Single Electrochemical Nano-Impacts of Synthetic Redox Phospholipid Liposomes
Authors: Lebegue Estelle , Barrière Frédéric , Bard Allen Joseph 
Published: July 2021

Electrochemical discrete nano-impacts at an ultramicroelectrode is a useful technique to detect one at a time single biological entities such as cells, bacteria, macromolecules, viruses, and synthetic or biological vesicles.[1-4] Especially, electrochemical detection of single liposome nano-impacts by recording electron transfer from the ultramicroelectrode to an encapsulated redox species is fully appropriate for studying their membrane permeability.[2,4] Since electron transfer do not readily occur across a bilayer lipid membrane, the electrolysis of the liposome redox active content after collision and membrane rupture or opening at the electrode surface provides insights on the membrane permeation mechanism. The vesicle membrane opening by electroporation is strongly dependent on lipid membrane properties, liposome content, vesicle size, temperature, electrode potential, the nature of the electrode and the concentration of redox species inside and outside the liposome.[2] For example, to observe the current spikes corresponding to the oxidation of ferrocyanide encapsulated inside phospholipid vesicles when they collide with a Pt ultramicroelectrode, the presence of an appropriate concentration of surfactant in solution is required.[4] In the absence of surfactant, we found that impact and adhesion of vesicles at the Pt ultramicroelectrode does not allow the electrolysis of their ferrocyanide content.[4] It is well established that the liposome membrane stability is strongly dependent on its lipid composition and external parameters such as temperature and pH but also depends on interactions with specific molecules able to weaken, permeabilize, and penetrate the lipid bilayer following different pathways. An understanding of these different interactions and especially the mechanism leading to the lipid membrane opening is still an active research field. To this end, chronoamperometry is a useful method to probe the liposomes membrane permeability and to understand vesicle fusion processes onto electrode surface. Especially, the electrochemical detection of single liposome collisions at ultramicroelectrodes is becoming an efficient and complementary tool for analyzing fundamental biological processes related to intracellular and extracellular electron transfers to discrete biological or artificial entities. Electrochemistry of single redox liposome nano-impacts at an ultramicroelectrode is a powerful method which consists in detecting the electrolysis of a redox probe encapsulated inside the liposome when it is released at the ultramicroelectrode after impact (or collision).[4]

To explore the factors influencing the vesicles membrane permeability, we investigated the electrochemical and electrocatalytic reaction of different aqueous redox species (potassium ferrocyanide and cobalt(II) nitrate) encapsulated inside synthetic DMPC (1,2-dimyristoyl-sn-glycero-3-phosphocholine) liposomes subjected to different experimental conditions (addition of surfactant, increase of temperature or presence of a redox probe in solution), by single collision detection on 10 μm diameter ultramicroelectrodes (Pt and carbon) as illustrated in Figure.[2] The influence of the presence of surfactant (Triton X-100) and the increase of solution temperature (20-70 °C) on the lipid membrane permeability has been investigated, and the results showed a similar effect of these two parameters with a maximum collision frequency reached for an optimal surfactant concentration (0.2 mM) and temperature (60 °C) respectively, at platinum or carbon ultramicroelectrodes.[2]

[1] Lebègue, E.; Costa, N.L.; Louro, R.O.; Barrière, F. Communication—Electrochemical Single Nano-Impacts of Electroactive Shewanella Oneidensis Bacteria onto Carbon Ultramicroelectrode. J. Electrochem. Soc.
2020, 167, 105501, doi:10.1149/1945-7111/ab9e39.

[2] Lebègue, E.; Barrière, F.; Bard, A.J. Lipid Membrane Permeability of Synthetic Redox DMPC Liposomes Investigated by Single Electrochemical Collisions. Anal. Chem.
2020, 92, 2401–2408, doi:10.1021/acs.analchem.9b02809.

[3] Dick, J.E.; Lebègue, E.; Strawsine, L.M.; Bard, A.J. Millisecond Coulometry via Zeptoliter Droplet Collisions on an Ultramicroelectrode. Electroanalysis
2016, 28, 2320–2326, doi:10.1002/elan.201600182.

[4] Lebègue, E.; Anderson, C.M.; Dick, J.E.; Webb, L.J.; Bard, A.J. Electrochemical Detection of Single Phospholipid Vesicle Collisions at a Pt Ultramicroelectrode. Langmuir
2015, 31, 11734–11739, doi:10.1021/acs.langmuir.5b03123.

Figure 1