2D LC Separation and Determination of Bradykinin in Rat Muscle Tissue Dialysate with On-Line SPE-HILIC-SPE-RP-MS

A 2D liquid chromatography (LC) system using hydrophilic interaction chromatography (HILIC) and reversed phase columns has been employed for comprehensive (LC × LC) separation of rat muscle tissue micro-dialysate. Incorporation of an on-line reverse-phase solid phase extraction (SPE) enrichment column in front of the first dimension enabled aqueous samples with high salt concentrations to be injected directly without compromising the chromatographic performance of the HILIC column. Since the SPE enrichment column allowed injection of large sample volumes (e.g. 450 μL), a capillary HILIC column (inner diameter 0.3 mm) could be employed instead of a larger column which is often used in the first dimension to load sufficient amounts of sample. The two chromatographic dimensions were connected using a column selector system with 18, 1.0 mm I.D. C₁₈ “transition” SPE columns. A PLRP C₁₈ column was used in the second dimension. The 2D LC system’s performance was evaluated with a tryptic digest mixture of three model proteins. Good trapping accuracy (HILIC→transition SPE→RP recovery >95%) and repeatability (within-and between day retention time RSDs of first and second dimension chromatography >1%) was achieved. A dialysis sample of rat muscle tissue was separated with the 2D system, revealing complexity and large differences in concentrations of the various compounds present, factors which could potentially interfere with the quantification and monitoring of two target analytes, arg-bradykinin and bradykinin. Subsequently, “Heart-cut” 2D LC-electrospray–mass spectrometry (ESI–MS) with post-column on-line standard injection was employed to monitor arg-bradykinin and bradykinin levels as a function of various muscle conditions. The method’s quantification precision was RSD = 3.4% for bradykinin.     

Collagen-binding peptide interaction with retinal tissue surfaces

One of the current challenges in treating age-related macular degeneration (AMD) is the surface modification of the retinal Bruch membrane. In this study, the collagen fibers of the inner collagenous zone of the Bruch membrane were identified as type I and type III. Subsequently, the adsorption of a collagen-binding peptide onto the inner collagenous zone surface was investigated. The collagen-binding peptide was able to bind specifically to the collagen fibers while maintaining the biological activity of the N-terminus biotin tag. These results indicate that the collagen-binding peptide may be used as an anchor to immobilize bioactive molecules on the inner collagenous zone surface of the Bruch membrane.     

Mixed adlayer of alkanethiol and peptide on GaAs(100): quantitative characterization by X-ray photoelectron spectroscopy

Homogeneous and mixed adlayers composed of an alkanethiol (1-octadecanethiol, ODT) and a peptide (CGISYGRKKRRQRRR) on GaAs(100) were formed in two different solvent systems: phosphate-buffered saline (PBS) and N,N-dimethylformamide (DMF). The chemical composition of each adlayer was characterized by X-ray photoelectron spectroscopy (XPS). The data showed that the makeup of the adlayer and its stability largely depends on the solvent used. Angle-resolved XPS also revealed that the adlayer thickness and tilt angles were different from values obtained from ellipsometry measurements and vastly varied between the two solvents used. The coverage data extracted from the XPS measurements indicated that homogeneous adlayers of peptide in PBS buffer form a multilayered film. Homogeneous alkanethiol adlayers exhibited monolayer coverage under all solvent treatments. Coadsorbed layers containing both alkanethiol and peptide have fractional monolayer coverage in both solvents.     

Circular dichroism study of enzymatic manipulation on magnetic and metallic DNA template nanowires

Circular dichroism spectroscopy (CD) was used to examine the mechanism of endonuclease clipping and ligation of the DNA template nanowires. The biomolecular manipulation of the DNA template is compared for both metallic (Au) and magnetic (Fe₂O₃ and CoFe₂O₄) nanowires. The dependence of nanoparticle (NP) concentration on enzymatic clipping and DNA ligation was studied, in addition to performing absorbance and thermal melting experiments. Low-NP concentration preserved and digested the DNA template structure. Yet, at higher NP concentrations, the DNA template began to denature before enzyme addition. It was also observed that ligation of the digested DNA occurred more efficiently at low-NP concentrations. These results provide significant information on structural alteration and biorecognition effectiveness of the DNA template after enzymatic manipulation.     

A convenient room temperature polycondensation toward hyperbranched AB2‐type all‐aromatic polyesters with phenol terminal groups

As a convenient alternative to the classical melt polycondensation the one‐pot solution polycondensation of suitable AB₂ monomers under mild conditions has been successfully adapted to hyperbranched all‐aromatic polyester with phenol terminal groups. The polymerization was performed in solution at room temperature directly using commercially available 3,5‐dihydroxybenzoic acid as monomer and 4‐(dimethylamino) pyridinium 4‐tosylate as catalyst to suppress the formation of N‐acylurea. Different carbodiimides as coupling agents were investigated to find the optimal esterification conditions. The polymers have been characterized extensively and were compared with their well‐known analogs synthesized in melt. The characterization was carried out by NMR spectroscopy, size exclusion chromatography, and asymmetric flow‐field flow fractionation as an alternative separation technique for multifunctional polymers. © 2009 Wiley Periodicals, Inc. J Polym Sci Part A: Polym Chem 47: 5158–5168, 2009     

Amphiphilic silicones prepared from branched PEO‐silanes with siloxane tethers

Amphiphilic silicones were prepared by the covalent incorporation of branched polyethylene oxide (PEO) via a siloxane tether. This was achieved by using six novel branched PEO‐silanes with varying siloxane tether lengths and PEO molecular weight (M_n). Each PEO‐silane was crosslinked via acid‐catalyzed sol–gel condensation with α,ω‐bis(Si‐OH)polydimethylsiloxane (PDMS) (M_n = 3000 g/mol) to yield six amphiphilic silicone films. Film surface hydrophilicity increased with siloxane tether length, particularly after exposure to an aqueous environment, indicating that the PEO segments were more readily driven to the surface. This effect was more pronounced for films prepared with PEO‐silanes containing lower M_n PEO segments. AFM was used to study surface reconstruction of films upon exposure to an aqueous environment. Adsorption of bovine serum albumin (BSA) and human fibrinogen (HF) proteins decreased with siloxane tether length, particularly after first exposing films to an aqueous environment. For a given siloxane tether length, relatively less BSA adsorbed onto films prepared with PEO‐silanes with lower M_n PEO segments whereas less HF adsorbed onto films prepared with PEO‐silanes with higher M_n PEO segments. © 2010 Wiley Periodicals, Inc. J Polym Sci Part A: Polym Chem 48: 4108–4119, 2010     

Separation of linear and star-shaped polystyrenes by phase distribution chromatography

The phase distribution chromatographic technique was optimized and applied for the separation of linear and star-shaped polystyrene (PS). For this purpose non-crosslinked, ultra high molecular weight PS coated on different supporting materials was used. The stability of the coating under chromatographic conditions was tested by thermo gravimetric analysis and microscopic techniques. The modification of different column packing materials was tested. Separation according to branching was indicated for different molecular weights of linear and star-shaped PS. The resolution of the separation was improved by changing the density of the stationary phase and the temperature. The separation results were supported by cloud point measurements and the determination of the critical conditions for linear and star-polymers at the same molecular weight.     

Chemical and technological characterization of medieval glass bracelets from South-East Bulgaria

The present paper deals with chemical and physicochemical characterization of seven glass fragments of medieval glass bracelets from South-East Bulgaria. Samples were investigated by Fourier-transform infrared spectroscopy (FT-IR), transmission electron microscopy (TEM) and differential thermal analysis (DTA). Gravimetric chemical analysis was used for silica assessment. Flame photometry was applied to determine sodium and potassium content while aluminium, calcium and magnesium were determined by flame atomic absorption spectrometry after dissolution using a mixture of acids. All investigated artefacts are soda-lime-silica glasses and when produced two different recipe norms were used. Important technological parameters of glass manufacturing were determined and discussed. This investigation throws light on the technological development and production of glass during medieval times.      

Bulgarian natural diatomites: modification and characterization

Natural Bulgarian diatomite modified by oxidation with sulfuric acid and H₂O₂ or by coating with manganese oxide was characterized considering its chemical composition, surface area, pore volume, and structure. Modified diatomites displayed larger surface area and pore volumes in comparison with untreated natural diatomite, which favored their sorption behavior. Sorption properties of diatomites towards Fe³⁺, Pb²⁺, Cu²⁺, Cd²⁺, Mn²⁺, Ni²⁺, Co²⁺, Cr³⁺, Pd²⁺, Ca²⁺, and Mg²⁺ were investigated and their sorption capacities were determined. Sorption properties of manganese oxide-modified diatomite were superior to those of diatomite modified by oxidation. Owing to its high sorption capacity towards Co²⁺, Ni²⁺, Pb²⁺, Cr³⁺, Fe²⁺, Cu²⁺, and Cd²⁺, the manganese oxide-modified diatomite is a promising low-cost sorbent for selective removal of milligram amounts of these toxic metal ions from contaminated water.