Separation of bilirubin from hemoglobin by recording derivative spectrophotometry

Recording derivative spectrophotometry is a technic for resolving overlapping absorbance curves by enhancing small changes in those curves. The derivative (slope) is a function sensitive to rapid changes in the curve and will amplify those regions while damping out slowly changing portions of curves. The method presented relies on a mathematical analysis of the composite absorption curve formed by bilirubin and hemoglobin. Data confirming the usefulness of this technic for reliably and quantitatively separating these two substances in aqueous solution are presented. Recording derivative spectrophotometry may be useful for analyzing amniotic fluid, urine, or other biologic fluids.