Preparation of Giant Vesicles Exhibiting Visible-light-induced Morphological Changes
Summary
The synthesis of ruthenium complex surfactants exhibiting photoisomerization in giant vesicles is described. The preparation and light irradiation of the giant vesicles are also described.
Abstract
We describe the preparation of giant vesicles that incorporate a photoresponsive ruthenium complex having two alkyl chains. The vesicles exhibited morphological changes when exposed to visible light. The ruthenium complex proximal-[Ru(L1)(L2)OH2](NO3)2, proximal–2 (L1 is 4′-decyloxy-2,2′;6′,2″-terpyridine, L2 is 2-(2′-(6′-decyloxy)-pyridyl)quinoline) was prepared by a thermal reaction of Ru(L1)Cl3 and L2, followed by removal of a chloride ligand. In an aqueous solution and vesicle dispersions, proximal–2 was reversibly photoisomerized to the distal isomer. Giant vesicles containing proximal–2 were prepared by hydration of phospholipid films containing proximal–2 in the dark at 80 °C. Giant vesicles were frequently found in the dispersions prepared from DOPC/proximal–2 rather than from DPPC/proximal–2 (DOPC is 1,2-dioleoyl-sn-glycero-3-phosphocholine, DPPC is 1,2-dipalmitoyl –sn-glycero-3-phosphocholine). The ratio of proximal–2 and DOPC in the vesicle preparation was varied from 5:100 to 20:100. The light-induced morphological changes were observed for proximal–2/DOPC in the presence of Na2SO4. However, they were highly suppressed in the presence of NaOH. Incubation of light-exposed vesicles at 45 °C in the dark induced reverse morphological changes. Morphological changes were observed under fluorescence microscopy using 635 nm (red) light. Rhodamine-DOPC [rhodamine-DOPC: 1,2-dioleoyl-sn-glycero-3-phos-phoethanolamine-N-(lissamine rhodamine B sulfonyl)] was used to fluorescently label the vesicles.
Introduction
Controlling the morphologies and shapes of macro- and meso-scale molecular assemblies by external stimuli has attracted considerable attention.1,2 In particular, the control of vesicle morphologies by remote stimuli such as light has potential applications for drug delivery.3 In this context, organic photochromic molecules with hydrophobic and hydrophilic moieties have been widely incorporated into liposomes and polymer vesicles.4,5,6,7,8 However, most of the assemblies require ultraviolet (UV) light to drive the morphological changes, and their applications are limited because UV light is strongly scattered in living tissues and induces DNA damage and cell death.
Alternatively, utilization of visible or near-infrared light in the phototherapeutic window (600-1000 nm) is more favorable because of abundant sunlight and its high transmission in tissues of living organisms. In this regard, ruthenium complexes with polypyridyl ligands are suitable visible-light-responsive surfactants. They exhibit a strong visible light absorption band (ε~104 M-1 cm-1) that induces ligand substitution9,10 and photoisomerization.11,12,13,14,15,16 Incorporation of the ruthenium complexes into vesicles will expand their applications because these complexes are also known as water oxidation catalysts17,18,19 and bioactive molecules.20,21 Recently, ruthenium complexes have been incorporated into vesicles.22,23,24 However, controlling morphologies of vesicles via visible-light absorption has remained challenging.
We have previously reported irreversible and reversible photoisomerization of mononuclear ruthenium aqua complexes having asymmetric bidentate ligands.25,26,27,28 Recently, we synthesized novel surfactants (proximal–2, see Figure 1) that exhibit visible-light photoisomerization equilibria with distal–2 by introducing an alkyl chain on each tridentate and bidentate ligand of the ruthenium aqua complex. Giant vesicles incorporating proximal–2 undergo morphological changes under the irradiation of visible light in the phototherapeutic window.29 Herein, we describe the detailed syntheses of ruthenium complexes and the preparation of giant vesicles. The protocols will enable researchers to prepare, characterize, and utilize light-responsive giant vesicles.
Figure 1: Ruthenium complex surfactants. Reversible photoisomerization equilibrium between proximal- and distal- type complex of 1 (top) and 2 (bottom). Please click here to view a larger version of this figure.
Protocol
Representative Results
Discussion
The ruthenium chloro complex proximal-[Ru(L1)(L2)Cl]+ was prepared by thermal synthesis of Ru(L1)Cl3 and a bidentate ligand L2 in the presence of triethylamine. The proximal isomer was the major product and a distal isomer and Ru(L1)22+ was a minor impurity. The crude product was purified with size-exclusion chromatography using methanol as an eluent. Coordinating solvents, such as wat…
Divulgations
The authors have nothing to disclose.
Acknowledgements
The authors have no acknowledgements.
Materials
| Triethylamine | Wako Pure Chemical Industries, Ltd. | 202-02646 | |
| Lithium Chloride | Wako Pure Chemical Industries, Ltd. | 125-01161 | |
| Chloroform | Kanto Chemical Co. Ltd. | 07278-03 | Used for vesicle preparation |
| Chloroform | Junsei Chemical Co. Ltd. | 28560-0382 | Used for ligand synthesis |
| Acetone | Junsei Chemical Co. Ltd. | 11265-0382 | |
| Ethanol | Junsei Chemical Co. Ltd. | 17065-0382 | |
| Ethyl Acetate | Junsei Chemical Co. Ltd. | 67150-0382 | |
| Hexane | Junsei Chemical Co. Ltd. | 31055-0382 | |
| Silica gel | Kanto Chemical Co. Ltd. | 37558-79 | 100-210 μm |
| 1-decanol | Tokyo Chemical Industry Co., Ltd. | D0031 | 25 mL |
| Potassium hydroxide | Kanto Chemical Co. Ltd. | 32344-00 | |
| Sodium hydrixude | Wako Pure Chemical Industries, Ltd. | 197-02125 | |
| Dimethyl sulfoxide (DMSO) | Kanto Chemical Co. Ltd. | 10378-00 | |
| d-DMSO | Sigma-Aldrich | 166290100 | |
| CD3OD | Kanto Chemical Co. Ltd. | 25221-43 | |
| d-Acetone | Kanto Chemical Co. Ltd. | 01054-43 | |
| D2O | Cambridge Isotope Laboratories, Inc. | DLM-4-100 | |
| Ruthenium chloride n-Hydrate | Wako Pure Chemical Industries, Ltd. | 183-00823 | |
| 2,2':6',2"-Terpyridine | Sigma-Aldrich | 234672-5G | |
| 0.1 mol/L Silver nitrate solution | Wako Pure Chemical Industries, Ltd. | 192-00855 | |
| Sodium sulfate | Kanto Chemical Co. Ltd. | 37280-00 | |
| 1,2 Dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) | Wako Pure Chemical Industries, Ltd. | 160-12781 | 100 mg, stored at -20°C |
| 1,2 Dioleoyl-sn-glycero-3-phosphocholine (DOPC) | Sigma-Aldrich | P6354-100mg | 100 mg, stored at -20°C |
| 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine-N-(carboxyfluorescein) (ammonium salt) | Avanti Polar Lipids, Inc. | Avanti 810332p | 5 mg, stored at -20°C |
| 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine-N-(lissamine rhodamine B sulfonyl) (ammonium salt) | Avanti Polar Lipids, Inc. | Avanti 810150c | 1 mg, stored at -20°C |
| Dextran gel | GE healthcare Japan | 17009010 | Sephadex LH-20 |
| Amber glass vial | Maruemu | 0407-06 | |
| Septum | Sigma-Aldrich | Z564648-100EA | |
| Heater | Advantech | DRM 320 DB | |
| Silicon film | AS ONE | 6-9085-03 | Thickness: 0.2 mm |
| Slide glass | Matsunami | S003130 | 76×26 mm, thickness: 0.8-1.0 mm |
| Cover glass | Matsunami | C218181 | 18×18 mm, thickness: 0.12-0.17 mm |
| Transfer pipette | Brand GMBH | 704774 | |
| Round-bottom flask | Vidtech | 1500-05 | |
| Sonicator | AS ONE | 1-4591-32 | |
| Optical power meter | OPHIR | ORION/PD P/N 1Z01803 | |
| Oil bath | Riko | MH-3D | |
| Magnetic stirrer | Riko | MSR-10 | |
| Diatomite | Wako Pure Chemical Industries, Ltd. | 537-02305 | Celite 545 |
| Evaporator | Yamato | RE-52 | |
| Glass funnel | Kiriyama | SB-21 | 10 mL, 21 mmφ |
| Bell jar | Kiriyama | VKB-200 | |
| Filter paper | Kiriyama | No.4 | 21 mmφ |
| Optical microscope | KEYENCE | VHX-5000 | |
| Confocal fluorescence microscope | Olympus | FV-1000 |
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