Image-based phenotyping to estimate anthocyanin concentrations in lettuce

Anthocyanins provide blue, red, and purple color to fruits, vegetables, and flowers. Due to their benefits for human health and aesthetic appeal, anthocyanin content in crops affects consumer preference. Rapid, low-cost, and non-destructive phenotyping of anthocyanins is not well developed. Here, we introduce the normalized difference anthocyanin index (NDAI), which is based on the optical properties of anthocyanins: high absorptance in the green and low absorptance in the red part of the spectrum. NDAI is determined as (I_red - I_green)/(I_red + I_green), where I is the pixel intensity, a measure of reflectance. To test NDAI, leaf discs of two red lettuce (Lactuca sativa) cultivars 'Rouxai' and 'Teodore' with wide range of anthocyanin concentrations were imaged using a multispectral imaging system and the red and green images were used to calculate NDAI. NDAI and other commonly used indices for anthocyanin quantification were evaluated by comparing to with the measured anthocyanin concentration (n = 50). Statistical results showed that NDAI has advantages over other indices in terms of prediction of anthocyanin concentrations. Canopy NDAI, obtained using multispectral canopy imaging, was correlated (n = 108, R² = 0.73) with the anthocyanin concentrations of the top canopy layer, which is visible in the images. Comparison of canopy NDAI from multispectral images and RGB images acquired using a Linux-based microcomputer with color camera, showed similar results in the prediction of anthocyanin concentration. Thus, a low-cost microcomputer with a camera can be used to build an automated phenotyping system for anthocyanin content.