality drinking water and the necessity for ultrapure water
for injectable grade drug formulations hitting the market
[1]. The presence of heavy metal ions such as Cd2?, Pb2?
and Hg2? in water is not desirable, since they are not
degradable and produce toxic effects at very low concen-
trations. Hence, the development of systems that can
scavenge these metals at trace levels is vital. This can be
achieved using macrocyclic scaffolds with chemical
& Fintan Kelleher
fintan.kelleher@ittdublin.ie
1
Molecular Design & Synthesis Group, Centre of Applied
Science for Health, Institute of Technology Tallaght,
Tallaght, Dublin 24, Ireland
2
Marine and Environmental Sensing Technology Hub, School
of Chemical Sciences, Dublin City University, Glasnevin,
Dublin 9, Ireland
3
Department of Science, Institute of Technology Tallaght,
Tallaght, Dublin 24, Ireland
123

J Incl Phenom Macrocycl Chem
functionalities capable of trapping ionic species and
removing them from aqueous media.
rigid backbone comprising aromatic rings linked by tight
methylene bridges, and therefore have the required archi-
tecture for entrapping small molecules and ions, and this
has been well documented in the literature.
Calix[4]arenes are an important class of macrocyclic
compounds which have been used extensively in
supramolecular chemistry for molecular recognition [2].
They were first prepared by Baeyer in 1872, as by-products
in the synthesis of baeklite [3]. Their synthesis was further
improved by Zinke and Ziegler in 1944 [4], and then early
conformational studies were published in 1955 by Corn-
forth [5]. Since Gutsche first published the synthesis of
calix[4]arene-based macrocycles [6], the investigation of
the complexation properties of these compounds has been
of intense interest for a wide variety of ionic species and
small organic guest molecules, through the formation of
host–guest or supramolecular complexes. This area is
subject to regular in-depth reviews [2].
Another way to constrain the flexibility of small mole-
cules is by incorporating a spirocyclic fusion into the
molecule [8]. This gives a defined conformation with
directionality to any ligating groups within the spirocyclic
moiety. Spirocycles have been used to constrain the con-
formation of flexible molecules, in particular peptides,
leading to an improvement in their chemical and biological
properties [9]. They are composed of two or more ring
hydrocarbons fused together by a quaternary carbon, and
often have N or O atoms as part of the ring. These het-
eroatoms are commonly included compounds designed to
act as synthetic hosts or ionophores, and are often linked to
carbonyl groups to provide more binding sites within a
host. This platform provides a highly-organised and rigid
unit for the incorporation of amine and amide groups that
have potential to bind specifically to small molecules or
guests, a key feature of any chemical sensor. This is the
first time macrocyclic cage-like compounds incorporating
spirocyclic functionalities have been prepared for potential
sensor application; where the calixarene provides a rela-
tively inflexible cavity large enough to accommodate metal
ions, and the spirocyclic antenna provide additional
directionality to provide improved interaction with the
guest metal ions.
While the study of calixarenes with metal ions has
attracted much attention, they have been less studied for
binding to transition metal ions. Much of the current
research into calixarenes focuses on the smallest and most
rigid of the family, namely the calix[4]arenes, since the
larger cavity-containing analogues are more flexible and
therefore not always suited to strong binding with guest
species, and are also generally less selective. Many
calix[4]arene compounds have been synthesised over the
last 40 years, with a variety of ligating groups to fine-tune
binding to an array of ionic and neutral guests in solution
and in the solid state [2]. Some of the functionalities
attached to the calixarene scaffold are large and while they
can facilitate binding can crowd the cavity and decrease the
binding strength. In order to overcome this potential issue
chemists have devised calix[4]arenes where only two of the
phenolic OH groups possess ligating groups. These are
referred to as 1,3-distal derivatised calix[4]arenes. In our
previous work, we have synthesised such 1,3-distal
derivatives which have been used in nanostructures capable
of selectively sensing Li? and Fe3? in aqueous solution [7].
Among the macrocyclic complexing agents for metal ions
in general, distal 1,3-substituted calix[4]arene-based iono-
phores currently occupy an important position [2b].
Pederson’s solvent extraction method, by using picrate
salts of metal ions, is a useful method for studying the
binding ability of macrocyclic molecules with metal ions
[10]. Herein, we report the synthesis of a series of
bis(spirolactam) 1,3-double-armed calix[4]arene receptors,
on the lower rim and their binding properties with heavy-
metal and transition-metal ions.
Results and discussion
Synthesis
Most early chemical sensors were based on small
molecules, with suitable ligating groups capable of binding
to the guest species, being detected and measured. How-
ever, these host systems are flexible, and they are not
selective for one particular guest species in particular. By
constraining the ligating groups in a ring system or within a
larger macromolecular structure then conformational free-
dom should be reduced and the ligating groups should then
bind more selectively to the guest molecule. If the
macromolecule has a set 3-D geometry and/or has a cavity
(like the calix[n]arenes) then this should pre-organise the
ligating groups within the host compound even more for
selective binding to the guest. The calixarene family has a
There are many literature reports of distal 1,3-bis(amido)-
linked calix[4]arenes with a variety of ligating groups for
binding to neutral, anionic and cationic guests [2]. In the
vast majority of these cases the ligating groups have a large
degree of conformational freedom. An underlying theme of
our research for many years has been on the introduction of
conformational constraint into flexible structures, in order
to improve their binding properties to, for example, guest
molecules. One method employed is the introduction of
spiro fused rings with defined conformational preferences
due to the constraints imposed by their structure. We have
reported previously on the synthesis, characterisation and
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J Incl Phenom Macrocycl Chem
binding properties of a number of proline-derived spiro-
lactams and diamines [11]. The spirolactams were syn-
thesised in good yields starting from L-proline (Fig. 1). It
was possible to easily vary the lactam ring N-substituent to
give a number of spirolactams which could be subse-
quently derivatised on the pyrrolidine nitrogen atom. It was
thought that conjugation of the spirolactam moiety in a 1,3-
distal manner to a calix[4]arene backbone, via a linker,
would give compounds with a varied cavity size, capable
of binding metal ions. Moreover, it might also be possible
to tune the cavity size by the choice of the lactam ring
N-substituent, in order to give selective binding to target
metal ions, such as heavy metal ions. The N-substituents
initially chosen contained an ester group with potential for
binding through the oxygen atoms, as well as two with
potential for alkene- or arene-cation binding (Fig. 1).
Literature syntheses of amide linked calix[4]arenes have
mainly used three different methods to create the amide
linker [2]. Firstly, aminolysis of an ester with an amine
gave good yields of the amides, but this method is
restricted to the use of primary amines. Secondly, coupling
of amines to a carboxylic acid moiety using a variety of
‘‘peptide coupling’’ methods gave the amides in good
yields, but again the vast majority of examples used pri-
mary amines or primary amino acids. The third method is
the direct reaction of an amine with an acid chloride.
McKervey reported this method for reactions involving
secondary amines [12]. Since the spirolactams being used
were secondary amines, and also highly sterically hindered,
it was felt that this method was likely to give the best
chance of success.
was found that in each case the NMR spectra (1H and 13C)
were very complicated because the ‘‘product’’ was a mix-
ture of diastereoisomers, since the spirolactams used were
racemic mixtures. All efforts to resolve the spirolactams
(1–3), using a number of resolution methods, to provide the
individual enantiomers were unsuccessful [14]. Therefore,
after further very careful column chromatography, small
amounts of single diastereoisomer compounds were iso-
lated, with much simplified NMR spectra. Thus far,
attempts to obtain crystals of any of these products suit-
able for X-ray analysis have been unsuccessful, so the
relative and absolute stereochemistries of the spiro centres
are currently not known.
The synthetic route to the dimethoxy calix[4]arene
1,3-distal substituted spirolactam compounds is also
shown in Scheme 1. The literature procedures of
Gutsche and McKervey were used to prepare the ter-
minal diester-dimethoxy 1,3-distal calix[4]arene using
NaH and MeI in THF [12, 13]. The ester groups were
subsequently hydrolysed using ethanolic NaOH, fol-
lowed by pH adjustment with 3 M HCl to give the
dicarboxylic acid-dimethoxy derivative 10. The product
was isolated in 98 % yield after purification by recrys-
tallisation from EtOH/H2O as a white crystalline solid.
Reaction with thionyl chloride to provide the diacid-
chloride-dimethoxy compound 11 was conducted using
the same conditions as described above and the yield
was quantitative. The dimethoxy-spirocyclic calixarenes
12, 13 and 14 were obtained in yields ranging from 45 to
66 % using the same reaction conditions outlined pre-
viously. When the Diamond Group published the syn-
thesis of compounds with methoxy groups in the
literature [15], they reported that the1H NMR spectrum
contained several broad signals too close together to
assess even using high field techniques, and used accu-
rate mass spectral data to determine that they had
obtained the correct compound. This was found to be
the case for the spirolactam-dimethoxy derivatives 12–14
also, so accurate mass spectral data (see Table 1), cou-
pled with IR data was used to show that reaction did
indeed provide compounds 12, 13 and 14. The broadness
of the NMR signals is most likely due to the loss of
intramolecular H-bonding at the phenolic oxygen posi-
tions, making the calixarene backbone more mobile in
solution with rapid conformational fluctuations occurring
The first set of target spirolactam-calixarenes were
synthesised as shown in Scheme 1. The literature proce-
dures of McKervey [12] and Gutsche [13] were used to
prepare the distal 1,3-terminal bis(carboxylic acid) com-
pound 4.
The 1,3-bis(acid chloride) substituted compound 5 was
prepared using a method analogous to that of McKervey
[12] where the acid was converted to the acid chloride by
heating the acid with thionyl chloride in toluene, at reflux
temperature. The less toxic solvent toluene was used in
place of benzene, which had been used in the published
method, without any reduction in yield. The bis(acid
chloride) 5 was then separately reacted with each spiro-
lactam (1–3) to give the desired compounds in 49–52 %
isolated yields, after column chromatography. However, it
as
a
result, something that has been previously
Fig. 1 Synthesis of proline-
derived spirolactam
trifluoroacetate salts 1–3 [11]
Ref. [11]
1 R = -CH2CO2Me
N
2
3
= -CH2CH=CH2
= -CH2Ph
TFA.
N
H
COOH
R
N
H
O
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J Incl Phenom Macrocycl Chem
i
4
OH
2
OR
O
O
R = H (4)
Me (10)
OH
ii
iii
2
2
OR
O
O
OR
O
O
R = H (5)
Me (11)
Cl
N
O
N
R'
R = H; R' = -CH2CO2CH3 (6)
-CH2CH=CH2 (7)
-CH2-Ph
(8)
R = Me; R' = -CH2CO2CH3 (12)
-CH2CH=CH2 (13)
-CH2-Ph
(14)
Reagents and Conditions: i) Refs. [12] and [13]; ii) Thionyl chloride, toluene, reflux temperature;
iii) 1,2 or 3, Et3N, DCM, rt.
Scheme 1 Reagents and conditions i Refs. [12] and [13]; ii Thionyl chloride, toluene, reflux temperature; iii 1,2 or 3, Et3N, DCM, rt
Metal picrate extraction studies
Table 1 HRMS results for the dimethoxy-spirolactam calixarene
compounds 12, 13 and 14
Since Diamond and McKervey developed a Na? ion
Compound
Calculated
Found
selective electrode using a calix[4]arene host compound
over 20 years ago, numerous calixarene derivatives con-
taining pendant functional groups (e.g. ketones, amides,
ethers, crown ethers and esters) have been incorporated in
an array of commercial sensor devices capable of sensing
alkali and alkaline earth metal guests for many applications
[17]. There are some calixarene-based sensors reported for
the soft heavy metal ions, like Ag?, Pb2?, and Hg2? which
display a strong affinity for soft coordination centres, i.e.
N, S, Se and P [18]. Calixarenes with azo and imine units
(12 1 Na)1
(13 1 H)1
(14 1 Na)1
1203.6606
1117.6946
1239.7125
1203.6609
1117.6993
1239.7126
Complexation with Na? from the glass vial in which the sample was
stored is a common feature for calixarenes [16]
documented for calixarenes [15]. A detailed NMR con-
formational study on these calix-spirolactams by VT
NMR will be undertaken and reported in the future.
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J Incl Phenom Macrocycl Chem
Table 2 Extraction (% E) of metal picrates with bis(spirolactam) calixarene hosts 6,7 and 8
Host Co2?
Ni2?
Cu2?
Zn2?
Cd2?
Pb2?
Li?
Na?
K?
Ca2?
Mg2?
6
7
8
27.9 (1.0) 36.6 (1.0) 24.8 (1.3) 23.5 (1.6) 29.0 (2.9) 63.5 (3.9) 19.7 (0.7) 10.0 (0.5) 19.6 (2.2) 22.4 (1.3) 22.1 (2.7)
0.5 (5.3)
0 (2.0)
4.0 (4.9)
1.0 (1.9)
3.9 (0.9) 56.7 (0.9)
5.0 (0.6) 12.9 (4.1) 15.8 (1.2) 11.2 (4.6)
3.4 (2.0)
20.5 (1.5)
8.0 (2.3) 74.9 (2.6) 26.1 (2.2) 19.2 (2.1) 75.3 (1.0) 25.2 (0.8) 24.6 (1.0) 18.7 (0.8) 29.4 (1.7) 27.3 (0.4)
n = 3, figures in brackets are relative standard deviation (RSD) values (0.4–5.3 % range)
very much in use today [10]. The spirolactam calix[4]arene
compounds were tested against transition heavy metal ion
picrates. Some of the smaller alkali and alkaline metal ion
picrates salts were used for comparison, since the
calix[4]arene cavity has always been selective for Na?, in
particular [20]. However, the selectivity is known to be
affected by the substituents present, particularly at the
narrower lower rim [21].
The metal picrate extraction experiments were per-
formed by using a 2.5 9 10-4 M host solution of
calix[4]arene compounds in dichloromethane and a picrate
solution in deionised water, according to the procedure
described by Pedersen [10]. The absorption of the extracted
picrate were measured from the aqueous layer at 355 nm
and compared with the blank experiment (with no
bis(spirolactam)-calix[4]arene host) of the appropriate
picrate. It was found that 1 h for shaking and then 2 h for
the separation of the two phases was sufficient for
achieving equilibrium. The results, expressed as a per-
centage extraction (%E) of cation are shown in Table 2 for
the spirolactam-calix[4]arenes 6, 7 and 8.
Fig. 2 % Extraction of the heavy metal divalent ion picrate salts
from aqueous solution into dichloromethane by spirolactam-calix[4]-
arene host compounds 6–8, with lower rim phenolic groups
have shown good binding towards metal ions such as Ca2?
and Ni2?, respectively [19]. In more recent years a number
of calixarene derivatives containing thioether, pyridyl,
benzothiazoyl and other heteroatomic arrangements have
been synthesised and tested as ionophores for heavy metals
[2c]. However, while they may bind very well to the guest
species, they are generally not always very selective
towards any one particular metal ion. This is a significant
drawback for commercial application in a sensor or in a
more traditional ion-selective electrode system.
In all cases it was found that the new host compounds
showed relatively weak extraction of the alkali and alkaline
earth metal ions. This would be expected, given the nature
of the pendant arms attached to the 1,3-distal phenolic
rings. Traditionally simple tetra and di-ester linkages were
more suited to extraction of those guest species [20, 21].
The trends for the extraction of the divalent heavy-metal
salts by the spirocyclic distal –OH calixarene host com-
pounds (6–8) are presented graphically in Fig. 2.
One of the most useful measures of a host compound’s
ability for use in an environmental or other type of
chemical/bio sensor application is its ability to selectively
interact with the sensing agent or guest species. When that
agent is an ionic guest it is frequently required to obtain
that measurement from an aqueous solution. However, the
trapping macrocycles are usually highly organic in nature
and hydrophobic. So, if the initial screening of host or
sensing compounds is carried out in a biphasic mixture it
can provide a strong indicator of how that host would
behave in a sensor. In addition, a quantitative measure of
the host–guest interaction that provides information on
selectivity or specificity of binding is vital to direct studies
on specific sensor application. The most effective method
that satisfies both of these key requirements is the picrate
extraction method devised by Pedersen in 1968 and still
The largest extraction capabilities for all three hosts
were displayed with Pb2? ion. The host with a benzyl
group attached at the 1,3-distal spirolactam calix[4]arene 8,
showed the highest % extraction values, of up to 75 %, for
both Pb2? and Cu2? ions. Host 7, with the pendant allyl
groups, demonstrated the highest overall selectivity for
Pb2? over all of the ions used in the study. The host
compound 6, with the pendant ester groups, demonstrated
the least selectivity but with good extraction levels for all
heavy metals studied.
When the two remaining –OH groups of 6–8 were
replaced by –OCH3 groups the extraction ability was
measured for a select number of metal ions. There was an
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J Incl Phenom Macrocycl Chem
with the pendant ester substituents on the spiroalactam unit,
at 86.5 % for Pb2?. The most selective of this group of
hosts was compound 14, with the pendant N-benzyl groups.
Like their –OH analogues, the –OMe compounds (12–14)
showed negligible extraction efficiency (\30 %) towards
the alkaline and alkali metal ions (data not shown).
There is a difference in sensitivity and selectivity for the
metal ions between the extractant compounds with phe-
nolic –OH groups (6–8) and those with –OCH3 groups (12–
14). The compilation of all the extraction results (Fig. 4)
points to preferential extraction of Pb2?, in all cases. The
recorded % extraction for Pb2? values are comparable to
heavily substituted lower rim calix[4]arenes published in
the literature [22] and other 1,3-distal substituted cal-
ixarene hosts [2]. There are reports of calix-crown com-
pounds with higher % extraction for Pb2? but they are not
as selective as the compounds reported in this study [23].
There may also be a role for the diastereoisomers of
compounds of this study, as different diastereoisomers
would be likely to have different effects on both the cavity
size and the directionality of the ligating groups of the
pendant arms. It is therefore highly probable that the
overall extraction capability and selectivity would be
influenced by which diastereoisomer was used. However,
this is speculative as the stereochemistry of the compounds
of this study are not currently known (vide supra) and pure
samples of the other diastereoisomers are not currently
available.
Table 3 Extraction (% E) of metal picrates with bis(spirolactam)
calixarene hosts 12,13 and 14
Host
Co2?
Ni2?
Cu2?
Zn2?
Cd2?
Pb2?
12
13
14
66.0
62.5
42.1
57.3
64.5
21.6
56.5
47.3
30.0
66.8
50.5
42.7
63.4
50.4
46.5
86.5
64.8
78.5
n = 3 (RSD values were \5 %, in all cases)
Fig. 3 % Extraction of the heavy metal divalent ion picrate salts
from aqueous solution into dichloromethane by spirolactam-calix[4]-
arene host compounds 12–14, with lower rim methoxy groups
overall increase in the % extraction, as shown in Table 3
for the dimethoxy spirolactam calix[4]arenes 12, 13 and
14, with the exception of Cu2? ion and host compound 14.
However, in all cases, 12–14 were found to be less
selective for one specific metal guest ion (Fig. 3). The
highest level of extraction was recorded for compound 12
Conclusions
In summary, we have developed a new class of double-
armed calix[4]arenes which have demonstrated efficient
binding for heavy metal ions over their alkali and alkaline
Fig. 4 Trend in % extraction by
metal ion for all host
compounds 6–8 and 12–14
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J Incl Phenom Macrocycl Chem
earth metal counterparts. Selectivity for Pb2? was achieved
using compound 7, which had an N-allyl substituent on the
lactam ring, and this selectivity is a key prerequisite for
application in sensor studies. The next step will be the
dispersion the spirocyclic calix[4]arenes into a membrane,
as a first step in the development of a commercial Pb2?
sensor. We are also currently synthesising calix[4]arenes
with different pendant spirocyclic arms in order to try to
find even more efficient and selective binders of Pb2? ions,
as well as giving an improved overall understanding of the
structure-extraction ability relationship for the spirocyclic
double-armed calix[4]arene compounds. The results from
all of these studies will be reported in due course.
software package. Solid samples were prepared as disper-
sions in KBr discs and liquid samples as an interspersed
film between polished sodium chloride plates.
Synthesis
5,11,17,23-Tetra-tert-butyl-25,27-dichloroformylmethoxy-
26,28-dihydroxycalix[4]arene (5) [12, 13]
To calix[4]arene diacid 4 [12, 13] (5.50 g, 7.19 mmol) in
toluene (140 ml) was added thionyl chloride (0.41 ml,
7.4 mmol) and the solution was heated at reflux tempera-
ture for 2 h. After cooling, the solution was concentrated in
vacuo to give a brown solid which was used without further
purification (5.72 g, 99 %). 1H NMR (CDCl3, d ppm) 7.07
(4H, s), 6.74 (4H, s), 6.14 (2H, s), 5.02 (4H, s), 4.29 (4H, d,
J = 12.0 Hz), 3.41 (4H, d, J = 12.0 Hz), 1.28 (18H, s),
0.96 (18H, s); 13C NMR (CDCl3, d ppm) 169.9, 149.8,
149.7, 142.3, 131.7, 129.2, 128.2, 127.9, 126.3, 125.4,
Experimental
General methodology
All solvents used for synthesis were either HPLC of AR
grade, purchased from Sigma-Aldrich, Labscan or Lennox.
The grade of petroleum ether used was 40–60°°C, unless
otherwise stated. All reactions requiring anhydrous condi-
tions were carried out under a nitrogen atmosphere. Flash
chromatography was performed using silica gel
78.7, 33.91; 33.90, 31.73, 31.70, 30.9; IR (KBr, cm-1
)
3502, 3049, 3025, 2962, 2905, 2868, 1813.
5,11,17,23-Tetra-tert-butyl-25,27-di(7-
methoxycarbonylmethyl-6-oxo-1,7-diaza-
spiro[4.4]nonane)-26,28-dihydroxycalix[4]arene (6)
˚
(0.040–0.063 mm) 60 A, purchased from Sigma-Aldrich.
All isolated products were dried under vacuum in advance
of characterisation.
To a solution of spirolactam trifluoroacetate salt 1 [11a]
(0.75 g, 2.32 mmol) in dichloromethane (50 ml) at 0 °C
was added triethylamine (3.25 ml, 23.2 mmol) and the
solution was stirred for 30 min at ambient temperature. To
this solution was added dropwise a solution of bis(acid
chloride) 5 (0.93 g, 1.16 mmol) in dichloromethane
(20 ml) and the solution was stirred at room temperature
for 5 days, then washed with 1 M HCl solution
(3 9 20 ml), brine (3 9 20 ml), dried over MgSO4 and
concentrated in vacuo. The resulting solid was purified by
column chromatography on silica gel in 5 % methanol/
dichloromethane to give an off white solid (0.69 g,
51.5 %). Further purification of this mixture of
diastereoisomers by column chromatography using 0.5 %
methanol/dichloromethane gave an off-white solid (0.14 g
(10 %). M.pt. 163–165 °C; Rf 0.26 (5 % methanol/
dichloromethane);1H NMR (CDCl3, d ppm) 7.41 (br s, 2H),
7.26 (CHCl3), 6.97 (s, 4H), 6.77 (s, 4H), 4.80–4.30 (m,
12H), 3.66 (s, 6H), 3.60–3.44 (m, 4H) 3.42–3.23 (m, 4H),
2.84–2.76 (m, 4H), 2.22–1.95 (m, 12H), 1.23 (s, 18H), 0.95
(s, 18H); 13C NMR (CDCl3, d ppm) 175.2, 169.4, 167.2,
152.9, 151.3, 149.3, 148.7, 132.7, 127.7, 125.8, 125.1,
76.4, 68.1, 52.1, 47.8, 44.9, 44.7, 36.6, 34.7, 34.2, 34.1,
31.8, 31.2, 29.7, 24.4; IR (KBr, cm-1) 3416, 2960, 2870,
1750, 1655; Microanalysis, C68H88N4O12Á0.3CHCl3
requires C, 68.98 %, H, 7.48 %, N, 4.71 %; found C,
Melting point analysis was performed using a Bibby
Stewart Scientific SMP1 melting point apparatus and are
uncorrected. Elemental analysis was carried out at
University College Dublin, Belfield using a CE 440 Ele-
mental Analyser. Mass Spectrometry analysis was carried
out at the Centre for Synthesis and Chemical Biology,
University College Dublin, Belfield.
1H and13C NMR spectroscopy was carried out on a Jeol
JNM-LA 300 FT-NMR 300 MHz spectrometer using either
CDCl3 or d-DMSO, using tetramethyl silane (TMS) as the
internal standard, unless otherwise stated. Chemical shifts
are expressed in parts per million (ppm, d), downfield from
the internal standard. Resolution is 0.18 Hz or 0.0006 ppm
for 1H-spectra (coupling constants at
0.62 Hz or 0.008 ppm for 13C.
0.36 Hz) and
Thin layer chromatography (TLC) was carried out with
the stated eluent system in a pre-saturated elution tank
using silica gel 60F254 (20 cm 9 20 cm 9 0.2 mm) alu-
minium backed plates. Preparative TLC was carried out
using silica gel 60F254 coated glass plates
(20 cm 9 20 cm 9 1000 lm). Compounds were detected
under ultraviolet light, by development in an iodine tank or
by dying with a potassium permanganate solution.
Infrared spectroscopy was carried out using a Nicolet
Avatar 320 FT-IR spectrometer running on the ‘‘Omnic’’
123

J Incl Phenom Macrocycl Chem
5,11,17,23-Tetra-tert-butyl-25,27-dicarboxymethoxy-
69.06 % H, 7.50 % N, 4.86 %; MS (ES?) calculated for
C68H88N4O12, [M ? H]? 1153.6, found [M ? H]? 1153.6.
26,28-dimethoxycalix[4]arene (10)
To
a
solution of 5,11,17,23-tetra-tert-butyl-25,27-di-
methyleneoxy-26,28-dihydroxycalix[4]
5,11,17,23-Tetra-tert-butyl-25,27-di(7-allyl-6-oxo-1,7-
ethoxycarbonyl
diaza-spiro[4.4]nonane)-26,28-dihydroxycalix[4]arene (7)
arene [12, 13, 15] (4.0 g, 4.7 mmol) in THF (40 ml) was
added sodium hydride (0.56 g, 23.5 mmol) and the mixture
was stirred for 30 min at ambient temperature. A solution of
iodomethane (0.65 ml, 10.34 mmol) in THF (5 ml) was then
added dropwise and the solution was stirred at room tem-
perature for 16 h. Water (5 ml) was added and the solution
was concentrated in vacuo. The solid residue was dissolved
in dichloromethane (30 ml), washed with 10 % aqueous HCl
solution (10 ml), water (3 9 10 ml), dried over MgSO4 and
concentrated in vacuo. 5,11,17,23-Tetra-tert-butyl-25,27-
diethoxycarbonyl methyleneoxy-26,28, dimethoxycalix[4]
arene was recovered from methanol as a white solid (3.12 g,
74 %). M.pt. 181–183 °C; Rf 0.69 (5 % ethyl acetate/pet-
roleum ether); IR (KBr, cm-1) 2961, 2821, 1763
Spirolactam derivatised calixarene 7 was prepared in a
similar manner to that described for 6 using trifluoroacetate
salt 2 [11a]. A mixture of diastereoisomers was recovered
after purification by column chromatography in 5 %
methanol/dichloromethane to give an off white solid
(0.62 g, 49 %). Further purification, as described above,
resulted in the isolation of one diastereoisomer as an off
white solid (0.18 g, 14 %). M.pt. 179–181 °C; Rf 0.32
(5 % methanol/dichloromethane); 1H NMR (CDCl3, d
ppm) 7.49 (s, 2H), 6.95 (s, 4H), 6.78 (s, 4H), 5.78–5.72 (m,
2H), 5.32–5.18 (m, 4H), 4.78 (d, 4H, J = 15.0 Hz), 4.55
(d, 4H, J = 15.0 Hz), 4.38–4.34 (m, 4H), 3.94–3.90 (m,
4H), 3.73 (m, 4H), 3.50–3.47 (m, 2H), 3.30–3.25 (m, 6H),
2.18–1.95 (m, 8H), 1.56 (H2 O), 1.22 (s, 18H), 0.97 (s,
18H); 13C NMR (CDCl3, d ppm) 173.6, 166.6, 151.1,
149.9, 147.3, 141.9, 133.6, 130.5, 132.7, 125.6, 125.0,
122.6, 74.7, 65.3, 49.8, 47.4, 43.5, 34.6, 34.3, 34.0, 31.6,
31.0, 27.6, 24.8; IR (KBr, cm-1) 3426, 2959, 2870, 1694,
1652, 1648, 901; Microanalysis, C68H88N4O8 requires C,
74.97 %, H, 8.14 %, N, 5.14 %; found C, 74.66 %, H,
7.54 %, N, 4.54 %; MS (ES?) calculated for C68H88N4O8,
expected [M ? H]? 1089.7, found [M ? H]? 1089.7.
To a suspension of 5,11,17,23-tetra-tert-butyl-25,27-di-
ethoxycarbonyl methyleneoxy-26,28, dimethoxycalix[4]
arene (3.0 g, 3.5 mmol) in ethanol (80 ml) was added
15 % aqueous NaOH solution (12 ml) and the solution was
heated to reflux temperature for 24 h, cooled and concen-
trated in vacuo. The resulting solid was suspended in H2O
(80 ml) and the pH of the mixture was adjusted to 1 using
3 M aqueous HCl solution. The resulting solid was isolated
by vacuum filtration, dissolved in dichloromethane
(50 ml), washed with 3 M HCl (10 ml), water
(3 9 10 ml), dried over MgSO4 and concentrated in vacuo.
Calix[4]arene 10 was recovered as a white solid from
ethanol/water (2.56 g, 98 %). M.pt. 259–261 °C; Rf 0.63
5,11,17,23-Tetra-tert-butyl-25,27-di(7-benzyl-6-oxo-1,7-
diaza-spiro[4.4]nonane)-26,28-dihydroxycalix[4]arene (8)
(10 % methanol/dichloromethane); IR (KBr, cm-1
3500–3200, 2962, 2869, 1760.
)
Spirolactam derivatised calixarene 8 was prepared in a
similar manner to that described for 6 using trifluoroacetate
salt 3 [11a]. A solid residue was purified was purified by
column chromatography on silica gel with 5 % methanol/
dichloromethane to give an off white solid (0.72 g, 52 %)
and further purified to give one diastereoisomer as an off
white solid (0.12 g, 9 %). M.pt. 264–266 °C; Rf 0.21 (5 %
methanol/dichloromethane);1H NMR (CDCl3, d ppm) 7.39
(s, 2H), 7.34–7.22 (m, 10H), 6.98 (s, 4H), 6.78 (s, 4H),
4.93–4.40 (m, 12H), 3.94–3.71 (m, 4H) 3.37–3.26 (m, 4H),
2.78–2.62 (m, 2H), 2.19–1.85 (m, 14H), 1.56 (H2 O), 1.24
(s, 18 H), 0.97 (s, 18 H); 13C NMR (CDCl3, d ppm) 173.6,
166.6, 151.5, 150.4, 146.9, 141.2, 136.3, 132.7, 132.1,
129.1, 128.7, 128.0, 127.8, 126.8, 127.7, 125.4, 73.9, 68.3,
47.6, 47.2, 43.2, 36.8, 34.4, 34.2, 33.8, 31.6, 31.0, 29.8,
24.4; IR (KBr, cm-1) 3421, 2961, 2868, 1697, 1655;
Microanalysis, C76H92N4O8Á3.5H2O requires C, 72.87 %,
H, 7.97 %, N, 4.47 %; found C, 72.71 %, H, 7.88 %, N,
4.94 %; MS (ES?) calculated for C76H92N4O8, expected
[M ? H]? 1189.7, found [M ? H]? 1189.7.
5,11,17,23-Tetra-tert-butyl-25,27-dichloroformylmethoxy-
26,28-dimethoxycalix[4]arene (11)
To a solution of calixarene 10 (2.0 g, 2.73 mmol) in
toluene (30 ml) was added thionyl chloride (0.15 ml,
2.8 mmol) and the solution was heated to reflux tempera-
ture for 2 h, cooled and concentrated in vacuo to give a
brown solid (2.08 g, 99 %). IR (KBr, cm-1) 2963, 2868,
1809.
5,11,17,23-Tetra-tert-butyl-25,27-di(7-methoxycarbonylmethyl-
6-oxo-1,7-diaza-spiro[4.4]nonane)-26,28-
dimethoxycalix[4]arene (12)
Spirolactam derivatised calixarene 12 was prepared in a
similar manner to that described for 6 using trifluoroacetate
salt 1 [11a]. A solid residue was purified by column
chromatography on silica gel in 5 % methanol/
123

J Incl Phenom Macrocycl Chem
dichloromethane to give an off white solid (0.19 g, 58 %).
M.pt. 146–148 °C; Rf 0.28 (5 % methanol/dichlor-
omethane); IR (KBr, cm-1) 2961, 2868, 1753, 1749, 1649;
HRMS (ES?) calculated for C70H92N4NaO12, expected
[M ? Na]? 1203.6606, found [M ? Na]? 1203.6609.
25 °C for 1 h. Mixing of 10 ml of metal picrate solution
with 10 ml of spirolactam-calix[4]arene solution was car-
ried out in stoppered plastic tubes. Complete mixing was
achieved using a mechanical water bath shaker for 1 h and
the biphasic solution was left standing for an additional
2 h. The water bath temperature was kept at a constant
25 °C throughout. The concentration of the picrate metal
ions remaining in the aqueous phase was then determined
spectrophotometrically at 355 nm. Blank experiments
showed that no picrate extraction occurred in the absence
of the spirolactam-calix[4]arene compounds. The average
of three samples is reported with the RSD of \5 %, in all
cases.
5,11,17,23-Tetra-tert-butyl-25,27-di(7-Allyl-6-oxo-1,7-
diaza-spiro[4.4]nonane)-26,28-dimethoxycalix[4]arene
(13)
Spirolactam derivatised calixarene 13 was prepared in a
similar manner to that described for 12 using trifluoroacetate
salt 2 [11a]. A solid residue was purified was purified by
column chromatography on silica gel with 5 % methanol/
dichloromethane to give 13 as an off white solid (0.17 g,
54 %). M.pt. 152–154 °C; Rf 0.36 (5 % methanol/dichlor-
omethane); IR (KBr, cm-1) 2960, 2873, 1690, 1651, 1649,
898; HRMS (ES?) calculated for C70H92N4O8, expected
[M ? H]? 1117.6946, found [M ? H]? 1117.6993.
The % extraction values of various ions, shown in Figs. 3
and 4, were calculated using the following equation:
ðA1 À AÞ
% E ¼ 100 Â
A0
where A0 is the absorbance of the 2.5 9 10-4 metal picrate
solution originally, A is the absorbance of the picrate salt in
the aqueous layer after extraction, A1 is the absorbance
from of the blank experiments in the absence of spirolac-
tam-calix[4]arene.
5,11,17,23-Tetra-tert-butyl-25,27-di(7-allyl-6-oxo-1,7-
diaza-spiro[4.4]nonane)-26,28-dimethoxycalix[4]arene
(14)
Spirolactam derivatised calixarene 14 was prepared in a
similar manner to that described for 12 using trifluoroac-
etate salt 3 [11a]. A solid residue was purified was purified
by column chromatography on silica gel with 5 % metha-
nol/dichloromethane to give 14 as an off white solid
(0.21 g, 66 %). M.pt. 232–234 °C; Rf 0.25 (5 % methanol/
dichloromethane); IR (KBr, cm-1) 2963, 2870, 1699, 1656;
HRMS (ES?) calculated for C78H96N4O8, expected
[M ? Na]?: 1239.7125, found [M ? Na]? 1239.7126.
Funding J.W. funded by Strand I of the Irish Government’s
National Development Plan Technological Sector Research Program
and the ITT Dublin PhD Continuation Fund. The work of L.L. was
supported by QUESTOR Centre, Belfast and Dublin City University.
Compliance with ethical standards
Conflict of Interest The authors declare that they have no conflict
of interest.
Binding studies
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