Piper betel leaf extract: anticancer benefits and bio-guided fractionation to identify active principles for prostate cancer management

Abstract

Plant extracts, a concoction of bioactive non-nutrient phytochemicals, have long served as the most significant source of new leads for anticancer drug development. Explored for their unique medicinal properties, the leaves of Piper betel, an evergreen perennial vine, are a reservoir of phenolics with antimutagenic, antitumor and antioxidant activities. Here, we show that oral feeding of betel leaf extract (BLE) significantly inhibited the growth of human prostate xenografts implanted in nude mice compared with vehicle-fed controls. To gain insights into the 'active principles', we performed a bioactivity-guided fractionation of methanolic BLE employing solvents of different polarity strengths using classical column chromatography. This approach yielded 15 fractions, which were then pooled to 10 using similar retention factors on thin-layer chromatographs. Bioactivity assays demonstrated that one fraction in particular, F2, displayed a 3-fold better in vitro efficacy to inhibit proliferation of prostate cancer cells than the parent BLE. The presence of phenols, hydroxychavicol (HC) and chavibetol (CHV), was confirmed in F2 by nuclear magnetic resonance, high-performance liquid chromatography and mass spectroscopy. Further, the HC containing F2 subfraction was found to be ~8-fold more potent than the F2 subfraction that contained CHV, in human prostate cancer PC-3 cells as evaluated by the 3-(4,5-dimethylthiazole-2-yl)-2,5-diphenyl tetrazolium bromide assay. Removing CHV from F2 remarkably decreased the IC50 of this fraction, indicating that HC is perhaps the major bioactive constituent, which is present to an extent of 26.59% in BLE. These data provide evidence that HC is a potential candidate for prostate cancer management and warrants further preclinical evaluation.

Piper Betel Leaf: A Reservoir of Potential Xenohormetic Nutraceuticals with Cancer-Fighting Properties

“…Other betel phytochemicals include allylpyrocatechol (APC; 2-hydroxychavicol), 4-hydroxycatechol, b-caryophyllene, methyl eugenol, carotenes, starch, diastases, and an essential oil containing hydroxychavicol (18,19). Hydroxychavicol, a phenolic compound quantitatively present at approximately 26% in betel leaves, has been shown to exert antiproliferative activity in prostate cancer (20). Hydroxychavicol has also been shown to impede cell-cycle progression of prostate cancer and oral KB carcinoma cells (20)(21)(22).…”

“…Hydroxychavicol, a phenolic compound quantitatively present at approximately 26% in betel leaves, has been shown to exert antiproliferative activity in prostate cancer (20). Hydroxychavicol has also been shown to impede cell-cycle progression of prostate cancer and oral KB carcinoma cells (20)(21)(22). Several reports indicate hydroxychavicol as an antimutagenic agent (12,23) as well as an effective inhibitor of cyclooxygenase (COX), platelet calcium signaling, and thromboxane B2 production (21).…”

“…Chavibetol (CHV), along with hydroxychavicol, acts as a radioprotectant (25), and exhibits substantial immunomodulatory and free radical scavenging activities (25). We have shown that CHV synergizes with hydroxychavicol to exert antiproliferative activity against human prostate cancer PC-3 cells (20). Betel leaf contains large amounts of safrole, which is rapidly degraded in the human body into dihydroxychavicol and eugenol, the antimutagenic agents, which are then excreted via the urine (26,27).…”

“…Betel leaf nutraceuticals target multiple cellular events and play a crucial role in inhibiting tumor growth, inflammation by decreasing the levels of cyclooxygenases, COX-1 and COX-2 (21) and stress-inducing events like lipid peroxidation. Furthermore, they activate detoxifying enzymes and upregulate the expression of ABCA1 (ATP-binding cassette transporter) and Liver X receptors, while inhibiting LDL oxidation (62), increase free radical scavenging, and induce apoptosis signified by increased levels of cleaved caspase-3 and PARP levels (20,63). The betel leaf constituents kill cancer cells by inducing oxidative DNA damage demonstrating their pro-oxidant role (29).…”

“…It was observed that when betel leaf extract was fed to rats bearing mammary tumors in the initiation phase, the tumor growth was prevented, while rats with fully developed mammary tumors failed to show any reduction/inhibition in tumor growth (52,53). We have recently reported that betel leaf extract exhibits significant efficacy in inhibiting growth of prostate cancer xenografts (20). These studies generate compelling foregrounds to warrant future investigation of chemopreventive efficacy of betel leaves and/or bioactive betel leaf constituents in appropriate animal models.…”