Evidence-Based Complementary and Alternative Medicine Diﬀusion method [21, 22]. The procured microorganism strains were inoculated in a Petri dish containing nutrient broth at 37◦C for 24 h and were referred as seeded broth. The density of bacterial suspension of the cultures was standardized turbidometrically to 500 000–1 000 000 colony forming units per milliliter (CFU/ml) at wavelength of 600 nm. The EI extracts were dissolved initially in DMSO, diluted to a concentration of 100 mg/ml and ﬁltered using 0.45 µm millipore ﬁlters. Sterile discs were impregnated with the extract solutions (0.05 ml from 100 mg/ml extract) and placed in inoculated agar. Streptomycin (10 µg/disc) was used as the standard. Controls were prepared using similar solvents but without extracts. The inoculated plates containing both the test and standard discs were incubated at 37◦C for 24 h. 2.5. Cytotoxicity Assay 2.5.1. Preparation of Extracts. To screen the extracts of EI, dried extracts of the plant were dissolved in 1 ml of DMSO to give a stock solution of extract at 10 mg/ml. All extracts were kept at 4◦C throughout the experiments. Stock solutions were further diluted in RPIM1640 (Sigma, MO, USA) to obtain ﬁnal concentrations of 0.469, 0.938, 1.875, 3.75, 7.5, 15 and 30 µg/ml. 2.5.2. Cell Culture Condition. MCF-7 human breast cancer cells and HT-29 human colon carcinoma cells were pur- chased from the American Type Culture Collection (ATCC), USA. Human T4-lymphoblastoid cell line, CEM-SS, used in this study were obtained from The NIH AIDS Research and Reference Reagent Program, USA. The cancer cells were grown at 37◦C in humidiﬁed CO2 incubator with 5% CO2 in RPMI-1640 media supplemented with 10% fetal bovine serum (Invitrogen Corp., Auckland, New Zealand). 2.5.3. Cell Growth Inhibition Assay. Cell suspension (5 × 105 cells/ml) was plated out into 96-well microtiter plate. Plant extracts were initially dissolved in DMSO as mentioned earlier, with the ﬁnal concentration of DMSO being 0.1% (v/v). Serial dilutions of the sample were prepared in RPIM1640. The cytoxicity proﬁles of the extracts were assessed using 3-[4,5-dimethylthiazol-2- yl]-2,5-diphenyltetrazolium bromide (MTT) microculture tetrazolium viability assay as described by Mosmann [23]. Thereafter, various concentrations of the plant extract samples were plated out in triplicates. Each plate included untreated cell controls and a blank cell-free control. After 68 h of incubation, MTT (5 µg/ml) was added to each well and re-incubated for further 4 h. Then, the media was removed and DMSO was added into each well to solubilize the formazan crystals. Finally, the absorbance was read at wavelength of 595 nm using a microtitre plate reader (Labsystems iEMS Reader MF) and the percentage cell viability was calculated with the appropriate controls taken into account. The concentration which inhibited 50% of cellular growth (IC50 value) was determined and the 3 Table 1: TPC in EI extracts. Extract TPC as GAE (mg/g extract) Hexane 210 ± 34b DCM 310 ± 83b EA 175 ± 12a MeTH 450 ± 210c Each value in the table is represented as mean ± SE (n = 3); diﬀerent letters in the same column indicate signiﬁcant diﬀerence (P < .05). inhibitory rate of cell proliferation was calculated by the following formula: Growth inhibition = ODcontrol − ODtreated ODcontrol × 100, (2) where OD is the optical density. Cytotoxicity of the sample towards the cancer cells was expressed as IC50 values (i.e. the EI extract concentration reducing the absorbance of treated cells by 50% in respect to untreated cells). 2.6. Statistical Analysis. Data were expressed as mean ± SD, and ANOVA test was used to analyze the diﬀerences between DPPH activities of EI extracts, with 0.05 as a level of signiﬁcance. Ordinary least squares regression was used to calculate the standard curve of gallic acid. Pearson correlation coeﬃcient was used to measure the relationship between DPPH, scavenging activity and TPC. 3. Results 3.1. Antioxidant Assays. Total phenolic content was deter- mined using Folin-Ciocalteu reagent and the results revealed that the MeTH of EI having the highest TPC (450 ± 210 GAE mg/g extract; P < .05; Table 1) when compared with DCM, hexane and EA extracts. Meanwhile, in the DPPH inhibition results (Figure 1), the MeTH has shown to have the most free radical scavenging activity as observed from the percentage inhibition of the DPPH absorption (77.7%). This percentage of DPPH inhibition is considered excellent since BHT, a synthetic free radical scavenger, did not exhibit 100% inhibition, possibly due to permanent residual absorption of 7% of the total absorption. The EA, hexane and DCM extracts were found to have less free radical scavenging activities (64.5, 47.19 and 40.83%, resp.) when compared with the MeTH. These results reveal that there is a strong and signiﬁcant correlation between TPC and DPPH free radical scavenging activity of the EI extracts (Pearson correlation coeﬃcient = 0.507; P < .05). 3.2. Antimicrobial Activity of EI. The antimicrobial eﬀects of EI extracts studied are summarized in Table 2. The antibac- terial activities of the diﬀerent extracts of EI were evaluated using both Gram-positive and Gram-negative bacteria. The highest antibacterial activity observed was obtained by the hexane extract on MRSA, while the DCM extract showed weak activity on P. aeruginosa. On the other hand, the EA