The production of inoculum is one of the hindrances in the large scale application of arbuscular mycorrhizal fungi (AMF). The objective of this work was to evaluate the effect of nutrient solutions with or without TrisHCl buffer, on sporulation of AMF. The experiment was carried out in a greenhouse, using a substrate with sand and vermiculite (1:1 v/v). Fifty spores of Gigaspora margarita, Scutellospora heterogama, and Glomus etunicatum were inoculated in Sorghum vulgare (sorghum) or Panicum miliaceum (fodder millet). The substrate received the following nutrient solutions: Hoagland with 3 μM P (S1); Long Ashton II with 15.9 μM P (S2) and Hoagland with 20 μM P (S3), with or without 50 mM of Tris-HCl buffer (pH 6.5); the control treatment, consisting of a soil + sand + vermiculite (2:1:1 v/v) substrate was irrigated with deionized water. Ten weeks after the beginning of the experiment sporulation did not differ in treatments with sorghum. Panicum miliaceum promoted higher sporulation of the AMF than sorghum, and differences among treatments with nutrient solutions were observed. Production of spores of G. margarita and S. heterogama increased significantly after addition of buffer in S1 and S2, while that of G. etunicatum was improved when the substrate was irrigated with S1 + buffer and S3 solutions. Solution S1 + buffer benefited sporulation of the three fungi. However, as observed, each AMF, host, and substrate system should be studied separately for establishment of the most favorable conditions for inoculum production.