Two compounds were usually employed 

 in the preparation of each series of stand- 

 ard samples. We discouraged the practice 

 of using stock solution more than once. The 

 practice of starting each series of standard 

 samples from the "dry" compounds rather than 

 from preserved stock solutions, and also of 

 using two different compounds to a series 

 of standard samples, enabled the analyst to 

 detect systematic errors that could reduce 

 the accuracy of his data. 



To correct for glassware and reagent 

 contamination, blanks consisting of "pure" 

 water were treated in the same manner as 

 unknown samples. Any color that developed 

 was due to impurities which were corrected 

 by subtracting the blank sample readings 

 from those of the regular samples. System- 

 atic errors can be introduced by using 

 a low quality distilled water in preparing 

 blank samples. We have found that a satis- 

 factory grade can be prepared by distilling 

 tap water two times in a borosilicate glass 

 still. The arrangement has been described 

 by Marvin and Lansford (1958). 



Phosphate 



We used the method of Robinson and 

 Thompson (1948a) for the determination of 

 inorganic phosphate, and that of Harvey 

 (1948) for the determination of total phos- 

 phate. To make total and inorganic analyses 

 on aliquots of the same sample, we used Har- 

 vey's method for both analyses, but, in this 

 case, did not digest the inorganic samples. 



The mechanics of the phosphate deter- 

 minations are as follows: 



Inorganic Only 



1. Remove frozen samples from cold 

 storage and thaw by placing in front of an 

 electric fan. Do not thaw more than 50 

 samples at a time. 



2. While samples are thawing, set up 

 enough 50-milliliter pyrex Erlenmeyer flasks 

 to take care of duplicate determinations 

 from each vial. 



3. With a 2-milliliter automatic 

 pipette, add 0.25 ml. of molybdate reagent 

 to each flask. 



4. When the samples have thawed and 

 warmed to room temperature, shake them 



vigorously. Clean each cap-glass junction 

 with a jet of distilled water, remove caps, 

 and dispense 25-milliliter portion into the 

 sample flasks with a large bore 25-milli- 

 liter graduated pipette. 



5. Add 1 drop of stannous chloride 

 reagent to the first sample flask and then 

 swirl the flask thoroughly. Thirty seconds 

 later, add a drop to the second flask and 

 swirl. Continue at 30-second intervals 

 until all flasks have been treated. 



6. Determine the density of the blue 

 color that will develop if there is any 

 phosphate present with a Fisher a.c. elec- 

 trophotometer equipped with a 650 A filter 

 and a 23-milliliter cell. Color-density 

 determinations are made 30 minutes after the 

 addition of the stannous chloride reagent. 

 If analyst cannot check a sample every 30 

 seconds, increase the time interval in item 

 5 as necessary. 



7. Convert the density units (-log T) 

 into concentration units (jig at PO^-P/l) by 



dividing by the slope ■ at^°po p/i obtained 



by running known standards along with regu- 

 lar samples. 



Reagents : 



a. Molybdate reagent. Add 1 

 part of a 10-percent ammonium molyb- 

 date solution to 3 parts of 18N H2SO4. 



b. Stannous chloride reagent. 

 Dissolve 1 gm. of stannous chloride 

 in 5 ml. of 12N HCl (if necessary, 

 heat gently until acid solution 

 clears) and dilute to 100 ml. with 

 triple-distilled water. 



Total and Inorganic 



1. Remove about 50 frozen samples at 

 a time from cold storage and thaw by placing 

 in front of an electric fan. 



2. While samples are thawing, set up 

 two sets of 50-milliliter pyrex Erlenmeyer 

 flasks. One set will be used for inorganic 

 and the other for organic analyses. 



3. With a 2-milliliter automatic 

 pipette, add 0.38 ml. of 18N HjSO^ to each 

 flask. Cover immediately with 30- or 50- 

 milliliter beakers. 



