Glyoxal bis (2-hydroxyanil) (GBHA) is known as a sensitive and relatively selective complexant for the spectrophotometric determination of calcium. The pink Ca-GBHA complex in alkaline solution can be measured in alcohol-water or chloroform medium [1,2,31. Accordingly, the colored complex is extracted immediately after being formed and measured before hydrolysis begins. In a recent modification [4] the stability of the colored extract has been increased up to a 1 hr period.In our method, the color is developed and measured in potassium hydroxide-absolute ethanol, the extraction stage thus being unnecessary. In the solvent suggested the color is stable for at least 24 hours.
ExperimentalApparatus and Reagents: Model SP 600 Unicam spectrophotometer was used to measure absorption. Certified Lange-Levy ;>ipets were used to deliver 50-and 250-~1 volumes. Calcium standards were prepared from calcium carbonate and hydrochloric acid. GBHA (O.40/0, Baker) was prepared in absolute ethanol. Alcoholic potassium hydroxide solution was made by dissolving 10.0 gm of carbonate-free potassium hydroxide in 100 m1 of absolute ethanol. It was stored in polyethylene bottles. All chemicals used were of reagent grade quality. Procedure: Transfer an aliquot of sample containing not more than 12~g of calcium to a 50 ml beaker. Evaporate the solution to complete dryness in an oven at 100°C; cool. Add 20.0 ml absolute ethanol, 250~l GBHA reagent solution, 50~l alcoholic potassium hydroxide (in that order) and mix after each addition. Transfer immediately to stoppered bottles of such volumes that the container is full of the solution with minimum air space. Set aside for 30 min. and then measure the optical density in covered 4-cm cells at 550~. Prepare a blank in the same manner, but omitting the calcium, and use as reference solution.