Variation in Assessment of Leaf Pigment Content from Vegetation Indices Caused by Positions and Widths of Spectral Channels

Vegetation indices (VIs) are a widely adopted and straightforward tool for non-contact estimation of chlorophyll and carotenoid content in plant leaves. However, VI-based method accuracy depends critically on instrument configuration and calibration procedures. This study aimed to evaluate the sensitivity of VI-based pigment assessment to variations in spectral channel parameters (central wavelength and bandwidth) as well as to changes in calibration details defined by the specific VI formula. Pigment content was measured in leaves of <i>Lactuca sativa</i> L. and <i>Cucumis sativus</i> L. at contrasting developmental stages using VI-based reflection spectroscopy across the 450-950 nm spectral range with various protocols and spectrophotometry as the reference method. VI values were calculated with varying central wavelength and widths of spectral bands, and across different VI formulas. Comparative analysis of the obtained measurements revealed that even minor shifts in central wavelengths of less than 20 nm or the use of an alternative index formula could lead to relative errors of 42-77% in the estimation of chlorophylls and carotenoids content, while changes in bandwidth had a much smaller impact, resulting in only 2-5% relative errors. Even with identical parameters of spectral channels, the choice of an appropriate VI and its regression model could introduce significant errors, ranging from 36% to 86%. These findings highlight the critical role of instrument specifications and calibration models in the VIs-based method accuracy and stability, as measurement errors can lead to suboptimal agronomic decisions. Moreover, our study underscores that comparing results from different sensors or platforms can be unreliable unless the channel parameters and calibration details are clearly specified. Therefore, standardization and transparency in VIs assignment is vital to ensure reproducibility and cross-compatibility in non-destructive pigment monitoring by using various devices.