Novel techniques for in vivo hemolysis studies in guinea pigs

The in vivo toxic-hemolytic studies using small experimental animals are complicated by difficulties in preventing hemolysis during repeated collection of blood specimens and in measuring hemoglobin concentration in small amounts of plasma sample. To solve these problems we tried to develope the new techniques for the in vivo hemolysis studies using guinea pigs. The hemolysis accident was minimized to 2.75 mg/dl by collecting the blood directly into heparinized microhematocrit tubes by small longitudinal incision in the auricular artery. The hemoglobin in a small amount of sample (10 microliters) was determined by the new analytical system using a microflow spectrophotometer with a modified cyanmethemoglobin method. The standard curve of the hemoglobin concentration in the system revealed a line of Y = 1.8X + 0.79 (r = 0.999), CV < 1% with a minimum detectable concentration of 1.25 mg/dl. By using the new techniques, it was found that the plasma hemoglobin concentration in normal animals were 7.27 +/- 0.44 mg/dl (mean +/- S.E.). The in vivo hemolytic activity of saponin was observed dose-dependently at doses of 30-50 mg/kg, i.v. in the guinea pigs. It is concluded that the present techniques are useful for in vivo hemolytic studies in small experimental animals such as guinea pigs.