Pure Oligomers Research Articles

Activated Bone Marrow-Derived Macrophages Eradicate Alzheimer's-Related Aβ42 Oligomers and Protect Synapses.

Impaired synaptic integrity and function due to accumulation of amyloid β-protein (Aβ42) oligomers is thought to be a major contributor to cognitive decline in Alzheimer's disease (AD). However, the exact role of Aβ42 oligomers in synaptotoxicity and the ability of peripheral innate immune cells to rescue synapses remain poorly understood due to the metastable nature of oligomers. Here, we utilized photo-induced cross-linking to stabilize pure oligomers and study their effects vs. fibrils on synapses and protection by Aβ-phagocytic macrophages. We found that cortical neurons were more susceptible to Aβ42 oligomers than fibrils, triggering additional neuritic arborization retraction, functional alterations (hyperactivity and spike waveform), and loss of VGluT1- and PSD95-excitatory synapses. Co-culturing neurons with bone marrow-derived macrophages protected synapses against Aβ42 fibrils; moreover, immune activation with glatiramer acetate (GA) conferred further protection against oligomers. Mechanisms involved increased Aβ42 removal by macrophages, amplified by GA stimulation: fibrils were largely cleared through intracellular CD36/EEA1+-early endosomal proteolysis, while oligomers were primarily removed via extracellular/MMP-9 enzymatic degradation. In vivo studies in GA-immunized or CD115+-monocyte-grafted APPSWE/PS1ΔE9-transgenic mice followed by pre- and postsynaptic analyses of entorhinal cortex and hippocampal substructures corroborated our in vitro findings of macrophage-mediated synaptic preservation. Together, our data demonstrate that activated macrophages effectively clear Aβ42 oligomers and rescue VGluT1/PSD95 synapses, providing rationale for harnessing macrophages to treat AD.

Rational protein design of Bacillus sp. MN chitosanase for altered substrate binding and production of specific chitosan oligomers

BackgroundPartially acetylated chito-oligosaccharides (paCOS) have a variety of potential applications in different fields, but to harness their benefits, pure paCOS or well-defined paCOS mixtures are essential. For example, if one could produce fully acetylated (A4) and fully deacetylated (D4) tetramers in abundance, all possible variants of tetrameric paCOS could be generated reliably from them. A promising approach for generating defined paCOS is by enzymatic depolymerization of chitosan polymers using chitosanases, since these enzymes’ subsite specificities directly influence the composition of the paCOS produced; however, enzymatic production of e.g. D4 is challenging because the substrate is generally hydrolyzed further by most chitosanases. To overcome this, chitosanases could potentially be engineered so that upon hydrolyzing chitosan, they are unable to hydrolyze certain substrates, leaving well-defined oligomers intact in the hydrolysate.ResultsFor this purpose, we performed rational protein engineering on the extensively studied GH 8 chitosanase CSN from Bacillus sp. MN. By specifically targeting residues with a predicted function in substrate binding, we created new muteins incapable of efficiently hydrolyzing the fully deacetylated tetramer D4, and we were able to demonstrate efficient large-scale production of D4 with an altered version of CSN. Furthermore, we were able to uncover differences in the substrate positioning and subsite specificities of the muteins, which result in altered paCOS mixtures produced from partially acetylated chitosan polymers, with possibly altered bioactivities.ConclusionThe value of protein engineering as a tool for the more efficient production of pure oligomers and potentially bioactive paCOS mixtures was demonstrated by the results and the suitability of specific muteins for the large-scale production of strictly defined, pure paCOS in a batch process was shown using the example of D4.

H-Bonded Duplexes based on a Phenylacetylene Backbone.

Complementary phenylacetylene oligomers equipped with phenol and phosphine oxide recognition sites form stable multiply H-bonded duplexes in toluene solution. Oligomers were prepared by Sonogashira coupling of diiodobenzene and bis-acetylene building blocks in the presence of monoacetylene chain terminators. The product mixtures were separated by reverse phase preparative high-pressure liquid chromatography to give a series of pure oligomers up to seven recognition units in length. Duplex formation between length complementary homo-oligomers was demonstrated by 31P NMR denaturation experiments using dimethyl sulfoxide as a competing H-bond acceptor. The denaturation experiments were used to determine the association constants for duplex formation, which increase by nearly 2 orders of magnitude for every phenol-phosphine oxide base-pair added. These experiments show that the phenylacetylene backbone supports formation of extended duplexes with multiple cooperative intermolecular H-bonding interactions, and together with previous studies on the mixed sequence phenylacetylene 2-mer, suggest that this supramolecular architecture is a promising candidate for the development of synthetic information molecules that parallel the properties of nucleic acids.

Controlling the aqueous solubility of PNIPAM with hydrophobic molecular units

The effect of co-polymerization of thermo-sensitive polymers with hydrophilic or hydrophobic co-monomers on the LCST and the structure of proximal water remains a fundamental and challenging problem. Here, we employ all-atom molecular dynamics simulations to investigate the aqueous solubility of a thermo-sensitive polymer, poly(N-isopropylacrylamide) (PNIPAM), a hydrophobic polymer polystyrene (PS) and block co-polymers of PNIPAM-co-PS. The simulations of pure oligomers of PNIPAM and PS and their co-polymers are conducted below and above the LCST of PNIPAM to elucidate the effect of increase in number of PS units in PNIPAM-co-PS co-polymers on the coil-to-globule transition of PNIPAM and structure of proximal water. Our simulations suggest that while the LCST of pure PNIPAM oligomers is strongly associated with the structural changes in the proximal water molecules present near PNIPAM, inclusion of PS units disrupts the interactions between PNIPAM and water and promotes the faster dehydration of PNIPAM chains above its LCST. This phenomenon is manifested by a coil-to-globule transition in PNIPAM chains present in PNIPAM-co-PS co-polymers at much shorter times as compared to the pure PNIPAM oligomers. Our results also reveal that with increase in the number of PS units in PNIPAM-co-PS co-polymers the coil-to-globule transition is accelerated.

Evaluating The Extent of Bio-Polyester Polymerization in Solid Wood by Thermogravimetric Analysis

In this study, the thermogravimetric analysis (TGA) technique has been applied to determine the extent of in situ polymerization achievable in solid wood on treating with bio-polyester oligomers. Low-molecular-weight oligomers of poly(lactic acid) (PLA), poly(glycolic acid) (PGA), poly(butylene succinate) (PBS), and poly(butylene adipate) (PBA) were impregnated and then thermally polymerized within solid wood to enhance the physical properties of wood. TGA revealed a similar degree of oligomer polymerization was achieved either with pure oligomers or within the wood structure. The influence of relatively acidic treatments, such as low-molecular-weight PGA oligomers, was observed to lead to degradation of the hemicellulose wood component. Polymerization of PLA and PGA oligomer treatments which penetrate the wood cell wall gave relatively lower wood thermal stability. Treatment and polymerization of lumen filling PBS and PBA oligomers contributed to increased wood thermal stability.

Solid-State Properties and Vibrational Circular Dichroism Spectroscopy in Solution of Hybrid Foldamers Stereoisomeric Mixtures.

Upon slow evaporation of a 1:1 diastereoisomeric mixture of Boc-(L-Phe-L-Oxd)2 -OBn (1; Boc=tert-butyloxycarbonyl; L-Oxd=trans-(4S,5R)-4-carboxy 5-methyloxazolidin-2-one, Bn=benzyloxycarbonyl) and Boc-L-Phe-L-Oxd-D-Phe-L-Oxd-OBn (2) in methyl tert-butyl ether, single crystals suitable for an X-ray diffraction study were obtained. In contrast, the two pure oligomers lead to the formation of amorphous solids under any crystallization conditions. The preferential conformation of both oligomers was fully elucidated in the solid phase and compared with the known conformation of Boc-(L-Phe-D-Oxd)2 -OBn (3). The preferred conformation of 1 ranges from a polyproline II (PPII) helix to β strands and we can gather that longer and more structured oligomers will form PPII helices. In contrast, compound 3 forms infinite antiparallel β-sheet structures; thus showing the strong effect of the reversal of the absolute configuration of the Oxd moieties on the secondary structure of these hybrid foldamers. The same outcome was retained in solution, as demonstrated by vibrational circular dichroism analysis. Finally, we have demonstrated that a 1:1 mixture of 1 and 2 leads to the formation of new materials with interesting properties that are missing from the two pure compounds, such as the tendency to form crystals, fibers, and globules, depending on the solvent.

Stereocomplexation between PLA-like substituted oligomers and the influence on the hydrolytic degradation

Stereocomplex formation was revealed between two PLA-like oligoester enantiomers. Two oligomeric enantiomers of each α-hydroxyisovaleric acid and lactic acid were synthesized and two part blends were prepared between all oligomer combinations. The blends were evaluated by DSC and XRD and a new crystal structure was detected in the case of d/l oligo-α-hydroxyisovaleric acid blend confirming stereocomplexation. Stereocomplexation was also verified for the D/L oligolactic acid blend. The two stereocomplexes had higher melting temperatures and increased enthalpy of melting compared to the pure oligomers. Hetero-stereocomplexation between different enantiomers of oligo(α-hydroxyisovaleric) acid and oligo(lactic acid) did not take place. The hydrolytic degradation rate was slower for the blends containing the more bulky and hydrophobic oligo(α-hydroxyisovaleric acid)s. Modifying PLA with this monomer could be used to reduce the degradation rate and at the same time modify the physical properties of PLA materials.

New ELISAs with high specificity for soluble oligomers of amyloid β-protein detect natural Aβ oligomers in human brain but not CSF

BackgroundSoluble oligomers of amyloid ß-protein (Aß) have been increasingly linked to synaptic dysfunction, tau alteration, and neuritic dystrophy in Alzheimer's disease (AD) and mouse models. There is a great need for assays that quantify Aß oligomers with high specificity and sensitivity. MethodsWe designed and validated two oligomer-specific (o-) enzyme-linked immunoassays (ELISAs) using either an Aß aggregate-selective monoclonal for capture and a monoclonal to the free N-terminus for detection, or the latter antibody for both capture and detection. ResultsThe o-ELISAs specifically quantified pure oligomers of synthetic Aß with sizes from dimers up to much larger assemblies and over a wide dynamic range of concentrations, whereas Aß monomers were undetectable. Natural Aß oligomers of similarly wide size and concentration ranges were measured in extracts of AD and control brains, revealing >1000-fold higher concentrations of Aß oligomers than monomers in the soluble fraction of AD cortex. The assays quantified the age-related rise in oligomers in hAPP transgenic mice. Unexpectedly, none of 90 human cerebrospinal fluid (CSF) samples gave a specific signal in either o-ELISA. ConclusionsThese new o-ELISAs with rigorously confirmed specificity can quantify oligomer burden in human and mouse brains for diagnostic and mechanistic studies and for AD biomarker development. However, our data raise the likelihood that the hydrophobicity of Aß oligomers makes them very low in number or absent in aqueous CSF.

Enhanced P-stereodependent stability of complexes formed by phosphorothioate oligonucleotides due to involvement of sulfur as strong hydrogen bond acceptor

Abstract The antisense or antigene properties are exerted not only by natural oligonucleotides, but also by their different analogs. Among them, phosphorothioate oligonucleotides (PS-DNA), in which the sugar-phosphate backbone is modified due to substitution of the sulfur atom for one of the nonbridging oxygens, are much more resistant toward nucleases and, simultaneously, maintain good hybridization properties. However, the substitution induces the P-chirality of dinucleoside phosphorothioate moiety and even short PS-DNA synthesized using standard chemical methods exist as a mixture of hundreds or thousands of diastereoisomers. Diastereomerically pure oligomers of [PS]-d(CG)4 and [PS]-d(GC)4 series, obtained using the oxathiaphospholane method, were investigated with respect to their ability to adopt the left-handed conformation at high sodium chloride concentration. Obtained data allow us to postulate the formation of a strong intramolecular hydrogen bond with anionic sulfur atom as an acceptor. Homopurine [All-RP-PS]-oligos, but not [All-SP-PS]-oligos, form with the RNA templates extremely stable triplex structures, so far not described in the literature. Most likely, the triplexes are stabilized by hydrogen bonds or water bridges with the participation of sulfur atoms of internucleotide linkage. Notably, target RNA molecules are "arrested" by properly designed [All-RP-PS]-DNA probes at submicromolar concentration, and as the result, they are not recognized by reverse transcriptase.

Characterization of synthetic and commercial trisiloxane surfactant materials

AbstractThe organosilicone surfactant Silwet L‐77® (L‐77), used as an agrochemical adjuvant, is a mixture comprised predominantly of [(CH3)3SiO]2(CH3)Si(CH2)3(OCH2CH2)nOCH3 oligomers (n = 3–16, average n ≈ 7.5). The commercially available L‐77 mixture was purified by reversed‐phase high‐performance liquid chromatography (HPLC) to obtain individual trisiloxane surfactant components. Pure oligomers (n = 3, 6 and 9) were also synthesized. Synthesis was achieved by hydrosilylation of monomeric ethoxylate monomethyl ether starting reagents. Pure hexa‐ and nona‐ethylene glycols were produced by condensation of smaller oligomers.Atmospheric‐pressure ionization mass spectrometry (MS) methods were used to characterize fully the commercial L‐77 product and synthesized or isolated components. The application of Fourier‐transform ion cyclotron resonance MS and online HPLC–electrospray ionization MS techniques to the analysis of this surfactant are described here. The application of these analytical techniques also enabled elucidation of the synthetic by‐products present in the commercial formulation.In addition, physico‐chemical properties specific to agrochemical uses, such as droplet spread areas on plant foliage and surface tension for the different oligomer solutions, are also reported. Copyright © 2004 John Wiley & Sons, Ltd.

New chemistry of oxophlorins (oxyporphyrins) and their π-radicals

Syntheses of novel 15-substituted-oxophlorins via the MacDonald condensation of diformyl-dipyrroketones and 5-substituted-dipyrromethanes are described. The electronic and steric features of the 15-substituent enable facile control over the oxidation potential of the oxophlorins. Introduction of an electron-withdrawing group efficiently minimizes the formation of oxophlorin π-radicals. Stabilization of neutral π radicals is promoted by hyperconjugation with a 15- tert-butyl group. A sterically induced stabilization of a novel non-aromatic tautomer of oxophlorin, the so-called “iso-oxophlorin” is demonstrated. These species exist also as 15-iso-oxophlorins upon complexation to divalent metals. Radical formation, enhanced by mild oxidants such as K 3FeCN 6, yielded pure oligomers and stereospecific supramolecular arrays by radical dimerizations taking place at the 10- and 10′-positions.

Mutual diffusion in aqueous solution of poly(ethyleneglycol) samples. Some comments on the effect of chain length and polydispersity

Mutual diffusion in aqueous solution of polydisperse poly(ethyleneglycol) samples (PEG) was analysed through Gouy interferometry with special attention to the molecular weight and concentration effects. For PEG samples with molecular weight higher than 1000 g mol-1 the diffusive trend on concentration is justified in thermodynamic terms. The diffusion properties of polydisperse samples and pure oligomers are compared. We emphasise that the refractive index gradient at the diffusion boundary in polydisperse PEG significantly deviates from a Gaussian distribution expected where the PEG monodisperse. The combination of mutual diffusion with literature intradiffusion and activity data allows testing of the Darken equation for each PEG sample. Because of the polydispersity the analysis of the velocity cross correlation coefficients (VCCs) or the distinct diffusion coefficients becomes critical.

Synthesis And Binding Study Of Phosphonate Analogues Of Pnas And Their Hybrids With Pnas

The preparation of monomers for the synthesis of phosphonate analogues of peptide nucleic acids containing the four natural nucleobases: thymine, cytosine, adenine and guanine, has been ccomplished. The monomers obtained were used for the automated online solid phase synthesis of pure phosphono-PNA oligomers as well as chimeras consisting of phosphono-PNA and PNA resudies. The hybridization properties of these oligonucleotide mimics to complementary DNA and RNA fragments were studied.

Modified Poly(2,6-dimethyl-1,4-phenylene ether)s Prepared by Redistribution

Redistribution of poly(2,6-dimethyl-1,4-phenylene ether) (PPE) with a phenolic compound yields a depolymerized polymer having the phenolic compound incorporated as the tail end. The molecular weight of the product is adjusted by use of the ratio of phenol over PPE repeating units. PPE oligomers and telechelics with a large variety of end groups, including reactive end groups, can be prepared in this way. Furthermore, block copolymers can be prepared by reacting a phenol terminated polymer in a redistribution with PPE. This is illustrated by the reaction of phenol terminated polystyrene with PPE, leading to polystyrene−PPE diblock copolymers. TMDPQ or copper/amine catalysts can be used as catalyst depending on the reacting phenolic compound, desired reaction time, and product purity. For a fast PPE redistribution with ortho-unsubstituted phenols, without any oxidation of the methyl substituents of the PPE chain, the use of TMDPQ is preferred. For the preparation of pure oligomers and block copolymers, a slow redistribution using copper catalysts in the absence of oxygen is preferred.

Uniform oligomers as reference materials for molecular weight determination of polymers

Uniform oligomers are special oligomers having no molecular weight distribution. They have been used as reference materials for polymer characterization, especially for accurate molecular weight determination in lower molecular weight ranges. These oligomers are used for different purposes. First, they are used in the calibration for chromatographic or spectroscopic data. Second, they are applied to identifying peaks appearing in chromatographies such as supercritical fluid chromatography (SFC) of oligomeric mixtures. Finally, they could give accurate instrumental or physicochemical constants used in polymer characterizations. The uniform oligomers have been prepared by using preparative chromatographies as separation methods. In some cases, step-wise syntheses have been applied together with the chromatographic methods. Recently, we prepared uniform oligostyrenes and oligo(oxyethylene)s whose molecular weights are greater than 1000 by using preparative size exclusion chromatography. More recently, using preparative SFC, some Japanese researchers have prepared high molecular weight and uniform oligomers such as oligo(methylmethacrylate)s. In this article, we briefly review recent progress mainly made by Japanese researchers in the field of preparation, analysis and applications of these pure oligomers.

Quantitative trace analysis of surfactant mixtures by reversed-phase high-performance liquid chromatography with refractometric detection

Reversed-phase partition liquid chromatography on an octyl column allowed the separation of complex non-ionic poly(ethylene oxide)-type (PEO) surfactant mixtures resulting from the condensation of ethylene oxide with saturated fatty alcohols. As these compounds have no chromophoric group, they were detected by differential refractometry. Accurate quantitation of each oligomer (C m E n) allowed the main characteristics of each non-ionic surfactant, i.e., the nature and percentage of the different alkyl chains (with m = number of carbons) and the average number of ethylene oxide units ( n ) to be obtained in one analysis. Preparative liquid chromatography was used to isolate pure oligomers with a higher degree of ethoxylation ( n =10, 11, 12, 14 and more) than the commercially available standards, in order to determine a wide range of refractometric response factors. It appeared that they are constant as a function of alkyl chain length (C 10-C 16 range) but that they vary significantly and non-linearly as a function of the degree of ethoxylation, n. It was found that neglecting the variation of response factors can result in a distortion of the average ethoxylation number and in an unsatisfactory quantitative analysis. This chromatographic method, involving a quantitative and reproducible trace enrichment procedure with liquid-solid extraction, allowed the analysis of very dilute PEO mixtures in water. The components of complex PEO mixtures in water were determined at concentrations as low as 0.5 mg 1 −1, without any distortion of the distribution, the detection limit being 0.25 μg 1 −1 for the less abundant oligomers.

Spectroscopic and electrical investigations of undoped and doped α-oligothiophene thin films

Chemically pure α-oligothiophene ( αnT, n = 4–8) were used in this study to prepare oligomeric thin film structures as model systems for polythiophene. The pure materials were characterized under UHV conditions with X-ray photoelectron spectroscopy (XPS), ultraviolet photoelectron spectroscopy (UPS) and high resolution electron-energy-loss spectroscopy (HREELS). The change of the electronic structure with increasing number of rings could be evaluated quantitatively. Furthermore, the solid-state doping effects of the oligomers α5T and α7T were investigated in situ for systematically varied concentrations (0–70%) of FeCl 3 as dopant. With increasing dopant concentration two additional loss structures were observed at energy losses below the bandgap transitions of the pure oligomers. Finally, four-point measurements were performed of co-sublimed thin films of p-doped α6T under UHV conditions. A conductivity of δ = 3·10 −1 (Ω cm) −1 was determined for the p-doped material at T = 300 K.

1,N6-etheno deoxy and ribo adenosine and 3,N4-etheno deoxy and ribo cytidine phosphoramidites. Strongly fluorescent structures for selective introduction in defined sequence DNA and RNA molecules.

Synthesis of 1,N6-etheno-2'-deoxyadenosine, 3,N4-etheno-2'-deoxycytidine, and further chemistry on both deoxy and ribo series etheno nucleosides produces the corresponding phosphoramidites. These novel phosphoramidites are introduced selectively, quantitatively, and at specific positions at single or multiple sites into DNA or RNA sequences. The purification and chemistry involved in the synthesis of these products has been optimized to achieve the purity in excess of 99%. The resulting phosphoramidites were tested for their ability to couple and produce poly deoxy and ribonucleotides by solid phase chemistry. The coupling efficiency achieved was greater than 99% per step. Due to the instability of these etheno compounds in acidic and basic medium, various criteria to obtain pure oligomers have been established. The selective introduction of these fluorescent nucleosides into defined sequence DNA and RNA molecule will greatly facilitate the structure-function studies of various RNAs, protein-RNA structures, and DNA-RNA based diagnostics applications. The characteristic and high fluorescent intensity (detection below 1 x 10(-9) M for adenosine sites and below 1 x 10(-7) M for cytidine sites) is particularly suited for the biochemical and biological research and product development applications. The usefulness of these etheno containing modified sequences as sequencing and amplification primers is demonstrated by their full participation in polymerase chain reaction experiments.

Preparation of trimers and tetramers of mixed sequence oligodeoxynucleoside methylphosphonates and assignment of configurations at the chiral phosphorus

Synthesis of stereoregular DNA methylphosphonates has been accomplished for homo-oligomers, but remains a formidable problem for oligomers of a defined antisense target sequence. In this work, four trimer and tetramer deoxynucleoside methylphosphonates of mixed sequence (dACA, dCCAA, dAGGG, and dGCAT) were prepared by block coupling of diastereomerically pure dimers with either monomers or other diastereomerically pure dimers. These oligomers were separated chromatographically into individual diastereomers, and the configurations of the chiral methylphosphonate linkages were assigned. Three types of methods were used to assign configuration of a new methylphosphonate linkage: preparation of the same diastereomer through multiple synthetic pathways, base hydrolysis, and acid hydrolysis. Hydrolysis of the diastereomerically pure oligomers into component dimers and monomers was followed by chromatographic comparison with control dimers of known configuration. In all cases studied, oligomers with R configurations displayed faster elution from silica gel than did oligomers with the respective S configuration. NMR spectra of individual diastereomers of dACA were studied, revealing characteristic differences in chemical shifts which may prove useful in configurational assignments of longer oligomers. Thus, these data provide a methodological basis for synthesis and configurational assignment of longer methylphosphonate oligomers to use as antisense probes.

Preparative isolation of oligomers with a terminal anhydrosugar unit by thermal degradation of chitin and cellulose

Chitin and cellulose have been depolymerized by thermal degradation in a high boiling and inert solvent to give water-soluble oligomers with a terminal anhydrosugar unit. Pure oligomers 1 and 3 up to the pentamers have been obtained by preparative gel permeation chromatography, and characterized by 1H-NMR, 13C-NMR and fast atom bombardment-mass spectrometry.