Increased Carbazole Alkaloid Accumulation in Clausena harmandiana Callus Culture by Treatments of Biocontrol Agent, Trichoderma harzianum and Bacillus subtilis.

Clausena harmandiana (Pierre) Guillaumin is the Thai medicinal plant that possessed several pharmacological activities. The main active constituents of this plant are the carbazole alkaloids, isolated from wild plants. However, the in vitro culture for production of carbazole alkaloids from this plant has never been reported. Therefore, we aimed to develop callus culture of C. harmandiana elicited with two biotic elicitors, Trichoderma harzianum and Bacillus subtilis, as a sustainable source of carbazole alkaloids. The callus treated with living B. subtilis (BL) at 0.1 and 1% (v/v) for 3 days accumulated 5-fold increased level of clausine K. The highest level reached 309.37 ± 34.84 μg/g DW. This treatment also showed a significant increase in both total phenolic content and antioxidant capacity, which support the optimum usage of this elicitor. Moreover, only callus treated with 1% (v/v) Trichoderma culture filtrate (CF) showed a significant increase in the total phenolic contents and antioxidant capacity. The correlation analysis also revealed the significant correlation between antioxidant capacity and total phenolic level, total flavonoids, and clausine K but not 7-methoxymukonal. The results from our study suggested the use of C. harmandiana callus with the Bacillus elicitors for high-level production of clausine K.