Bamboo-cultured cells (BCCs) were produced under three phytohormone conditions. BCC lignin was then isolated and characterized by heteronuclear single-quantum coherence-nuclear magnetic resonance (HSQC-NMR) analysis. HSQC-NMR analysis revealed that all three BCC lignin samples were composed of guaiacyl (G), syringyl (S), oxidized syringyl (S′), and p -hydroxyphenyl (H) units. p -Coumaric acid ( p CA) and ferulic acid (FA) were identified as well. Main lignin substructures, including β - O -4, β -5, and β - β , were also detected. However, β - O -4/ α - O -4, spirodienone, dibenzodioxocin, or tricin structures were absent in the BCC lignin. The BCC lignin contained higher proportions of H, FA, and β -5 structures, but less proportions of S, S′, and β - O -4 structures than the mature bamboo lignin. The removal of auxin 2,4-dichlorophenoxyacetic acid (2,4-D) from the subculture medium promoted G unit formations. Nevertheless, it suppressed H and p CA unit formations. Cytokinin 6-benzyladenine (BA) promoted H and β - β structure formations as well but suppressed β - O -4 formations than in the mature bamboo and BCC lignin produced under phytohormone free conditions.