Detection of nonderivatized peptides in capillary electrophoresis using quenched phosphorescence.
A capillary electrophoresis detection technique for (small) peptides is presented, i.e. quenched phosphorescence, a method that is generally applicable and does not require chemical derivatization. For this purpose, a novel phosphorophore, 1-bromo-4-naphthalenesulfonic acid (BrNS), was synthesized. BrNS has sufficient water solubility and provides strong phosphorescence at room temperature over a wide pH range. The detection is based on the dynamic quenching of the BrNS phosphorescence background signal by electron transfer from the amino group of the peptides at pH 9.5-10. For the di- and tripeptides Val-Tyr-Val, Val-Gly-Gly, Ala-Ser, Gly-Asn, Gly-Ala, and Gly-Tyr, detection limits in the range of 5-20 microg/L were obtained. The novel technique is even a good alternative for the (limited) group of peptides containing tyrosine and, thus, exhibiting native fluorescence as well as strong UV absorption: using Gly-Tyr, Val-Tyr-Val, methionine enkephalin, and human angiotensin II as test compounds, quenched phosphorescence detection was found to compare favorably with absorption detection at 190- and 266-nm laser-induced fluorescence detection, as performed with a recently developed, small-size, quadrupled Nd:YAG laser.
Kuijt
J
J
Department of Analytical Chemistry and Applied Spectroscopy, Vrije Universiteit, Amsterdam, The Netherlands.
van Teylingen
R
R
Nijbacker
T
T
Ariese
F
F
Brinkman
U A
UA
Gooijer
C
C
D000804:Angiotensin II / Q000032:analysis; D019075:Electrophoresis, Capillary / Q000379:methods*; D004744:Enkephalin, Methionine / Q000032:analysis; D006801:Humans; D008163:Luminescent Measurements; D010455:Peptides / Q000032:analysis*
D010455:Peptides; D000804:Angiotensin II; D004744:Enkephalin, Methionine