Separation of phenolic compounds and corresponding glucuronides by coupled-column micellar reversed-phase liquid chromatography |
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Authors: | B Lu M Koimur D Westerlund |
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Institution: | (1) Analytical Pharmaceutical Chemistry, Uppsala University Biomedical Cetre, P.O. Box 574, 751 23 Uppsala, Sweden |
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Abstract: | Summary An isocratic HPLC technique for separation of phenolic compounds and corresponding glucuronides in urine is developed. Sample
pre-treatment, often a tedious and rate-limiting factor, was eliminated by use of a coupled column system. Spiked urine samples
were injected directly into a C4 precolumn and a selected fraction was transferred on-line from the precolumn to a silanized C18 analytical column in the backflush mode. Analyte peak enrichment was attained by employing mobile phases of different elution
strengths. The weaker mobile phase (7% v/v acetonitrile) was used to strongly retain the analytes on the precolumn while most
of the polar endogenous compounds were washed to waste. Elution and transfer of the trapped analytes from the precolumn to
the analytical column was achieved by introducing a stronger mobile phase (20% v/v acetonitrile) with the aid of a switching
valve. The use of cetyltrimethylammonium bromide as counter ion and micellar agent in the mobile phase involved a high selectivity
for the analytes relative to the urine matrix components and allowed simultaneous analysis of the glucuronides and parent
compounds without the need of gradient elution. The system demonstrated a good repeatability on spiked urine samples.
Who passed away July 21, 1996 |
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Keywords: | Micellar liquid chromatography Coupled-column system Direct injection of urine samples Phenolic compounds Glucuronide conjugates |
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