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In this study, a simple, rapid, and solvent-free method for quantitative determination of (+)-Δ(3)-carene metabolites released from larvae of Spodoptera litura was developed using headspace solid-phase microextraction (HS-SPME) coupled with gas chromatography-mass spectrometry (GC-MS). Further, we calibrated and validated the HS-SPME method for the quantitation of the loss of substrates owing to their biotransformation by the larvae. (+)-Δ(3)-Carene metabolites were extracted at 25°C for 30 min using an SPME fiber over a period of 24 h; the SPME fiber used was made of divinylbenzene-carboxen-polydimethylsiloxane. This technique was used to analyze the time course of the larval headspace, and the results of this analysis were used to propose a metabolic pathway. An external calibration curve was used for the quantification of (+)-Δ(3)-carene metabolites from the larval headspace. The total release volume of the larvae was calculated at 24 % of the dosage. Moreover, the biotransformation by S. litura began 2 h after it was injected with (+)-Δ(3)-carene. The method was validated by calculating the limit of detection (LOD), limit of quantification (LOQ), accuracy, precision, and linearity. The LOD and LOQ corresponded to signal/noise ratios of 3 and 10, respectively. The LODs ranged from 0.002 to 0.003 nmol/mL, and the LOQs ranged from 0.007 to 0.009 nmol/mL. This method was sensitive enough to quantitate the (+)-Δ(3)-carene metabolites released from the Spodoptera larvae. The developed SPME method can have wide applications in various in vivo larval metabolite studies.

Quantitative Evaluation of (+)-Δ3-Carene Metabolites from Living Larvae of Spodoptera litura by Headspace Solid-phase Microextraction