Elevated CO2 improves growth, modifies anatomy, and modulates essential oil qualitative production and gene expression in Lippia alba (Verbenaceae)

Carbon dioxide (CO2) concentrations have grown in recent decades and will continue to increase during this century, affecting plant physiology and development. Aiming to evaluate the effect of CO2 elevation on growth, anatomy, essential oil qualitative production and expression of genes related to biosynthesis of these compounds, three chemotypes of Lippia alba (BGEN-01, BGEN-02 and BGEN-42) were cultivated in vitro. Firstly, we focused on the effects of gas exchange in the essential oil profile by comparing three CO2 exchange rates: 14, 21 and 25 µL L−1 s−1 CO2. Continuing, in addition to the previous 14 and 25 µL L−1 s−1 CO2 treatments, plants were placed into acrylic chambers with continuous forced air at 360 and 1000 μL L−1 of CO2; an additional control without allowing gas exchange was added inside the chambers, totaling five treatments with 6 replicates. After 45 days, essential oil profile, histochemical, stomatal density, growth evaluation analyses and transcript analysis were performed. The enrichment with CO2 enhanced plant dry and fresh weight, total chlorophylls and carotenoids in BGEN-01 and BGEN-02, and also increased stomatal density and lignin content for all chemotypes. The multivariate analysis showed that the essential oil profile varied, not only among the different chemotypes, but also within BGEN-01 and BGEN-02 treatments. The qualitative production was different in the treatments with forced air renovation and CO2 enrichment. Regarding the gene expression analyses, Farnesyl pyrophosphate synthase (LaFPPS) and Geranyl pyrophosphate synthase (LaGPPS) did not vary, except for the treatments with forced air ventilation (360 and 1000 µL L−1) in the BGEN-01, which had LaFPPS upregulated. Geraniol synthase (LaGES) was upregulated in all BGEN-02 treatments and for BGEN-01 treatments with 360 and 1000 µL L−1 CO2. Nerolidol/Linalool synthase (LaNES/LIS) was upregulated only in the BGEN-01, in the 360 and 1000 µL L−1 CO2 treatments. These findings provide a better understanding of how CO2 regulates secondary metabolites production, providing a basis to clarify the pathway regulation, further enabling the targeted production of essential oils with greater economic and industrial interest.