Maximum carboxylation rate (Vcmax) is a key parameter to characterize the forest carbon cycle process. Hence, monitoring the Vcmax of different forest types is a hot research topic at home and abroad, and hyperspectral remote sensing is an important technique for Vcmax inversion. Moso bamboo is a unique forest type with a high carbon sequestration capacity in subtropical regions, but the lack of Vcmax data is a major limitation to accurately modeling carbon cycling processes in moso bamboo forests. Our study area was selected in the moso bamboo forest carbon sink research base in Shanchuan Township, Anji County, Zhejiang Province, China, which has a typical subtropical climate and is widely distributed with moso bamboo forests. In this study, the hyperspectral reflectance and V25cmax (Vcmax converted to 25 °C) of leaves of newborn moso bamboo (I du bamboo) and 2- to 3-year-old moso bamboo (II du bamboo) were measured at different canopy positions, i.e., the top, middle and bottom, in the moso bamboo forest. Then, we applied a discrete wavelet transform (DWT) to the obtained leaf hyperspectral reflectance to construct the wavelet vegetation index (WVI), analyzed the relationship between the WVI and moso bamboo leaf V25cmax, and compared the WVI to the existing hyperspectral vegetation index (HVI). The ability of the WVI to characterize the moso bamboo V25cmax was interpreted. Finally, the partial least squares regression (PLSR) method was used to construct a model to invert the V25cmax of moso bamboo leaves. We showed the following: (1) There are significant leaf V25cmax differences between I du and II du bamboo, and there are also significant leaf V25cmax differences between the top, middle and bottom canopy positions of I du bamboo. (2) Compared to that with the HVI, the detection wavelength of the V25cmax of the WVI expanded to the shortwave infrared region, and thus the WVI had a higher correlation with the V25cmax. The absolute value of the correlation coefficient between the V25cmax of I du bamboo and SR2148,2188 constructed by cD1 was 0.75, and the absolute value of the correlation coefficient between the V25cmax of II du bamboo and DVI2069,407 constructed by cD2 was 0.67. The highest absolute value of the correlation coefficient between V25cmax and WVI at the three different canopy positions was also 13–21% higher than that with the HVI. The longest wavelength used by the WVI was 2441 nm. (3) The validation accuracies of the V25cmax inversion models constructed with the WVI as a variable were all higher than those of the models constructed with the HVI as a variable for all ages and positions, with the highest R2 value of 0.97 and a reduction of 20–60% in the root mean square error (RMSE) value. After the wavelet decomposition of the hyperspectral reflectance of moso bamboo leaves, the low-frequency components contained no noise, and the high-frequency components highlighted the original spectral detail features. The WVI constructed by these components increases the wavelength range of V25cmax interpretation. Therefore, the V25cmax retrieval model based on the WVI encompasses different resolutions and levels of spectral characteristics, which can better reflect the changes in bamboo leaves and can provide a new method for the inversion of the V25cmax of moso bamboo forests based on hyperspectral remote sensing.