PHOSPHORUS 279 



to separate P^- from the extraneous mass of a biological sample merely 

 for counting purposes. 



Chemistry. Standard methods for estimation of piiosi)horus in various 

 biological materials have been described in the literature (P-2 to P-4). 

 Usually an aliciuot of the ash solution can be used for radioassay, or the 

 entire ash solution can be counted and then used for chemical estimation 

 of phosphorus. Plant and animal samples can be dry-ashed at 500 to 

 600°C, usually with the addition of magnesium nitrate, and the ash dis- 

 solved in dilute HCl. Wet-ashing may be done with 30 per cent Il2C)2 in 

 H2SO4 solution. The H2O2 must be phosphorus-free and must be com- 

 pletely removed from the final ash solution. The latter can be accom- 

 plished by adding distilled water and evaporating to white fumes in the 

 final step. The phosphorus in an ahquot of the ash solution can be esti- 

 mated colorimetrically by various modifications of the Fiske Subbarow 

 method (P-3) : An aliquot of the ash solution, usually containing of the 

 order of 0.2 mg P, is mixed in order with a molybdate reagent and an 

 aminonaphthol sulfonic acid reagent and is then diluted for colorimetric 

 evaluation after a standard time interval. Plasma is treated with tri- 

 chloroacetic acid, and the filtrate used directly for development of the 

 molybdenum blue color as above. 



Typical Methods. In studies with dairy cows (P-5), 40 mc P*^ was 

 injected intravenously, and casein containing 2 juc/g was prepared from 

 milk collected during the 3 days postinjection. For counting, 0.1-ml 

 samples of milk were pipetted onto disks of lens paper, dried on an alu- 

 minum disk, and fixed with a spray of an alcoholic shellac dispersion. For 

 ordinary distribution studies the dosage for cattle might range from 2 to 

 10 mc. Labeled casein was used to study the digestion of this compound 

 by the calf and also to estimate the fecal phosphorus excretion of calves; 

 27 to 72 fxc of P^--labeled casein was fed to 55- to 140-lb animals, and 

 radioactivity was determined on both fresh and dry-ashed samples (P-6, 

 P-7). In tissue distribution studies with swine, 1.5 to 5 mc was injected 

 intravenously into 25- to 180-lb animals (P-8). For the estimation of 

 endogenous fecal phosphorus in dairy cattle (P-9), about 1.3 mc P^- was 

 injected intravenously twice daily for 23 days. In work with poultry 

 about 300 /xc was given orally to laying hens (P-10), and about 20 /xc to 

 chicks (P-11). One to ten microcuries is adequate for most studies with 

 mice and rats. In a study of P'^'- in the embryo and larva of frogs, fer- 

 tilized eggs were maintained in solutions of 65 to 550 /xc/750 ml (P-12). 

 The utilization of P^^ by developing chick embryos has been followed by 

 injections of 0.2 to 5 /xc per egg (P-13, P-14). 



liabeled mosquitoes were produced by treatment of late third — and 

 fourth — instar larvae with about 0.1 /xc P^^ per larva in 1 ml water (P-15). 

 Adult mosquitoes were tagged by allowing them to feed on a rabbit that 



