Determination of free morphinen and human haemoglobin Binding in vitro / 中国药理学通报

ABSTRACT

AIM To develope a reversed phase high performance liquid chromatography method to determine free concentrations of morphine (M) in the solution of human haemoglobin (Hb). To study the binding of M to haemoglobin, and evaluate the binding parameters of M to Hb. METHODS An ultrafiltration technique was used to recover morphine from the samples. Morphine was analyzed using a kromasil column (150 mm?4 6 mm) and a mobile phase of 0 1% tyiethylamine methanol (75∶25,v/v). The mobile phase pH was adjusted to 7 0 by phosphoric acid. The detection was set at 283 nm. RESULTS The ultrafiltration recovery of morphine was 98 5%. The Hb binding of M was concentration dependent of Hb and M. There were single typed binding sites for M to human Hb. The parameters determined were 4 1 for N and 340 mol?L -1 for K when the concentration of M and Hb were added from 8 50?10 -5 ～1 17?10 -2 mol?L -1 and 1 29?10 -4 ～8 57?10 -4 mol?L -1 respectively. CONCLUSION An ultrafiltration technique has proved to be simple and rapid for free drug determination. It is suitable for drug protein binding study.