Schleichera oleosa (Molk.)Oken Callus Induced BAP and 2,4 D In vitro

Schleichera oleosa (Lour.) Oken is a plant that it can used as drug candidate. S. oleosa has several health benefits including as anti-cancer, anti-oxidant


INTRODUCTION
Schleichera oleosa (Lour.)Oken is a wild plant that it commonly found in the forests of Nusa Tenggara, Seram Island, Moluccas, Java, Celebes, Bali, and Kai Island.This tree grows well in tropical regions, resistant to drought, or in dry season (Heyne, K., 1987).Based on the result of the researcher (Bhatia et al., 2013) it can be seen that S. oleosa (Lour.)Oken has a several benefits, including as anti-cancer, antioxidant, anti-microbial, biodiesel, phytoremediation, and animal feeds.Khan, et al., (2016) added that this S. oleosa (Lour.)Oken has an efficacy as anti-inflammatory.
S. oleosa (Lour.)Oken is a new discovery source as drug products to be developed.As conventionally, the secondary metabolite from the plant contains as several active compounds through extraction process from the plant organ.However, the uses of the plant for compound production as continuously influences the availability of this plant species.Thus, the large scale cultivation is still needed, in addition to obtaining active compounds in the extraction, isolation, and purification processes that require a high costs.Some compounds which are obtained synthetically require high costs because the active structure is complex.Therefore, it is necessary to develop alternative methods of extracting plants to obtain active compounds in this plant (Chattopadhyay et al., 2002).
One of step for increasing the secondary metabolite through increase the drug compounds from the plants is from biotechnology, especially through in vitro culture plant.This in vitro culture has a big potential, because it has a benefits for nature compound production as reliable (Vanisree et al., 2004).In vitro culture technique has great potential, because it has the advantage of being able to produce natural compounds continuously and reliably (Vanisree et al., 2004) so that with tissue culture techniques, apart from increasing secondary metabolites, S. oleosa is also propagated with good plant quality.
The modification of culture media is an important thing for increasing secondary metabolite content in various plants.The cell or callus proliferation induction need the addition of exogenous plant growth regulator from Auxin and Cytokines group which can be added to the media, either single or combination with this two types of PGR at appropriate concentrations.2,4-D growth regulator is an auxin group which is often added to the callus induction media (Nagasawa & Finer, 1988)2,4-D which is combined with cytokinins (BAP or kinetin) 2,4-D treatment in combination with cytokinins (BAP or citin) greatly influenced the callus development (Thao et al., 2003).

RESULTS and DISCUSSION
Based on the observation result from the callus induction of S. oleosa (Lour.)Oken with the addition of 2,4-D and BAP treatments in various concentrations that it give a real effect.This is evidenced by the result of the Analysis of Variance (ANOVA) that it presented in Table 1.Based on ANOVA results, it was shown that the observation result between the interaction of 2,4-D and BAP have a real effect to all variables observation, such as: day of emerging callus, percentage of callus formation, and weight of callus (table 1).This is shown from the results of the F-count of day emerging callus, percentage of emerging callus, and weight of callus is bigger than Ftable of 5%, as well as the significance value (<0,05) which means there is an effect on all type observations.So, it can be further tested by DMRT 5% test.The following DMRT 5% test results are presented in table 2.

DISCUSSION
The result of DMRT 5% in table 4.2 showed that the interaction between 2,4-D and BAP has significantly affect for the three parameters, including the day of emerging callus, percentage of callus formation, and weight of callus for S. oleosa (Lour.)Oken callus induction.Day of emerging callus at concentration 0,5 mg/L BAP without the addition of 2,4-D can accelerate callus formation which is 32 DAP.
The percentage of callus formation variable, the combination of 1 mg/L 2,4-D and 0,5 mg/L BAP gave the effective percentage is 83,33% (Rahayu W.P, 2003)the callus formation stage is influenced by the division, elongation, and the development of cells.Auxin also has a function for the callus formation, because the addition of auxin can influence the cell wall permeability, so air, macro and micro substances, as well as organic and inorganic molecules in the media can be absorbed into the cells.
In weight of callus varibale, the combination between 2 mg/L 2,4-D and 0,5 mg/L BAP produces callus with the optimum weight.Physiologically, the fresh weight of callus has two material contents, namely polysaccharides and water.The fresh weight callus has a high water content in the callus.The speed of callus cells in dividing themselves to multiply the callus cell mass, thus affecting the fresh weight of callus production (Andaryani, 2010).
The best texture that used to produce secondary metabolite is compact texture.(Sugiyarto & Kuswandi, 2014) states that the texture of compact callus cells is a callus texture that has a tight cell composition.So, the bonds between cells are also getting stronger and not eaily separated.Besides, the size of the vacuole in a large cell also allows the callus can store more water content.So, the fresh wet callus is also getting callus.
Most of the green calluses from S. oleosa (Lour.)Oken leaves as explant initially have light green color.However, at last observation the S. oleosa (Lour.)Oken leaves have dark and brownish color.The green color of callus has chlorophyll content in the callus tissue.Due of several factors, the process of metabolism has excess phenol compounds causes the media to turn yellow, so the plant will die (PYD et al., 2012).Phenols formed in callus cell induction.In this study, as a form to wounds caused by slicing is done.If phenol are formed to experience oxidation, it can cause brown color on the callus known as browning.Changes the color of callus from green to brown indicates the growth and development of the callus entering a stationary phase (aging) which then the callus will be dead (Purnamaningsih & Ashrina, 2011).

CONCLUSION
Based on the result of the Callus induction of S. oleosa (Lour.)Oken with the addition of 2,4-D and BAP showed different effects on several bservation variables.The day of emerging callus variable gave the optimal result at 0,5 mg/L BAP, which is incubated for 32 day after planting.Besides, the weight of callus variable is 1,1458 grams in dose 2 mg/L BAP dan 0,5 mg/L 2,4D .While the percentage of callus formation at concentration 0,5 mg/L 2,4-D + 0,5 mg/L BAP is 83,33%.

Table 1 .
The result of Analysis of Variance (ANOVA)

Table 2 .
The result of DMRT 5% of the interaction between 2,4 Dichloropenoxyacetic Acid and Benzyl Amino Purine with various concentrations of Callus Induction of Schleichera oleosa (Lour.)Oke