LABORATORY MANUAL

FOR INSTRUMENTAL CHEMISTRY

Curriculum 2019

Author:
Prof. Dr.rer.nat. Agus Taftazani

Layout:
Isnaini Kholilurrohmi, S.Pd.

Yogyakarta, 1 September 2019

DEPARTMENT OF CHEMISTRY
FACULTY OF MATHEMATICS AND NATURAL SCIENCES
UNIVERSITAS ISLAM INDONESIA
 Laboratory Manual for Instrumental Chemistry

ANALYSIS USING NUCLEAR MAGNETIC RESONANCE (NMR)

I. EXPERIMENTAL OBJECTIVE
1. The students are able to perform quantitative analysis using NMR.
2. The students are able to master the technique of molecule identification
using NMR.

II. THEORITICAL BACKGROUND

NMR spectroscopy is one of the techniques for identifying molecular structures.
The information obtained is in the form of a spectrum (a curve of absorbance
frequency vs intensity) when the analyte is placed in a magnetic field (Bσ) and
exposed to radiation at a radio frequency (FR). The spectrum of the sample depends
on the presence of atomic nuclei and the molecular structure of the analyte.
Although most commonly used for characterizing organic molecules containing 1H
and 13C nuclei, other atomic nuclei with nonzero spin can be identified for
determining chemical structure, relative configuration, etc., without damaging the
analyte.
The NMR instrument consists of the following components:
1) Magnet
2) Coil
3) FR transmitter
4) FR receiver
5) Data processing
6) Control unit
7) Shim system

Figure 1.1 NMR Component

The spin of atomic nuclei tends to align with the external magnetic field (along
the z-axis), with a slight thermodynamic excess of parallel spins resulting in total
magnetization in the sample. When radio frequency (RF) is applied, the atomic nuclei
become excited (rotating to the x and y planes). The nuclei undergo precession
(orientation change from the rotation axis) and relax back to their equilibrium state
through lattice relaxation (T1) and spin-spin relaxation (T2) processes, resulting in
oscillation that decreases sinusoidal waves called a free-induction decay (FID).

Curriculum 2019 Master of Chemistry Study

Program
Faculty of Mathematics and Natural
 Laboratory Manual for Instrumental Chemistry

Figure 1.2. basic principle of NMR illustration

There are three basic types of information that can be obtained from 1H
NMR spectra.
1. The type of protons present in the sample.
The 1H NMR spectroscopy is based on the interaction of protons with a
magnetic field. Protons interact with the magnetic field of the spectrometer and
also with the local field (electron density around them). Since the local
electronic environment depends on the functional groups, protons of the same
type will resonate at the same frequency (e.g., alkyl, alcohol, aldehyde). The
resonance frequency appears as a peak where the position of the peak is often
referred to as chemical shift (δ).
2. The 2nd basic type of information that can be collected from 1H NMR spectra is
the number of protons in each type of proton present in the sample. The area
under each peak (i.e., its integral) is proportional to the number of protons
contributing to the signal. There are 3 important points to consider: (a) the
integral is a relative number, which means that symmetry in the molecule needs
to be taken into account; (b) exchangeable protons (-OH, -NH2) have low
integrals; (c) carbon always has 4 bonds. The use of integrals can aid in the
correct identification of functional groups.
3. The 3rd basic type of information that can be obtained is the connectivity of the
protons. NMR signals can appear as a single peak (singlet) or be split into 2
(doublet), 3 (triplet), 4 (quartet), etc. peaks grouped closely together. This is
referred to as multiplicity, coupling pattern, or splitting. Multiplicity of a peak is
determined by the number of neighboring protons. Therefore, it can determine
the proton connectivity in the molecule. For the important case of proton (spin =
1/2), peak splitting follows the following equation:
Multiplicity = n+1
Where n = equivalent H in proximity to the proton signal.

NMR analysis can be used for both quantitative analysis and identification.

III. EQUIPMENT
a. Nuclear Magnetic Resonance (NMR)
b. 5 mL and 1 mL volumetric flasks
c. Dropping pipette

Curriculum 2019 Master of Chemistry Study

Program
Faculty of Mathematics and Natural
 Laboratory Manual for Instrumental Chemistry

IV. MATERIALS
a. Sodium Acetat
b. Eugenol
c. Vanilin

V. METHOD
Determining the concentration of sodium acetate by standard addition.
1. Prepare an unknown concentration solution of sodium acetate as the sample.
2. Prepare 5 mL of 100 mM stock solution of sodium acetate in D2O.
3. Pipette 0.4 mL of the sample solution into 6 1 mL volumetric flasks.
4. Calculate the volume of 100 mM sodium acetate stock solution that must be
added to each flask to create standard concentrations of 0, 10, 20, 30, 30, 40,
and 50 mM.
For example, in the 50 mM standard solution = 1 mL x (50 mM/100 mM) = 0.5 mL
5. Pipette the calculated amount of each stock solution into the respective 1 mL
volumetric flask.
6. Dilute with D2O.
7. Transfer 0.5 mL of each solution from the volumetric flask to a separate NMR
tube. Label each tube accordingly.
8. Determine the concentration of the sample from the NMR spectra results.

Identification of molecular structure.

Identification of molecular structure is done by comparing NMR spectra for eugenol
and vanillin compounds.

Curriculum 2019 Master of Chemistry Study

Program
Faculty of Mathematics and Natural
 Laboratory Manual for Instrumental Chemistry

X-RAY DIFFRACTION (XRD)

I. EXPERIMENTAL OBJECTIVES
Students are able to understand the working principles of XRD instrument and
are able to characterize various materials through a diffractogram interpretation.

II. THEORETICAL BACKGROUND

X-ray diffraction (XRD) is one of a techniques used to determine structure of a
crystalline solid which is also called as a method of powder X-ray diffraction (X-ray
powder diffraction), as shown in Figure 1.1. Samples in the form of crystalline solid
powder with a diameter size from 10 -7 - 10-4 m are placed on a glass plate. The X-rays
are obtained from electrons which come out from a hot filament in a vacuum
condition at high voltage, with a high speed collide with a metal surface, commonly
made of copper (Cu).

Figure 2.1 X-ray powder diffraction

The X-rays collide with crystalline solid samples, then difract lights in all
directions in accordance to Bragg's Law. A detector moves with a constant angular
velocity to detect X-ray beam which has been diffracted by the sample. Sample
powder or crystalline solids have a lattice fields which are arranged randomly with
various possible orientations, as well as the crystalline particles contained therein.
Each set of this lattice plane has several specific angular orientation angles, so that
the X-ray diffraction meets the Bragg's Law:
nλ = 2 dsin
θ with ; n : diffraction order (1,2,3, ...)
λ : length of the X-rays

Curriculum 2019 Master of Chemistry Study

Program
Faculty of Mathematics and Natural
 Laboratory Manual for Instrumental Chemistry

d : distance grating
θ : diffraction angle
Outcome form of difractometer could be in the form of analog or digital data..
Recording of the analog data is in the form of graph of lines which are recorded per
minute sync, with detector angle 2θ per minute, so that the x-axis is equivalent to the
angle 2θ. On the other hand, digital recording informs the intensity of X-rays towards
the amount of light intensity per second.
The generated diffractogram pattern is a sequence of diffraction peaks with a
relative intensity varies along a specific 2θ value. Magnitude of the relative intensity
of those peak series depend on the number of existing atoms or ions, and their
distribution within the material unit cell. Diffraction pattern of each distinctive
crystalline solid depends on the crystal lattice, unit parameters and X-ray
wavelengths used. Thetefore, the possibility that the same diffraction pattern is
generated from different crystalline solids is very small (Warren, 1969).

III. EQUIPMENT
Instrument of X-Ray Diffraction (XRD)

IV. MATERIALS
Various samples of solids

V. METHOD
a. Study interpretation of the following data:
Here is a reflection of wurtzite ZnO phase. Evidence of the existing wurtzite
phase is on the top of Cu-Kα (2θ) = 31.3670; 34.0270; 35.8596; 47.1635;
56.2572;
62.5384. These peaks related to reflection: 100, 002, 101.102, 110, and 103. To
ensure that these peaks are corresponding each other, we can clarify through
comparison intenitas (I / Io) of peaks: 3:31 (1), 3:13 (0.86) 2.93 (0.84). If we add
the data diffraction angles (θ), n and λ into Bragg’s equation, then we will obtain
a value of lattice spacing (d). Highest reflection (101) is the highest peak and
gives a percentage of 100% (1). The intensity of peak heights 002 and 100 were
86% (0.86) and 84% (0.84) compared to the highest peak, respectively.

Curriculum 2019 Master of Chemistry Study

Program
Faculty of Mathematics and Natural
 Laboratory Manual for Instrumental Chemistry

b. Discuss and interpret the following

diffractogram
1) The results of material Platinum (Pt) analysis

Peaks of reflection is at 2θ = 39.6; 47.4; 67.1; 81.2 and 83.6 °, correspond to

the reflection (111), (200), (220), (311) and (222) correspond to (JCPDS Card
04-0802). Comparison intensity: 2,265 (1); 1.962 (0.53); 1.1826 (0.33)
Prove the conformity of reflection to the intensity ratio.

Curriculum 2019 Master of Chemistry Study

Program
Faculty of Mathematics and Natural
 Laboratory Manual for Instrumental Chemistry

2) Result of the following minerals analysis:

Determine whether this mineral:

a. Pt
b. Ag
c. Pd
The following data is a reflection of each reference:
Pt: 2θ = 39.6; 47.4; 67.1; 81.2 and 83.6 °, according to the reflection (111),
(200), (220), (311) and (222) correspond to (JCPDS Card 04-0802).
Ag: 2θ = 38.14; 44.77; 64.57; 77.37; and 81.69o in accordance with the
reflection of (111), (200), (220), (311) and (222)
Pd: 2θ = 40.118; 46.658 and 68.119 according to the reflection of (111),
(200), (220).

VI. Worksheet
Determine whether results of this analysis are the result of XRD analysis of:
a. TiO2 Anatase
b. Rutile TiO2
c. Al2O3

Curriculum 2019 Master of Chemistry Study

Program
Faculty of Mathematics and Natural
 Laboratory Manual for Instrumental Chemistry

PARTICLE SIZE ANALYZER

I. EXPERIMENTAL OBJECTIVE
Students can understand the working principle of the instrument and the
particle size analyzer can determine the particle size of various samples

II. THEORITICAL BACKGROUND

The particle size an important ingredient for known or determined because it
will affect the properties of the material, for example a powder or droplet with a size
of 2- 5 μm will be more easily shaped aerosol thus more easily penetrated the target,
the smaller particles will easily dissolve and cause the viscosity of the suspension
more high. There are several methods of measuring particle include a method of
enrichment (shieving), microscopy, sedimentation, laser diffraction and dynamic light
scaterring (DLS).
The principle instrument particle size analyzer with DLS method is to measure
the Brownian motion of particles of matter and the medium and correlate with
particle size. Brownian motion is the movement of particles of matter and particles
medium that never stops and is random. Brownian motion is highly dependent on
particle size and particle terdisfusi cause the medium.
The working principle of this instrument is to illuminate laser light at a sample
and analyzing fluctuations in the intensity of light scattered at the scattering angle θ
by fast photon detector. At the time of the laser light comes, the light scattered by
the particles. When the phase of the light scattered equal, then the intensity (I) of
high light, whereas if a different phase / opposite the low intensity because of the
mutually exclusive consequently scattered light intensity fluctuates.

III. EQUIPMENT
Particle Size Analyzer (PSA) method DLS type Horiba

IV. MATERIALS
Various samples of material

V. METHOD
a. Sample preparation according to the instructions
b. Determine the size of the particles using a particle size analyzer Horiba brand
c. Record the work instruction tool
d. Print the results and interpret

VI. DATA ANALYSIS

Of the output device that has been printed, specify the size of the particles of the
sample.

Curriculum 2019 Master of Chemistry Study

Program
Faculty of Mathematics and Natural
 Laboratory Manual for Instrumental Chemistry

X-RAY FLOURESCENCE (XRF)

I. EXPERIMENTAL OBJECTIVE
1. After following this lab, students are able to perform characterization of
materials using XRF (X-Ray flourescence).
2. After following this lab, students are able to analyze specific elements or atoms
within a material.

II. THEORITICAL BACKGORUND

XRF analysis tecnique used is based on the identification and enumeration of the
characteristic x-rays of the events that occurred in a sample of the photoelectric
effect. The photoelectric effect occurs because the electrons in the target atom
(sample) exposed to high-energy rays (X-rays). If the energy is higher than the
binding energy of electrons in the orbital K, L, or M atom target, it will be the target
atom's electrons out of orbit so that the target atom will undergo electron vacancy.
This vacancy will be filled by an electron from the outer orbital followed by the
release of energy in the form of a specific energy x-rays (discrete). Discrete spectral
type that occurs depends on the electron transfer that occurs in the material atom.
This spectrum is known as the characteristic x-ray spectra. XRF spectrometry utilizing
a specific x-rays emitted by the sample to be analyzed subsequently arrested
detector element content in the sample.
Output data can be in the form of a graph of intensity versus X-ray energy of the
specific content of the element in the sample or plot between elements (content
elements) and intensity as shown in Figure 5.1 (J. Anal. Chem. 2012, 84, 10260-

10266)
Figure 5.1 Example of output XRF
III. EQUIPMENT
XRD (X-Ray Fluorescence)

IV. MATERIALS
Material solid or powder samples to be analyzed

V. METHOD
Interpret the compound obtained from the output data is a graph of intensity versus
energy x-ray of the specific content of the element in the sample or plot between
elements (content elements) and intensity.

Curriculum 2019 Master of Chemistry Study

Program
Faculty of Mathematics and Natural
 Laboratory Manual for Instrumental Chemistry

VI. WORKSHEET
a. Based on the following data mentioned element content and the intensity of the
elements contained in the sample (J. Anal. Chem. 2014, 86, 10892-10899)

Figure 5.2 Full spectrum (right) obtained by applying the multi-image acquisition on
an archeological pottery (left)

b. Based on the following data mentioned element content and the intensity of the
elements contained in the sample. (J. Chem. Educ., 2011, 88, 819-821)

Figure 5.3 Sample XRF spectrum of NaCl: x denotes the experimental points and
the solid lines are Gaussian fits. Note Argon KR peak is from atmospheric
background

c. Based on the following data mentioned element content and the intensity of the
elements contained in the sample. (Energy Fuels 2015, 29, 1669-1685)

Curriculum 2019 Master of Chemistry Study

Program
Faculty of Mathematics and Natural
 Laboratory Manual for Instrumental Chemistry

Figure 5.4 Typical X-ray fluorescence spectrum recorded from corn stover. Note
that both x and y axes are linear in the square root. (a) Full spectrum with the Rh
excitation and Compton lines. (b) Zoom on low energy part showing the equidistant
spacing of the fluorescence lines on the square-root energy axis (Moseley’s law).
The Rh peaks are caused by incident Rh Kα and Kβ X-rays being scattered in
the sample via Compton or Rayleigh (elastic) scattering

d. Based on the following data mentioned element content and the intensity of the
elements contained in the sample. (J. Environ. Sci. Technol. 2011, 45, 5491-
5497)

Curriculum 2019 Master of Chemistry Study

Program
Faculty of Mathematics and Natural
 Laboratory Manual for Instrumental Chemistry

Figure 5.5. μXRF spectra, 2D Debye rings (20 μm 20 μm spot size) and 1D X-ray
diffraction patterns of a region of the CHDS-045 dust sample with (a) high Pb XRF
count and (b) medium to low Pb XRF count. HC = hydrocerussite
[(PbCO3)2•Pb(OH)2)]; Pb = Pb metal; K = kaolinite; Go = goethite; C = cerussite
(PbCO3); Ca = calcite.

e. Based on the following data mentioned element content and the intensity of the
elements contained in the sample. (J. Agric. Food Chem. 2015, 63, 2406-2412)

Figure 5.6 Overlapped XRF spectra of (a) all standards and samples of table
salt (RS), marine salt (MS), and low-sodium salt (LS) and (b) 0−4 keV region.

Curriculum 2019 Master of Chemistry Study

Program
Faculty of Mathematics and Natural
 Laboratory Manual for Instrumental Chemistry

ELECTROLYSIS FOR DETERMINATION OF AVOGADRO NUMBER (NA)

I. EXPERIMENTAL OBJECTIVES
1. Students can arrange electrochemistry cell (elektrolysis)
2. Students can determine the change of theoretical electrode mass
stoichiometrically.
3. Students can determine the Avogadro number.

II. THEORETICAL BACKGROUND

One of an important thing in chemistry is Avogadro Number (NA) which is found
to be 6.02 x 1023 particle/mole. There are some methods available to determine NA
and the most appropriate method is X-ray crystallography. However in this
experiment, NA will be determined using electrolysis method. In the electrolysis cell,
electrical energy used originates from the external DC Power Supply in order to
trigger the inspontaneous redox reaction or it can be stated that the alteration of
electrical energy into chemical energy is occurred in the cell. Michael Faraday in the
year of 1834 mentioned the First Faraday Law “that the number of moles of materials
which is deposited or released from an electrode is equal to the number of electrical
charges passing the cell”. Coulomb (given in symbol C) is an international unit to state
the electrical charge. The value of coulomb is equal to Ampere-second.
C=Axs
In this practical activity, plate of Zn and Cu will be used as electrodes placed in a
beaker filling with sulphuric acid. In this electrolysis cell, Cu plate will function as the
anode and Zn plate as the cathode. By determining the average electric current
which is used in the reaction, in which all of the Cu ion formed are Cu(II), then it can
be determined the number of atom in one mole of Cu and its Avogadro number.

Figure 6.1 Electrolysis cell

III. EQUIPMENT AND MATERIALS

a. DC Power Supply 1.5 volt
b. Electrical wire
c. Alligator clips (4)

Curriculum 2019 Master of Chemistry Study

Program
Faculty of Mathematics and Natural
 Laboratory Manual for Instrumental Chemistry

d. Sulphuric acid 1 M
e. Cu plate 6. Zn plate
f. Distilled water
g. Beaker glass 250 mL (2)
h. Analytical balance
i. Abrasive in sheet

IV. METHOD
a. Use abrasive sheet to clean the Cu plate which will be applied as anode and Zn
plate as cathode.
b. Weight the Cu plate using analytical balance. Note as initial mass of the plate.
c. Fill about ¾ part of beaker glass with sulphuric acid 1 M (Caution: carefull in
handling the acid, use gloves)
d. Arrange the two electrodes to the DC Power Supply using electrical wire as it can
be seen in the figure 16.1. (do not soak the electrode into the acid before this
step)
e. Add the electrodes into the acid. Make sure that the two electrodes are soaked
completely.
f. Turn on the DC Power Supply and observe the appered current. Make sure that
the current is in the range of 0.2-0.6 A. If it has not been in the range, then set
the DC Power Supply. After properly fix, turn off the DC Power Supply. The
electrolysis cell is then ready to be run.
g. Turn on the Power Supply, observe the obtaining current, and perform the
electrolysis for 3 min.
h. After completed, take the two electrodes briefly from the acid.
i. Rinse the Cu plate with distilled water and dry.
j. Weight the plate, note as the final mass of Cu.
k. Repeat the previous steps in order to obtain the 2nd and the 3rd trials.

V. Data Analysis
Parameter 1st Trial 2nd Trial 3rd Trial
Initial mass of Cu plate (gram)
Final mass of Cu plate (gram)
Mass alteration of the
electrode (gram)
Electrical current (A)
Electrolysis time (s)
VI. Discussion
a. Calculate the total charges (in coulomb) passing the cell for each repeatition.
b. Use the previous answer to calculate the number of electrons involved in the
electrolysis for each trial (1,602 x l0-19 coulomb/e).

Curriculum 2019 Master of Chemistry Study

Program
Faculty of Mathematics and Natural
 Laboratory Manual for Instrumental Chemistry

c. Determine the number of Cu atoms which is lost in each trial. Remember that
the electrolysis process releases 2 electrons to produce 1 Cu2+ ion.
d. Calculate the number of Cu atom/g Cu which is lost on the anode for each trial
(atom/gram). (mass of the Cu atom lost is equal to the mass of the produced Cu
(II)).
b. calculate the number of Cu atom in 1 mole Cu for each trial using the molar mass
of Cu (atom/gram x gram/mole= atom/mole). (Ar = 63.54 gram/mol). Compare
the experimental results with the Avogadro number and calculate the % error!

Curriculum 2019 Master of Chemistry Study

Program
Faculty of Mathematics and Natural