Laser-induced fluorescence Dotaz Zobrazit nápovědu
Interestingly, even though the absorption maximum of prepared capped carbon quantum dots (CQDs) is 210 nm and the emission maximum is 392 nm, using capillary electrophoresis with laser-induced fluorescence detection (CE-LIF) with excitation wavelength of 470 nm and long pass emission filter (510 nm) a signal was observed. Application of separation technique reviled presence of two different species, which corresponded to two well-resolved peaks present in the electropherogram. This fact is probably caused by presence of particles of different sizes.
Ellipticine (5,11-dimethyl-6H-pyrido[4,3-b]carbazole), an alkaloid isolated from Apocynaceae plants, exhibits an antitumor activity, which is exceptionally high against several specific types of tumors. Ellipticine is also interesting as an anticancer drug as it has limited side effects and lacks of hematological toxicity. Various methods to study intercalating activity of this drug have been developed. However, to our best knowledge, capillary electrophoresis (CE) as a technique combining high separation resolution with various detection options has never been used for these purposes. In this study, a novel separation method based on CE with laser-induced fluorescence (CE-LIF) detection has been developed for the determination of ellipticine and for the monitoring of ellipticine-DNA interaction. Sodium acetate (50 mM, pH 4.5) was used as a background electrolyte and LIF detection at λ(ex) = 488 nm. The limit of detection for ellipticine was determined to be 5 × 10⁻⁸ M. A total of 20% dimethyl sulfoxide was found optimal as sample solvent. Additionally, intercalation of ellipticine into the double-stranded DNA was investigated. Signal corresponding to ellipticine was decreasing and a new peak appeared and was growing. It can be concluded that CE-LIF is a method applicable to in vitro studies of ellipticine-DNA complexes.
- MeSH
- dimethylsulfoxid chemie MeSH
- DNA krev chemie izolace a purifikace metabolismus MeSH
- elektroforéza kapilární metody MeSH
- elipticiny chemie metabolismus farmakologie MeSH
- erytrocyty chemie MeSH
- fluorescenční spektrometrie metody MeSH
- kur domácí MeSH
- limita detekce MeSH
- lineární modely MeSH
- zvířata MeSH
- Check Tag
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
A new sensitive capillary electrophoretic method with laser-induced fluorescence (LIF) was developed for quantitation of glutathione (GSH) in biological samples. Eosin-5-maleimide was used to label the GSH molecule and the formed conjugate was separated in a 15 mM 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid electrolyte at pH 7.0 in less than 3 min. The conjugate was detected with an in-house built LIF system, utilizing an inexpensive 515 nm diode laser module. Studies were performed to optimize the derivatization (the ratio of reagent to analyte, the reaction time, pH, etc.) and separation conditions. Sensitive detection of GSH at concentrations as low as 0.18 nM was obtained. The method was applied in the analysis of biological fluids (exhaled breath condensate, saliva) and was found to be suitable for determination of GSH in these samples at trace levels below 1 nM. To the best of our knowledge, this is the first report on determination of GSH in exhaled breath condensate by capillary electrophoresis (CE).
- MeSH
- dechové testy MeSH
- elektroforéza kapilární metody MeSH
- fluorescence MeSH
- glutathion analýza MeSH
- lasery * MeSH
- lidé MeSH
- sliny chemie MeSH
- tělesné tekutiny chemie MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
In this work, a sensitive capillary electrophoresis method with laser induced fluorescence detection for determination of malondialdehyde in various biological fluids was developed. Malondialdehyde reacts with thiobarbituric acid under optimized conditions of pH=2, reaction time of 60min and temperature of 90°C, yielding an adduct that can be separated in a 50mM sodium borate background electrolyte at pH 9. The separation of the formed adduct was accomplished in less than 6min with limit of detection of 1.1nM due to the use of 532nm laser module, exactly matching the maximum excitation wavelength of the formed adduct. The developed method offers unprecedented sensitivity and was for the first time used for analysis of malondialdehyde in exhaled breath condensate. The method proved to be also applicable to other samples of biological fluids, such as blood plasma and saliva.
- MeSH
- dechové testy metody MeSH
- elektroforéza kapilární metody MeSH
- fluorescence MeSH
- lasery * MeSH
- lidé MeSH
- malondialdehyd analýza MeSH
- sérum metabolismus MeSH
- sliny metabolismus MeSH
- vydechnutí * MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
Capillary electrophoresis with laser-induced fluorescence detection (CE-LIF) was applied to separation and sensitive determination of red food colorants. Diode pumped frequency-doubled Nd:YAG laser (532 nm) was used as an excitation source in a laboratory-built CE-LIF system. For highly fluorescent erythrosine B (E127), an extrapolated limit of detection (LOD) of 0.4 ng mL(-1) (S/N=3) was achieved. Extrapolated LODs of other tested red additives, such as carmoisine, E122 (0.5 microg mL(-1)); amaranth, E123 (0.2 microg mL(-1)); ponceau 4R, E124 (0.3 microg mL(-1)) and red 2G, E128 (0.3 microg mL(-1)) were about one-order lower compared to results obtained with CE with absorbance detection in UV/vis (CE-UV/vis). The main advantages of using CE-LIF for analysis of food samples are high selectivity and minimization of matrix effect. To our knowledge, this is the first use of CE-LIF for determination of red food colorants.
Quantum dots (QDs) are one of the most promising nanomaterials, due to their size-dependent characteristics as well as easily controllable size during the synthesis process. They are promising label material and their interaction with biomolecules is of great interest for science. In this study, CdTe QDs were synthesized under optimal conditions for 2 nm size. Characterization and verification of QDs synthesis procedure were done by fluorimetric method and with CE. Afterwards, QDs interaction with chicken genomic DNA and 500 bpDNA fragment was observed employing CE-LIF and gel electrophoresis. Performed interaction relies on possible matching between size of QDs and major groove of the DNA, which is approximately 2.1 nm.
- MeSH
- DNA chemie metabolismus MeSH
- elektroforéza kapilární metody MeSH
- fluorescenční spektrometrie metody MeSH
- kur domácí MeSH
- kvantové tečky chemie metabolismus MeSH
- sloučeniny kadmia chemie metabolismus MeSH
- telur chemie metabolismus MeSH
- zvířata MeSH
- Check Tag
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Doxorubicin (DOX) is an effective antitumor drug employed for treatment of a wide range of cancers types such as neuroblastoma, osteosarcoma, breast and esophageal carcinomas. On the other hand, the cumulative dose is restricted (300-550 mg/m(2)) and its amount administered to a patient has to be closely controlled due to its cardiotoxicity. To understand the mechanisms of the DOX side effects as well as to reveal the ways how to reduce its adverse impact on cardiomyocytes, the interactions with particular components of the blood and tissues have to be studied in greater detail. In this work, microdialysis technique was optimized to extract DOX from samples and subsequently monitor its interaction with BSA. Finally, the microdialysis probe was connected on-line to the LIF detector to ensure the real-time detection. The best flow rate was 1 μL/min and after 120 min of microdialysis 28% of the DOX was dialyzed out from the sample. The results from investigation of the DOX-BSA interaction indicate that the interaction occurs in less than 30 min, causing marked decrease in the amount of DOX extracted by microdialysis.
- MeSH
- doxorubicin chemie MeSH
- fluorescence MeSH
- lidé MeSH
- limita detekce MeSH
- mikrodialýza MeSH
- sérový albumin hovězí chemie MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Analysis of PCR fragments for applications, such as screening of nucleotide polymorphisms, detection of somatic mutations, or quantification of reverse-transcription PCR products, becomes central in clinical research as well as preventive testing, diagnostic screening, and pharmacogenomic genotyping. A variety of CE techniques, utilizing great potential of multicapillary-array sequencers, is now commonly applied in prevention, diagnosis, and treatment of a wide range of genetic diseases (cancer, cardiovascular, and neurodegenerative diseases, etc.). Costs of fluorescently labeled primers is often a major factor in large-scale projects requiring mutation analysis in hundreds or thousands of samples. In the present paper we introduce a simple approach of detecting unlabeled DNA fragments through intercalation without a need for adding intercalator to the separation polymer matrix. The dye is only added to the anode reservoir, and mixing with the separated DNA fragments takes place upon its migration opposite to the direction of the CE separation. Using two common intercalating dyes (ethidium bromide and SYBR Green II) we present this method as a tool for routine PCR detection and quantification.
