Alkaline Reduction Research Articles

Secretion of a lactosaminoglycan-containing glycoprotein by peri-implantation sheep conceptuses

Sheep conceptuses from day 16 of pregnancy we cultured in the presence of [ 3H]glucosamine and [ 14C]leucine and a high-molecular-weight glycoprotein (HMWG) secreted into the culture medium was purified by a combination of anion-exchange and gel filtration chromatography. The HMWG was found to have a molecular weight between 800 000 and 900 000 and to be highly resistant to digestion with pronase. Characteristics of the carbohydrate portion of the purified glycoprotein were examined by selective chemical and enzymatic digestions and lectin binding studies. Mild alkaline reduction was ineffective in disassociating carbohydrate chains from the protein core. Furthermore, the protein was resistant to both O-glycanase and peptide: N-glycanase F. Harsh alkaline reduction caused the release of carbohydrates, however. After pronase digestion of these products, three molecular weight classes of carbohydrates were resolved by Sephadex G-25 chromatography. Two lines of evidence indicate that the HMWG contains lactosaminoglycan components. The intact molecule and two of the molecular weight classes of carbohydrates resolved by harsh alkaline reduction bind Datura stramonium lectin. Binding of HMWG to lectin could be partially inhibited by N-acetyllactosamine and completely inhibited by a mixture of N, N′-diacetylchitobiose and N, N′ , N″-triacetylchitotriose. Secondly, digestion with endo-β-galactosidase causes the release of 16% of the [ 3H]glucosamine from the intact molecule. Therefore, the HMWG of the sheep conceptus is the first reported example of secretion of lactosaminoglycan-containing glycoprotein by peri-implantation embryos.

4] Metal-free chromatographic media

The three major considerations in reducing metal-gel binding are (1) the type of gel to be used, (2) the ionic strength of the eluent, and (3) alkaline reduction of the gel. Each of these parameters should be considered in advance of an experiment. A method is offered for determining the optimal gel type, gel treatment, and eluent for a critical metal-free chromatography experiment.

A nuclear specific glycoprotein representative of a unique pattern of glycosylation.

Whole rat liver nuclei were reacted with UDP-[14C]galactose in the presence of bovine beta(1----4) galactosyltransferase. The reaction mixture was electrophoresed on a reducing sodium dodecyl sulfate-polyacrylamide gel. Autoradiograms of the gel demonstrated a major labeled broad band migrating with an apparent molecular weight of 65,000-66,000. A number of other less prominently labeled bands were also present. The labeled 65,000-66,000 band when cut from the gel and subjected to alkaline reduction while in the gel matrix exclusively yielded a 14C-labeled disaccharide that co-migrated with a [14C]Gal-GlcNAcol standard in descending paper chromatography. Treatment of this disaccharide with beta-galactosidase (beta-D-galactoside galactohydrolase; EC 3.2.1.23) from Aspergillus niger removed all the [14C]galactose label. Treatment of the labeled 65,000-66,000 polypeptide with Endoglycosidase F, however, did not remove the [14C]galactose label. Western transfer blots of sodium dodecyl sulfate-polyacrylamide gel electrophoresis gels performed with horseradish peroxidase-labeled succinyl wheat germ agglutinin, a lectin specific for GlcNAc, on unlabeled nuclei revealed a dominant band at 63,000-64,000. Subjecting 14C-labeled nuclei to this procedure resulted in a shift of the major horseradish peroxidase-labeled succinyl wheat germ agglutinin band to 65,000-66,000. The shifted band was coincident with the [14C]galactose band as visualized on an autoradiogram. A survey of other rat tissue nuclei revealed the same spectrum of [14C]galactose acceptor proteins with a dominant 65,000-66,000 galactose-labeled band.

Isolation and characterization of proteoglycans and glycosaminoglycans from human chondrosarcoma

The proteoglycans and glycosaminoglycans of four human chondrosarcomas with different degrees of malignancy (I–III) have been studied. The hydrodynamic size of proteoglycan subunits and the tissue concentration of total glycosaminoglycans decreased with increasing grade of malignancy. The glycosaminoglycan distribution pattern of all chondrosarcomas showed a similar ratio of chondroitin-4-sulfate: chondroitin-6-sulfate but an increasing portion of keratan sulfate from grade I (6.5%) to grade III (19.2%). Determinations of the molecular weight ( M r values) of glycosaminoglycans were made after 3H labeling by alkaline reduction of proteoglycans in the presence of NaB 3H 4. The M r of [ 3H]chondroitin sulfate isomeres decreased markedly from grade I (35,500) to grade III (15,100) while the chain length of [ 3H]keratan sulfate showed minor variations ( M r 5600–6200). The previously reported decrease in the molecular weight of keratan sulfate with increasing degree of malignancy ( FS. Pal, W. Strider, R. Margolis, G. Gallo, S. Lee-Huang, and L. Rosenberg, 1978, J. Biol. Chem. 253, 1279–1289) was not observed.

The measurement of glycosylated albumin by reduction of alkaline nitro-blue tetrazolium

A method is described for the measurement of glycosylated albumin in human plasma. Purification of albumin is achieved by a simple one-step Chromatographie method, adapted for use as a batch analysis technique. The glycosylated albumin fraction is subsequently measured by a modification of the fructosamine assay (alkaline reduction of nitroblue tetrazolium). The method correlates with the plasma fructosamine assay ( r = 0.85; p < 0.001) and with an electrophoretic method for the measurement of HbA 1 ( r = 0.78; p < 0.001).

Native and chemically modified porin channels from Salmonella typhi Ty2 in planar lipid bilayers

Native porins, from Salmonella typhi Ty2 outer membrane, and porins alkylated with pyridoxal phosphate (Plp) were studied in planar lipid bilayers. The conductance of bilayers exposed to native or chemically modified porins increases in discrete jumps. Conductance histograms for native porins displayed two major peaks at 1.7 and 6.7 nS (in 0.5 M KCl). On the other hand, Plp-treated porins exhibited a single major peak at 1 nS. The relation between bilayer conductance and native porin concentration was linear. However, this relation became logarithmic in the presence of modified porins. The results support the notion that alkaline reduction of S. typhi Ty2 porins with Plp dissociates porin channel trimers in a reversible fashion.

O-linked oligosaccharides of mouse egg ZP3 account for its sperm receptor activity

Previously, we reported that ZP3, one of three different glycoproteins present in the mouse egg's zona pellucida, serves as a sperm receptor. Furthermore, small glycopeptides derived from egg ZP3 retain full sperm receptor activity, suggesting a role for carbohydrate, rather than polypeptide chain in receptor function. Here, we report that removal of O-linked oligosaccharides from ZP3 destroys its sperm receptor activity, whereas removal of O-linked oligosaccharides has no effect. A specific size class of O-linked oligosaccharides, recovered following mild alkaline hydrolysis and reduction of ZP3, is shown to possess sperm receptor activity and to bind to sperm. The results presented strongly suggest that mouse sperm bind to eggs via O-linked oligosaccharides present on ZP3.

33] In Vitro synthesis of C15–C60 polyprenols in a cell-free system of Myxococcus fulvus and determination of chain length by high-performance liquid chromatography

Publisher Summary This chapter describes the biosynthetic formation of 14 C-labeled C 15 -C 60 polyprenols that may be used as standards, and their separation and chain length determination by High-Performance Liquid Chromatography (HPLC). Phosphorylated polyprenols may also be analyzed in this way when converted to their corresponding alcohols by alkaline phosphatase, hydrolysis, or reduction with LiAIH 4 , because handling of the free alcohols is much easier than of phosphorylated compounds. A number of graphs are obtained, demonstrating the homologous behavior of the compounds. Because of this and the overlapping separation patterns of the different graphs an unequivocal assignment of chain length can be made, although relatively few standards are available. The slight curvature of these graphs is explained by the nonideality of partition on reversed phase columns.

Combined removal of Cr, Cd, and Ni from wastes

AbstractResults of pilot‐plant tests on treating electroplating wastes by alkaline reduction, precipitation, and upflow filtration.

Isolation and characterization of a large, neurite-associated glycoconjugate from neuroblastoma cells.

A high molecular weight glycoconjugate has been isolated from neurite-producing neuronal tumor cells in culture and has been designated as I(0) based on its elution characteristics in gel filtration chromatography. This molecule cannot be found in a variety of nonneuronal cells. I(0) is found in the substratum-attached material or cell fraction of neurite-producing neuroblastoma cells, depending upon culture conditions. It is found in the substratum-bound fraction of B104 rat neuroblastoma cells during serum starvation and in the EGTA-detached cell fraction of B104 cells grown in chemically defined N2 medium. It occurs only in the cell fraction of the human neuroblastoma line Platt. Examination of behavioral variants of the B104 rat line further strengthens the association of I(0) with neurite production; the constitutive neurite-producing E(R)B9 variant contains I(0) while the non-neurite-producing E(R)A11 variant does not. I(0) is large, eluting in the void volume of sepharose-CL2B columns. Radioiodination of intact cells with lactoperoxidase shows I(0) to be a cell surface component. Metabolic radiolabeling studies show that it contains a high proportion of polysaccharide to protein, does not contain mannose, and is unsulfated. Alkaline borohydride reduction release two size classes of large polysaccharide chain. The alkaline reduction results, along with the mannose incorporation studies, show the presence of O-glycosidic linkages and few, if any, N-linkages. Resistance to nitrous acid deamination, insensitivity to glycosaminoglycan lyases, and the absence of sulfation, indicate that I(0) does not contain the glycosaminoglycans hyaluronic acid, chondroitin-, dermatan-, or heparin- sulfates. Affinity column chromatography reveals high binding affinity of I(0) to polyornithine and no binding to gelatin (collagen) or the glycosaminoglycans hyaluronate and heparin. These studies describe a unique high molecular weight glycoconjugate on the surface of neurite-producing neuroblastoma cell lines from two species.

Synthetic studies on terpenoids. Part 6. Synthesis of (±)-4,13-dimethyl-13-vinylpodocarp-8(14)-en-4α- and -4β-ols

The podocarpa-8,11,13-trien-4-one (6) was converted into the corresponding 4-methylpodocarpatrien-4-ol (7) with MeLi. Birch reduction of the alcohol (7) with Li–liquid NH3 yielded the podocarpenone (8) and the podocarpadienones (9) and (10). Condensation of (8) with ethyl cyanoacetate followed by conjugate addition of Me2CuLi afforded the 13-cyano(ethoxycarbonyl)methyl ester (12) which on alkaline hydrolysis, esterification and reduction with LiAIH4 afforded the corresponding 13-hydroxyethyl compound. Conversion of the latter, into its tosylate and elimination of the resulting tosylate by alumina gave 4,13-dimethyl-13-vinylpodocarp-8(14)-en-4β-ol (1). Dehydration of 13-methoxycarbonylmethyl-4,13-dimethylpodocarp-8(14)-en-4-ol furnished the 4-methylene compound (16) which on epoxidation and then reduction with LiAIH4 produced alcoholic material whose transformation to the tosylate followed by its elimination with alumina yielded the 4,13-dimethyl-13-vinylpodocarp-8(14)-en-4α-ol (2). Hydrolysis of compounds (1) and (2) with PtO2 in ethanol afforded the corresponding 13-ethyl compounds (3) and (4) respectively.

Chemical Linkage of Pine Polysaccharides to Lignin

Abstract Methylation analysis was used to investigate the bonds to lignin of the carbohydrates remaining after enzymatic hydrolysis and alkaline reduction of ball-milled loblolly pine wood and red pine compression wood. The carbohydrates exist as oligomeric chains with degrees of polymerization of 7–14. Approximately one sugar unit per oligomer chain is bonded to lignin. Bonding at C-6 of the hexose units is favored, and the arabinose is bonded exclusively at C-5. Galactan and arabinan are structurally of the 1–4 and 1–5 linked types respectively, characteristic of the so-called “pectic group substances.”

Preparation of a flocculant based on polyamides

AbstractA flocculant based on waste oligomers from the polycaprolactam production was synthesized by alkaline reduction with Ni/Al alloy. Both the reduced oligoamide and the sodium aluminate are precipitated by methanol. An oligomer, absorbed by a mineral carrier, is isolated as final product. The modified oligoamide was characterized by elemental analysis, IR spectra, and was shown to contain hydroxyl groups. The prepared flocculant may be used for the clarification of natural dilute (1%) clay suspensions and wastewaters. It accelerates the precipitation 35 times, giving a compact precipitate, which favors the decanting of the clarified liquid. The optimal concentration of the flocculant is 1.8 g/liter (corresponding to 0.06 g/liter oligomer).

Rapid diagnosis of carbon monoxide poisoning and quantitative determination of CO-Hb in blood

A quick diagnosis of carbon monoxide poisoning is a very difficult clinical problem. This paper describes a photometric method which permits a reliable diagnosis in a time of 7 to 10 minutes. Only 0.3 ml whole blood are necessary for the total procedure. Carboxyhemoglobin changes in absorption maximum slightly after reduction, whereas Oxyhemoglobin shows a significant change in extinction. Measurement of the hemolysate at the same wave length before and after alkaline reduction results in large differences in the extinctions (very large for Oxyhemoglobin and very small for Carboxyhemoglobin). Calibration curves are prepared by calibrating rest-extinction defined as: (formula: see text) to percent Carboxyhemoglobin. This curve can also be produced in mathematical way (as control e.g.). In practice there are the following steps: blood collecting (from fingerpad), hematolysis, first photometric measurement, reduction, second measurement, then reading the result from the calibration curve. The method can be automated (as demonstrated with the "Gilford System") providing the same results as with the manual method. All steps of quality control are given in full detail; as is the preparation of the hemolysates with various percentages of Carboxyhemoglobin.

Studies on the chemical nature of the lipidic J blood-group substance of cattle.

Total lipids extracted from J-positive cattle serum, erythrocytes or spleen exhibit J blood-group activity. The J subsance is concentrated in a lipid fraction obtained by column chromatography. Following mild alkaline hydrolysis or reduction with complex hydrides (LiAlH4, LiBH4), the J activity remains detectable in this lipid fraction even though all acyl ester groups have been destroyed as revealed by ester group determination. This disagrees with the suggestion that fatty acyl esters are essential for J activity. This was confirmed by experiments with a water-soluble J-active product prepared by ozone treatment of glycosphingolipids from bovine spleen. The results of these experiments are in favour of a glycosphingolipid containing anunusually lang oligosaccharide chain. Furthermore, it appears that the terminal moiety of the J determinant is not necessarily an N-acetyl galactosamine unit as suggested previously.

Combined gas chromatography-chemical ionization mass spectrometry of sphingolipids. I. Glucosyl sphingosine, ceramides and cerebrosides of the spleen in Gaucher's disease

Trimethylsilylated glucosyl sphingosine, ceramides and glucocerebrosides were analysed by combined gas chromatography (GC)-chemical ionization (CI) mass spectrometry. Isobutane, methane and ammonia were used as reactant gases for chemical ionization. Essentially the same fragment ions were detected in the spectra with the different reactant gases. Purified glucocerebrosides isolated from the spleen of a patient with Gaucher's disease were clearly separated into their 8 molecular species by gas chromatography. Three other minor components were detected by spectrometry, and these 11 molecular species of glucocerebrosides from the spleen of the patient with Gaucher's disease have been analysed. The ceramides obtained by periodate oxidation and then alkaline reduction of the glucocerebrosides were also separated into 11 molecular species by GC-CI mass spectrometry. Molecular weight could be determined using the major fragment ion of m/ e (M +−90) in the spectra of ceramides and cerebrosides. The chemical ionization method is useful for structural analyses and determination of the molecular species of sphingoglycolipids.

Modified curcumin and dianthrimide methods for the determination of boron in the presence of high levels of nitrate and organics

Modifications have been made to the curcumin and 1,1′-dianthrimide methods for boron analysis so that they may be used in the presence of up to 700 mg l −1 NO − 3 and 500 mg l −1 organic carbon. In the curcumin method, nitrate is removed by alkaline reduction using a slurry of aluminum powder. In the dianthrimide method, nitrate up to 1000 mg l −1 NO − 3 is removed in the dehydration-with-sulfuric-acid step; organics are most conveniently removed by treatment of the dehydrated sample with solid potassium persulfate. With dianthrimide, an automated procedure is used for the colour formation and measurement steps.

ChemInform Abstract: SYNTHESIS OF MONO‐ AND DI‐FUNCTIONAL SILICON CONTAINING DERIVATIVES OF LONG CHAIN FATTY ACID ESTERS

AbstractDie Monochlorsilane (I) addieren sich an die ungesättigten längerkettigen Fettsäureester (II) unter Bildung von (III).

Synthesis of mono‐ and di‐functional silicon containing derivatives of long chain fatty acid esters

AbstractThe addition products of some monochlorosilanes to methyl oleate and undecanoate were hydrolyzed and condensed to yield the corresponding difunctional disiloxanes, which were, inturn, transformed by alkaline hydrolysis or reduction to the corresponding diacids or diols. The addition products of dichlorosilanes gave on hydrolysis and condensation cyclosiloxanes but not linear polymers, as shown by mol wt data and IR spectra, while the corresponding addiiton products of trichlorosilances gave on similar treatment rubbery cross‐linked polymers. In contrast to the silanol intermediates, obtained on hydrolysis of methylchlorosilanes, which condensed easily to the corresponding disiloxanes, the corresponding bulkier phenyl chlorosilanes gave stable silanols which showed a lower tendency to condense. Some mono‐and di‐alkoxy, phenoxy, and acetoxy silyl derivatives of the long chain fatty acid esters also were synthesized.

Detection and identification of intermediate and final products of electrochemical reactions by means of the rotating ring-disc electrode method

This paper discusses the principles of identification of intermediate and final products of electrochemical reactions on the basis of kinetic information obtained by means of the rotating ring-disc electrode method. Most important information about the nature of the products may be obtained by investigating the relationship between the ring collection efficiency for the appropriate product and the electrode rotation speed, disc potential, concentration of the initial reactant in the solution, temperature and pH of the solution. This approach was used for identification of the products of nitroferrocene alkaline reduction. It was demonstrated that the cathodic reduction of this compound involves formation of anion-radicals of nitroferrocene, unstable ferrocenylhydroxylamine and amino-ferrocene.