Tissue culture and elicitor applications for sustainable secondary metabolite enhancement in Mentha longifolia (L.) Huds

• Elicitor treatments (MeJA, SA, YE) significantly enhanced biomass and secondary metabolite accumulation in-vitro . • MeJA is most effective in increasing phenolics, flavonoids, and essential oil as well as upregulation of biosynthetic genes. • The study demonstrates a sustainable tissue culture-based strategy for reliable production of bioactive metabolites from M. longifolia . Mentha longifolia (L.) Huds., an aromatic herb from the Lamiaceae family, is well known for its bioactive compounds and high demand in the food, cosmetic, and pharmaceutical industries. To overcome several challenges due to overharvesting, disease, and climate change, a dependable, sustainable production method for the important secondary metabolites is necessary, and steady metabolite production under regulated circumstances may be achievable through plant tissue culture. M. longifolia callus cultures were used in this research to evaluate the effects of elicitors, methyl jasmonate (MeJA), salicylic acid (SA), and yeast extract (YE) on biomass accumulation and metabolite synthesis. Murashige and Skoog (MS) medium fortified with 1 mg/l 2,4-D was used to produce calli formed from leaves, which were then subjected to varying concentrations of elicitor. HPLC, quantitative analysis of phenolics, flavonoids, alkaloids, and essential oils were evaluated. The findings indicated that MeJA, at 100 µM supplementation, significantly increased biomass (dry weight increased by 65.12%) and metabolite accumulation, producing the significant levels of phenolics (95.3 ± 4.8 mg GAE/g DW), flavonoids (59.7 ± 3.5 mg QE/g DW), and essential oil content (3.28 ± 0.4% w/w). Both SA and YE increased the accumulation of metabolites, although SA (150 µM) was the most efficient at accumulating alkaloids (2.1 ± 0.1 mg/g DW). In response to the elicitors, the key secondary metabolites biosynthetic genes (PAL, CHS, TDC, and HMGR) were found to be upregulated, with MeJA eliciting (∼5 fold) the highest transcriptional response significantly. All the findings indicate that the use of elicitors, particularly MeJA, showed promise in augmenting the in-vitro production of important secondary metabolites from M. longifolia .