Coupling of acetonitrile deproteinization and salting-out extraction with acetonitrile stacking for biological sample clean-up and the enrichment of hydrophobic compounds (porphyrins) in capillary electrophoresis.
A new sample pretreatment approach in CE was developed for concurrent biological sample clean-up and the concentration of hydrophobic compounds based on the combination of ACN deproteinization with salting-out extraction. Further enhancement in concentration detection sensitivity was achieved by coupling (offline) salting-out extraction with an online CE sample enrichment technique known as "ACN stacking". By optimizing the pH of salting-out extraction, a number of model compounds (hydrophobic porphyrins with clinical significances), i.e. zinc-protoporphyrin, protoporphyrin, and coproporphyrin (CP) III and I, can be efficiently extracted from the aqueous sample into a smaller volume organic solvent (ACN) phase and an enrichment factor of ca. 100 can be obtained. The pressure injection of the enriched ACN phase (containing ca.1% NaCl) into the CE capillary at 10% capillary volume resulted in additional concentration of the various hydrophobic porphyrins, allowing for a combined enrichment factor of ca.1000 to be obtained. Calibration curves obtained for the determination of a pair of positional isomers with significant diagnostic value, urinary CPIII and CPI, were found to be linear between 10-300 ng/mL (with R2 = 0.999), and LODs (absorbance detection at 400 nm) were ca. 0.8 ng/mL (1.1 nmol/L of CPIII or CPI). Based on a single salting-out extraction, intraday precisions (nine consecutive injections) for both CPIII and CPI (at spiked concentrations of 10-300 ng/mL into urine) in terms of migration time and peak area were found to be within the range of 0.2-0.5 and 0.8-2.9%, respectively.
Li
Qi
Q
Department of Chemistry, Hong Kong Baptist University, Kowloon Tong, Hong Kong, P R China.
Huie
Carmen W
CW
D000097:Acetonitriles / Q000737:chemistry; D053000:Analytic Sample Preparation Methods; D019075:Electrophoresis, Capillary / Q000379:methods* / Q000592:standards*; D004962:Estranes / Q000737:chemistry; D006801:Humans; D057927:Hydrophobic and Hydrophilic Interactions; D009570:Nitriles / Q000737:chemistry; D011166:Porphyrins / Q000652:urine*; D011506:Proteins / Q000737:chemistry*; D012965:Sodium Chloride / Q000737:chemistry
D000097:Acetonitriles; D004962:Estranes; D009570:Nitriles; D011166:Porphyrins; D011506:Proteins; C084683:3-hydroxy-5-estrane-17-carbonitrile; D012965:Sodium Chloride; C032159:acetonitrile