Evaluation location: Georgia, United States
Plants were grown in the field in Griffin, GA in 2017. All fruit (ripe) of each accession were harvested from individual (single) plants. Fruit were placed in foil trays and dried at 80oC for 48 hours, packaged, and sent to the University of Maine for analysis.
Analysis of Capsaicinoids in Dried Capsicum Fruit – Methods and Materials
Sample Preparation. Modified from Collins et al, 1995.
1. Crush dried fruit into a coarse powder packed by messaging plastic sample packet between fingers (bag kept sealed).
2. Weigh entire powder content into 50 mL glass, srew-top centrifuge tube.
3. Add HPLC-grade acetonitrile to tube at appropriate volume (10:1 (mL:g) solvent:sample).
4. Place samples in sand-filled block heater, pre-heated to 80 degrees, C for 4 hours. Swirl samples manually every hour.
5. Remove samples from heat and allow to cool to room temperature.
6. Further extract the samples with a ploytron tissue homogenizer at medium speed for 1 minute each.
7. Filter each sample through 0.45 um nylon syringe filter into HPLC sample vial and store at 5 degrees C until analysis by HPLC.
Sample Analysis. In-house HPLC method
1. Standard Preparation:
a. Analytical Standards Capsaicin (99%) & Dihydrocapsaicin (97%) purchased from Sigma Aldrich.
b. Stock solutions of each prepared using HPLC-grade acetonitrile as a diluent
c. Seven-point standard curve (0.5 ug/mL – 500 ug/mL) prepared for each compound diluting appropriate stock volume with HPLC-grade acetonitrile
2. HPLC Conditions:
a. HPLC System: Agilent 1100/1200 series with pump, autosampler, degasser, diode array detector and column heater.
b. Column: Phenomenex Prodigy C18, 4.6 x 250 mm, 5 micron.
c. Mobile Phase: 70:30 (acetonitrile:water), isocratic, 1.0 mL/min.
d. Detector: DAD, 280 nm
Collins, M. D., Wasmund, L. M. and Bosland, P. W. 1995. Improved method for quantifying capsaicinoids in Capsicum using high-performance liquid chromatography. HortScience 30(1):137-139.