1314
erythro
trilobacin (7)
trilobalicin (8)
bullatalicin (9)
new acetogenins.38 Their quantitative estimation is now routine using HPLC coupled with tandem mass spectrometry (HPLC/MS/MS), and these methods are quite successful in screening studies; for example, 44 acetogenins, including four new ones, were detectable in 2 μg samples of crude methanolic extracts of Rollinia mucosa.39 Structural determinations of the acetogenins have been facilitated by carefully studying the fragments produced during electron impact-mass spectrometry (EI-MS); for example, by analyzing the trimethylsilyl (TMS) and deutero-TMS derivatives, using EIMS and fast atom bombardment mass spectrometry (FABMS), the positions of the hydroxyl groups along the hydrocarbon chain can be determined.
The acetogenins are waxy, do not crystallize readily, and, thus, are not readily amenable to X-ray crystallographic analyses, so a combination of 1H and 13C nuclear magnetic resonance (NMR) methods has evolved to aid in their structural elucidations. Synthetic single-THF and adjacent bis-THF models40,41 have been indispensable. Three of my graduate students (Kent Rupprecht, Matt Reiser, and Yu-Hua Hui) worked closely with the group of Thomas Hoye at the University of Minnesota to apply Mosher ester methods to solve the absolute stereochemistry of the carbinol centers, especially those flanking the THF subunits.42 Zhe-Ming Gu devised micromethods for preparing formaldehyde acetals that, upon 1H NMR analyses, identify erythro/threo relationships among vicinal and nearby diols.43 Such strategies for structural elucidation, the revisions of erroneous and incomplete structures published in the literature, a brief mention of several synthesis methods, and the biological effects are covered in our review papers previously cited. New methods for the syntheses of the acetogenins and their congeners continue to be developed,44 and it is reassuring that our structures, as proposed from the spectral methods employed, have been confirmed, so far. Asimicin (1), bullatacin (6), and trilobacin (7) have all been synthesized.45,46 Composition of matter patents have been assigned to Purdue University protecting the rights to commercialization of many of the acetogenins that we have found, especially those that show cytotoxic selectivity toward pancreatic and prostate carcinomas.47-51
Previous phytochemical studies of Asimina triloba (paw paw) have led to the isolation of oil, lipids, fatty acids, and proteins from the fruits and seeds, tannins, sitosterol, caffeic acid, several flavonoids (procyanidin, quercetin, quercetin 3-glycoside, quercetin 3-rutinoside, and quercetin 3-glycoside-7-glucoside), and a number of alkaloids (asimine, which was reported to be emetic, analobine, which was once used as a medicine, coreximine, anolobine, asimilobine, isocorydine, liriodenine, and norushinsunine).52 During our bioactivity-directed work, we isolated four nonacetogenin compounds that were bioactive; these were N-p-coumaroylyramine, N-trans-feruloylyramine, (+)-syringaresinol, and squamolone,52,53 but the acetogenins are, without a doubt, the major bioactive components of paw paw.
Following our isolation of asimicin (1) from the bark and seeds of paw paw,18 continued work by Geng-Xian Zhao,52-58 Kan He,59,60 Mi-Hee Woo,61-65 and Eun-Jung Kim66,67 led to the isolation and characterization of 49 additional acetogenins from the extracts, monitoring the fractionations with the brine shrimp test. The majority (29) of the paw paw acetogenins represent the adjacent bis-THF type of compounds and can be organized into three major subtypes. The asimicin (1) subtype (asimicin, asimin, asiminacin, asiminecin, asiminocin, asimilobin, parviflorin, 2,4-cis- and trans-asimicinones, asimitrin, asitribin, asimenins A and B, and 10-hydroxyasimicin) includes 14 compounds. The bullatacin (6) subtype (bullatacin, 2,4-cis- and 2,4-trans-bullatacinones, bullatetrocin, bullatin, 30R- and 30S-bullanin, squamocin, and motrilin) includes nine compounds. The trilobacin (7) subtype (trilobacin, trilobin, 10-hydroxytrilobacin, 2,4-cis- and 2,4-trans-trilobacinones, and 4-hydroxytrilobin) includes six compounds. Most of these compounds within the subtypes differ from the parent compounds only in the addition of another hydroxyl group or a repositioning of the hydroxyl from C-4 to another position further down the chain. Only one nonadjacent bis-THF acetogenin, trilobalicin (8), has been reported, but bullatalicin (9), a nonadjacent bis-THF acetogenin originally found in Annona bullata,68 is apparent upon HPLC/MS/MS analyses of the extracts of the fruit and twigs and in the extracts of zebra swallowtail butterflies, which eat the leaves of paw paw and sequester the acetogenins as a defense against predation.69 Some 24 mono-THF (single ring) acetogenins have been found in the paw paw extracts; these are 2,4-cis- and 2,4-trans-annonacin-A-ones, 2,4-cis- and 2,4-trans-gigantetracinones, annonacin A, annonacin (3), 16,19-cis-murisolin, murisolin A, 2,4-cis- and 2,4-trans-murisolinones, gigantetrocin A, 2,4-cis- and 2,4-trans-gigantetrocin A-ones, 2,4-cis- and 2,4-trans-isoannonacins, asitrilobins A–D, annomontacin, xylomatacin, asitricin, and 2,4-cis- and 2,4-trans-asitricinones. Several additional acetogenins can be detected using HPLC/MS/MS; for example, previously unidentified peaks at m/z 620 are probably dehydro analogues of the several C37 bis-THF compounds that carry three hydroxyls. The roots have not been fractionated so far, and their potent bioactivity suggests that they might yield something new.
Our initial work with Eli Lilly and Company (Greenfield, IN) and the USDA (Peoria, IL) demonstrated that the paw paw acetogenins are potent in inhibiting a number of agricultural pests: mosquito larvae, two-spotted spider mites, Mexican bean beetles, striped cucumber beetles, European corn borers, melon or cotton aphids, blowfly larvae, and a nematode (Caenorhabditis elegans).[17-20] More recently the group of Cortes[70] evaluated the antifeedant and insecticidal effects of squamocin and annonacin (3) against three additional insect species. A number of experiments at AgriDyne Inc. (Salt Lake City, UT) are worthy of mention.[71] With Colorado potato beetles, foliar sprays of paw paw extract showed excellent results, with concentrations as low as 250 ppm being quite effective. Against white flies on cotton leaves, the paw paw extract and