To maximize the use of wheat straw as a feedstock for biofuels and other biorefinery products, the structure of lignin-carbohydrate complexes (LCCs) was characterized by injection of 13C isotope-labeled xylose into living wheat straw. Afterwards, lignin-carbohydrate complexes were extracted from the harvested straw by the Björkman method. The extracted LCCs were chemically characterized by Fourier transform-infrared spectroscopy (FT-IR), sugar composition, molecular weight analysis, 13C-NMR, and HSQC. The results showed that LCCs in wheat straw were particularly enriched with xylan and exhibited narrow polydispersity (Mw/Mn < 1.5). NMR analysis showed that the lignin was linked with the carbohydrates through γ-ester, phenyl glycoside, and benzyl ether bonds. In addition to S, G, and H lignin units, p-coumarate and ferulate were also in the LCCs. The substructures in lignin were β-O-4′, β-β’, and β-5′. Quantitative data analysis of 13C-NMR combined with HSQC showed that the lignin in the LCCs of wheat straw contained guaiacyl (G), syringyl (S), and p-hydroxyphenyl (H) units in the proportion of 5:4:1 (S:G:H). The main lignin substructure, β-O-4′, comprised 71.64% of the isolated lignin. The total LCC linkages (the sum of phenyl glycoside, γ-ester and benzyl ether bonds) were 210.86 /100C9 in 13C-LCC, which was dominated by phenyl glycoside linkages, followed by γ-ester bonds and minor amounts of benzyl ether bonds. Lignin and xylan in the LCCs of wheat straw were mainly linked by benzyl ether bonds and phenyl glycoside linkages.