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Logo of nihpaAbout Author manuscriptsSubmit a manuscriptHHS Public Access; Author Manuscript; Accepted for publication in peer reviewed journal;
Chem Commun (Camb). Author manuscript; available in PMC 2010 May 10.
Published in final edited form as:
PMCID: PMC2866091

meso-Octamethylcalix[4]pyrrole: an old yet new transmembrane ion-pair transporter

The first example of lipid bilayer membrane transport of a salt by a calix[4]pyrrole is reported.

The development of ditopic ion-pair receptors has recently attracted much interest.1 Ditopic receptors that simultaneously bind anion and cation pairs have advantages in terms of substrate affinity and selectivity over monotopic hosts. Moreover, in systems where the bound cation and anion carry the same charge, the supramolecular host–salt complex formed will be neutral. This gives the complex optimal solubility in organic solvents, an important advantage when applying these systems in extraction processes or as transmembrane carriers for charged species. On the other hand, to the best of our knowledge there is only one example of a synthetic molecule capable of simultaneous transmembrane ion-pair transport across a lipid bilayer,2 and no examples of natural products capable of salt binding and transport, although related formal H+/Cl co-transport is a common mechanism in both natural (prodigiosins)3 and synthetic chloride carriers.4

Calix[4]pyrrole derivatives have been extensively studied as anion receptors over the past few years.5 These compounds can be made in high yield in one step and are easy to functionalise, making them attractive binding motifs for an-ionic guests. More recently their potential as ion-pair receptors has been recognised.6 Preorganisation of the macrocycle into a cone conformation upon anion binding allows the inclusion of large, charge diffuse cations such as caesium or organic cations such as imidazolium or pyridinium into the aromatic “cup”, as demonstrated both in solution and solid state studies.7 These findings led to the application of meso-octamethylcalix[4]pyrrole as an extractant for halide salts from aqueous to organic solutions.8,9 Herein we report the first example of transmembrane transport in 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) and POPC–cholesterol vesicles of a salt by meso-octamethylcalix[4]pyrrole.

Calix[4]pyrrole derivatives 1–5 (Fig. 1) were synthesised by condensation of pyrrole and the appropriate ketone according to literature procedures.10 In order to study the transport properties of these macrocycles we prepared unilamellar POPC vesicles11 loaded with CsCl and suspended them in an external NaNO3 solution. A sample of the respective calix[4]pyrrole 1–5 (2% molar carrier to lipid) was added as a DMSO solution and the resultant Cl efflux monitored using a chloride selective electrode.12 After five minutes the vesicles were lysed by addition of detergent and the final reading of the electrode used to calibrate 100% release of chloride. The results are shown in Fig. 2.

Fig. 1
Structures of calix[4]pyrroles 1–5.
Fig. 2
Chloride efflux promoted upon addition of 1 (■), 2 (○), 3 (▲), 4 (□), and 5 (●) (2% molar carrier to lipid) to unilamellar POPC vesicles ...

Under these conditions meso-octamethylcalix[4]pyrrole 1 showed excellent transport activity inducing the release of virtually all of the encapsulated chloride within 5 min. Different substitution at the meso-position of the macrocycle resulted in a dramatic loss of transport activity with only the tetraspirocyclohexyl derivative 5 showing detectable activity as a carrier. There is no immediate explanation for the lack of activity of compounds 2–4, although the possibility that poor solubilities of these derivatives under the conditions of the assay hamper the incorporation of the macrocycles into the phospholipid bilayer cannot be ruled out. We then explored the cation selectivity of the transport process and similar assays using NaCl, KCl and RbCl loaded vesicles were carried out (Fig. 3). Lack of chloride efflux in these experiments is evidence that supports the hypothesis that chloride is transported selectively as a calixpyrrole-bound ion-pair with caesium. These assays also rule out the possibility of Cl efflux being promoted by simple disruption of the vesicles by the macrocycle. We also wished to investigate the influence of the composition of the external medium on the transport activity shown by 1. For this purpose CsCl loaded vesicles were suspended in buffered Na2SO4 solution (Fig. 3). The result shows that carrier activity is essentially maintained and is independent of the external anion, further supporting a CsCl co-transport mechanism for this process (Scheme 1).

Fig. 3
Chloride efflux promoted upon addition of 1 (2% molar carrier to lipid) to unilamellar POPC vesicles loaded with 488 mM CsCl (■), or RbCl (○), or KCl (▲), or NaCl (□), 5 ...
Scheme 1
Proposed mechanism for chloride efflux promoted by calix[4]pyrrole 1.

We screened the transport activity of 1 through a range of carrier concentrations (20–0.1 μM, 2–0.01 mol% carrier to lipid) repeating the transport assays using different amounts of macrocycle (Fig. 4). Almost complete chloride efflux is obtained with loadings of 2–0.1% molar carrier to lipid within the first five minutes of the experiment; meanwhile the transport activity clearly decreases with lower carrier concentrations, although detectable activity is observed even at 0.01% molar carrier to lipid (0.1 μM) concentrations.

Fig. 4
Chloride efflux promoted upon addition of 1 (20–0.1 μM; 2–0.01% molar carrier to lipid) to unilamellar POPC vesicles loaded with 488 mM CsCl, 5 mM phosphate buffer, pH 7.2, ...

Finally, in order to shed light into the transport mechanism of the process we performed transport assays using vesicles composed of phospholipid and cholesterol. The results, shown in Fig. 5, clearly indicate a significant reduction in the transport activity of 1 when the vesicle bilayer composition includes cholesterol. Incorporation of the steroid to the bilayer membrane significantly reduces its fluidity and this result indicates that a carrier mechanism is likely for this process.13

Fig. 5
Chloride efflux promoted upon addition of 1 (2% molar carrier to lipid) to unilamellar POPC vesicles (■), or POPC–cholesterol (70: 30 molar ratio) (●) vesicles loaded ...

In conclusion, we have demonstrated that meso-octamethylcalix[ 4]pyrrole mediates the selective transport of an ion pair (CsCl) through model phospholipid bilayers. Efforts aimed towards producing efficient ion-pair carriers with different selectivities are currently underway in our laboratories.

Supplementary Material



PAG thanks the EPSRC for a postdoctoral fellowship (CCT). RQ thanks the Spanish Ministerio de Educación y Ciencia for a Ramon y Cajal contract and Jose Castillejo grant and the Royal Society of Chemistry for a Journals Grant. JLS acknowledges NIH grant GM 58907 for support.


This communication is dedicated to an inspiring leader in the supramolecular chemistry arena, Professor Seiji Shinkai, on the occasion of his 65th birthday.

Electronic supplementary information (ESI) available: Synthesis and characterisation data of compound 4. See DOI: 10.1039/b814988g

Notes and references

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5. (a) Sessler JL, Gale PA, Cho WS. Synthetic Anion Receptor Chemistry. ch 5 Royal Society of Chemistry; Cambridge: 2006. (b) Gale PA, Hursthouse MB, Light ME, Sessler JL, Warriner CN, Zimmerman R. Tetrahedron Lett. 2001;42:6759–6762. (c) Gale PA, Anzenbacher P, Jr, Sessler JL. Coord Chem Rev. 2001;222:57–102. (d) Gale PA, Král V, Sessler JL. Chem Commun. 1998:1–8.
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7. (a) Sessler JL, Gross DE, Cho WS, Lynch VM, Schmidtchen FP, Bates GW, Light ME, Gale PA. J Am Chem Soc. 2006;128:12281–12288. [PubMed] (b) Bates GW, Gale PA, Light ME. CrystEngComm. 2006;8:300–302. (c) Bates GW, Gale PA, Light ME. Supramol Chem. 2008;20:23–28. (d) Gross DE, Schmidtchen FP, Antonius W, Gale PA, Lynch VM, Sessler JL. Chem–Eur J. 2008;14:7822–7827. [PubMed]
8. Wintergerst MP, Levitskaia TG, Moyer BA, Sessler JL, Delmau LH. J Am Chem Soc. 2008;130:4129–4139. [PubMed]
9. For an example of a calix[4]pyrrole decorated polymer see Aydogan A, Coady DJ, Lynch VM, Akar A, Marquez M, Bielawski CW, Sessler JL. Chem Commun. 2008:1455–1457. [PubMed]
10. Gale PA, Sessler JL, Král V, Lynch V. J Am Chem Soc. 1996;118:5140–5141. and references cited therein.Jubb J, Scoles L, Jenkins H, Gambarotta S. Chem–Eur J. 1996;2:767–771.
11. A chloroform solution of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) (20 mg mL−1) (Genzyme) or a 70: 30 POPC–cholesterol mixture was evaporated using a rotary evaporator and the lipid film obtained was dried under high vacuum for at least 2 h. The lipid film was rehydrated by addition of a metal chloride (MCl) salt solution (488 mM MCl and 5 mM phosphate buffer, pH 7.2) and careful vortexing. The lipid suspension was then subjected to nine freeze–thaw cycles and twenty-nine extrusions through a 200 nm polycarbonate nucleopore membrane using a LiposoFast Basic extruder (Avestin, Inc.) to obtain unilamellar vesicles. The vesicles were dialysed against a NaNO3 solution (488 mM NaNO3 and 5 mM phosphate buffer, pH 7.2) or Na2SO4 (162 mM Na2SO4 and 5 mM phosphate buffer, pH 7.2) to remove unencapsulated MCl salts.
12. Koulov AV, Lambert TN, Shukla R, Jain M, Boon JM, Smith BD, Li HY, Sheppard DN, Joos JB, Clare JP, Davis AP. Angew Chem, Int Ed. 2003;42:4931–4933. [PubMed]In this study the following procedure was used: unilamellar vesicles (200 nm mean diameter) composed of POPC or POPC–cholesterol containing an encapsulated solution of 488 mM MCl and 5 mM phosphate buffer pH 7.2, were suspended in a solution of 488 mM NaNO3 and 5 mM phosphate buffer pH 7.2, or 162 mM Na2SO4 and 5 mM phosphate buffer pH 7.2, for a final lipid concentration of 1 mM. A DMSO solution of the carrier molecule, typically 10 μL to avoid influence of the solvent molecules in the assay, was added and the chloride release from vesicles was monitored using an Accumet chloride selective electrode for 7 min. At a time (t) = 5 min the vesicles were lysed with detergent (polyoxyethylene (8) lauryl ether) to release all chloride ions; the resulting value was considered to represent 100% release and used as such.
13. Smith BD, Lambert TN. Chem Commun. 2003:2261–2268. [PubMed]