The scope of the protocol is the quantification of pigments in pasture samples in the context of proximal and remote sensing-based studies. The protocol describes a standard procedure for the destructive sampling of mixed pasture covers for the estimation of biophysical variables (biomass, leaf area index, canopy water content, specific leaf area, specific leaf weight), pigments, and nutrients without discrimination by species or functional types and the laboratory procedure for the samples pretreatment and quantification of pigments contents (Chlorophyll a, Chlorophyll b, and carotenoids) in the collected samples. The procedure allows the quantification of pigments in herbaceous covers per fresh or dry mass as well as per sampling area. The determination of pigments is performed by a cold extraction procedure with 80% v/v aqueous acetone followed by a spectrophotometric determination.