This study examined the effects of exogenous citrulline (control, 1 and 2 mM) and water availability (100%, 50%
and 25% WA) on antioxidant attributes and essential oil constituents of Hyssopus officinalis L. in two successive
harvests. Hyssop tolerantly responded to water deficiency by well-promoted antioxidant enzymes (i.e., superoxide
dismutase, and catalase), strong DPPH-scavenging activity, and increasing polyphenols; however, the
essential oil content was negatively reduced by water stress. External citrulline further increased the activity of
antioxidant enzymes. Citrulline application at 2 mM under severe water stress could also improve essential oil
(EO) content in the first and second harvests by about 15 and 30%, respectively. Furthermore, under severe
drought, citrulline at 2 mM could obtain the highest yield of isopinocamphone (47%) as the main component of
EO. The results showed the high potential of this novel applied metabolite agent to be used in a well-fulfilled
production of this medicinal plant.