Anticancer potential of musa × paradisiaca as cervical carcinoma and malignant melanoma

This study was performed to assess the anticancer potential of Musa × pardisiaca L. It has several traditional benefits due to the presence of various bioactive compounds. Ethyl acetate extract of leaves was prepared and characterized by Gas Chromatography-Mass spectrometry (GC-MS). Compounds ( 1-40 ) were isolated from leaves, out of which two major compounds i.e. eugenol and phytol are potentially responsible for anticancer activity and oxidative stress. In vitro cytotoxicity of extract showed significant inhibition of cells in cervical carcinoma and malignant melanoma when treated by MTT assay against Hela and A375 cell lines. Ethyl acetate extract of musa × paradisiaca L. shown anticervical carcinoma and anti-malignant melanoma activity in our study.


Introduction
Carcinoma cervix is a type of cancer that occurs in the cells of the cervix, the lower part of the uterus that connects it to the vagina. The incidence and mortality rate due to this cancer have declined in the developed countries over past few decades but still remains to be the most common gynecological cancer and fourth most common malignancy in women across developing countries [1]. In India, it is second most common cancer in females after breast cancer, with around a lakh new cases identified annually [2]. The highest incidence was recorded in Papumpare, Aizawl, Mizoram, accounting for a total of 27% of cancer cases amongst female in India [3,4]. It accounted for 0.3% of all cancer patients and 27% of all female cancer patients seen [4]. Infection with human papillomavirus (HPV) is associated with more than 90% cases of cervix cancer [5].
Surgery, radiotherapy or chemotherapy, alone or in combination [6], can be used as therapeutic modalities for carcinoma cervix [7]. Malignant melanoma is the most dangerous form of skin cancer [8], being fifth most common tumor type among men and sixth in women in the United States [9]. In addition, malignant melanoma is 1.4 times more common among men than in women. The incidence of both non-melanoma and melanoma skin cancers have been increasing over the past decades and are reported maximum in Australia, New Zealand, Switzerland, Denmark and Norway [10]. Ultraviolet (UV) radiation and number of moles more than 100 is the greater risk for melanoma [11].
Surgery is the definitive treatment depending on number of factors, including the type of cancer, size of the lesion, anatomic location, available resources and patient preferences for early-stage melanoma, keeping or leaving medical management generally reserved as adjuvant treatment for advanced melanoma [12]. Moreover, nowadays doctors are exploring plant extracts to treat cancer, hypothesizing their traditional use because they can be used even in palliative care as well as to reduce the side effects associated with cancer treatment. The National Cancer Institute (NCI) has collected about 35,000 plant samples from 20 countries and has screened around 114,000 extracts for anticancer activity. Over 3000 species of plants with potential antitumor activity have been reported [13]. Approximately 60% of anticancer compounds and 75% drugs for infectious disease come from natural products or their derivatives [14]. One of them is Musa × paradisiaca L.
commonly known as Plantain belonging to family musaceae (banana family) [15]. Native to the Mediterranean region. It possesses several medicinal properties which include antilithiatic, antibacterial, antidiabetic, antiulcer, anti-diarrheal, hypocholesterolaemic, hepatoprotective, anti-snake venom, wound healing, hair growth promoting, anti-fungal and anti-menorrhagia activity [16]. due to the presence of anti-cancer activity Musa × paradisiaca L can be prove effective against cervical carcinoma and malignant melanoma [17].

Evaluation of antioxidant activity by DPPH radical scavenging assay
The antioxidant activity of ethyl acetate extract of Musa × paradisiaca L. leaf and standard ascorbic acid was evaluated on the basis of the radical scavenging effect of the stable 1,1-diphenyl-2-picrylhydrazyl (DPPH) free radical activity by modified DPPH assay [19]. In DPPH assay DPPH act as free radical scavenger as it accepts hydrogen (H) from the scavenging molecule and changes color from purple to yellow. The absorbance of the sample was measured by UV spectroscopy at 517 nm [20]. The absorbance was noted and % inhibition was calculated as follows:

Preparation of test solutions
For cytotoxicity studies, 32 mg/ml stocks were prepared using methanol and serially three-fold dilutions were prepared from 320 µg/ml to 10 µg/ml using DMEM plain media for treatment.

Cell lines and culture medium
All the cell lines were procured from ATCC. Stock cells were cultured in DMEM, supplemented with 10% inactivated Fetal Bovine Serum (FBS), penicillin (100 IU/ml), streptomycin (100 µg/ml) in a humidified atmosphere of 5% CO 2 at 37 o C until confluent.
The cells were dissociated with cell dissociating solution (0.2% trypsin, 0.02% EDTA, 0.05% glucose in PBS). The viability of the cells was checked and centrifuged. Further, 50,000 cells per well were seeded in a 96 well plate and incubated for 24 h at 37 o C, in 5% CO 2 incubator Or with 5% CO 2 atmosphere.

Procedure
The monolayer cell culture was trypsinized and the cell count was adjusted to 1.0 × 10 5 cells/ml using respective media containing 10% FBS. To each well of the 96 well microtiter plate, 100 µl of the diluted cell suspension (50,000 cells/well) was added. After 24 h, when a partial monolayer was formed, the supernatant was flicked off, washed the monolayer once with medium and 100 µl of different test concentrations of test drugs were added on to the partial monolayer in microtiter plates. The plates were then incubated at 37 o C for 24 h in 5% CO 2 atmosphere. After incubation, the test solutions in the wells were discarded and 100 µl of MTT (6 mg/10 ml of MTT in PBS) was added to each well. The plates were incubated for 4 h at 37 o C in 5% CO 2 atmosphere. The supernatant was removed and 100 µl of dimethyl sulphoxide (DMSO) was added and the plates were gently shaken to solubilize the formed formazan. The absorbance was measured using a microplate reader at a wavelength of 590 nm. The percentage growth inhibition was calculated using the following formula and concentration of test drug needed to inhibit cell growth by 50% (IC 50 ) value is generated from the dose-response curves for each cell    Growth inhibition against Hela cell line Growth inhibition against A375 cell lines