Doppler shift spectroscopy (DSS) diagnostics, performed on several neutral beam injectors based on positive ion sources, shows the appearance of a shifted peak corresponding to a hydrogen bearing impurity species (mostly due to water vapour: H2O). However, for DSS experiments based on negative ion sources, we are reporting the detection of this peak for the first time. This peak appears in the DSS spectrum when the observations are recorded with a high-speed turbo molecular pump (pressure during beam pulse ~1 × 10−4 mbar) and disappear when a high-speed cryo pump is used (pressure during beam pulse ~1 × 10−5 mbar). A detailed study on this peak using DSS and a residual gas analyser (RGA) was carried out in a ROBIN (RF operated beam source in India for negative ion research) test stand. In these investigations, it was established that the appearance of the peak is due to emissions from fast neutrals produced from the dissociation of fast hydrogen bearing impurity ions such as H2O−/OH−, extracted with energy E. For the present experiments, negative ions of H2O can be formed either by dissociative attachment of (H2O−)* leading to the formation of H−, O− and OH− fragments or via ion–molecule reactions between H−/O− with a H2O molecule. In this work, a study on these formation mechanisms is presented. There is some evidence that the amount of impurity present in the ion source affects the ratio of the main species. An estimation of relative impurity content in the ion source is done using the intensity ratio of a fast hydrogen peak and a hydrogen peak originating from the negative ion impurity. For these computations, the Balmer-alpha excitation cross-section at low energies (~1.5 keV) was reviewed, since the published data is available only for higher energy ranges, and suitable cross sections are used.