PPDF ICB - UFRJ
LPNBio - Chemistry Laboratory of Bioactive Natural Products





Summary of the 5 most relevant articles in the group

Schmourlo G, Mendonça-Filho RR, Alviano CS, Costa SS. Screening of antifungal agents using ethanol precipitation and bioautography of medicinal and food plants. Journal of Ethnopharmacology, v. 96, n.3, p. 563-568, 2005.
Abstract: In the search for bioactive compounds, bioautography and ethanol precipitation of macromolecules (proteins, polysaccharides, etc.) of plant aqueous extracts were associated in an antifungal screening. Thus, the supernatants, precipitates (obtained by ethanol precipitation) and aqueous extracts were investigated of medicinal and fruit bearing plants used against skin diseases by the Brazilian population. The agar diffusion and broth dilution methods were used to assess the activity against three fungi: Candida albicans, Trichophyton rubrum and Cryptococcus neoformans. The results, evaluated by the diameter of the inhibition zone of fungal growth, indicate that six plant species, among the 16 investigated, showed significant antifungal activity. The minimal inhibitory concentration (MIC) was determined on plant extracts that showed high efficacy against the tested microorganisms. The most susceptible yeast was Trichophyton rubrum and the best antifungal activity was shown by Xanthosoma sagittifolium supernatant. The bioautography was performed only for the aqueous extracts and supernatants of those plants that showed antifungal activity against Candida albicans and Cryptococcus neoformans, using n-butanol/acetic acid/water (BAW) 8:1:1 to develop silica gel TLC plates. Clear inhibition zones were observed for aqueous extracts of Schinus molle (R(f) 0.89) and Schinus terebinthifolius (R(f) 0.80) against Candida albicans, as for supernatant of Anacardium occidentale (R(f) 0.31) against Cryptococcus neoformans. The separation of macromolecules from metabolites, as in the case of Anacardium occidentale, Solanum sp. and Xanthosoma sagittifolium, enhances antifungal activity. In other cases, the antifungal activity is destroyed, as observed for Momordica charantia, Schinus molle and Schinus terebinthifolius.

This is the most cited article group ( average of 10 citations / year ) and was one of the magazine's Top 25 articles Journal of Ethnopharmacology in the year of its publication. The article focused on the use of a strategy adopted in our laboratory to facilitate biomonitored fractionation of a plant extract which has been greatly appreciated in the scientific community . The ethanol precipitation to divide a crude extract in two fractions, each with a different chemical profile and hence a biological activity profile also different. The adoption of this strategy has been very helpful in saving time of the steps of a biomonitored project.



Muzitano MF, Cruz EA, de Almeida AP, Da Silva SA, Kaiser CR, Guette C, Rossi-Bergmann B, Costa SS. Quercitrin: an antileishmanial flavonoid glycoside from Kalanchoe pinnata. Planta Medica, v. 72,  n1, p. 81-83, 2006.
Abstract: Quercitrin (quercetin 3-O-alpha- L-rhamnopyranoside), one of the constituents of the biologically active aqueous extract obtained from Kalanchoe pinnata, is demonstrated to be a potent antileishmanial compound (IC (50) approximately 1 microgram/mL) with a low toxicity profile. This is the first time that antileishmanial activity is demonstrated for a flavonoid glycoside.
Este artigo faz parte de uma série de publicações da linha de pesquisa desenvolvida na busca de substâncias potencialmente terapêuticas para leishmaniose, uma doença negligenciada, que afeta milhões de pessoas no mundo. Os estudos foram realizados em modelo de leishmaniose cutânea em camundongos infectados com Leishmania amazonensis em parceria com a equipe da Profa. Bartira Rossi-Bergmann (IBCCF - UFRJ).



Muzitano MF, Tinoco LW, Guette C, Kaiser CR, Rossi-Bergmann B, Costa SS. The antileishmanial activity assessment of unusual flavonoids from Kalanchoe pinnata. Phytochemistry, v. 67, n. 18, p. 2071-2077, 2006.

Abstract: The importance of flavonoids for the antileishmanial activity of Kalanchoe pinnata was previously demonstrated by the isolation of quercitrin, a potent antileishmanial flavonoid. In the present study, the aqueous leaf extract from the medicinal plant K. pinnata (Crassulaceae) afforded a kaempferol di-glycoside, named kapinnatoside, identified as kaempferol 3-O-α-L-arabinopyranosyl (1 → 2) α-L-rhamnopyranoside (1). In addition, two unusual flavonol and flavone glycosides already reported, quercetin 3-O-α-L-arabinopyranosyl (1 → 2) α-L-rhamnopyranoside (2) and 4',5-dihydroxy-3',8-dimethoxyflavone 7-O-β-D-glucopyranoside (3), have been isolated. Their structures were determined via analyses of mono and bi-dimensional 1H and 13C NMR spectroscopic experiments and HR-MALDI mass spectra. Because of its restricted occurrence and its abundance in K. pinnata, flavonoid (2) may be a chemical marker for this plant species of high therapeutic potential. The three flavonoids were tested separately against Leishmania amazonenis amastigotes in comparison with quercitrin, quercetin and afzelin. The quercetin aglycone - type structure, as well as a rhamnosyl unit linked at C-3, seem to be important for antileishmanial activity.

In this article it described the structure of a novel flavonoid isolated from the aqueous extract of K. pinnata and the report of two other unusual flavonoids . Flavonoids were tested separately in Leishmania amazonensis amastigotes compared to quercitrin , quercetin and afzelin previously isolated . The study allowed to advance understanding of the role of flavonoid K. pinnata in in vitro assay of leishmaniasis .



Muzitano MF, Bergonzi MC, De Melo GO, Lage CLS, Bilia AR, Vincieri FF, Rossi-Bergmann B, Costa SS. Influence of cultivation conditions, season of collection and extraction method on the content of antileishmanial flavonoids from Kalanchoe pinnata. Journal of Ethnopharmacology, v. 133, p. 132-137, 2011.

Abstract: Ethnopharmacological relevance: Leaves from Kalanchoe pinnata (Lamarck) Persoon (Crassulaceae) are popularly used for healing wounds. Its antileishmanial properties are established in experimental animals, and its active flavonoid components have been identified. Aim of the study: In this study, we attempted to standardize the extract from K. pinnata leaves by evaluating the influence of season of harvest, sunlight exposure and method of extraction on antileishmanial flavonoids content. Materials and Methods: HPLC-DAD-MS was used to identify and quantify the active antileishmanial flavonoids in different extracts. ANOVA test for analyses of variance followed by the Tukey test of multiple comparisons were used in the statistical analysis. The antileishmanial potential was assessed by the activation of nitric oxide production by murine macrophage using the Griess method. Results: We demonstrated that active flavonoids were significantly more abundant when the leaves were collected in the summer, and that aqueous extraction at 50 °C allowed the highest flavonoid extraction. The benefit of sunlight exposure was confirmed in plants cultivated under direct sunlight when compared with those that grown under shade. Under sunny conditions the yield of the most active antileishmanial favonoid quercitrin was increased by 7-fold. All aqueous extracts tested were capable to enhance the macrophage nitric oxide production. However, hot aqueous extract from leaves collected in summer exhibited the higher activity, in agreement with HPLC-DAD-MS analysis tendency. In addition, with the aim of reducing the individual chemical variations of the plant constituents and optimizing the production of the active extract, it was obtained in vitro monoclonal KP specimens that were easily adapted to field conditions and were able to produce antileishmanial flavonoids. Conclusion: Our study reports the better conditions of cultivation, harvest and extraction protocol for obtaining a K. pinnata extract exhibiting the highest antileishmanial activity. Additionally, we propose the flavonoids quercetin 3-0-α-L-arabinopyranosyl (1 → 2)-α-L-rhamnopyranoside and quercitrin, as satisfactory chemical markers for standardization purposes.


This article represents an important step towards a possible application of K. pinnata in the treatment of leishmaniasis. The study had the participation of researchers from the University of Florence (Italy ) in the section on the analysis of HPLC -DAD -MS. It is reported the seasonal effects on the production of bioactive flavonoid , the best extraction method thereof and the production of monoclonal seedlings to obtain plant material with a controlled profile of production of flavonoids with anti- leishmania action. Although the article was published just over two years , it has been cited 8 times in publications , according to the Scopus .

De Almeida AP, Miranda MMFS, Simoni IC, Wigg MD, Lagrota MHC, Costa SS. Flavonol monoglycosides isolated from the antiviral fractions of Persea americana (Lauraceae) leaf infusion. Phytotherapy Research, v. 12, p. 562-567, 1998.
Abstract: An infusion of Persea americana leaves (Lauraceae) strongly inhibited herpes simplex virus type 1 (HSV-1), Aujeszky's disease virus (ADV) and adenovirus type 3 (AD3) in cell cultures. Its fractionation, guided by anti-HSV-1 and ADV assays, allowed the isolation and identification of two new flavonol monoglycosides, kaempferol and quercetin 3-O-α-D-arabinopyranosides, along with the known kaempferol 3-O-α-L-rhamnopyranoside (afzelin), quercetin 3-O-α-L-rhamnopyranoside (quercitrin), quercetin 3-O-β-gluco-pyranoside and quercetin. The known quercetin 3-O-β-galactopyranoside was identified in a mixture. Afzelin and quercetin 3-O-α-D-arabinopyranoside showed higher activity against acyclovir-resistant HSV-1. Chlorogenic acid significantly inhibited the HSV-1 replication without any cytotoxicity. However, all the substances tested were less active than the infusion or fractions. The same substances did not affect ADV replication. Chemical structures were elucidated by the analysis of UV, 1H and 13C NMR data, mainly by APT, homo and heteronuclear COSY experiments. The configuration of the D-arabinose unit, not usual in natural plant products, was established on the basis of the optical rotation of the free sugar obtained after acid hydrolysis.

This was one of the first articles of our group focused on obtaining bioactive substances in a biomonitored study of a plant aqueous extract . The study was part of an interdisciplinary project in partnership with a team of virology of IMPPG - UFRJ and the Biological Institute of São Paulo. The fractionation guided by HSV- 1 and adenovirus type 3 trials led to the isolation and identification of phenolic compounds with high inhibitory activity of HSV-1 acyclovir-resistant . Inhibition of drug -resistant virus in clinical use in cases of herpes infections makes the flavonoids and the leaf extract of the avocado tree ( Persea americana ) a potential source for development of new drugs .

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