Natural Polyesters: Dufour's Gland Macrocyclic Lactones Form Brood Cell Laminesters in Colletes Bees.

Bees in the genus Colletes make their brood cells in the ground and coat them with a highly resistant, waterproof, transparent membrane. This membrane is a polyester constructed mainly from 18-hydroxyoctadecanoic acid and 20-hydroxy-eicosanoic acid, which are stored as their corresponding lactones in the Dufour's gland of the bee. When lining the cells, the bee secretes its glandular content, and the membrane is apparently a product of polycondensation reaction of its contents. This appears to be the first report of a naturally occurring linear polyester. The term laminester (lamina approximately layer + ester) for this class of compounds is proposed.

Defense in thrips: forbidding fruitiness of a lactone.

Expulsion of anal fluid from the upturned abdomen was demonstrated to serve a defensive function in the thrips Bagnalliella yuccae. An allomone in the anal exudate was identified as gamma-decalactone, a fruity-smelling compound that repelled potential predators. Chemical defenses may contribute to the ability of thrips to maintain large aggregations.

Alkaloid from fire ant venom: identification and synthesis.

An alkaloid, trans-2-methyl-6-n-undecylpiperidine (solenopsin A), has been isolated from the venom of the fire ant Solenopsis saevissima. The structure has been confirmed by an unambiguous synthesis.

Anthophora Bees: Unusual Glycerides from Maternal Dufour's Glands Serve as Larval Food and Cell Lining.

The Dufour's gland of Anthophora abrupta, a solitary bee, secretes a complex mixture of liquid triglycerides containing one long-chain and two shortchain fatty acids. This is applied inside the earthen brood cells and added to the provision, where it is converted, perhaps by enzymes from the bee's saliva or gut, to solid diglycerides that are later eaten by the bee larvae. This use of Dufour's gland secretion as food and its nutritive function are reminiscent of the royal jelly secreted by honey bees.

Juvenile hormone: identification of an active compound from balsam fir.

A sesquiterpenoid ester with high juvenile hormone activity for Pyrrhocoris apterus (L.) was isolated from balsam fir, Abies balsamea (L.) Miller, and identified as the methyl ester of todomatuic acid.

Proteins in a nonvenomous defensive secretion: biosynthetic significance.

In common with many arthropods, the true bug, Leptoglossus phyllopus, when disturbed, emits a two-phase secretion that consists of an organic phase and an aqueous phase. The organic phase is a mixture of highly reactive low-molecular-weight compounds, analogous to those produced by other arthropods, and is deterrent to many kinds of predators. The aqueous phase, heretofore ignored in most analyses of arthropod defensive secretions, contains proteins. Even though the secretion is not injected, the proteins enzymatically catalyze the derivation of the most reactive components within the impermeable cuticular storage reservoir and, thus, constitute part of the defensive system that appears to be commonly used by arthropods producing irritating chemicals.

Caste-specific compounds in male carpenter ants.

Three caste-specific substances new to arthropod glandular secretions occur in the mandibular glands of male ants of five species in the genus Camponotus. These volatile compounds, which are not found in alate females or workers, have been identified as methyl 6-methyl salicylate, 2,4-dimethyl-2-hexenoic acid, and methyl anthranilate. The free acid has not been described previously.

Sex pheromones in snakes.

The majority of pheromones identified to date are insect pheromones, which are volatile in nature. Identification of nonvolatile pheromones have been relatively rare, especially in vertebrates. Male and female garter snakes use pheromones to mediate sexual behavior. The female sex attractiveness pheromone of the Canadian red-sided garter snake, Thamnophis sirtalis parietalis, consists of a novel series of nonvolatile saturated and monounsaturated long-chain methyl ketones, whereas the male sex recognition pheromone contains squalene. These compounds were isolated, identified, and partially synthesized, and field tests show them to be biologically active.

Identification of a neuropeptide hormone that regulates sex pheromone production in female moths.

A pheromone biosynthesis activating neuropeptide (PBAN) hormone that controls sex pheromone production in female moths was identified from the brain-subesophageal ganglion complexes of the adult corn earworm, Heliothis zea. PBAN has 33 amino acid residues and a molecular weight of 3900. Its amino acid sequence has no significant homology with any of the fully characterized peptide hormones. The synthetic peptide, at a dose of between 2 and 4 picomoles, induced production of a normal quantity of sex pheromone in ligated H. zea females. The peptide also induced pheromone production in six other species of moths, thus indicating that this or similar peptides may be responsible for the regulation of pheromone production in moths.

Human alveolar macrophage-derived chemotactic factor for neutrophils. Stimuli and partial characterization.

The presence of neutrophils within the lung is a characteristic feature of a variety of lung diseases. To evaluate the potential role of alveolar macrophages in modulating the migration of neutrophils to the lung, normal human alveolar macrophages obtained from volunteers by bronchopulmonary lavage, were exposed for various periods of time in vitro to heat-killed microorganisms, and noninfectious particulates, immune complexes, and the macrophage supernates were evaluated for chemotactic activity. The microorganisms, noninfectious particulates, and immune complexes were chosen as stimuli for alveolar macrophages because these stimuli are representative of a spectrum of pathogenic agents that cause neutrophil accumulation in the lower respiratory tract. After incubation with each of these stimuli, alveolar macrophages released low molecular weight (400-600) chemotactic factor(s) (alveolar macrophage-derived chemotactic factor[s] [AMCF]) with relatively more activity for neutrophils than monocytes or eosinophils. Checker-board analysis of the AMCF revealed that the factor was primarily chemotactic and not chemokinetic for neutrophils. The selectivity for neutrophils vs. monocytes could not be explained by a selective deactivation of monocytes, because the AMCF was more potent in deactivating neutrophils than monocytes. Partial characterization of AMCF demonstrated it was heterogeneous with the following features: (a) stable to heating at 56 and 100 degrees C for 30 min; (b) stable over a pH range of 1.0 to 12.0 for 60 min; (c) stable after exposure to trypsin, papain, chymotrypsin, collagenase, and elastase; (d) partially inhibited by serum chemotactic factor inhibitor(s); (e) two major isoelectric points (pI 7.6 and 5.2); and (f) partially extractable into ethyl acetate, ether, and hexane. Although AMCF was, at least, partially lipid in nature, it did not appear to be similar to previously described lipid chemotactic factors (e.g., hydroxy-derivatives of 5,8,10,14-eicosatetraenoic acid); analysis by gas chromatography-mass spectrophotometry of AMCF extracted into ethyl acetate did not reveal the presence of 5,8,10,14-eicosatetraenoic acid. The macrophage supernates containing the AMCF also stimulated normal human neutrophils to release lysozyme and lactoferrin but not lactate dehydrogenase. These studies suggest that a wide variety of potentially pathogenic stimuli induce normal alveolar macrophages to generate a low molecular weight chemotactic factor(s) that preferentially attracts neutrophils. Because alveolar macrophages are normal residents of alveoli, it is likely that by releasing this factor(s) macrophages play a significant role in amplifying the inflammatory processes seen in many acute and chronic lung diseases.

Refsum's disease: characterization of the enzyme defect in cell culture.

Refsum's disease (heredopathia atactica polyneuritiformis, HAP) is an inherited neurological disorder associated with storage of the branched-chain fatty acid, phytanic acid (3,7,11,15-tetramethylhexadecanoic acid). Cultured fibroblasts derived from skin biopsies of HAP patients did not contain elevated levels of phytanate, yet showed rates of phytanate-C-(14)C oxidation less than 3% of those seen in cells from control subjects. Cells of control subjects converted phytanate to alpha-hydroxyphytanate, to pristanate (the [n-1] homologue of phytanate) and to 4,8,12-trimethyltridecanoate, compounds previously identified as intermediates on the major pathway for phytanate metabolism in animals, providing the first direct evidence that this same oxidative pathway is operative in human cells. None of these breakdown products could be found after incubation of phytanate with HAP cells. Labeled alpha-hydroxyphytanate and labeled pristanate were oxidized at normal rates by HAP cells. Oxidation of the latter proceeded at normal rates both when added to the medium at very low tracer levels and at levels 100 times greater. Phytanate was incorporated into and released from lipid esters at normal rates by HAP cells. Elevated levels of free phytanate in the medium were no more toxic to HAP cells than to control cells over the 48- to 72-hr exposures involved in these studies, as evidenced by morphologic criteria and by ability to oxidize labeled palmitate. These findings are consistent with the hypothesis that the cells from HAP patients are deficient in a single enzyme involved in the alpha-hydroxylation of phytanate, while the enzymes involved in later steps are present at normal or near-normal levels.

Studies on the metabolic error in Refsum's disease.

Studies utilizing mevalonic acid-2-(14)C and D(2)O as precursors failed to provide evidence for an appreciable rate of endogenous biosynthesis of phytanic acid in a patient with Refsum's disease. Orally administered tracer doses of phytol-U-(14)C were well absorbed both by seven normal control subjects (61 to 94%) and by two patients with Refsum's disease (74 and 80%). The fraction of the absorbed dose converted to (14)CO(2) in 12 hours was 3.5 and 5.8% in Refsum's disease patients and averaged 20.9% in seven control subjects. Labeled phytanic acid was demonstrated in the plasma of both control subjects and patients given phytol-U-(14)C, establishing phytol in the diet as a potential precursor of phytanic acid. This labeled phytanic acid had disappeared almost completely from the plasma of the seven control subjects by 24 to 48 hours, whereas it persisted at high concentrations in the plasma of the two patients for many days. We conclude that the phytanic acid accumulating in Refsum's disease is primarily of exogenous origin and that patients with Refsum's disease have a relative block in the degradation of phytanic acid and possibly other similar branched-chain compounds. This may relate to a deficiency in mechanisms for release of phytanic acid from stored ester forms or, more probably, to reactions essential to oxidative degradation of the carbon skeleton.

ω-Phonetoxin-IIA: a calcium channel blocker from the spider Phoneutria nigriventer.

A peptide with neurotoxic effect on mammals, purified from the venom of the spider Phoneutria nigriventer, was studied regarding its primary structure and its effects on voltage-gated calcium channels. The peptide, named ω-phonetoxin-IIA, has 76 amino acids residues, with 14 Cys forming 7 disulphide bonds, and a molecular weight of 8362.7 Da. The neurotoxicity is a consequence of the peptide's blocking effects on high-voltage-activated (HVA) calcium channels. N-type HVA calcium channels of rat dorsal root ganglion neurons are blocked with affinity in the sub-nanomolar concentration range. The toxin also blocks L-type channels of rat ß pancreatic cells, with an affinity 40 times lower. Although not studied in detail, evidence indicates that the toxin also blocks other types of HVA calcium channels, such as P and Q. No effect was observed on low-voltage-activated, T-type calcium channels. The significant homologies between ω-phonetoxin-IIA and the peptides of the ω-agatoxin-III family, and the overlapping inhibitory effects on calcium channels are discussed in terms of the structure-activity relationship.

Electrospray mass spectrum of a per(onio)-substituted benzene: retention of coulombic charge upon collisionally activated decomposition.

The hexakis (4-diemthylaminopyridyl) benzene hexacation of 1 is investigated as an extreme example of the ability of electrospray ionization to allow transfer of small multivalent ions to the gas phase. The hexacationized benzene ring ions are stabilized by forming gas phase complexes with two to five trifluoromethanesulfonate counterions. MS/MS analysis reveals that their fragmentation takes place by loss of neutrals such as trifluoromethanesulfonic acid and 4-dimethylaminopyridine; no rupture of the benzene or pyridine rings was observed in spite of accumulation of positive charge in a restricted geometry.

Iron carboxylate oxygen-centered-triangle complexes detected during electrospray use of organic acid modifiers with a comment on the finnigan TSQ-700 electrospray inlet system.

Use of infusion methods rather than high-performance liquid chromatography allowed us to confirm the observation that solutions of propionic acid-isopropanol restore sensitivity lost due to trifluoroacetic acid in electrospray mass spectra of basic substances, particularly peptides. In this work, when propionic acid-isopropanol was used, we detected an abundant ion with m/z 622 that shifted to m/z 538 when we substituted acetic acid-methanol for the propionic acid-isopropanol. Via accurate mass measurement and tandem mass spectrometry the origin of the ion was identified as the complex Fe3O(O2CR)6(L)0-3, where L is one of several ligands from solvent or water. The grounding arrangement of the Finnigan TSQ-700 electrospray source produces electrolytic currents that may accentuate the abundance of this complex and specifically produces observable gas bubbles that adversely affect the spray stability.

Structural assignment of isomeric 2-(2-quinolinyl)-1H-indene-1,3(2H)-dione mono- and disulfonic acids by liquid chromatography electrospray and atmospheric pressure chemical ionization tandem mass spectrometry.

Positionally isomeric 2-(2-quinolinyl)-1H-indene-1,3(2H)-dione mono- and disulfonic acids give rise to similar electrospray ionization (ESI) and atmosphere pressure chemical ionization (APCI) mass spectra, which show very abundant MH(+) ions and negligible fragmentation. The MH(+) ions of these isomeric acids exhibit notably different behavior under collision-induced dissociation (CID) conditions. The acids with a sulfonic group at position 8' in the quinoline moiety, adjacent to the N-atom, exhibit highly abundant [MH - H(2)SO(3)](+) ions (m/z 272 for the mono- and m/z 352 for the disulfonic acids), which are of lower abundance in the CID spectra of isomers with the SO(3)H group at other positions, remote from the nitrogen atom. The latter isomers undergo efficient eliminations of SO(3) and HSO(3). The isomeric diacids with one SO(3)H group at position 4 of the indene-1,3(2H)-dione moiety, adjacent to one of the carbonyl groups, undergo highly efficient elimination of H(2)O. Mechanistic pathways, involving interactions between adjacent groups, are proposed for the above regiospecific fragmentations. Pronounced different behavior has been also observed in negative ion tandem mass spectrometric measurements of the sulfonic acids. The distinctive behavior of the isomeric acids was strongly pronounced when the measurements were performed with an ion trap mass spectrometer (LCQ), and much less so with a triple-stage quadrupole instrument (TSQ).

Separation of alkaloids by pH-zone-refining counter-current chromatography.

pH-Zone-refining counter-current chromatography was applied to the separation of alkaloids from a crude extract of Crinum moorei using a multilayer coil planet centrifuge. After methyl tert.-butyl ether and water were equilibrated, triethylamine (5-10 mM) was added to the organic phase and hydrochloric acid (5-10 mM) to the aqueous phase. The separation was performed by eluting the column with either the organic phase (displacement mode) or the aqueous phase (reverse-displacement mode) while the other phase was used as the stationary phase. From 3 g of the extract, crinine, powelline and crinamidine were separated in 2.5-7 h with minimum overlapping.

Countercurrent chromatographic analysis of ovalbumin obtained from various sources using the cross-axis coil planet centrifuge.

The present studies have been conducted to investigate the cause of an unusually broad peak of ovalbumin obtained by countercurrent chromatography (CCC) reported earlier [K. Shinomija et al., J. Chromatogr., 644 (1993) 215]. A series of CCC experiments using our prototyte of the cross-axis coil planet centrifuge revealed that commercial ovalbumin products were classified into two groups: group A formed two peaks of ovalbumin at pH 7.0 and 5.8, while group B showed a relatively sharp single peak in a broad range of pH. Electrophoresis indicated that the group A ovalbumin consisted of both natural and denatured products: the natural ovalbumin is a monomer (Mr 45 000) whereas the denatured products form dimers (Mr 90 000). The abnormally broad peak obtained from the group A ovalbumin at pH 9 is apparently caused by the heterogeneity of the sample protein.

2-Hydroxypropyldigitonin: synthesis and properties of preparations differing in degree of substitution.

Reaction of propylene oxide with digitonin in strong aqueous alkali yielded water soluble, nontoxic solubilizers of drugs and hormones, the molecular composition of which could be fully deduced from plasma desorption mass spectrometry data. Preparations with average degrees of substitution ranging from 0.7 to 4.5 were investigated, and all the distributions of degrees of substitution observed were relatively narrow and symmetrical; thus, the reactivity of primary and secondary hydroxyls of saponin and of its conversion products had to be quite similar. The effects of the average degree of substitution of 2-hydroxypropyldigitonins on the formation of micelles and on the solubilization of four drugs into aqueous solutions were found to be minor. On the other hand, the ability to hemolyze erythrocytes decreased sharply with an increasing average degree of substitution.

Reaction of cyclodextrins with propylene oxide or with glycidol: analysis of product distribution.

Reaction of cyclomalto-hexaose, -heptaose, or -octaose with propylene oxide in strong aqueous alkali gave products in which distribution of the degrees of substitution was relatively narrow and nearly symmetrical, and increased with the average degree of substitution. When an equimolar mixture of cyclomalto-hexaose, heptaose, or -octaose was used, the average degrees of substitution of all three carbohydrates were close to each other. These findings indicate that the reactivities of the hydroxyl groups of cyclomalto-hexaose, -heptaose, or -octaose, and of all their (2-hydroxypropyl) ethers formed in the reactions, are quite similar. Reaction of cyclomaltoheptaose with glycidol also yielded a product having a narrow distribution of degree of substitution, but which was slightly skewed towards the higher degrees. Thus, as it proceeds, this etherification leads to products having higher reactivity towards the epoxide.