Über eine titrimetrische Mikrobestimmung des Cholesterins mit Hilfe der Blutkörperchenhämolyse und ihre Anwendung auf Serum

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Abstract
About 5 ml. of human blood are placed in a graduated centrifuge tube containing 30 mg. Na citrate. The plasma is removed, the red cells thoroughly washed with 09% NaCl soln., and suspended in such an amt. of 0.9% NaCl that the total vol. is 10 times the amt. of blood placed in the centrifuge tube. The digitonin soln. is prepared by dissolving 150 mg. in 0.9% NaCl-M/15 phosphate buffer soln., pH 7 (3:1). One ml. of the digitonin soln. is placed in a small test tube, which is kept in the H2O bath at 40[degree]. A 2 ml. micro burette is filled with the red cell suspension and the suspension added to the test tube drop by drop until a turbidity which persists 2-5 min. appears. Hemolysis ceases because of the combination of the digitonin with the cholesterol of the red cells. Since the digitonin soln. usually has a titcr of 46.7 [gamma] cholesterol/ ml., if 1 ml. of the digitonin soln. requires 0.87 ml. of the cell suspension for the titration, then the suspension contains 46.7/0.87 = 53.7 [gamma] cholesterol/ml. Since the dilution of original red cells is 10 times, the cells contain 53.7 mg. % cholesterol. Free cholesterol in serum and body fluids is detd. by determining how much an aliquot of diluted serum or fluid decreases the hemolysis. The amt. of cholesterol is detd. by back titration of the excess digitonin. Total cholesterol is detd. in a similar manner after saponification of the extracts of the fluids. The method requires simple apparatus, is rapid, is accurate within [plus or minus] 5-10%, and can detect 1 [gamma] cholesterol. Free cholesterol may-be detd. in 15 min., and total cholesterol in 1 hr. Values for cholesterol obtained agree well with those reported in the literature.