STABILIZATION OF THE PHOSPHATE RATIO OF SEA WATER 



BY FREEZING 



By Albert W. Collier, Fishery Research Biologist 

 and Kenneth T. Marvin, Chemist 



It is the accepted practice in oceanographic 

 investigations to make all chemical analyses, ex- 

 cept salinity determinations, at sea. This is neces- 

 sary because there has been no way in which 

 samples could be stabilized for the relatively long 

 period intervening between the collection of sam- 

 ples and the return to base. Only a limited num- 

 ber of analyses can be performed at sea, as a 

 rolling, vibrating ship is not suitable for the deli- 

 cate instruments required to make many of the 

 chemical analyses. The chemists assigned to 

 oceanographic investigations have thus been forced 

 to make determinations under conditions unfav- 

 orable to producing the best quality or the greatest 

 quantity of work. 



Since the program of the Gulf Fishery Investi- 

 gations of the United States Fish and Wildlife 

 Service necessitated expansion of chemical investi- 

 gation into the fields of organic chemistry and of 

 biochemistry, it was essential that we find some 

 method of overcoming the difficulties to making 

 chemical analyses at sea. We desired to increase 

 the precision and quality of work and to take 

 advantage of the gain in productive manpower 

 inherent in chemists working ashore, so as to 

 make an intensive study of the phosphorus econ- 

 omy of the sea, reexamine the nitrate problem, 

 and study the organic compounds present in sea 

 water. 



This paper reports the first of a series of exper- 

 iments to test the stability of certain organic and 

 inorganic complexes in sea water after the sam- 

 ples have been quick-frozen. The analysis for 

 inorganic phosphorus is universally used in oce- 

 anography and marine biology and is very sensi- 

 tive to changes in biological activity. Accordingly, 

 we selected it as the first of a series for testing 

 and reporting. 



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TECHNIQUES OF FREEZING AND 

 ANALYSIS 



Sea water was collected from Offat's Bayou, a 

 small inlet on the bay side of Galveston Island, 

 and from the open beach of the Gulf of Mexico 

 near the laboratory. Normally, the waters of 

 Offat's Bayou are richer in phosphate than are 

 the waters of the Gulf of Mexico. The bottles 

 found most satisfactory for this work were 4-ounce 

 prescription bottles with molded plastic screw 

 caps. They withstood the freezing well with only 

 occasional breakage. 



Freezing was accomplished by immersing the 

 sample bottles in a bath of dry ice and ethyl alco- 

 hol. Preliminary tests demonstrated that ordi- 

 nary refrigeration did not stabilize the materials, 

 and that even freezing in the ice-tray section of 

 an ordinary refrigerator was too slow. The dry- 

 ice-and-alcohol bath made an easily portable 

 arrangement, and froze the samples quickly 

 enough to be effective. 



Conditions aboard ship made certain modifica- 

 tions of this method necessary. Samples to be 

 frozen were collected in glass culture tubes (25 x 

 200 millimeters) fitted with molded plastic screw 

 caps. To speed the freezing of sea-water sam- 

 ples and to save time on shipboard, the sample 

 tubes were prerinsed three times with distilled 

 water and thoroughly drained in the shore labora- 

 tory before the ship sailed. Before the samples 

 were put in the freezing bath the tubes were 

 tightly sealed at the junction of cap and tube 

 with plastic insulating tape. This prevented con- 

 tamination by the freezing solution or by the 

 water of the bath during thawing. 



Freezing was done in a vat of 38 percent 

 ethylene glycol held at an average temperature of 

 — 5° F. in a commercial quick-freeze cabinet in- 



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