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Galphimia glauca produces compounds denominated galphimines (galphimine‐A, galphimine‐B and galphimine‐E). Due to their important anxiolytic activity, we initiated in vitro cultures of the species with the purpose of developing a biotechnological process for obtaining galphimines. In this work, we stimulated the biosynthesis and excretion of galphimines with two‐phase batch‐type cell suspension cultures of G. glauca. The effect of nutritional variation and the 2,4‐dichlorophenoxy acetic acid added to Murashige & Skoog(MS) culture medium was evaluated. Later, we evaluated the effect of the stimulation with calcium and methyl jasmonate (MeJ). The greatest production of galphimine‐B (3.39 × 10−5 g/L day−1) was obtained on day 40 of kinetics, and induced by a treatment containing concentrations of nitrates and phosphate that are double of those normally used in MS medium, without sucrose but with added 2,4‐dichlorophenoxy acetic acid (4 mg/L). Time of galphimine‐B biosynthesis diminished due to the effect of MeJ in combination with calcium, and induced the excretion (100%) of galphimine‐B (6.35 × 10−5 g/L day−1) into the culture medium. Thus, the use of calcium and MeJ comprises a viable alternative to stimulate the production and excretion of galphimine‐B and galphimine‐A in batch‐type cultures of G. glauca in modified MS medium. Once optimized, the production of the anxiolytic compounds can be scaled up to the industrial level.

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