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Indomethacin in Serum

Indomethacin was analyzed using Asahipak ODP-50 6E (a column for reversed phase chromatography). Indomethacin is a steroid-free, anti-inflammation drug used for rheumatism, inflammation of arthritis and podagra and is known to have strong side effects on the central nervous system and digestive organs. This is an example of a high sensitive analysis of indomethacin by using the principle that it converts to a fluorescent substance by alkaline hydrolysis at high temperature.
1) Use an ODP column with an eluent of pH10.
2) Set a heating coil at postcolumn.
3) Only one pump is necessary. Since the eluent is alkaline, postcolumn mixing is unnecessary.
4) The indomethacin is converted into fluorescent substance by alkaline hydrolysis in the heating coil, a highly selective and sensitive analysis is possible.
(Sample pre-treatment)
1) Add 20 μL of CH3CN to 20 μL of serum
2) Stir (2 to 3 sec)
3) Centrifuge (10,000G , 2min)
4) Skim supermatant and inject

Sample : Indomethacin in Serum

Column : Shodex Asahipak ODP-50 6E (6.0mmI.D. x 150mm)
Eluent : 25mM Sodium phosphate buffer(pH10.0)/CH3CN=65/35
Flow rate : 1.0mL/min
Detector : Fluorescence detecotr(Ex.298nm , Em.375nm)
Column temp. : Room temp. Post
Column : Heating coil at 140°C is followed with cooling coil reaction at 15°C
H. Kubo, Y.Umiguchi, T.Kinoshita, Chromatographia Vol.33, No.7/8, 1992