Peng Fu, Katherine Meacham‐Hensold, Kaiyu Guan, Jin Wu, and Carl Bernacchi

The lack of efficient means to accurately infer photosynthetic traits constrains understanding global land carbon fluxes and improving photosynthetic pathways to increase crop yield. Here, we investigated whether a hyperspectral imaging camera mounted on a mobile platform could provide the capability to help resolve these challenges, focusing on three main approaches, that is, reflectance spectra‐, spectral indices‐, and numerical model inversions‐based partial least square regression (PLSR) to estimate photosynthetic traits from canopy hyperspectral reflectance for 11 tobacco cultivars. Results showed that PLSR with inputs of reflectance spectra or spectral indices yielded an R² of ~0.8 for predicting V _cmax and J _max, higher than an R² of ~0.6 provided by PLSR of numerical inversions. Compared with PLSR of reflectance spectra, PLSR with spectral indices exhibited a better performance for predicting V _cmax (R² = 0.84 ± 0.02, RMSE = 33.8 ± 2.2 μmol m⁻² s⁻¹) while a similar performance for J _max (R² = 0.80 ± 0.03, RMSE = 22.6 ± 1.6 μmol m⁻² s⁻¹). Further analysis on spectral resampling revealed that V _cmax and J _max could be predicted with ~10 spectral bands at a spectral resolution of less than 14.7 nm. These results have important implications for improving photosynthetic pathways and mapping of photosynthesis across scales.