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Assessment of in vitro methodologies to determine topical and transdermal delivery of the flavonoid quercetin

VICENTINI, Fabiana Testa Moura de Carvalho; GEORGETTI, Sandra Regina; BENTLEY, Maria Vitória Lopes Badra; FONSECA, Maria José Vieira
Fonte: Universidade de São Paulo, Faculdade de Ciências Farmacêuticas Publicador: Universidade de São Paulo, Faculdade de Ciências Farmacêuticas
Tipo: Artigo de Revista Científica
ENG
Relevância na Pesquisa
75.78%
To be effective against the oxidative damages induced by UVB irradiation in the skin, the drug needs to release from the formulation in which it was incorporated and reach the skin layers where the ROS are generated. Thus, it is very important the development of a robust and sensitive methodology to extract and quantify in different skin layers the antioxidant agent delivered from topical formulations. Therefore, in the present work suitable methods to extract and quantify quercetin in skin samples and receptor phase after in vitro penetration studies were developed. The results demonstrated that the recovery from two different layers of skin, the SC and [E+D], using two different methods of quantification (DPPH• assay and HPLC, respectively), was 93.8 % when the quercetin spiked dose was 50 µg/mL, 100.4 % when it was 100 µg/mL and 89.9 % for 250 µg/mL and the average recovery of the quercetin extraction from receptor phase when dichloromethane was used as extractor solvent was 96%. These results demonstrate that the described methods have a potential application to in vitro skin penetration studies of quercetin, since it showed to be accurate and sensitive.; Para ser efetiva contra os danos oxidativos induzidos pela radiação UVB na pele...

Estabilidade e estudo de penetração cutânea in vitro da rutina veiculada em uma emulsão cosmética através de um modelo de biomembrana alternativo; Stability and in vitro penetration study of rutin incorporated in a cosmetic emulsion through an alternative model biomembrane

BABY, André Rolim; HAROUTIOUNIAN-FILHO, Carlos Alberto; SARRUF, Fernanda Daud; TAVANTE-JÚNIOR, Carlos Roberto; PINTO, Claudinéia Aparecida Sales de Oliveira; ZAGUE, Vivian; ARÊAS, Elizabeth Pinheiro Gomes; KANEKO, Telma Mary; VELASCO, Maria Valéria R
Fonte: Divisão de Biblioteca e Documentação do Conjunto das Químicas da Universidade de São Paulo Publicador: Divisão de Biblioteca e Documentação do Conjunto das Químicas da Universidade de São Paulo
Tipo: Artigo de Revista Científica
POR
Relevância na Pesquisa
75.9%
A rutina é empregada como antioxidante e na prevenção da fragilidade capilar. Estudos de penetração in vitro através da pele humana seria a situação ideal, entretanto, há dificuldades de sua obtenção e manutenção de sua viabilidade. Entre os demais modelos de membrana, a muda de pele de cobra se apresenta como estrato córneo puro, fornecendo barreira similar ao humano e é obtida sem a morte do animal. Os objetivos desta pesquisa foram desenvolver e avaliar a estabilidade de uma emulsão cosmética contendo rutina e, como promotor de penetração cutânea, o propilenoglicol; e avaliar a penetração e a retenção cutânea in vitro da referida substância ativa da formulação, empregando um modelo de biomembrana alternativo. A emulsão foi desenvolvida com rutina e propilenoglicol, ambos a 5,0% p/p. Quantificou-se a rutina das emulsões por espectrofotometria a 361,0 nm, método previamente validado. A penetração e retenção cutânea in vitro foram realizadas em células de difusão vertical com muda de pele de cobra de Crotalus durissus, como modelo de biomembrana alternativo, e água destilada e álcool etílico absoluto 99,5% (1:1), como fluido receptor. O experimento foi conduzido em um período de seis horas...

Fluorometric quantification of protoporphyrin IX in biological skin samples from in vitro penetration/permeation studies

ROSSETTI, Fábia Cristina; DEPIERI, Lívia Vieira; TEDESCO, Antônio Cláudio; BENTLEY, Maria Vitória Lopes Badra
Fonte: Universidade de São Paulo, Faculdade de Ciências Farmacêuticas Publicador: Universidade de São Paulo, Faculdade de Ciências Farmacêuticas
Tipo: Artigo de Revista Científica
ENG
Relevância na Pesquisa
85.83%
A fluorometric analytical method was developed for quantification of protoporphyrin IX (PpIX) in skin samples and receptor phase solution after in vitro cutaneous penetration/permeation studies. Analytical conditions used were: excitation and emission wavelengths: 400 nm and 632 nm; bandwidth: 0.5 nm; excitation and emission slits: 10/10. PpIX was recovered from two different layers of skin, the stratum corneum (SC) and the epidermis plus dermis ([E+D]), by vortex homogenization, probe and bath sonication, using DMSO as an extraction solvent. The detection and quantification limits were 0.002 and 0.005 μg/mL, respectively. The assay was linear from 0.005 - 0.5 μg/mL. The within-day and between-day assay precision and accuracy in DMSO and receptor phase solution were each studied at the two concentration levels 0.04 and 0.2 μg/mL, and 0.01 and 0.08 μg/mL, respectively. The coefficients of variation and deviation from the theoretical values were lower than 5%. The skin recovery of PpIX from SC and [E+D] layers using two different concentrations (0.5 and 1.0 μg/mL) were all above 90.0%. The method described has potential application to in vitro penetration/permeation studies of PpIX using porcine skin as a biological membrane model.; Um método analítico por espectrofluorimetria foi desenvolvido para quantificar a protoporfirina IX (Pp IX) em amostras de pele e fase receptora após a realização de testes in vitro de penetração/permeação cutâneas. As condições analíticas utilizadas foram: comprimentos de onda de excitação e emissão: 400 nm e 632 nm; largura de banda: 0...

Stability and in vitro penetration study of rutin incorporated in a cosmetic emulsion through an alternative model biomembrane

BABY, Andre Rolim; HAROUTIOUNIAN-FILHO, Carlos Alberto; SARRUF, Fernanda Daud; TAVANTE JR., Carlos Roberto; PINTO, Claudineia Aparecida Sales de Oliveira; ZAGUE, Vivian; AREAS, Elizabeth Pinheiro Gomes; KANEKO, Telma Mary; VELASCO, Maria Valeria Robles
Fonte: UNIV SAO PAULO, CONJUNTO QUIMICAS Publicador: UNIV SAO PAULO, CONJUNTO QUIMICAS
Tipo: Artigo de Revista Científica
POR
Relevância na Pesquisa
75.87%
Rutin is employed as antioxidant and to prevent the capillary fragility and, when incorporated in cosmetic emulsions, it must target the action site. In vitro cutaneous penetration studies through human skin is the ideal situation, however, there are difficulties to obtain and to maintain this tissue viability. Among the membrane models, shed snake skin presents itself as pure stratum corneum, providing barrier function similar to human and it is obtained without the animal sacrifice. The objectives of this research were the development and stability evaluation of a cosmetic emulsion containing rutin and propylene glycol (penetration enhancer) and the evaluation or rutin in vitro cutaneous penetration and retention from the emulsion, employing an alternative model biomembrane. Emulsion was developed with rutin and propylene glycol, both at 5.0% w/w. Active substance presented on the formulation was quantified by a validated spectrophotometric method at 361.0 nm. Rutin Rutin cutaneous penetration and retention was performed in vertical diffusion cells with shed snake skin of Crotalus durissus, as alternative model biomembrane, and distilled water and ethanol 99.5% (1:1), as receptor fluid. The experiment was conducted for six hours...

Development and validation of HPLC method for imiquimod determination in skin penetration studies

PAULA, Daniel De; MARTINS, Carolina Azenha; BENTLEY, Maria Vitoria Lopes Badra
Fonte: JOHN WILEY & SONS LTD Publicador: JOHN WILEY & SONS LTD
Tipo: Artigo de Revista Científica
ENG
Relevância na Pesquisa
75.78%
A simple, rapid and sensitive analytical procedure for the measurement of imiquimod in skin samples after in vitro penetration studies has been developed and validated. In vitro penetration studies were carried out in Franz diffusion cells with porcine skin. Tape stripping technique was used to separate the stratum corneum (SC) from the viable epidermis and dermis. Imiquimod was extracted from skin samples using a 7:3 (v/v) methanol:acetate buffer (100 mm, pH 4.0) solution and ultrasonication. Imiquimod was analyzed by H-PLC using C(8) column and UV detection at 242 ran. The mobile phase used was acetonitrile:acetate buffer (pH 4.0, 100 mM):diethylamine (30:69.85:0.15, v/v) with flow rate 1 mL/min. Imiquimod eluted at 4.1 min and the running time was limited to 6.0 min. The procedure was linear across the following concentration ranges: 100-2500 ng/mL for both SC and tape-stripped skin and 20-800 ng/mL for receptor solution. Intra-day and inter-day accuracy and precision values were lower than 20% at the limit of quantitation. The recovery values ranged from 80 to 100%. The method is adequate to assay imiquimod from skin samples, enabling the determination of the cutaneous penetration profile of uniquimod by in vitro studies. Copyright (C) 2008 John Wiley & Sons...

Liberação e permeação in vitro de produtos transdérmicos: um estudo metodológico de aparatos e de condições experimentais; In vitro release and permeation transdermal products: evaluation of methods and apparatus

Praça, Fabíola Silva Garcia
Fonte: Biblioteca Digitais de Teses e Dissertações da USP Publicador: Biblioteca Digitais de Teses e Dissertações da USP
Tipo: Tese de Doutorado Formato: application/pdf
Publicado em 24/08/2010 PT
Relevância na Pesquisa
75.62%
Liberação transdérmica de fármacos apresenta várias vantagens na terapêutica quando comparada com administração oral ou parenteral. Não existe até o momento nenhum método previsto na Farmacopéia Brasileira para realizar testes de liberação de fármacos em adesivos transdérmicos, outros compêndios oficiais como Farmacopéia Americana, Britânica e Européia, descrevem o aparato de pás sobre disco, o cilindro rotatório e o suporte recíproco. Atualmente a literatura descreve diversos tipos de células de difusão para liberação transdérmica das quais a célula de difusão de Franz tem sido a mais empregada tanto para adesivos transdérmicos como formas semi-sólidas, utilizada no desenvolvimento farmacotécnico, caracterização biofarmacêutica e controle de qualidade. A proposta deste estudo tem por objetivo estipular critérios para a escolha mais adequada do equipamento e metodologias in vitro na avaliação da liberação transdérmicas de fármacos, utilizando a nicotina como fármaco modelo. Para tal, foram empregados ensaios in vitro de liberação e retenção cutânea utilizando métodos de pás sobre disco e método de célula de difusão de Franz modificada em sistema estático e fluxo contínuo. A validação dos fatores que influenciam a taxa de liberação in vitro da nicotina foram fundamentais para escolha do meio receptor...

Avaliação in vitro da permeabilidade cutânea da rutina em emulsões cosméticas; In vitro evaluation of rutin cutaneous permeability from cosmetic emulsions

Baby, André Rolim
Fonte: Biblioteca Digitais de Teses e Dissertações da USP Publicador: Biblioteca Digitais de Teses e Dissertações da USP
Tipo: Tese de Doutorado Formato: application/pdf
Publicado em 10/09/2007 PT
Relevância na Pesquisa
65.9%
A rutina é empregada como antioxidante e na prevenção da fragilidade capilar. Pode ser veiculada em emulsões tópicas adequadas para atingir o local de ação. Estudos de penetração in vitro através da pele humana seria a situação ideal, entretanto, há dificuldades de sua obtenção e manutenção de sua viabilidade. Entre os demais modelos de membrana, a muda de pele de cobra apresenta-se como estrato córneo puro, fornecendo barreira similar ao humano e é obtida sem a morte do animal. Os objetivos desta pesquisa foram: (1) desenvolver e avaliar a estabilidade de emulsões cosméticas, contendo rutina e promotores de penetração cutânea, tais como, uréia (U), isopropanol (IP) e propilenoglicol (PG); (2) avaliar a liberação da referida substância ativa das emulsões e; (3) avaliar a penetração e a retenção cutânea in vitro da rutina da formulação de melhor desempenho. Emulsões foram desenvolvidas com rutina a 5,0% p/p e U, IP e PG, associados ou não e em proporções distintas, segundo planejamento fatorial com dois níveis com ponto central. Quantificou-se a rutina das emulsões por espectrofotometria a 361,0 nm, método previamente validado. A liberação da rutina nas formulações foi realizada em células de difusão vertical com membrana de acetato de celulose e água destilada e álcool etílico absoluto 99...

Avaliação da penetração cutânea de nanocápsulas de isotretinoína por tape stripping in vitro em pele humana e suína; Assessment of cutaneous penetration of isotretinoin-loaded nanocapsules by tape stripping in vitro in human and pig skin

Bettoni, Clarissa Cassini
Fonte: Universidade Federal do Rio Grande do Sul Publicador: Universidade Federal do Rio Grande do Sul
Tipo: Dissertação Formato: application/pdf
POR
Relevância na Pesquisa
65.91%
Objetivos: objetivo geral deste trabalho foi avaliar a penetração cutânea da isotretinoína nanoencapsulada e livre utilizando técnicas de microdiálise e tape stripping in vitro. Métodos: A viabilidade da utilização da técnica de microdiálise para avaliar o perfil farmacocinético da isotretinoína após aplicação tópica foi investigada através da determinação da recuperação relativa (RR) in vitro por diálise e retrodiálise. A influência da concentração, do fluxo de perfusão e a ligação do fármaco à tubulação das sondas de microdiálise foram investigadas. A metodologia analítica para quantificação do fármaco em microdialisado foi validada. Em seguida, as penetrações cutâneas da isotretinoína livre e nanoencapsulada incorporada em géis hidrofílicos foram comparadas através da técnica de tape stripping in vitro em células de difusão de Franz utilizando pele humana e pele de porco. Para garantir a integridade das formulações, a estabilidade físico-química das mesmas foi avaliada. Os resultados de penetração cutânea foram comparados com os resultados in vivo obtidos em trabalho prévio do grupo de pesquisa. Resultados e Conclusões: Um método analítico simples e rápido para a determinação de isotretinoína em microdialisado foi validado de acordo com o FDA. A RR mostrou-se concentração independente e observou-se que há diferenças significativas entre as RR avaliadas pelos dois métodos utilizados...

Cyclosporin A in skin samples from in vitro penetration studies may be assayed by a simple HPLC method

Lopes,Luciana B.; Bentley,Maria Vitória Lopes Brada
Fonte: Divisão de Biblioteca e Documentação do Conjunto das Químicas da Universidade de São Paulo Publicador: Divisão de Biblioteca e Documentação do Conjunto das Químicas da Universidade de São Paulo
Tipo: Artigo de Revista Científica Formato: text/html
Publicado em 01/12/2005 EN
Relevância na Pesquisa
95.78%
A simple High Performance Liquid Chromatographic method with UV detection was developed for quantification of Cyclosporin A (CysA) in skin samples after in vitro penetration studies. The peptide was recovered from two different layers of skin, the stratum corneum and the epidermis plus dermis ([E+D]), by vortex homogenization and bath sonication in an organic solvent (methanol). Recovery of CysA from skin was about 85%, and CysA was estimated by HPLC using a RP-18 column, UV detection at 210 nm and acetonitrile:water (67:33 v/v) as the mobile phase. The quantification limit was 150ng/mL. The assay was linear from 0.15-500 mg/muL. The within-day and between-day assay precision and accuracy were studied at three concentration levels (1, 10 and 20 mug/mL). The coefficients of variation and deviation from the theoretical values were lower than 5%. The method described has a potential application to in vitro penetration studies of CysA using porcine skin as a biological membrane model.

Assessment of in vitro methodologies to determine topical and transdermal delivery of the flavonoid quercetin

Vicentini,Fabiana Testa Moura de Carvalho; Georgetti,Sandra Regina; Bentley,Maria Vitória Lopes Badra; Fonseca,Maria José Vieira
Fonte: Universidade de São Paulo, Faculdade de Ciências Farmacêuticas Publicador: Universidade de São Paulo, Faculdade de Ciências Farmacêuticas
Tipo: Artigo de Revista Científica Formato: text/html
Publicado em 01/06/2009 EN
Relevância na Pesquisa
75.75%
To be effective against the oxidative damages induced by UVB irradiation in the skin, the drug needs to release from the formulation in which it was incorporated and reach the skin layers where the ROS are generated. Thus, it is very important the development of a robust and sensitive methodology to extract and quantify in different skin layers the antioxidant agent delivered from topical formulations. Therefore, in the present work suitable methods to extract and quantify quercetin in skin samples and receptor phase after in vitro penetration studies were developed. The results demonstrated that the recovery from two different layers of skin, the SC and [E+D], using two different methods of quantification (DPPH• assay and HPLC, respectively), was 93.8 % when the quercetin spiked dose was 50 µg/mL, 100.4 % when it was 100 µg/mL and 89.9 % for 250 µg/mL and the average recovery of the quercetin extraction from receptor phase when dichloromethane was used as extractor solvent was 96%. These results demonstrate that the described methods have a potential application to in vitro skin penetration studies of quercetin, since it showed to be accurate and sensitive.

Fluorometric quantification of protoporphyrin IX in biological skin samples from in vitro penetration/permeation studies

Rossetti,Fábia Cristina; Depieri,Lívia Vieira; Tedesco,Antônio Cláudio; Bentley,Maria Vitória Lopes Badra
Fonte: Universidade de São Paulo, Faculdade de Ciências Farmacêuticas Publicador: Universidade de São Paulo, Faculdade de Ciências Farmacêuticas
Tipo: Artigo de Revista Científica Formato: text/html
Publicado em 01/12/2010 EN
Relevância na Pesquisa
85.8%
A fluorometric analytical method was developed for quantification of protoporphyrin IX (PpIX) in skin samples and receptor phase solution after in vitro cutaneous penetration/permeation studies. Analytical conditions used were: excitation and emission wavelengths: 400 nm and 632 nm; bandwidth: 0.5 nm; excitation and emission slits: 10/10. PpIX was recovered from two different layers of skin, the stratum corneum (SC) and the epidermis plus dermis ([E+D]), by vortex homogenization, probe and bath sonication, using DMSO as an extraction solvent. The detection and quantification limits were 0.002 and 0.005 μg/mL, respectively. The assay was linear from 0.005 - 0.5 μg/mL. The within-day and between-day assay precision and accuracy in DMSO and receptor phase solution were each studied at the two concentration levels 0.04 and 0.2 μg/mL, and 0.01 and 0.08 μg/mL, respectively. The coefficients of variation and deviation from the theoretical values were lower than 5%. The skin recovery of PpIX from SC and [E+D] layers using two different concentrations (0.5 and 1.0 μg/mL) were all above 90.0%. The method described has potential application to in vitro penetration/permeation studies of PpIX using porcine skin as a biological membrane model.

In vitro skin penetration of clobetasol from lipid nanoparticles: drug extraction and quantitation in different skin layers

Silva,Luís Antônio Dantas; Taveira,Stephânia Fleury; Lima,Eliana Martins; Marreto,Ricardo Neves
Fonte: Universidade de São Paulo, Faculdade de Ciências Farmacêuticas Publicador: Universidade de São Paulo, Faculdade de Ciências Farmacêuticas
Tipo: Artigo de Revista Científica Formato: text/html
Publicado em 01/12/2012 EN
Relevância na Pesquisa
75.74%
Clobetasol propionate (CP) is a potent topical corticosteroid that causes several cutaneous and systemic side effects. In the present work, CP was encapsulated in nanostructured lipid carriers (NLCs) to increase drug retention in the outer skin layers and improve the safety of topical therapy. NLCs were prepared using a microemulsion technique with a mixture of lecithin, taurodeoxycholate, stearic acid, and oleic acid. In vitro penetration studies were performed in a modified Franz-type diffusion cell, and porcine ears were used as a model of human skin. A simple and sensitive liquid chromatographic method was developed and validated for clobetasol determination in different skin layers. NLCs presented uniform size distribution, high zeta potentialand entrapment efficiency values (> 98%). The analytical procedure was validated according to FDA guidelines. Clobetasol recoveries from skin samples were higher than 85%, with no interference of skin components and NLC ingredients. In experiments, after 6 h, a higher drug accumulation in the stratum corneum arising from NLCs compared to aqueous CP solution was observed. Thus, the NLCs demonstrated high potential for targeting CP to the skin and ensuring drug accumulation in the stratum corneum.

In vitro penetration of fresh and vitrified swine oocytes by homologous spermatozoa using different incubation systems

Macedo J??nior, Milton C.; Deschamps, Jo??o Carlos; Lucia J??nior, Thomaz; Bordignon, Jonas; Serret, Carolina Gon??alves; Rambo, Gissele; Pivato, Ivo; Schmitt, Eduardo
Fonte: Elsevier Publicador: Elsevier
Tipo: Artigo de Revista Científica
EN_US
Relevância na Pesquisa
95.72%
The present study consisted of two experiments. In the first one, ejaculates from four boars were used to compare in vitro penetration (IVP) rates of fresh and vitrified swine oocytes by homologous spermatozoa in four treatments: fresh oocytes in conventional incubation (CO2 incubator) (FC), vitrified oocytes in conventional incubation (VC), fresh oocytes in submarine (bag) incubation (FS) and vitrified oocytes in submarine incubation (VS). The IVP rates for FC, VC, FS and VS were 46.5, 44.3, 36.9 and 33.1%, respectively. Analysis through Chi-square tests identified no differences in IVP rates between FC and VC and between FS and VS (P > 0.05), but IVP rate for FC was greater (P < 0.05) than those for both FS and VS. Besides IVP rate for VC did not differ (P > 0.05) from those for FC and FS, but it was greater than that for VS (P < 0.05). Logistic regression analysis identified differential effects of treatments dependant on individual boars. The second experiment evaluated the influence of semen storage period on the semen quality of the two boars associated with greater IVP rate in the first experiment. Semen quality was estimated by IVP rate using the VC treatment and by the following methods: sperm motility, sperm morphology...

Teste de penetra????o esperm??tica em o??citos in vitro e fertilidade in vivoap??sinsemina????o heterosp??rmica em su??nos; In vitro penetration test using oocytes and in vivo fertility after heterospermic insemination in swine

MACEDO JUNIOR, Milton Carvalho
Fonte: Universidade Federal de Pelotas; Biotecnologia; Programa de P??s-Gradua????o em Biotecnologia; UFPel; BR Publicador: Universidade Federal de Pelotas; Biotecnologia; Programa de P??s-Gradua????o em Biotecnologia; UFPel; BR
Tipo: Tese de Doutorado Formato: application/pdf
POR
Relevância na Pesquisa
95.78%
The knowledge of the reproductive potential of a male is of economic and reproductive importance, mainly if used in programs of artificial insemination. The conventional tests for semen quality evaluation do not have the capacity to measure the fertilizing potential of a sample, although they indicate if that one is fertile or not. The in vitro penetration test, appears in this context as an alternative laboratorial test to categorize of males capacity of fecundation, as mimics in vitro what happens in vivo. However this test has its use limited for for difficult execution and high cost. With the objective to simplify the test, the present work evaluated alternatives for reduction of the execution time, use of cryopreserved oocytes in different methods and its association with in vivo fertility through heterospermic insemination and following diagnostic fertility. It was concluded that: 1) It is possible to use the incubation system BOTTLE, to replace the conventional incubation system, eliminating the need for using expansive CO2 incubator; 2) Oocytes can be cryopreserved in CRIOVIAL, together with the medium of fecundation and mineral oil, eliminating the use of stereoscopic lupa to manipulate oocytes in the day of the execution of the test; 3) It is viable to reduce co-incubation time of oocytes and sperm for 6 hours...

Development of in vitro embryo production systems for red deer (Cervus elaphus) - Part 1. Effect of epithelial oviductal monolayers and heparin on in vitro sperm motility and penetration of in vitro matured oocytes

Berg, D.; Thompson, J.; Asher, G.
Fonte: Elsevier Science BV Publicador: Elsevier Science BV
Tipo: Artigo de Revista Científica
Publicado em //2002 EN
Relevância na Pesquisa
65.88%
In vitro fertilisation (IVF) protocols for red deer have yielded low fertilisation rates, with no embryo development beyond the eight-cell stage when heparin was used as the in vitro capacitation agent. As this low fertilisation rate may result from reduced motility, the present study investigated the use of red deer oviduct epithelial cell monolayers (COEM) and conditioned medium (Cm) from the monolayers to maintain red deer sperm motility in vitro. A second experiment compared the fertilisability of red deer sperm pre-incubated for 4-12h on COEM or for 4h in TALP medium supplemented with 20 microg of heparin.COEM was superior in maintaining red deer sperm motility compared with either Sp-TALP alone or Cm (P<0.05). COEM sustained sperm motility at levels comparable to the initial motility over the 24h period. The motility of sperm incubated in Sp-TALP and Cm was similar and had declined to less than 10% by 4h and no motile sperm were observed by 8h. Overall, the penetration rates of in vitro red deer oocytes were low (5-28%) regardless of sperm treatment. Sperm pre-incubated on COEM penetrated more oocytes than sperm incubated with heparin (P<0.001). Penetration rates were similar for 4-12h pre-incubation of sperm on COEM (P>0.50). Penetration rates were greater across all treatments when both sperm and oocytes were co-incubated for 24h compared to 12h (P<0.001). There were no differences in penetration rates among the four donor stags used in the study. It was concluded that COEM sustains red deer sperm motility in vitro during the 24h observation period. Pre-incubating sperm on COEM does increase sperm penetration rates compared with heparin alone...

In vitro penetration of swine oocytes by homologous spermatozoa: Distinct systems for gamete’s co-incubation and oocyte’s cryopreservation

Macedo Júnior, Milton C.; Lucia Júnior, Thomaz; Rambo, Gissele; Ferreira Filho, E. B.; Rosa, A. P.; Fabiane, C.; Cabral, M.; Deschamps, João Carlos
Fonte: Elsevier Publicador: Elsevier
Tipo: Artigo de Revista Científica
EN_US
Relevância na Pesquisa
95.72%
In vitro penetration (IVP) of swine oocytes by homologous spermatozoa was evaluated in two experiments using four boars as semen donors. In experiment 1, the IVP rate and the number of penetrating spermatozoa (PSP) were compared using three co-incubation systems for vitrified oocytes and fresh sperm: (1) 35mL petri dishes in a CO2 incubator, (2) 35mL petri dishes in bags (submarine system) and (3) glass flasks partially submerged in water bath with the same gas mixture used for the bag system. Mean PSPwas 8.2±10.1 and the IVP ratewas 90.5%. The PSP differed across all systems (P = 0.0006): 15.5±0.5 for flasks, 6.3±0.4 for CO2, and 3.9±0.4 for bags. The IVP rate for flasks (95.0%) was greater (P = 0.01) than for CO2 and bags (90.8% and 85.0%, respectively), but it did not differ between flasks and CO2 for three boars (P > 0.05). In experiment 2, co-incubation was done as described for glass flasks in experiment 1. The IVP rate and PSP were compared for cryopreserved oocytes: either vitrified in open pulled straws (OPS), or frozen in cryotubes. Mean PSP was 5.4±6.5 and IVP ratewas 89.6%. Both PSP and IVP ratewere greater (P < 0.0001) for oocytes frozen in cryotubes (7.0±0.3% and 95.8%, respectively) than those frozen in OPS (3.7±0.3% and 83.4%...

Fluorometric quantification of protoporphyrin IX in biological skin samples from in vitro penetration/permeation studies

Rossetti, Fábia Cristina; Depieri, Lívia Vieira; Tedesco, Antônio Cláudio; Bentley, Maria Vitória Lopes Badra
Fonte: Universidade de São Paulo. Faculdade de Ciências Farmacêuticas Publicador: Universidade de São Paulo. Faculdade de Ciências Farmacêuticas
Tipo: info:eu-repo/semantics/article; info:eu-repo/semantics/publishedVersion; ; ; ; ; ; Formato: application/pdf
Publicado em 01/12/2010 ENG
Relevância na Pesquisa
75.81%
Um método analítico por espectrofluorimetria foi desenvolvido para quantificar a protoporfirina IX (Pp IX) em amostras de pele e fase receptora após a realização de testes in vitro de penetração/permeação cutâneas. As condições analíticas utilizadas foram: comprimentos de onda de excitação e emissão: 400 nm e 632 nm; largura de banda: 0,5 nm; fendas de excitação e emissão: 10/10. A PpIX foi extraída de amostras de estrato córneo (EC) e da epiderme sem estrato córneo + derme ([E+D]) através da agitação em vórtex e sonicação por haste e banho, utilizando-se o DMSO como solvente extrator. O limite de detecção e quantificação foram, respectivamente, de 0,002 e 0,005 μg/mL. O método mostrou-se linear da faixa de 0,005 - 0,5 μg/mL. A precisão e exatidão intra e inter-ensaio em DMSO e na fase receptora foram validadas utilizando-se duas concentrações distintas, respectivamente, de 0,004 e 0,2 μg/mL, e 0,01 e 0,08 μg/mL. Os valores de coeficiente de variação e o desvio do valor teórico foram inferiores a 5%. A recuperação da PpIX das camadas da pele (EC e [E+D]) utilizando-se duas concentrações distintas (0,5 e 1,0 μg/mL) foram todas acima de 90,0%. O método descrito pode ser utilizado para determinação da PpIX após estudos de penetração/permeação cutânea in vitro utilizando pele de porco como modelo de membrana.; A fluorometric analytical method was developed for quantification of protoporphyrin IX (PpIX) in skin samples and receptor phase solution after in vitro cutaneous penetration/permeation studies. Analytical conditions used were: excitation and emission wavelengths: 400 nm and 632 nm; bandwidth: 0.5 nm; excitation and emission slits: 10/10. PpIX was recovered from two different layers of skin...

Assessment of in vitro methodologies to determine topical and transdermal delivery of the flavonoid quercetin

Vicentini, Fabiana Testa Moura de Carvalho; Georgetti, Sandra Regina; Bentley, Maria Vitória Lopes Badra; Fonseca, Maria José Vieira
Fonte: Universidade de São Paulo. Faculdade de Ciências Farmacêuticas Publicador: Universidade de São Paulo. Faculdade de Ciências Farmacêuticas
Tipo: info:eu-repo/semantics/article; info:eu-repo/semantics/publishedVersion; ; ; ; ; ; Formato: application/pdf
Publicado em 01/06/2009 ENG
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Para ser efetiva contra os danos oxidativos induzidos pela radiação UVB na pele, é necessário que o ativo seja liberado da formulação na qual foi incorporado e alcance as camadas da pele onde são geradas as EROS. Desta forma, torna-se de grande importância o desenvolvimento de métodos eficazes e sensíveis para extrair e quantificar, nas diferentes camadas de pele, o agente antioxidante liberado de formulações tópicas. No presente trabalho foram desenvolvidos métodos adequados para extrair e quantificar a quercetina em amostras de pele e na fase receptora após estudos de penetração cutânea in vitro. Os resultados demonstraram que a recuperação das camadas de pele, EC e [E+D], quando do uso de duas diferentes metodologias de quantificação (ensaio de DPPH• e CLAE, respectivamente), foi de 93,8 % quando aplicada uma dose de 50 µg/mL de quercetina, 100,4 % para 100 µg/mL e 89,9 % para 250 µg/mL e a recuperação média da extração da quercetina da fase receptora, quando do emprego de diclorometano como solvente extrator, foi de 96 %. Tais resultados demonstram que os métodos descritos têm grande potencial de aplicação em estudos de penetração in vitro já que apresentaram exatidão e sensibilidade.; To be effective against the oxidative damages induced by UVB irradiation in the skin...

Cyclosporin A in skin samples from in vitro penetration studies may be assayed by a simple HPLC method; Análise por CLAE da ciclosporina contida em amostras de pele provenientes de estudos de penetração cutânea in vitro

Lopes, Luciana B.; Bentley, Maria Vitória Lopes Brada
Fonte: Universidade de São Paulo. Faculdade de Ciências Farmacêuticas Publicador: Universidade de São Paulo. Faculdade de Ciências Farmacêuticas
Tipo: info:eu-repo/semantics/article; info:eu-repo/semantics/publishedVersion; Artigo Avaliado pelos Pares Formato: application/pdf
Publicado em 01/12/2005 ENG
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95.82%
A simple High Performance Liquid Chromatographic method with UV detection was developed for quantification of Cyclosporin A (CysA) in skin samples after in vitro penetration studies. The peptide was recovered from two different layers of skin, the stratum corneum and the epidermis plus dermis ([E+D]), by vortex homogenization and bath sonication in an organic solvent (methanol). Recovery of CysA from skin was about 85%, and CysA was estimated by HPLC using a RP-18 column, UV detection at 210 nm and acetonitrile:water (67:33 v/v) as the mobile phase. The quantification limit was 150ng/mL. The assay was linear from 0.15-500 mg/muL. The within-day and between-day assay precision and accuracy were studied at three concentration levels (1, 10 and 20 mug/mL). The coefficients of variation and deviation from the theoretical values were lower than 5%. The method described has a potential application to in vitro penetration studies of CysA using porcine skin as a biological membrane model.; Neste estudo foi desenvolvido um método simples utilizando Cromatografia Líquida de Alta Eficiência (CLAE) com detecção no UV para quantificação de Ciclosporina A (CysA) em amostras de pele após estudos de penetração cutânea in vitro. O peptídeo foi extraído do estrato córneo e da epiderme sem estrato córneo + derme ([E+D]) através de agitação em vórtex e sonicação utilizando um solvente extrator (metanol). A recuperação da CysA de ambas as camadas da pele foi de 85%...

Stability and in vitro penetration study of rutin incorporated in a cosmetic emulsion through an alternative model biomembrane; Estabilidade e estudo de penetração cutânea in vitro da rutina veiculada em uma emulsão cosmética através de um modelo de biomembrana alternativo

Baby, André Rolim; Haroutiounian-Filho, Carlos Alberto; Sarruf, Fernanda Daud; Tavante-Júnior, Carlos Roberto; Pinto, Claudinéia Aparecida Sales de Oliveira; Zague, Vivian; Arêas, Elizabeth Pinheiro Gomes; Kaneko, Telma Mary; Velasco, Maria Valéria R
Fonte: Universidade de São Paulo. Faculdade de Ciências Farmacêuticas Publicador: Universidade de São Paulo. Faculdade de Ciências Farmacêuticas
Tipo: info:eu-repo/semantics/article; info:eu-repo/semantics/publishedVersion; Artigo Avaliado pelos Pares Formato: application/pdf
Publicado em 01/06/2008 POR
Relevância na Pesquisa
65.9%
Rutin is employed as antioxidant and to prevent the capillary fragility and, when incorporated in cosmetic emulsions, it must target the action site. In vitro cutaneous penetration studies through human skin is the ideal situation, however, there are difficulties to obtain and to maintain this tissue viability. Among the membrane models, shed snake skin presents itself as pure stratum corneum, providing barrier function similar to human and it is obtained without the animal sacrifice. The objectives of this research were the development and stability evaluation of a cosmetic emulsion containing rutin and propylene glycol (penetration enhancer) and the evaluation of rutin in vitro cutaneous penetration and retention from the emulsion, employing an alternative model biomembrane. Emulsion was developed with rutin and propylene glycol, both at 5.0% w/w. Active substance presented on the formulation was quantified by a validated spectrophotometric method at 361.0 nm. Rutin cutaneous penetration and retention was performed in vertical diffusion cells with shed snake skin of Crotalus durissus, as alternative model biomembrane, and distilled water and ethanol 99.5% (1:1), as receptor fluid. The experiment was conducted for six hours, at 37.0 ± 0.5 ºC with constant stirring of 300 rpm. Spectrophotometry at 410.0 nm...