Bioactivity-guided isolation and structural characterization of the antifungal compound, plumbagin, fromNepenthes gracilis

Abstract

Context: Despite several phytochemical studies of Nepenthes gracilis Korth (Nepenthaceae), the biological activities of this pitcher plant remain to be explored. Objective: This study evaluates the antifungal activity of N. gracilis extracts, isolates, and characterizes its bioactive compound and evaluates the cytotoxicity of the isolated compound. Materials and methods: Fresh leaves of N. gracilis were sequentially extracted. The fungistatic and fungicidal activities of the extracts were evaluated against six species of fungi of medical importance using a colorimetric broth microdilution method. The most active extract was fractionated by liquid-liquid partitioning and further purified by a preparative thin layer chromatography. Structural elucidation was carried out using FT-IR, GC-MS, and NMR. Cytotoxicity testing against rhesus monkey kidney epithelial cells (LLC-MK2) was assessed by a neutral red uptake (NRU) assay. Results: The hexane extract, which showed the lowest minimum inhibitory concentration (MIC) and minimum fungicidal concentration (MFC), both at 20 lg/mL against Candida albicans, Issatchenkia orientalis, and Trichophyton mentagrophytes, was subjected to bioactivity-guided fractionation. The isolated compound exhibited potent activity with the MIC values ranging from 2 to 31 lg/mL against all the fungi. The active compound was identified as plumbagin (5-hydroxy-2-methyl-naphthalene-1,4-dione). The 50% cytotoxicity concentration (CC 50 ) of plumbagin was 0.60 lg/mL. Discussion and conclusion: The selectivity indices of plumbagin against all the fungi were less than 1.0, indicating that plumbagin is more toxic to mammalian than fungal cells. This study provides information on the antifungal properties of N. gracilis leaf extracts, as well as the antifungal and cytotoxicity properties of plumbagin.

Ethyl acetate extract of Nepenthes adrianii x clipeata induces antiproliferation, apoptosis, and DNA damage against oral cancer cells through oxidative stress

“…Several kinds of solvent extractions of Nepenthes plants showed diverse biological activities. For example, methanolic extract of N khasiana leaf and hexane extract from N gracilis displayed antifugal activity. Ethanolic extract of N thorelii roots displayed antimalarial effect .…”

“…In China, N mirabilis is regarded as a TCM for the treatment of cough, fever, hypertension, and urinary system infection . Recent pharmacological studies revealed the extracts of Nepenthes plants have antibacterial, anti‐inflammatory, antifungal, antimalarial, and α‐glucosidase inhibitory activities. Furthermore, phytochemical studies reported that the major constituents of Nepenthes plants included flavonoids, naphthalene glucosides, naphthoquinones, steroids, triterpenoids, and phenolic compounds .…”

Ethnobotanical, Phytochemical, and Pharmacological Properties of Nepenthes Species: A Review

“…The compound plumbagin isolated showed strong antifungal activity with the MIC values between 2 and 31 µg/ml against all of the tested fungi [32]. The pitcher liquid extract of N. khasiana contained droserone, 5-0-methyldroserone, and purified naphthoquinones which exhibited antifungal activities when tested against Aspergillus sp.…”

“…The viability of LLC-MK2 cells treated with different plumbagin concentrations were measured by NRU assay. Moreover, a 50% cytotoxicity concentration of plumbagin at 0.60 µg/ml was observed [32]. Acute cytotoxicity study showed that the N. khasiana methanolic extract of leaf neither demonstrated any toxicity nor mortality at a 3000 mg/kg bw dose level in rats [13].…”

“…A liquid from the pitcher of N. khasiana was found to contained 5-0-methyldroserone and droserone [31]. Plumbagin was isolated from the fresh leaves of N. gracilis [32]. A phytochemical analysis of pitcher and leaf extract of N. khasiana Hook showed the existence of flavonoids, glycoside, tannins, alkaloids, phytosterols, and saponins [13,23,24].…”