펩타이드가 봉입된 가변형 에토좀의 피부투과 개선효과
Transformer-lipid carriers for improving skin delivery of peptide
- 주제(키워드) Palmitoyl pentapeptide , Transformer-Ethosome , Skin permeation , Collagen synthesis , and Deformability
- 발행기관 아주대학교
- 지도교수 박영준
- 발행년도 2018
- 학위수여년월 2018. 2
- 학위명 박사
- 학과 및 전공 일반대학원 약학과
- 실제URI http://www.dcollection.net/handler/ajou/000000026773
- 본문언어 영어
- 저작권 아주대학교 논문은 저작권에 의해 보호받습니다.
초록/요약
The purpose of the present study was to prepare flexible lipid carriers loaded with palmitoyl pentapeptide (PPP) to improve its skin delivery and provide an anti-skin aging effect in the absence of any abnormal skin responses. PPP is a palmitoylated derivative of pentapeptide (Lys-Thr-Thr-Lys-Ser, KTTKS). Palmitoylation provides the peptide with a more desirable partition coefficients, an important consideration for skin absorbance. However, by itself, PPP has limitations with respect to its instability on or in the skin, and has poor penetration and permeation across the skin due to its high molecular weight (802.05 Da), and net charge under the physiological conditions. PPP-loaded conventional liposomes (PPP-CLs), ethosomes (PPP-ESs), and transformer-ethosomes (PPP-TESs) were prepared by a modified extrusion method using phosphatidylcholine (PC) mixed with different ratios of cholesterol or fatty acids. The physicochemical characteristics of palmitoyl pentapeptide-loaded transformer-ethosomes (PPP-TESs) and their skin permeation properties were evaluated. PPP-CLs, PPP-ESs, and PPP-TESs had mean diameters < 150 nm with a homogeneous particle population, a net positive surface charge, and an encapsulation efficiency (EE) of above 97 %. DSC thermograms showed that the incorporation of capric acid (CA), linoleic acid (LA), and myristic acid (MA) into the lipid bilayers altered the transition temperature of the lipid bilayers and also improved the flexibility. The flexibility values of the TESs containing MA, CA, and LA increased in an inverse proportion to the transition temperature of the formulations. Skin permeation of PPP-TESs through both of an artificial membrane and human cadaver skin was much higher than that seen for PPP-CLs and PPP-ESs. In particular, TESs inserted with MA (TESs-MA) was found to be the most effective in improving skin permeation. Moreover, confocal laser scanning microscopy (CLSM) showed that rhodamine-6G-loaded TES-MA (R6G-TES-MA) penetrated into the deeper skin layer. The anti-aging effect of PPP-TES was assessed in normal human dermal fibroblasts (nHDFs) by measuring the mRNA expression of collagen type I alpha 1 chain (COL1A1) and matrix metalloproteinase-1 (MMP-1). nHDFs treated with PPP-lipid carriers had increased type I procollagen level, but decreased MMP-1 levels. In a clinical study, we observed that application of a PPP-lipid carriers solution increased skin elasticity and also decreased facial wrinkles, without abnormal skin responses. These results demonstrated the potential of TESs to enhance skin delivery, indicating that they can provide an anti-skin aging effect both in vitro and in clinical study. Conclusively, TES could be a promising candidate for the transdermal delivery of PPP as well as other drugs.
more목차
1. Introduction 1
1.1. Skin structure and its penetration route 1
1.1.1. Skin structure, function, and its cells 1
1.1.2. Routes of drug penetration through the skin 4
1.2. Skin aging 6
1.2.1. Intrinsic versus extrinsic skin aging 7
1.3. Peptides for anti-skin aging 10
1.3.1. Signal peptides 10
1.3.2. Neurotransmitter-affecting peptides 11
1.3.3. Carrier peptides 11
1.4. Palmitoyl pentapeptide 11
1.5. Skin penetration enhancement systems 13
1.5.1. Drug and vehicle interaction 15
1.5.2. Horny layer modification and removal 16
1.5.3. Electrical techniques 18
1.5.4. Nanocarriers for transdermal drug delivery 18
A. Nanoemulsions (NEs) 19
B. Solid lipid nanoparticles (SLNs) 19
C. Nanostructured lipid carriers (NLCs) 20
D. Niosomes 20
1.6. Lipid bilayer systems for transdermal drug delivery 21
1.6.1. Conventional liposomes (CLs) 21
1.6.2. Ethosomes (ESs) 22
1.6.3. Deformable liposomes (Transfersomes) 22
2. Objectives of the study 24
3. Materials and methods 26
3.1. Materials 26
3.2. Preperation of lipid carriers 26
3.2.1. Preperation of PPP-loaded conventional liposomes (CLs) 26
3.2.2. Preperation of PPP-loaded ethosomes (ESs) 27
3.2.3. Preperation of PPP-loaded transformer-ethosomes (TESs) 27
3.3. Characterization of PPP lipid carriers 29
3.3.1. Transmission Electron Microscopy (TEM) 29
3.3.2. Particle size, particle distribution and zeta potential measurement 29
3.3.3. Differential scanning calorimetry (DSC) 29
3.4. Encapsulation efficiency and loading capacity 33
3.5. Determination of the flexibility properties 33
3.6. In vitro drug release 34
3.7. Skin permeation study 34
3.7.1. PPP solubility 34
3.7.2. Artificial membrane (Strat-MTM) 35
3.7.3. Human cadaver skin 35
3.7.4. Cumulative amount 36
3.7.5. Confocal laser scattering microscopy (CLSM) 37
3.8. CLSM images of cross-sections of human skin 38
3.9. Cell viability 38
3.10. In vitro efficacy 39
3.11. Human efficacy 41
3.12. Stability studies 42
3.13. HPLC analysis and method validation 42
3.14. Statistical analysis 43
4. Results and discussion 44
4.1. Characterization of PPP-lipid carriers 44
4.1.1. Morphology by TEM 44
4.1.2. Paticle size, distribution, and surface charge by DLS 44
4.1.3. Entrapment efficiency and loading capacity 46
4.2. Thermal properties of PPP-lipid carriers 52
4.3. Flexibility of PPP-lipid carriers 55
4.4. In vitro drug release 58
4.5. Skin permeation of PPP-lipid carriers 58
4.5.1. Solubility of PPP 58
4.5.2. Artificial membrane (Strat-MTM) 61
4.5.3. Human cadaver skin 70
4.6. Skin penetration study using rhodamine 6G-loaded lipid carriers 74
4.7. Cell viability 78
4.8. In vitro anti-skin aging activity 78
4.9. Anti-aging effect of PPP-lipid carriers in human subjects 81
4.10. Stability 86
5. Conclusions 90
6. References 91
국문 초록 111
