Effects of plant growth regulators and carbohydrates on callus induction and proliferation from leaf explant of Lippia multiflora Moldenke (Verbenacea)

Lippia multiflora Moldenke is a tropical plant used in traditional medicine for many diseases treatment (malaria, hyperthermia, blood pressure, etc.). The aqueous extract and essential oils obtained from the leaves have pharmacological, pesticide and cosmetic properties. This study aims to develop a protocol of callus induction and proliferation from leaf explants of L. multiflora. Murashige and Skoog (1962)basal medium was respectively supplemented with different concentrations (0,5; 2,5 ; 5 and 10 µM)of the plant growth regulators 2,4-Dichlorophenoxyacetic acid (2,4-D), 1-Naphthaleneacetic acid (NAA)and Picloram (PIC). The best auxin was then combined respectively with three types of cytokinins [kinetin (KIN), benzylaminopurine (BAP)and zeatin (ZEA)]. The best hormonal combination was used to test the effect of type and concentration of carbohydrate on callus formation. The results showed that the medium supplemented with 5µM PIC expressed a callus induction rate and a fresh and dry weight of callus significantly higher than the other tested auxins. Medium supplemented with 5 µM PIC + 25 µM KIN was more suitable for callus induction and proliferation. Among the carbohydrates, the maximum rate of callus formation was expressed by the leaf explants on medium containing 3 per cent fructose. The protocol for callus induction developed in this study would allow the in vitro production of essential oil of L. multiflora from cell suspension cultures.