INSTRUMENTAL ANALYSIS I

EXPERIMENT 1: QUANTITATIVE ANALYSIS USING ULTRAVIOLET-VISIBLE (UV-VIS)

SPECTROPHOTOMETRY

MATEUS S.E
201705714
INTRODUCTION
Ultraviolet–visible spectroscopy or ultraviolet-visible spectrophotometry (UV-Vis or UV/Vis)
refers to absorption spectroscopy or reflectance spectroscopy in the ultraviolet-visible spectral
region (about 190-820nm).The Ultraviolet-visible absorption based on molecules containing π-
electrons or non-bonding electrons (n-electrons) which can absorb the energy in the form of
ultraviolet or visible light to excite these electrons to higher anti-bonding molecular orbitals. The
more easily excited the electrons (i.e. lower energy gap between the HOMO and the LUMO), the
longer the wavelength of light it can absorb. The basic principle of quantitative absorption
spectroscopy lies in comparing the extent of absorption of a sample solution with that of a set of
standards under radiation of a selected wavelength through the application of Beer-Lambert law.
The Beer-Lambert law states that the absorbance of a solution is directly proportional to the
concentration of the absorbing species in the solution and the path length. Thus, for a fixed path
length, UV/Vis spectroscopy can be used to determine the concentration of the absorber in a
solution. It is necessary to know how quickly the absorbance changes with concentration.
Equation:
A = ԑbC

Where A is the absorbance, C is the concentration in mol.dm-3

absorptivity. A plot of A versus C over a given concentration range is called a calibration curve. At
low solute concentrations, the calibration curve is linear and should pass through the origin. Calibration
curves are extensively used to establish the signal response of an instrument before a sample of
unknown concentration can be analyzed

AIM
The purpose of this experiment was to estimate the precision and accuracy of the determination of
phenol using a UV-Vis spectrophotometer and to determine the phenol concentration in an nknown
solution.

EXPERIMENTAL
APPARATUS:
- 0.0941 g of phenol
- Spectrophotometer
- Distilled water
- Quartz Cuvette
- Volumetric flasks
- Small beaker
- Pipette filler & pipettes

PROCEDURE:
A spectrophotometer was used in this experiment, which is an instrument containing a monochromatic,
a device which produces a light beam containing wavelengths in a narrow band around a selected
wavelength, and a means of measuring the ratio of that beams intensity as it enters and leaves the
cuvette. A 0.0100M phenol solution was prepared by weighing 0.0941g of the phenol in a small beaker
and a small amount of water was added, than this solution was transferred to a 100mL volumetric flask,
which was than diluted to the mark with distilled water. From the stock solution prepared, 25mL was
pipetted into a 250ml volumetric flask and diluted to the mark with water. Set up of the
spectrophotometer was done according to certain parameters and recorded the absorbance of the first
dilution. Re-diluted the first diluted solution by transferring 150mL into 250mL volumetric flask and
scanned it with UV (absorbance recorded). A series of standard phenol solutions were further prepared
and measured the absorbance.

RESULTS & DISCUSSION

Λmax = 270 nm
= 0.921

TABLE 1
V1=Vol. standard 10.00 15.00 20.00 25.00 30.00 35.00 40.00 45.00
solution (mL)
Vol. dH2O added 40.00 35.00 30.00 25.00 20.00 15.00 10.00 5.00
(mL)
Absorbance 0.212 0.287 0.550 0.466 0.576 0.631 0.732 0.807

C2=Concentration 0.0012 0.0018 0.0024 0.0030 0.0036 0.0042 0.0048 0.0054

(mol/L)

ABSORBANCE VS. CONCENTRATION (MOLE/LITRE)

0.9
0.8
0.7
Absorbance

0.6
0.5
0.4
0.3
y = 133.79x + 0.0911
0.2
0.1
0
0 0.001 0.002 0.003 0.004 0.005 0.006

Concentration (mol/L)

From the graph y = 133.79x + 0.0911

0.921 = 133.79x + 0.0911

0.921-0.0911 = 133.79x

x = 0.921-0.0911/133.79

x = 0.0062 M
DISCUSSION
As observed in Table 1, the concentrations of the standard solutions were computed and tabulated.
A graph of absorbance vs concentration was plotted and an equation of y = 133.79x + 0.0911 was
obtained, were y and x are the absorbance and the concentration of the unknown in mol/L. The
concentration was determined as 0.0062 M. There might have been some experimental errors
encountered such as sample preparation were if two samples are prepared so that one carries along
a greater concentration of insoluble particulates, then additional scattering will lead to an apparent
greater absorption. Furthermore the precision of measurement is poor because the transmitted
intensity is so low, and the calibration curve linearity is poor due to stray light. The effect of stray
light can be reduced by taking the readings at a wavelength where the absorbance is lower or by
using a non-linear calibration curve fitting technique.

CONCLUSION
From above we can conclude that UV/Vis spectroscopy is best method which routinely used in
analytical chemistry for the quantitative determination of different analytes, such as transition
metal ions, highly conjugated organic compounds, and biological macromolecules. Spectroscopic
analysis is commonly carried out in solutions but solids and gases may also be studied. The
presence of an analyte gives a response assumed to be proportional to the concentration. For
accurate results, the instrument's response to the analyte in the unknown should be compared with
the response to a standard; this is very similar to the use of calibration curves.