Improvements in the Methods for Calcium Determination in Biological Material

It is well known that the principal disadvantage of any method for the determination of calcium involving precipitation as calcium oxalate lies in washing the precipitate. The solubility of the precipitate in water or in dilute ammonium hydroxide is appreciable and, in centrifuging, a small amount of calcium oxalate tends to float on the surface of the liquid and is lost in decanting. The satisfactory results obtained by the oxalate method are due largely to the compensation of errors, the loss of the precipitate of calcium oxalate being balanced by incomplete removal of ammonium oxalate in the washings. It is apparent that a washing fluid which would remove the excess of oxalate solution with no appreciable solution of the precipitate and which prevents any of the precipitate from floating on the surface of the liquid would increase the accuracy of the method. It has been found that a mixture of 2 per cent ammonia in equal parts of alcohol, ether, and distilled water meets these requirements, provided that the ammonium oxalate used for precipitation is not too concentrated. 0.1 M ammonium oxalate is substitutued for the usual saturated ammonium oxalate (4 per cent). This is about one-third the usual concentration but insures an adequate oxalate concentration for complete precipitation of calcium oxalate. Two washings with the ammoniacal alcohol-ether-water mixture are needed but there is no appreciable loss of calcium with three washings. Three washings are not sufficient when more concentrated ammonium oxalate is used. In applying the new washing solution to calcium determinations we have used Kramer and Tisdall’s (1) permanganate titration,