Avaliação da estabilidade e permeação cutânea in vitro e ex vivo de ácido retinoico em emulsão e dispersão líquida para uso domiciliar e em peeling
Data
2018-04-26
Autores
Raminelli, Ana Claudia Pompeu 
Orientadores
Leonardi, Gislaine Ricci
Tipo
Tese de doutorado
Título da Revista
ISSN da Revista
Título de Volume
Resumo
O uso tópico do ácido retinoico (AR) em concentrações de 0,025, 0,05 e 0,1% é o tratamento padrão ouro para o fotoenvelhecimento. Porém, o seu emprego como agente para peeling químico superficial, em formulações com concentrações mais elevadas, necessita de mais estudos para comprovação de eficácia e segurança. Desta forma, este estudo propôs a avaliação da estabilidade de formulações com AR para uso domiciliar (0,05%) e em peeling (5,0%), compostas por diferentes veículos e antioxidantes, já que não há relatos na literatura, e estudo de permeação in vitro através de pele de orelha de suíno e ex vivo em pele de antebraço humano com diagnóstico de fotoenvelhecimento. Para tal, uma metodologia utilizando cromatografia líquida de alta eficiência com detecção no ultravioleta-visível (CLAE-UV-vis) foi desenvolvida e validada. O método foi seletivo, exato, preciso e responsivo para analisar a estabilidade das formulações. As formulações em creme de poliacrilamida foram as mais estáveis, apresentando teor de AR por 180 dias à temperatura de 40,0 ± 2,0 ºC. Além da estabilidade, a permeação in vitro através de células de difusão de Franz foi estudada, utilizando-se pele de orelha de suíno entre o compartimento doador e receptor. Para as formulações a 5,0%, o veículo alcoólico promoveu maior permeação do ativo em camadas mais profundas da epiderme e derme, inclusive sendo detectada concentração do AR na solução receptora. Quando avaliado para uso domiciliar, este veículo promoveu maior retenção do ativo no estrato córneo, sendo que a quantidade encontrada na epiderme e derme foi similar para o veículo emulsionado ou alcoólico. A permeação cutânea ex vivo do AR em emulsão foi avaliada através da quantificação em fragmentos de pele do antebraço humano que receberam a formulação a 5,0% em um antebraço e a preparação a 0,05% em outro. Para isso, outra metodologia foi desenvolvida e validada através de cromatografia líquida acoplada a espectrômetro de massas (LC-MS/MS), a qual mostrou-se sensível, seletiva, precisa e exata. As biópsias da região tratada e da área controle foram realizadas após a permanência das formulações por 6 horas. A maior concentração média de AR foi encontrada no estrato córneo através da aplicação do AR a 5,0%, sendo 5 vezes maior que a concentração média quantificada nos fragmentos com AR a 0,05%, responsável pela ação cutânea superficial. A análise estatística (p<0,05) apontou diferença significativa para as concentrações de AR contidas no estrato e na epiderme / derme. Neste tecido, a concentração de AR foi cerca de 2,5 vezes maior para a formulação a 5,0%, que para a formulação a 0,05%. Por fim, a comparação entre o tratamento da pele com AR a 5,0% por 6 horas e por 4 meses mostrou que o AR foi em torno de 5 vezes maior no tratamento por 6 horas. Porém, através destes resultados pode-se constatar que quando as voluntárias foram submetidas ao tratamento prolongado observou-se um acúmulo de AR nos tecidos cutâneos.
Topical use of retinoic acid (RA) is the gold standard treatment for photoaging, which is formulated at concentrations of 0.025, 0.05 and 0.1%. At the same time, the treatment as agent for superficial chemical peeling, with higher concentrations, requires further studies to evaluate efficacy and safety. Therefore, this study aimed to verify the chemical stability of formulations with RA for daily use (0.05%) and peeling (5.0%), with different antioxidants, since there are no reports of these studies in the literature, and study of permeation in vitro through porcine ear skin and ex vivo in human forearm skin diagnosed with photoaging. For that, a methodology using High Performance Liquid Chromatography with Ultraviolet detector (HPLC-UV) was developed and validated. The method was selective, accurate, and responsive to analyze the stability of the formulations. The polyacrylamide cream formulations were the most stable, having acceptable concentration of RA for 180 days at a temperature of 40.0 ± 2.0 ºC. In vitro permeation through Franz diffusion cells was studied using pig ear skin between the donor and recipient compartment. For the 5.0% formulations, the alcoholic vehicle promoted greater permeation of the drug in deeper layers of the epidermis and dermis, even being detected concentration of the drug in the receptor solution. When evaluated for daily use, this vehicle promoted greater retention of the RA in the stratum corneum, and the amount found in the epidermis and dermis was similar for the cream or alcoholic vehicle. Ex vivo skin permeation of the RA in emulsion was assessed by quantifying skin fragments of the human forearm which received the formulation at 5.0% in one forearm and the preparation at 0.05% in another. For this, another methodology was developed and validated by liquid chromatography coupled to mass spectrometer (LC-MS / MS), which was sensitive, selective, accurate and accurate. The biopsies of the treated region and the control area were performed after the formulations had remained for 6 hours. The highest mean RA concentration was found in the stratum corneum through the application of 5% RA, 5 times higher than the mean concentration quantified in the fragments with 0.05% RA, responsible by superficial skin action. Statistical analysis (p< 0.05) showed a significant difference for the concentrations of RA contained in the stratum and in the epidermis / dermis. In this tissue, the RA concentration was about 2.5-fold higher for the 5.0% formulation than for the 0.05% formulation. Finally, the comparison between skin treatment with 5.0% RA for 6 hours and 4 months have shown that RA was around 5 times greater in the treatment for 6 hours. However, through these results it can be seen that when the volunteers underwent the prolonged treatment an accumulation of RA in the cutaneous tissues was observed.
Topical use of retinoic acid (RA) is the gold standard treatment for photoaging, which is formulated at concentrations of 0.025, 0.05 and 0.1%. At the same time, the treatment as agent for superficial chemical peeling, with higher concentrations, requires further studies to evaluate efficacy and safety. Therefore, this study aimed to verify the chemical stability of formulations with RA for daily use (0.05%) and peeling (5.0%), with different antioxidants, since there are no reports of these studies in the literature, and study of permeation in vitro through porcine ear skin and ex vivo in human forearm skin diagnosed with photoaging. For that, a methodology using High Performance Liquid Chromatography with Ultraviolet detector (HPLC-UV) was developed and validated. The method was selective, accurate, and responsive to analyze the stability of the formulations. The polyacrylamide cream formulations were the most stable, having acceptable concentration of RA for 180 days at a temperature of 40.0 ± 2.0 ºC. In vitro permeation through Franz diffusion cells was studied using pig ear skin between the donor and recipient compartment. For the 5.0% formulations, the alcoholic vehicle promoted greater permeation of the drug in deeper layers of the epidermis and dermis, even being detected concentration of the drug in the receptor solution. When evaluated for daily use, this vehicle promoted greater retention of the RA in the stratum corneum, and the amount found in the epidermis and dermis was similar for the cream or alcoholic vehicle. Ex vivo skin permeation of the RA in emulsion was assessed by quantifying skin fragments of the human forearm which received the formulation at 5.0% in one forearm and the preparation at 0.05% in another. For this, another methodology was developed and validated by liquid chromatography coupled to mass spectrometer (LC-MS / MS), which was sensitive, selective, accurate and accurate. The biopsies of the treated region and the control area were performed after the formulations had remained for 6 hours. The highest mean RA concentration was found in the stratum corneum through the application of 5% RA, 5 times higher than the mean concentration quantified in the fragments with 0.05% RA, responsible by superficial skin action. Statistical analysis (p< 0.05) showed a significant difference for the concentrations of RA contained in the stratum and in the epidermis / dermis. In this tissue, the RA concentration was about 2.5-fold higher for the 5.0% formulation than for the 0.05% formulation. Finally, the comparison between skin treatment with 5.0% RA for 6 hours and 4 months have shown that RA was around 5 times greater in the treatment for 6 hours. However, through these results it can be seen that when the volunteers underwent the prolonged treatment an accumulation of RA in the cutaneous tissues was observed.
Descrição
Citação
RAMINELLI, Ana Cláudia Pompeu. Avaliação da estabilidade e permeação cutânea in vitro e ex vivo de ácido retinoico em emulsão e dispersão líquida para uso domiciliar e em peeling. 2018. 173 f. Tese (Doutorado em Medicina Translacional) - Escola Paulista de Medicina, Universidade Federal de São Paulo, São Paulo, 2018.