β-carotene and astaxanthin levels detected by HPLC in wild-type and lemon T. kanzawai.

For each treatment 30 females were collected in three replicates and homogenized in 1 ml of acetone. The homogenate was filtrated using a glass syringe with a membrane possessing a pore size of 0.45 μm. The filtrate was dried under a nitrogen gas flow and dissolved in 300 μl of methanol. Five μl of the solution was used for HPLC analysis. Carotenoids were quantified by monitoring the absorbance at 450 nm. External calibration curves were constructed with authentic standards.