Estrogenic and mutagenic activities of Crotalaria pallida measured by recombinant yeast assay and Ames test
TL;DR: Considering the excellent estrogenic activity performed by stigmasterol in the RYA associated with the absence of mutagenic activity when evaluated by the Ames test, stig masterol becomes a strong candidate to be used in hormone replacement therapy during menopause.
Abstract: Crotalaria pallida Ailton is a plant belonging to the Fabaceae family, popularly known as “rattle or rattlesnake” and used in traditional medicine to treat swelling of the joints and as a vermifuge. Previous pharmacological studies have also reported anti-inflammatory, antimicrobial and antifungal activities. Nevertheless, scientific information regarding this species is scarce, and there are no reports related to its possible estrogenic and mutagenic effects. Thus, the purpose of the present study was to investigate the estrogenic potential of C. pallida leaves by means of the Recombinant Yeast Assay (RYA), seeking an alternative for estrogen replacement therapy during menopause; and to reflect on the safe use of natural products to assess the mutagenic activity of the crude extract from C. pallida leaves, the dichloromethane fraction and stigmasterol by means of the Ames test. The recombinant yeast assay with the strain BY4741 of Saccharomyces cerevisiae, was performed with the ethanolic extract, dichloromethane fraction and stigmasterol isolated from the leaves of C. pallida. Mutagenic activity was evaluated by the Salmonella/microsome assay (Ames test), using the Salmonella typhimurium tester strains TA100, TA98, TA97 and TA102, with (+S9) and without (-S9) metabolization, by the preincubation method. All samples showed estrogenic activity, mainly stigmasterol. The ethanolic extract from C. pallida leaves showed mutagenic activity in the TA98 strain (-S9), whereas dichloromethane fraction and stigmasterol were found devoid of activity. Considering the excellent estrogenic activity performed by stigmasterol in the RYA associated with the absence of mutagenic activity when evaluated by the Ames test, stigmasterol becomes a strong candidate to be used in hormone replacement therapy during menopause.
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TL;DR: A comprehensive summary of more than 450 estrogenic chemicals including estrogenic endocrine disruptors is provided here to understand the complex and profound impact of estrogen action.
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...Stigmasterol (E) ER/Traditional medicine Boldrin et al. (2013) (R)...
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Cites background from "Estrogenic and mutagenic activities..."
...The raw leaves are very useful for abortion since it has stigmasterol which affects estrogenic activity (Boldrin et al., 2013)....
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TL;DR: The leaves of Tridax procumbens were screened for the presence of bioactive molecules and the results showed that they had high flavonoids, alkaloids, hydroxycinnamates, tannins and phytosterols, moderate benzoic acid derivatives and lignans, and low carotenoids contents as discussed by the authors.
Abstract: The leaves of Tridax procumbens were screened for the presence of bioactive molecules. They had high flavonoids, alkaloids, hydroxycinnamates, tannins and phytosterols, moderate benzoic acid derivatives and lignans, and low carotenoids contents. Thirty nine known alkaloids (mainly akuammidine, 68.756%), twenty three known flavonoids (mainly 17.593% kaempferol and 12.538% (-)-epicatechin), five known carotenoids (mainly lutein, 62.608%), four known benzoic acid derivatives (mainly ferulic acid, 46.091%), two phytosterols (mainly stigmasterol, 80.853%) and six known lignans (mainly galgravin, 77.326%) were detected. Also detected were caffeic acid and tannic acid. The medicinal properties of the flavonoids, phytosterols, alkaloids, tannins, hydroxicinnamates, carotenoids, benzoic acid derivatives and lignans that were present in the leaves were discussed herein and proposed to be explored for their potential medicinal values. The great number of potentially active nutrients and their multifunctional properties make Tridax procumbens a perfect candidate for the production of health-promoting food and food supplements.
28 citations
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...Stigmasterol has anti-osteoarthritic, anti-hypercholestrolemic, cytotoxicity, antitumor, thyroid inhibiting, hypoglycemic, anti-mutagenic, antioxidant, anti-inflammatory, anticonvulsant and sedative [69], chemopreventive, anticancer, estrogenic [70] activities....
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TL;DR: The findings demonstrate the utility of the yeast recombinant assay for analyzing complex natural samples and stress the necessity of a panel of different yeast systems to adequately describe endocrine‐disruptor activities.
Abstract: Endocrine disruptors exert physiological effects at very low concentrations. Surface waters present often a mixture of high concentrations of low-potency disruptors and low amounts of very powerful ones, making their chemical analysis complicated and expensive. We developed a recombinant yeast assay (RYA) for estrogenic compounds using 96-well microtiter plates. This assay is based on three yeast strains, transformed with self-propagating plasmids. One strain contains an expression plasmid for the human estrogen hormone receptor and an appropriate reporter; it detects estrogenic and antiestrogenic activities. The two other yeast strains, one expressing the human progesterone receptor and a second based on the yeast activator Gal4p, served to analyze the nature of antiestrogenic activities. We applied this technique to water samples from two tributaries of the Llobregat river (NE Spain) as well as from four sewage treatment plants discharging on them. Our results indicate that the efficiency of sewage treatment plants for eliminating estrogenic compounds varied notably, being in at least one case completely inefficient. We also observed a prevalence of an inhibitory activity all through the two rivers; this inhibition was hormone specific. These results were consistent to previously obtained chemical analyses of the same samples, although chemical and in vivo analyses showed rather different levels of sensitivity for some compounds. Our findings demonstrate the utility of the yeast recombinant assay for analyzing complex natural samples; at the same time, they stress the necessity of a panel of different yeast systems to adequately describe endocrine-disruptor activities.
158 citations
"Estrogenic and mutagenic activities..." refers methods in this paper
...Benjamin Piña (CSIC, Barcelona, Spain), was transformed together with plasmids pH5HE0 and pVitBX2 [15]....
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...pallida leaves was investigated using the RYA (Recombinant Yeast Assay), in which recombinant yeast is utilized as an experimental model and the transcription of a reporter gene depends on the presence in the midst of compounds capable of binding to the estrogen receptor [15]....
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TL;DR: This chapter discusses the yeast transcription cassettes of several types used for the expression of cloned genes, which include vectors with regulatable promoters and protein fusions to coding region for another protein, such as β-galactosidase.
Abstract: Publisher Summary It is necessary to separate a gene from its own promoter to control expression of its product in different conditions or to produce the product to a greater or lesser degree than normal. This chapter discusses the yeast transcription cassettes of several types used for the expression of cloned genes. The first type includes vectors with regulatable promoters; GAL, PGK, and ADH1 respond to the carbon source, and PH05 responds to the inorganic phosphate concentration. These vectors allow the expression of the cloned gene to be turned on and off by changes in the growth medium. Another class creates a protein fusion to an amino-terminal signal sequence so that the protein are secreted from the cell. A third kind of cassette makes protein fusions of the cloned gene to the coding region for another protein, such as β-galactosidase, or to a small antigenic epitope. Therefore, proteins for which there is no antibody or convenient assay available can be tagged with a heterologous enzymatic activity or antigen.
141 citations
"Estrogenic and mutagenic activities..." refers background in this paper
...The expression plasmid pH5HE0 contained the human estrogen hormone receptor gene HE0 [19], cloned into the constitutive yeast expression vector pAAH5 [20]....
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01 Jan 2011
TL;DR: The aim of this study is to identify and characterize the bioactive principle from the aerial parts of the plant, which has wide folk medicinal use.
Abstract: General phytochemical screening of the aerial parts of Ageratum conyzoides (Asteraceae) revealed the presence of steroids, terpenes, phenolic compounds, saponins, fatty acids, alkaloids. The aim of this study is to identify and characterize the bioactive principle from the aerial parts of the plant. It has wide folk medicinal use. For isolation of the compound, the dried aerial parts powder of Ageratum conyzoides was subjected to hot extraction with petroleum ether; this extract was saponified with alcoholic KOH and subjected to chromatography. Isolated compound were purified by chloroform. The isolation and purification afforded white crystalline powder which was subjected to physical, chemical and spectral identification by IR, 1H‐NMR, 13C‐NMR and GC‐MS. The compound was concluded as stigmasterol and β‐sitosterol.
139 citations
"Estrogenic and mutagenic activities..." refers background in this paper
...The 1H NMR spectrum showed characteristic signals of hydrogen steroid [18]....
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...records in the literature [18] allowed us to identify it as stigmasterol....
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TL;DR: In vitro concentrations of genistein >5 microM are defined as non-physiological, and thus "high" doses, in contrast to much of the previous literature, in order to distinguish genotoxic versus potentially beneficial in vitro effects of natural dietary products such as Genistein.
137 citations
"Estrogenic and mutagenic activities..." refers background in this paper
...These constituents are also related to the reduction of cancer incidence, particularly estrogen-dependent cancers, such as breast cancer, cardiovascular diseases and osteoporosis [11-14]....
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Patent•
26 Jan 2001
TL;DR: In this article, the human nuclear hormone receptors (NHREC) and polynucleotides which identify and encode NHREC were used for diagnosing, treating or preventing disorders associated with aberrant expression of NHREC.
Abstract: The invention provides human nuclear hormone receptors (NHREC) and polynucleotides which identify and encode NHREC. The invention also provides expression vectors, host cells, antibodies, agonists, and antagonists. The invention also provides methods for diagnosing, treating or preventing disorders associated with aberrant expression of NHREC.
109 citations