Aqueous-acetone extraction improves the drawbacks of using dimethylsulfoxide as solvent for photometric pigment quantification in Quercus ilex leaves

Abstract

Aim of study: We evaluated the use of dimethylsulfoxide (DMSO) for the photometric determination of chlorophyll (Chla and Chlb) and carotenoids in Quercus ilex L. leaves by comparative analysis using aqueous-acetone extraction. Area of study: a Q. ilex dehesa in Las Majadas del Tietar, Cáceres, Spain Material and methods: Q. ilex leaves were sampled during two vegetative periods. Field SPAD-502 Chlorophyll measurements and photometric chlorophyll determinations were performed. Two procedures were used: extraction of intact foliar discs at 65ºC with DMSO and fine foliar powder with cold aqueous-acetone. Main results: DMSO produced Chlb overestimation and different fitting performance for SPAD vs pigment calibrations (R2=0.64, RMSE=0.20, p<0.0001 for Chla (µg/cm2); R2=0.33, RMSE=0.23, p<0.0001 for Chlb (µg/cm2) and R2=0.50, RMSE=0.23, p<0.0001 for carotenoids (µg/cm2)). Aqueous-acetone provided more accurate predictions (R2=0.90, RMSE=0.16, p<0.0001 for Chla and R2=0.91, RMSE=0.16, p<0.0001 for Chlb, R2=0.90, RMSE=0.02, p<0.0001 for carotenoids) and mean ratio Chla/Chlb=3.6 inside the range for sun exposed leaves. Research highlights: Oxidizing conditions and polyphenol concentrations in Q. ilex leaves generated brown colorations in the DMSO extraction procedure, interfering with the photometric measurements in the red-orange region. Aqueous-acetone extraction was free from interference. DMSO should be avoided for pigment determination in Q. ilex leaves or when comparing different tree species.