Highly sensitive high-performance liquid chromatography for the measurement of malondialdehyde in biological samples.
A highly sensitive method for the measurement of malondialdehyde as thiobarbituric acid-malondialdehyde complex by reversed-phase high-performance liquid chromatography with fluorescence detection in biological samples is described. As samples, 20 microliters of rat plasma or 10% (w/v) liver homogenate mixed with 2.0% thiobarbituric acid in 2 M sodium acetate buffer containing 0.05% butyl hydroxytoluene (pH 3.5) were heated at 95 degrees C for 45 min to give the complex. The complex, extracted with n-butanol, was chromatographed on a system equipped with a reversed-phase column, and the eluted peak was monitored with a fluorescence detector (excitation wavelength 515 nm, emission wavelength 553 nm). The mobile phase was a acetonitrile-water (2:8, v/v) under isocratic conditions at ambient temperature, and a single analysis was done in ca. 4 min. The minimum detection level for malondialdehyde was as low as 0.05 pmol. The n-butanol extract was stable at least for 3 days. The simple mobile phase, the extremely sensitive detection limit, and the stability of the complex make this system applicable to routine clinical analysis with a small amount of tissue or biopsy sample.
Fukunaga
K
K
Department of Food Science and Technology, Faculty of Fisheries, Hokkaido University, Japan.
Suzuki
T
T
Takama
K
K
D000818:Animals; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D008099:Liver / Q000737:chemistry; D008297:Male; D008315:Malondialdehyde / Q000032:analysis* / Q000097:blood; D051381:Rats; D017208:Rats, Wistar; D012680:Sensitivity and Specificity; D013050:Spectrometry, Fluorescence; D013858:Thiobarbiturates
D013858:Thiobarbiturates; D008315:Malondialdehyde; C029684:thiobarbituric acid