In this study, a detailed characterization of earthworm low molecular size humic substances (LMS) was performed and these substances were used to study their effect on the nitrate influx in roots, tissue nitrate content, and expression of maize genes putatively involved in nitrate uptake in maize (Zea mays L.). The results show that the humic fraction with low molecular size used in this study is endowed with the characteristic structural network described for most humic substances so far isolated and confirm the presence of IAA in this fraction. The results also show that the LMS fraction of humic substances stimulates the uptake of nitrate by roots and the accumulation of the anion at the leaf level. Moreover, the analysis of the expression of genes encoding two putative maize nitrate transporters (ZmNrt2.1 and ZmNrt1.1) and of two maize H(+)-ATPase isoforms (Mha1 and Mha2) show that these substances may exert direct effects on gene transcription in roots, as shown for the Mha2 gene, and long-distance effects in shoots, as observed for the ZmNrt2.1 gene.