Ewa Kłodzińska

Effect of zeta potential value on bacterial behavior during electrophoretic separation

The aggregation and/or adhesion of bacterial cells is a serious disadvantage of electrophoretic separations. In this study, physicochemical surface characteristics of bacteria were measured to establish their role in bacterial adhesion and aggregation on the basis of electrophoretic behavior of different clinical strains of Gram‐positive Staphylococcus aureus and Gram‐negative Escherichia coli bacteria. The number and the shape of peaks obtained on the electropherograms were connected with the zeta potential measurements and in‐line microscope observation using specially designed CE fluorescence stereomicroscope setup. These results suggest that the lower the zeta potential, the higher the number of smaller peaks detected. The direct microscopic observation of electrophoretic movement proved the presence of many small aggregates originating from individual or clustered bacterial cells. On the other hand, lower zeta potential was also observed for dead bacterial cells, which suggested that some of the peaks can be attributed to viable cells while the other to the dead ones.

Identification of volatile organic compounds secreted from cancer tissues and bacterial cultures

The early cancer diagnosis increases the possibility of total recovery. The infection of Helicobacter pylori is associated with gastric cancer, the second most common cancer in the world. The determination of volatile organic compounds (VOCs) excreted by stomach tissue and bacteria culture has been investigated. Solid-phase microextraction (SPME) was used for preconcentration and the determination was accomplished by gas chromatography coupled with mass spectrometry (GC/MS). The samples of tissue were taken from five patients (ten samples) with stomach cancer and normal (non-cancerous) segments from other parts of the stomach were used as a control. Eighteen compounds were identified in stomach tissue and seven of them were present both in healthy and cancer tissue. These compounds assumed to be endogenous and acetone ratio (AR) was calculated for ethanol, butane, carbon disulfide, 1-propanol, 2-butanone and 2-pentanone. The data shows that amount of 1-propanol and carbon disulfide in the gaseous composition is higher in cancer tissue than in normal tissue. Eight compounds were identified both in bacteria and tissue. These data suggest that bacteria present in the stomach might cause the increase in the concentration of 1-propanol and carbon disulfide in emission from cancer tissue.

Electrokinetic Detection and Characterization of Intact Microorganisms

Achievements in bacteria analysis with electromigration techniques may improve medical diagnoses, detection of food contamination, and sterility testing. (To listen to a podcast about this feature, please go to the Analytical Chemistry website at pubs.acs.org/ac.)

Coupling of solid-phase microextraction continuous bed (monolithic) capillaries with capillary zone electrophoresis for direct analysis of drugs in biological fluids†

Hyperlink robust biocompatible solid‐phase microextraction (SPME) devices were prepared using continuous bed (monolithic) restricted‐access media (RAM) as the SPME capillary insert. The RAM‐based SPME approach was able to simultaneously separate proteins from a biological sample, while directly extracting the active components of caffeine, paracetamol and acetylsalicylic acid from the drug NeoCitramonum. The devices were interfaced with a CZE system and fully automated analysis for sample preconcentration, desorption, separation and quantification of analytes was evaluated. Comparative study of in‐line coupled SPME–CZE using RAM and RP capillary inserts was carried out. Using an SPME (RAM) insert, the calculated caffeine, paracetamol and acetylsalicylic acid LODs in a bovine plasma sample were 0.3, 0.8 and 1.9 ng/mL, respectively.

Differentiation of Staphylococcus aureus strains by CE, zeta potential and coagulase gene polymorphism

Staphylococcus aureus is a common cause of infection in both hospitals and the community, and it is becoming increasingly virulent and resistant to antibiotics. Possibilities of fast, sensitive and cheap determination of these pathogenic bacteria are extremely important in antimicrobial therapy. In the present study, CE with chemically modified capillary and zeta potential measurements were used for differentiation of three different clinical strains of S. aureus. The data presented in this contribution suggested that electrophoretic behavior and the values of zeta potential should be very useful in distinguishing between closely related strains, which exhibited coagulase gene/protein polymorphism. Understanding the differences between S. aureus strains could help to improve our knowledge about S. aureus pathogenecity and to monitor for and respond to emergence of more virulent strains.

Determination of pathogenic bacteria by CZE with surface‐modified capillaries

The importance of electromigration techniques in molecular biology and medicine is increasing rapidly, especially in systematic studies on proteomes and metabolomes. Staphylococcus aureus and Escherichia coli are bacterial species most frequently encountered in human infections, and many serious illnesses can be observed in the hospital environment. In this contribution we proposed a CE method with different modification of internal capillary surface and with monolithic beds as a selective material for determination of bacteria in clinical samples. The electrophoretic separation depends on the differential mobility of bacteria in the capillary and selective interactions between bacterial cells and stationary phases (modified surface, monolithic beads). Proposed procedures could become an effective tool for diagnosis of certain diseases caused by S. aureus and E. coli as well as Proteus vulgaris.

Migration of bacteria through a monolith

The separation of bacteria by electromigration techniques was a subject of several of our previous papers. This contribution presents the results of investigation of the porosity of the monolithic bed and migration of Staphylococcus aureus cells through it. The gigaporous monolith was thermally synthesized using glycidyl methacrylate, triethylene glycol dimethacrylate and trimethylolpropane trimethacrylate as the monomers in the presence of porogen solvent containing 1-decanol, polyethylene glycol and 2-methoxyethanol. The porous properties were evaluated by inverse size-exclusion chromatography (ISEC) using a wide range of polystyrene standards of different molecular weights. The results have shown, that large pores (ca. 300 nm) dominate in the monolithic bed structure, however much larger flow-through pores must also be present as ca. 1 microm sized S. aureus bacteria were able to migrate through the bed.

Comparison of Several Extraction Methods for the Isolation of Benzoic Acid Derivatives from Melissa officinalis

Abstract Several extraction techniques, such as Soxhlet extraction, solid phase extraction using molecularly imprinted polymer, matrix solid phase dispersion, and supercritical fluid extraction were evaluated for the isolation and purification of phenolic compounds, e.g., benzoic acids from natural samples of Melissa officinalis. The extracts of benzoic acids were analyzed by high performance liquid chromatography (HPLC) in reversed phase modus (C18 column) and under gradient elution (acetonitrile/0.074 mol/L formic acid). The results showed that the recovery rates of gallic acid, p‐hydroxybenzoic acid, protocatechuic acid, gentisic acid, vanillic acid, and syringic acid from biological materials by MSPD was equivalent with and, in fact, higher than that of conventional extraction methods.

Combination of capillary electrophoresis, PCR and physiological assays in differentiation of clinical strains of Staphylococcus aureus.

Fast, sensitive and cheap determination of pathogenic bacteria is extremely important in many branches, for example biotechnology, quality control, analysis of samples and antimicrobial therapy. The development and application of analytical techniques in practice could provide new possibilities in this regard. The bacterial pathogen Staphylococcus aureus is responsible for a significant amount of human morbidity and mortality. Rapid and sensitive determination is therefore very important. In the present study, novel methods, based on capillary zone electrophoresis and (as confirmation of these results) molecular analysis of a part of the coag gene, were developed for identification and differentiation of three S. aureus strains. The electrophoretic measurements rely on the differential mobility of bacteria in the fused silica capillary under the direct current electric field. To perform coagulase gene typing, the repeated units encoding hypervariable regions of the S. aureus gene were amplified using the PCR technique followed by restriction enzyme digestion and analysis of restriction fragment length polymorphism patterns as well as sequencing. Finally, the results of electrophoretic measurements with molecular analysis were compared.

A study of interactions between bacteria and antibiotics by capillary electrophoresis.

To assess the bacteria–antibiotic interactions in patients with postoperative wound infections, a simple electrophoretic test was performed. To estimate the effectiveness of the antibiotic therapy and to prepare 3‐day profiles of bacteria “quantity” in biological samples, CE was used. As our team demonstrated earlier, the method is easy and fast, sample pretreatment is not necessary, and it is characterized by high selectivity. Finally, the statistically optimal and significant results of the CZE test analysis for detection of Escherichia coli cells was established for migration time lower than 3.5 min. The obtained sensitivity and specificity amounted to 89.5 and 100%, respectively. It is the first application of CZE in the study of medical therapy.