Phosphate Exp

EXP 2: DETERMINATION OF PHOSPHATES
- Spectrophotometric Determination of Phosphates in Water Background Controlled

concentration of phosphorus in water is critical for a stable ecosystem. Aquatic plants are
dependent on a certain amount of phosphorus in order to survive; however, excess
phosphorus leads to eutrophication, the over enrichment of a water body with nutrients. This
can lead to a surge in biomass growth manifesting as an algal bloom that blocks sunlight from
other aquatic plants and starves the water of oxygen adversely affecting the environment.
The EPA Water -
Quality Criteria recommends a maximum phosphorus concentration of 100 µg/L in rivers
and streams and 25 µg/L in lakes in order to prevent eutrophication from occurring.
- Phosphorus is also found in tap water where it is added in order to inhibit corrosion of
piping - a thin layer of phosphorus builds up on the inside of the pipes and prevents any
metals from seeping into the water.
- A maximum phosphorus concentration of 0.14 µg/L is mandated by the California Water
Resources Control Board.
- In this experiment, you will measure phosphorus concentrations in water from several
locations by spectrophotometry with a molybdate complex that turns dark blue in the
presence of phosphorus.
- The phosphorus cycle is a simple method to comprehend the role phosphorus plays in the
environment. Since phosphorus has limited methods to enter vapor form, the cycle
predominately moves through organisms and ground/water phases.
- Phosphorus is essential to various biological compounds in organisms, such as DNA and
proteins.
When organisms die, the organic phosphorus taken up by the organism returns to the soil.
Various bacteria and fungi initiate processes that then break down the phosphorus which is
either picked up by ground water or rainfall or swept into rivers and lakes, or it may build up as
sediment in a mineralization process leading to phosphorus rich rocks and minerals that may
erode later on. Alternatively, plants can uptake the phosphorus from the soil directly.
- Phosphorus is present mainly in the form of phosphates. These phosphates exist in three
forms.
Organic phosphates exist in biological systems and play a role in the life cycle of the organism.
An example would be adenosine triphosphate (ATP) that provides energy to cells.
- Metaphosphates are found in inorganic structures such as metals and salts.
- Orthophosphates are the simplest form of phosphate. Unlike organic phosphates and
metaphosphates which are not readily available, orthophosphates are reactive forms of
phosphate in water and can be directly measured.
- Phosphorus can seep into the environment from both natural sources and man-made
sources.
- Natural sources include rocks, minerals, and sediment while man-made sources include
fertilizers and detergents. Many US states have banned phosphate detergents leading to
most detergent companies reformulating their products.
- Phosphate-based fertilizers are especially effective at growing crops and are therefore used
on many fields.
Any rainfall over a field will drain into a body of water and lead to high concentrations of
phosphorus. This can be extremely problematic in agricultural regions such as the central
valley where a significant portion of land gets fertilized causing large increases in phosphates
in surrounding rivers and streams
Reagents required
1. Ammonium molybdate (0.005M). Dissolve 6.1793g of Ammonium heptamolybdate4H 2O in
1L of 0.4M nitric acid. Weigh 0.616 of Ammonium molybdate in 100ml of 0.4M
HNO3
2. Ascorbic acid (0.7% w/w). Dissolve 7.00g of Ascorbic acid in 1l of water and add 10ml
of glycerine. Weigh 0.7g Ascorbic acid in 100ml, top to the mark then add
1ml of glycerine.
3. i. Dissolve 0.014477g of potassium dihydrogen phosphate in 1l of distilled water =
1000ppm.
ii.Prepare 50ppm in 50ml volumetric flask from the 1000ppm using the dilution equation
(CcVc=CdVd) (stock solution).
A. COLOUR FORMATION (DIAZOTIZATION)

NB: FOR ALL THE STANDARDS PREPARED AND WATER SAMPLES
- i. - Add 3mls of molybdate solution to 3.0ml of ascorbic acid solution in a 50ml
volumetric flask and shake to mix.
- prepare Phosphate working standard solutions (10, 8, 6, 4, 2, 1mg/ml) from 50ppm
using the dilution equation into the 50ml volumetric flask in step i and top up to the
mark using distilled water. Mix well and allow to stand for 10 – 15 minutes.
ii. –pipette 10ml of each of the water sample provided and repeat step i above.
B: Determine the wavelength of maximum absorbance of Phosphate (λmax)
Using solution no 2;
- Fill one cuvette approximately ¾ full with deionized water (blank).
- Fill a second cuvette approximately ¾ full with solution no 2.
- Set the wavelength at 450nm and increase by a range of 4 until you reach 460nm ensuring
each time you change the wavelength you zero the spect with the blank (distilled water) and
record absorbance as a function of wavelength. (Consult a demonstrator).
– plot a graph of absorbance against wavelength. Make sure you get enough points to describe
any maxima accurately.
C. Establish a standard absorption curve from which calibration graph can be

plotted.
- Set the instrument to the wavelength of maximum absorbance (λmax) obtained from step B.
now measure the absorbance of each of your solutions in this wavelength (starting with the
lowest concentration) and also that of the water samples.
Discussion:
- Plot a calibration curve of absorbance vs phosphates concentration and use it to calculate
the phosphates concentration in the samples provided.
- Comment on the chemistry involved in the reaction.

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EXP 2: DETERMINATION OF PHOSPHATES

- Spectrophotometric Determination of Phosphates in Water Background Controlled

A. COLOUR FORMATION (DIAZOTIZATION)

C. Establish a standard absorption curve from which calibration graph can be

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