Guidelines for quantifying leaf chlorophyll content via non‐destructive spectrometry

Abstract Premise Leaf chlorophyll is a fundamental bioindicator used in several fields; however, we lack clear guidelines for optimizing sampling efforts and producing comparable studies. Methods We estimated the leaf chlorophyll content of 10 plant species using nondestructive spectrometry methods. We stored half of the leaves at 4°C and half at room temperature under similar light levels to assess the role of storage in the chlorophyll degradation rate. Results The chilled mature leaves maintained a chlorophyll content within 5% of the original value for ~1.5 d, while the chlorophyll content of unrefrigerated mature leaves decreased rapidly, indicating that their chlorophyll content should be measured within 4 h. When refrigerated, the chlorophyll content of the expanding leaves remained within 5% of the original level for at least 5 d, but we suggest analyzing them within 3 d. In mature leaves, 73% of the variation in chlorophyll content is at the species level, 15% is between individuals, and the variation within leaves is negligible (<1%). Measuring one mature leaf from eight individuals was sufficient to provide a species chlorophyll estimate within 5% of the true value at least 80% of the time. Discussion We advise researchers to prioritize sampling more individuals rather than repeating measures within leaves or individuals. Our findings will help researchers to optimize their time and research efforts, and to obtain more robust ecological data.