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[reaction: see text] Dichlorocyclopropanation of (-)-O-menthyl acrylate under conditions of phase-transfer catalysis (CHCl3, KOH, tetramethylammonium bromide), with sonication, gives excellent yields (85-94%) of the corresponding dichlorocyclopropanecarboxylate ester compared to thermal conditions (90 degrees C, 56%). No diastereoselectivity was observed, but one isomer was isolated pure by fractional crystallization. The measured kinetic isotope effect (initial rate (CHCl3)/rate (CDCl3) approximately 1.7) suggests deprotonation of CHCl3 as the rate-limiting step.

Glucosinolates are a class of secondary metabolites with important roles in plant defense and human nutrition. Here, we characterize a putative UDP-glucose:thiohydroximate S-glucosyltransferase, UGT74B1, to determine its role in the Arabidopsis glucosinolate pathway. Biochemical analyses demonstrate that recombinant UGT74B1 specifically glucosylates the thiohydroximate functional group. Low Km values for phenylacetothiohydroximic acid (approximately 6 microm) and UDP-glucose (approximately 50 microm) strongly suggest that thiohydroximates are in vivo substrates of UGT74B1. Insertional loss-of-function ugt74b1 mutants exhibit significantly decreased, but not abolished, glucosinolate accumulation. In addition, ugt74b1 mutants display phenotypes reminiscent of auxin overproduction, such as epinastic cotyledons, elongated hypocotyls in light-grown plants, excess adventitious rooting and incomplete leaf vascularization. Indeed, during early plant development, mutant ugt74b1 seedlings accumulate nearly threefold more indole-3-acetic acid than the wild type. Other phenotypes, however, such as chlorosis along the leaf veins, are likely caused by thiohydroximate toxicity. Analysis of UGT74B1 promoter activity during plant development reveals expression patterns consistent with glucosinolate metabolism and induction by auxin treatment. The results are discussed in the context of known mutations in glucosinolate pathway genes and their effects on auxin homeostasis. Taken together, our work provides complementary in vitro and in vivo evidence for a primary role of UGT74B1 in glucosinolate biosynthesis.

2-Aryl-1-N-carboalkoxyenamines (enamides) are selectively reduced to the corresponding 2-arylethylamine carbamates by Et3SiH in the presence of CF3COOH in excellent yields. The reduction proceeds by addition of hydride at C-1 and the rate-limiting step involves proton transfer from CF3COOH. This reduction is useful for preparation of isotopically labeled arylethylamines.

A chemoselective conversion of bastadin-4 to the important Ca2+ channel modulator bastadin-5 (1a) has been achieved using cationic hydrogenation (Et3SiH, TFA, 60%). Specifically deuterated bastadin-5 (1b, >95 at. %) was prepared following this method and the simplified 1H NMR H-5/H2-6 spin system of 1b exploited to study temperature-dependent macrocyclic ring dynamics.

2-Aryl-1-N-carboalkoxyenamines (enamides) are selectively reduced to the corresponding 2-arylethylamine carbamates by Et3SiH in the presence of CF3COOH in excellent yields. The reduction proceeds by addition of hydride at C-1 and the rate-limiting step involves proton transfer from CF3COOH. This reduction is useful for preparation of isotopically labeled arylethylamines.

An unprecedented 24-N-imidazolyl steroidal alkaloid, amaranzole A, was isolated from a tropical sponge, Phorbas amaranthus. The structure was solved by interpretation of MS, NMR, including a novel exciton-coupled CD spectrum due to an allyl imidazole, and comparison with model compounds.

Bastadin-5, a brominated macro-dilactam from the marine sponge Ianthella basta, enhances release of Ca2+ from stores within the sarcoplasmic reticulum (SR) of muscle and nonmuscle cells by modulating RyR1/FKBP12 complex. Analogues of bastadin-5 present desirable targets for SAR studies to shed light on the gating mechanism and locus of bastadin-5 binding on these heteromeric channels that mediate essential steps in early coupling of membrane excitation to Ca2+ signaling cascades. Simple, ring-constrained analogues of bastadin-5 were synthesized from substituted benzaldehydes in a convergent manner, featuring an efficient S(N)Ar macroetherification, and evaluated in an assay that measures [3H]-ryanodine that is known to correlate with the functional open state of the Ca2+ channel. The simplified 14-membered ring, atropisomeric analogue (+/-)-7, like bastadin-5, enhanced ryanodine binding to the RyR1/FKBP12 complex (EC50 11 microM), however, unexpectedly, the corresponding achiral 18-membered ring analogue 14 potently inhibited binding (IC50 6 microM) under the same conditions. Structure-activity relationships of both families of cyclic analogues showed activity in a ryanodine binding assay that varied with substitutions of the Br atom on the trisubstituted aryl ring by various functional groups. The most active analogues were those that conserved the dibromocatechol ether moiety that corresponds to the 'western edge' of the bastadin-5 structure. These data suggest that cyclic analogues of bastadin-5 interact with the channel complex in a complex manner that can either enhance or inhibit channel activity.

Glucosinolates are a class of secondary metabolites with important roles in plant defense and human nutrition. Here, we characterize a putative UDP-glucose:thiohydroximate S-glucosyltransferase, UGT74B1, to determine its role in the Arabidopsis glucosinolate pathway. Biochemical analyses demonstrate that recombinant UGT74B1 specifically glucosylates the thiohydroximate functional group. Low Km values for phenylacetothiohydroximic acid (approximately 6 microm) and UDP-glucose (approximately 50 microm) strongly suggest that thiohydroximates are in vivo substrates of UGT74B1. Insertional loss-of-function ugt74b1 mutants exhibit significantly decreased, but not abolished, glucosinolate accumulation. In addition, ugt74b1 mutants display phenotypes reminiscent of auxin overproduction, such as epinastic cotyledons, elongated hypocotyls in light-grown plants, excess adventitious rooting and incomplete leaf vascularization. Indeed, during early plant development, mutant ugt74b1 seedlings accumulate nearly threefold more indole-3-acetic acid than the wild type. Other phenotypes, however, such as chlorosis along the leaf veins, are likely caused by thiohydroximate toxicity. Analysis of UGT74B1 promoter activity during plant development reveals expression patterns consistent with glucosinolate metabolism and induction by auxin treatment. The results are discussed in the context of known mutations in glucosinolate pathway genes and their effects on auxin homeostasis. Taken together, our work provides complementary in vitro and in vivo evidence for a primary role of UGT74B1 in glucosinolate biosynthesis.

Two new sulfate monoesters of hemibastadins-1 and -2 were isolated from the marine sponge Ianthella basta (Pallas) from Guam. A third new compound was tentatively assigned the structure 34-O-sulfatobastadin-9. The 1-O-sulfatohemibastadins-1 and –2 were antagonists of the RyR1-FKBP12 Ca2+ channel under conditions where the known compound bastadin-5 exhibits potent agonism (EC50 2μM).

The sponge Phorbas amaranthus from Florida contains the new ring A-contracted steroids, phorbasterones A-D, and the known anthosterones A and B. The structures of phorbasterones A-D were determined by interpretation of their spectroscopic data. Phorbasterones show moderate cytotoxicity against HCT-116 tumor cells.

Two new chlorocyclopropane macrolides, phorbasides A and B, have been characterized from the sponge Phorbas sp. that previously yielded phorboxazoles A and B. We describe the assignment of the absolute configuration of the trans-chlorocyclopropane ring that exploits a CD Cotton effect arising from hyperconjugation to the ene-yne chromophore. Phorbasides and callipeltoside A share the same macrolide configurations but, unexpectedly, opposite cyclopropane configurations.

An unprecedented 24-N-imidazolyl steroidal alkaloid, amaranzole A, was isolated from a tropical sponge, Phorbas amaranthus. The structure was solved by interpretation of MS, NMR, including a novel exciton-coupled CD spectrum due to an allyl imidazole, and comparison with model compounds.