Titrimetric estimation of calcium with disodium ethylene diamine tetra-acetate (E.D.T.A., sodium edetate), as performed on serum (Baron and Bell, 1957), cannot be applied directly to most other biological specimens because of their high concen-tration of phosphate. A high ratio of phosphate to calcium in the material titrated causes a drawn-out and inaccurate endpoint. A rapid procedure for calcium determination has been devised which gives accurate results after separation of phos-phate. METHOD Reagents (1) Deionized or glass-distilled water must be used throughout all stages of the procedure. (2) For Ashing.-N-HCl is used. (3) For Phosphate Separation (Horner, 1955).-(a) 48 % morpholine nitrate: 50 o v/v HNO3 is slowly added to 28 ml. morpholine at 0 ° C. until the solution is neutral or slightly acidic to B.D.H. universal indicator paper (about 40 ml. acid is required). The solution is then diluted to 100 ml. with water. (b) The morpholine nitrate-nitric acid reagent is a 1:1 mixture of 48 morpholine nitrate and 25% v/v HNO3. (c) 66 % w/v sodium tungstate Na2WO42H2O. (4) For Titration.-The following reagents are made up: (a) 40 % w/v NaOH. (b) Glycine buffer pH 12.66 of 20 ml. of 0.1
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Baron et al. (1959) studied this question.