A new detector for macromolecular separations is described. The detector counts individual macromolecules (molecular weights greater than about 10,000) and reports counts per second. The chromatographic effluent is electrosprayed, neutralized, and swept to the detector by a stream of air. The detector is a condensation particle counter that detects individual particles by light scattering from droplets condensed on the particles. When used as the detector for a size exclusion separation of proteins, the detector has a linear range of 4 orders of magnitude with detection limits as low as 0.1 microgram/mL. The detector can be directly interfaced (no makeup flow) with effluent flows as low as 10 nL/min. A Monte Carlo model based on size measurements of the electrosprayed droplets correctly predicts the observed detector behavior.

A new liquid chromatography instrument is described that employs small-diameter capillary columns and an optically controlled sample gating procedure to separate a mixture of fluorescein isothiocyanate (FITC) labeled amines in as few as 6 s. Efficiency of the separation, expressed as the number of theoretical plates, is observed to increase linearly with column length and thus analysis time. The combination of short analysis time and automated sample introduction allows signal averaging to be used to enhance the precision of the measurement.

Abstract. The performance of etched borosilicate capillary columns with a bonded octadecyl stationary phase has been examined. Plate heights for three test compounds in columns with inner diameters between 6.5 and 14.6 pm were measured as a function of mobile phase velocity. The reduced plate height vs. reduced velocity curves were then fit to the Golay equation using nonlinear regression analysis. It was found that in addition to longitudinal diffusion in the mobile phase, longitudinal diffusion in the stationary phase was a significant contributor to broadening at low mobile phase flow rates. At higher flow rates, resistance to mass transfer in the stagnant mobile phase in the porous glass produced by the etching procedure became measurable.

The preparation of open tubular liquid chromatographc columns with small inner diameters ( 5 2 0 pm) from Pyrex 7740 borosilicate glass is described. The surface area of the inner wall of the capillary is increased by etching with a series of acidic and basic solutions. Dimethyloctadecylchlorosilane is then reacted with the column wall to create a bonded stationary phase layer. The resulting columns exhibit good retention and high efficiency. The effects of each step in the etching process are examined and the silanization reaction is optimized.