Abstract
Pennyroyal (Mentha pulegium L.) is a valuable medicinal plant, renowned for its diverse biological and therapeutic effects, which are largely attributed to its wide range of secondary metabolites, such as terpenoids and phenolic compounds. Currently, water deficit, as a major growth-limiting factor for many medicinal plants, poses one of the most significant challenges in plant production. A factorial experiment, based on a completely randomized design (CRD) with three replications, was conducted to assess the effects of foliar application of 5-aminolevulinic acid (ALA) on the growth, physiological attributes, and biochemical responses of M. pulegium under a gradient of drought stress intensities. The experimental treatments comprised four levels of drought stress (100%, 75%, 50%, and 25% of field capacity) and four concentrations of 5-aminolevulinic acid (ALA) (0, 5, 10, and 15 µM). The accumulation of malondialdehyde (MDA), hydrogen peroxide (H₂O₂), proline, glycine betaine, and total phenols, as well as the activities of catalase (CAT), peroxidase (POD), and superoxide dismutase (SOD) enzymes, were elevated in response to increasing drought intensity. A decline in relative water content, photosynthetic pigment levels, stomatal conductance, Rubisco enzyme activity, and photosynthetic rate with increasing stress levels resulted in reduced plant growth. Foliar application of ALA significantly mitigated the accumulation of hydrogen peroxide and lipid peroxidation (as measured by malondialdehyde content) in pennyroyal plants. This protective effect was mediated by an enhancement in the accumulation of antioxidant compounds, including total phenols and essential oils, coupled with increased activity of antioxidant enzymes such as catalase, peroxidase, and superoxide dismutase. Furthermore, ALA application, particularly at a concentration of 10 µM, alleviated the detrimental effects of water deficit on photosynthesis by enhancing photosynthetic pigment content, stomatal conductance, and Rubisco enzyme activity. The enhanced drought tolerance observed in pennyroyal following ALA application is primarily attributed to a significant boost in the plant's antioxidant capacity. This enhanced capacity effectively protects the photosynthetic apparatus from oxidative damage. Consequently, the resultant improvement in photosynthetic efficiency supports better growth performance under drought stress conditions.