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The in-chlorophyll centre waveband (ICCW) (640–680 nm) is the specific chlorophyll (Chl) absorption band, but the reflectance in this band has not been used as an optimal index for non-destructive determination of plant Chl content in recent decades. This study develops a new spectral index based solely on the ICCW for robust retrieval of leaf Chl content for the first time. A glasshouse experiment for solution-culture of one chlorophyll-deficient rice mutant and six wild types of rice genotypes was conducted, and the leaf reflectance (400–900 nm) was measured with a high spectral resolution (1 nm) spectrophotometer and the contents of chlorophyll a (Chl a ), chlorophyll b (Chl b ) and chlorophyll a+b (Chl t ) of the rice leaves were determined. It was found that the reflectance curves from 640 nm to 674 nm and from 675 nm to 680 nm of the low-chlorophyll mutant leaf were drastically steeper than that of the wild types in the ICCW. The new index based on the reflectance variation within ICCW, the difference of the first derivative sum within the ICCW (DFDS_ICCW), was highly sensitive (r = −0.77, n = 93,  P <0.01) to Chl t  while the mean reflectance (R_ICCW) in the ICCW became insensitive ( r  = −0.12, n = 93,  P >0.05) to Chl t  when the leaf Chl t  was higher than 200 mg/m 2 . The best equations of R-ICCW and DFDS_ICCW yielded an RMSE of 78.7, 32.9 and 107.3 mg/m 2 , and an RMSE of 37.4, 16.0 and 45.3 mg/m −2 , respectively, for predicting Chl a , Chl b  and Chl t . The new index could rank in the top 10 for prediction of Chl a  and Chl t  as compared with the 55 existing indices. Additionally, most of the 55 existing Chl-related VIs performed robustly or strongly in simultaneous prediction of leaf Chl a , Chl b  and Chl t .

Reflectance Variation within the In-Chlorophyll Centre Waveband for Robust Retrieval of Leaf Chlorophyll Content