AMMONIUM MOLYBDATE SOLUTION

Prepare as described in Part 11.2.1, Section E 1.

53

54 A PRACTICAL HANDBOOK OF SEAWATER ANALYSIS 2 . SULPHURIC ACID SOLUTIO N

Prepare as described in Part 11 .2 .1, Section E 2 .

3 . ASCORBIC ACID SOLUTIO N

Prepare as described in Part 11 .2 .1, Section E 3 . 4 . POTASSIUM ANTIMONYL-TARTRATE SOLUTION Prepare as described in Part 11 .2 .1, Section E 4 . 5 . MIXED REAGEN T

Prepare as described in Part 11 .2 .1, Section E 5 . 6 . ISO-BUTANOL

Wash good quality iso-butanol with two small aliquots of distilled water, about 150 ml water to 1000 ml iso-butanol . Dry the alcohol over potassium carbonate and distill, collecting the fraction boiling between 105 C and 109 C (corrected) .

7 . GLASS REDISTILLED WATE R 8 . ABSOLUTE ETHANOL

F. EXPERIMENTAL

PROCEDURE

1 . Warm the samples to a temperature between 15 C and 30 C (Note a) . 2 . Measure 200- :L2 ml of sample with a graduated cylinder and pour into a 250-ml separatory funnel .

3 . To the sample in funnel add 20-!-1 ml of mixed reagent from a 25-ml graduated cylinder and mix at once .

4 . After 10 min and within the next 2 hr add 28-!-0 .2 ml iso-butanol and shake the funnel for 60 sec (Note b) .

5 . Allow the funnel contents to stand for 5 min and separate off the lower aqueous phase .

6 . Run the iso-butanol fraction into a clean, dry 10-ml graduated cylinder allowing 10 sec for the funnel to drain .

7 . Make the alcoholic extract up to 10-}0 .1 ml with absolute ethanol and mix by swirling (Note c) .

8 . Measure the extinction of the extract in a dry 10-ml, 10-cm absorption cell against iso-butanol at a wavelength of 6900 A . If a filter-type absorptiometer is used, choose a filter having a maximum transmission in the region of 7000 A .

9 . Correct the measured extinction by subtracting the reagent blank (see Sect . G) . Calculate the phosphate concentration in microgram atoms of phosphate phosphorus per liter (µg-at P/liter) from the expression :

µg-at P/liter = corrected extinction x F

where F is a factor obtained as described in Section H . Report results to three significant figures . Silicon and arsenic cause negligible interference .

NOTES

(a) The method appears to have no significant temperature coefficient (less than 0.2% per

1I.2.II. REACTIVE PHOSPHORUS (LOW LEVELS) 55 1 C) between 15 C and 30 C but it is wisest to have samples at a temperature within this range.

(b) Erratic results may occur with less than 60 sec vigorous shaking. The extracted colour is stable in the alcoholic phase for at least 2 hr. Iso-butanol is more soluble in distilled water than in sea water and 28 ml has been chosen to give a recoverable extract volume not exceeding 9.5 ml in water of 33%o salinity.

(c) The addition of ethanol removes water droplets.

G. DETERMINATION OF BLANKS 1. CELL-TO-CELL BLANKS

When both sample and reference cells are filled with iso-butanol the extinction of one against the other should be 0.000. Due to slight optical defects a slight positive or negative value may be found. This is allowed for when reagent blanks are subtracted but the value should be determined.

2. REAGENT BLANKS

Carry out the method exactly as described in Section F, paragraphs 2-9 inclusive, using redistilled water in place of the sample. The reagent blank corrected for cell-to-cell blank should not exceed 0.100. If the blank exceeds this amount either the molybdate reagent or the iso-butanol is suspect. The reagent blank should be determined for each batch of samples measured.

H. CALIBRATION

1. STANDARD PHOSPHATE SOLUTION

Dissolve 0.816 g of anhydrous potassium dihydrogen phosphate, KHZPO„

in 1000 ml of distilled water. Store in a dark bottle with 1 ml of chloroform. The solution is stable for many months.

1 m1- 6.0 µg-at P

Dilute 5 ml of this solution to 1000 ml with distilled water. Store in a dark bottle with 1 ml of chloroform. Fresh dilute standards should be made up once a week.

1 ml - 0.03 µg-at P

or 1 ml - 0.15 µg-at P/liter in 200-ml water sample

2. PROCEDURE

Prepare four standards consisting of 1.0 ml of dilute phosphorus solution and 200 ml of redistilled water. Prepare two blanks using redistilled water. Carry out the determination exactly as described in Section F, paragraphs 2-9.

Calculate the factor F from the expression:

F 0.15 E8 -Eb

where E. is the mean extinction of the four standards and Eb is the mean extinc-tion of the two blanks (not corrected for cell-to-cell blanks). The value for F is independent of salinity, but for most precise work in determining phosphate concen-trations in sea water use a synthetic sea water in place of redistilled water as a larger volume of extract is obtained. The value for F should be near 0.375.

11.3. DETERMINATION OF TOTAL PHOSPHORUS INTRODUCTION

In this method the total phosphorus, in a filtered or unfiltered sample, is mineralized by evaporating the sample to fumes with perchloric acid. We have had no experimental evidence to indicate that any form of combined phosphorus in sea water can withstand this treatment without being mineralized and brought into solution. The method is taken, basically, from the procedure described by Hansen and Robinson (J. Marine Res., 12: 31, 1953) but there have been several major modifications designed to give a better "blank" determination, to facilitate the routine use of the method, and to eliminate any interference from arsenate.

METHOD A. CAPABILITIES

Range: 0.08-6 pg-at/liter

1. PRECISION AT THE 2.5 MG-AT/LITER LEVEL

The correct value lies in the range:

Mean of n determinations ±0.1 3/& gg-at/liter.

2. PRECISION AT THE 0.5 MG-AT/LITER LEVEL

The correct value lies in the range:

Mean of n determinations ±-0.08/ni Ag-at/liter.

B. OUTLINE OF METHOD

A 50-ml sample of sea water is evaporated with perchloric acid. Chloride is replaced by perchlorate and much of the arsenic is volatilized. The residue is heated and any organic matter is oxidized, liberating phosphorus as inorganic phosphate.

The total phosphate is then determined, after adding 50 ml of water, as described in Part 11.2.

C. SPECIAL APPARATUS AND EQUIPMENT

Use 125-ml Pyrex Erlenmeyer flasks. Three small dents, 1-2 mm deep, are made symmetrically around the lip of each flask which is covered with a 2-inch diam coverglass.

Clean Pyrex Erlenmeyer flasks have been found to give no phosphate con-tamination. Flasks should be retained especially for the present method, covered by aluminium foil, and not put into general circulation. Before their initial use, they should be cleaned by fuming after the addition of about 30 ml of water, 20 ml of concentrated hydrochloric acid, and 10 ml of perchloric acid solution. The perchloric acid is refluxed vigorously in each flask for about 1 hr. A blank determination (see Sect. G below) should then be carried out on each flask before it is finally put into use. The flasks are rinsed thoroughly with distilled water after each determination and stored dry.

Antibumping devices consisting of carborundum granules, sized about -1—*

57

58 A PRACTICAL HANDBOOK OF SEAWATER ANALYSIS

inch and cleaned by fuming with perchloric acid for 1 or 2 hr and then rinsing thoroughly with hot distilled water. The granules should only be handled with forceps and two used with each flask. The granules and 2-inch coverglasses should be boiled for a few minutes in 50% v/v hydrochloric acid before use or after a prolonged shutdown period.

D. SAMPLING PROCEDURE AND SAMPLE STORAGE

If the total phosphorus is to be determined on Millipore-filtered samples the filtration must take place immediately after the samples are taken from the sea.

It appears to be impossible to prevent a very small, but detectable, pick-up of phosphorus from glassware over a sufficient period of time but Pyrex boiling tubes, 200 x 25 mm holding about 80-90 ml, have proved satisfactory for storing samples for total phosphate for a period exceeding a month. The tubes are cleaned thoroughly in hot chromic—sulphuric acid for several hours, rinsed thoroughly with distilled water and conditioned by being allowed to stand full of slightly acidic water (1 drop of concentrated hydrochloric acid) for several months. When not in use the tubes should be kept nearly full of acidic water and covered with a clean square of Parafilm or aluminium foil. In no circumstances should a cork or rubber stopper be used as either can introduce contamination. This water is emptied, and the tube drained as dry as possible, immediately before the sample is placed in it for storage.

Exactly 50 ml of sample is pipetted into the tube and 1-2 drops of concentrated hydrochloric added to acidify the seawater sample during storage. The tubes should be re-stoppered immediately with Parafilm sheeting. The very slight phosphorus pick-up which may still occur during a longer cruise ^(Ca. 0.1 btg-at P/liter or less) can be minimized by storing the boiling tubes in a deep-freeze and by assessing a mean blank correction. This is obtained by storing synthetic sea water of known total phosphorus content in three or four tubes selected at random and redetermining the total phosphorus when the batch of samples is analysed.

Probably the best technique in phosphorus analysis is to do any filtrations within 1 lr of taking samples (which are kept cool and dark) and then quickly deep-freeze filtrates to —20 C in polyethylene bottles. All analyses should be carried out immediately the samples are quick-thawed and no refreezing and thawing should be permitted.

E. SPECIAL REAGENTS