Novel and simple headspace in-tube microextraction coupled with capillary electrophoresis

Lee, H.R.,Cho, S.M.,Kim, J.,Chung, D.S. Elsevier 2014 Journal of chromatography Vol.1346 No.-

In liquid phase microextraction, high enrichment factors can be obtained using an acceptor phase of small volume. By hanging an acceptor drop at the separation capillary tip, single drop microextraction (SDME) can be in-line coupled with capillary electrophoresis (CE). The small surface-to-volume ratio of the drop enables high enrichment factors to be obtained in a short time. One practical issue in SDME is how to keep the drop attached to the capillary stable. Here, we present novel but extremely simple in-tube microextraction (ITME) using the liquid inside the capillary as an acceptor phase, without forming a drop at the capillary tip. As a first example, ITME has been combined with headspace (HS) extraction. Simply by placing a capillary filled with a basic run buffer in the HS above an acidic donor solution, volatile acidic analytes were extracted into the acceptor phase in the capillary. After extraction, electrophoresis of the extracts in the capillary was carried out. Owing to the robust nature of the acceptor phase, the extraction temperature and time ranges of HS-ITME can be extended significantly, compared to HS-SDME. The enrichment factors for chlorophenols in a standard solution were up to 1100 under an optimal HS-ITME condition of 80<SUP>o</SUP>C for 15min and the limits of detections (LODs) obtained by monitoring the absorbance at 214nm were about 4nM. The whole procedures of HS-ITME-CE were carried out automatically using built-in programs of a commercial CE instrument.

Affinity chromatography and capillary electrophoresis for analysis of the yeast ribosomal proteins

( Miriam S. Goyder ),( Keith R. Willison ),( David R. Klug ),( Andrew J. Demello ),( Oscar Ces ) 생화학분자생물학회 (구 한국생화학분자생물학회) 2012 BMB Reports Vol.45 No.4

We present a top down separation platform for yeast ribosomal proteins using affinity chromatography and capillary electrophoresis which is designed to allow deposition of proteins onto a substrate. FLAG tagged ribosomes were affinity purified, and rRNA acid precipitation was performed on the ribosomes followed by capillary electrophoresis to separate the ribosomal proteins. Over 26 peaks were detected with excellent reproducibility ( 0.5% RSD migration time). This is the first reported separation of eukaryotic ribosomal proteins using capillary electrophoresis. The two stages in this workflow, affinity chromatography and capillary electrophoresis, share the advantages that they are fast, flexible and have small sample requirements in comparison to more commonly used techniques. This method is a remarkably quick route from cell to separation that has the potential to be coupled to high throughput readout platforms for studies of the ribosomal proteome. [BMB reports 2012; 45(4): 233-238]

Practical sample pretreatment techniques coupled with capillary electrophoresis for real samples in complex matrices

Jarvas, Gabor,Guttman, Andras,Mię,kus, Natalia,Bą,czek, Tomasz,Jeong, Sunkyung,Chung, Doo Soo,Pä,toprstý,, Vladimir,Masá,r, Mariá,n,Hutta, Milan,Datinská,, Vladim Elsevier 2020 Trends in analytical chemistry Vol.122 No.-

<P><B>Abstract</B></P> <P>By coupling a sample pretreatment technique of sample clean up and enrichment power with capillary electrophoresis (CE) of high-performance separation, the task of analyzing trace analytes in a complex matrix such as a biological sample can be carried out successfully with ease. This review aims for providing an overview of strategies to couple sample pretreatment techniques with capillary and related microscale (e.g., microchip) electrophoresis, practically adoptable in an automatic manner, without requiring serious modification of existing instruments to install sophisticated interfaces. In-line sample pretreatment techniques based on liquid phase microextraction performed before sample injection and on-line sample preconcentration techniques performed during or after sample injection are discussed with emphasis on the applicability to samples of high conductivity, commonly encountered for biological samples. An overview of the recent developments in microfluidic immobilized enzymatic microreactors which fit excellently to microchip CE is also given.</P> <P><B>Highlights</B></P> <P> <UL> <LI> Recent advances and major trends in sample pretreatment for capillary electrophoresis are summarized. </LI> <LI> In-line and on-line sample pretreatment techniques are discussed with emphasis on biological samples. </LI> <LI> We provide an overview of strategies to couple sample pretreatment techniques with capillary and microchip electrophoresis. </LI> </UL> </P>

Optimization of TILLING System Based on Capillary Electrophoresis for Targeted Selection of Pepper Gene Mutants

강한솔,김상훈,이상우,김세원,류재혁,김진백,염선인,강시용,조영득 한국원예학회 2018 Horticulture, Environment, and Biotechnology Vol.59 No.3

Mutation breeding is largely performed by forward and reverse genetics-based approaches. In the latter, Target Induced Local Lesions IN Genomes (TILLING) can be used to screen mutants harboring mutations in a target gene whose function can be predicted with available functional genomic information. In TILLING, it is necessary to use an analytical system for accurate and efficient screening of target gene mutants among several randomly mutagenized lines. Therefore, we optimized a capillary electrophoresis-based TILLING system that can quickly and easily analyze many samples and confirmed its performance by applying it to pepper mutation populations. First, optimal conditions concerning heteroduplex DNA formation, type and concentration of endonucleases, and concentration, pooling depth, and size of PCR amplicons were determined in an analytical system using a capillary electrophoresis apparatus without labeled primers. The results showed that CelII endonuclease (Surveyor™) had higher sensitivity than CelI. While the recommended concentration in the manufacturer’s manual was 1×, we obtained higher sensitivity when the amount of PCR product and concentration of the endonucleases were at 180 ng and 0.5× for CelII and 120 ng and 2× for CelI. Meanwhile, the maximum pooling depth for DNA from individuals in mutant populations inversely correlated to PCR amplicon size. The limitation of DNA pooling was 4× for a 1872-bp amplicon, while at least 16× pooling was possible for a 672-bp amplicon. Using this optimized system, TILLING was performed on 2615 individuals from the mutant populations developed in a Korean pepper landrace, “Yuwolcho,” to screen lines bearing a mutation in CCS (capsanthin/capsorbin synthase). We were able to select a mutant line with a substituted base pair in the translational start codon of CCS. The capillary electrophoresis-based TILLING system optimized in this study is expected to be useful in practical applications for reverse genetics-based breeding and functional genomics studies.

Analysis of Health-related Microbes by Capillary Electrophoresis

Moon, Byoung-Geoun,Kim, Yong-Seong Korean Chemical Society 2003 Bulletin of the Korean Chemical Society Vol.24 No.8

Analysis of health-related microbes called probiotics was performed by capillary electrophoresis. A rapid and easy characterization for two important probiotics, Saccharomyces cerevisiae and Enterococcus feacalis, was obtained in the running buffer containing poly(ethylene oxide). Quantitation of probiotic (Saccharomyces cerevisiae) shows a good linearity between the peak area versus the concentration of microbe. From the comparison of electropherograms of antidiarrhea, it was found that capillary electrophoresis could be employed for the quality control and quality assurance for the production of a medicine containing the probiotics.

Capillary electrophoresis(CE)를 이용한 천궁의 원산지 판별

김정현 ( Kim Jeong Hyeon ),김은영 ( Kim Eun Yeong ),정경숙 ( Jeong Gyeong Sug ),류미라 ( Lyu Mi La ) 한국응용생명화학회 2003 Applied Biological Chemistry (Appl Biol Chem) Vol.46 No.4

Optimal extraction, separation and capillary rinsing conditions for capillary electrophoresis (CE) were established to discriminate the geographical origin of ligusticum root (Ligusticum wallichii) using 113 samples (domestic sample n=62, foreign sample n=51). Ligusticum root wat extracted with 30% ethanol and separated on a uncoated fused-silica (50㎛×27㎝) capillary. Conditions for optimal analysis include: temperature, 40℃; voltage, 10 kV; and pressure injection time, domestic and foreign samples were 5 sec and 2 sec, respectively. The optimal separation buffer was 0.1 M phosphate buffer (pH 2.5) containing 15 mM iminodiacetic acid with 40% methanol. Under the optimal conditions established for CE, the ratio of specific peak area (peak LW-1) to other peak area (peak LW-5) was effective in discrimination geographical origin of ligusticum root. The mean accuracy for correct discrimination of geographical origin of domestic and foreign ligusticum roots were 65% and 63%, respectively.

Application of capillary electrophoresis to amino acid sequencing of peptide

Kim, Nam Jung,Kim, Jung Ho,Lee, Kong-Joo 梨花女子大學校 藥學硏究所 1995 藥學硏究論文集 Vol.- No.5

Separations of twenty phenylthiohydantoin (PTH) amino acids, and amino acid derivatives, resulting from Edman degradation of peptides and proteins, were optimized for peptide sequencing by capillary electrophoresis. Manual sequencing of angiotensin II was performed by Edman degradation and capillary electrophoresis of the PTH amino acid obtained after each cycle. The results were compared with those of an automated conventional protein sequencer. Interfacing capillary electrophoresis with Edman degradation provides an additional option for protein sequencing.

Capillary Electrophoresis에 의한 Carvedilol 및 그 대사체의 거울상이성질체 분리

명승운,조천호,구소현 한국분석과학회 2006 분석과학 Vol.19 No.4

A capillary electrophoresis method for the separation of carvedilol and its metabolites enantiomersconcentration and capillary temperature for enantiomer resolution were investigated. Best results were obtainedby 15 mM carboxymethyl-β-cyclodextrin in the run bufer. Also the effect of capillary temperature on efficiencywas assessed. The optimized method was applied for separation of chiral carvedilol and its three metabolites.

Sensitive analysis of amino acids with carrier-mediated single drop microextraction in-line coupled with capillary electrophoresis

Choi, J.,Choi, K.,Kim, J.,Ahmed, A.Y.B.H.,Al-Othman, Z.A.,Chung, D.S. Elsevier 2011 Journal of chromatography Vol.1218 No.41

In order to analyze amino acids sensitively without derivatization, we have developed carrier-mediated single drop microextraction (SDME). Nonane-1-sulfonic acid was added to an acidic sample donor solution as a carrier to form neutral ion pair complexes with amino acids. The ion pair complexes were extracted to the organic phase, covering a drop of an aqueous basic acceptor phase hanging at the tip of a capillary, and then back-extracted to the basic acceptor phase, where both the amino acids and the carrier have negative charges and the ion pair complexes are broken. The resulting extract of enriched amino acids was injected into the capillary and analyzed by capillary electrophoresis. With 20-min SDME with agitation of the donor phase, enrichment factors of four aromatic amino acids were up to 120-fold, yielding the LOD of 70-500nM. The linear dynamic ranges for corrected peak areas were 1-100μM with linear correlation coefficients larger than 0.9959. With internal standardization, the intraday RSDs of migration times and corrected peak areas were 0.01-0.04% and 2.0-3.7%, respectively. The capabilities of sample cleanup including desalting and preconcentration of carrier-mediated SDME were demonstrated with the analysis of human urine after minimal pretreatment of acidification and centrifugation.

Application of capillary electrophoresis to amino acid sequencing of peptide

Kim, Nam Jung,Kim, Jung Ho,Lee, Kong-Joo 이화여자대학교 생명과학연구소 1995 생명과학연구논문집 Vol.6 No.-

Separations of twenty phenylthiohydantoin (PTH) amino acids, and amino acid derivatives, resulting from the Edman degradation of peptides and proteins, were optimized for peptide sequencing by capillary electrophoresis. Manual sequencing of angiotensin Ⅱ was performed by Edman degradation and capillary electrophoresis of the PTH amino acid obtained after each cycle. The results were compared with those of an automated conventional protein sequencer. Interfacing capillary electrophoresis with Edman degradation provides an additional option for protein sequencing.