Online capillary liquid-liquid electroextraction of peptides as fast pre-concentration prior to LC-MS.
In this research paper, we show that capillary electroextraction (cEE) is capable of fast online peptide concentration and that it can be coupled online to LC-MS to result in a fast and sensitive method. Electroextraction takes place when an electrical field is applied in a two-phase liquid-liquid system. Sample molecules in the organic phase migrate very fast into the aqueous phase and are concentrated in a small zone. In this work, cEE of peptides is developed and coupled online to LC-MS via a switching valve. Comparison of 10 min of cEE-LC-MS with a normal LC-MS injection showed more than 100-fold increased peak heights. Of five model peptides, good calibration curves in the range of 0.05-5 μmol/L were obtained. The linearity was good (R(2) values between 0.984 and 0.996) and RSD between 5% at the highest to 25% at the lowest concentration (n=3). The LOD of bradykinin, angiotensin I-converting enzyme inhibitor and angiotensin I was in the low nmol/L range. Analysis of a tryptic digest of eight model proteins resulted in more than 170 peptides, without bias for pI or hydrophilicity. Urine analysis is demonstrated, resulting in an LOD around 0.04 μmol/L urine for tryptic cytochrome C peptides spiked to urine and an increase of 42% in the number of chromatographic peaks compared with the conventional LC-MS. In summary, cEE-LC-MS is a fast electrophoresis-driven sample preconcentration technique that is quantitative, able to extract a wide peptide range and applicable to bioanalysis.
Division of Analytical Biosciences, Leiden/Amsterdam Center for Drug Research, Leiden University, The Netherlands. p.lindenburg@chem.leidenuniv.nl
Petrus W
PW
Lindenburg
Remco
R
Seitzinger
F W Alexander
FW
Tempels
Ubbo R
UR
Tjaden
Jan
J
van der Greef
Thomas
T
Hankemeier
D000818:Animals; D002417:Cattle; D005591:Chemical Fractionation / Q000379:methods*; D002645:Chickens; D002853:Chromatography, Liquid / Q000379:methods*; D019075:Electrophoresis, Capillary / Q000379:methods*; D006801:Humans; D016014:Linear Models; D013058:Mass Spectrometry / Q000379:methods*; D008958:Models, Molecular; D010446:Peptide Fragments / Q000032:analysis / Q000302:isolation & purification* / Q000378:metabolism / Q000652:urine; D011506:Proteins / Q000378:metabolism; D012680:Sensitivity and Specificity
D010446:Peptide Fragments; D011506:Proteins