Poster Topical Area: Methods and Protocols
Location: Hall D
Poster Board Number: 662
Avocatin B is a mixture of two 17-carbon polyhydroxylated fatty alcohols (PFAs): avocadene (1,2,4-trihydroxyheptadec-16 ene; C17H34O3) and avocadyne (1,2,4-trihydroxyheptadec-16-yne; C17H32O3). First discovered in avocado seed, their quantity in pulp is unknown. Analytical methods to detect avocado seed PFAs have utilized NMR and GC-MS; both of which are limited in quantitative capacity, sensitivity and accuracy. The objective of this study was to develop and validate a liquid chromatography-mass spectrometry (LC-MS) method to detect and quantify amounts of avocadene and avocadyne in Hass avocado seed and pulp matter.
A commercially available standard of avocatin B was used to ascertain exact mass of avocadene and avocadyne molecular ions (MS) and their subsequent fragmentation (MS/MS) in positive electrospray ionization mode (ESI) in a high resolution quadrupole-orbitrap mass spectrometer. The fragmentation peaks [M+H-H2O]+ of avocadene and avocadyne were most intense and enabled the development of a sensitive high mass resolution UHPLC-MS method for quantification.
The method has a reliable and linear response range of 0.1-50 µM (0.03-17.2 ng/µL) for both avocadene and avocadyne (r2>0.990) with a lower limit of quantitation (LLOQ) of 0.1 µM. The intra-and inter-assay accuracy and precision of the quality control (QC) samples at LLOQ showed ≤18.2% percentage error and ≤14.4% coefficient of variation (CV). The intra-and inter-assay accuracy and precision for QC samples at low and high concentrations were well below 10% percentage error and CV. This method was successfully applied to quantify avocadene and avocadyne in total lipid extracts of Hass avocado pulp and seed matter. Results showed that the average amount in the dry weight of one Hass avocado pulp and seed, respectively, was 59.05±16.40 mg and 60.35±30.23 mg for avocadyne and 117.50±52.53 mg and 70.92±31.69 mg avocadene.
Avocatin B possesses potent anti-cancer activity by selectively targeting and eliminating leukemia stem cells. This work demonstrates that avocadene and avocadyne are found in appreciable quantities in pulp and seeds of Hass avocados; the most cultivated and highly consumed variety of avocados in the world.
N.A., is supported by a Mitacs Accelerate scholarship. J.J.A.H is supported by an NSERC Post-Graduate Doctoral Scholarship. K.D.S., is supported by a Canada Research Chair in Nutritional Lipidomics. P.A.S., is supported by grants from Hass Avocado Board (CA, USA), and University of Guelph.
University of Guelph
Kitchener, Ontario, Canada