- Email: [email protected]
Antifungal study of the resinous exudate and of meroterpenoids isolated from Psoralea glandulosa (Fabaceae)
Journal of Ethnopharmacology 144 (2012) 809–811
Contents lists available at SciVerse ScienceDirect
Journal of Ethnopharmacology journal homepage: www.elsevier.com/locate/jep
Ethnopharmacological communication
Antifungal study of the resinous exudate and of meroterpenoids isolated from Psoralea glandulosa (Fabaceae) Alejandro Madrid a,n, Luis Espinoza a, Cesar Gonza´lez a, Marco Mellado a, Joan Villena b, Rocı´o Santander c, Viviana Silva d, Iva´n Montenegro a,n a
˜a No. 1680, Valparaı´so, Chile Departamento de Quı´mica, Universidad Te´cnica Federico Santa Marı´a, Av. Espan Centro de Investigaciones Biome´dicas (CIB), Escuela de Medicina, Universidad de Valparaı´so, Av. Hontaneda No. 2664, Valparaı´so, Chile c ´gica, Universidad de Santiago de Chile, Santiago, Chile Laboratorio de Carnes, Facultad Tecnolo d ´lica de Valparaı´so, Valparaı´so, Chile Instituto de Quı´mica, Pontificia Universidad Cato b
a r t i c l e i n f o
abstract
Article history: Received 30 July 2012 Received in revised form 5 October 2012 Accepted 14 October 2012 Available online 23 October 2012
Ethnopharmacological relevance: Psoralea glandulosa L. (Fabaceae) is a medicinal resinous shrub used in Chilean folk medicine as antiseptic in treatment of infections and skin diseases caused by bacteria and fungus. Aim of the study: To evaluate the in vitro antifungal activity of the resin and the active components from P. glandulosa against clinical yeast isolates. Materials and methods: Active compounds were obtained of the resinous exudate from aerial parts of P. glandulosa. Eight species of yeast were exposed to the resin and two major compounds. Minimum inhibitory concentration (MIC80) was determined according to the standard broth microdilution method. Results: Bakuchiol and 3-hydroxy-bakuchiol demonstrated potent activity with the MIC80 ranging from 4 to 416 and 0.125 to 16 mg/mL, respectively. The resin had some degree of antifungal activity. Conclusions: The overall results provided important information for the potential application of the 3-hydroxy-bakuchiol from P. glandulosa in the therapy of serious infection and skin diseases caused by clinical yeast. & 2012 Elsevier Ireland Ltd. All rights reserved.
Keywords: Antifungal activity Meroterpenoids Psoralea glandulosa Resinous exudate
1. Introduction
2. Materials and methods
Psoralea glandulosa L. (Fabaceae) is a medicinal resinous shrub used in Chilean folk medicine. It is known by the vernacular names ‘‘culen’’, ‘‘cule’’ and ‘‘hualhua’’. It is also known by the Aymara name of ‘‘Wallikaya’’ (Girault, 1984), which is commonly used in the Mapuche communities (Hoffmann et al., 1992). The genus Psoralea is represented in Chile by culen which is characterized by producing resinous exudate from its glandular trichome that covers the surface of leaves and stems, for that reason its name is glandulosa (Riedemann and Aldunate, 2001). P. glandulosa is commonly used in folk medicine as vulnerary and for hemorrhoids (Montes and Wilkomirsky, 1987). It acts as antiseptic in treatment of infections and skin diseases caused by bacteria and fungus (Hoffmann et al., 1992; Rozzi, 1984). In this communication, the antifungal activity of the resinous exudate of P. glandulosa and of the two major meroterpenoids isolated from the plant’s resinous exudate are presented.
2.1. Plant material, extraction of the resinous exudate and pure compounds
n
Corresponding authors. Tel: þ 56 32 2654328; fax: 56 32 2654782. E-mail address: [email protected] (A. Madrid).
0378-8741/$ - see front matter & 2012 Elsevier Ireland Ltd. All rights reserved. http://dx.doi.org/10.1016/j.jep.2012.10.027
P. glandulosa was collected during the flowering season (November 2011) in Lo Orozco (Regio´n de Valparaı´so, Chile; 3311300 S, 7112500 W). A voucher specimen (No. Pg-11112) was deposited at the Herbarium of the Natural Products Laboratory, ‘‘Dr. Herbert Appel A’’, Department of Chemistry, Universidad Te´cnico Federico Santa Marı´a, Valparaı´so, Chile. Aerial parts of fresh P. glandulosa (300 g) were extracted by immersion in dichloromethane (1 L) for 30 s at room temperature and the filtered solution was then concentrated under reduced pressure to give the resin extract (5.54 g). The resin (1.01 g) was submitted to column chromatography (silica gel Merck 200– 300 mesh) and eluted with EtOAc–hexane mixtures of increasing polarity as a developing system. 4-[(1E,3S)-3-ethenyl-3,7-dimethylocta-1,6-dienyl]phenol (bakuchiol, 701.3 mg), isolated as a colorless oil, and 4-[(1E,3S)-3ethenyl-3,7-dimethylocta-1,6-dienyl] benzene-1,2-diol (3-hydroxybakuchiol, 85.7 mg), isolated as a yellow oil, were identical in all
810
A. Madrid et al. / Journal of Ethnopharmacology 144 (2012) 809–811
respects (Fourier transform infrared spectroscopy, 1H and 13C nuclear magnetic resonance, [a]D) with the compounds described previously (Erazo et al., 1990 and Labbe´ et al., 1996).
Table 1 Antifungal activity of the resinous exudate and of meroterpenoids isolated from P. glandulosa. Compounds
2.2. Antifungal activity determination The crude dichloromethane P. glandulosa extract and meroterpenoids were assayed against the strains of Candida albicans ATCC 7978 and Candida parasilopsis ATCC 22019 tested belonging to the collection of the Microbiology Clinic Laboratory, Facultad de Medicina, Universidad Austral de Chile and including strains of clinical isolates obtained from Hospital Base de Valdivia have been tested. The collection included C. albicans sp. 10935, Candida dubliniensis 3240, Candida glabrata 10912, Candida guilliermondii 2204, Candida lusitaniae 2305 and Candida tropicalis 9841. All isolates were obtained from blood or other normally sterile sites and represented individual infectious episodes. The isolates were identified by standard methods (Cassel and Vargas, 2006) and stored as water suspensions until being used in the study. Prior to being tested, each isolate was passaged at least twice onto potato dextrose agar (Remel, Lenexa, Kansas) and Chromagar Candida medium (Becton Dickinson and Company, Sparks, MD) to ensure purity and viability. The antifungal susceptibility testing was performed in accordance with the guidelines in (NCCLS, 2002) document M27-A2 by using RPMI 1640 medium, inoculums of 0.5 103 to 2.5 103 cells/mL, and incubation at 35 1C. MICs were determined visually, after 24 h of incubation, as the lowest concentration of drug that caused a significant diminution ( Z50% inhibition) of growth below control levels (Pfaller et al., 2006). The Minimum Inhibitory Concentration (MIC) was determined by microdilution method for yeast (NCCLS/CLSI, 2008) M27-A3. Briefly, cultures of all yeast were placed on Sabouraud Dextrose Agar (SDA) and incubated for 24–72 h at temperature 37 1C. Colonies of this culture were suspended in sterile 0.85% NaCl and the inoculum was standardized according to the scale of 0.5 McFarland (1–5 106 CFU/mL). The antifungal test was performed in 96-well plates. Yeast strains were prepared in sterile water and were diluted in RPMI 1640 medium (except in the sterility control). Resin and compounds were dissolved in dimethylsulfoxide (DMSO) at final concentrations of 16 to 0.03 mg/mL. The MIC80 determination was conducted with approximately 0.5–2.5 103 CFU/mL of the microorgnism in each well. The plates were incubated at 37 1C for 24–48 h and measured the absorbance at 540 nm (Pfaller et al., 2008). The MIC endpoint was calculated as the lowest concentration giving rise to an inhibition of growth equal to or greater than 80% of that of the growth control (MIC80), similar to the visual endpoint criterion recommended for yeast (NCCLS).
3. Results and discussion The antifungal activities of the resinous exudates of P. glandulosa, bakuchiol and 3-hidroxi-bakuchiol are summarized in Tables 1 and 2. Research on resinous exudates from medicinal plants has allowed the identification of several terpenes as the compounds responsible for their antimicrobial activity (Mendoza et al., 1997; Urzu´a et al., 2006). This resinous exudate is very rich in bakuchiol, which was identified as the active antidermatophytic being reported (Lau et al., 2010). Until now, the effect of antifungal activity of the resinous exudate in human pathogenic yeast had not been reported.
P. glandulosa resin Bakuchiol 3-hydroxy-bakuchiol Fluconazole Ketoconazole Itraconazole DMSO
Antifungal testa (MIC80, lg/mL) at 24 h 1
2
3
4
5
6
7
8
416 416 16.0 0.25 1.0 4 i
0.5 2.0 0.5 0.5 1.0 0.5 i
16.0 16.0 4.0 0.25 1.0 0.5 i
4.0 0.25 2.0 0.25 1.0 2.0 i
16.0 16.0 4.0 0.125 1.0 0.25 i
8.0 16.0 4.0 0.5 0.5 1.0 i
0.5 0.125 0.125 0.5 0.5 4.0 i
2.0 2.0 1.0 0.25 0.5 0.5 i
i, inactive. a 1,Candida glabrata (10912); 2, Candida parasilopsis (ATCC 22019); 3, Candida tropicalis (9841); 4, Candida lusitaniae (2305); 5, Candida albicans (ATCC 7978); 6, Candida albicans sp. (10935); 7, Candida guilliermondii (2204); 8, Candida dublinensis (3240).
Table 2 Antifungal activity of the resinous exudate and of meroterpenoids isolated from P. glandulosa. Compounds
P. glandulosa resin Bakuchiol 3-hydroxy-bakuchiol Fluconazole Ketoconazole Itraconazole DMSO
Antifungal testa (MIC80, lg/mL) at 48 h 1
2
3
4
5
6
7
8
416 416 16.0 8.0 1.25 8.0 i
16.0 416 8.0 0.5 1.25 1.0 i
4 16 4 16 4.0 1.0 1.25 1.25 i
8.0 416 2.0 0.5 1.25 4.0 i
16.0 416 8.0 0.25 1.25 0.5 i
16.0 416 8.0 0.5 1.0 2.0 i
1.0 8.0 0.125 1.0 1.0 8.0 i
4.0 4.0 2.0 1.0 1.0 2.0 i
i, inactive. a 1, Candida glabrata (10912); 2, Candida parasilopsis (ATCC 22019); 3, Candida tropicalis (9841); 4, Candida lusitaniae (2305); 5, Candida albicans (ATCC 7978); 6, Candida albicans sp. (10935); 7, Candida guilliermondii (2204); 8, Candida dublinensis (3240).
It is noteworthy that 3-hydroxy-bakuchiol exhibited a very strong activity against the majority of strains, specially, C. guilliermondii. Similarly, bakuchiol showed a less antifungal activity than the parent 3-hydroxy-bakuchiol, which possesses one more hydroxyl group. For that reason, we think that the antifungal activity of the resinous exudate is due to a synergistic effect triggered by its constituents, mainly bakuchiol and 3-hydroxy-bakuchiol. In conclusion, the vernacular medicinal use of P. glandulosa as an antiseptic in treatment of infections and skin diseases is validated in the present communication by our findings that 3-hydroxy-bakuchiol and bakuchiol, isolated from the resinous exudate of the aerial parts of this plant, are effective against all tested fungi except one human pathogen (C. glabrata). In addition, a more convenient method for obtaining large amounts of these compounds is described, clearly superior to the original procedure (Labbe´ et al., 1996), which could only isolate bakuchiol and 3-hydroxy-bakuchiol in small amounts of time from the leaves and stems of P. glandulosa. From the resinous exudate of the same plant we have been able to isolate much larger amounts of an 8.2:1 mixture of the two compounds. The fact that these readily available compounds have also proved to be bioactive at relatively low concentrations clearly justifies our interest in pursuing the synthesis and biological studies of their derivatives in the future.
A. Madrid et al. / Journal of Ethnopharmacology 144 (2012) 809–811
Acknowledgements The authors thank DGIP of Universidad Te´cnica Federico Santa Marı´a for financing (Project DGIP No. 13.11.36, PAC 2010–2012 for A.M. and PIIC 2012 for I.M.), and support to this research and graphic designer Reinaldo Madrid V. References Cassel, E., Vargas, R., 2006. Experiments and modeling of the Cymbopogon winterianus essential oil extraction by steam distillation. Journal of the Mexican Chemical Society 50, 126–129. CLSI, Clinical and Laboratory Standards Institute Document M27-A3, third ed., Wayne Ed., NCCLS: Pennsylvania, 2008, 28(14), pp. 1–25. Erazo, S., Garcia, R., Delle monache, F., 1990. Bakuchiol and other compounds from Psoralea glandulosa. Revista Latinoamericana de Quı´mica 21, 62. Girault, L., 1984. Kallawaya, Gue´risseurs itine´rants des Andes. Recherche sur les pratiques me´dicinales et magiques. E´ditions de l0 Orstom, Paris, 245. Hoffmann A., Farga C., Lastra J. and Veghazi E., 1992. Plantas Medicinales de uso comu´n en Chile, Fundacio´n Claudio Gay, Santiago de Chile, pp. 79–82. Labbe´, C., Faini, F., Coll, J., Connolly, J., 1996. Bakuchiol derivatives from the leaves of Psoralea glandulosa. Phytochemistry 42, 1299–1303. Lau, K., Fu, L., Wong, C., Wong, Y., Lau, C., Han, S., Chan, P., Fung, P., Lau, C., Huiy, M., Leung, P., 2010. Two antifungal components isolated from Fructus Psoraleae
811
and Folium Eucalypti Globuli by bioassay-guided purification. The American Journal of Chinese Medicine 38, 1005–1014. Mendoza, L., Wilkens, M., Urzua, A., 1997. Antimicrobial study of the resinous exudates and of diterpenoids and flavonoids isolated from some Chilean Pseudognaphalium (Asteraceae). Journal of Ethnopharmacology 58, 85–88. Montes, M., Wilkomirsky, T., 1987. Medicina Tradicional Chilena. Editorial Universitaria, Santiago de Chile, p. 108. National Committee for Clinical Laboratory Standards. 2002. Reference Method for Broth Dilution Antifungal Susceptibility Testing for Yeasts. Approved Standard M27-A2, second ed., National Committee for Clinical Laboratory Standards, Wayne, PA. Pfaller, M.A., Boyken, L., Hollis, R.J., Messer, S.A., Tendolkar, S., Diekema, D.J., 2006. In vitro susceptibilities of Candida spp. to caspofungin: four years of global surveillance. Journal of Clinical Microbiology 44, 760–763. Pfaller, M.A., Boyken, L., Hollis, R.J., Messer, S.A., Tendolkar, S., Diekema, D.J., 2008. In Vitro susceptibility of invasive isolates of Candida spp. to anidulafungin, caspofungin, and micafungin: six years of global surveillance. Journal of Clinical Microbiology 46, 150–156. Riedemann, P., Aldunate, G., 2001. Flora Nativa de Valor Ornamental. Chile, Zona Centro Identificacio´n y propagacio´n. Editorial Andre´s Bello, Santiago de Chile, pp. 70–71. Rozzi, S., 1984. Las Plantas, Fuente de Salud. Pı´a Sociedad de San Pablo, Santiago de Chile, p. 104. Urzu´a, A., Jara, F., Tojo, E., Wilkens, M., Mendoza, L., Rezende, M.C., 2006. A new antibacterial clerodane diterpenoid from the resinous exudate of Haplopappus uncinatus. Journal of Ethnopharmacology 103, 297–301.
Contents lists available at SciVerse ScienceDirect
Journal of Ethnopharmacology journal homepage: www.elsevier.com/locate/jep
Ethnopharmacological communication
Antifungal study of the resinous exudate and of meroterpenoids isolated from Psoralea glandulosa (Fabaceae) Alejandro Madrid a,n, Luis Espinoza a, Cesar Gonza´lez a, Marco Mellado a, Joan Villena b, Rocı´o Santander c, Viviana Silva d, Iva´n Montenegro a,n a
˜a No. 1680, Valparaı´so, Chile Departamento de Quı´mica, Universidad Te´cnica Federico Santa Marı´a, Av. Espan Centro de Investigaciones Biome´dicas (CIB), Escuela de Medicina, Universidad de Valparaı´so, Av. Hontaneda No. 2664, Valparaı´so, Chile c ´gica, Universidad de Santiago de Chile, Santiago, Chile Laboratorio de Carnes, Facultad Tecnolo d ´lica de Valparaı´so, Valparaı´so, Chile Instituto de Quı´mica, Pontificia Universidad Cato b
a r t i c l e i n f o
abstract
Article history: Received 30 July 2012 Received in revised form 5 October 2012 Accepted 14 October 2012 Available online 23 October 2012
Ethnopharmacological relevance: Psoralea glandulosa L. (Fabaceae) is a medicinal resinous shrub used in Chilean folk medicine as antiseptic in treatment of infections and skin diseases caused by bacteria and fungus. Aim of the study: To evaluate the in vitro antifungal activity of the resin and the active components from P. glandulosa against clinical yeast isolates. Materials and methods: Active compounds were obtained of the resinous exudate from aerial parts of P. glandulosa. Eight species of yeast were exposed to the resin and two major compounds. Minimum inhibitory concentration (MIC80) was determined according to the standard broth microdilution method. Results: Bakuchiol and 3-hydroxy-bakuchiol demonstrated potent activity with the MIC80 ranging from 4 to 416 and 0.125 to 16 mg/mL, respectively. The resin had some degree of antifungal activity. Conclusions: The overall results provided important information for the potential application of the 3-hydroxy-bakuchiol from P. glandulosa in the therapy of serious infection and skin diseases caused by clinical yeast. & 2012 Elsevier Ireland Ltd. All rights reserved.
Keywords: Antifungal activity Meroterpenoids Psoralea glandulosa Resinous exudate
1. Introduction
2. Materials and methods
Psoralea glandulosa L. (Fabaceae) is a medicinal resinous shrub used in Chilean folk medicine. It is known by the vernacular names ‘‘culen’’, ‘‘cule’’ and ‘‘hualhua’’. It is also known by the Aymara name of ‘‘Wallikaya’’ (Girault, 1984), which is commonly used in the Mapuche communities (Hoffmann et al., 1992). The genus Psoralea is represented in Chile by culen which is characterized by producing resinous exudate from its glandular trichome that covers the surface of leaves and stems, for that reason its name is glandulosa (Riedemann and Aldunate, 2001). P. glandulosa is commonly used in folk medicine as vulnerary and for hemorrhoids (Montes and Wilkomirsky, 1987). It acts as antiseptic in treatment of infections and skin diseases caused by bacteria and fungus (Hoffmann et al., 1992; Rozzi, 1984). In this communication, the antifungal activity of the resinous exudate of P. glandulosa and of the two major meroterpenoids isolated from the plant’s resinous exudate are presented.
2.1. Plant material, extraction of the resinous exudate and pure compounds
n
Corresponding authors. Tel: þ 56 32 2654328; fax: 56 32 2654782. E-mail address: [email protected] (A. Madrid).
0378-8741/$ - see front matter & 2012 Elsevier Ireland Ltd. All rights reserved. http://dx.doi.org/10.1016/j.jep.2012.10.027
P. glandulosa was collected during the flowering season (November 2011) in Lo Orozco (Regio´n de Valparaı´so, Chile; 3311300 S, 7112500 W). A voucher specimen (No. Pg-11112) was deposited at the Herbarium of the Natural Products Laboratory, ‘‘Dr. Herbert Appel A’’, Department of Chemistry, Universidad Te´cnico Federico Santa Marı´a, Valparaı´so, Chile. Aerial parts of fresh P. glandulosa (300 g) were extracted by immersion in dichloromethane (1 L) for 30 s at room temperature and the filtered solution was then concentrated under reduced pressure to give the resin extract (5.54 g). The resin (1.01 g) was submitted to column chromatography (silica gel Merck 200– 300 mesh) and eluted with EtOAc–hexane mixtures of increasing polarity as a developing system. 4-[(1E,3S)-3-ethenyl-3,7-dimethylocta-1,6-dienyl]phenol (bakuchiol, 701.3 mg), isolated as a colorless oil, and 4-[(1E,3S)-3ethenyl-3,7-dimethylocta-1,6-dienyl] benzene-1,2-diol (3-hydroxybakuchiol, 85.7 mg), isolated as a yellow oil, were identical in all
810
A. Madrid et al. / Journal of Ethnopharmacology 144 (2012) 809–811
respects (Fourier transform infrared spectroscopy, 1H and 13C nuclear magnetic resonance, [a]D) with the compounds described previously (Erazo et al., 1990 and Labbe´ et al., 1996).
Table 1 Antifungal activity of the resinous exudate and of meroterpenoids isolated from P. glandulosa. Compounds
2.2. Antifungal activity determination The crude dichloromethane P. glandulosa extract and meroterpenoids were assayed against the strains of Candida albicans ATCC 7978 and Candida parasilopsis ATCC 22019 tested belonging to the collection of the Microbiology Clinic Laboratory, Facultad de Medicina, Universidad Austral de Chile and including strains of clinical isolates obtained from Hospital Base de Valdivia have been tested. The collection included C. albicans sp. 10935, Candida dubliniensis 3240, Candida glabrata 10912, Candida guilliermondii 2204, Candida lusitaniae 2305 and Candida tropicalis 9841. All isolates were obtained from blood or other normally sterile sites and represented individual infectious episodes. The isolates were identified by standard methods (Cassel and Vargas, 2006) and stored as water suspensions until being used in the study. Prior to being tested, each isolate was passaged at least twice onto potato dextrose agar (Remel, Lenexa, Kansas) and Chromagar Candida medium (Becton Dickinson and Company, Sparks, MD) to ensure purity and viability. The antifungal susceptibility testing was performed in accordance with the guidelines in (NCCLS, 2002) document M27-A2 by using RPMI 1640 medium, inoculums of 0.5 103 to 2.5 103 cells/mL, and incubation at 35 1C. MICs were determined visually, after 24 h of incubation, as the lowest concentration of drug that caused a significant diminution ( Z50% inhibition) of growth below control levels (Pfaller et al., 2006). The Minimum Inhibitory Concentration (MIC) was determined by microdilution method for yeast (NCCLS/CLSI, 2008) M27-A3. Briefly, cultures of all yeast were placed on Sabouraud Dextrose Agar (SDA) and incubated for 24–72 h at temperature 37 1C. Colonies of this culture were suspended in sterile 0.85% NaCl and the inoculum was standardized according to the scale of 0.5 McFarland (1–5 106 CFU/mL). The antifungal test was performed in 96-well plates. Yeast strains were prepared in sterile water and were diluted in RPMI 1640 medium (except in the sterility control). Resin and compounds were dissolved in dimethylsulfoxide (DMSO) at final concentrations of 16 to 0.03 mg/mL. The MIC80 determination was conducted with approximately 0.5–2.5 103 CFU/mL of the microorgnism in each well. The plates were incubated at 37 1C for 24–48 h and measured the absorbance at 540 nm (Pfaller et al., 2008). The MIC endpoint was calculated as the lowest concentration giving rise to an inhibition of growth equal to or greater than 80% of that of the growth control (MIC80), similar to the visual endpoint criterion recommended for yeast (NCCLS).
3. Results and discussion The antifungal activities of the resinous exudates of P. glandulosa, bakuchiol and 3-hidroxi-bakuchiol are summarized in Tables 1 and 2. Research on resinous exudates from medicinal plants has allowed the identification of several terpenes as the compounds responsible for their antimicrobial activity (Mendoza et al., 1997; Urzu´a et al., 2006). This resinous exudate is very rich in bakuchiol, which was identified as the active antidermatophytic being reported (Lau et al., 2010). Until now, the effect of antifungal activity of the resinous exudate in human pathogenic yeast had not been reported.
P. glandulosa resin Bakuchiol 3-hydroxy-bakuchiol Fluconazole Ketoconazole Itraconazole DMSO
Antifungal testa (MIC80, lg/mL) at 24 h 1
2
3
4
5
6
7
8
416 416 16.0 0.25 1.0 4 i
0.5 2.0 0.5 0.5 1.0 0.5 i
16.0 16.0 4.0 0.25 1.0 0.5 i
4.0 0.25 2.0 0.25 1.0 2.0 i
16.0 16.0 4.0 0.125 1.0 0.25 i
8.0 16.0 4.0 0.5 0.5 1.0 i
0.5 0.125 0.125 0.5 0.5 4.0 i
2.0 2.0 1.0 0.25 0.5 0.5 i
i, inactive. a 1,Candida glabrata (10912); 2, Candida parasilopsis (ATCC 22019); 3, Candida tropicalis (9841); 4, Candida lusitaniae (2305); 5, Candida albicans (ATCC 7978); 6, Candida albicans sp. (10935); 7, Candida guilliermondii (2204); 8, Candida dublinensis (3240).
Table 2 Antifungal activity of the resinous exudate and of meroterpenoids isolated from P. glandulosa. Compounds
P. glandulosa resin Bakuchiol 3-hydroxy-bakuchiol Fluconazole Ketoconazole Itraconazole DMSO
Antifungal testa (MIC80, lg/mL) at 48 h 1
2
3
4
5
6
7
8
416 416 16.0 8.0 1.25 8.0 i
16.0 416 8.0 0.5 1.25 1.0 i
4 16 4 16 4.0 1.0 1.25 1.25 i
8.0 416 2.0 0.5 1.25 4.0 i
16.0 416 8.0 0.25 1.25 0.5 i
16.0 416 8.0 0.5 1.0 2.0 i
1.0 8.0 0.125 1.0 1.0 8.0 i
4.0 4.0 2.0 1.0 1.0 2.0 i
i, inactive. a 1, Candida glabrata (10912); 2, Candida parasilopsis (ATCC 22019); 3, Candida tropicalis (9841); 4, Candida lusitaniae (2305); 5, Candida albicans (ATCC 7978); 6, Candida albicans sp. (10935); 7, Candida guilliermondii (2204); 8, Candida dublinensis (3240).
It is noteworthy that 3-hydroxy-bakuchiol exhibited a very strong activity against the majority of strains, specially, C. guilliermondii. Similarly, bakuchiol showed a less antifungal activity than the parent 3-hydroxy-bakuchiol, which possesses one more hydroxyl group. For that reason, we think that the antifungal activity of the resinous exudate is due to a synergistic effect triggered by its constituents, mainly bakuchiol and 3-hydroxy-bakuchiol. In conclusion, the vernacular medicinal use of P. glandulosa as an antiseptic in treatment of infections and skin diseases is validated in the present communication by our findings that 3-hydroxy-bakuchiol and bakuchiol, isolated from the resinous exudate of the aerial parts of this plant, are effective against all tested fungi except one human pathogen (C. glabrata). In addition, a more convenient method for obtaining large amounts of these compounds is described, clearly superior to the original procedure (Labbe´ et al., 1996), which could only isolate bakuchiol and 3-hydroxy-bakuchiol in small amounts of time from the leaves and stems of P. glandulosa. From the resinous exudate of the same plant we have been able to isolate much larger amounts of an 8.2:1 mixture of the two compounds. The fact that these readily available compounds have also proved to be bioactive at relatively low concentrations clearly justifies our interest in pursuing the synthesis and biological studies of their derivatives in the future.
A. Madrid et al. / Journal of Ethnopharmacology 144 (2012) 809–811
Acknowledgements The authors thank DGIP of Universidad Te´cnica Federico Santa Marı´a for financing (Project DGIP No. 13.11.36, PAC 2010–2012 for A.M. and PIIC 2012 for I.M.), and support to this research and graphic designer Reinaldo Madrid V. References Cassel, E., Vargas, R., 2006. Experiments and modeling of the Cymbopogon winterianus essential oil extraction by steam distillation. Journal of the Mexican Chemical Society 50, 126–129. CLSI, Clinical and Laboratory Standards Institute Document M27-A3, third ed., Wayne Ed., NCCLS: Pennsylvania, 2008, 28(14), pp. 1–25. Erazo, S., Garcia, R., Delle monache, F., 1990. Bakuchiol and other compounds from Psoralea glandulosa. Revista Latinoamericana de Quı´mica 21, 62. Girault, L., 1984. Kallawaya, Gue´risseurs itine´rants des Andes. Recherche sur les pratiques me´dicinales et magiques. E´ditions de l0 Orstom, Paris, 245. Hoffmann A., Farga C., Lastra J. and Veghazi E., 1992. Plantas Medicinales de uso comu´n en Chile, Fundacio´n Claudio Gay, Santiago de Chile, pp. 79–82. Labbe´, C., Faini, F., Coll, J., Connolly, J., 1996. Bakuchiol derivatives from the leaves of Psoralea glandulosa. Phytochemistry 42, 1299–1303. Lau, K., Fu, L., Wong, C., Wong, Y., Lau, C., Han, S., Chan, P., Fung, P., Lau, C., Huiy, M., Leung, P., 2010. Two antifungal components isolated from Fructus Psoraleae
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and Folium Eucalypti Globuli by bioassay-guided purification. The American Journal of Chinese Medicine 38, 1005–1014. Mendoza, L., Wilkens, M., Urzua, A., 1997. Antimicrobial study of the resinous exudates and of diterpenoids and flavonoids isolated from some Chilean Pseudognaphalium (Asteraceae). Journal of Ethnopharmacology 58, 85–88. Montes, M., Wilkomirsky, T., 1987. Medicina Tradicional Chilena. Editorial Universitaria, Santiago de Chile, p. 108. National Committee for Clinical Laboratory Standards. 2002. Reference Method for Broth Dilution Antifungal Susceptibility Testing for Yeasts. Approved Standard M27-A2, second ed., National Committee for Clinical Laboratory Standards, Wayne, PA. Pfaller, M.A., Boyken, L., Hollis, R.J., Messer, S.A., Tendolkar, S., Diekema, D.J., 2006. In vitro susceptibilities of Candida spp. to caspofungin: four years of global surveillance. Journal of Clinical Microbiology 44, 760–763. Pfaller, M.A., Boyken, L., Hollis, R.J., Messer, S.A., Tendolkar, S., Diekema, D.J., 2008. In Vitro susceptibility of invasive isolates of Candida spp. to anidulafungin, caspofungin, and micafungin: six years of global surveillance. Journal of Clinical Microbiology 46, 150–156. Riedemann, P., Aldunate, G., 2001. Flora Nativa de Valor Ornamental. Chile, Zona Centro Identificacio´n y propagacio´n. Editorial Andre´s Bello, Santiago de Chile, pp. 70–71. Rozzi, S., 1984. Las Plantas, Fuente de Salud. Pı´a Sociedad de San Pablo, Santiago de Chile, p. 104. Urzu´a, A., Jara, F., Tojo, E., Wilkens, M., Mendoza, L., Rezende, M.C., 2006. A new antibacterial clerodane diterpenoid from the resinous exudate of Haplopappus uncinatus. Journal of Ethnopharmacology 103, 297–301.