YACHAY TECHNOLOGY UNIVERSITY

SCHOOL OF CHEMICAL SCIENCES AND

ENGINEERING

Applications of Ultraviolet -Visible

Molecular Absorption Spectrometry

PRACTICE 1:
Quantitative Determination of Hexavalent Chromium [Cr(vi)]
by a Diphenyl carbazide Colorimetry Method

INSTRUMENTAL ANALYSIS II

Professor Lola de Lima

STUDENTS:

 NATHALY JEMINA GONZÁLEZ BAÑO

 JOSÉ DAVID PULLAS CHIRIBOGA
 MARÍA EMILIA HERNÁNDEZ PÁEZ

07-October-2023
 QUANTITATIVE DETERMINATION OF HEXAVALENT
CHROMIUM [CR(VI)] B BY A DIPHENYLCARBAZIDE
COLORIMETRY METHOD

Nathaly González[1], Emilia Hernández[1], David Pullas [1].

1
School of Chemical Sciences and Engineering, Yachay Tech University.

ABSTRACT

Keywords:

RESUMEN

Palabras clave:

1. Introduction.  Acetone (analytical reagent grade)

 Potassium dichromate stock solution
of 1000 ppm Cr(VI)
2. Objectives  Potassium dichromate standard
a.General Objetives solution of 100 ppm Cr(VI)
 Sulfuric acid, 10% (v/v)
 250 mg 1,5-diphenylcarbazide
b.Particular Objetives b.Experimental Procedure

3. Methodology
a. Reagents and Material
 Distillated water
 Acetone (analytical reagent grade)
 Potassium dichromate stock solution
of 1000 ppm Cr(VI)
 Potassium dichromate standard
solution of 100 ppm Cr(VI)
 Sulfuric acid, 10% (v/v) c. Calibration
 250 mg 1,5-diphenylcarbazide
 Calibrations

Reagents and Material

 Distillated water
 data were interpolated towards the "x" axis
(figure 1).

Fig.1: Spectrum between 400 and 800

nm (A/nm).

It was determined that said length is

 Analysis in Samples equal to approximately 545.5 nm, this value is
d.Preliminar calculations very close to the theoretical value of
wavelength equal to 545 nm at which the
absorbance of the product obtained from the
reaction of hexavalent chromium with
diphenylcarbazide in a acid solution is
4. Results and Discussion
measured (Cr-DPC).

A measurement error associated with

the interpolation was determined equal to -0.5
a. Determination of the working
nm, which is considered not to affect the
wavelength.
veracity of the results obtained.
It is known that hexavalent chromium
b. Determination of the optimum
at acidic pH is orange in color, and its molar
complexing solution volume.
absorptivity coefficient is relatively low, so to
determine its presence in a sample, it is reacted
Experimentally, the trial-error method
with the diphenylcarbazide complex since this
was used to determine the optimal value of
allows, at the same concentration, to obtain a
complex. Three flasks were prepared to which
much bigger sign. In this way, when creating a
1 ml of standard solution and a volume of
calibration curve, it will have a high sensitivity
DPC equal to 1, 3 and 5 ml respectively were
and greater slope which, in other words,
added. After 10 minutes (for full color
allows for lower detection limits and thus
development), the absorbance was determined
minimizes errors related to interpolation
at 545.5 nm.
calculations.
Table 3.1. Complexing Solution for
A calibration curve was made
(absorbance/wavelength in nm) and the data Optimization
were fitted following a linear model, thus Volume complexing Absorbance
complying with Beers' law. Therefore, to solution (mL)
determine the working wavelength, the 1 0.3040
maximum point of the curve was taken and the
3 0.3161
 5 0.3678

The absorbance results presented in

Table 3.1 have a systematic error that is
believed to be due to the following reasons:

• At 1ml: this error could have been

due to not stirring the solution enough.

• At 5ml: this error could have been

since this value was determined after 12
minutes and not after 10 minutes of training,
as in the other cases.

Analyzing table 3.1. and considering

that DPC must be placed in excess to ensure
that all the hexavalent chromium is complexed
or converted to Cr-DPC (violet-red color), it
was determined that a volume equal to 3ml
should be used, which corresponds to a value
of absorbance equal to 0.3161.
To support these results, the maximum
point of the calibration curve in figure 1 was
taken and the data were interpolated towards
the "y" axis, thus obtaining an absorbance
equal to approximately 0.325, a value that, in
fact, is more like the absorbance obtained by
using a volume equal to 3 ml than in the other
two cases.
c. Determining complex formation time
and stability.
The complex (3 ml) was allowed to
stand for 10 minutes and then the absorbance
was measured at 545.5 nm.
Table 3.2. Complex Formation Time and
Stability
Time (min) Absorbance
2 0.7033
4 0.7103
6 0.7189
8 0.7230
10 0.7254
12 0.7249
14 0.7258
16 0.7264
18 0.7270
20 0.7147
Fig.2.: Kinetics plot, A versus t in seconds.
Analyzing figure 2, the kinetics of the
complex can be observed. At 840-1020
seconds or approximately 14-17 minutes the
absorbance value is relatively constant
(0.7258) which can be interpreted as that at 14
minutes the complex has finished forming,
until 17 minutes it remains stable and from
there it begins. to decompose up to 20 min.
Therefore, the optimal time to perform the
measurements is 14-17 minutes.
These results can be supported with
annex 8.2, which shows a table with the
kinetic data of the complex in seconds
generated by the spectrophotometer.
d. Determining concentration interval.
To determine the values of Cmin and
Cmax by using Beer's law, the range in which
the spectrophotometer measures the
absorbance of a solution at a given wavelength
was consulted, in this case it is 0.1 for Cmin
and 2, 0 Cmax.
So,
ε = A / (b * c)
ε = (0.7258 / (mol/L * M * cm)) / (1 cm * 100
ppm) = 0.0726 M
C𝑚𝑖𝑛 = A / (ε * b) = 0.1 / (0.0726 M * 1 cm)
= 1.3774 * 51.996 g/mol / 1,000 = 0.0716 ppm
 C𝑚𝑎𝑥 = A / (ε * b) = 2.0 / (0.0726 M * 1 cm)
= 27.5482 * 51.996 g/mol / 1,000 = 1.4324
ppm

Table 3.3. Interval of concentrations

Wavelengths (nm) Absorptivity Cmin Cmax
coefficient (mg L −1 ) (mg L −1 )
545.5 0.1 0.0716 -
545.5 2.0 - 1.4324

Therefore, the interval of

Fig.4: Graph of the residuals.
concentrations allowed for the working
wavelength is (0.0716-1.4324) ppm.

Analyzing the residuals (figure 4), the

e. Preparation of calibration curve. largest points at 0.2 ppm and 1 ppm are
discarded to improve the value of R² from
Table 3.4. Calibration curve. Concentration of the stock solution: ( ± 0.9828
)ppm to 0.9996 and thus have a better fit of
Standard Volume stock solution Final Volume Concentration
the data.
(± mL) (±mL) (mg L −1 )
1 0.05 25
2 0.1 25
3 0.15 25
4 0.20 25
5 0.25 25
6 0,3 25

The results obtained in table 3.4. were

obtained using the formula: Vc* [ x]c = Vd*[x
]d. The absorbances of the six standards were
measured 3 times to reduce the error Fig.5: Improved calibration curve (A/c in
associated with their measurement. ppm).

To create the calibration curve at In Figure 5 it can be seen that the

545.5 nm (figure 3), the Excel tool was used, curve has a strong positive linear trend and
and the linear regression method was also that the value of R² is equal to 0.9996, which
applied to analyze the values of A and c (ppm) represents that the variables are directly
obtained (annex 8.3.). proportional and have a high veracity value.

In addition, it is analyzed that the

curve has a high sensitivity and greater slope
than that of figure 3 which, in other words,
allows us to have lower detection limits and
thus minimize the error related to interpolating
the data to determine the chromium
concentration in the sample.

3.4.4 Analysis in Samples:

Fig.3: Calibration curve (A/c in ppm). • Std. Residual Error (Syx)= √((Σ(y -
ŷ)²) / (n - 2))
 Σ (y - ŷ)² = 0.1152 (as mentioned in
Annex 8.4).

n = # standards * # measurements of
each standard = 4 * 3 = 12. Since y¯ and yo are practically the same so all
that term is 0, and thus reduce the error of the
Syx = √(0.1152 / (12 - 2)) = 0.1073. calculation.

• F-statistic = ((R²) / (1 - R²)) * ((n - 2) So:

/ 1)
So/co=0.1073/0.62309*√(1/12+1/3=0.1112
F-statistic = ((0.9996 ²) / (1 - 0.9996
²)) * ((12 - 2) / 1) = (0.9992 / 0.0079) * 2 = The results obtained reveal a highly
1264.81 positive situation in the regression model. The
Syx value, which represents the average
dispersion of the data around the regression
line, is notably low, indicating an accurate fit
Table 3.5. Interval of concentrations of the model. This finding is reinforced by a
Coefficients Estimate Std. Error t value Pr(>t)high coefficient of determination (R²) of
Intercept 0.034057143 0.00316634 10.755992 0.9996, which confirms the model's ability to
8.11882E-07
make highly accurate predictions.
1 Furthermore, the high value of F equal to
Slope 0.623090476 0.00392736 158.6536438 2.43114E-18
1264.8101 supports the global relevance of the
3 regression and suggests that the model is
Model Residual 0.1073 statistically significant in explaining the
standard error variability in the data, in addition the
interpolation error equal to 0.11 is also low.
R-squared 0.99960287 0.00402435 - -
Together, these results validate the
effectiveness of the present regression model
F-statistic 1264.81 - - in predicting the Cr concentration in ppm.

Since the calibration curve (figure 5) 3.4.4.2 Absorbance measurement of samples:

on the y-axis ranges from 0.28-0.76
absorbance, it is intended that when diluting Table 3.6. Sample dissolution
the sample they fall at the centroid equal to Sample volume Final volume Dilution
0.52 absorbance, because, at this point a lower ( ± mL) (± mL) Factor
related error is visually recognized to the 25 100 4:1
interpolation of the data towards the x-axis,
resulting in a concentration equal to 0.76 ppm
(figure 6).
Table 3.7. Sample Analysis Results
Dilution Dilution Absorbance
Factor 1
1 4:1 0.4911
2 4:1 0.4911
3 4:1 0.4970

The average of A1, A2 and A3 (table 3.7.) at

Fig.6.:Calibration curve (A/c in ppm). 545.5nm is obtained and substituted into the
equation of the line:
 Interpolation error calculation
equation:
 · A= (0.4911+0.4911+0.4970)/3
=0.4931
· y = mx+b
b = 0.4931 - (0.623090476 * 0.76 ppm) =
0.4931 - 0.4735487618 ppm = 0.01955123824
ppm
· FD = Vf/Va =25 ml /4 ml =
6.25
 Concentration = b * FD = 0.0195 ppm
* 6.25 = 0.1219 ppm Cr (VI)
0.1219 mg/L Cr * 0.1 L = 0.0122 mg Cr (VI)
0.0122 mg Cr/100g muestra = 1.219*10^-4
ppm Cr (VI) en la muestra.
5. Conclusions
• In this lab, the diphenylcarbazide
method in spectrophotometry was successfully
applied to quantitatively determine the
hexavalent chromium content in a sample.
This method is based on the formation of a
colored complex between hexavalent
chromium and diphenylcarbazide in an acid
solution.
• A strong linear relationship was
established between absorbance and
hexavalent chromium concentration in a series
of standard solutions. This relationship
allowed the construction of an accurate
calibration curve that was successfully used to
quantify the chromium concentration in an
unknown sample.
• The method was shown to be highly
selective for the determination of hexavalent
chromium and is not significantly affected by
the presence of other substances in the
samples, as long as the concentrations of
interferences, such as molybdenum and
mercury, are maintained for below certain
limits.
• The optimal working wavelength
was determined as 545.5 nm, where maximum
absorbance was observed. This choice of
wavelength ensures highly accurate and
reproducible measurements.
6. Recommendations
• Since UV spectrophotometry
equipment analyzes monochromatic light, it is
essential to focus on identifying the peak point
for accurate concentration determination by
interpolation. This guarantees reliable
measurements.
• For future applications of this
method, it is recommended to implement
rigorous quality control. This should include
preparation of standard solutions and
verification of the precision and accuracy of
the spectrophotometry equipment used. These
procedures are essential to obtain reliable and
accurate results.
• If the samples of interest contain
substances that could interfere with the
diphenylcarbazide reaction, it is advisable to
carry out a further evaluation of the selectivity
of the method. If necessary, sample
pretreatment methods should be considered to
eliminate or reduce interferences.
• It is essential to keep a detailed
record of all experimental data and
calculations performed during practice. This
ensures the preparation of a complete and
accurate scientific report that supports the
results obtained.
• It is suggested to explore the
application of this method in real water
samples, especially those related to water
quality in industrial or environmental settings.
This will allow the concentration of hexavalent
chromium to be determined and compliance
with current water quality standards to be
assessed.
7. References
[1] D.A. Skoog, F.J. Holler, and S.R. Crouch.
Principles of Instrumental Analysis.
Cengage Learning, 2017. ISBN:
9781337468039.
 URL:https://books.google.com.ec/books?

id=D13EDQAAQBAJ.

[2] SW-846 Test Method 7196A: Chromium,

Hexavalent (Colorimetric).

URL:https://www.epa.gov/hw-sw846/sw-

846-test-method-7196a-chromium-

hexavalent-

colorimetric#:~:text=SW846%20Test%20

8. Anexes

8.1 Safety data Sheet

Annex 8.2: Kinetics of the complex in

seconds.
Annex 8.3.: Data used in figure 3.

c (ppm) A
0,2 0,2014
0,2 0,2036
0,2 0,2072
0,4 0,2794
0,4 0,2786
0,4 0,2791
0,6 0,4108
0,6 0,4108
0,6 0,4109
0,8 0,5376
0,8 0,5372
0,8 0,5352
1 0,727
1 0,7258
1 0,7258
1,2 0,7775
1,2 0,7794
1,2 0,78

Annex 8.4.: Calculation of Σ (y - ŷ)²) and Σ

(X-X̄ )².