Accurate monitoring of the chlorophyll content of tea leaves is of great significance to the nutritional status and growth of tea trees. Thus, a method for rapid and nondestructive detection of chlorophyll content of leaves is proposed on the basis chlorophyll fluorescence spectroscopy technology. The chlorophyll fluorescence collection device is used to collect the spectrum of tea leaves and measure the relative chlorophyll content. The Savitzky-Golay (S-G) smoothing method is used to preprocess the spectrum, which can eliminate a large number of noise signals. The proposed method is compared with the traditional method. Experimental results show that the proposed method can effectively eliminate irrelevant variables, and the optimization of the model can achieve better results. The partial least square model established after simplifying the variables has a correlation coefficient of 0.96 on the prediction set, and a root mean square error of 0.87. The correlation coefficient on the model set is 0.96, and the root mean square error is 0.95. The fluorescence spectroscopy and chemometric methods can provide a quick and easy analysis method for the quantitative analysis of tea leaf chlorophyll content.