ANALYSIS OF SAMPLES ABSORBANCES

USING ULTRAVIOLET-VISIBLE (UV-VIS) SPECTROSCOPY

BY:

I PUTU RAIWATA MERTANJAYA (0813031019)

CHEMISTRY EDUCATION DEPARTMENT

FACULTY OF MATHEMATICS AND SCIENCES

GANESHA UNIVERSITY OF EDUCATION

SINGARAJA

2011

ANALYSIS OF SAMPLES ABSORBANCES

USING ULTRAVIOLET-VISIBLE (UV-VIS) SPECTROSCOPY

I. Experiment Objective

1. Determine the copper content in the metal ore.

2. Determine the absorbance of Ethyl-p-methoxycinnamate (EPMS).

3. Determine the absorbance of natural indicator in acidic and basic solutions.

II. Theory

Many modern technologies depend on the metal. Therefore, it has become imperative

for a chemist to analyze metal ore to determine its content and develop a method for the

determination of metals that have commercial value. One of the important metal in

technological and economic value is copper. Copper is widely used for conductors, water

pipes, and a mixture of various other metals, known as the alliance (such as brass, bronze,

and silver coins). Spectrophotometry is widely used in laboratory analysis. Most laboratories

that require identification and determination of organic compounds and inorganic compounds

(eg pharmaceuticals, fertilizer, mining, etc.) using a spectrophotometer. In this trial will be

determined percentage of the mass of copper in copper ore using spectroscopic techniques.

Spectroscopy is the study of interaction of light (electromagnetic radiation) with

matter (atoms and molecules). If certain wavelengths of light by an adsorbed atom or

molecule, then the resulting absorption spectrum. Electronic structure of a species or a critical

molecule absorption of light by species or molecule. The color complex compounds depends

on the metal involved and the number of d orbitals has, associated with the oxidation state.

However, there are several compounds of transition group has no d orbitals but the

compounds are colored. The color is caused by electronic transitions involving the valence

electrons in the other. Physicochemical analysis method based on spectral data is known as

spectrometry. Several spectrophotometric methods is very important is ultraviolet (UV),

infrared (IR), nuclear magnetic resonance (NMR), and mass. Ultraviolet and visible

spectrophotometry can provide information about the chromophore group, which covalently

unsaturated group contained in the molecule. The main components of spectrophotometer

tool shown in the following scheme.

Here are the types of UV-Vis:

(c) Double beam (DB)-in-time

Absorption of light by molecules in the ultraviolet and visible spectral regions depend

on the electronic structure of molecules. Absorption of this energy is quantized, namely the

elevation of the orbital electrons in the ground state (ground state) to the orbital with higher

energy (excited state = excited state), causing changes in the electronic energy of molecules,

Beam Source Monochromator Sample Detector

Screen

namely the transition of valence electrons in the molecule. Various electronic energy levels or

electronic transition can be summarized as follows:

Figure 1. Electronic Energy Levels

Transition of n * requires less energy than a transition * or * .

Transition of n * (R band) of a single chromophore groups (such as carbonyl

or nitro), the characteristic bands ( maks <100, 250, -350 nm).

Transition of * (K band) for molecules with conjugated systems (such as

butadiene, mesitil oxide, and aromatic molecules with substitutions:-styrene,

benzaldehid, or asetofenon), the characteristic bands ( maks <104; 200-400 nm

wavelength).

Transition of * (B band). Benzenoid is characteristic of aromatic

molecules or heteroaromatik, the characteristic bands at ( maks 102 - <5000; 230-

270nm).

Transition of * (band E). Etilenic is typical for aromatic structure with

substitution auksochrom, tape characteristics on ( maks > 104; 180-200nm).

In analyzing the use of UV-Vis spektrofotrometer should note the following:

a. Formation of colored compounds

This step is conducted if the analyzed compounds not absorbing visible region. In

this case the compound must be converted into other compounds that can do the

absorption or reacted with reagent so as to absorb visible light.

b. Selection of wavelength

Wavelengths required in a quantitative analysis by spectrophotometry is the

wavelength corresponding to maximum absorbance (peak absorption). This is

caused by changes in absorbance for each unit of concentration is greatest at the

maximum wavelength, it will obtain the maximum sensitivity as well.

c. Preparation of calibration curve

For calibration curves, created a standard solution with various concentrations of

the unknown. Absorbance of standard solution is measured, then plots the

absorbance (A) against concentration (C), the curve formed is called the

calibration curve.

Ultraviolet and visible spectra is a picture of the position and intensity of absorption.

The position of absorption related to the radiation wavelength (λ), where the energy is equal

to the energy required for electronic transitions, while the absorption intensity (transmittance

or absorbance) depends on two factors: the ability of interaction between radiation energy

and electronic systems, as well as differences between ground state state and excited state.

The intensity of absorption quantitatively expressed as Lambert-Beer equation:

A = ε b c = - log T = - log0I

I= log

I

I 0

C b I

Ilog-

0

Where, A is the absorbance.

T is transmittance.

I is the intensity of light emitted by the solution in the cell.

I0 is the intensity of light emitted by the solvent in the cell at I the same.

Log is logarithm to base 10.

ε is the coefficient of extinction of the absorbing species or constant comparison (cm-1

M-1

).

b is the length through which the light solution (typically 1 cm).

C is the concentration of absorbing species in units of mol L-1

(M).

Lambert-Beer law states the relationship between the intensity of light absorbed by

the concentration and the thick solution through which the beam. If a beam of light with a

certain length is passed in a solution containing absorbent material, where some rays will be

absorbed and transmitted light sbagian. Simply put, Lambert-Beer law can be shown in the

following scheme.

Figure 2. Light absorption by the solution with a concentration of C

Intensity of light or radiation with P0 that passes through a medium-thick b containing

a solution with concentration C, will result in reduced intensity of P so that P < P0. The

relationship between transmission, heavy trailers and can be stated as follows:

Log AkbCP

P0

Where P0 and P is the initial light intensity / beam fell and the light transmitted. A is the

absorbance, b for kuvet thickness, C for concentration, and k is a constant which depends on

the concentration used. If C in g / L, constant called absorptivity (a) and if C in mol / L,

constant called the molar absorptivity (ε). Based on this, Lambert-Beer law can be written in

two forms, namely:

A = a, b C (g / L) or A = ε b C (mol / L)

Transmittance is the fraction of the transmitted power falls by an example. If A = log (P0 / P)

then A = log 1 / T, where T = (P0 / P).

Absorption spectrum graph is most often described as the transmittance (T) or absorbance

(A) against concentration (C). By changing the concentration, the absorbance will change at

each wavelength (λ). Graph can be described as follows:

Beam

source Sample

P0 P

b

Slope = εb

C (mg/L)

A

Graphic of relation between absobance and concenration

Information obtained from ultaviolet and visible spectra:

1. Maximum wavelength ( maks )

a. Quantitative analysis, all measurements are based on, but to compound that

has been known to frequently used approach to Woodward-Fieser.

b. Qualitative analasis: less informative.

2. Maximum molar Absorptivitas ( maks ); qualitative analysis, namely information

type tape / electronic transitions (plays a role in elelusidasi structure).

Copper Cu is one element of the transition period to the fourth element in the periodic

system. Copper metal has the electron configuration Ar (3d10

4s1) To achieve stability of

copper metal releases electrons to form compounds. One of the properties of copper that can

conduct electricity well but is less reactive. Cu in small amounts are essential for life, but

would be toxic in large amounts, especially for bacteria, algae, and fungi. Among the many

copper compounds used as pesticides are alkaline acetate, carbonate, chloride, hydroxide, and

sulfate. Commercially important compounds that are CuSO4. In addition in agriculture,

CuSO4 also used for battery and plating, the manufacture of other copper salts petroleum,

rubber and steel industries.

Ethyl-p-methoxycinnamate (EPMS) is one of the active ingredients found in

sunscreen lotions.

C (mg/L)

T

Graphic of relation between absobance and transmittance

O C 2H 5

H

H

O

CH 3O

EPMS Structure

Usually the ethyl-p-methoxycinnamate can be isolated from natural materials from

Koempheria galanga (Kencur). These compounds can be isolated using solvent extraction

using organic solvents.

Sunscreen lotions are usually used to minimize ultraviolet radiation. The range of

ultraviolet radiation is divided into two: UV A and UV B. UV-B which has a wavelength

range 280-320 nm is responsible for the skin damage. Therefore, sunscreen lotion contains

several active compounds that can absorb ultraviolet radiation rays.

Ethy-p-methoxycinnamate crystal has a melting point of about 48o-49

oC. These

crystals can be obtained by isolate it from the natural ingredients contained which in such as

kencur.

In isolation experiments EPMS can be used sokhlet extraction method. Separation

principle is based on the distribution ratio of solute in two solvents that do not dissolve each

other. Kencur-paste, put in sokhlet tool that has been wrapped with filter paper. Extraction

sokhlet discontinued when the suspected substance in the circulation that will be extracted are

exhausted, this is indicated by no color change of solvent after passing through the sample.

The organic solvent used in isolation EPMS using soxhlet extraction method is solvent ether.

Ethy-p-methoxycinnamate crystals obtained usually is mixed with impurities.

Purification can be done by recrystallization using ethanol and proceed with the test crystal

melting point EPMS.

Erythrina crista galli or red dadap is a kind of tree tribe members Fabaceae

(Leguminosae). Plants are often used as live fences and shade it has many other designations.

Medium-sized tree, reaching 15-20 m tall and 50-60 cm. Sections of bark are still young and

fine vertical stripes of green, gray, brown or whitish; stem usually with a small outboard

spikes (1-2 mm) black. Similar umbrella or rounded canopy gap, abort leaves in the dry

season.

The compound leaves bear three leaves, green to light green, leaf axis with 10-40 cm

long stalks. Child leaves inverted egg round, triangular, up to a rhombus shape with blunt tip;

child leaves the tip of the largest in size, 9-25 × 10-30 cm.

The flowers are arranged in cone-shaped bunches, in addition to or at the end of the

bare twigs, usually appear when the leaves fall, attracts many birds coming to pollinate. The

crown is red orange to dark red; flag from 5.5 to 8 × 8 cm, short nails, no white striped. Pod

thick and dark, narrowed between seeds, 15-20 cm × 1.5-2 cm, containing 50-10 eggs egg-

shaped seed, brown, red or purple shiny.

Erythrina crista galli often used as shade trees in coffee farms and cocoa, or the

propagation of trees for black pepper, betel nut, vanilla, or yam tubers. Also good to use as a

living fence poles. This plant produces a light wood (BJ 0.2 to 0.3), soft and white, good for

making floats, packing crates, frame, and children's toys. The wood pulp is also an ingredient,

but less well-used as a wood fire because a lot of smoke.

The leaves of young Erythrina crista galli can be used as a vegetable. The leaves are

nutritious augment maternal milk, making sleep more soundly, and together with interest

thereon for the launch period. The fluid extract of leaves mixed with honey drink to cure

worms; Erythrina crista galli leaf juice mixed with castor oil is used to cure dysentery. The

leaves are heated Erythrina crista galli used as a poultice to relieve rheumatism. Bark

Erythrina crista galli has efficacy as a laxative, laxative sputum urine and diluent.

Has a protein content (and nitrogen) is high, the leaves Erythrina crista galli also used

as feed for livestock or green manure. Erythrina crista galli tree of medium size, which

trimmed 3-4 times a year, can produce 15-50 kg of green feed in a year. So far, the leaves are

known Erythrina crista galli not toxic (toxic) for ruminants. Erythrina crista galli root

symbiosis with Brady rhizobium bacteria bind nitrogen from the air, and improve soil

fertility.

III. CHEMICAL AND CHEMICAL APPATARUS

3.1 Chemical Apparatus

3.2 Chemicals

No Chemical Apparatus Size Measure Amount

1 Volumetric pipette 5 mL 1

2 Beaker glass 100 mL, 250 mL 2

3 Volumetric flask 50 mL, 100 mL 1

4 Pipette - 2

5 Analitical balance - 1

6 Spatula - 1

7 Funnel - 1

8 Stirring rod - 1

9 Filer - 1

10 Spectrofotometry UV-Vis 1

No Chemicals Amount

1 CuSO4.5H2O 2.4948 grams

2 Aquades 500 mL

3 Ethanol 95% 500 mL

IV. PROCEDURES AND RESULTS

a. Determinination of Cu content in sample

No Procedures Results

Part 1

1. A total of 2.495 grams

CuSO4.5H2O weighed and then

dissolved into a little distilled

water. After it is inserted into a 100

mL volumetric flask and added

distilled water until a limit (0.1 M

CuSO4 solution).

CuSO4.5H2O powder blue

CuSO4.5H2O who weighed at 2.4948 g

0.1 M CuSO4 solution in the form of a

solution of blue

2. A total of 100 mL of 0.1 M CuSO4

solution was diluted to a

concentration of 0.02, 0.04, 0.06,

0.08 M added with distilled water.

Initial solution is diluted to a standard solution of

0.02, 0.04, 0.06, 0.08 M color is fading.

3. Standard solution which has diluted

the measured absorbance using

UV-visible spectrophotometer.

Konsentrasi larutan

standar Cu2+

(M)

Absorbansi

0,02 0,224

0,04 0,470

0,06 0,724

0,08 0,969

4. From these data the standard

calibration curve is made of copper

which states the relationship

between concentration and

absorbance.

y = 12.44x - 0.025R² = 1

0

0.5

1

1.5

0 0.05 0.1

Ab

sorb

anti

on

Concentration

Standard Calibration Curve of Copper

Series1

Linear (Series1)

4 Kelopak bunga dadap merah 100 grams

5 NaOH 10 mL

6 HCl 10 mL

7 Serbuk PP 1 gram

8 Kristal EPMS 1 gram

Part 2

1. Copper ore weighed about - about

1.01 grams and was placed in 100

mL beaker and add 10 mL of

concentrated nitric acid and 6 drops

of concentrated sulfuric acid.

Reaction expected until the copper

dissolves completely in

concentrated nitric acid.

The mass of copper that weighed 1.0150

grams.

Copper after the soluble form of the blue

solution.

2. The mixture is inserted into a 250

mL volumetric flask and then

added distilled water to mark the

limit. After that, 25 mL samples

were taken and diluted to 50 mL.

After the solution becomes diluted color is fading.

3. The samples were diluted in the

measured absorbance using UV-

visible spectrophotometer. Molar

concentration of samples was

determined through a calibration

curve in part I.

Konsentrasi larutan

standar Cu2+

(M)

Absorbansi

Sample 0,371

b. Determination of EPMS and nattural indicator absorbances

No Procedures Results

1 Sedikit kristal EPMS dilarutkan

dalam pelarut etanol. Kristal EPMS berupa kristal berwarna putih

Larutan EPMS berupa larutan bening tidak

berwarna

2 Rekam spectrum larutan EPMS

berikut dengan menggunakan etanol

95% sebagai sampel referensi dan

ukur absorbansinya.

Larutan

EPMS

Absorbansi

pada λ maks

1,834

c. Determination of indicators absorbance

No Procedures Results

Bagian 1

1 Serbuk PP dilarutkan dalam pelarut

air. Serbuk PP berupa serbuk berwarna putih

Larutan PP berupa larutan bening tidak

berwarna

2 Rekam spectrum masing – masing

larutan berikut dengan

menggunakan air sebagai sampel

referensi

Larutan E : Larutan PP

Larutan F : Larutan PP + NaOH

Larutan G : Larutan PP + HCl

Larutan

E

Larutan

F

Larutan

G

Absorbansi

pada λ maks

0,130 0,407 0,079

Bagian 2

1 Kelopak bunga dari bunga Dadap

Merah digerus kemudian hasil Kelopak bunga Dadap Merah berwarna

merah padam

gerusan disaring. Ekstrak bunga

Dadap Merah dilarutkan dengan

etanol 95%

Ekstrak bunga Dadap Merah berupa larutan

berwarna merah kecoklatan

2 Rekam spectrum masing – masing

larutan berikut dengan menggunakan

etanol 95% sebagai sampel referensi

dan ukur absorbansinya.

Larutan H : Ekstrak bunga Dadap

Merah

Larutan I : Ekstrak bunga Dadap

Merah + NaOH

Larutan J : Ekstrak bunga Dadap

Merah + HCl

Larutan

H

Larutan

I

Larutan

J

Absorbansi

pada λ maks

0,036 0,444 0,008

V. ANALYSIS AND DISCUSSIONS

a. Analysis of copper content in sample

In these experiments used UV-Vis spectrophotometer to determine the amount of

absorption wavelength of a colored complex compounds. UV-Vis spectrophotometer uses

two light sources are deuterium lamp (D) for ultraviolet light and tungsten light (W) for

visible light. The sample to be measured absorbance is inserted into the sample container

(kuvet). Sample container or kuvet is made of material which could continue to absorb light

and no light. Kuvet usually is made of glass or silica. Then the light source is passed through

an existing gap in the monochromatic, where the light is directed separately through reference

and sample with a rotating mirror that allows the first light in one direction, then in the other

direction. Both light and then turn to change direction with a continuous rotation mirror and

see the light detector and then the other one repeatedly. Electrical signals that change is

processed, converted to digital, and compared and programmable calculations performed.

Monochromatic wavelength is set to transfer a motor driven or for the measurement

wavelength is determined by the operator.

In this lab analysis of copper metal content of the sample solution CuSO4.

Copper Content Analysis of Solvent Samples CuSO4

First of all done recording the spectra distilled water in a reference cell and a solution

of A to D in the sample cell. Solvent A to D is a solution containing cupri ion Cu (H2O)42+

,

where the data is obtained as follows:

A B C D

Concentration (C) 0.02 0.04 0.06 0.08

Absorbance 0.224 0.470 0.724 0.969

Wavelength (nm) 800 800 800 800

From the table above can be made to the absorbance relationship against concentration

as follows:

Curve 1. Curve relation to the concentration of the sample solution absorbance

From the above curve can be obtained straight line equation: y = 12.44x - 0025, where

the value 12.44 represents the slope of the curve.

A = mC + a

A = 12.44 C - 0.025

0.371 = 12.44 C - 0.025

12.44 C = 0.371 + 0.025

C = 0.0318 M for 50 mL (after the sample was diluted from 25 mL to 50

mL)

Solution concentration before dilution is:

V1.M1 = V2.M2

50 mL x 0.0318 M= 25 mL x M2

M2 = 0.0637 M

So the initial molarity (in volume 250 mL) is 0.0637 M. So the number of moles in

the initial solution is:

Mol Cu = M x V

= 0.0637 M x 0.25 L

= 0,015925 mol

Mass Cu in sample = mole x molar mass

y = 12.44x - 0.025R² = 1

0

0.2

0.4

0.6

0.8

1

1.2

0 0.02 0.04 0.06 0.08 0.1

Ab

sorb

ance

Concentration

Standard Calibration Curve of Copper

Series1

Linear (Series1)

= 0.015925 mol x 63,5

= 1,0112 grams

% Cu in sample = 𝑚𝑎𝑠𝑠 𝑜𝑓 𝐶𝑢

𝑀𝑎𝑠𝑠 𝑜𝑓 𝑠𝑎𝑚𝑝𝑙𝑒× 100%

= 1,0112 𝑔𝑟𝑎𝑚𝑠

1,0150 𝑔𝑟𝑎𝑚𝑠 × 100%

= 99,63 %

b. Analysis of EPMS absorbance

EPMS is an active compound in a sunscreen lotion. Uptake EPMS can be measured

by using UV-Vis. The results show two peaks at a wavelength of 307 nm and 228.5 nm.

EPMS absorption at a wavelength of 228.5 nm is 0.770 and for a wavelength of 307 nm

absorbance is 1.834. EPMS can absorb at a wavelength of 307 nm. This suggests that

EPMS can absorb UV-B so that used as a sunscreen lotion. For the data can be seen in

this following image.

c. Analysis of natural indicator absorbance

PP indicator absorption in the atmosphere of acids and bases can be determined by

using UV-Vis. Uptake of PP without the addition of acid indicator or base can be seen in

the picture below.

From this picture can be seen there are three peaks at a wavelength of 229, 276, and

342.5 nm with absorption, respectively, 0.396, 0.163, and 0.130. Addition of acid in the

PP is not too much change summit, but the uptake of PP changed. This does not provide

color changes in the solution of PP. PP acid absorption in the atmosphere can be seen in

the following figure.

Uptake of PP at 228.5, 276, and 342 nm respectively are 0.329, 0.108, and 0.079.

Addition of base change in the indicator PP peak. Originally there were three peaks, but

with the addition of alkali to make 2 pieces of the peak. This shows the changes in the

structure of PP under alkaline conditions. These structural changes cause the PP red

under alkaline conditions. PP absorption in alkaline conditions can be seen in the

following figure.

PP uptake was detected in two peaks at a wavelength of 370 and 553 nm. Uptake of

each peak was 0.082 and 0.407.

In this experiment also tested the absorption of red dadap flower extract as a natural

indicator sample. Absorption of red dadap extract without the addition of acid or base

brownish red in color, shows 3 pieces of the peak at a wavelength of 230, 283, and 663

nm. Uptake of each peak is 2.003, 1.166, 0.036. Figure absorption red dadap extract

attached.

Addition of acid to extract red dadap provide 4 pieces of the peak at a wavelength of

225, 284, 532, and 665 with respective absorption peaks are 1.316, 0.635, 0.077, and

0.008. The addition of these top causes dadap extract red color changed from red to

brownish red. Figure absorption dadap extract of red in acidic form.

Addition of bases in extracts of red dadap provide 2 pieces of the peak at a

wavelength of 244 and 579 with respective absorption peaks are 2.458 and 0.444. This

peak reduction causes red dadap extract color changes from red brown to greenish

brown. Figure absorption red dadap extract under alkaline conditions attached.

VI. CONCLUSION

1. Standard curve absorbance relationship with concentration can be made from data

obtained using UV-Vis spectroscopy and copper metal content can be searched,

which acquired levels of copper in copper ore at 99.63%.

2. EPMS can absorb UV-B at wavelengths of 307 with the uptake of 1.834.

3. Dadap red flower extract may be used as a natural indicator because it can change

color in acidic and alkaline.

REFFERENCES

Ismono. 1978. Cara-cara Optik Dalam Analisis Kimia. Diktat. Bandung: Jurusan Kimia ITB

Fessenden, Ralph dan Joan Fesseanden. 1982. Kimia Organik Jilid 1. Jakarta : Erlangga.

Khopkar, S.M. 1990. Basic Concepts of Analytical Chemistry. Terj. Saptoraharjo, A. Konsep

Dasar Kimia Analitk. Jakarta: UI-Press.

Petrucci. 1985. Kimia Dasar Prinsip dan Terapan Modern. Jakarta : Erlangga. Diakses

tanggal 24 April 2008 dari http://www.Kebonkembang.com/neo.

Tim Pembina Ilmu Kimia. 2004. Modul Pembinaan Pembelajaran Kimia Bidang Ilmu Kimia

Fisika. Bali : Jurusan Kimia FMIPA Universitas Udayana.