*2.3. Phytotoxic Activity of P. undulata EOs*

The EOs of both Saudi and Egyptian ecotypes of *P. undulata* showed significant phytotoxic activity against seed germination and seedling growth of the noxious weed *B. pilosa* (Figure 3). At the highest concentration (100 μL L−1), EOs of Saudi ecospecies showed inhibition of germination, shoot growth, and root growth of *B. pilosa* by 66.67%, 74.59%, and 83.47%, respectively, while the Egyptian species showed inhibition values of 86.67%, 79.23%, and 94.17%, respectively (Figure 3). Based on the IC50, the Saudi ecospecies showed IC50 values of 72.83, 72.84, and 44.55 μL L−<sup>1</sup> regrading germination, shoot growth, and root growth of *B. pilosa*, respectively. However, the Egyptian ecospecies showed IC50 values of 42.42, 65.71, and 40.70 μL L<sup>−</sup>1, respectively (Figure 3).

**Figure 3.** Phytotoxic effect of the EOs extracted from the aerial parts of both Saudi and Egyptian ecotypes of *P. undulata* on the (**a**) germination of seeds, (**b**) shoot growth, and (**c**) root growth of the weed *Bidens pilosa*. Different letters on each line mean significant differences (one-way randomized blocks ANOVA). Data are mean value (*n=3*), and the bars represent the standard error. \* *p* < 0.05, \*\* *p* < 0.01.

It was evident that the Egyptian ecospecies were more effective against *B. pilosa* than Saudi ecospecies, which could be ascribed to the variation in the quality and quantity of the chemical composition of the EO. In this study, the Egyptian ecospecies were richer in oxygenated compounds than the Saudi ones. EOs rich in oxygenated compounds have been reported to possess more activity [38–41]. The phytotoxic activity of the EO from Egyptian ecospecies might be attributed to its major compounds such as spathulenol, hexahydrofarnesyl acetone, *α*-bisabolol, and τ–cadinol. Additionally, the Saudi ecospecies had *β*-pinene, isoshyobunone, 6-epi-shyobunol, *α*-pinene, and *α*-terpinolene as major compounds. Moreover, τ–cadinol was identified as a major compound in the EO of *Cullen plicata*, where it showed strong phytotoxic activity against *B. pilosa* and *Urospermum picroides* [38]. Additionally, τ–cadinol was reported in a high concentration of the EO of *Rhynchosia minima*, which showed significant allelopathic activity against *Dactyloctenium aegyptium* and *Rumex dentatus* [42]. However, *α*-bisabolol, as a major compound of the Egyptian ecospecies in the present study, has not been reported to possess phytotoxicity; therefore, further study is recommended for its characterization as an allelochemical compound.

In the Egyptian ecospecies, the major compound, spathulenol (30.86%), has also been reported as major compounds of EOs with substantial phytotoxic activity such as *Launaea mucronata* [26], *Xanthium strumarium* [37], *Eucalyptus camaldulensis* [43], *Teucrium arduini* [44], and *Symphyotrichum squamatum* [25]. Moreover, hexahydrofarnesyl acetone (18.12%), was determined in a high concentration of the EO, which exhibited strong phytotoxicity such as *Heliotropium curassavicum* [23], *Launaea nudicaulis*, *Launaea mucronata* [26], and *Bassia muricata* [36].

Otherwise, the main compound in the EO of Saudi ecospecies, *β*-pinene (21.14%), has been reported as the main compound of EOs of various plants that have exhibited phytotoxic activity such as *Schinus terebinthifolius* [45], *Symphyotrichum squamatum* [25], *Pinus brutia*, *Pinus pinea* [46], *Lavandula angustifolia* [44], and *Heterothalamus psiadioides* [47]. The other major compounds of the Saudi ecospecies have also been reported in EOs with significant phytotoxicity [1,46,48]. Additionally, the present data showed that the roots were more sensitive to the EO than shoots since roots were directly exposed to the EO. Moreover, root cells have more permeability than the cells of the shoot [22,38].

Results also indicated that the EOs of Saudi and Egyptian ecospecies showed more inhibitory activity against the weed *D. aegyptium* than *B. pilosa* (Figure 4).

**Figure 4.** Phytotoxic effect of the EOs extracted from the aerial parts of both Saudi and Egyptian ecotypes of *P. undulata* on the (**a**) germination of seeds, (**b**) shoot growth, and (**c**) root growth of the weed *Dactyloctenium aegyptium*. Different letters on each line mean significant differences (one-way randomized blocks ANOVA). Data are mean value (*n=3*) and the bars represent the standard error. \* *p* < 0.05, \*\* *p* < 0.01.

At the highest concentration of the Saudi EOs (100 μL L<sup>−</sup>1), the *D. aegyptium* seedling growth was completely inhibited. However, the germination was reduced by 93.33%, while the Egyptian ecospecies showed 96.67%. Based on the IC50 values, the EO of the Saudi ecospecies showed IC50 values of 48.61, 50.49, and 62.92 μL L−<sup>1</sup> for germination, shoot growth, and root growth of *D. aegyptium*, respectively, while the Egyptian ecospecies attained IC50 values of 38.84, 46.59, and 51.87 μL L<sup>−</sup>1, respectively.
