Experiment 5 UV VIS Post Lab

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Experiment 5 – Analysis of Dye

Formulation and Chromophore


Combination (UV-Vis)
Post-Lab Report

Group 9: - B11006011 蔡美先


- B11006012 Nicholas Wahjudi (Writer)
- B11006111 龐兆恩
- B11006112 Elizabeth Loid Urtecho

I. Experimental Results and Analysis


Week 1
1. 5 ml of 100 pm dye solution for a total volume of 50 ml
Dye final concentration : 10 ppm.
The wavelength (λ1) of the highest absorbance in the spectrum of dye
solution in volumetric flask 9: 615 nm
The wavelength (λ2) of the highest absorbance in the spectrum of dye
solution in volumetric flask 1: 435 nm

Table 1. Result of 1st week experiments


Flask 1 2 3 4 5 6 7 8 9
pH 1.65 4.49 6.18 6.50 7.14 7.60 7.90 9.01 11.33
[A] at λ1 0 0 0.079 0.187 0.385 0.557 0.650 0.825 0.808
[A] at λ2 0.323 0.378 0.337 0.331 0.260 0.248 0.194 0.320 0.214
[In-]/[HIn] -- 0 0.108 0.301 0.910 2.219 4.114 -48.5 --

Aa : 0 Ab : 0.808
2. From the spectrum of the measuring bottle (9), find the wavelength at which
the absorption is maximum as the detection wavelength (λ1), and use pH
versus absorption degree [A] Drawing. Such as schematic diagram (1).
Fig 1. pH vs [A] graph

pH vs [A]
0.9
0.8
0.7
0.6
[A] at λ1

0.5
0.4
0.3
0.2
0.1
0
1.65 4.49 6.18 6.5 7.14 7.6 7.9 9.01 11.33
pH

3. Plotting the pH value as horizontal axis and log([In-] / [HIn]) on the vertical
axis to find the linear relationship.

Table 2. Preparation to plot pH vs log([In-] / [HIn])


Flask 5 6 7 8
pH 7.14 7.60 7.90 9.01
[ln-] / [Hln] 0.910 2.219 4.114 -48.5
Log [ln-] / [Hln] -0.041 0.346 0.614 Error

Fig 2. Log 10 (ln-/Hln) vs pH value at 615 nm wavelength graph

Log 10 (ln-/Hln) vs pH value at 615


nm wavelength
1.5
Log 10 (ln- / Hln)

1 y = 0.3275x - 0.3487
R² = 0.9891
0.5

0
7.14 7.6 7.9 9.01
-0.5
pH

pKa = pH - Log [ln-] / [Hln]


y = 0.03275x – 0.3487 , y = 0 , x = 10.647
pKa of bromothymol blue is 7.10, then percentage of error is:
10.647−7.1
× 100% = 47.3 %
7.1

Week 2
1. For acidic environment, pH is constant, and the dye concentration changes.

Table 3. Result and Calculation for week 2 (acidic solution)


Flask 1 2 3 4
100 ppm 2 3 4 5
bromothymol blue
(mL)
Final 4 6 8 10
Concentration
(ppm)
[C] (mole/L) 6.4 x 10-6 9.6 x 10-6 1.28 x 10-5 1.6 x 10-5
[A] 0.0588 0.0786 0.0914 0.1271
[ln-] / [Hln] -1.7 -1.95 -1.89 -2.25
X 2.428 2.052 2.124 1.8
[ln-]* 1.554 x 10-5 1.970 x 10-5 2.718 x 10-5 2.88 x 10-5
[Hln]* -9.14 x 10-6 -1.010 x 10-5 -1.439 x 10-5 -1.28 x 10-5
Aa: 0.323 Ab: 0.214

Calculation:
[C]
Mw: 624.38 g/mol
𝐶 = 4 𝑚𝑔/𝐿 × 1 𝑔/1000 𝑚𝑔 × 1 𝑚𝑜𝑙 /624.38 𝑔 = 6.4 × 10-6
𝐶 = 6 𝑚𝑔/𝐿 × 1 𝑔/1000 𝑚𝑔 × 1 𝑚𝑜𝑙 /624.38 𝑔 = 9.6 × 10-6
𝐶 = 8 𝑚𝑔/𝐿 × 1 𝑔/1000 𝑚𝑔 × 1 𝑚𝑜𝑙 /624.38 𝑔 = 1.28 × 10-5
𝐶 = 10 𝑚𝑔/𝐿 × 1 𝑔/1000 𝑚𝑔 × 1 𝑚𝑜𝑙 /624.38 𝑔 = 1.6 × 10-5

To calculate [ln-] / [Hln]:


For A:0.0588
[ln-] / [Hln] = (A-Aa)/(Ab-A) = -1.7
For A: 0.0786
[ln-] / [Hln] = (A-Aa)/(Ab-A) = -1.95
For A: 0.0914
[ln-] / [Hln] = (A-Aa)/(Ab-A) = -1.89
For A: 0.1271
[ln-] / [Hln] = (A-Aa)/(Ab-A) = -2.25

To calculate [X]:
Hln ↔ H+ + ln-
1-x x x
(1-x) x x

[ln-] / [Hln] = x/(1-x)

For : x/(1-x)= -1.7 , x = 2.428


For : x/(1-x)= -1.95 , x = 2.052
For : x/(1-x)= -1.89 , x = 2.124
For : x/(1-x)= -2.25 , x = 1.8

To Calculate [ln-]* :
[ln-]* = x[C]
(1) [ln-]* = x[C] = 2.428 x 6.4 x 10-6 = 1.554 x 10-5
(2) [ln-]* = x[C] = 2.052 x 9.6 x 10-6 = 1.970 x 10-5
(3) [ln-]* = x[C] = 2.124 x 1.28 x 10-5 = 2.718 x 10-5
(4) [ln-]* = x[C] = 1.8 x 1.6 x 10-5 = 2.88 x 10-5

To Calculate [Hln]* :
[Hln]* : (1-x)[C]
(1) [Hln]* : (1-x)[C] = (1-2.428) x 6.4 x 10-6 = -9.14 x 10-6
(2) [Hln]* : (1-x)[C] = (1-2.052) x 9.6 x 10-6 = -1.010 x 10-5
(3) [Hln]* : (1-x)[C] = (1- 2.124) x 1.28 x 10-5 = -1.439 x 10-5
(4) [Hln]* : (1-x)[C] = (1-1.8) x 1.6 x 10-5 = -1.28 x 10-5
Fig 3. Results for absorption at maximum wavelength λ2

Fig 4. Molar Concentration vs Abs at Maximum wavelength λ2 graph

Concentration vs Absorption at Maximum


Wavelength λ2
0.14
0.12 y = 0.0274x - 0.0099
R² = 0.946
0.1
0.08
Abs

0.06
0.04
0.02
0
0 6.4 9.6 12.8 16
Concentration * 10^-6 (mol/L)

Slope: 0.0274 , 𝜀2=274 L/mol.cm


2. For basic environment, pH is constant, and the dye concentration changes.

Table 4. Result and calculation for basic solution


Flask 5 6 7 8
100 ppm 2 3 4 5
bromothymol blue
(mL)
Final 4 6 8 10
Concentration
(ppm)
[C] (mole/L) 6.4 x 10-6 9.6 x 10-6 1.28 x 10-5 1.6 x 10-5
[A] 0.0103 0.0194 0.0260 0.0394
[ln-] / [Hln] 0.0129 0.0246 0.0332 0.0513
X 0.0127 0.0241 0.0321 0.0488
[ln-]* 8.128 x 10-8 2.313 x 10-7 4.109 x 10-7 7.808 x 10-7
[Hln]* 6.319 x 10-6 9.368 x 10-6 1.239 x 10-5 1.522 x 10-5
Aa: 0 Ab: 0.808

Calculation:
[C]
Mw: 624.38 g/mol
𝐶 = 4 𝑚𝑔/𝐿 × 1 𝑔/1000 𝑚𝑔 × 1 𝑚𝑜𝑙 /624.38 𝑔 = 6.4 × 10-6
𝐶 = 6 𝑚𝑔/𝐿 × 1 𝑔/1000 𝑚𝑔 × 1 𝑚𝑜𝑙 /624.38 𝑔 = 9.6 × 10-6
𝐶 = 8 𝑚𝑔/𝐿 × 1 𝑔/1000 𝑚𝑔 × 1 𝑚𝑜𝑙 /624.38 𝑔 = 1.28 × 10-5
𝐶 = 10 𝑚𝑔/𝐿 × 1 𝑔/1000 𝑚𝑔 × 1 𝑚𝑜𝑙 /624.38 𝑔 = 1.6 × 10-5

To calculate [ln-] / [Hln]:


For A: 0.0103
[ln-] / [Hln] = (A-Aa)/(Ab-A) = 0.0129
For A: 0.0194
[ln-] / [Hln] = (A-Aa)/(Ab-A) = 0.0246
For A: 0.0260
[ln-] / [Hln] = (A-Aa)/(Ab-A) = 0.0332
For A: 0.0394
[ln-] / [Hln] = (A-Aa)/(Ab-A) = 0.0513

To calculate [X]:
Hln ↔ H+ + ln-
1-x x x
(1-x) x x

[ln-] / [Hln] = x/(1-x)

For : x/(1-x)= 0.0129 , x = 0.0127


For : x/(1-x)= 0.0246 , x = 0.0241
For : x/(1-x)= 0.0332 , x = 0.0321
For : x/(1-x)= 0.0513 , x = 0.0488

To Calculate [ln-]* :
[ln-]* = x[C]
(5) [ln-]* = x[C] = 0.0127 x 6.4 x 10-6 = 8.128 x 10-8
(6) [ln-]* = x[C] = 0.0241 x 9.6 x 10-6 = 2.313 x 10-7
(7) [ln-]* = x[C] = 0.0321 x 1.28 x 10-5 = 4.109 x 10-7
(8) [ln-]* = x[C] = 0.0488 x 1.6 x 10-5 = 7.808 x 10-7

To Calculate [Hln]* :
[Hln]* : (1-x)[C]
(5) [Hln]* : (1-x)[C] = (1-0.0127) x 6.4 x 10-6 = 6.319 x 10-6
(6) [Hln]* : (1-x)[C] = (1-0.0241) x 9.6 x 10-6 = 9.368 x 10-6
(7) [Hln]* : (1-x)[C] = (1- 0.0321) x 1.28 x 10-5 = 1.239 x 10-5
(8) [Hln]* : (1-x)[C] = (1-0.0488) x 1.6 x 10-5 = 1.522 x 10-5
Fig 5. Results for absorption at maximum wavelength λ1

Fig 6. Molar Concentration vs Abs at Maximum wavelength λ1 graph

Molar Concentration vs Absorption at


Maximum wavelength λ1
0.045
0.04 y = 0.0094x - 0.0092
0.035 R² = 0.99
0.03
0.025
Abs

0.02
0.015
0.01
0.005
0
0 6.4 9.6 12.8 16
Concentration * 10^-6 (mol/L)

Slope: 0.094 , 𝜀1: 0.094 L/mol.cm


II. Conclusion

In conclusion, the experiment focused on the analysis of dye formulation and


chromophore combination using UV-Vis spectroscopy has yielded valuable
insights into the absorption characteristics of the tested substances. By employing
the UV-Vis spectrometer, we were able to explore how different dye formulations
and chromophore combinations interact with light in the ultraviolet and visible
regions. The experiment demonstrated the significance of pH and concentration in
influencing absorption patterns within the UV-vis spectrum. Leveraging knowledge
from the Instrumental Analysis class, particularly the key intersection point at 500
nm, enabled a more precise execution of the experiment and facilitated the
computation of findings.

From my experience I think this experiment is very cool because I get to learn
about this new machine, the UV-Vis spectrometer and the pH meter. Furthermore,
the things that I learnt during my instrumental analysis classes help me a lot to do
this experiment. During the experiment, we also still dealing with some problems
and error when we use the instruments. I think those error is caused by our own
negligence during mixing the mixture. Also, I think this is one of the longest
experiments that I have done this semester.

The worst things that happen when we do this experiment is we have done some
error during the experiment, that makes our results in the reports very strange too.
I think it is because we didn’t change the range to 615 nm, instead we still use 500
nm.

III. Question and Discussion


1. If the maximum wavelength is not determined in the second week, and different
concentrations are measured, what will be the impact on the high concentration
absorbance?
• If the maximum absorption wavelength is not determined or taken to be
smaller than the original maximum absorption wavelength, it will cause an
error in Beer’s law, thus the line will not be straight for the relationship of pH
and the absorbance. The greater the absorbance deviation is, the greater the pH
value is.

2. In the experiment, when the dye concentration is constant and the pH value is
changed, there is an obvious intersection point near the wavelength of 500 nm
in the spectrum diagram. What is the physical significance of this point?
• A reagent called bromothymol blue is used to determine the weak acids and
bases in a pH range of 6.4 to 8.0. In an acidic environment, it turns yellow,
while in a basic environment, it turns blue. The figure below shows that at a
wavelength of 500 nm, there is a clear intersection point. This indicates that
the absorbance remains constant at all pH values, indicating that the molar
absorptivity of [In-] and [Hln] is equal.
IV. References
[1] College, S. M. (n.d.), LibreTexts Chemistry, Detection and Absorption of
Ultraviolet Light (Experiment):
https://chem.libretexts.org/Ancillary_Materials/Laboratory_Experiments/Wet_Lab
_Experiments/General_Chemistry_Labs/Online_Chemistry_Lab_Manual/Chem_9
_Experiments/04%3A_Detection_and_Absorption_of_Ultraviolet_Light_(Experi
ment

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