# Positively Charged Lipid as Potential Tool to Influence the Fate of Ethosomes
## 4 Conclusions  
The role of cationic charges on vesicular systems represents a new trend for topical delivery of active compounds, which needs to be better clarified. In this attempt cationic endosomal systems made with small amounts of cationic lipids have been realized in this study and compared with data recorded from conventional ethosomal formulations.  
The results obtained showed that the addition of small quantities of DOTAP to conventional ethosomes allows to realize positively charged systems. In detail, the systems made of PL90G: DOTAP 10: 1 or 10: 0.5 molar ratio proved to be the most suitable for a possible topical skin application. In our physico-chemical and technological formulation studies, these systems showed average sizes below 300 nm, a low polydispersity index (below 0.2) and a net positive surface charge (over +50 mV) conferring a certain stability to the system. Further characterization studies performed using Turbiscan Lab analysis confirmed the stability of the realized formulations, thus recording values of \(\Delta\)Backscattering within \(\pm\)1. The storage of samples over one month also confirmed the great stability of the realized samples as no marked variations in size and polydispersity index occurred during this period. Cationic ethosomes presented good encapsulation efficacy of hydrophilic and lipophilic model compounds, comparable to that recorded from conventional ethosomes. In detail, the encapsulation of oil red o or rhodamine b was found to be above 50% of the amount added during the preparation. Toxicity studies carried out on NCTC2544 cells and on healthy volunteers finally proved the suitability of these systems for cutaneous application as they were able to maintain cell viability above 70% within 72 h. Concerning preparation made of DOTAP (10:0.5 and 10:1 molar ratio) they also reported great safety profiles on NCTC 2544 cells at 72 h over lipid concentration of 1.0 \(\upmu\)g/mL. Finally, no relevant irritation phenomena occurred after 48 h from their application to the forearm of healthy human volunteers.  
Thus, we can conclude that cationic ethosomes made of PL90G:DOTAP at a 10:1 molar ratio and 10:0.5 molar ratio represent promising systems for further permeation studies, which will allow us to investigate the influence of surface charge on the fate of such deformable systems.  
The following are available online at [[https://www.mdpi.com/article/10](https://www.mdpi.com/article/10)]([https://www.mdpi.com/article/10](https://www.mdpi.com/article/10)) 3390/app11157060/s1, Figure S1: Delta transmission (\(\Delta\)T) and delta backscattering (\(\Delta\)BS) profiles of conventional ethosomes (formulation A). Figure S2: Delta transmission (\(\Delta\)T) and delta backscattering (\(\Delta\)BS) profiles of probe-loaded ethosomal formulations.  
Conceptualization, D.P.; methodology, D.P. and M.F. investigation, A.M. and M.C.C.; data curation, A.M. and D.T.; writing--original draft preparation, A.M.; writing--review and editing, M.C.C.; supervision, D.P. All authors have read and agreed to the published version of the manuscript.  
This research received no external funding.  
The study was conducted according to the guidelines of the Declaration of Helsinki and approved by the Ethics Committee of the Magna Gracia University of Catanzaro (protocol code 391 of 19 December 2019).  
Informed Consent Statement: Informed consent was obtained from all subjects involved in the study.  
The data presented in this study are available on request from the corresponding author.  
The authors declare no conflict of interest.