Production of anti-inflammatory compounds in calli and cells in suspension of Tilia americana var. mexicana

Key message Calli and cells in suspension of Tilia americana produce coumarins and triterpenes compounds with anti-inflammatory activity. Tilia americana var. mexicana is a species identified in Mexico as in danger of extinction. This tree is used in Mexican traditional medicine to treat diseases related to inflammation and nervous sufferings; tiliroside, iso-quercetin, rutin, and quercetin-3-beta-d-glucoside are the compounds identified with anxiolytic properties. Calli from apical and axillary buds established in Murashige and Skoog (MS) medium with thidiazuron (TDZ) and indole butyric acid (IBA) produced anxiolytic compounds and scopoletin; the methanolic extracts of calli and tree leaves inhibited the edema formation in the mouse ' s ear produced with a diester of phorbol (TPA). In this study, calli and cells in suspension of T. americana were established from leaf explants cultivated in MS medium with 2,4-dichlorophenoxiacetic acid (2,4-D, 2.0 mg L-1) plus kinetin (Kin, 0.5 mg L-1). The best maximal growth rate (mu(max)) and doubling time (Dt) of cells in suspension were obtained with the inoculum of 4.5%. The methanolic extract (TaCCSMeOH) from callus and cells in suspension and its dichloromethane fraction (TaFCCSD) inhibited the inflammatory process produced by TPA. The anti-inflammatory compounds found were the coumarins: scopoletin (2), esculetin (3), fraxin (4) and esculin (7) and the triterpenes: beta-sitosterol (1), ursolic acid (5) and betulinic acid (6); quercetin-3-beta-d-glucoside was also detected. The anti-inflammatory effect of TaFCCSD and betulinic acid (6) increased with increasing doses, with an effective dose for 50% of inhibition (ED50) of 0.215 mg/ear and ED50 of 0.14 mg/ear, respectively.