As the main structural component of tobacco cell wall, lignin content is an important factor affecting the safety of tobacco smoking. However, it is time‐consuming to quantify lignin by conventional wet chemical analysis methods. In this work, a 13C multiCP/MAS NMR spectral analysis method for tobacco lignin was established. The multiCP/MAS NMR sequence was optimized for tobacco lignin. The optimized nuclear magnetic sequence parameters were 9 CP cycles of 1.5 ms, repolarization time of 0.7 s, and a total acquisition time of 130 min. Subsequently, TMSP was used as the internal standard substance to establish the working curve, and the correlation coefficient was 0.9946. The relative standard deviation (RSD, n = 5) was 3.26%. This method was applied to the determination of lignin content in different types of tobacco samples. The relative error in the determination of lignin content by this method did not exceed 4.46% compared to the results of the chemical method. The results showed that the 13C multiCP/MAS NMR spectral analysis method had the advantages of accuracy and rapidity, which provided a new technical means for the quantitative study of tobacco cell wall substances.