| Ethosomes are novel liposomes with high content of alcohol(20%～50%,w/w)added to the basis of traditional liposomes,which have the characteristic of high fluidity,high encapsulation rate,and strong skin permeability.However,the high fluidity of ethosomes can lead to poor stability and drug leakage,limiting their practical application.Therefore,it is necessary to adopt certain modification methods to improve the performance of ethosomes.Ceramide 3(Ceramide 3,Cer3)is a common lipid in the stratum corneum,which can reduce the fluidity of the carrier.Therefore,Cer3 was used for the first time to modify the ethosomes to improve its stability in this study.In addition,the single-encapsulated ethosomes have been extensively reported,but the double-encapsulated ethosomes are rarely studied.Therefore,in this topic,lipid soluble active substance--retinol palmitate(RP)and water-soluble active substance--carnosine(Car)were further selected for encapsulation,and the stability mechanism and physicochemical properties of the carrier were explored.Finally,in order to reduce the irritation caused by ethanol in the carrier,RP-Car binary cerethosomes were constructed using a combination of propylene glycol and ethanol,and the in vitro transdermal,antioxidant,and cytotoxic properties of them were investigated.The main research results of this topic are as follows:Firstly,the optimal preparation conditions were determined based on appearance,particle size,PDI,and encapsulation efficiency as indicators:absolute ethanol volume fraction was33.3%,soybean lecithin(PC60)content was 25 mg/mL,Cer3 content was 1 mg/mL,cholesterol content was 1 mg/mL,RP content was 2 mg/mL,Car content was 50 mg/mL,hydration temperature was 45℃,and hydration time was 20 minutes.The encapsulation efficiency of RP was(92.14±0.88)%,the encapsulation efficiency of Car was(30.56±0.53)%,the particle size was(64.8±2.3)nm,and the PDI was(0.143±0.019)under these conditions.TEM results showed that the drug-loaded ethosomes were multilayer vesicles before and after Cer3modification,but the particle size of ethosomes was reduced by Cer3 modification.FTIR results showed that the drug was well encapsulated in the carrier.FTIR results showed that the drug was well encapsulated.The phase transition temperature of drug-loaded ethosomes increased after Cer3 modification in DSC analysis.Secondly,the stability of drug-loaded ethosomes before and after Cer3 modification was compared,and the results showed that the stability of drug-loaded ethosomes after Cer3modification was improved;After storage at 4℃,25℃,and 45℃for 120 days,there were no significant changes in the appearance and particle size of RP-Car cerethosomes.The FTIR and changes in membrane microviscosity of RP-Car cerethosomes with different Cer3 contents indicated that the hydrogen bonding between Cer3 and cholesterol enhanced the molecular interaction in the bilayer membrane,and the membrane microviscosity increased,thus enhancing the stability.The transdermal properties of drug-loaded ethosomes before and after Cer3 modification were compared by Franz diffusion cell method.HPLC,CLSM and Raman imaging results showed that the transdermal penetration of RP-Car cerethosomes increased;The skin accumulation of RP increased from(0.194±0.004)μg/cm~2 to(0.245±0.001)μg/cm~2,and the skin permeability of RP increased from 0μg/cm~2 to(1.834±0.047)μg/cm~2.The skin accumulation of Car increased from(15.612±0.596)μg/cm~2 to(18.079±0.236)μg/cm~2,and skin permeability of Car increased from(29.822±0.119)μg/cm~2 to(63.501±0.476)μg/cm~2.ATR-FTIR revealed the transdermal mechanism of RP-Car cerethosomes as follows:the carrier could significantly enhance the skin hydration and loose keratin in the skin,as well as replenish the intercellular lipids in the stratum corneum and change them from ordered to disordered.RP-Car cerethosomes showed higher antioxidant activity compared to single encapsulated RP or Car ethosomes,and Cer3 modification reduced cytotoxicity to HaCat to some extent.Finally,in order to further improve the stability of the carrier and reduce irritation,RP-Car binary cerethosomes were prepared using an ethanol/PG ratio of 6:4,and the encapsulation efficiency of the water-soluble active substance Car increased to(47.20±0.71)%.TEM results showed that RP-Car binary cerethosomes was a regular spherical vesicle with a particle diameter of about 80 nm.Compared with RP-Car cerethosomes,the stability of RP-Car binary cerethosomes was improved due to the increase in microviscosity of the hydrophilic region of the bilayer phospholipid head by PG;Moreover,the high viscosity of PG delayed drug release and resulted in more drug deposition in the skin.The skin accumulation of RP and Car increased to(0.273±0.002)μg/cm~2 and(22.521±0.280)μg/cm~2,respectively.The skin permeability of RP and Car decreased to 0μg/cm~2 and(20.559±0.120)μg/cm~2,respectively.In addition,the antioxidant properties of RP-Car binary cerethosomes were basically unchanged,and the cytotoxicity was decreased. |
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