Cytotoxic Activity of Gambier Leave (Uncaria gambir) Ethyl Acetate Extract on Mouse Embryonic Fibroblast Cell (NIH-3T3) using MTT Assay

Abstract

Background: Uncaria gambir or gambier is one of the plants widely found in Indonesia. Gambier is locally known as an antioxidant and antibacterial agent because it has high catechin content. Ethyl acetate extract of gambier leaves has been investigated to contain the highest catechin content than other extraction solvents. Fibroblasts are often used in biomaterial viability and toxicity tests because they have a highly reproducible growth rate and biological response. NIH-3T3 is commonly used as a substitute for human gingival fibroblasts. However, no study has been conducted on the cytotoxic activity of gambier extract on fibroblast cells. Objective: The aim of this study is to evaluate whether the cytotoxic activity of gambier ethyl acetate extract (GEE) exerts on NIH-3T3 cell lines using MTT assay. Methods: The cytotoxic activity of gambier extract was evaluated in three incubation periods. The cytotoxicity test was conducted using an ethyl acetate extract of gambier (Uncaria gambir Roxb.) leaves. The NIH-3T3 cell was treated by GEE in ten concentrations (0, 2.5, 5, 10, 25, 50, 100, 250, 500, and 1000 ppm) for 24-, 48, and 72-hour incubation periods. Cell viability was determined with MTT (3-4,5-dimethylthiazol-2yl -2,5-diphenyltetrazolium bromide) assay. The data were analyzed statistically using SPSS based on ANOVA, followed by Tukey HSD post hoc with p<0.05 and ANOVA paired sample T-test with p<0.05, and the CD50 value was measured by Sigma Plot software. Results: GEE at 2.5, 5, 10, 25, 50, 100, and 250 ppm have cell viability >80%, and cell viability was observed to be increased based on the incubation period. GEE at 1000 ppm significantly decreased the cell viability from GEE at 0 ppm in 24-, 48-, and 72-hour incubation periods (23.83%, 30.14%, and 19.02%, respectively). Moreover, GEE at 500 ppm became toxic by significantly decreasing the cell viability in 48- and 72-hour incubation (40.43% and 23.03%, respectively). The CD50 value of GEE at 24-, 48-, and 72-hour incubation was found to be 578.03 ppm, 488.63 ppm, and 470.70 ppm, respectively. Conclusion: GEE at 2.5, 5, 10, 25, 50, 100, and 250 ppm were not found to be toxic to NIH-3T3 cells for 24-, 48-, and 72-hour incubation periods.