Search

Bird feather lipids are usually attributed to the oily secretion product of the uropygial (preen) gland. We have observed, however, that feathers exhibit a strong reaction with osmium tetroxide (OsO4), even after treatment with detergents. This leads us to postulate the existence of endogenous feather lipids distinct from preen gland lipids. In order to substantiate our hypothesis, we investigated down feathers from a 1-day-old chicken as their uropgygial gland is not functionally active. The results confirmed the osmiophilic reaction, which was concentrated in the center of barbs and strongly reduced after lipid extraction. In these lipid extracts, we identified using thin layer chromatography, cholesterol, various ceramides, glycolipids, phospholipids, and fatty acids, which closely resembled the lipid composition of the water barrier in the chicken-cornified epidermal envelope. This composition is clearly distinct from chicken uropygeal gland secretion (UGS) known to consist of fatty alcohols as part of aliphatic monoester waxes and of free, predominantly saturated, fatty acids. A filter assay showed a strong reactivity between OsO4 and the fatty acids C18:1 and C18:2 and with feather lipid extracts, but not with UGS. These observations were confirmed by gas chromatography detecting unsaturated fatty acids including C18:1 and C18:2 as well as cholesterol exclusively in chicken feathers. Our results indicate that (1) endogenous lipids are detectable in chicken feathers and distinct from UGS and (2) in analogy to the morphogenesis of the cornified envelope of chicken feather lipids that may have derived from cellular feather-precursors, apparently enduring the specific cell death during developmental feather cornification. Electronic supplementary material The online version of this article (10.1007/s00709-020-01544-7) contains supplementary material, which is available to authorized users.

Osmium tetroxide and TMEDA form stable crystalline adducts with alkenes. The structure of liquid alkenes can be determined through X-ray analysis of these derivatives. Osmium, a heavy atom, facilitates the crystallographic analysis and the determination of the absolute configuration using common Mo X-ray sources. The utility of this method for assigning structures and absolute configurations was demonstrated on a number of unsaturated substrates that include simple alkenes, enones, enol ethers, and silyl enol ethers.

Super-resolution correlative light and electron microscopy (SR-CLEM) is a powerful approach for imaging specific molecules at the nanoscale in the context of the cellular ultrastructure. Epon epoxy resin embedding offers advantages for SR-CLEM, including ultrastructural preservation and high quality sectioning. However, Epon embedding eliminates fluorescence from most fluorescent proteins. We describe a photocontrollable fluorescent protein, mEosEM, that can survive Epon embedding after osmium tetroxide (OsO4) treatment for improved SR-CLEM.

Staining compounds containing heavy elements (electron dyes) can facilitate the visualization of DNA and related biomolecules by using TEM. However, research into the synthesis and utilization of alternative electron dyes has been limited. Here, we report the synthesis of a novel DNA intercalator molecule, bis‐acridine uranyl (BAU). NMR spectroscopy and MS confirmed the validity of the synthetic strategy and gel electrophoresis verified the binding of BAU to DNA. For TEM imaging of DNA, two‐dimensional DNA origami nanostructures were used as a robust microscopy test object. By using scanning transmission electron microscopy (STEM) imaging, which is favored over conventional wide‐field TEM for improved contrast, and therefore, quantitative image analysis, it is found that the synthesized BAU intercalator can render DNA visible, even at the single‐molecule scale. For comparison, other staining compounds with a purported affinity towards DNA, such as dichloroplatinum, cisplatin, osmium tetroxide, and uranyl acetate, have been evaluated. The STEM contrast is discussed in terms of the DNA–dye association constants, number of dye molecules bound per base pair, and the electron‐scattering capacity of the metal‐containing ligands. These findings pave the way for the future development of electron dyes with specific DNA‐binding motifs for high‐resolution TEM imaging.

SUMMARY Leaves of Lolium multiflorum Lam. and Zea mays L. of different ages were prepared for TEM by a number of cytochemical methods. After post-fixation in potassium ferricyanide with osmium tetroxide the peripheral reticulum in both mesophyll and bundle sheath chloroplasts of maize stain very clearly. Post-fixation with uranyl acetate, copper and lead salts also stains the chloroplast envelope rather than the internal thylakoid system. Differences between the mesophyll and bundle sheath chloroplasts of maize and between those of ryegrass are revealed after post-fixation in zinc iodide with osmium tetroxide. After post-fixation in potassium iodide with osmium tetroxide the loculi stain less intensely than in zinc iodide. Malachite green added to glutaraldehyde at pH 7 0 enhances membrane staining in general and reveals large lipid drops in the stroma. At pH below 6 0 only the 'A' space is stained. The 'A' space and the loculus are stained when ethidium bromide is added to glutaraldehyde. The different reactions of the chloroplast membranes probably reflect differences in chemical composition.

Combined analysis of immunostaining for various biological molecules coupled with investigations of ultrastructural features of individual cells is a powerful approach for studies of cellular functions in normal and pathological conditions. However, weak antigenicity of tissues fixed by conventional methods poses a problem for immunoassays. This study introduces a method of correlative light and electron microscopy imaging of the same endocrine cells of compact and diffuse islets from human pancreatic tissue specimens. The method utilizes serial sections obtained from Epon-embedded specimens fixed with glutaraldehyde and osmium tetroxide. Double-immunofluorescence staining of thick Epon sections for endocrine hormones (insulin and glucagon) and regenerating islet-derived gene 1 α (REG1α) was performed following the removal of Epoxy resin with sodium ethoxide, antigen retrieval by autoclaving, and de-osmification treatment with hydrogen peroxide. The immunofluorescence images of endocrine cells were superimposed with the electron microscopy images of the same cells obtained from serial ultrathin sections. Immunofluorescence images showed well-preserved secretory granules in endocrine cells, whereas electron microscopy observations demonstrated corresponding secretory granules and intracellular organelles in the same cells. In conclusion, the correlative imaging approach developed by us may be useful for examining ultrastructural features in combination with immunolocalisation of endocrine hormones in the same human pancreatic islets.

Knowledge about the three-dimensional fine structure of human heart, as a crucial vital organ of the body, is not only fascinating from the scientific or educational points of view, but has a very important clinical impact. Therefore, we decided to create a three-dimensional atlas of fine structure of the human heart. Tissue samples from ten human hearts were rinsed in phosphate-buffer solution, fixed by glutaraldehyde buffered solution, and post-fixed in osmium tetroxide solution. A gentle dehydration with ethanol in different concentration and drying at the critical point of CO2 were applied as next procedures. Non-conductive specimens were galvanized with thin gold layer and observed in scanning electron microscope. In this study, we present the three-dimensional ultrastructural architecture of the human heart from patients after myocardial infarction, end-stage failing heart as well as without apparent cardiac abnormalities at the time of autopsy. The results are presented as a histological atlas. Its images illustratively describe the fine structure of endocardium including the morphology of Purkyně (Purkinje) fibres, spatial arrangements of cardiac muscle cells inside myocardium or the arrangements of adipose tissue of epicardium. We present also figures of the ultrastructure of papillary muscles, intercalated discs as well as the connective tissue scars after myocardial infarction. The scanning electron microscopy could be a reliable technique to perform a more complete morphological data and to improve our knowledge of some pathological changes of tissues and cells, not always detected by conventional morphological examinations.

Intracellular accumulation of excessive glycogen or lipid in the liver occurs as a manifestation of metabolic dysfunction or drug-induced toxicity in humans or animals. The identification of individual components (lipids, glycogen, or water) involved is essential to the diagnosis of disease and pathogenesis. The aqueous fixatives and processing solvents used in paraffin embedding for routine hematoxylin and eosin stains remove both lipid and glycogen, leaving clear vacuoles of which the precise content is unknown in routine hematoxylin and eosin-stained sections. Traditional methods for detecting intracellular lipids and glycogen require two separate techniques, such as periodic acid-Schiff (PAS) reaction, with or without diastase digestion for glycogen, and osmium tetroxide (OsO4) post-fixation for lipid and utilization of different tissue sections. We describe here, for the first time, a novel OsO4/PAS dual-staining technique that permits a simple color-coded detection and direct visualization of both l...

The influence of an electric field on an isolated channel or nanopore separating two compartments filled with electrolytes produces a constant ion flux through the pore. Nucleic acids added to one compartment traverse the pore, and modulate the current in a sequence-dependent manner. While translocation is faster than detection, the α-hemolysin nanopore (α-HL) successfully senses base modifications in ssDNA immobilized within the pore. With the assistance of a processing enzyme to slow down translocation, nanopore-based DNA sequencing is now a commercially available platform. However, accurate base calling is challenging because α-HL senses a sequence, and not a single nucleotide. Osmylated DNA was recently proposed as a surrogate for nanopore-based sequencing. Osmylation is the addition of osmium tetroxide 2,2’-bipyridine (OsBp) to the C5–C6 pyrimidine double bond. The process is simple, selective for deoxythymidine (dT) over deoxycytidine (dC), unreactive towards the purines, practically 100% effective, and strikingly independent of length, sequence, and composition. Translocation of an oligodeoxynucleotide (oligo) dA10XdA9 via α-HL is relatively slow, and exhibits distinct duration as well as distinct residual current when X = dA, dT(OsBp), or dC(OsBp). The data indicate that the α-HL constriction zone/β-barrel interacts strongly with both OsBp and the base. A 23 nucleotide long oligo with four dT(OsBp) traverses 18-times slower, and the same oligo with nine (dT+dC)(OsBp) moieties traverses 84-times slower compared to dA20, suggesting an average rate of 40 or 180 μs/base, respectively. These translocation speeds are well above detection limits, may be further optimized, and clear the way for nanopore-based sequencing using osmylated DNA.

Gemcitabine analogues with a lipophilic 4-N-alkyl chain bearing a terminal β-keto sulfonate moiety suitable for fluorination compatible with 18F-radiolabeling have been explored. Displacement of p-toluenesulfonylamino in protected 4-N-tosylgemcitabine with 1-amino-10-undecene gave 4-N-(10-undecenyl)-3',5'-di-O-benzoyl-2'-deoxy-2',2'-difluorocytidine. Oxidation of the terminal double bond in the latter with OsO4/NMO afforded 4-N-(10,11-dihydroxyundecanyl) derivative. Regioselective sulfonation of primary hydroxyl followed by oxidation of secondary hydroxyl with Collin's reagent yielded desired β-keto sulfonate analogues 8 or 9. Subsequent displacement of the mesylate or tosylate group with KF in the presence of Kryptofix 2.2.2. or 18-crown-6 ether followed by deprotection with NH3/MeOH gave 4-N-(11-fluoro-10-oxoundecanyl)-2'-deoxy-2',2'-difluorocytidine 11.

Fluorescent proteins facilitate a variety of imaging paradigms in live and fixed samples. However, they lose their fluorescence after heavy fixation, hindering applications such as correlative light and electron microscopy (CLEM). Here we report engineered variants of the photoconvertible Eos fluorescent protein that fluoresce and photoconvert normally in heavily fixed (0.5-1% OsO4), plastic resin-embedded samples, enabling correlative super-resolution fluorescence imaging and high-quality electron microscopy.

A novel and convenient strategy for iodine labeled glycopeptide molecular probe and purification was developed. The fluorine rich bi-functional coupling agent, 4-tris(2-perfluorohexylethyl)stannylbenzoate succinimidyl ester, was successfully synthesized via 5 steps starting from the fluorous Grignard reagent. It was purified by a simple and fast isolation using perfluorinated hexanes (FC-72). The "cold" iodine labeled yield for the coupling agent was as high as 92% within 15 min. The iodine-labeled product was only in organic fractions as we expected. It was shown that there was only one major peak in organic fractions according to HPLC. Finally, the iodine-labeled coupling agent was applied to label glycopeptide and afforded a high yield of 87% within 30 min.

A range of cyclic allylic trichloroacetamides has been synthesised. Dihydroxylation utilising catalytic osmium tetroxide and quinuclidine-N-oxide monohydrate as the re-oxidant in dichloromethane yields diols with good levels of syn selectivity. (C) 2000 Elsevier Science Ltd.

Single-stranded DNA and RNA aptamers are able to bind to specific target molecules with high affinity and are widely used in many routine molecular-diagnostic laboratories. Developing methods for direct detection of specific targets using aptamers could be challenging. As a central protease of hemostasis, thrombin is widely used as targeted molecule for testing new analytical approaches. To develop an electrochemical method for the detection of thrombin, our collaborator uses osmium tetroxide, 2,2' bipyridine as an electrochemical tag that can be detected at an electrode surface. However, conjugation of the osmium to thrombin can potentially change the binding interface, diminishing the binding affinity of the aptamer to the thrombin analyte. To gain insight into the molecular basis of the thrombin-aptamer interaction, we investigated systems of hydrated thrombin and osmium-thrombin in the presence of DNA aptamer with 29 nucleotides. In particular, we conducted both canonical simulations and umbrella sampling with exchange of umbrellas to calculate the binding energy of the aptamer with thrombin and osmium-thrombin. We found that the orientations of thrombin-aptatmer complex rapidly relax from native conformation into another more energetically favourable conformations. Our results also indicate that, as a result of conjugating osmium complex to thrombin, the binding affinity of aptamer for osmium-thrombin conjugate significantly decreased in relation to thrombin/aptamer system. Therefore, we redesigned aptamers to regain the lost affinity for the osmium-thrombin conjugate to accommodate the osmium tags. We then calculate and compare the binding free energy of computer-optimized aptamers in the present of thrombin and osmium-thrombin conjugate. Overall, our results reveal in atomistic detail a variety of redesigned aptamers that may have enhanced binding affinity for osmium-thrombin conjugate. Our next step is to validate our computer-optimized aptamers in the presence of thrombin and osmium-thrombin using experimental methods.

With the recent advances in electron microscopy (EM), computation, and nanofabrication, the original idea of reading DNA sequence directly from an image can now be tested. One approach is to develop heavy atom labels that can provide the contrast required for EM imaging. While evaluating tentative labels for the respective nucleobases in synthetic oligodeoxynucleotides (oligos), we developed a streamlined CE protocol to assess the label stability, reactivity, and selectivity. We report our protocol using osmium tetroxide 2,2'-bipyridine (Osbipy) as a thymidine (T) specific label. The observed rates show that the labeling process is kinetically independent of both the oligo length, and the base composition. The conditions, i.e. temperature, optimal Osbipy concentration, and molar ratio of reagents, to promote 100% conversion of the starting oligo to labeled product were established. Hence, the optimized conditions developed with the oligos could be leveraged to allow osmylation of effectively all Ts in ssDNA, while achieving minimal mislabeling. In addition, the approach and methods employed here may be adapted to the evaluation of other prospective contrasting agents/labels to facilitate next-generation DNA sequencing by EM.

We demonstrate an approach for the electrochemical detection of genetically modified maize in real maize flour samples by means of square-wave voltammetry (SWV). After labeling the asymmetric PCR-amplified targets with osmium tetroxide bipyridine [OsO4(bipy)], they were hybridized with immobilized oligonucleotide probes on gold electrodes. We could detect the maize genes ivrp and SSIIb in near isogenic maize. The transgene cryIa/b and the MON810 specific fragment were detected in all transgenic maize samples, down to a content of 0.6% of MON810 in mixed samples. While it was possible to detect all sequences in the samples containing 100% near isogenic or respectively transgenic maize after a hybridization time of less than 10 min, a hybridization time of 30 min was necessary for the detection of the genetic modifications in samples containing only 0.9 to 0.6% of transgenic maize. No significant detection of the transgene cryIa/b or MON810 was possible when only 0.5% of transgenic maize was present in the sample, most likely due to insufficient amplification of the template DNA. Keywords: Electrochemical DNA sensor, Hybridization, Gold electrode, Osmium tetroxide bipyridine, Real sample, Maize MON810

Niemann-Pick disease (types A and B), or acid sphingomyelinase deficiency, is an inherited deficiency of acid sphingomyelinase, resulting in intralysosomal accumulation of sphingomyelin in cells throughout the body, particularly within those of the reticuloendothelial system. These cellular changes result in hepatosplenomegaly and pulmonary infiltrates in humans. A knockout mouse model mimics many elements of human ASMD and is useful for studying disease histopathology. However, traditional formalin-fixation and paraffin embedding of ASMD tissues dissolves sphingomyelin, resulting in tissues with a foamy cell appearance, making quantitative analysis of the substrate difficult. To optimize substrate fixation and staining, a modified osmium tetroxide and potassium dichromate postfixation method was developed to preserve sphingomyelin in epon-araldite embedded tissue and pulmonary cytology specimens. After processing, semi-thin sections were incubated with tannic acid solution followed by staining with toluidine blue/borax. This modified method provides excellent preservation and staining contrast of sphingomyelin with other cell structures. The resulting high-resolution light microscopy sections permit digital quantification of sphingomyelin in light microscopic fields. A lysenin affinity stain for sphingomyelin was also developed for use on these semi-thin epon sections. Finally, ultrathin serial sections can be cut from these same tissue blocks and stained for ultrastructural examination by electron microscopy.

The oxidation of ethylene by osmium tetroxide (OsO4), osmyl hydroxide (OsO2(OH)2) and osmyl chloride (OsO2Cl2) have been studied by hybrid density functional theory at the B3LYP/LACVP∗ level of theory. It was found that in the reaction of OsO4 with ethylene, the [3 + 2] addition pathway leading to a five-membered metallacycle intermediate is more favorable than the [2 + 2] addition to form a four-membered metallaoxetane and that the reaction leads ultimately to the formation of diols without epoxide formation, in agreement with earlier works on the subject. The reaction of ethylene with osmyl hydroxide (OsO2(OH)2), which is released when the ‘cyclic’ esters formed from [3 + 2] reaction of OsO4 with ethylene undergo hydrolysis, was found to be less favorable, kinetically and thermodynamically, than the [3 + 2] reaction of OsO4 with ethylene. In the reaction of OsO2Cl2 with ethylene it was found that the [2 + 2] addition pathway to form a four-membered metallacycle is more favorable than the [3 + 2] addition to form the five-membered metallacycle intermediate. It was also found that the formation of an epoxide precursor from the reaction of OsO2Cl2 with ethylene, though possible, is a very unfeasible reaction kinetically and therefore epoxide is not a likely product in the reaction. The results of this study indicates that the preference of the [3 + 2] pathway in the addition of metal oxides across C C bonds, which has been established for osmium tetroxide, is not pervasive in all osmium oxo complexes as publications on the subject seem to suggest.

A new chemoentrapment strategy for recycling osmium in the catalytic olefin cleavage reaction is reported. The new strategy utilizes KOH/ i -PrOH to generate water-soluble Os(VI) species as a recyclable metal catalyst after the oxidative cleavage reaction. For the recycling of the catalyst, NaIO 4 –NaClO 2 was found to be the best combination of secondary oxidants and acetonitrile–water was chosen as an optimal solvent for the best recycling results. The new method allows for an efficient recycling of osmium in the reactions involving mono- and di-substituted olefins with 1 mol % of OsO 4 without any significant side reactions and loss of yield.

Preserving the structural arrangement of the components of a bacterial infection process within a plant for microscopy study is a technical challenge because of the different requirements of each component for optimal preservation and visualization. We used low temperature scanning electron microscopy (cryo-SEM), anhydrous fixation at ambient temperature and freeze-substitution for transmission electron microscopy to examine fractured and sectioned strawberry leaves infected with Xanthomonas fragariae. Cryo-SEM images of fractured samples showed the bacterial colonization of mesophyll air spaces in the leaf, limited by the vascular bundles and the orientation and packing of bacteria in extracellular polysaccharide. Transmission electron microscopy of samples fixed using osmium tetroxide dissolved in FC-72 solvent at ambient temperature showed that the entire plant/bacteria/extracellular polysaccharide system was preserved in situ, and showed plasmolysis of mesophyll cells and disruption of organelles. In freeze-substitution samples, osmium tetroxide in FC-72 solvent gave superior preservation of the extracellular polysaccharide as compared to a conventional cocktail. In addition, strands believed to be xanthan were preferentially contrasted to show their density and orientation around the bacterial cells. We conclude that anhydrous fixation using osmium tetroxide in FC-72 at ambient temperature gave the best preservation of the entire system, and freeze-substitution using this same fixative enhanced the visualization of strands in the biofilm.

The Golgi method has been used for over a century to describe the general morphology of neurons in the nervous system of different species. The “single-section” Golgi method of Gabbott and Somogyi (1984) and the modifications made by Izzo et al. (1987) are able to produce consistent results. Here, we describe procedures to show cortical and subcortical neurons of human brains immersed in formalin for months or even years. The tissue was sliced with a vibratome, post-fixed in a combination of paraformaldehyde and picric acid in phosphate buffer, followed by osmium tetroxide and potassium dicromate, “sandwiched” between cover slips, and immersed in silver nitrate. The whole procedure takes between 5 and 11 days to achieve good results. The Golgi method has its characteristic pitfalls but, with this procedure, neurons and glia appear well-impregnated, allowing qualitative and quantitative studies under light microscopy. This contribution adds to the basic techniques for the study of human nervous tissue with the same advantages described for the “single-section” Golgi method in other species; it is easy and fast, requires minimal equipment, and provides consistent results.

An efficient synthetic route of L-hamamelose was successfully accomplished starting from D-ribose. L-Hamamelose was synthesized in 42% overall yield with six reaction steps via a stereoselective Grignard reaction, a stereoselective crossed aldol reaction and a controlled oxidative cleavage of the double bond of a vinyl diol compound. During the oxidative cleavage of the double bond of the vinyl diol compound with osmium tetroxide and NaIO(4), an over-oxidative cleavage of alpha-hydroxyl aldehyde generated from ring opening of the first cleaved product, formyl lactol, did not occur, probably due to the stability of the lactol form. A plausible mechanism for the stereoselective crossed aldol reaction was suggested. The final target compound, L-hamamelose can play a very important role as a chiral building block in synthesizing a wide variety of enantiopure compounds.

Electron microscopy revealed the presence of a 600-1500 A thick layer of polysaccharide on the surface of human corneal endothelial cells. The surface layer was visualized by combined fixation and staining in a mixture of ruthenium red and osmium tetroxide. The coating material was stable for at least 39 h post mortem and was retained on disintegrating cells.

Conspicuous cytoplasmic granules are reported in a magnetotactic multicellular prokaryote named ‘ Candidatus Magnetoglobus multicellularis’. Unfortunately, this microorganism, which consists of an assembly of gram-negative bacterial cells, cannot yet be cultivated, limiting the biochemical analysis of the granules and preventing in vitro studies with starvation/excess of nutrients. In this scenario, light and electron microscopy techniques were used to partially address the nature of the granules. Besides magnetosomes, three types of inclusions were observed: small (mean diameter = 124 nm) polyhydroxyalkanoate-like (PHA) granules, large (diameters ranging from 0.11 to 2.5 μm) non-PHA lipid granules, and rare phosphorus-rich granules, which probably correspond to polyphosphate bodies. The PHA granules were rounded in projection, non-reactive with OsO 4 , and suffered the typical plastic deformation of PHAs after freeze fracturing. The nature of the large granules, consisting of round globular structures (mean diameter = 0.76 μm), was classified as non-PHA based on the following data: (a) multilayered structure in freeze-fracture electron microscopy, typical of non-PHA lipids; (b) Nile blue fluorescence imaging detected non-PHA lipids; (c) imidazole buffered osmium tetroxide and ruthenium red cytochemistry stained the globules, which appeared as electron-dense granules instead of electron lucent as PHAs do. Most likely, ‘ Candidatus Magnetoglobus multicellularis’ stores carbon mainly as unusual lipid granules, together with smaller amounts of PHAs.

The effect of high glucose concentrations on the ultrastructure and the production of docosahexaenoic acid (DHA) by Aurantiochytrium mangrovei MP2 was investigated at 25°C with orbital shaking. The cultured cells were separated into a floating and a bottom layer after centrifugation during harvest; therefore, the ultrastructure and DHA level were also analyzed separately. Cell size generally increased with glucose concentrations, whereas the overall DHA production (mg/l) increased 19% when the glucose concentration was raised from 6% to 10% w/v. Biomass and DHA production increased significantly, but not linearly, at the floating layer and decreased at the bottom layer in elevated glucose concentrations. Also, the lipid bodies of the cells in the floating layer were more heavily stained in osmium tetroxide than those in the bottom, suggesting that the cells in the floating layer may contain greater amount of unsaturated fatty acids.

This paper presents a new approach to electrochemical sensing of DNA damage, using osmium DNA markers and voltammetric detection at the pyrolytic graphite electrode. The technique is based on enzymatic digestion of DNA with a DNA repair enzyme exonuclease III (exoIII), followed by single-strand (ss) selective DNA modification by a complex of osmium tetroxide with 2,2'-bipyridine. In double-stranded DNA possessing free 3'-ends, the exoIII creates ss regions that can accommodate the electroactive osmium marker. Intensity of the marker signal measured at the pyrolytic graphite electrode responded well to the extent of DNA damage. The technique was successfully applied for the detection of (1) single-strand breaks (ssb) introduced in plasmid DNA by deoxyribonuclease I, and (2) apurinic sites generated in chromosomal calf thymus DNA upon treatment with the alkylating agent dimethyl sulfate. The apurinic sites were converted into the ssb by DNA repair endonuclease activity of the exoIII enzyme. We show that the presented technique is capable of detection of one lesion per approximately 10(5) nucleotides in supercoiled plasmid DNA.

Hydrogen peroxide and nicotinamide adenine dinucleotide (NADH) may be determined amperometrically using screen-printed electrodes chemically modified with iron(III) hexacyanoosmate(II) (Osmium purple) in flow injection analysis (FIA). The determination is based on the exploitation of catalytic currents resulting from the oxidation/reduction of the modifier. The performance of the sensor was characterized and optimized by controlling several operational parameters (applied potential, pH and flow rate of the phosphate buffer). Comparison has been made with analogous complexes of ruthenium (Ruthenium purple) and iron (Prussian blue). Taking into account the sensitivity and stability of corresponding sensors, the best results were obtained with the use of Osmium purple. The sensor exhibited a linear increase of the amperometric signal with the concentration of hydrogen peroxide in the range of 0.1–100 mg L −1 with a detection limit (evaluated as 3 σ ) of 0.024 mg L −1 with a R.S.D. 1.5% for 10 mg L −1 H 2 O 2 under optimized flow rate of 0.4 mL min −1 in 0.1 M phosphate buffer carrier (pH 6) and a working potential of +0.15 V versus Ag/AgCl. Afterwards, a biological recognition element – either glucose oxidase or ethanol dehydrogenase – was incorporated to achieve a sensor facilitating the determination of glucose or ethanol, respectively. The glucose sensor gave linearity between current and concentration in the range from 1 to 250 mg L −1 with a R.S.D. 2.4% for 100 mg L −1 glucose, detection limit 0.02 mg L −1 (3 σ ) and retained its original activity after 3 weeks when stored at 6 °C. Optimal parameters in the determination of ethanol were selected as: applied potential +0.45 V versus Ag/AgCl, flow rate 0.2 mL min −1 in 0.1 M phosphate buffer carrier (pH 7). Different structural designs of the ethanol sensor were tested and linearity obtained was up to 1000 mg L −1 with a maximum R.S.D. of 5.1%. Applications in food analysis were also examined.

This review summarizes the function, structure, and chemistry of the myelin sheaths of axons: discusses the mechanisms of several staining techniques using inorganic compounds, dyes, and antibodies; and provides technical instructions for 13 staining methods for normal and degenerating myelin. Myelin has alternating hydrophobic and hydrophilic layers. The former contain the hydrocarbon tails of lipid molecules and hydrophobic segments of transmembrane proteins; the latter contain the anionic heads of polar lipids and also basic proteins. Aqueous fixatives allow some myelin lipids to be retained in paraffin sections. Stains for normal myelin include histochemical reactions that detect choline-containing phospholipids (Baker's chromation-acid hematein) or double bonds (unsaturation) in the fatty acid components of lipids (osmium tetroxide or palladium chloride reduction; the pseudoplamal reaction and DAB fluorescence, for oxidized double bonds). Traditional fat stains (Sudan dyes) stain normal myeli...

A general method for the solid-phase synthesis of carboxy-functionalized peptides by oxidative cleavage of alkynes is presented. Clean and quantitative conversion is enabled by the addition of bases, such as DABCO and HMTA, to the classical OsO4/NaIO4 mixture. The utility of the reaction is further illustrated by the synthesis of oxamic acids.

Immunogold labeling of cryosections according to Tokuyasu (Tokuyasu KT. A technique for ultracyotomy of cell suspensions and tissues. J Cell Biol 1973;57:551-565), is an important and widely used method for immunoelectron microscopy. These sections are cut from material that is chemically fixed at room temperature (room temperature fixation, RTF). Lately in many morphological studies fast freezing followed by cryosubstitution fixation (CSF) is used instead of RTF. We have explored some new methods for applying immunogold labeling on cryosections from high-pressure frozen cells (HepG2 cells, primary chondrocytes) and tissues (cartilage and exocrine pancreas). As immunolabeling has to be carried out on thawed and stable sections, we explored two ways to achieve this: (1) The section fixation method, as briefly reported before (Liou W et al. Histochem Cell Biol 1996;106:41-58 and Möbius W et al. J Histochem Cytochem 2002;50:43-55.) in which cryosections from freshly frozen cells were stabilized in mixtures of sucrose and methyl cellulose and varying concentrations of glutaraldehyde, formaldehyde and uranyl acetate (UA). Only occasionally does this method reveal section areas with excellent cell preservation and negatively stained membranes like Tokuyasu sections of RTF material. (Liou et al.) (2) The rehydration method, a novel approach, in which CSF with glutaraldehyde and/or osmium tetroxide (OsO4) was followed by rehydration and cryosectioning as in the Tokuyasu method. Especially, the addition of UA and low concentrations of water to the CSF medium favored superb membrane contrast. Immunogold labeling was as efficient as with the Tokuyasu method.

The endomembranous system of Serrasalmus spilopleura oocyte secondary growth was analysed using structural and ultrastructural cytochemical techniques. In vitellogenic oocytes, the endoplasmic reticulum components, the nuclear envelope intermembranous space, some Golgi dictiossomes, lysosomes, yolk granules, regions of the egg envelope and sites of the follicle cells react to acid phosphatase detection (AcPase). The cortical alveoli, some heterogeneous cytoplasmic structures, regions of the egg envelope, and sites of the follicle cells are strongly contrasted by osmium tetroxide and zinc iodide impregnation (ZIO). The endoplasmic reticulum components, some vesicles, and sites of the follicle cells also react to osmium tetroxide and potassium iodide impregnation (KI). The biosynthetic pathway of lysosomal proteins, such as acid phosphatase, required for vitellogenesis, involves the endoplasmic reticulum, Golgi complex, vesicles with inactive hydrolytic enzymes, and, finally, lysosomes. In S. spilopleura oocytes at secondary growth, the endomembranous system takes part in the production of the enzymes needed for vitellogenesis, and in the metabolism of yolk exogenous components (AcPase detection). The endomembranous system compartments also show reduction capacity (KI reaction) and are involved in the metabolism of proteins rich in SH-groups (ZIO reaction).

We have utilized protective oligonucleotides to modify DNA fragments with osmium tetroxide complexes without compromising their ability to hybridize with immobilized thiol-linked probe-SAMs on gold electrodes. Due to reversible voltammetric signals of Os(VI/IV), this method allowed sensitive electrochemical hybridization detection of short (25 bases) and long (120 bases) thymine-containing DNA targets. The detection limit was 3.2 nM for the long target. We found an optimum 40 degrees C hybridization temperature for the short target. No interference by noncomplementary DNA was observed. At least 10 repetitive hybridization experiments at the same probe-SAM were possible with thermal denaturation in between. Such use of protective strands could be useful also for other types of DNA recognition and even for other DNA-modifying agents. Moreover, it is possible to produce electrochemically active oligonucleotides (targets and reporter probes) in ones own laboratory in a simple way.

We have discovered that the immunoreactivity of the fluorophore Alexa Fluor 488 survives glutaraldehyde and osmium tetroxide fixation and epoxy resin embedding and etching. We have developed new localization methods that for the first time take advantage of this property. The antigen is localized in cryosections using suitable primary antibody and an Alexa Fluor 488-conjugated secondary antibody. Cryosection fluorescence can be photographed for later correlation with electron microscopy (EM) findings. The sections are then further fixed with glutaraldehyde and OsO4, if desired and flat-embedded in epoxy resin. Semi-thin sections are etched completely with sodium ethoxide, whereas thin sections are partially etched. Alexa Fluor 488 is then localized with rabbit anti-Alexa Fluor 488 and goat anti-rabbit conjugated to Alexa Fluor 488 [light microscopy (LM)] or to colloidal gold (EM). A second antigen may also be localized using Alexa Fluor 568. When used without postfixation, these methods produce high-resolution semi-thin, or even thin, sections that retain a high level of fluorescence for LM observations. These methods allow highly sensitive immunolocalizations in tissue while preserving cell fine structure through traditional fixation and epoxy embedding. In demonstration of the methods, we describe the localization of the thiazidesensitive sodium/chloride cotransporter and the epithelial sodium channel in rat kidney.

In the present paper, we report the results we obtained using several cytochemical techniques to analyze the infective larva of Wuchereria bancrofti. An imidazole osmium tetroxide solution was used to visualize unsaturated fatty acids. A highly contrasted material forming a continuous structure was observed on the larval surface and over the epicuticle. A strong reaction was observed on the esophagus and also on the inner secreted material. Carbohydrates containing vic-glycol groups were not observed on the cuticle of the third-stage larva of W. bancrofti submitted to the Thiéry technique. Using a panel of eight gold-labeled lectins, we found that the cuticle exhibited slight labeling with all lectins used, indicating residues of N-acetyl-D: -glucosamine, N-acetyl-galactosamine, D: -galactose, D: -manose, and L: -fucose. Surface anionic sites were visualized by using cationized ferritin particles. Treatment with trypsin partially inhibited the reaction, whereas the treatment with chondroitinase ABC, a specific enzyme for glycosaminoglycans, completely abolished the labeling with cationic particles.

The UV‐VIS spectrophotometric methods for the determination of Os(VIII) (as OsO4) and Os(IV) (as OsCl6 2− complex) in their mixtures were developed. Quercetin (Q), a flavonoid compound, was used as a chromogenic reagent. Both direct and derivative spectrophotometry can be employed for the determination of Os(VIII). The calculation of the first‐derivative spectrum of the examined mixture and the use of the signal at 285.1 nm allows reaching a better detection limit (0.01 µg mL−1 Os) as compared with direct spectrophotometry (0.1 µg mL−1 Os). Relative standard deviations of the results are in the range of 0.87%–4.65% and 0.45%–1.15% for direct and derivative mode, respectively. Selective redox reaction of OsO4 with Q under the conditions used (0.05 M HCl, 1×10−4 M Q, 15 min heating at 70°C) makes the basis of its determination in mixtures with the OsCl6 2− complex. Quercetin does not react with the OsCl6 2− complex. The signals of the OsCl6 2− complex can be isolated from the examined mixtures by t...

Chlorosomes are the light harvesting structures of green photosynthetic bacteria. Each chlorosome from green sulfur bacteria houses hundreds of thousands of bacteriochlorophyll molecules in addition to smaller amounts of chlorobiumquinone and carotenoids. In electron microscopy studies, chlorosomes exhibit different appearances depending on the fixation method used. Fixation with osmium tetroxide results in electron-transparent chlorosomes. Fixation with potassium permanganate results in clearly delineated electron-dense chlorosomes. This fixation method features an electron-transparent area in the interior of the chlorosome. In addition to electron density patterns that can be considered compositions of rod-shaped elements, chlorosomes exhibit a striation pattern that is oriented parallel to the longitudinal axis. Treatment with osmium tetroxide followed by potassium permanganate treatment results in a more diffused density distribution that outlines connecting elements between the chlorosome and the cytoplasmic membrane, and connecting elements between the cytoplasmic membrane and the outer membrane, which act as a diffusion barrier for electron density.

[reaction: see text]. The use of 2-O-propargyl ethers as protecting groups in 4,6-O-benzylidene-protected mannopyranosyl donors bearing either bulky silyl groups or glycosidic linkages on O3 overcomes the poor stereoselectivity achieved with the corresponding 2-O-benzyl ethers, due to a reduction of steric buttressing. Deprotection is conducted by treatment with potassium tert-butoxide followed by catalytic osmium tetroxide and N-methylmorpholine N-oxide.

Aqueous solutions of osmium tetroxide (OsO4) have been injected into arthritic knees for the past 45 years to chemically destroy diseased tissue, in a procedure termed “chemical synovectomy.” Arthritis is an inflammatory disease. The primary inflammatory chemical species are the superoxide anion radical (O2 −) and nitric oxide ( NO), which combine to form the peroxynitrite anion (ONOO−). Here we show that OsO4 does not react with ONOO− but very efficiently catalyzes the dismutation of O2 − to O2 and H2O2. Using the pulse-radiolysis technique, the catalytic rate constant has been determined to be (1.43 ± 0.04) × 109 M−1 s−1, independent of the pH in the 5.1–8.7 range. This value is about half that for the natural Cu,Zn-superoxide dismutase (Cu,Zn-SOD). Per unit mass, OsO4 is about 60 times more active than Cu,Zn-SOD. The catalytically active couple is OsVIII/OsVII, OsVIII oxidizing O2 − to O2 with a bimolecular rate constant of k = (2.6 ± 0.1) × 109 M−1 s−1 and OsVII reducing it to H2O2 with a bimolecular rate constant of (1.0 ± 0.1) × 109 M−1 s−1. Although lower valent osmium species are intrinsically poor catalysts, they are activated through oxidation by O2 − to the catalytic OsVIII/OsVII redox couple. The OsVIII/OsVII catalyst is stable to biochemicals other than proteins and peptides comprising histidine, cysteine, and dithiols.

Osmium tetroxide is reduced by molecular hydrogen in the presence of ligands in both polar and nonpolar solvents. In CHCl3 containing pyridine (py) or 1,10-phenanthroline (phen), OsO4 is reduced by H2 to the known Os(VI) dimers L2Os(O)2(mu-O)2Os(O)2L2 (L2 = py2, phen). However, in the absence of ligands in CHCl3 and other nonpolar solvents, OsO4 is unreactive toward H2 over a week at ambient temperatures. In basic aqueous media, H2 reduces OsO4(OH)n(n-) (n = 0, 1, 2) to the isolable Os(VI) complex, OsO2(OH)4(2-), at rates close to that found in py/CHCl3. Depending on the pH, the aqueous reactions are exergonic by deltaG = -20 to -27 kcal mol(-1), based on electrochemical data. The second-order rate constants for the aqueous reactions are larger as the number of coordinated hydroxide ligands increases, k(OsO4) = 1.6(2) x 10(-2) M(-1) s(-1)k(OsO4(OH)-) = 3.8(4) x 10(-2) M(-1) s(-1)k(OsO4(OH)2(2-)) = 3.8(4) x 10(-1) M(-1) s(-1). The observation of primary deuterium kinetic isotope effects, k(H2)/k(D2) = 3.1(3) for OsO4 and 3.6(4) for OsO4(OH)-, indicates that the rate-determining step in each case involves H-H bond cleavage. Density functional calculations and thermochemical arguments favor a concerted [3+2] addition of H2 across two oxo groups of OsO4(L)n and argue against H* or H- abstraction from H2 or [2+2] addition of H2 across one Os=O bond. The [3+2] mechanism is analogous to that of alkene addition to OsO4(L)n to form diolates, for which acceleration by added ligands has been extensively documented. The observation that ligands also accelerate H2 addition to OsO4(L)n highlights the analogy between these two reactions.

Two series of chlorin p6 13,15-cycloimides that differ in their substituents at the nitrogen atom of the additional six-membered ring were synthesized. The compounds of the first series have a hydroxyl, alkoxyl, or acyloxy group at the 13,15-cycloimide nitrogen and those of the second series, residues of aliphatic alcohols. The cycloimides synthesized are satisfactorily stable and display an intensive light absorption maximum at 710-718 nm. Treatment of the cycloimides with sodium periodate in the presence of osmium tetroxide and with the Vilsmeier reagent resulted in the formation of 3-formyl- and 3-(2-formylvinyl)derivatives, respectively. The conversion of vinyl into formyl group or 2-formylvinyl group leads to an additional bathochromic shift of the long-wave maximum by 30 nm on an average. An extra hydroxy group was introduced at position 18 of the macrocycle to increase the cycloimide hydrophilicity. The English version of the paper: Russian Journal of Bioorganic Chemistry, 2004, vol. 30, no. 5; see also http: // www.maik.ru.

Site-specific chemical modification is a useful technology in characterisation of proteins, but the number of chemical probes of the protein structure reacting with proteins under mild conditions in aqueous solutions is rather limited. Here we studied the reaction of osmium tetroxide, 2,2′-bipyridine (Os,bipy) with several peptides using capillary zone electrophoresis (CZE) and matrix-assisted laser desorption-ionisation–time-of-flight mass spectrometry (MALDI–TOF MS). Both techniques showed formation of a stable complex of Os,bipy with tryptophan residues. In CZE peaks with different migration times and UV-Vis spectra were observed. MALDI–TOF MS showed the formation of a product with characteristic isotopic pattern corresponding to the presence of osmium atom. Oxidation of cysteine and methionine side chains to cysteic acid and methionine sulfone by Os,bipy was detected by CZE and confirmed by MALDI–TOF and post-source decay (PSD) mass spectra. PSD showed specific shifts of molecular weights of the peptides and their fragments after the derivatisation. We believe that Os,bipy may become a useful agent in the characterisation of proteins.

[reaction: see text] A concise total synthesis of 1-epiaustraline 3 is described that utilizes a diastereoselective Birch reduction of an electron-deficient pyrrole and a chelation-controlled vinyl Grignard addition to an aldehyde to introduce the C7 stereocenter. The C1 and C2 stereocenters were set through an OsO(4)-catalyzed dihydroxylation.

Dufour glands of Apis mellifera and Melipona bicolor were studied under light and transmission electron microscopy, using the cytochemical techniques of mercury bromophenol blue for protein detection, imidazole-buffered osmium tetroxide selective staining of unsaturated lipids, lanthanum nitrate for intercellular junction identification and zinc-iodide-osmium tetroxide for cytoplasmic endomembrane visualization. The results in both species corroborated the lipid nature of the gland secretion and showed in A. mellifera the poverty of the synthetic machinery in the worker gland cells in comparison with the queen, as expected by previous biochemical analyses. The pathway of the exogenous compounds of the secretion is intracellular, since substances can penetrate the cell folds and intercellular junctions, but their access to the gland lumen is barred by the apical intercellular junctions.

Adherens junctions (AJs) are cell-cell and cell-matrix junctions that are known to comprise the transmembrane and cytoplasmic components linked to the f-actin cytoskeleton. Although the presence of AJs han been confirmed in normal human epidermis, previous studies immunolocalizing AJ-related antigens have been controversial. The purpose of this study was to produce a more precise molecular mapping of AJs and their constituents in relation to desmosomes in normal human epidermal keratinocytes. Using an electron microscope (EM) method to optimally fix plasma membranes. AJ structures were typically seen as a narrowing of the intercellular space between two keratinocytes that was distinct from desmosomes and gap junctions. Such structures were consistently found more frequently in the upper epidermis than in the basal layer. Immunogold electron microscopy showed an absence of the AJ components (E-cadherin and beta-catenin) from desmosomal areas but they were present at interdesmosomal areas at sites of close membrane association. Conversely, the desmosomal components plakoglobin and plakophilin 1 were restricted only to the outer attachment plaque of the desmosome. These results further confirm that AJs have a distinct molecular composition and distribution from desmosomes and that they regularly occur between desmosomes along the keratinocyte plasma membrane to provide alternative cell-cell adhesion mechanisms.

For in vivo NMR studies, starting from pyrroles, a series of fluorinated porphyrins were synthesized by following the MacDonald reaction conditions. Upon reaction with osmium tetroxide, a fluorinated porphyrin containing four trifluoromethyl groups (12 fluorine units) was converted into the related chlorin and bacteriochlorin which exhibited long-wavelength absorptions at 652 and 720 nm, respectively. All compounds produced good singlet oxygen production efficiency. A comparative study of nine porphyrins with and without fluorine substituents indicated no adverse effects of the presence of fluorinated groups in the photophysical properties of the porphyrins, chlorins or bacteriochlorins. The first and second one-electron reduction potentials (vs SCE) of the investigated compounds range between −1.29 and −1.49 V and between −1.66 and −1.84 V in PhCN containing 0.1 M TBAP. UV–visible spectroelectrochemical data suggested the formation of π-anion and π-cation radicals upon the first reduction and first oxidation. The in vivo 19F MR study of a representative fluorine labeled compound with twelve equivalent fluorines confirmed the presence of the fluorine labeled sensitizer in mouse (C3H/HeJ) implanted with RIF tumors on mouse foot dorsum by inoculating 2×105 cells (the studies were repeated on four tumored mice to confirm the feasibility and reproducibility). All fluorinated compounds were found to be quite effective in vitro. In a comparative intracellular localization study with Rhodamine-123 in RIF tumor cells, the most soluble porphyrin containing two propionic ester side chains was found to localize in mitochondria as well as the related chlorin and bacteriochlorin.

Carotenoids as well as their metabolites and oxidation products stimulate gap junctional communication (GJC) between cells, which is thought to be one of the protective mechanisms related to cancer-preventive activities of these compounds. Increased intake of lycopene by consumption of tomatoes or tomato products has been epidemiologically associated with a diminished risk of prostate cancer. Here, we report a stimulatory effect of a lycopene oxidation product on GJC in rat liver epithelial WB-F344 cells. The active compound was obtained by complete in vitro oxidation of lycopene with hydrogen peroxide/osmium tetroxide. For structural analysis high performance liquid chromatography, gas chromatography coupled with mass spectrometry, ultraviolet/visible-, and infrared spectrophotometry were applied. The biologically active oxidation product was identified as 2,7,11-trimethyl-tetradecahexaene-1,14-dial. The present data indicate a potential role of lycopene degradation products in cell signaling enhancing cell-to-cell communication via gap junctions.

Acrolein, a known mutagen, undergoes reaction in vitro under physiological conditions with both 2 ′ -deoxyguanosine and native DNA to give rise to exocyclic adducts of the 5,6,7,8-tetrahydropyrimido[1,2- a ]purine-10(3 H )-one class having an hydroxy group at either the 6 or the 8 position. Previously we have shown that the 8-hydroxy derivative in a bacterial system has very low mutagenicity probably because in double-stranded DNA this residue exists in the open-chain aldehydic form [ N 2 -(3-oxopropyl)-2 ′ -deoxyguanosine] ( 3 ). To continue our investigation in this area, we needed ample supplies of the 6-hydroxy isomers. This current paper describes high-yield simple methods for the synthesis in bulk of the 6-hydroxy adduct 1 and its incorporation into DNA oligomers. The basic methods for the synthesis of the adduct 1 , involve 1-substitution of dG derivatives with a 3-butenyl group, dihydroxylation of the olefin with osmium tetroxide and N -methylmorpholine N -oxide, then diol cleavage with periodate ion after incorporation of the 1-(3,4-diacetoxybutyl)-2 ′ -deoxyguanosine into oligomeric DNA.

Respiration and ion regulation are the two principal functions of teleostean gills. Mainly found in the gill filaments of fish, mitochondria-rich cells (MRCs) proliferate to increase the ionoregulatory capacity of the gill in response to osmotic challenges. Gill lamellae consist mostly of pavement cells, which are the major site of gas exchange. Although lamellar MRCs have been reported in some fish species, there has been little discussion of which fish species are likely to have lamellar MRCs. In this study, we first compared the number of filament and lamellar MRCs in air-breathing and non-air-breathing fish species acclimated to freshwater and 5 g NaCl L(-1) conditions. An increase in filament MRCs was found in both air-breathing and non-air-breathing fish acclimated to freshwater. Lamellar MRCs were found only in air-breathing species, but the number of lamellar MRCs did not change significantly with water conditions, except in Periophthalmus cantonensis. Next, we surveyed the distribution of MRCs in the gills of 66 fish species (including 29 species from the previous literature) from 12 orders, 28 families, and 56 genera. Our hypothesis that lamellar MRCs are more likely to be found in air-breathing fishes was supported by a significant association between the presence of lamellar MRCs and the mode of breathing at three levels of systematic categories (species, genus, and family). Based on this integrative view of the multiple functions of fish gills, we should reexamine the role of MRCs in freshwater fish.

Clustered DNA damage, where two or more lesions are located proximally to each other, is frequently induced by ionizing radiation. Individual base lesions within a cluster are repaired by base excision repair. In this study we addressed the question of how thymine glycol (Tg) within a cluster would affect the repair of opposing lesions by human cell extracts. We have found that Tg located opposite to an abasic site does not affect cleavage of this site by apurinic/apyrimidinic (AP) endonuclease. However, Tg significantly compromised the next step of the repair. Although purified DNA polymerase beta was able to incorporate the correct nucleotide (dAMP) opposite to Tg, the rate of incorporation was reduced by 3-fold. Tg does not affect 5'-sugar phosphate removal by the 2-deoxyribose-5-phosphate (dRP) lyase activity of DNA polymerase beta, but further processing of the strand break by purified DNA ligase III was slightly diminished. In agreement with these findings, although an AP site located opposite to Tg was efficiently incised in human cell extract, only a limited amount of fully repaired product was observed, suggesting that such clustered DNA lesions may have a significantly increased lifetime in human cells compared with similar single-standing lesions.

Oxidation of 5-acetamido-4,8-anhydro-1,2,3,5-tetradeoxy-D-glycero-D-ido-non-1-enitol [3-C-(2-amino-2-deoxy-beta-D-glucopyranosyl)-1-propene] was studied to search for preparative routes to aminodeoxy didehydro nonulosonic acid derivatives. Since only moderate chiral induction was observed with osmium tetroxide dihydroxylation as well as with peracid epoxidation, the catalytic asymmetric dihydroxylation conditions were applied to give the stereocontrolled formation of 1,2-propanediol derivatives. The structures of these diastereoisomeric 1,2-propanediol derivatives were determined by X-ray crystallographic analyses. The formation of diastereoisomeric 1,2-propanediols also varied with the nature of 2-substituent on the aminodoexy glycosyl moiety. Thus 5-acetamido-4,8-anhydro-3,5-dideoxy-D-erythro-L-ido-nonitol [(2S)-3-C-(2-acetamido-2-deoxy-beta-D-glucopyranosyl)-1,2-propanediol] was obtained predominantly up to 70% from 3-C-(2-acetamido-2-deoxyglycosyl)-1-propene by the use of ADmixbeta reagent. The (2S)-propanediol derivative was transformed in a five-step reaction sequence to 2,3-didehydro-2,7-dideoxy-N-acetylneuraminic acid.