Publication details

CIEF separation, UV detection, and quantification of ampholytic antibiotics and bacteria from different matrices

Authors

HORKÁ Marie VYKYDALOVÁ Marie RŮŽIČKA Filip ŠALPLACHTA Jiří HOLÁ Veronika DVOŘÁČKOVÁ Milada KUBESOVÁ Anna ŠLAIS Karel

Type Article in Periodical
Magazine / Source Analytical and Bioanalytical chemistry
MU Faculty or unit

Faculty of Medicine

Citation
Web http://link.springer.com/article/10.1007%2Fs00216-014-8053-8
Doi http://dx.doi.org/10.1007/s00216-014-8053-8
Field Microbiology, virology
Keywords Capillary isoelectric focusing with UV detection; Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry; Ampholytic antibiotics; Staphylococcus epidermidis; Human whole blood
Description The effect of antibiotics on the microbial cells and concentration of antibiotics in the human body is essential for the effective use of antimicrobial therapy. The capillary isoelectric focusing is a suitable technique for the separation and the detection of bacteria, and amphoteric substances from nature. However, the determination of isoelectric points of ampholytic antibiotics by conventional techniques is time consuming. For this reason, capillary isoelectric focusing seems to be appropriate as a simple and reliable way for establishing them. The separation conditions for the capillary isoelectric focusing of selected ampholytic antibiotics with known isoelectric points and pK (a)s, ampicillin (pI 4.9), ciprofloxacin (pI 7.4), ofloxacin (pI 7.1), tetracycline (pI 5.4), tigecycline (pI 9.7), and vancomycin (pI 8.1), were found and optimized in the suitable pH ranges pH 2.0-5.3, 2.0-9.6, and 9.0-10.4. The established values of isoelectric points correspond with those found in the literature except tigecycline. Its pI was not found in the literature. As an example of a possible procedure for direct detection of both ampholytic antibiotics and bacteria, Staphylococcus epidermidis, in the presence of culture media or whole human blood, was found. The changes of the bacterial cells after their treatment with tetracycline were confirmed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. Capillary isoelectric focusing allows the fast and simple determination of isoelectric points of relevant antibiotics, their quantification from the environment, as well as studying their effectiveness on microorganisms in biological samples.
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