A study of complex protein mixtures obtained from biological samples by MS demands proper purification and separation technique. The method of divergent flow isoelectric focusing (DF IEF) promises improvement of sample preparation in proteomic studies. DF IEF was carried out in a separation channel with increasing width. The channel was cut out from a polyester nonwoven web. DC voltage (800 V) was brought to two pairs of electrodes situated on the channel sides. Amphoteric compounds, including proteins, drift through the channel carried by flow (18-25 ml/h) in streamlines given by their isoelectric points. The pH gradient (3-10) and its stability during analysis have been monitored with colored low-molecular mass pI markers. Separated fractions were collected in ten microvials and further analyzed by MS. The suggested method was used for separation and purification of crude protein extract from barley grain, malt, and beer. Collected fractions of separated proteins were characterized by MALDI-MS. Desalting during IEF enhanced significantly the quality of mass spectra. It also simplified monitoring of post-translational modifications and protein changes occurring during malting and brewing. Results have shown the real potential of the suggested DF IEF device lay-out as an efficient preparative tool for separation and purification of complex protein mixtures for further analyses.
- MeSH
- Time Factors MeSH
- Equipment Design MeSH
- Electrodes MeSH
- Financing, Organized MeSH
- Isoelectric Focusing methods instrumentation MeSH
- Hydrogen-Ion Concentration MeSH
- Analytic Sample Preparation Methods methods MeSH
- Molecular Weight MeSH
- Proteins analysis MeSH
- Proteomics MeSH
- Reproducibility of Results MeSH
- Sensitivity and Specificity MeSH
- Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization methods instrumentation MeSH
This paper describes laboratory preparation, characterization and antibacterial activity testing of ZnO/kaoline composites. ZnO/kaoline composites with 50 wt.% of ZnO were laboratory prepared, dried at 105 °C and calcined at 500 °C. XRPD analysis revealed that thermal treatment caused the phase transformation of Zn containing precursor into ZnO. Scanning and transmission electron microscopy techniques were used for characterization of morphology of the prepared samples. A standard microdilution test was used for evaluation of antibacterial activity using four common human pathogens (Staphylococcus aureus, Escherichia coli, Enterococcus faecalis and Pseudomonas aeruginosa). Daylight was used for induction photocatalytically based antibacterial activity. Second possible explanation of antibacterial activity of ZnO/kaoline could be the presence of biologically available forms of zinc. During the antibacterial activity assays the ZnO/kaoline composites exhibited antibacterial activity, where differences in an onset of the antibacterial activity and activity against bacterial strains were observed. The highest antibacterial activity was observed against S. aureus, where the lowest value of minimum inhibitory concentration was determined equal to 0.41 mg/ml.
- MeSH
- Anti-Bacterial Agents chemistry pharmacology MeSH
- X-Ray Diffraction MeSH
- Enterococcus faecalis drug effects MeSH
- Escherichia coli drug effects MeSH
- Kaolin chemistry MeSH
- Catalysis MeSH
- Nanocomposites chemistry ultrastructure MeSH
- Zinc Oxide chemistry MeSH
- Pseudomonas aeruginosa drug effects MeSH
- Staphylococcus aureus drug effects MeSH
- Light MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
Preparation of nanocomposite kaolinite/TiO(2), using hydrolysis of titanyl sulfate in the presence of kaolin was addressed. A variable (kaolin)/(titanyl sulfate) ratio has been used in order to achieve the desired TiO(2) content in prepared nanocomposites. Calcination of the composites at 600 °C led to the transformation of the kaolinite to metakaolinite and to origination of metakaolinite/TiO(2) composites. The prepared samples were investigated using X-ray fluorescence spectroscopy, X-ray powder diffraction, Fourier transform infrared spectroscopy, scanning electron microscopy, thermogravimetry and diffuse reflectance spectroscopy in the UV-VIS region. Structural ordering of TiO(2) on the kaolinite particle surface was modeled using empirical force field atomistic simulations in the Material Studio modeling environment. Photodegradation activity of the composites prepared was evaluated by the discoloration of Acid Orange 7 aqueous solution.
- MeSH
- Azo Compounds chemistry radiation effects MeSH
- Coloring Agents chemistry radiation effects MeSH
- Benzenesulfonates chemistry radiation effects MeSH
- Photolysis MeSH
- Kaolin chemistry MeSH
- Environmental Pollutants chemistry radiation effects MeSH
- Nanocomposites chemistry MeSH
- Computer Simulation MeSH
- Surface Properties MeSH
- Industrial Waste prevention & control MeSH
- Titanium chemistry MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
This study aimed to prepare a bioactive acrylic material by adding different types of glasses. Commercially available polymerized acrylic resin was mixed with 10% of four different types of glasses in the powder form and cured. Flexural strength, sorption, and solubility of the samples were tested according to ISO 20795-1:2013. The total number of samples used in the tests were 60. The materials were placed in artificial saliva of pH 4 and 7, and elution was performed for 0, 1, 28, and 42 days. The collected samples were analyzed using inductively coupled plasma atomic emission spectrometry to detect Ca, P, and Si ions and using ion chromatography to detect F ions. The materials obtained after modification with glasses showed lower compressive strength compared with pure polymethyl methacrylate but met the standard requirements. Two glass types showed higher solubility values compared with the value defined by the ISO standard. Biomin C and S53P4 released Ca, P, and Si ions, respectively, after 42 days in artificial saliva. Acrylic resins modified with 10% Biomin C and S53P4 glasses can be a valuable source of Ca and P ions under acid conditions for 28 and 42 days.
- MeSH
- Acrylic Resins * chemistry MeSH
- Saliva, Artificial MeSH
- Polymethyl Methacrylate * chemistry MeSH
- Powders MeSH
- Materials Testing MeSH
- Publication type
- Journal Article MeSH
Thin layers of chitosan (positively charged)/sodium hyaluronate (negatively charged)/nonwoven fabrics were constructed by polyelectrolyte multilayer pad-dry-cure technique. Pure chitosan (CS) was isolated from shrimp shell and immobilized onto nonwoven fabrics (NWFs) using citric acid (CTA) as cross linker and solvent agents through a pad-dry-cure method. The prepared thin layer of chitosan citrate/nonwoven fabrics (CSCTA/NWFs) were consequently impregnated with hyaluronan (CSCTA/HA/NWFs) in the second path through a pad-dry-cure method. Chitosan/hyaluronan/nonwoven fabrics wound dressing was characterized by different techniques such as FTIR-ATR, TGA and SEM. The antibacterial activity and the cytotoxicity of the dressing sheets were evaluated against Escherichia coli (E. coli) and Streptococcus aureus (S. aureus), mouse fibroblast (NIH-3T3) and keratinocytes (HaCaT) cell lines, respectively. The cell-fabrics interaction was also investigated using fluorescence microscope, based on live/dead staining assay of 3T3 cells. The healing properties of the new wound dressing were evaluated and compared with the control sample.
- MeSH
- Anti-Bacterial Agents chemistry therapeutic use MeSH
- NIH 3T3 Cells MeSH
- Chitosan chemistry therapeutic use MeSH
- Escherichia coli drug effects pathogenicity MeSH
- Wound Healing drug effects MeSH
- Hyaluronic Acid chemistry therapeutic use MeSH
- Humans MeSH
- Mice MeSH
- Bandages microbiology MeSH
- Staphylococcus aureus drug effects pathogenicity MeSH
- Textiles microbiology MeSH
- Animals MeSH
- Check Tag
- Humans MeSH
- Mice MeSH
- Animals MeSH
- Publication type
- Journal Article MeSH
Rapid and correct production of generic solid dosage forms requires a large amount of analytical data and conclusions. Modern analytical techniques have a good resolution and accuracy and allow obtaining a lot of information about the original product. Scanning electron microscopy (SEM) is used for observation and assessing individual layers, core and surface of solid dosage forms. Fourier transform infrared (FTIR) spectroscopy mapping allows determining the distribution and characterization of individual components in a solid dosage form. However, the samples prepared by common way, using scalpel or tablet splitter, are not good enough. It was the reason for development of a new and better method of sample preparation, which uses microtome. Well-prepared samples analyzed by SEM and FTIR mapping allow to determine a solid dosage form formulation, excipient content and distribution of excipient and active pharmaceutical ingredient.
External application of seabuckthorn oil (Hippophae rhamnoides L.) is difficult due to its liquid state in spite of its benefits for damaged skin. In order to overcome this inadequacy the semisolid emulsion with seabuckthorn oil was prepared. Previous research showed that this emulsion possessing an enhanced structure with liquid crystals showed a higher wound healing potential than seabuckthorn oil. The aim of this investigation was to characterize suitability of this emulsion for topical use. The emulsion was prepared by combining emulsifiers that form liquid crystals. Two different quantities of seabuckthorn oil were incorporated. Samples were prepared with 10% and 40% of seabuckthorn oil. Organoleptic characteristics were estimated visually and by smearing samples on a thin glass plate. Type of emulsion was determined by a conductometric method, while a pH value of the emulsion was measured by a pH meter. Samples of seabuckthorn emulsion were orange, semisolid, shiny, easily spreadable on skin, and the smear on the glass plate was homogeneous. There was an absence of smell and the emulsion could be rinsed by water after the application on skin, which is a desired characteristic of oil/water emulsions. Results of an electrical conductivity confirmed that an outer phase is water. Samples possesed an acceptable pH value for an external topical use. This research confirmed that constituents and a method used were suitable for preparing semisolid emulsion with seabuckthorn oil. Organoleptic properties, a pH value and a type of obtained emuslion appear to be adequate for topical use.
- MeSH
- Administration, Topical MeSH
- Chemistry Techniques, Analytical methods MeSH
- Electric Conductivity MeSH
- Emulsions chemical synthesis MeSH
- Financing, Organized MeSH
- Phytotherapy MeSH
- Hippophae MeSH
- Wound Healing MeSH
- Hydrogen-Ion Concentration MeSH
- Conductometry statistics & numerical data MeSH
- Rhamnus MeSH
- Fruit chemistry MeSH
- Plant Preparations chemistry therapeutic use MeSH
- Research MeSH
The binding ability of a drug to plasma proteins influences the pharmacokinetics of a drug. As a result, it is a very important issue in new drug development. In this study, affinity capillary electrophoresis, capillary electrophoresis with frontal analysis, and Hummel Dreyer methods with internal and external calibration were used to study the affinity between bovine serum albumin and salicylic acid. The binding constant was measured by all these approaches including the equilibrium dialysis, which is considered to be a reference method. The comparison of results and other considerations showed the best electrophoretic approach to be capillary electrophoresis-frontal analysis, which is characterized by the high sample throughput with the possibility of automation, very small quantities of biomacromolecules, simplicity, and a short analysis time. The mechanism of complex formation was then examined by capillary electrophoresis with frontal analysis. The binding parameters were determined and the corresponding thermodynamic parameters such as Gibbs free energy ΔG(0), enthalpy ΔH(0), and entropy changes ΔS(0) at various temperatures were calculated. The results showed that the binding of bovine serum albumin and salicylic acid was spontaneous, and that hydrogen bonding and van der Waals forces played a major role in the formation of the complex.
Cíle studie: Cílem studie bylo vytvořit protokol pro zpracování vzorků moči pro získání aminokyselinových profilů pomocí iontově výměnné chromotografie s detekcí ve viditelném spektru (IEC/Vis) na základě tvorby komplexů aminokyselin s ninhydrinem a následné zkrácení času analýzy pro klinicky zajímavé molekuly (sarkosin či taurin), které mohou byt využity jako potenciální nádorové biomarkery. Typ studie: Metodická Materiál a metody: Vzorky moči, odebrané pacientům s diagnostikovaným karcinomem prostaty (n = 500) byly shromažďovány po dobu 1 roku a uskladňovány při teplotě -80°C. Pro vlastní analýzu aminokyselinových profilů byly vzorky připraveny kyselou hydrolýzou v mikrovlnném reaktoru. Za optimalizovaných podmínek (80 W, 120°C, 25 bar, 105 min) bylo 500 μl vzorku, smíchaného s 500 μl 35% kyseliny chlorovodíkové zhydrolyzováno na výsledný produkt, který byl následně ředěn pufrem sodíkového cyklu (0,2 mol/l NaCl, 60 mmol/l C6H807, 1,5 mmol/l N3Na a 0,4 % S(CH2CH2OH)2). Po centrifugaci (25 000 g při 4°C, 20 min) byly vzorky neutralizovány (0,6 mol/l NaOH) a analyzovány iontově-výměnnou kapalinovou chromatografií s postkolonovou derivatizací ninhydrinem využívající optickou detekci při vlnových délkách λ = 440 a 570 nm. Pro detekci sarkosinu a taurinu byly vzorky připraveny odpařením 250 μl vzorku na dusíkové odparce (40 min, teplota dusíku 60°C, tlak 1 bar) a následně resuspendovány pufrem sodíkového cyklu (250 μl). Výsledky a závěr: Analýzou aminokyselinových profilů (n = 500) bylo zjištěno, že lze docílit získání důležitých informací z neinvazivně odebraných vzorků moči. Použití dusíkové odparky redukuje manipulaci se vzorkem a zlepšuje opakovatelnost analýzy. Zkrácená analýza sarkosinu a taurinu ze vzorků odpařených na dusíkové odparce může sloužit jako metoda pro citlivou a nízkonákladovou analýzu klinických vzorků.
Objective: The aim of our study was to prepare a protocol for pre-treatment of clinical urinary samples, subsequently employed for acquirement of amino acid profiles via ion exchange chromatography with detection in visible spectrum (IEC/ Vis). Further for clinically interesting biomolecules as sarcosine and taurine the optimization was carried out to shorten the analysis time. Design: Methodological Material and methods: Urine specimens from patients diagnosed with prostate cancer (n = 500) were collected within 1 year and stored in -80°C. Samples were processed for analysis of amino acid profiles through acidic hydrolysis in a microwave reactor. Using optimized conditions (80 W, 120°C, 25 bar, 105 min), 500 μL sample, mixed with 500 μL of 35% hydrochloric acid hydrolyzed. That resulted in product, which was subsequently diluted in buffer of sodium cycle (0.2 mol/L NaCl, 60 C6H807 mmol/L, 1.5 mmol/L and 0.4% N3Na S(CH2CH2OH)2). After centrifugation (25000 g at 4°C, 20 min), the samples were neutralized (0.6 mol/l NaOH) and analyzed using ion-exchange liquid chromatography with post-column derivatization with ninhydrin, using the detection wavelength λ = 440 and 570 nm (IEC/Vis). For analysis of sarcosine and taurine, the samples were prepared by evaporation of 250 μL of sample employing a nitrogen evaporator (40 min, temperature 60°C. Nitrogen pressure of 1 bar) and resuspended with buffer of sodium cycle (250 μL). Results and conclusion: The analyses of amino acid profiles offer interesting clinical information not only in non-invasively collected urinary samples, but also in other organic matrices. Employment of nitrogen evaporator for sample pre-treatment leads to reduction of manipulation with sample and its combination with shortened analyses times of sarcosine and taurine may serve as a sensitive and low-cost method for analysis of clinical specimens.
- MeSH
- Early Diagnosis * MeSH
- Chromatography, Ion Exchange * methods instrumentation utilization MeSH
- Humans MeSH
- Biomarkers, Tumor analysis MeSH
- Prostatic Neoplasms * diagnosis MeSH
- Oxidative Stress MeSH
- Mass Screening economics utilization MeSH
- Prostate-Specific Antigen * analysis MeSH
- Sarcosine * analysis urine MeSH
- Spectrophotometry, Ultraviolet methods instrumentation utilization MeSH
- Taurine * analysis urine MeSH
- Check Tag
- Humans MeSH
- Publication type
- Research Support, Non-U.S. Gov't MeSH
Determination of an antihyperlipidemic drug simvastatin (SIM) was carried out using a carbon paste electrode bulk-modified with multiwalled carbon nanotubes (MWCNT-CPE). Scanning electrochemical microscopy (SECM), scanning electron microscopy (SEM), and atomic force microscopy (AFM) were used for the characterization of the prepared electrodes. Different electrodes were prepared varying in mass percentage of MWCNTs to find out the optimum amount of MWCNTs in the paste. The MWCNT-CPE in which the mass percentage of MWCNTs was 25% (w/w) was found as the optimum. Then, the prepared electrode was used in a mechanistic study and sensitive assay of SIM in pharmaceutical dosage form and a spiked human plasma sample using differential pulse voltammetry (DPV). The prepared electrode shows better sensitivity compared to the bare carbon paste and glassy carbon electrode (GCE). The detection limit and the limit of quantification were calculated to be 2.4 × 10-7 and 8 × 10-7, respectively. The reproducibility of the electrode was confirmed by the low value of the relative standard deviation (RSD% = 4.8%) when eight measurements of the same sample were carried out. Determination of SIM in pharmaceutical dosage form was successfully performed with a bias of 0.3% and relative recovery rate of 99.7%. Furthermore, the human plasma as a more complicated matrix was spiked with a known concentration of SIM and the spiking recovery rate was determined with the developed method to be 99.5%.
- MeSH
- Electrodes * MeSH
- Humans MeSH
- Microscopy, Electron, Scanning MeSH
- Nanotubes, Carbon chemistry ultrastructure MeSH
- Simvastatin chemistry MeSH
- Carbon chemistry MeSH
- Check Tag
- Humans MeSH
- Publication type
- Journal Article MeSH
