Fundamental aspects of aerosol-based light-scattering detectors for separations.
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Biomedical subjects
Publications and source records attributed to J A Koropchak.
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Condensation nucleation light scattering detection (CNLSD) was coupled with a pressurized capillary electrochromatography (pCEC) system using an electrospray interface. Supplementary pressure from a high-pressure pump was used to stabilize the electrospray and electrochromatography processes. Hydrodynamic injections were made with a 20 nl injection valve, and the inherent dead volume from the valve was successfully minimized, such that plate numbers in the range of 120,000 to 350,000/m were observed. Selectivity tuning using both pressure and voltage with the pressurized capillary electrochromatography system was demonstrated. Good reproducibility, comparable sensitivities for a wide range of compounds, including carbohydrates, and limits of detection down to the 50 ng/ml level, corresponding to 1-2 pg levels, were determined without the need for derivatization, demonstrating that condensation nucleation light scattering detection is a sensitive, universal detection method for pressurized capillary electrochromatography.
A sensitive method of analysis for biogenic amines, putrescine, cadaverine, histamine and an amino acid precursor, histidine is described herein using ion-exchange chromatography and condensation nucleation light scattering detection. The method was successfully used for the analysis of biogenic amines in fish samples. The method offers a number of advantages: fast elution of analytes with no need for mobile phase conductivity suppression, no derivatization and no electrochemical activity for the analyte's detection. The 3 sigma detection limits for these compounds were found to range from 8 to 20 ng/ml.
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An improved method for coupling capillary electrophoresis (CE) with condensation nucleation light scattering detection (CNLSD) is described. The method employs an electrospray aerosol source followed by aerosol particle neutralization within a weak plasma established by a polonium-210 alpha emitter. Sensitive, universal detection of underivatized proteins, peptides, and amino acids is demonstrated. The system performance was significantly affected by the operational mode of the electrospray source, characterized by the physical appearance of the droplet at the end of the electrospray capillary. With a so-called pulsating mode, relatively low backgrounds were obtained with 10 mM ammonium acetate or 10 mM ammonium acetate/10 mM ammonia CE buffers using one diffusion screen, allowing detection of proteins at single microgram per milliliter levels and amino acids and peptides at submicrogram per milliliter levels. Linearity of response, expressed as peak height or peak area; mass limits of detection (LODs) for proteins, peptides, and amino acids at the picogram level, corresponding to femtomole levels of peptides and amino acids or subfemtomole levels of proteins; and better detectability compared to UV absorbance at 214 and 200 nm were demonstrated. The separation efficiencies obtained with the CE-electrospray-CNLSD system are much higher than those obtained for a previously reported pneumatic nebulizer-based system and in the range of approximately 20,000-160,000 plates/m using the pulsating electrospray mode. With careful adjustment of the electrospray voltage, a different operating mode, called the silver bullet mode, could be established for which higher signals, lower background and background noise, and higher separation efficiencies were observed compared to the pulsating mode. The lower background levels observed with the silver bullet mode eliminated the need for the use of a diffusion screen for background control with the buffer employed. With the silver bullet mode without a diffusion screen, linear response and LODs at the 15 ng/mL level, corresponding to subpicogram or 1-2 fmol levels of underivatized peptides and amino acids, and plate numbers in the range of 65,000-220,000 plates/m were estimated.