Calcium precipitation from mammalian physiological salines (Ringer solutions) and the preparation of high [Ca] media

The maximum nonprecipitating concentration of calcium in a bicarbonate-buffered Krebs-Henseleit-type mammalian physiologic saline at 25 degrees C has been established as 10 mM in the presence of 1 mM phosphate, and as 18 mM when phosphate is absent. At 38 degrees C, the respective limits are 5.6 mM with the phosphate, and 16 mM in its absence. At its physiological concentration of 1.6 mM, calcium precipitation also appeared in solutions of pH greater than 9. When phosphate was present, the precipitate appeared immediately, but redissolved once pH was reduced below 7.8; in the absence of phosphate, precipitation of calcium carbonate became visible only after 3-4 min, but this precipitate would not redissolve even at pH 7.4. As carbonated stock bicarbonate solution gradually loses CO2 and becomes alkaline, a protocol for mixing physiologic salines using alkaline bicarbonate stock was developed. Calcium precipitation can be avoided if, before calcium is added, the bicarbonate-containing bulk solution is gassed with 5% CO2 for 15 min at a flow of 0.5 liter/min per liter of final solution.