Journal of Chromatography B, 947–948 (2014) 23–31
Contents lists available at ScienceDirect
Journal of Chromatography B journal homepage: www.elsevier.com/locate/chromb
In vitro drug release and ex vivo percutaneous absorption of resveratrol cream using HPLC with zirconized silica stationary phase Hudson Caetano Polonini a,∗ , Carina de Almeida Bastos b , Marcone Augusto Leal de Oliveira b , Carla Grazieli Azevedo da Silva c , Carol Hollingworth Collins c , Marcos Antônio Fernandes Brandão a , Nádia Rezende Barbosa Raposo a a
Núcleo de Pesquisa e Inovac¸ão em Ciências da Saúde (NUPICS), Federal University of Juiz de Fora, 36036-900 Juiz de Fora, MG, Brazil Grupo de Química Analítica e Quimiometria (GQAQ), Federal University of Juiz de Fora, 36036-900 Juiz de Fora, MG, Brazil c Institute of Chemistry, University of Campinas, PO Box 6154, 13083-970 Campinas, SP, Brazil b
a r t i c l e
i n f o
Article history: Received 6 September 2013 Received in revised form 2 December 2013 Accepted 3 December 2013 Available online 17 December 2013 Keywords: In vitro drug release Percutaneous absorption Transdermal dosage form Resveratrol Factorial design Validation studies
a b s t r a c t Since the designs of optimal formulations for resveratrol permeation via the skin are lacking, the aim of this study was to establish the profile of resveratrol permeability into and across human skin. For that, a laboratory-made chromatographic column was used (Zr-PMODS), with its performance being compared to a traditional C18 column. In vitro drug release was conducted with polysulfone membranes, and the flux (JS ) was 30.49 g cm−2 h−1 ), with a lag time (LT ) of 0.04 h, following a pseudo-first-order kinetics. For ex vivo percutaneous absorption using excised female human skin, the kinetic profile was the same, but JS was 0.87 g cm−2 h−1 and LT was 0.97 h. From the initials 49.30 g applied to the skin, 9.50 g were quantified in the receptor medium, 20.48 g was retained at the stratum corneum (do not account as permeated) and 21.41 g was retained at the viable epidermis + dermis (account as permeated), totalizing 30.90 g of resveratrol permeated after 24 h of application (62.6%). From these results, one can conclude that a person using the 1-g emulsion dose released by the pump containing 20 mg of resveratrol will have, theoretically, 12.53 mg of it liberated into his bloodstream, gradually and continuously for 24 h. © 2013 Elsevier B.V. All rights reserved.
1. Introduction Resveratrol (3,5,4-trihydroxystilbene) (Fig. 1) is a naturally occurring non-flavanoid phenolic compound produced by some spermatophytes, such as grapes [1]. It is found in high concentrations in many red grape skins and their wines [2], with the trans-resveratrol isomer being more biologically active [3]. The substance has been reported to possess antioxidant, neuroprotective, antiphotoaging and antiviral activities, and it seems that it also plays a role in the prevention and reduction of pathological processes such as inflammation, cancer and heart diseases [4]. However, it has poor oral bioavailability, which creates a dilemma between its great in vitro efficacy and its low in vivo effect [5], mainly because it is extensively metabolized in the body [6].
∗ Corresponding author at: Federal University of Juiz de Fora, Rua José Lourenc¸o Kelmer, s/n, Campus Universitário, 36036-900, Juiz de Fora, MG, Brazil. Tel.: +55 32 21023809; fax: +55 32 21023809. E-mail addresses: [email protected] (H.C. Polonini), [email protected] (M.A.L.d. Oliveira), [email protected] (M.A.F. Brandão), [email protected] (N.R.B. Raposo). 1570-0232/$ – see front matter © 2013 Elsevier B.V. All rights reserved. http://dx.doi.org/10.1016/j.jchromb.2013.12.005
On the other hand, topically applied resveratrol has been shown to possess more pronounced properties, namely: it is antiproliferative and chemopreventive against skin carcinogenesis [7]; gives sunprotection against skin damage from ultraviolet B (UVB) exposure [8]; is antimicrobial against dermatophytes and herpes simplex virus [1,9]; and activates estrogen receptors [10]. Thus, skin delivery of resveratrol is paramount for the effective insertion of the substance into pharmaceuticals, either topical or transdermal. According to literature reports, the stationary phase most commonly used for resveratrol analysis is the octadecyl-bonded silica (C18), whether in nutraceuticals [11], grape fruits [12], wines [13] or biological samples [14]. However, to the best of the authors’ knowledge transdermal emulsions containing resveratrol for human use have not been investigated so far, although one previous study have reported encouraging data for solutions and hydrogels [15]. Since the designs of optimal formulations for resveratrol permeation via the skin are lacking [15], the aim of this study was to establish the profile of resveratrol permeability into and across human skin. For that, a laboratory-made chromatographic column was used (Zr-PMODS), with its performance being compared to a commercial C18 column.
24
H.C. Polonini et al. / J. Chromatogr. B 947–948 (2014) 23–31
Fig. 1. Resveratrol chemical structure.
2. Materials and methods 2.1. Reagents, standard and materials The acetonitrile (ACN) used in the preparation of the mobile phase was HPLC grade, and sodium chloride (NaCl), calcium chloride (CaCl2 ), magnesium sulfate (MgSO4 ), and potassium dihydrogen phosphate (KH2 PO4 ) were of analytical grade, all from Sigma-Aldrich (USA). Ultrapure water (H2 O) obtained in an AquaMax-Ultra 370 Series (Young Lin, Korea) (18.2 M cm resistivity at 25 ◦ C and 1100 m) because in vivo it is vascularized, and thus the drug within it is able to reach the bloodstream [26]. As ex vivo skin has its microcirculation obliterated, the dermis can retain compounds that would penetrate in vivo [29]. For determining the skin retention of resveratrol, the authors performed the microtome cuts. The results for the cutaneous retention experiments can be seen in Fig. 4, which also offer a good picture of the whole permeation experiment in terms of resveratrol absorption. Fig. 4 shows that a relatively high amount of resveratrol was retained within the stratum corneum (SC, ∼100 m) and the other epidermal layers. Despite this retention, the drug had good permeation, as the largest amount of drug were retained into the viable epidermis (between 100 and 200 m) and into the beginning of the connective tissue. All drug contained at the viable epidermis is supposed to permeate, once the main barrier of skin, the SC, was already surpassed by the drug, then that amount can be accounted as a drug “possible to permeate”, on the contrary of the drug found at the dermis layers, that can be accounted as “already permeated”. In summary, from the initials 49.30 g applied to the skin, 9.50 g were quantified in the receptor medium, 20.48 g was retained at the SC (do not account as permeated) and 21.41 g was retained at the viable epidermis + dermis (account as permeated), totalizing 30.90 g of resveratrol permeated after 24 h of application (62.6%). On the other hand, 100.65 g were lost to the spreader during the application. It is possible that this would not be lost in vivo because the spreader in such a case would be the finger of the person performing the application. However, this study considered that this person could wash his or her hands after the application, which would result in the true loss of this amount of applied drug. The percutaneous absorption is determined by its physicochemical properties, in particular the molar mass and lipophilicity, and more lipophilic substances possess higher fluxes, in a general manner, as do neutral molecules. The partition coefficients for substances between SC and water (log PSC/water ) can be estimated as: log PSC/water = −0.024 + 0.59 log Poctanol/water [30], which confirms that the partitioning of a drug into the SC from an aqueous
Table 8 Mass balance for the transdermal permeation of resveratrol. Parameter
Result
Theoretical drug amount applied to skin (g)a Real drug amount applied to skin (g)b Total permeated amount after 24 h (g)c Drug retained in stratum corneum (g) Total recovery (%)d Percentage of permeation per dose (%)e
150.0 49.3 30.9 20.5 102.2 62.6
Total amount of cream placed in the donor chamber (4.0 mg cm−2 ). Total amount (a) minus the retained amount in the spreader. c Drug in the receptor medium + viable epidermis + dermis. d Percent of drug quantified in the spreader, the skin and the receptor medium, related to the total amount applied (a). e Mathematical estimate of the total drug that would permeate the skin accordingly to the amount applied [(c/b) × 100]. a
b
solution is directly related to its lipophilicity. Since resveratrol in its non-ionic form has a high lipophilicity (log Poctanol/water = 3.32), this explains the good percutaneous absorption it possesses, in the neutral condition. This satisfactory performance of the product is also quite likely to occur because of a synergistic effect of the liposomal vehicle, which increased resveratrol permeation capacity. Fig. 3 and Table 8 show the mass balance of the study and present some data that allow for some inferences. The acceptable recovery rate lies between 85 and 115%. Using the percentage of permeation by dose (62.6%), one can conclude that a person using the 1-g emulsion dose released by the pump containing 20 mg of resveratrol will have, theoretically, 12.53 mg of it liberated into his bloodstream, gradually and continuously for 24 h. To confirm that, the next step is to perform in vivo pharmacokinetic/pharmacodynamics studies. Acknowledgments The authors would like to thank the Brazilian agencies CNPq, FAPESP and FAPEMIG, for financial support; the companies Fagron and Ortofarma for reagents and infrastructure; Dr. Cristiano Ramos for supplying the skin; and Drs. Ângela Gollner and Sônia Cupolilo (Pathology Service, Hospital UniversitárioUFJF) for skin processing. H.C Polonini would like to thanks CAPES/Rede Nanobiotec-Brasil (Edital 04/CII-2008) and Programa Ciência sem Fronteiras (245781/2012-9) for the scholarships granted.
H.C. Polonini et al. / J. Chromatogr. B 947–948 (2014) 23–31
References [1] J.J. Docherty, J.S. Smith, M.M. Fu, T. Stoner, T. Booth, Antiviral Res. 61 (2004) 19–26. [2] R.A. Baxter, J. Cosmet. Dermatol. 7 (2008) 2–7. [3] F. Orallo, Curr. Med. Chem. 13 (2006) 87–98. [4] J.A. Santos, G.S.G. Carvalho, V. Oliveira, N.R.B. Raposo, A.D. da Silva, Recent Pat. Food, Nutr. Agric. 5 (2013) 144–153. [5] S. Sale, R.D. Verschoyle, D. Boocock, D.J.L. Jones, N. Wilsher, K.C. Ruparelia, G.A. Potter, P.B. Farmer, W.P. Steward, A.J. Gescher, Br. J. Cancer 90 (2004) 736–744. [6] J.A. Baur, D.A. Sinclair, Nat. Drug Discovery 5 (2006) 493–506. [7] M.H. Aziz, S. Reagan-Shaw, J. Wu, B.J. Longley, N. Ahmad, FASEB J. 19 (2005) 1193–1195. [8] H.C. Polonini, L.L. Lima, K.M. Gonc¸alves, A.M. do Carmo, A.D. da Silva, N.R. Raposo, Bioorg. Med. Chem. 15 (2013) 964–968. [9] M.M.Y. Chan, Biochem. Pharmacol. 63 (2002) 99–104. [10] J.L. Bowers, V.V. Tyulmenkov, S.C. Jernigan, C.M. Klinge, Endocrinology 141 (2000) 3657–3667. [11] D. Rossi, A. Guerrini, R. Bruni, E. Brognara, M. Borgatti, R. Gambari, S. Maietti, G. Sacchetti, Molecules 17 (2012) 12393–12405. [12] A.M. Giuffrè, Afr. J. Agric. Res. 8 (2013) 224–229. ´ A. Petrovic, ´ S. Jovic, ´ L. Gojkovic´ Bukarica, M. Bursac, ´ J. Cvejic, ´ [13] M. Atanackovic, Food Chem. 131 (2012) 513–518. [14] G. Singh, R.S. Pai, V. Pandit, J. Adv. Pharm. Technol. Res. 3 (2012) 130–135. [15] C.F. Hung, Y.K. Lin, Z.R. Huang, J.Y. Fang, Biol. Pharm. Bull. 31 (2008) 955–962. [16] C.G.A. da Silva, C.H. Collins, J. Chromatogr. A 1260 (2012) 81–87. [17] C.G.A. da Silva, C.H. Collins, J. Chromatogr. A 1232 (2012) 248–256. [18] H.C. Polonini, M.A.L. de Oliveira, A.O. Ferreira, N.R.B. Raposo, L.N. Grossi, M.A.F. Brandão, J. Braz. Chem. Soc. 22 (2011) 1263–1270.
31
[19] ICH, ICH—International Conference on Harmonization of Technical Requirements for registration of Pharmaceuticals for Human Use, in: Guideline on Validation of Analytical Procedure-Methodology (Q2BR1), Geneva, 2005. [20] INMETRO—Brazilian National Institute of Metrology, Standardization and Industrial Quality, Orientacc¸ões sobre Validacc¸ão de Métodos de Ensaios Químicos, DOQ-CGCRE-008, 2003. [21] B. Baert, J. Boonen, C. Burvenich, N. Roche, F. Stillaert, P. Blondeel, J. van Boxclaer, B. de Spiegeleer, J. Pharm. Pharm. Sci. 13 (2010) 218–230. [22] B.T. Veering, A.G. Burm, J.H. Souverijn, J.M. Serree, J. Spierdijk, Br. J. Clin. Pharmacol. 29 (1990) 201–206. [23] A.R. Baby, C.A. Haroutiounian-Filho, F.D. Sarruf, C.A. Pinto, T.M. Kaneko, M.V. Velasco, Drug. Dev. Ind. Pharm. 35 (2009) 272–282. [24] FDA—Food and Drug Administration, Guidance for Industry on Nonsterile Semisolid Dosage Forms—Scale-up and Postapproval Changes: Chemistry, Manufacturing, and Controls, In vitro Release Testing and In Vivo Bioequivalence Documentation, FDA—Food and Drug Administration, U.S. Department of Health and Human Services, Center for Drug Evaluation and Research (CDER), 1997. [25] H.S. Chiong, Y.K. Yong, Z. Ahmad, M.R. Sulaiman, Z.A. Zakaria, K.H. Yuen, M.N. Hakim, Int. J. Nanomed. 8 (2013) 1245–1255. [26] J. Reichling, U. Landvatter, H. Wagner, K.H. Kostka, U.F. Schaefer, Eur. J. Pharm. Biopharm. 64 (2006) 222–228. [27] A. Olejnik, J. Goscianska, I. Nowak, J. Pharm. Sci. 101 (2012) 4032–4045. [28] R. Sharma, R.B. Walker, K. Pathak, Ind. J. Pharm. Ed. Res. 45 (2011) 25–31. [29] J.M. Sallovitz, P. Nejamkin, A.L. Lifschitz, G.L. Virkel, F.A. Imperiale, C.E. Lanusse, J. Vet. Pharmacol. Ther. 35 (2012) 184–192. [30] M.S. Roberts, W.J. Pugh, J. Hadgraft, Int. J. Pharm. 132 (1996) 23–42.
Contents lists available at ScienceDirect
Journal of Chromatography B journal homepage: www.elsevier.com/locate/chromb
In vitro drug release and ex vivo percutaneous absorption of resveratrol cream using HPLC with zirconized silica stationary phase Hudson Caetano Polonini a,∗ , Carina de Almeida Bastos b , Marcone Augusto Leal de Oliveira b , Carla Grazieli Azevedo da Silva c , Carol Hollingworth Collins c , Marcos Antônio Fernandes Brandão a , Nádia Rezende Barbosa Raposo a a
Núcleo de Pesquisa e Inovac¸ão em Ciências da Saúde (NUPICS), Federal University of Juiz de Fora, 36036-900 Juiz de Fora, MG, Brazil Grupo de Química Analítica e Quimiometria (GQAQ), Federal University of Juiz de Fora, 36036-900 Juiz de Fora, MG, Brazil c Institute of Chemistry, University of Campinas, PO Box 6154, 13083-970 Campinas, SP, Brazil b
a r t i c l e
i n f o
Article history: Received 6 September 2013 Received in revised form 2 December 2013 Accepted 3 December 2013 Available online 17 December 2013 Keywords: In vitro drug release Percutaneous absorption Transdermal dosage form Resveratrol Factorial design Validation studies
a b s t r a c t Since the designs of optimal formulations for resveratrol permeation via the skin are lacking, the aim of this study was to establish the profile of resveratrol permeability into and across human skin. For that, a laboratory-made chromatographic column was used (Zr-PMODS), with its performance being compared to a traditional C18 column. In vitro drug release was conducted with polysulfone membranes, and the flux (JS ) was 30.49 g cm−2 h−1 ), with a lag time (LT ) of 0.04 h, following a pseudo-first-order kinetics. For ex vivo percutaneous absorption using excised female human skin, the kinetic profile was the same, but JS was 0.87 g cm−2 h−1 and LT was 0.97 h. From the initials 49.30 g applied to the skin, 9.50 g were quantified in the receptor medium, 20.48 g was retained at the stratum corneum (do not account as permeated) and 21.41 g was retained at the viable epidermis + dermis (account as permeated), totalizing 30.90 g of resveratrol permeated after 24 h of application (62.6%). From these results, one can conclude that a person using the 1-g emulsion dose released by the pump containing 20 mg of resveratrol will have, theoretically, 12.53 mg of it liberated into his bloodstream, gradually and continuously for 24 h. © 2013 Elsevier B.V. All rights reserved.
1. Introduction Resveratrol (3,5,4-trihydroxystilbene) (Fig. 1) is a naturally occurring non-flavanoid phenolic compound produced by some spermatophytes, such as grapes [1]. It is found in high concentrations in many red grape skins and their wines [2], with the trans-resveratrol isomer being more biologically active [3]. The substance has been reported to possess antioxidant, neuroprotective, antiphotoaging and antiviral activities, and it seems that it also plays a role in the prevention and reduction of pathological processes such as inflammation, cancer and heart diseases [4]. However, it has poor oral bioavailability, which creates a dilemma between its great in vitro efficacy and its low in vivo effect [5], mainly because it is extensively metabolized in the body [6].
∗ Corresponding author at: Federal University of Juiz de Fora, Rua José Lourenc¸o Kelmer, s/n, Campus Universitário, 36036-900, Juiz de Fora, MG, Brazil. Tel.: +55 32 21023809; fax: +55 32 21023809. E-mail addresses: [email protected] (H.C. Polonini), [email protected] (M.A.L.d. Oliveira), [email protected] (M.A.F. Brandão), [email protected] (N.R.B. Raposo). 1570-0232/$ – see front matter © 2013 Elsevier B.V. All rights reserved. http://dx.doi.org/10.1016/j.jchromb.2013.12.005
On the other hand, topically applied resveratrol has been shown to possess more pronounced properties, namely: it is antiproliferative and chemopreventive against skin carcinogenesis [7]; gives sunprotection against skin damage from ultraviolet B (UVB) exposure [8]; is antimicrobial against dermatophytes and herpes simplex virus [1,9]; and activates estrogen receptors [10]. Thus, skin delivery of resveratrol is paramount for the effective insertion of the substance into pharmaceuticals, either topical or transdermal. According to literature reports, the stationary phase most commonly used for resveratrol analysis is the octadecyl-bonded silica (C18), whether in nutraceuticals [11], grape fruits [12], wines [13] or biological samples [14]. However, to the best of the authors’ knowledge transdermal emulsions containing resveratrol for human use have not been investigated so far, although one previous study have reported encouraging data for solutions and hydrogels [15]. Since the designs of optimal formulations for resveratrol permeation via the skin are lacking [15], the aim of this study was to establish the profile of resveratrol permeability into and across human skin. For that, a laboratory-made chromatographic column was used (Zr-PMODS), with its performance being compared to a commercial C18 column.
24
H.C. Polonini et al. / J. Chromatogr. B 947–948 (2014) 23–31
Fig. 1. Resveratrol chemical structure.
2. Materials and methods 2.1. Reagents, standard and materials The acetonitrile (ACN) used in the preparation of the mobile phase was HPLC grade, and sodium chloride (NaCl), calcium chloride (CaCl2 ), magnesium sulfate (MgSO4 ), and potassium dihydrogen phosphate (KH2 PO4 ) were of analytical grade, all from Sigma-Aldrich (USA). Ultrapure water (H2 O) obtained in an AquaMax-Ultra 370 Series (Young Lin, Korea) (18.2 M cm resistivity at 25 ◦ C and 1100 m) because in vivo it is vascularized, and thus the drug within it is able to reach the bloodstream [26]. As ex vivo skin has its microcirculation obliterated, the dermis can retain compounds that would penetrate in vivo [29]. For determining the skin retention of resveratrol, the authors performed the microtome cuts. The results for the cutaneous retention experiments can be seen in Fig. 4, which also offer a good picture of the whole permeation experiment in terms of resveratrol absorption. Fig. 4 shows that a relatively high amount of resveratrol was retained within the stratum corneum (SC, ∼100 m) and the other epidermal layers. Despite this retention, the drug had good permeation, as the largest amount of drug were retained into the viable epidermis (between 100 and 200 m) and into the beginning of the connective tissue. All drug contained at the viable epidermis is supposed to permeate, once the main barrier of skin, the SC, was already surpassed by the drug, then that amount can be accounted as a drug “possible to permeate”, on the contrary of the drug found at the dermis layers, that can be accounted as “already permeated”. In summary, from the initials 49.30 g applied to the skin, 9.50 g were quantified in the receptor medium, 20.48 g was retained at the SC (do not account as permeated) and 21.41 g was retained at the viable epidermis + dermis (account as permeated), totalizing 30.90 g of resveratrol permeated after 24 h of application (62.6%). On the other hand, 100.65 g were lost to the spreader during the application. It is possible that this would not be lost in vivo because the spreader in such a case would be the finger of the person performing the application. However, this study considered that this person could wash his or her hands after the application, which would result in the true loss of this amount of applied drug. The percutaneous absorption is determined by its physicochemical properties, in particular the molar mass and lipophilicity, and more lipophilic substances possess higher fluxes, in a general manner, as do neutral molecules. The partition coefficients for substances between SC and water (log PSC/water ) can be estimated as: log PSC/water = −0.024 + 0.59 log Poctanol/water [30], which confirms that the partitioning of a drug into the SC from an aqueous
Table 8 Mass balance for the transdermal permeation of resveratrol. Parameter
Result
Theoretical drug amount applied to skin (g)a Real drug amount applied to skin (g)b Total permeated amount after 24 h (g)c Drug retained in stratum corneum (g) Total recovery (%)d Percentage of permeation per dose (%)e
150.0 49.3 30.9 20.5 102.2 62.6
Total amount of cream placed in the donor chamber (4.0 mg cm−2 ). Total amount (a) minus the retained amount in the spreader. c Drug in the receptor medium + viable epidermis + dermis. d Percent of drug quantified in the spreader, the skin and the receptor medium, related to the total amount applied (a). e Mathematical estimate of the total drug that would permeate the skin accordingly to the amount applied [(c/b) × 100]. a
b
solution is directly related to its lipophilicity. Since resveratrol in its non-ionic form has a high lipophilicity (log Poctanol/water = 3.32), this explains the good percutaneous absorption it possesses, in the neutral condition. This satisfactory performance of the product is also quite likely to occur because of a synergistic effect of the liposomal vehicle, which increased resveratrol permeation capacity. Fig. 3 and Table 8 show the mass balance of the study and present some data that allow for some inferences. The acceptable recovery rate lies between 85 and 115%. Using the percentage of permeation by dose (62.6%), one can conclude that a person using the 1-g emulsion dose released by the pump containing 20 mg of resveratrol will have, theoretically, 12.53 mg of it liberated into his bloodstream, gradually and continuously for 24 h. To confirm that, the next step is to perform in vivo pharmacokinetic/pharmacodynamics studies. Acknowledgments The authors would like to thank the Brazilian agencies CNPq, FAPESP and FAPEMIG, for financial support; the companies Fagron and Ortofarma for reagents and infrastructure; Dr. Cristiano Ramos for supplying the skin; and Drs. Ângela Gollner and Sônia Cupolilo (Pathology Service, Hospital UniversitárioUFJF) for skin processing. H.C Polonini would like to thanks CAPES/Rede Nanobiotec-Brasil (Edital 04/CII-2008) and Programa Ciência sem Fronteiras (245781/2012-9) for the scholarships granted.
H.C. Polonini et al. / J. Chromatogr. B 947–948 (2014) 23–31
References [1] J.J. Docherty, J.S. Smith, M.M. Fu, T. Stoner, T. Booth, Antiviral Res. 61 (2004) 19–26. [2] R.A. Baxter, J. Cosmet. Dermatol. 7 (2008) 2–7. [3] F. Orallo, Curr. Med. Chem. 13 (2006) 87–98. [4] J.A. Santos, G.S.G. Carvalho, V. Oliveira, N.R.B. Raposo, A.D. da Silva, Recent Pat. Food, Nutr. Agric. 5 (2013) 144–153. [5] S. Sale, R.D. Verschoyle, D. Boocock, D.J.L. Jones, N. Wilsher, K.C. Ruparelia, G.A. Potter, P.B. Farmer, W.P. Steward, A.J. Gescher, Br. J. Cancer 90 (2004) 736–744. [6] J.A. Baur, D.A. Sinclair, Nat. Drug Discovery 5 (2006) 493–506. [7] M.H. Aziz, S. Reagan-Shaw, J. Wu, B.J. Longley, N. Ahmad, FASEB J. 19 (2005) 1193–1195. [8] H.C. Polonini, L.L. Lima, K.M. Gonc¸alves, A.M. do Carmo, A.D. da Silva, N.R. Raposo, Bioorg. Med. Chem. 15 (2013) 964–968. [9] M.M.Y. Chan, Biochem. Pharmacol. 63 (2002) 99–104. [10] J.L. Bowers, V.V. Tyulmenkov, S.C. Jernigan, C.M. Klinge, Endocrinology 141 (2000) 3657–3667. [11] D. Rossi, A. Guerrini, R. Bruni, E. Brognara, M. Borgatti, R. Gambari, S. Maietti, G. Sacchetti, Molecules 17 (2012) 12393–12405. [12] A.M. Giuffrè, Afr. J. Agric. Res. 8 (2013) 224–229. ´ A. Petrovic, ´ S. Jovic, ´ L. Gojkovic´ Bukarica, M. Bursac, ´ J. Cvejic, ´ [13] M. Atanackovic, Food Chem. 131 (2012) 513–518. [14] G. Singh, R.S. Pai, V. Pandit, J. Adv. Pharm. Technol. Res. 3 (2012) 130–135. [15] C.F. Hung, Y.K. Lin, Z.R. Huang, J.Y. Fang, Biol. Pharm. Bull. 31 (2008) 955–962. [16] C.G.A. da Silva, C.H. Collins, J. Chromatogr. A 1260 (2012) 81–87. [17] C.G.A. da Silva, C.H. Collins, J. Chromatogr. A 1232 (2012) 248–256. [18] H.C. Polonini, M.A.L. de Oliveira, A.O. Ferreira, N.R.B. Raposo, L.N. Grossi, M.A.F. Brandão, J. Braz. Chem. Soc. 22 (2011) 1263–1270.
31
[19] ICH, ICH—International Conference on Harmonization of Technical Requirements for registration of Pharmaceuticals for Human Use, in: Guideline on Validation of Analytical Procedure-Methodology (Q2BR1), Geneva, 2005. [20] INMETRO—Brazilian National Institute of Metrology, Standardization and Industrial Quality, Orientacc¸ões sobre Validacc¸ão de Métodos de Ensaios Químicos, DOQ-CGCRE-008, 2003. [21] B. Baert, J. Boonen, C. Burvenich, N. Roche, F. Stillaert, P. Blondeel, J. van Boxclaer, B. de Spiegeleer, J. Pharm. Pharm. Sci. 13 (2010) 218–230. [22] B.T. Veering, A.G. Burm, J.H. Souverijn, J.M. Serree, J. Spierdijk, Br. J. Clin. Pharmacol. 29 (1990) 201–206. [23] A.R. Baby, C.A. Haroutiounian-Filho, F.D. Sarruf, C.A. Pinto, T.M. Kaneko, M.V. Velasco, Drug. Dev. Ind. Pharm. 35 (2009) 272–282. [24] FDA—Food and Drug Administration, Guidance for Industry on Nonsterile Semisolid Dosage Forms—Scale-up and Postapproval Changes: Chemistry, Manufacturing, and Controls, In vitro Release Testing and In Vivo Bioequivalence Documentation, FDA—Food and Drug Administration, U.S. Department of Health and Human Services, Center for Drug Evaluation and Research (CDER), 1997. [25] H.S. Chiong, Y.K. Yong, Z. Ahmad, M.R. Sulaiman, Z.A. Zakaria, K.H. Yuen, M.N. Hakim, Int. J. Nanomed. 8 (2013) 1245–1255. [26] J. Reichling, U. Landvatter, H. Wagner, K.H. Kostka, U.F. Schaefer, Eur. J. Pharm. Biopharm. 64 (2006) 222–228. [27] A. Olejnik, J. Goscianska, I. Nowak, J. Pharm. Sci. 101 (2012) 4032–4045. [28] R. Sharma, R.B. Walker, K. Pathak, Ind. J. Pharm. Ed. Res. 45 (2011) 25–31. [29] J.M. Sallovitz, P. Nejamkin, A.L. Lifschitz, G.L. Virkel, F.A. Imperiale, C.E. Lanusse, J. Vet. Pharmacol. Ther. 35 (2012) 184–192. [30] M.S. Roberts, W.J. Pugh, J. Hadgraft, Int. J. Pharm. 132 (1996) 23–42.
