Chlorophyll-deficient tea plant exhibits a significantly higher accumulation of free amino acids (FAAs) than normal tea plants. This study focused on the impact of leaf color and the developmental stage on FAAs in six tea germplasms while maintaining all other conditions. The total FAAs content initially increased as the leaf matured during the one-bud-two-leaves (1B2L) and one-bud-three-leaves (1B3L) stages in green germplasms, then decreased or stabilized in the one-bud-four-leaves (1B4L) stage. In contrast, chlorotic germplasms showed continuous FAAs’ content increase from 1B2L to 1B4L, thus being significantly positively correlated with total chlorophyll content. Interestingly, ethylamine content decreased with leaf maturation in both chlorotic and green germplasms, thus showing a significant negative correlation with L-theanine content only in chlorotic germplasms. Comparative RNA-seq analysis linked FAAs accumulation in chlorotic germplasm’s 1B3L to photosynthesis inhibition and in 1B4L to nitrogen assimilation promotion. Feeding experiments revealed higher L-theanine synthesis and degradation abilities in chlorotic shoots versus green shoots, with synthesis efficiency exceeding degradation efficiency. Overall, this study uncovers a developmental-specific FAAs accumulation pattern in chlorotic germplasms and offers novel insights into the precise regulation by leaf color and developmental stage.