Murraya koenigii is a well-known Indian medicinal herb, and a carbazole alkaloid (mahanine) from this plant causes apoptosis in cancer cells. Here, we investigated how seasonal and geographical variations influence carbazole alkaloids composition and medicinal property of this plant against cancer cells in vitro and in vivo. Leaflets were collected from various places in different seasons for three years. A mahanine-enriched fraction (MEF) was prepared in two steps using ethanol and water. The best plant was selected based on the highest percent of mahanine. MEF prepared from leaflets of nine different locations showed a different concentration of identified markers (mahanine, mahanimbine, and koenimbine) which exhibited differential reduced metabolic activity against ovarian cancer, mahanine being the best. Our systematic study revealed that mahanine content was highest during September–December. Interestingly, MEF from southern part (tropical zone) exhibited 43 ± 2.5% mahanine compared to 2.7 ± 1.3% in northeastern part (subtropical zone) with five folds higher activity against PA1. Moreover, MEF reduced metabolic activity of sixteen cancer cell lines from nine different origins and significantly reduced tumor mass in lung and ovarian cancer xenograft models. Taken together, this is the first report demonstrating the marker’s content in these leaflets is highly dependent on location/season. A positive correlation between biological activity and mahanine concentration was established in MEF. Such a comprehensive study suggests that the selection of location and suitable season for collection of any plant materials with biologically active stable markers in sufficient quantity play a decisive role in determining the fate of their medicinal property.
Murraya koenigii is well documented in the Indian ancient medical text “Charaka Samhita.” The carbazole alkaloid “mahanine” from this plant exhibited anticancer activity against several cancers. Here, we have taken a comprehensive study to standardize the method for the preparation of a mahanine-enriched fraction (MEF) with the highest yield and defined markers. Our optimized method produced MEF having the highest amount of mahanine, a major marker, with excellent in vitro antiproliferative activity against ovarian and breast cancer cells as evidenced by decreased cell viability by MTT assay. Moreover, it exhibited condensed and fragmented nuclei by DAPI staining and increased annexin V-/PI-stained cells after MEF treatment, indicating apoptosis. It also exhibited good efficacy in ovarian and breast cancer syngeneic mice models, with an ED50 of 300 mg/kg body weight (BW). MEF is stable up to 40°C for ≥3 months. Its biological activity remains unchanged at a wide range of pH (1-10) for up to ~3 hours, indicating a safe oral route of administration. Additionally, the comparative pharmacokinetics of MEF and mahanine in rats showed a 31% higher bioavailability of mahanine in MEF-fed rats compared to rats fed with mahanine alone. Furthermore, mice fed with MEF at 5000 mg/kg BW single dose, 300-1500 mg/kg BW/day for 14 days, and 300 mg/kg BW/day for 28, 90, and 180 days for subacute, subchronic, chronic studies, respectively, did not show any significant clinical signs of toxicity, behavioral changes, mortality, organ weights, serum biochemistry, and hematological parameters indicating no/minimum toxicity for up to 180 days. To the best of our knowledge, this is the first report showing the pH/temperature stability and chronic toxicity studies of MEF along with in vivo efficacy against breast cancer. Taken together, our study will enhance the commercial value of this highly potential medicinal plant and will be helpful as a reference material for its clinical development.
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