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Environmental & Plant Biology
Mailing address: Porter 315

Ascorbic Acid Method for Anion Exchange Membranes

By Jared L. DeForest

For Anion Exchange Membranes (i.e., resin strips)

96-well plate method

Reagents

Solution A (2.5 M H2SO4)

In 250 ml volumetric flask

  • Fill flask half-way with DI water.
  • 35 ml Sulfuric acid (18M H2SO4)
  • Mix well, bring to volume with DI water

Solution B

In 250 ml volumetric flask

  • Fill flask half-way with DI water.
  • 10 g Ammonium molybdate
  • Mix well, bring to volume with DI water.

Solution C

In 100 ml volumetric flask

  • 50 ml DI water
  • 0.2728 g Antimony potassium tartrate
  • Bring to volume with DI water.

Working Solution

Use within a day or flush with N2 for cold storage.

In 50 ml volumetric flask

  • 25 ml Solution A
  • 7.5 ml Solution B
  • 0.264 g Ascorbic acid (1 M C6H8O6)
  • 2.5 ml Solution C
  • Bring to volume with DI water.

Working Standard

In 100 ml volumetric flask

  • 50 ml 0.5 M HCl (i.e. matrix)
  • 2.5 ml Certified standard P solution (1,000 ppm)
  • Bring to volume to make a 25 ppm working standard.

Standard Dilutions

In 15 ml centrifuge tube

  • 1 ppm 0.4 ml standard, 9.6 ml matrix
  • 5 ppm 2.0 ml standard, 8.0 ml matrix
  • 10 ppm 4.0 ml standard, 6.0 ml matrix

Procedure

In a clear 96-well plate (2:1 dilution)

  • 200 碌l Solution (in 0.5 M HCl) using the single channel electronic pipette into 4 wells.
  • 100 碌l Using the 8-channel pipette, add working solution directly into the well to mix the solution.
  • Using the same procedure as above, add the three (3) standards to the last three columns in the plate. Incubate between 18 and 24 hours (Make sure standards are linear). Read absorbance at 880 nm.

Charging AEM (Anion Exchange Membranes)

In 250 ml Flask

  • 150 ml   0.5 M NaHCO3
  • AEM strips
  • Rapidly shake for 15 minutes.
  • Remove strips and place AEM in another flask.
  • Rinse AEM with DI water three (3) times.
  • Immediately use or store at 5掳C.

In Situ Soil Phosphorus (Field deployment)

Prep Work

  • Use a large syringe needle to poke a hole in a corner.
  • Thread Firewire finishing line and tie. Have at least 20 cm of loose line.
  • Place AEM inside a plastic bag with the treads taped on the outside of the bag.
  • Keep moist and cool.

In the Field

  • Remove O horizon by hand.
  • Using a 1鈥 putty knife, insert into the soil at an angle to a vertical depth of 5 cm.
  • Slide the AEM into the hole and press the soil back to make contact.
  • Tie the end of the line to a pin flag.
  • Keep in the field between one (1) and two (2) weeks.
  • Place each individual AEM, or grouped within a plot (3), into a small bottle or plastic bag. Keep moist with DI water & remove the thread.

Extraction

Clean off any loose debris with DI water (squirt bottle).

In 150 ml Flask

Place each AEM (or group of AEM) in the flask.

  • 25 ml 0.5 M HCl
  • Shake for four (4) hours
  • Analyze extract for anions (i.e. Phosphate) 鈥 See above.

Readily Available Soil Phosphorus (Resin P)

In 50 ml centrifuge tube

  • 10 g Field fresh soil
  • 25 ml DI water
  • 1 Charged AEM strip
  • Shake for four (4) hours.
  • Take out strip and rise to remove debris, and place into another 50 ml centrifuge tube.
  • Follow extraction procedure.

Created: 04/26/11 Revised: 10/10/11