Rapid and sensitive analysis of azadirachtin and related triterpenoids from Neem ( Azadirachta indica) by high-performance liquid chromatography–atmospheric pressure chemical ionization mass spectrometry

Abstract

Based on reversed-phase high-performance liquid chromatography (RP-HPLC) and atmospheric pressure chemical ionization (APCI) mass spectrometry, a HPLC–MS method was developed to permit the rapid qualitative and quantitative analysis of azadirachtin and related tetranortriterpenoids from seeds and tissue cultures of Neem ( Azadirachta indica). APCI+ standard scanning mass spectra of the major Neem triterpenoids were recorded and utilized to select suitable ions for selected ion monitoring (SIM). Transitions for selective reaction monitoring (SRM) were based on MS–MS experiments. Using SIM, major Neem triterpenoids were detected in callus culture material and seed kernels of A. indica. The limit of detection for azadirachtin in extract samples (∼1 ng ml −1 or 10 pg in SIM mode) was determined to be (with respect to injected absolute amounts) approximately 1000-times lower than values quoted in the literature for existing HPLC methods (∼200 ng ml −1 or 10 ng). In addition to high sensitivity, the HPLC–MS method is able to tolerate minimal sample preparation and purification, dramatically reducing total analysis time.