Serum alpha- and gamma-tocopherols, retinol, retinyl palmitate, and carotenoid concentrations in captive and free-ranging bottlenose dolphins (Tursiops truncatus)

Concentrations of retinol, retinyl palmitate, beta-carotene, alpha-carotene, cryptoxanthin, lutein, lycopene, alpha-tocopherol, and gamma-tocopherol were measured in blood samples collected from 15 captive and 55 free-ranging bottlenose dolphins (Tursiops truncatus). From June 1991 to June 1994, blood samples were collected from captive animals residing at two locations; at Seven Seas (Brookfield Zoo, Brookfield, IL) and Hawk's Cay (Marathon Key, FL). Blood samples were collected from free-ranging animals from June 1991 to June 1996. Retinol levels were not significantly different between captive dolphin groups. However, Seven Seas animals had higher (P < 0.01) serum retinol concentrations compared to free-ranging animals (0.061 vs 0.041 microgram/ml). Retinyl palmitate was not detected in the serum of captive or free-ranging dolphins. Alpha-tocopherol levels were significantly (P < 0.05) higher for Seven Seas dolphins (16.4 micrograms/ml) than for Hawk's Cay (13.0 micrograms/ml) and free-ranging dolphins (12.5 micrograms/ml). Gamma-tocopherol concentrations were similar among captive and free-ranging dolphins. Free-ranging dolphins showed levels of circulating carotenoids (lutein and beta-carotene) while the captive animals did not. Additional carotenoids (lycopene, alpha-carotene and cryptoxanthin) were analyzed but not detected in any samples. Serum vitamin differences between captive and free-ranging dolphins may reflect the natural diet or indicate some potential biological or nutritional status significance.     

Influence of dietary vitamins A and E on serum alpha- and gamma-tocopherols, retinol, retinyl palmitate and carotenoid concentrations in Humboldt penguins (Spheniscus humboldti)

Serum retinol, retinyl palmitate, beta-carotene, cryptoxanthin, lutein, alpha-tocopherol and gamma-tocopherol were measured in 18 captive Humboldt penguins (Spheniscus humboldti) prior to and following the removal of Columbia River (CR) smelt (Thaleichthys pacificus) from the diet. Dietary vitamin A was reduced from 59.8 to 13.5 IU g-1 (dry matter basis) when CR smelt was removed from the diet. Minimal changes were noted in dietary vitamin E. Serum samples Without-CR smelt had significantly lower circulating retinol (1.19 +/- 0.09 vs. 1.94 +/- 0.08 micrograms ml-1) and retinyl palmitate (0.033 +/- 0.012 vs. 0.105 +/- 0.004 microgram ml-1) compared to samples With-CR. The Without-CR smelt diet resulted in increased serum alpha-tocopherol from 26.4 +/- 0.94 to 39.1 +/- 3.72 micrograms ml-1. More serum samples taken Without-CR smelt had detectable levels of gamma-tocopherol than those With-CR smelt. Serum lutein was higher for the samples taken Without versus With-CR smelt. Serum cryptoxanthin did not differ. beta-Carotene was not detected. Data indicate that high levels of dietary vitamin A can affect circulating levels of retinol, retinyl palmitate and vitamin E. Thus, dietary vitamin A and the interrelationship between vitamins A and E should be considered when assessing captive penguins.     

Plasma tocopherol,retinol, and carotenoid concentrations in free-ranging Humboldt penguins (Spheniscus humboldti) in Chile

Plasma retinol and α-tocopherol concentrations were measured in heparinized blood samples collected from 51 free-ranging adult Humboldt penguins (Sphenicus humboldti) residing at two colonies off the Chilean coast. Thirty samples were collected in April 1992 from penguins inhabiting the Ex-islote de los Pájaros Niños in Algarrobo, Chile. In September 1992, 21 samples were collected from birds inhabiting Isla de Cachagua, Chile. Samples were assayed for retinol, retinyl palmitate, α-tocopherol, γ-tocopherol, lutein, β-cryptoxanthin, lycopene, α-carotene, and β-carotene. Retinol, α-tocopherol, and lutein were detected in all samples, while lycopene and γ-tocopherol were not detected in any. A significantly higher percentage of samples had detectable levels of retinyl palmitate and α-carotene in April (P < 0.001): for β-cryptoxanthin the percentage was higher in September (P < 0.001). Plasma concentrations of α-tocopherol and lutein were higher in September. Alpha-tocopherol concentrations were 1,877.1 ± 99.0 (SEM) μg/dl in April compared to 2.289 ± 122.3 μg/dl in September (P < 0.05); lutein concentrations were 4.16 ± 0.43 μg/dl in April vs. 10.68 ± 1.02 μg/dl in September (P < 0.001). Retinol concentrations were not significantly different (117 ± 8.0 μg/dl in April vs. 105.3 ± 7.6 μg/dl in September). Both physiologic changes associated with season, and the change in locale may have contributed to the differences seen in the assay means and the number of samples with detectable levels. © 1996 Wiley-Liss, Inc.     

Mitochondrial and nuclear DNA analyses reveal fine scale geographic structure in bottlenose dolphins (<Emphasis Type="Italic">Tursiops truncatus</Emphasis>) in the Gulf of Mexico

There is a need for biological information to support current stock designations of bottlenose dolphins (Tursiops truncatus) in the Gulf of Mexico. The existence of many inshore, resident “communities” raises questions as to the relationship these dolphins may hold with dolphins inhabiting neighboring inshore and coastal areas. In this study, population subdivision was examined among four resident, inshore bottlenose dolphin stocks (Sarasota Bay, FL, Tampa Bay, FL, Charlotte Harbor, FL and Matagorda Bay, TX) and one coastal stock (1–12 km offshore) in the Gulf of Mexico. Evidence of significant population structure among all areas was found on the basis of both mitochondrial DNA (mtDNA) control region sequence data and nine nuclear microsatellite loci. Estimates of relatedness showed no population contained a significantly high number of related individuals, while separate AMOVAs for males and females indicated that both sexes exhibit a significant level of site philopatry. Results presented here provide the first genetic evidence of population subdivision between the coastal Gulf of Mexico and adjacent inshore areas along the central west coast of Florida. Such strong genetic subdivision is surprising given the short geographical distance between many of these areas and the lack of obvious geographic barriers to prevent gene flow. These findings support the current, separate identification of stocks for bottlenose dolphins inhabiting the eastern coastal and inshore areas of the Gulf of Mexico.     

Mycobiota associated with larval mines of Thrypticus truncatus and T. sagittatus (Diptera, Dolichopodidae) on waterhyacinth, Eichhornia crassipes, in Argentina

Thrypticus truncatus is a candidate agent for biocontrol of waterhyacinth; the larvae of this diptera mine in the petioles and feed on the phloem in the vascular bundles. The mycobiota associated with T. truncatus and T. sagittatus mines was investigated during two surveys undertaken in the spring and autumn in the Delta of the Paraná River, Argentina. Isolations were made from the mines and larval feeding points, as well as from the larvae, following dissection of the petioles, and plated onto agar. Young and upper parts of the petioles without Thrypticus mines were used as controls. Twenty eight fungal species were isolated from the mines. Pestalotiopsis guepinii, Mucor attenuatus, Phoma tropica, Achlya americana, Fusarium avenaceum, Cladosporium cladosporioides, Clonostachys rosea, Epicoccum purpurascens, Plectosporium tabacinum, Alternaria alternata, and Acremonium sp. were the most common fungi associated with mines and feeding points. Cladosporium cladosporioides, Cytospora sp., Mucor attenuatus, and Phoma tropica were associated with the larval body. The list of fungi in mines was compared with bibliographic information to determine if the species are known pathogens on waterhyacinth or other plant species. This is the first study on mycobiota associated with T. truncatus and T. sagittatus mines in waterhyacinth petioles in Argentina.     

Anti-inflammatory properties of α- and γ-tocopherol

Natural vitamin E consists of four different tocopherol and four different tocotrienol homologues (α, β, γ, δ) that all have antioxidant activity. However, recent data indicate that the different vitamin E homologues also have biological activity unrelated to their antioxidant activity. In this review, we discuss the anti-inflammatory properties of the two major forms of vitamin E, α-tocopherol (αT) and γ-tocopherol (γT), and discuss the potential molecular mechanisms involved in these effects. While both tocopherols exhibit anti-inflammatory activity in vitro and in vivo, supplementation with mixed (γT-enriched) tocopherols seems to be more potent than supplementation with αT alone. This may explain the mostly negative outcomes of the recent large-scale interventional chronic disease prevention trials with αT only and thus warrants further investigation.     

Preparation, characterization and antifungal properties of 2-(α-arylamino phosphonate)-chitosan

In this paper, 20 kinds of different 2-(α-arylamino phosphonate)-chitosan (2-α-AAPCS) were prepared by different Schiff bases of chitosan (CS) reacted with di-alkyl phosphite in benzene solution. The structures of the derivatives (2-α-AAPCS) were characterized by FT-IR spectroscopy and elemental analysis. In addition, the antifungal activities of the derivatives against four kinds of fungi were evaluated in the experiment. The results indicated that all the prepared 2-α-AAPCS had a significant inhibiting effect on the investigated fungi when the derivatives concentration ranged from 50 to 500 μg mL⁻¹. Furthermore, the antifungal activities of the derivatives increased with increasing the molecular weight and concentration. And the antifungal activities of the derivatives were affected by their dimensional effect and charge density. Besides, the rule and mechanism of the antifungal activities of them were discussed in this paper.     

Vitamin A and E tissue distribution with comparisons to organochlorine concentrations in the serum, blubber and liver of the bowhead whale (Balaena mysticetus)

Vitamin A and E concentrations were determined in liver (n = 51), blubber (n = 23) and serum (n = 53) of subsistence-hunted bowhead whales (Balaena mysticetus), between 1998 and 2001. Retinol and alpha-tocopherol were the major forms of vitamins A and E detected, respectively. Liver contained the highest mean concentrations of vitamin A, followed by epidermis, blubber, and serum. Liver also contained the highest mean concentration of vitamin E, followed by serum, epidermis, and blubber. Stratification of retinol and tocopherol was examined throughout the blubber cores collected. Retinol concentrations were significantly higher in the epidermis than in the deeper blubber layers. Tocopherol concentrations were similar for epidermis and the intermediate layer of blubber. Both the epidermis and the intermediate layer of blubber had significantly higher tocopherol concentrations than the innermost and outermost blubber layers. Vitamin A and E concentrations were investigated with respect to gender and reproductive status of females (males, non-pregnant females, pregnant females), age groups and season of harvest. Certain persistent organic contaminants are known to have a negative effect on retinol concentration in serum of pinnipeds and cetaceans. Bowhead whales have relatively low concentrations of organochlorines (OCs) in comparison to other mysticete species. The relationships between serum, liver and blubber retinol and serum and blubber OC concentrations were examined with no significant correlations noted.     

Immunoreactivity of subunits of the (Na + K)-ATPase Cross-reactivity of the α,α+ and β forms in different organs and species

The immunologic cross-reactivity of the α and α+ forms of the large subunit and the β subunit of the (Na⁺ + K⁺)-ATPase from brain and kidney preparations was examined using rabbit antiserum prepared against the purified holo lamb kidney enzyme. As previously reported by Sweadner ((1979) J. Biol. Chem. 254, 6060–6067) phosphorylation of the large subunit of the (Na⁺ + K⁺)-ATPase in the presence of Na⁺, Mg²⁺, and [γ-³²P]ATP revealed that dog and, very likely, rat brain contain two forms of the large subunit (designated α and α+) while dog, rat, and lamb kidney contain only one form (α). The cross-reactivity of the α and α+ forms in these preparations was investigated by resolving the subunits by SDS-polyacrylamide gel electrophoresis. The separated polypeptides were transferred to unmodified nitrocellulose paper, and reacted with rabbit anti-lamb kidney serum, followed by detection of the antigen-antibody complex with ¹²⁵I-labeled protein A and autoradiography. By this method, the α and α+ forms of rat and dog brain, as well as the α form found in kidney, were shown to cross-react. In addition, membranes from human cerebral cortex were shown to contain two immunoreactive bands corresponding to the α and α+ forms of dog brain. In contrast, the brain of the insect Manduca sexta contains only one immunoreactive polypeptide with a molecular weight intermediate to the α and α+ forms of dog brain. The β subunit from lamb, dog and rat kidney and from dog and rat brain cross-reacts with anti-lamb kidney (Na⁺ + K⁺)-ATPase serum. The mobility of the β subunit from dog and rat brain on SDS-polyacrylamide electrophoresis gels is greater than the mobility of the β subunit from lamb, rat or dog kidney.     

High-Yield Expression and Purification of Human Interferon α-1 in Pichia pastoris

For several years, interferon α-1, also known as interferon α-D, has been studied for treatment of various viral diseases, such as hepatic fibrosis caused by hepatitis B, herpes simplex virus keratitis, and bovine respiratory diseases in calves. Currently, recombinant human interferon α-D (rHuIFNαD) is expressed intracellularly in Escherichia coli or secreted by Bacillus subtilis and Saccharomyces cerevisiae. In this report, we describe the process of obtaining a relatively high-yield secretion of biologically active recombinant rHuIFNαD using the Pichia pastoris system. The process produced as high as 0.7 mg of purified protein per 20 ml of shake culture of rHuIFNαD with better bioactivity than the commercially available rHuIFNαD molecule produced in E. coli.     

The sequence and phylogenesis of the α-globin genes of Barbary sheep (Ammotragus lervia), goat (Capra hircus), European mouflon (Ovis aries musimon) and Cyprus mouflon (Ovis aries ophion)

In order to investigate the polymorphism of α-globin chain of hemoglobin amongst caprines, the linked ^Iα and ^IIα globin genes of Barbary sheep (Ammotragus lervia), goat (Capra hircus), European mouflon (Ovis aries musimon), and Cyprus mouflon (Ovis aries ophion) were completely sequenced, including the 5′ and 3′ untranslated regions. European and Cyprus mouflons, which do not show polymorphic α globin chains, had almost identical α globin genes, whereas Barbary sheep exhibit two different chains encoded by two nonallelic genes. Four different α genes were observed and sequenced in goat, validating previous observations of the existence of allelic and nonallelic polymorphism. As in other vertebrates, interchromosomal gene conversion appears to be responsible for such polymorphism. Evaluation of nucleotide sequences at the level of molecular evolution of the ^Iα-globin gene family in the caprine taxa suggests a closer relationship between the genus Ammotragus and Capra. Molecular clock estimates suggest sheep-mouflon, goat-aoudad, and ancestor-caprine divergences of 2.8, 5.7, and 7.1 MYBP, respectively.     

Genome Analysis inNeurospora crassa;Cloning of Four Lociarginine-1(arg-1),methionine-6(met-6),unknown-7(un-7), andribosome production-1(rip-1) and Associated Chromosome Walking

We have cloned fourNeurospora crassagenes by complementation analysis. Cloned genes include thearginine-1(arg-1),methionine-6(met-6),unknown-7(un-7), andribosome production-1(rip-1) loci. Chromosome walks were initiated in ordered cosmid libraries from the cloned loci. A total of about 700 kb of theNeurosporagenome is covered in these walks.     

Conversion of α-methyltropate to optically active α-phenylpropionate by tropate-degrading Rhodococcus sp. KU1314

Microorganisms which can assimilate tropate were screened from soil. Among them, we found a microorganism which has an ability to convert α-methyltropate to optically active α-phenylpropionate, and it was identified as Rhodococcus sp. KU1314. Substrate specificity of the microorganism has been studied. When the aryl group was phenyl, 4-methoxyphenyl and 2-naphthyl, the substrate gave optically active α-propionate in good yields. To estimate the reaction mechanism, some compounds considered to be the intermediates were subjected to the reaction. Both enantiomers of α-methyltropate were converted to (R)-α-phenylpropionate with almost the same enantiomeric excess (68 and 72% from R-and S-enantiomers, respectively) and yield (605 and 48% from R-and S-enantiomers, respectively).     

Cell wall α-1,3-glucans from a biocontrol isolate of Rhizoctonia: immunocytolocalization and relationship with α-glucanase activity from potato sprouts

The interface between plants and pathogens plays an important role in their interaction. Studies of fungal cell walls are scarce and previous results show the existence of α-1,3-glucans in addition to ß-glucans. In addition, α-1,3-glucans are not present in plant cell walls, and α-glucanase activity in plants has not been described before. In a previous work, we purified and characterized an α-1,3-glucan from a binucleated, non-pathogenic Rhizoctonia isolate, which induces plant defence responses. Therefore, in order to study the architecture of the fungal cell wall, and the accessibility and localization of the α-glucan elicitor, we prepared an antibody against the α-1,3-glucan and analysed its localization by TEM. Immunolocalization showed the presence of the α-1,3-glucan in the intercellular spaces and along the cell walls, mainly on the inner layers. This result, and the presence of the α-1,3-glucan in the liquid culture medium in which binucleated non-pathogenic Rhizoctonia was grown, confirmed that the α-glucan had been secreted. The α-1,3-glucan was also immunocytolocalized on potato sprouts tissue elicited with the glucan; gold particles were observed in vacuoles and close to the plasmalemma. In addition, α-glucanase activity in potato sprouts was detected using cell wall glucans from the pathogenic isolate R. solani AG-3 as substrates; whereas, when cell wall glucans from non-pathogenic isolates were used, no α-glucanase activity was detected. Our results suggest that the presence of α-1,3-glucans could be associated with the formation and integrity of the cell wall and also with plant–fungi interactions. This is the first report to describe α-glucanolytic activity in plants.     

Purification, biochemical characterisation and partial primary structure of a new α-amylase inhibitor from Secale cereale (rye)

Plant α-amylase inhibitors show great potential as tools to engineer resistance of crop plants against pests. Their possible use is, however, complicated by the observed variations in specificity of enzyme inhibition, even within closely related families of inhibitors. Better understanding of this specificity depends on modelling studies based on ample structural and biochemical information. A new member of the α-amylase inhibitor family of cereal endosperm has been purified from rye using two ionic exchange chromatography steps. It has been characterised by mass spectrometry, inhibition assays and N-terminal protein sequencing. The results show that the inhibitor has a monomer molecular mass of 13 756 Da, is capable of dimerisation and is probably glycosylated. The inhibitor has high homology with the bifunctional α-amylase/trypsin inhibitors from barley and wheat, but much poorer homology with other known inhibitors from rye. Despite the homology with bifunctional inhibitors, this inhibitor does not show activity against mammalian or insect trypsin, although activity against porcine pancreatic, human salivary, Acanthoscelides obtectus and Zabrotes subfasciatus α-amylases was observed. The inhibitor is more effective against insect α-amylases than against mammalian enzymes. It is concluded that rye contains a homologue of the bifunctional α-amylase/trypsin inhibitor family without activity against trypsins. The necessity of exercising caution in assigning function based on sequence comparison is emphasised.     

Properties of γ-aminobutyraldehyde dehydrogenase from Escherichia coli

γ-Aminobutyraldehyde dehydrogenase from Escherichia coli K-12 has been purified and characterized from cell mutants able to grow in putrescine as the sole carbon and nitrogen source. The enzyme has an M_r of 195 000±10 000 in its dimeric form with an M_r of 95 000±1000 for each subunit, a pH optimum at 5.4 in sodium citrate buffer, and does not require bivalent cations for its activity. K_m values are 31.3±6.8 μM and 53.8±7.4 μM for Δ-1-pyrroline and NAD⁺, respectively. An inhibitory capacity for NADH is also shown using the purified enzyme.     

Microbial hydroxylation of 16α,17α-dimethyl-17β-(l-oxopropyl)androsta-l,4-dien-3-one to rimexolone by Curvularia lunata AS 3.4381

A novel synthetic route to rimexolone, a corticosteroid for treatment of ocular inflammation without significant elevation of intraocular pressure, was described. An investigation has been undertaken of the microbial transformation of 16α,17α-dimethyl-17β-(l-oxopropyl)androsta-l,4-dien-3-one by microorganisms known to hydroxylate conventional steroids, using Curvularia lunata AS 3.4381 gave rimexolone, the product of 11β-hydroxylation, respectively. The target compound was characterized with reference substance rimexolone by TLC, HPLC, elemental analysis, MS, IR, and NMR.     

Antibacterial activity of α-mangostin against vancomycin resistant Enterococci (VRE) and synergism with antibiotics

alpha-Mangostin, isolated from the stem bark of Garcinia mangostana L., was found to be active against vancomycin resistant Enterococci (VRE) and methicillin resistant Staphylococcus aureus (MRSA), with MIC values of 6.25 and 6.25 to 12.5 microg/ml, respectively. Our studies showed synergism between alpha-mangostin and gentamicin (GM) against VRE, and alpha-mangostin and vancomycin hydrochloride (VCM) against MRSA. Further studies showed partial synergism between alpha-mangostin and commercially available antibiotics such as ampicillin and minocycline. These findings suggested that alpha-mangostin alone or in combination with GM against VRE and in combination with VCM against MRSA might be useful in controlling VRE and MRSA infections.