Abstract
The purpose of this research was to investigate the effect of crude extracts from the root of Stemona tuberosa Lour. on the replication of Autographa californica multiple nucleopolyhedrovirus (AcMNPV). Cytotoxicity of crude hexane, dichloromethane and ethanol extracts from the root of S. tuberosa was tested against Spodoptera frugiperda cell line (Sf9) using MTT assay. The cytotoxic effect, represented as CC50 (µg/ml) was observed after 48 and 96 h. It was shown that dichloromethane extract was more toxic than hexane and ethanol extracts and 96-h CC50 for the dichloromethane extract was 1,708.98 µg/ml. Crude dichloromethane extract at the concentration of 31.25 µg/ml was then tested on AcMNPV. The extract was added after 1 h post-infection of AcMNPV at the multiplicity of infection (MOI) of 2, in Sf9 cell line cultivated in vitro. No significant difference between the percentage of infected cells in the control and the test sample with crude dichloromethane extract was found. The average number of polyhedra (OBs/ml) in the control (5.11×106±0.63 OBs/ml) was not significantly different from the test sample (4.19×106±0.31 OBs/ml). However, there was significant difference between the average virus titer (budded virus, BV) in the control (2.06×108±0.71 PFU/ml) and the test samples (2.65×108±0.79 PFU/ml). Crude dichloromethane extract (31.25 µg/ml) did not toxic to Sf9 cells but it could enhance AcMNPV replication in Sf9 cell line cultivated in vitro. It could be concluded that S. tuberosa can be a good candidate for developing the insect virus production in vitro for controlling insect pests. Key words: AcMNPV, crude extract, cytotoxicity test, by 3-(4, 5- dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) assay.
Highlights
In Thailand, Stemona species are known as Non Tai Yak
Based on 96 h exposure results obtained from hexane, dichloromethane and ethanol extracts from S. tuberosa against Sf9 cell line, the CC50 values were >2,000, 1,708.98, and >2,000 μg/ml, respectively
Among the crude extracts tested, the dichloromethane extract gave the highest toxic to Sf9 cells. The cytotoxicity of these extracts can be categorized using the classification of the cytotoxicity of natural ingredients reported by Gad Shayne (1999) and Shirazi et al (2004), we can conclude that these extracts had partially nontoxic to Sf9 cell line, because of the CC50 were more than 1,000 μg/ml (Table 2)
Summary
In Thailand, Stemona species are known as Non Tai Yak. Stemona spp. can be used as insect repellent and insecticides such as scabicide and pediculocide (Greger, 2006; Chanmahasathien et al, 2011). Profiles could be grouped into four types represented as the major component by stenine-type Stemona alkaloids such as tuberostemonine and neotuberostemonine or by non-stenine types such as croomine and stemoninine. The biological activities of Stemona alkaloids were insecticidal and larvicidal activities (Brem et al, 2002; Kaltenegger et al, 2003; Xu et al, 2006)
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