Abstract: 【Objective】To regulate the color changes of tobacco leaves during the yellowing stage of flue-curing，and investigate the effects of CO₂ concentration on chlorophyll degradation in tobacco leaves，which could provide technical support for the quality and color optimization of tobacco leaves during the flue-curing process.【Method】Middle leaves of Zhongyan 100 tobacco were used as experimental materials. During the yellowing stage，CO₂ gas regulation treatments were applied during the former 1/3 of the steady-temperature phase at each temperature point（36 ℃ for 3 h；38 ℃ for 8 h；40 ℃ for 4 h；42 ℃ for 6 h）. Three CO₂ concentration treatment group were set：（1.0±0.1）%（GR1），（1.5±0.1）% （GR2），（2.0±0.1）%（GR3），with conventional flue-curing serving as the control（CK）. The yellowing degree，color parameter values，chlorophyll content，chlorophyll-degrading enzyme activities and chlorophyll derivative content of tobacco leaves in each group were measured.【Result】The GR1 and GR2 gas regulation groups accelerated the yellowing process of tobacco leaves during the flue-curing yellowing stage，as well as the increase in the color parameters L*，a* and b*. At the end of 38 ℃，the degree of yellowing reached 87.39% and 92.28% respectively，and the total chlorophyll degradation rates at the later stage of yellowing（end of 42 ℃）were 85.95% and 85.54%，significantly higher than that of the GR3 gas regulation group and CK（P<0.05）. During the yellowing stage，the GR1 and GR2 gas regulation groups maintained higher enzyme activity levels of chlorophyllase（CLH），magnesium-dechelatase（MDCase）and pheophorbide hydrolase（PPH）for a longer period. Moreover，the peak activity of CLH and MDCase occurred earlier，and the activity of MDCase before the end of 38 ℃ and PPH activity during the entire yellowing period were consistently ranked as GR2> GR1>CK>GR3. The GR1 and GR2 gas regulation groups also demonstrated faster accumulation and conversion of chlorophyll a，pheophytin b and demagnesified chlorophyllin a during the yellowing stage，leading to a great increase in chlorophyll degradation efficiency. In contrast，the GR3 gas regulation group delayed the chlorophyll degradation process，hindering the color transformation of the tobacco leaves. After the yellowing stage，the leaves remained visibly green，and the total chlorophyll degradation rate was only 71.90%，indicating a certain negative impact.【Conclusion】Maintaining a CO₂ concentration of（1.0±0.1）% to（1.5±0.1）% during the yellowing stage of flue-curing effectively promotes chlorophyll degradation，ensure adequate yellowing of tobacco leaves，avoid the retention of green color on the surface of tobacco leaves after flue-curing ，and thereby improve flue-curing quality.