チオバルビツール酸法による血清過酸化脂質の測定：比色法と蛍光法の比較並びに二価鉄の影響

Serum lipid peroxides were measured by a thiobarbituric acid (TBA) method. TBA color reactions carried out under the conditions with acetic acid (TBA-AA) or hydrochloric acid (TBA-HCl) systems, and then TBA value was detected by colorimetric ([OD]) and fluorometric ([FR]) measurements. Effects of ferrous ion, ferric ion and EDTA on the both systems were examined. Equal TBA value dependent upon concentrations of MDA (tetraethoxypropane) were obtained on both systems detected by [OD], but the relative fluorescence of TBA-AA system was higher than that of TBA-HCl system. Fluorometric determination was suitable for low TBA value, and colorimetric one was for high TBA value. Good correlation between TBA[OD] values and TBA[FR] values of serum from patients was estimated, but it was suggested that effects of serum substance were differ on the both systems. Effects of iron on TBA-AA and TBA-HCl systems were different significantly, particularly high Fe-TBA values obtained from TBA-AA system in the presence of ferrous ion. The fact suggests that serum lipid peroxidation is induced by ferrous ion during a heating of TBA color reaction. Same effect of ferric ion in TBA-AA system was observed. The increment of TBA values by ferrous and ferric ions was inhibited by a addition of EDTA. It was considered that the difference effect of iron on the both systems was due to pH and acetic acid concentration. The relationship of TBA values and Fe-TBA values of serum from patients was no correlated, and the various inhibition ratios of EDTA for the TBA value was calculated. It is suggested that Fe-TBA value is possible to detemine the qualitative difference of serum lipid peroxides.

患者血清を試料に脂質過酸化物をTBA法を用い、酢酸酸性下(TBA－AA系）、塩酸酸性下（TBA-HCl系）で加熱反応させ、比色法（TBA[OD]値）と蛍光法（TBA[FR]値）で計測した。併せてFe(++)（Fe-TBA値）、Fe(+++)、EDTAの影響を検討した。両系においてMDAの検量を行った結果、TBA[OD]値は同一数値が得られたが、TBA[FR]値は異なって示された。低TBA値では蛍光法が、高TBA値では比色法が適している結果をみた。しかし、患者血清の定量からはTBA[OD]値、TBA[FR]値で相関は得られたものの、両測定方法に関与する血清内在因子の影響は異なることが示唆された。TBA-AA,TBA-HCl両系に対するFeの影響は異なり、特にTBA-AA系では著しいFe-TBA値が得られ、Fe誘導脂質過酸化反応が加熱発色反応中に誘起されていることが示されたが、EDTA添加でその誘導は抑制された。また、TBA-AA系ではFe(++),Fe(+++)で同様の影響がみられたが、TBA-HCl系では異なった影響がみられた。両系のTBA値、Fe-TBA値の相違の原因としてpH、酢酸の影響が考えられた。患者血清でTBA値とFe-TBA値に相関は得られず、Fe(++)誘導反応に相違がみられ、また、EDTAの抑制作用も血清試料で異なることから、Fe-TBA値で血清の質的な相違が計測できる可能性が示唆された。