Supplemental Information
ANTIPREDATOR ACTIVITY AND ENDOGENOUS BIOSYNTHESIS OF DEFENSIVE SECRETION IN LARVAL AND PUPAL Delphastus catalinae (HORN) (COLEOPTERA: COCCINELLIDAE)
STEPHEN T. DEYRUP1, LAURA E. ECKMAN2, ELEANOR E. LUCADAMO2, PATRICK H. McCARTHY2, JACQUELINE C. KNAPP2, SCOTT R. SMEDLEY2
1 Department of Chemistry and Biochemistry, Siena College, Loudonville, NY 12211 USA
2 Department of Biology, Trinity College, Hartford, CT 06106 USA
Correspondence to: Scott Smedley, email: ; phone: 860-297-2229; fax: 860-297-2538
Relative Configuration of (E,E)-2,6-diacetoxygermacra-1(10),4-diene (1)
Signals representing the olefinic protons H-1 and H-5 displayed NOESY crosspeaks with each other as well as with signals for aliphatic protons H-3β, and H-8β, allowing assignment of these groups to the same side of the ring-system. The opposite face of the ring-system was assigned based on resonances from the protons of olefinic methyl groups H3-14 and H3-15, which showed NOESY correlations to those of H-2, H-6 and H-7 as well as to each other. These correlations also allowed assignment of both double bond configurations as E. In addition, the large coupling constant between protons H-2 and H-3β supports this configurational assignment. Subsequent comparison of 1H and 13C chemical shift data and 1H J-values of 1 to literature data of the known (E,E)-6β-acetoxy-2β-hydroxygermacra-1(10),4-diene (Barrero et al. 1998) and (E,E)-2b,6b-dihydroxygermacra-l(l0),4-diene (Sanz et al. 1991) revealed a remarkably high degree of similarity, further reinforcing the assignment of the relative configuration as shown in 1. The absolute configuration of 1 was not determined due to limited sample quantity (350 µg).
Table S1. NMR spectral data for (E,E)-2,6-diacetoxygermacra-1(10),4-diene (1)
Position / δH (mult., JHH)a / δCb / HMBC (HàC#)a / NOESY (HàH)c1 / 5.04 (br d, 11) / 128.9 / 9, 14 / 3β, 5, 8β, 9β
2 / 5.60 (dt, 5.5, 11) / 69.6 / 1, 10, 16 / 3α, 14, 15
3α / 2.53 (dd, 11, 5.5) / 44.8 / 1, 2, 4, 5, 15 / 2, 3β, 15
3β / 2.18 (t, 11) / 1, 2, 4, 5, 15 / 1, 3α, 5
4 / – / 131.5 / – / –
5 / 5.13 (br d, 7.2) / 133.7 / 2d, 3, 4, 15 / 1, 3β, 8β
6 / 5.53 (dd, 7.2, 2.1) / 72.6 / 4, 5, 7, 8, 18 / 7, 12, 13, 15
7 / 0.94 (m) / 48.7 / 8α, 8β, 9α, 14, 15
8α / 1.50 (m) / 30.7 / 10 / 7, 8β, 9α, 9β
8β / 1.91 (m) / 1, 5, 8α, 9β
9α / 1.68 (dt, 5, 14) / 36.1 / 1, 10 / 7, 8α, 9β
9β / 2.45 (br d, 14) / 1, 7, 8, 10, 14 / 1, 8α, 8β, 9α
10 / – / 140.3 / – / –
11 / 1.58 (m) / 32.0 / 7 / 8α, 12, 13
12 / 0.93 (d, 6.7) / 21.0 / 7, 11, 13 / 6, 11
13 / 0.98 (d, 6.7) / 21.1 / 7, 11, 12 / 6, 11
14 / 1.79 (br s) / 22.9 / 1, 9, 10 / 2, 7, 15
15 / 1.59 (br s) / 17.7 / 3, 4, 5 / 2, 3α, 6, 7, 14
16 / – / 170.8 / – / –
17 / 1.99 (s) / 21.4 / 16
18 / – / 170.7 / – / –
19 / 2.00 (s) / 21.3 / 18
a Recorded at 600 MHz in methylene chloride-d2. b Determined from HMQC and HMBC data.
c Recorded in acetone-d6. d Weak four-bond correlation.
Fig. S1. 1H NMR spectrum of 1 at 600 MHz in acetone-d6
Fig. S2. dqfCOSY spectrum of 1 at 600 MHz in acetone-d6
Fig. S3. NOESY spectrum of 1 at 600 MHz in acetone-d6
Fig. S4. dqfCOSY of crude pupal surface wash at 600 MHz in methylene chloride-d2
Fig. S5. HMBC of crude pupal surface wash at 600 MHz in methylene chloride-d2
Fig. S6. HMQC of crude pupal surface wash at 600 MHz in methylene chloride-d2
Fig. S7. HPLC-MS chromatograms recorded for the 13C-glucose labeling study. A) A representative HPLC-MS trace for pupal surface wash of the “unlabeled” treatment group. B) A representative HPLC-MS trace for pupal surface wash of the “labeled” treatment group. C) Structure of (E,E)-2,6-diacetoxygermacra-1(10),4-diene (1), which eluted at the peaks indicated by the red arrows.
Fig. S8. Portions of the mass spectrum that display the isotopic ratios for (E,E)-2,6-diacetoxygermacra-1(10),4-diene (1) from the HPLC-MS 13C-glucose labeling study. A) A portion of the mass spectrum of 1 from the “unlabeled” treatment group. B) A portion of the mass spectrum of 1 from the “labeled” treatment group. C) Structure of 1 along with a summary of the isotopic ratios.
Fig. S9. HPLC-MS chromatograms recorded for the 13C-glucose labeling study. A) A representative HPLC-MS trace for pupal surface wash of the “unlabeled” treatment group. B) A representative HPLC-MS trace for pupal surface wash of the “labeled” treatment group. C) Structure of catalipyrone H, which eluted at the peaks indicated by the red arrows.
Fig. S10. Portions of the mass spectrum that display the isotopic ratios for catalipyrone H from the HPLC-MS 13C-glucose labeling study. A) A portion of the mass spectrum of catalipyrone H from the “unlabeled” treatment group. B) A portion of the mass spectrum of catalipyrone H from the “labeled” treatment group. C) Structure of catalipyrone H along with a summary of the isotopic ratios.
Fig. S11. HPLC-MS chromatograms recorded for the 13C-glucose labeling study. A) A representative HPLC-MS trace for pupal surface wash of the “unlabeled” treatment group. B) A representative HPLC-MS trace for pupal surface wash of the “labeled” treatment group. C) Structure of catalipyrone I, which eluted at the peaks indicated by the red arrows.
Fig. S12. Portions of the mass spectrum that display the isotopic ratios for catalipyrone I from the HPLC-MS 13C-glucose labeling study. A) A portion of the mass spectrum of catalipyrone I from the “unlabeled” treatment group. B) A portion of the mass spectrum of catalipyrone I from the “labeled” treatment group. C) Structure of catalipyrone I along with a summary of the isotopic ratios.
Fig. S13. HPLC-MS chromatograms recorded for the 13C-glucose labeling study. A) A representative HPLC-MS trace for pupal surface wash of the “unlabeled” treatment group. B) A representative HPLC-MS trace for pupal surface wash of the “labeled” treatment group. C) Structure of catalipyrone J, which eluted at the peaks indicated by the red arrows.
Fig. S14. Portions of the mass spectrum that display the isotopic ratios for catalipyrone J from the HPLC-MS 13C-glucose labeling study. A) A portion of the mass spectrum of catalipyrone J from the “unlabeled” treatment group. B) A portion of the mass spectrum of catalipyrone J from the “labeled” treatment group. C) Structure of catalipyrone J along with a summary of the isotopic ratios.
Fig. S15. 1H NMR spectrum of the crude extract of adult Delphastus catalinae, at 600 MHz in acetone-d6
Fig. S16. 1H NMR spectrum of the crude extract of adult whiteflies, at 600 MHz in methylene chloride-d2
Fig. S17. 1H NMR spectrum of the crude extract of immature whiteflies, at 600 MHz in methylene chloride-d2
Fig. S18. 1H NMR spectrum of the crude extract of D. catalinae pupae reared on a diet of whiteflies infesting tobacco, at 600 MHz in acetone-d6
Fig. S19. 1H NMR spectrum of the crude extract of D. catalinae pupae reared on a diet of whiteflies infesting collards, at 600 MHz in acetone-d6
References
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2. Sanz JF, García-Sarrión A, Marco JA (1991) Germacrane derivatives from Santolina chamaecyparissus. Phytochemistry 30: 3339-3342