Lignification of the stem in zinnia provides its mechanical properties due to xylem formation, which depends on the stage of plant development and is responsible for the transport of water and minerals. The study was aimed at the lignin deposition, anatomical traits, biochemical markers of lignification, as well as the genetic regulation of this process in zinnia stem cross sections of different age during their radial growth. The anatomical traits were assessed on cross sections. The content of lignin (Cysteine-assisted sulfuric method (CASA) and the thioglycolic acid (TGA) methods), the spectrum of phenolics (by thin layer chromatography (TLC)), the total activity and the variety of class III peroxidases were determined. The expression level of genes regulating phenylpropanoids and lignin biosynthesis were assessed. We suggest that time-specific and organ-specific lignification is determined by the metabolism of phenolic compounds and depends on the expression of genes of the phenylpropanoid pathway. It was shown that in the hypocotyl, during xylem ring formation, lignification was associated with increased expression of phenylalanine ammonia-lyase (PAL) and cinnamyl alcohol dehydrogenase (CAD) genes responsible for the early stages of the phenylpropanoid pathway, and with the rise of class III peroxidases activity, including cationic isoforms. This caused increased content and diversity of phenolics in mature hypocotyl. In epicotyl, which is younger than the hypocotyl, the proportion of ferulic acid among phenolics increased, which could be considered as a marker of lignification in it. The high expression level of CAD and the activity of peroxidases, including anionic isoforms, led to accumulation of lignin. Thus, the hypocotyl and epicotyl, being characterized by different ages, differed in spectrum of phenolics, isoforms of class III peroxidases, expression of the PAL, cinnamate 4-hydroxylase (C4H), peroxidases III class (PRX), and laccase (LAC) genes, and lignin content. © 2023 by the authors.