We describe a high-throughput method for estimating cell-wall chemistry traits using analytical pyrolysis. The instrument used to perform the high-throughput cell-wall chemistry analysis consists of a commercially available pyrolysis unit and autosampler coupled to a custom-built molecular beam mass spectrometer. The system is capable of analyzing approximately 42 biomass samples per hour. Lignin content and syringyl to guaiacol (S/G) ratios can be estimated directly from the spectra and differences in cell wall chemistry in large groups of samples can easily be identified using multivariate statistical data analysis methods. The utility of the system is demonstrated on a set of 800 greenhouse-grown poplar trees grown under two contrasting nitrogen treatments. High-throughput analytical pyrolysis was able to determine that the lignin content varied between 13 and 28% and the S/G ratio ranged from 0.5 to 1.5. There was more cell-wall chemistry variation in the plants grown under high nitrogen conditions than trees grown under nitrogen-deficiency conditions. Analytical pyrolysis allows the user to rapidly screen large numbers of samples at low cost, using very little sample material while producing reliable and reproducible results.