2.1. Floral scent composition of Nymphaea subg. Hydrocallis

The six species and two subspecies of Nymphaea subg. Hydrocallis investigated are all native to Brazil and three of them are known to be associated with pollinator cyclocephaline scarabs (Fig. 1; Table 1).

To the human nose, flowers of the seven investigated taxa of Nymphaea subg. Hydrocallis were remarkably fragrant during the consecutive evenings of the pistillate ($) and staminate (#) phases of anthesis. While the flowers of N. rudgeana and N. gardneriana emitted a pungent, fermented fruity odor with solvent-like reminiscents, the scents of the remaining species all bore a strong, uncharacteristic solvent-like odor (Table 1).

The chemical analysis showed that the seven studied taxa emitted floral volatiles in different quantities and compositions. A total of 22 compounds were identified in the analyzed samples, ranging in molecular weight from 102 [methyl butanoate] (1) to 192 [benzyl 2-methylbutanoate] (2) (Fig. 2; Table 2). The identified volatile compounds belong to three of the seven compound classes proposed by Knudsen et al. (2006): aliphatics (9), C5-branched chain compounds (5) and aromatics (8).

The number of compounds per species ranged from only two in N. lingulata, N. amazonum and N. tenerinervia, to 12 in N. lasiophylla (Table 2). Dominant compounds reaching an overall relative percentage content of at least 10% in the analyzed scent samples were methyl hexanoate (3) in N. rudgeana (36.3–77.8% $; 81.0–97.7% #) and N. gardneriana (21.9% $); methyl 2-methylbutanoate (4) in N. lasiophylla (0.8–12.5% $; 0.4–18.0% #) and N. gardneriana (74.0% $); (methoxymethyl)benzene (5) in N. rudgeana (20.2–61.7% $; 1.2–15.7% #), N. lasiophylla (81.6–95.1% $; 75.1–97.2% #) and N. lingulata (99.8% $; 99.8–99.9% #); and anisole (6) in N. tenerinervia (98.9% $), N. amazonum subsp. amazonum (99.7–100.0% $; 89.0– 100.0% #) and N. amazonum subsp. pedersenii (99.9% $; 99.9% #) (Figs. 2 and 3; Table 2).

* Data

from Wiersema (1987) and Amaral (2013).

† Data

from Moore and Jameson (2013).

No marked differences in floral scent composition between pistillate and staminate phases of anthesis could be evidenced, with the sole exception of the analyzed samples of N. rudgeana ($, n = 4; #, n = 2), in which the emission of (methoxymethyl)benzene (5) was dramatically reduced during the staminate phase (Fig. 3; Table 2).

The rates of floral scent emission differed between species (PERMANOVA: df = 6, F = 46.709, R 2 = 0.9, P = 0.031) and phases of anthesis (PERMANOVA: df = 1, F = 5.043, R 2 = 0.016, P = 0.031). The interaction between these two factors had no significant effect (PERMANOVA: df = 4, F = 1.477, R 2 = 0.019, P = 0.262), meaning that variations of scent between the pistillate and the staminate phases showed the same pattern for each species: floral scent discharge was higher at the pistillate phase (865 ± 194 µg h – 1, n = 16) than at the staminate phase (392 ± 97 µg h – 1, n = 16).

Among the different species from subg. Hydrocallis, only N. amazonum subsp. amazonum (144 ± 29 µg h – 1, n = 11), N. lasiophylla (436 ± 54 µg h – 1, n = 6), and N. rudgeana (1654 ± 138 µg h – 1, n = 8) exhibited significantly different rates of floral scent emission. N. lingulata (497 ± 99 µg h – 1, n = 3) and N. amazonum subsp. pedersenii (389 ± 56 µg h – 1, n = 2) did not show any significant difference in floral scent emission rates when compared to any of the other sampled species, and only a single sample of N. tenerinervia (121 µg h – 1) and N. gardneriana (275 µg h – 1) each were analyzed.