Bio-guided fractionation of the antimutagenic activity of methanolic extract from the fruit of Randia echinocarpa (Sessé et Mociño) against 1-nitropyrene
Highlights
► Bio-guided characterization of antimutagenic compounds of Randia echinocarpa fruit. ► Methanolic extract shows weak antimutagenicity. ► Hexanic fraction of the methanolic extract is a strong antimutagen. ► The main components are β-sitosterol and palmitic and linoleic acids. ► The fruit of Randia echinocarpa could be considered as nutraceutical.
Introduction
Nowadays, cancer disease represents one of the most important public health concerns. This illness of multifactor etiology can affect every organ and system in the body, and the symptomatology and treatment responses are highly variable. Consequently, early detection and prevention are the best strategies for disease control and new therapeutic/preventive agents are needed (Thun, DeLancey, Center, Jemal, & Ward, 2010).
Plants are the main source of bioactive compounds (Cragg, Grothaus, & Newman, 2009), especially of anticancer compounds. Mexico has a large floristic diversity but it is practically unknown (SEMARNAT, 2008).
Randia is considered as a Neotropical genus with 60 to 70 species; it is distributed from Southern United States to South America and Mexico represents the area of the highest diversity with 33 species (Bye et al., 1991, Lorence and Nee, 1987).
Randia echinocarpa Sessé et Mociño (Rubiaceae) is a shrub or small tree of 2–6 m high, distributed along the Pacific Ocean Shore of Mexico. The fruit of R. echinocarpa is known as “papache”. The pulp of the ripe fruit is dark, with a large number of rounded seeds (see Fig. S1 as supplementary data), the isolated compounds from the fruit of R. echinocarpa are mannitol, β-sitosterol, and quinovic, oxoquinovic, ursolic and oleanolic acids (Bye et al., 1991). R. echinocarpa fruit has a sweet/bitter flavor, it is mainly consumed by people of rural areas as food/medicine, and only commercialized in Mexican traditional markets (Cortés, 2000).
R. echinocarpa has been employed in Mexican traditional remedies for cancer, malaria and diabetes; as well as for kidney, pulmonary, circulatory and gastro-intestinal ailments (Bye et al., 1991, Cortés, 2000). These ethnopharmacological properties have not been scientifically confirmed. On the other hand, a positive effect has been experimentally demonstrated for an aqueous extract of the fruit, it improves the cicatrizing speed in skin of wounded rats (Pérez, Pérez, Pérez-González, & Vargas, 1993); negative effects are also registered for the oral administration of this extract in rats, increased diuretic activity and urolith formation (Vargas Solis & Pérez Gutiérrez, 2002). In a previous report, an aqueous extract of R. echinocarpa fruit (500 μg/tube) showed a moderate antimutagenic activity in the microsuspension assay with YG1024 strain, PI of 32 and 53% at 50 and 100 ng/tube of 1-NP, respectively; remarkably, the extract acts by desmutagenic and bioantimutagenic mechanisms (Santos-Cervantes, Ibarra-Zazueta, Loarca-Pina, Paredes-Lopez, & Delgado-Vargas, 2007).
R. echinocarpa plants were widely distributed along the State of Sinaloa and nowadays their populations have been diminished. Based on the traditional uses of R. echinocarpa, it could be considered an underutilized plant and an improved knowledge about its chemical composition and biological activities will contribute to use this natural resource under conditions of ecological sustainability (Jaenicke & Höschle-Zeledon, 2006).
The objective of this study was to characterize the responsible compounds of the antimutagenicity in the fraction with the highest activity obtained from the crude methanolic extract of R. echinocarpa fruit (ME).
Section snippets
Plant material
The R. echinocarpa fruits were collected from the municipality of Badiraguato, Sinaloa, México. The plant material was identified and a voucher specimen (var 9035) was deposited in the herbarium of the Faculty of Agronomy, Autonomous University of Sinaloa.
Seeds were eliminated and fruit pulp was freeze dried (VirTis 25 EL, VirTis Co., Gardiner, NY). Dried pulp, highly hygroscopic, was crushed by hand and immediately frozen at − 80 °C until required.
Chemical reagents
Silica gel 60 (particle size 0.035–0.070 mm,
Results
The yield for the ME was high (80.67%) and liquid–liquid partition produced a higher yield of polar (AqF, 76.40%) than non polar fractions (Fig. 1).
In agreement with the dose–response curve for the 1-NP mutagenicity against YG1024, doses of 50 and 100 ng/tube of 1-NP were chosen for the antimutagenicity assays (Fig. 2). 1-NP was not toxic in the range of concentrations tested; a uniform lawn of bacteria (micro-colonies) was observed in every plate of the mutagenic assay and the number of
Discussion
In a previous report, we established the antioxidant and antimutagenic activity of an aqueous extract of R. echinocarpa fruit, the antimutagenic PI at 500 μg/tube was 32% (50 ng/tube 1-NP) and 53% (100 ng/tube 1-NP) (Santos-Cervantes et al., 2007). These values were lower than the registered in this study for the HF, 75% and 84%, respectively. In addition, CF and EAF were positive antimutagens (Table 2) with values similar to those reported for the aqueous extract of papache fruit (
Conclusions
The fruit of papache contains compounds with antimutagenic activity. The antimutagenicity was highest in the non-polar fractions (HF, HF1–1 and HF1–2) which were enriched with sterols (campesterol and β-sitosterol, HF1–1) and fatty acids (palmitic and linoleic, HF1–2). Previous reports have shown antimutagenicity, antitumoral and anticarcinogenic properties for these compounds; thus, papache antimutagenicity could be associated with these sterols and fatty acids. R. echinocarpa fruit has been
Abbreviations
- 1-NP
1-nitropyrene
- AFB1
aflatoxin B1
- AqF
aqueous fractions
- CF
chloroformic fraction
- DMAB
3,2′-dimethyl-4-aminobiphenyl
- DMSO
dimethylsulfoxide
- EA
ellagic acid
- EAF
ethyl acetate fraction
- EtOAc
ethyl acetate
- HAE
hot aqueous extract of Hibiscus sabdariffa
- HF
hexanic fraction
- ME
crude methanolic extract of papache
- MeChlF
methylene chloride fraction of the methanol extract of Gleditsia sinensis thorns
- MeOH
methanol
- MI
mutagenic index
- MNNG
N-methyl-N′-nitro-N-nitroguanidine
- NPD
4-nitro-o-fenilenediamina
- PI
percentage of inhibition of
Acknowledgments
Authors acknowledge the financial support from Consejo Nacional de Ciencia y Tecnologia (CONACyT), Consejo Estatal de Ciencia y Tecnologia del Estado de Sinaloa (CECyT) and from Universidad Autónoma de Sinaloa (PROFAPI); the identification of R. echinocarpa by Dr. Rito Vega-Aviña, Faculty of Agronomy, Universidad Autónoma de Sinaloa (UAS); and technical assistance by Dr. José Ángel López-Valenzuela.
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