Objectives Direct challenge of cortical serotonergic (5-hydroxytryptamine, 5-HT) availability by tryptophan depletion test (TDT) was used to assess the hypothesized inverse relationship between central 5-HT function and loudness dependence of auditory evoked potentials (LDAEPs). Gender must be taken into particular account here, since there are gender differences in 5-HT brain synthesis, with women reacting more strongly to TDT. Methods In a double-blind, controlled cross-over study, 16 healthy females were ingested two highly concentrated amino acid mixtures with (+TRP) or without TRP (−TRP). While monitoring TRP levels and mood states, the AEP of different loudness stimuli were recorded, followed by dipole source analysis. Results Under the −TRP condition, free plasma TRP levels decreased by 81.10% (±5.14). Most of the loudness change rates of the relevant N1/P2 tangential dipole activities were significantly increased under −TRP, but calculated LDAEP did not differ significantly between treatments. LDAEP and states of mood were not correlated. Conclusions Despite strong TRP depletion, the results did not reach sufficient evidence that LDAEP is a valid biological marker of central 5-HT activity in females when using TDT. This agrees with the literature and supports the view that LDAEP indicates predominantly biological vulnerability in predisposed individuals. Copyright © 2007 John Wiley & Sons, Ltd.