Sniffer bees as a reliable tool for Andrographis paniculata detection

Honey bees of Apis mellifera could be trained to be highly reliable sniffers for the detection of Andrographis paniculata using the classical Pavlovian conditioning training method with high success rate, > 80% based on the proboscis extension reflex as a positive response to the presence of the herb. The success rate of sniffer bees was found to be in a temperature dependent manner, but not significantly affected by the heating duration (5–110 min). The variance of 7.7% success rate was observed for the heating temperature ranged 50–120 °C with the highest success rate (92.7%) at 100 °C. This could be due to the content of signature compounds released from the heated herbal samples. Three signature compounds such as dihydroactinidiolide, apiol and 6,10,14-trimethyl-2-pentadecanone were proposed to be the volatile marker of the herb since their concentrations changed in accordance with the temperature profile and success rate of sniffer bees. The volatile compounds were extracted by divinylbenzene and carboxen coated polydimethylsiloxane fiber in the headspace of solid phase micro-extraction before analyzed by GC–MS for identification. Almost 50% success rate could be achieved using the minimum amount of 20 mg herbal samples. High selectivity of the sniffer bees has also been proven by no response to another morphologically similar herb, Clinacanthus nutans which was also heat-treated in the similar manner. The sniffer bees also showed to exhibit 80% success rate to detect A. paniculata mixed with 50% C. nutans as interference in a mixture