Ga. Schultz et Jn. Alexander, INCORPORATION OF A RAM CELL IN A MICROCOLUMN FOR ON-COLUMN DETECTION OF RADIOLABELED MOLECULES FOR LC-RAM-ESI-MS, The Journal of microcolumn separations, 10(5), 1998, pp. 431-437
The objective of this work was to design a nondestructive solid scinti
llation radioactivity monitor (RAM) cell for the detection of radiolab
eled molecules for use with packed liquid chromatography (LC) microcol
umns coupled to electrospray ionization-mass spectrometry (ESI-MS). To
take advantage of the increased mass sensitivity offered by microcolu
mns coupled to ESI-MS, the contributions to band broadening due to the
solid scintillant was minimized by incorporation of the cell within t
he microcolumn, resulting in on-column detection of radiolabeled molec
ules. Our work shows that the contributions to band broadening and RAM
sensitivity depend on the particle size of the solid scintillant, cel
l volume, and cell diameter. Our results show that a solid scintillati
on cell packed with silanated lithium glass with a mean particle size
of 9.6 +/- 2.6 mu m gives a 24% increase in peak area compared to a ce
ll packed with a mean particle size of 21.0 +/- 2.5 mu m. Our results
show that a. 508-mu m-internal-diameter (i.d.) cell provided a 15% inc
rease in peak area compared to a 762-mu m-i.d. cell. This work demonst
rates the feasibility of detection of radiolabeled molecules using mic
rocolumns. We have developed a 1.2-mu L volume on-column RAM cell for
use with 250-mu m-i.d. packed microcolumns which can detect less than
200 radioactive disintegrations per minute (dpm) (1 dpm = 60 Bq) for a
1-min-wide chromatographic peak. Further development of solid scintil
lants with a mean particle size of 3-5 mu m and spherical shape will p
rovide additional sensitivity and a decrease in the band broadening co
ntributed by the RAM cell. Additionally, such particles may be slurrie
d with stationary phase and pack:ed into microcolumns for characteriza
tion of on-column dispersion processes. The development of an on-colum
n RAM solid scintillation cell for microcolumns combined with ESI-MS w
ill have applications including the characterization of radiolabeled m
olecules and their metabolites and the determination of ligand-protein
binding sites through the use of radiolabeled ligands combined with o
n-line protein digestion using microcolumn LC-RAM-ESI-MS. (C) 1998 Joh
n Wiley & Sons, Inc.