Phosphate-AscorbicAcid

Ammonium molybdate and potassium antimonyl tartrate react in acid medium with orthophosphate to form a heteropoly acid - phosphomolybdic acid - that is reduced to intensely colored molybdenum blue by ascorbic acid. This method for measuring phosphate is based on APHA Standard Method 4500-P E

Reagents and Equipment

• Assembled colorimeter

• Cuvettes

• 5 mL screw capped tubes

• Sulfuric acid, (14% v/v), Labchem, Cat # LC256002

• Potassium antimonyl tartrate solution, Labchem, Cat # LC187201

• Ammonium molybdate solution (4%), Labchem, Cat # LC112101

• Ascorbic acid, Labchem, Cat # LC115309

• 1mL adjustable micropipette

• Distilled water. Optional, for diluting water sample

Colorimeter set-up

• Wavelength: 625 nm. Use Red led (colorimeter RGB board).

• Phosphate standard data: download a data file or prepare your own phosphate standard data

• Before measuring samples, calibrate the colorimeter against water or a distilled water sample developed with ascorbic acid reagent as described below

Method

Step 1: Prepare fresh ascorbic acid reagent

• In a beaker dissolve 0.88 g of ascorbic acid in 50 mL distilled water;

• In a separate beaker add the reagents in the following order:

  1. 25 mL sulfuric acid solution;

  2. 2.5 mL potassium antimonyl tartrate solution;

  3. 7.5 mL ammonium molybdate solution;

  4. 15 mL ascorbic acid solution (prepared in step 1);

• Mix the ascorbic acid reagent. The reagent is stable for 4 hours.

Step 2: Sample testing

• Fill an empty sample bottle to the 5 mL line with your water sample

• Add 800 µL of test reagent solution, cap and invert several times to mix

• Note the time since adding the reagent to your water sample. When it has been at least 5-10 minutes, pour some of your sample into a cuvette, place in the colorimeter and click measure.The program will return the phosphate concentration in ppm.