Chemistry IA Proposal

RQ: How does the extent of oxidation of the dye affect the rate of degradation of trypan
blue, an azo dye, when exposed at varied lengths of time to sodium persulfate under
the presence of UV light (from a UV lamp) using UV-Vis spectroscopy method?

Motivation:
Trypan blue is used to stain tissues in cataract surgeries to facilitate the removal of eye
tissue. As an aspiring ophthalmologist, I’ve wondered if there is a quick way to break
down the dye in stained tissues after the surgery is completed.

Background info:
Azo dyes
Azo dyes are synthetic organic dyes with the functional group R−N=N−R′, in which R and R′ are
commonly Aryl and substituted Aryl groups. They are part of a family of Azo compounds that
contain a C-N=N-C linkage.[1] Most azo dyes contain only one azo group, but some can contain
two or three azo groups, namely diazo dyes and triazo dyes respectively. Azo dyes make up up
to 70% of all dyes used in many food and textile industries.[2]

Trypan Blue
Trypan Blue (C34H24N6Na4O14S4) is a water soluble diazo dye. Trypan blue has many
uses, such as treating African trypanosomiasis and also as a visual aid by staining
microscopic cell tissue in testing for cell viability.[3] It is also an important dye widely
used in ophthalmological surgery methods to stain the cornea and conjunctiva.
However, its safety for intraocular application has been debated for decades. There
have been reports that indicate observations of damage to retinal tissue after prolonged
use and high doses of Trypan blue. Thus, in this investigation I would like to find out
how the extent of trypan blue being oxidised by Sodium persulfate would affect its rate
of photocatalytic degradation by UV light.

Sodium persulfate
Sodium persulfate is the inorganic sodium salt with the formula Na2S2O8 of
peroxydisulfuric acid, H2S2O8, an oxidising agent. Sodium persulfate is used industrially
as a bleach or as detergent as it is a strong oxidising agent in chemistry, such as in the
Elbs persulfate oxidation reactions[4] It is mainly used as a radical initiator for
polymerisation reactions for some polymers containing styrene.[5]
Use of UV lamp
Na2S2O8 is able to react with Trypan blue without UV light but the process would be too
slow to yield concrete results opposed to with the UV lamp. Under UV light conditions,
the persulfate ion degrades quickly to produce oxygen. S2O8-2 + H2O ——> 2HSO4- +
1/2O2. The O2 reacts with the azo dye thus oxidising it. However O2 is a poor oxidising
agent. In the presence of UV light, sodium persulfate undergoes photolysis where the
weak O-O bond is broken down. Photolysis is a chemical process in which molecules
are broken down into smaller units in the presence of light.[6] Na2S2O8 is broken down
into hydroxyl radicals, H-O°, that are highly reactive and readily degrade Trypan blue.
The UV lamp serves as a catalyst in the photolysis of sodium persulfate and provides an
alternative pathway by allowing hydroxyl radicals to react with the dye instead of O2
molecules.

Hypothesis:
The longer the exposure of trypan blue to a fixed concentration of sodium persulfate;
the more oxidised the dye, the harder it is to degrade trypan blue.

Chemical and apparatus list:

Apparatus Name Uncertainty

Electronic Weighing balance ±0.001g

250cm3 Volumetric flask ±0.15cm3

500cm3 Volumetric flask ±0.25cm3

100cm3 Measuring cylinder ±0.5cm3

25cm3 Pipette ±0.06cm3

100cm3 Beaker ±5cm3

500cm3 Beaker ±25cm3

UV-Vis spectrophotometer ±0.005A

Cuvettes -

Stopwatch ±0.01s

UV Lamp -
 Chemical name Chemical formula IUPAC Name

Trypan blue (s) C34H24N6Na4O14S4 (3Z,3'Z)-3,3'-[(3,3'-dimethyl

biphenyl-4,4'-diyl)di(1Z)hyd
razin-2-yl-1-ylidene]bis(5-a
mino-4-oxo-3,4-dihydronap
hthalene-2,7-disulfonic

Sodium persulfate (s) Na2S2O8 Sodium peroxodisulfate

Sodium thiosulfate Na2S2O3 Sodium thiosulfate

Table of variables and control:

Independent variable The duration of exposure to a fixed
250cm3 of Sodium persulfate is varied as
such:
1 minute, 10 minutes, 20 minutes, 30
minutes, 40 minutes.

Dependent variable The rate of degradation of Trypan blue,

as seen in the absorbance against time of
the uv vis spectrophotometer.

Constant variable(s) -Concentration and volume of Sodium

persulfate added to each trial (250cm3)
-Concentration and volume of trypan blue
in each trial (50cm3)
-Light intensity of UV lamp

Procedure:
Step 1: Determine the concentration and the volume of sodium persulfate standard
solutions needed for five trials for each length of time. Use the table of calculations
below to find the mass of solid sodium persulfate to dissolve in distilled water.

Determining Sodium persulfate standard solution

C1V1=C2V2 C1 x 10 = 0.01 x 250

 C1 = 0.25M

Step 2: Weigh the mass of the powder in a beaker on the electronic weighing balance.
The mass of the empty beaker should not be included in obtaining the mass of solid
sodium persulfate.
Step 3: After weighing, empty all the solids into the 250cm3 volumetric flask then dilute
it with distilled water.
Step 4: Weigh the mass of trypan blue on the electronic weighing balance. Using the
equations No. of mol= concentration x volume and mol= mass/Mr, determine the mass
of trypan blue to form a 0.01M solution in 50cm3.

Data collection:
Step 1: Use a pipette to measure 100cm3 of Sodium persulfate solution into a 250cm3
beaker.
Step 2: Pipette 20cm3 of trypan blue solution from the 100cm3 beaker and empty it into
the 250cm3 beaker with the sodium persulfate.
Step 3: Start the stopwatch instantaneously.
Step 4: Add the quenching reagent, sodium thiosulfate, to stop the reaction at the 1
minute mark. Draw out a sample of the mixture into a cuvette.
Step 5: Use the UV-Vis spectrophotometer to record the absorbance data. Plot data in a
relevant table.
Step 6: Repeat steps 1-5 for the rest of the lengths of time (10,20,30,40 minutes) with 5
samples each.