Relationship between Stereoisomerism and Biological Activity of Pyrethroids

Higher homologous acids of chrysanthemic acid described in the previous papers were esterified with (±)-allethrolone. The toxicity of these esters and the related compounds were evaluated by the topical application method to Musca domestica vicina Macq. The allethronyl homochry-santhemate (entry Nos. 4, 5) was shown to be toxic and the dextrorotatory form was far more toxic than the laevorotatory one. Further elongation of the ester linkage resulted in a loss of toxicity. The cyclobutane carboxylic acid ester (entry No. 10) was shown to be toxic and so, the cyclopropane ring might be replaced to some extent by the cyclobutane ring, provided the other requirements were fulfiled. However, further elongation of the ester linkage also reduced the toxicity. The lactones (entry Nos. 12–16) obtained by the hot sulfuric acid treatment were non-toxic.     

Synthesis and Insecticidal Activity of Isobutenyl- and Isobutyl-cyclopropanecarboxylates

Several isobutenyl- and isobutyl-cyclopropanecarboxylates were synthesized. The insecticidal activities against the housefly of their 5-benzyl-3-furylmethyl or allethronyl esters were tested. Among the substituents on the cyclopropane ring, a methyl group cis to the ester linkage has been found to be essential for toxicity against the housefly and a trans-isobutenyl group greatly enhances the toxic activity.     

Studies on the Syntheses of the Pyrethrin Analogues and their Biological Activities

The separation of (±) -2,2-dimethyl-3- (3′,4′-methylenedioxyphenyl) -cyclopropane-1-carboxylic acid into the geometrical isomers and the assignment of their configurations were achieved. Of the two isomers, the (±) -trans-acid, which was found more toxic when esterified with (±) -allethrolone, was resolved by means of an optically active α-phenylethylamine salt into (+) - and (-) -enantiomers. (IR:3R) -Configuration was assigned to the (+) -trans-acid and (IS:3S) -configuration to the (-) -trans-acid. The bioassay revealed that the (±) -allethrolone ester with the (+) -trans-acid, which belongs to the same optical series as the natural chrysanthemum acids, was the most toxic against common houseflies, as was the case with other pyrethroids.     

Second Generation of cycloSal‐Pronucleotides with Esterase‐Cleavable Sites: The ”Lock‐In”‐Concept

A conceptual extension of the cycloSal‐pronucleotide approach is presented. The characteristic feature of the new cycloSal‐derivatives of the anti‐HIV active nucleoside analogue d4T 1 is the incorporation of an enzymatically cleavable carboxylic ester moiety with the intention to trap the triesters inside cells (”lock‐in”‐concept). CycloSal‐triesters bearing different ester groups in the 3‐or 5‐position of the cycloSal‐moiety are described. Surprisingly, only acetyl‐and levulinyl esters are cleaved readily in CEM cell extracts while alkyl esters were found to be stable. Nevertheless, in in‐vitro anti‐HIV assays most of the compounds achieve the thymidine–kinase bypass, thus proving that they act at least as nucleotide delivery systems.     

Relationship between Stereoisomerism and Biological Activity of Pyrethroids

(+)-trans-Homochrysanthemic acid, when boiled in dilute sulfuric acid, gives (+)-trans-ε-hydroxy-dihydrohomochrysanthemic acid, m.p. 176–7°, together with (+)-δ, δ-dimethyl-γ-isobutenyl-δ-valerolactone. The formation of optically active lactone from (+)-trans-homochrysanthemic acid provides another cogent evidence for the structure of the lactone previously deduced on the racemic compound.

The Arndt-Eistert reaction of the homo-acids give further higher homologues such as (±)-,(+)-trans-β-(3-isobutenyl-2, 2-dimethylcyclopropane-1)-propionic acids and (±)-cis-3-isobutenyl-2, 2 dimethylcyclobutane-1-acetic acid. Both trans-acids, in boiling dilute sulfuric acid, give the same (±)-γ-(1′, 1′, 4′-trimethyl-pent-2′-enyl)-butyrolactone together with the corresponding hydroxy-acids, optically inactive and active, respectively.

Complete resolution of (±)-trans-homochrysanthemic acid and (±)-trans-β-(3-isobutenyl-2, 2-dimethycyclopropane-1)-propionic acid was achieved by means of optically active α-phenylethylamine.     

Synthesis of 3-O-β-N-Acetylglucosaminyl Cyclic Tetrasaccharide through a Lysozyme-catalyzed Transfer Reaction

Egg white lysozyme was found to catalyze the transfer of N-acetylglucosamine to cyclo{→6)-α-D-Glcp-(1→3)-α-D-Glcp-(1→6)-α-D-Glcp-(1→3)-α-D-Glcp-(1→} (CTS). Structural analysis showed that the transfer product was3-O-β-N-acetylglucosaminyl CTS, cyclo{→6)-α-D-Glcp-(1→3)-α-D-Glcp-(1→6)-[β-GlcNAc-(1→3)]-α-D-Glcp-(1→3)-α-D-Glcp-(1→}. This branched saccharide is anticipated to be a model compound of the sugar chains of glycoproteins.     

Juvadecene: Discovery of a juvenile hormone mimic in the plant,Macropiper excelsum

A compound with significant insect juvenile hormone activity was isolated from the plant, Macropiper excelsum. The chemical structure was determined by spectral methods to be 1-(3,4-methylenedioxyphenyl)-trans-3-decene(l), and confirmed by synthesis. The hormonally active substance applied topically to last (fifth)-instar nymphs of the milkweed bug (Oncopeltus fasciatus) induced a supernumerary metamorphosis at 30 μg. Higher doses were toxic.     

Rapid identification ofRhizobium species based on cellular fatty acid analysis

As understanding of the evolutionary relationships between strains and species of root nodule bacteria increases the need for a rapid identification method that correlates well with phylogenetic relationships is clear. We have examined 123 strains ofRhizobium: R. fredii (19),R. galegae (20),R. leguminosarum (22),R. loti (17),R. meliloti (21), andR. tropici (18) and six unknowns. All strains were grown on modified tryptone yeast-extract (TY) agar, as log phase cultures, scraped from the agar, lysed, and the released fatty acids derivatized to their corresponding methyl esters. The methyl esters were analysed by gas-chromatography using the MIDI/Hewlett-Packard Microbial Identification System. All species studied contained 16:0, 17:0, 18:0 and 19cyclo_w9C fatty acids but onlyR loti andR tropici produced 12:0 3 OH,13:0 iso 3 OH,18:1_w9C and 15:0 iso 3 OH,17:0 iso 3 OH and 20:2_w6,9C fatty acids respectively. Principal component analysis was used to show that strains could be divided into clusters corresponding to the six species. Fatty acid profiles for each species were developed and these correctly identified at least 95% of the strains belonging to each species. A dendrogram is presented showing the relationships betweenRhizobium species based on fatty acid composition. The data base was used to identify unknown soil isolates as strains ofRhizobium lacking a symbiotic plasmid and a bacterium capable of expressing a symbiotic plasmid fromR. leguminosarum asSphingobacterium spiritovorum.     

Competition between cis,trans and Cyclopropane Fatty Acid Formation and its Impact on Membrane Fluidity

The impact of cis, trans and cyclopropane fatty acids on membrane fluidity was investigated using batch‐grown Pseudomonas putida P8 and Comamonas testosteroni ATCC 17454. A major difference observed between the two investigated strains is the absence of the ability to synthesize trans‐unsaturated fatty acids in Comamonas. When grown exponentially at 30 °C, a shift to 35 °C increased the trans/cis ratios of the fatty acids of P. putida P8 from 0 to 0.81 and 0 to 1.07, in lipid extracts and cell hydrolyzates, respectively. After prolonged growth followed by nutrient deprivation for 48 h, both at 30 °C, trans fatty acids were absent, but the cyclo/cis ratios rose from 0.1 to 1.55 in lipid extracts, and from 0.1 to 1.54 in cell hydrolyzates. C. testosteroni ATCC 17454 contained no cyclo fatty acids when harvested in the exponential phase after 6 h, whereas after 72 h cultivation, the cyclo/cis ratios rose to 0.49 and 0.47, in lipid extracts and cell hydrolyzates, respectively. Trans fatty acids were never observed in this strain. Increased cyclo/cis and trans/cis ratios correlated with decreased fluidity measured by the fluorescence anisotropy of 1,6‐diphenyl‐1,3,5‐hexatriene (DPH) intercalated in the bilayers of liposomes and by Fourier Transform Infrared (FTIR) spectroscopy of lipids prepared from the cells. The specific effect of cyclopropane fatty acids on membrane fluidity was much smaller than that of trans fatty acids. FTIR‐measurements of intact cells of P. putida P8 confirmed the high potency of trans fatty acids to decrease the fluidity. In cells with induced cyclopropane fatty acid synthesis, the membranes remained more fluidized, indicating the lower importance of these fatty acids for homeoviscosis.     

SYNTHESIS,HYDROLYSIS AND ANTI-EBV ACTIVITY OF A SERIES OF 3′-MODIFIED cycloSal-BVDUMP PRONUCLEOTIDES

A series of cycloSal-BVDUMP phosphate triesters has been prepared. The prototype compound was 3-methyl-cycloSal-BVDUMP 2. Furthermore, a series of 3′-O-acyl-modified derivatives having carboxylic acids with different lipophilicity or a L-configurated α-amino acid (phenylalanine) was prepared. The hydrolysis properties in phosphate buffer PBS as well as in PBS containing pig liver esterase (PLE) will be described. Finally, the biological activity against EBV has been determined.     

Conformation of dioxopiperazines,II. CNDO/2 quantum mechanical calculations on conformational preferences in cyclo (glycyl-L-alanyl), cyclo(glycyl-L-valyl), and both epimers of cyclo-di-(alanyl)

Conformational energy calculations, aimed at verification of the suitability of the semiempirical molecular orbital CNDO/2 method for conformational elucidations in cyclic dipeptides formed from amino acids with aliphatic side chains, have been carried out. The results obtained for four dioxopiperazines [DOP; cyclo(Glycyl-L-Alanyl), cyclo(Glycyl-L-Valyl), and both epimers of cyclo-di-(Alanyl)] point out very good agreement with experimental premises. The latter include (1) the preference of the cis-peptide bonds for being nonplanar, which results in twisted-boat conformations of the DOP ring; (2) greater stability of conformers with a side chain oriented axially over those with a side chain oriented equatorially; (3) the preference of cyclo(Gly-Val) for assuming a folded conformation with one of the side chain γ-methyl groups sticking over the DOP ring.     

Studies on Abscisic Acid

Methyl α-ionylideneacetates were oxidized with selenium dioxide to a mixture of methyl 3′-keto-β-ionylideneacetates and a small amount of methyl 4′-keto-α-ionylidene-acetates followed by treatment with active manganese dioxide. By a similar oxidation methyl 3′-keto-β-ionylideneacetates were prepared from methyl β-ionylidene acetates. Methyl 4′-keto-α-ionylideneacetates were obtained by oxidation of methyl α-ionylideneacetates with tert-butyl chromate. Dehydrobromination of methyl bromoionylideneacetate, obtained by bromination of methyl 2-trans-α-ionylideneacetate with N-bromosuccinimide, gave a mixture of methyl 2-trans-dehydro-β-ionylideneacetate and methyl 2-cis-dehydro-β-ionylideneacetate. The growth inhibitory activities of these sesquiterpene carboxylic acids and keto esters on rice seedlings were tested.     

CycloSaligenyl Pronucleotides of 5-Iodo and 5-Trifluoromethyl-1-(2-deoxy-β-D-ribofuranosyl)-2,4-difluorobenzene Mimics of Thymidine: Synthesis and Evaluation of this Pronucleotide Monophosphate Delivery System for Compounds with Potential Anticancer Activity

Abstract

A group of unnatural 1-(2-deoxy-β-D-ribofuranosyl)-2,4-difluorobenzenes possessing a 5-I or 5-CF₃ substituent, that were originally designed as thymidine mimics, were coupled via their 5′-OH group to a cyclosaligenyl (cycloSal) ring system having a variety of C-3 substituents (Me, OMe, H). The 5′-O-cycloSal-pronucleotide concept was designed to effect a thymidine kinase-bypass, thereby providing a method for the intracellular delivery and generation of the 5′-O-monophosphate for nucleosides that are poorly phosphorylated. The 5′-O-cycloSal pronucleotide phosphotriesters synthesized in this study were obtained as a 1:1 mixture of two diastereomers that differ in configuration (S _P or R _P) at the asymmetric phosphorous center. The (S _P)- and (R _P)-diastereomers for the 5′-O-3-methylcycloSal- and 5′-O-3-methoxycycloSal derivatives of 1-(2-deoxy-β-D-ribofuranosyl)-2,4-difluoro-5-iodobenzene were separated by silica gel flash column chromatography. This class of cycloSal pronucleotide compounds generally exhibited weak cytotoxic activities in a MTT assay (CC₅₀ values in the 10^?3 to 10^?4 M range), against a number of cancer cell lines (143B, 143B-LTK, EMT-6, Hela, 293), except for cyclosaligenyl-5′-O-[1′-(2,4-difluoro-5-iodophenyl)-2′-deoxy-β-D-ribofuranosyl]phosphate that was more potent (CC₅₀ values in the 10^?5 to 10^?6 M range), than the reference drug 5-iodo-2′-deoxyuridine (IUDR) which showed CC₅₀ values in the 10^?3 to 10^?5 M range.     

Design and synthesis of biologically active peptides: a 'tail' of amino acids can modulate activity of synthetic cyclic peptides

In earlier work, we synthesized a cyclic 9-amino acid peptide (AFPep, cyclo[EKTOVNOGN]) and showed it to be useful for prevention and therapy of breast cancer. In an effort to explore the structure–function relationships of AFPep, we have designed analogs that bear a short ‘tail’ (one or two amino acids) attached to the cyclic peptide distal to its pharmacophore. Analogs that bore a tail of either one or two amino acids, either of which had a hydrophilic moiety in the side chain (e.g., cyclo[EKTOVNOGN]FS) exhibited greatly diminished biological activity (inhibition of estrogen-stimulated uterine growth) relative to AFPep. Analogs that bore a tail of either one or two amino acids which had hydrophobic (aliphatic or aromatic) side chains (e.g., cyclo[EKTOVNOGN]FI) retained (or had enhanced) growth inhibition activity. Combining in the same biological assay a hydrophilic-tailed analog with either AFPep or a hydrophobic-tailed analog resulted in decreased activity relative to that for AFPep or for the hydrophobic-tailed analog alone, suggesting that hydrophilic-tailed analogs are binding to a biologically active receptor. An analog with a disrupted pharmacophore (cyclo[EKTOVGOGN]) exhibited little or no growth inhibition activity. An analog with a hydrophilic tail and a disrupted pharmacophore (cyclo[EKTOVGOGN]FS) exhibited no growth inhibition activity of its own and did not affect the activity of a hydrophobic-tailed analog, but enhanced the growth inhibition activity of AFPep. These results are discussed in the context of a two-receptor model for binding of AFPep and ring-and-tail analogs. We suggest that tails on cyclic peptides may comprise a useful method to enhance diversity of peptide design and specificity of ligand–receptor interactions.     

Distribution of C22-, C24- and C26-α-Unit-Containing Mycolic Acid Homologues in Mycobacteria

There are three mycolic acid homologues with C₂₂-, C₂₄- and C₂₆-α-units in Mycobacterium. In order to reveal the composition and distribution of these homologues in each subclass and molecular species of mycolic acids and to compare them with the composition of constitutive non-polar fatty acids (free and bound forms), we have separated non-polar fatty acids and each subclass of mycolic acids from 21 mycobacterial species by thin-layer chromatography, and analyzed non-polar fatty acid methyl esters by gas chromatography (GC) and the cleavage products of methyl mycolate by pyrolysis GC. We further performed mass chromatographic analysis of trimethylsilyl (TMS) ether derivatives of mycolic acid methyl esters by monitoring [B-29]⁺ ions (loss of CHO from the α-branched-chain structure of mycolic acids) of m/z 426, 454 and 482 which are attributed to C₂₂-, C₂₄- and C₂₆-α-units of TMS ether derivatives of methyl mycolates, respectively, (Kaneda, K. et al, J. Clin. Microbiol. 24: 1060-1070, 1986). By pyrolysis GC, C_22:0, C_24:0 and C_26:0 fatty acid methyl esters generated by the C₂-C₃ cleavage of C₂₂-, C₂₄- and C₂₆-α-unit-containing mycolic acid methyl esters, respectively, were detected. Their proportion was almost the same among subclasses of mycolic acids in every Mycobacterium and also similar to the proportion of constitutive non-polar C_22:0, C_24:0 and C_26:0 fatty acids. By mass chromatography, the composition and distribution of C₂₂- and C₂₄-α-unit-containing homologues were revealed to be similar between α- and α'-mycolic acids in every Mycobacterium. We further analyzed in detail M. vaccae and demonstrated that the mass chromatogram of C₂₂-α-unit-containing homologue was analogous in shape to that of the C₂₄-α-unit-containing one, with the latter mass chromatogram being up-shifted from the former by two carbon numbers, in every subclass of α-, α'-, keto and dicarboxy mycolic acids. The present study suggests that the compositions of three homologues of both mycolic acids and constitutive non-polar fatty acids, which are characteristic to each mycobacterial species, may reflect the proportion of the amount of free C_22:0, C_24:0 and C_26:0 fatty acids synthesized in the cell. It is further demonstrated that intermolecular condensation of two fatty acids which become α- and β-units of mycolic acids will occur independently of the carbon chain length or kinds of polar moieties of fatty acids.     

Novel Glycerolipids and Glycolipids from the Surface Lipids of Nicotiana benthamiana

The surface lipids of Nicotiana benthamiana contained novel glycerolipids and several varieties of glycolipids. As glycerolipids, the triacylglycerol, 1,3-diacylglycerol, and 1,2-diacylglycerol types of glycerolipids were isolated and identified. Each lipid contained acetyl, 16–methylheptadecanoyl, and 18–methylnonadecanoyl moieties. The acetylated position of each lipid was determined by 2D-NMR, using the HMBC technique. The structures were 1,3-di-O-acetyl-2-O-acylglycerol, 1-O-acetyl-3-O-acylglycerol, and 1-O-acetyl-2-O-acylglycerol. As glycolipids, one glucose ester and four types of sucrose esters were isolated and identified. These glycolipids contained acetic acid and such branched short-chain fatty acids as 5-methylhexanoic, 4-methylhexanoic, 6-methylheptanoic, and 5-methylheptanoic acids. The structure of the glucose ester was 3,4-di-O-acyl-α-D-glucopyranose. The structures of the sucrose esters were 6-O-acetyl-4-O-acyl-α-D-glucopyranosyl-(3-O-acyl)-β-D-fructofuranoside, 4-O-acyl-α-D-glucopyranosyl-(3-O-acyl)-β-D-fructofuranoside, 3,4-di-O-acyl-α-D-glucopyranosyl-β-D-fructofuranoside, and 6-O-acetyl-α-D-glucopyranosyl-β-D-fructofuranoside.     

A 13C spin-lattice relaxation study of dipeptides containing glycine and proline: mobility of the cyclic proline side chain.

Molecular dynamics of the cyclic dipeptides cyclo(Gly-L -Pro), cyclo-(L-Pro-L -Pro), and cyclo(L-Pro-D-Pro) and the linear dipeptides L-Pro-Gly and cis and trans Gly-L -Pro were studied in neutral aqueous solution by ¹³C nuclear magnetic resonance. Spinlattice relaxation times (T₁) were determined for each individual carbon atom. The correlation times, τ, were derived from a semiquantitative analysis of the T₁ data. The correlation times of the proline ring carbons, β, γ, and δ suggest that the cyclic dipeptides have more restriction of conformational freedom in the proline ring than the linear dipeptides. This effect is most pronounced on the γ carbon.     

Triterpene hexahydroxydiphenoyl esters and a quinic acid purpurogallin carbonyl ester from the leaves of Castanopsis fissa

Triterpene hexahydroxydiphenoyl (HHDP) esters have only been isolated from Castanopsis species, and the distribution of these esters in nature is of chemotaxonomical interest. In this study, the chemical constituents of the leaves of Castanopsis fissa were examined in detail to identify and isolate potential HHDP esters. Together with 53 known compounds, 3,4-di-O-galloyl-1-O-purpurogallin carbonyl quinic acid (1) and 3,24-(S)-HHDP-2α,3β,23,24-tetrahydroxytaraxastan-28,20β-olide (2) were isolated and their structures were elucidated by spectroscopic and chemical methods. The polyphenols of the leaves were mainly composed of galloyl quinic acids, triterpenes HHDP esters, ellagitannins and flavonol glycosides. In particular, the isolation yields of 1,3,4-trigalloyl quinic acid and compound 2 were 1.53% and 0.27%, respectively, from the fresh leaves. The presence of lipid soluble HHDP esters of oleanane-type triterpenes as one of the major metabolites is an important chemotaxonomical discovery. Lipase inhibition activities and ORAC values of the major constituents were compared. The triterpene HHDP ester showed moderate lipase inhibition activity and myricitrin gave the largest ORAC value.     

Phosphoramidite Chemistry for the Synthesis of cycloSal-Pro-Nucleotides

Abstract

An alternative synthesis of 3-methyl-cycloSal-nucleotides 3–6 using phosphor-amidite chemistry is described. This protocol clearly shows advantages for the cycloSal-introduction into cytosine containing nucleoside analogues.     

PIGMENTS,FATTY ACIDS,AND STEROLS OF THE TOXIC DINOFLAGELLATE GYMNODINIUM CATENATUM1

Cultures and field samples of the toxic dinoflagellate Gymnodinium catenatum Graham from Tasmania, Australia, were analyzed for pigment, fatty acid, and sterol composition. Gymnodinium catenatum contained the characteristic pigments of photosynthetic dinoflagellates, including chlorophyll a, chlorophyll c₂, and the carotenoids peridinin, dinoxanthin, diadinoxanthin, diatoxanthin, and β,β-carotene. In midlogarithmic and early stationary phase cultures, the chlorophyll a content ranged 50–72 pg · cell^?1, total lipids 956–2084 pg · cell^?1, total fatty acids 426–804 pg · cell^?1, and total sterols 8–20 pg · cell^?1. The major fatty acids (in order of decreasing abundance) were 16:0, 22:6(n-3), and 20:5(n-3) (collectively 65–70% of the total fatty acids), followed by 16:1(n-7), 18:2(n-6), and 14:0. This distribution is characteristic of most dinoflagellates, except for the low abundance (<3%) of the fatty acid 18:5(n-3), considered by some authors to be a marker for dinoflagellates. The three major sterols were 4α-methyl-5α-cholest-7-en-3β-ol, 4α,23,24-trimethyl-5α-cholest-22E-en-3β-ol (the dinoflagellate sterol, dinosterol), and 4α,23,24-trimethyl-5α-cholest-7-en-3β-ol. These three sterols comprised about 75% of the total sterols in both logarithmic and early stationary phase cultures, and they were also found in high proportions (22–25%) in natural dinoflagellate bloom samples. 4-Desmethyl sterols, which are common in most microalgae, were only present in trace amounts in G. catenatum. The chemotaxonomic affinities of G. catenatum and the potential for using specific signature lipids for monitoring toxic dinoflagellate blooms are discussed.