In vitro germination and plantlet establishment of Labisia pumila (Bl.) F. Vill. M. Hartinie * , G. Azlan Jualang School of Science and Technology, Universiti Malaysia Sabah, Locked Bag 2073, 88999 Kota Kinabalu, Sabah, Malaysia Received 8 June 2007; received in revised form 16 July 2007; accepted 18 July 2007 Abstract In vitro seeds germination and plantlet establishment of Labisia pumila were studied in this report. The seeds obtained from the mature fruits of L. pumila were sterilized and cultured on Murashige and Skoog (MS) solid media supplemented with 1–3 mM of 6-benzylaminopurine (BAP) and 3% (w/v) sucrose. The presence of BAP in the medium significantly affects seeds germination. High percentage of seeds germi- nation (up to 90%) was successfully achieved after 2 weeks of culture on medium supplemented with 2 mM BAP. Up to 70% of explants produced shoots through direct regeneration from newly emerged epicotyls after 5 weeks of culture. The average of 8.1  1.0 shoots per explant obtained on media treated with 2 mM BAP. Seedlings were further transferred to growth media fortified with different types of cytokinin. Result observed after 12 weeks showed that medium supplemented with 1 mM zeatin (ZEA) promote the highest growth with an average of 2.9  1.0 cm shoot length and 7.7  3.2 leaves per explant after 12 weeks. In addition, medium added with 2 mM BAP and supplemented with 3–4% (w/v) of sucrose promote the best growth i.e., 3.0  0.6 shoots per explant, 2.27  0.2 cm length and 4.3  0.5 leaves per explant. # 2007 Elsevier B.V. All rights reserved. Keywords: Medicinal plant; Micropropagation; Myrsinaceae; Plant growth regulators 1. Introduction Labisia pumila (Bl.) F. Vill. (Myrsinaceae) is a small herbaceous plant locally known as ‘‘Kacip Fatimah’’ or ‘‘Akar Fatimah’’ (Burkill, 1966; Jamia and Houghton, 2000) distributed in the rain forest lowlands and hill of Malaysia, Thailand, Indochina, Philippines and New Guinea (Stone, 1988; Jamia and Houghton, 2000; Wiart and Wong, 2002; Ong, 2004). Traditionally, this species has been used by locals as protective medicine before and after childbirth (Jamia and Houghton, 2000). Other medicinal uses are treatment for dysentery, flatulence, dysmenorrhoea and gonorrhoea (Burkill, 1966; Rozihawati et al., 2003). For this reason, this species was extensively used in preparation of herbal product and highly demanded for the manufacturing of commercial product (Jamia et al., 2003; Ezumi et al., 2006). The Malaysian market for herbal and natural product has been estimated to be worth RM4.55 billion of which 90% of the raw materials used was imported. Therefore, cultivation and harvesting are critical aspects of herbal production with only a small percentage is cultivated, with some still on a trial basis (Drewe, 1998; Chang and Rasadah, 2004; Ramlan and Sivakumar, 2006). The growth rate of L. pumila in the natural habitat is very slow (Mohd Noh et al., 2002) and normally it was propagated through seeds or stem cuttings but seeds are difficult to obtain due to depletion of mother plants. In addition, there has so far very little attempt to cultivate this plant (Indu Bala and Ng, 2000; Rozihawati et al., 2003). Hence, the development of an in vitro protocol will be of a great importance for production of planting material to offset the pressure on the natural populations. The in vitro plantlet was vital because the establishment of cultures from explants grown outdoors is difficult due to infection of microorganisms and external harsh conditions (Maliro and Kwapata, 2000; Bhattacharya and Khuspe, 2001). This paper described the protocol for establishment of in vitro plantlet of L. pumila through in vitro seeds germination, shoots proliferation and shoot tips culture of juvenile aseptic seedlings. www.elsevier.com/locate/scihorti Scientia Horticulturae 115 (2007) 91–97 Abbreviations: BAP, 6-benzylaminopurine; KIN, kinetin (6-furfurylami- nopurine); MS, Murashige & Skoog; 1962; PGRs, Plant growth regulators; ZEA, zeatin. * Corresponding author. Tel.: +60 88 320000; fax: +60 88 435324. E-mail address: hartinie@gmail.com (M. Hartinie). 0304-4238/$ – see front matter # 2007 Elsevier B.V. All rights reserved. doi:10.1016/j.scienta.2007.07.011