Papilio machaon subsp. hippocrates C. Felder et R. Felder 1864
Creators: name: Nishidono, Yuto
name: Niwa, Kotaro
name: Kitajima, Aoi
name: Watanabe, Shiro
name: Tezuka, Yasuhiro
name: Arita, Masanori
name: Takabayashi, Junji
name: Tanaka, Ken
Publisher: name: Zenodo
Year: 2021
Subjects: Biodiversity
Taxonomy
Animalia
Arthropoda
Insecta
Lepidoptera
Papilionidae
Papilio
Papilio machaon
P. machaon hippocrates

description: 2.1. Chemical analyses of the regurgitant of P. machaon hippocrates larvae   
 Larval regurgitant naturally contains large amounts of components derived from damaged plants. However, elicitor compounds derived from larvae are considered to be steadily produced regardless of diet. Therefore, we analyzed the regurgitant of larvae feeding on two different plant species (A. acutiloba or G. littoralis (Apiaecae)). Among the plants in the Apiaecae family, both A. acutiloba and G. littoralis are particularly susceptible to damage by the larvae of P. machaon hippocrates. Positive and negative total ion current (TIC) chromatograms of the regurgitant of P. machaon hippocrates larvae that had infested either A. acutiloba or G. littoralis are shown in Fig. 1A and B. To detect the compounds in the regurgitant comprehensively, LC–MS analyses in both positive and negative ionization modes were performed. The reported [M+H] + and [M H] ions of volicitin and its related compounds, such as N -linolenoyl-L-glutamine and N -linolenoyl-L-glutamic acid, were monitored to examine the presence of the compounds (Fig. 1). The peaks were annotated as shown in Tables S1 and S 2. In the mass chromatograms of both ionization modes, volicitin and its related compounds were not detected. These results were consistent with those of a previous report (Mori and Yoshinaga, 2011). In contrast, high concentrations of FAs were detected in the regurgitant as compared with the concentrations of other specialized metabolites in the leaves (Fig. 1A–D). The structures of fatty acids in the regurgitant were elucidated as α- LA, linolenic acid, oleic acid, palmitic acid, and stearic acid, based on the retention time and mass fragmentation patterns of authentic compounds by GC–MS analysis (Fig. 2). Compositions of FAs in the regurgitant of larvae parasitized on A. acutiloba or G. littoralis and those of the respective leaves are shown in Table 1. Among the FAs accumulated in the regurgitant of P. machaon hippocrates larvae, the enhancement rate of α- LA was remarkable. It is known that FAs in butterfly larvae are derived from the diet and the FA composition varies among the species of the insect and the host plants. For example, Morpho peleides Kollar, 1850 accumulate more polyunsaturated FAs (Wang et al., 2006), whereas Plodia interpunctella (Hübner, 1813) and Cadra cautella (Walker, 1863) accumulate more saturated and monounsaturated FAs (Subramanyam and Cutkomp, 1987).   
 Since FACs are biosynthesized from FAs accumulated in the body (Pare´et al., 1998), FACs and polyunsaturated FAs, such as α- LA, are often detected simultaneously (Alborn et al., 2003; Mori et al., 2003). Although P. machaon hippocrates larvae accumulate α- LA, no FACs were detected in the regurgitant. Regarding the elicitor activity of FACs, it has been reported that only FACs with the α- LA moiety are active (Alborn et al., 1997). In P. machaon hippocrates larvae, biosynthesis of FACs from accumulated α- LA may render the host plants unsuitable for feeding. Therefore, it is reasonable to assume that the absence of FACs is appropriate for the survival of P. machaon hippocrates, a highly evolved species in the order Lepidoptera (Muto-Fujita et al., 2017; Richard and Guedes, 1983). Conversely, considering the coevolution of insects and plants, it is reasonable to assume that plants will also evolve to respond to compounds accumulated in insects. Among the compounds detected in the regurgitant of P. machaon hippocrates larvae, we focused on α- LA because its bioenhancement rate is characteristically high, and it is generally accepted that the biosynthesis of JA begins with the oxygenation of free α- LA (Creelman and Mullet, 1997).
description: Published as part of Nishidono, Yuto, Niwa, Kotaro, Kitajima, Aoi, Watanabe, Shiro, Tezuka, Yasuhiro, Arita, Masanori, Takabayashi, Junji & Tanaka, Ken, 2021, α-Linolenic acid in Papilio machaon larvae regurgitant induces a defensive response in Apiaceae, pp. 1-7 in Phytochemistry (112796) (112796) 188 on page 2, DOI: 10.1016/j.phytochem.2021.112796, http://zenodo.org/record/8259205