(1)Analytical Chemistry / Instrumentation Fundamentals of Analytical Chemistry Classification of analytical methods and advantages of instrumental methods Paper No

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Analytical Chemistry / Instrumentation

Fundamentals of Analytical Chemistry

Classification of analytical methods and advantages of instrumental methods

Paper No. : 01Fundamentals of Analytical Chemistry

Module :02 Classification of analytical methods and advantages of instrumental methods

Principal Investigator: Dr.NutanKaushik, Senior Fellow

The Energy and Resouurces Institute (TERI), New Delhi Co-Principal Investigator: Dr. Mohammad Amir, Professor of Pharm. Chemistry,

JamiaHamdard University, New Delhi

Paper Coordinator: Prof. Rajeev Jain, Professor of Chemistry, Jiwaji University, Gwalior

Content Writer: Prof. Rajeev Jain, Professor of Chemistry, Jiwaji University, Gwalior

Content Reviwer: Dr. NimishaJadon, Assistant Professor, Jiwaji University, Gwalior

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Classification of analytical methods and advantages of instrumental methods Description of Module

Subject Name Analytical Chemistry / Instrumentation Paper Name Fundamentals of Analytical Chemistry

Module Name/Title Classification of analytical methods and advantages of instrumental methods

Module Id 02

Pre-requisites Objectives Keywords

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Classification of analytical methods and advantages of instrumental methods Paper: Fundamentals of Analytical Chemistry

Module 2

Classification of analytical methods and advantages of instrumental methods

Analytical chemistry can be divided into areas called qualitative analysis and quantitative analysis. Qualitative analysis deals with the identification of substances. It is concerned with what elements or compounds are present in a sample. Quantitative analysis is concerned with the determination of how much of a particular substance is present in a sample. Another classification of quantitative analysis may be based upon the size of the sample available for analysis. When a sample weighing more than 0.1 g is available, the analysis is spoken of as macro; semi-micro analyses are performed on samples of perhaps 10 to 100 mg; micro analyses deal with samples weighing from 1 to 10 mg; and ultramicro analyses involve samples on the order of a microgram.

Classical Chemical Analysis:

Quantitative chemical analysis started with the application of techniques of gravimetric procedures. In a gravimetric determination, a known volume of sample solution is treated with a suitable reagent which quantitatively precipitates the desired constituent present in the sample solution. The precipitate which is of known concentration is filtered, washed, dried and weighed. For example, an excess of dilute sulphuric acid is added to a given solution containing barium ions. The precipitate of barium sulphate formed is filtered, washed, dried and weighed. From the weight of barium sulphate the quantity of barium in the given solution is calculated.

Advantages offered by gravimetry are:

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 it is accurate and precise when using modern analytical balances

 possible sources of error are readily checked, since filtrates can be tested for completeness of precipitation

 precipitates may be examined for presence of impurities

 it is an absolute method

 it is inexpensive.

Another group of techniques was soon developed in which quantitative analysis was achieved by measuring volume of solutions, hence it was called volumetric. To the sample solution of unknown concentration, a solution of known concentration is gradually added till the reaction between them is just complete as shown by some indicator. The volume of the sample and reagent solutions, the concentration of the reagent solution are also known so the concentration of the given sample solution can be calculated. For example, a known volume of hydrochloric acid solution whose concentration is to be determined is taken in a conical flask and a few drops of phenolphthalein solution are added as indicator.

A solution of sodium hydroxide of known concentration is gradually added through a burette until the solution in the flask becomes just pink. The volume of sodium hydroxide solution added is recorded and from this, the concentration of given hydrochloric acid is calculated.

This process is called titration and the determination is termed titrimetric determination.

Advantages of titrimetry are:

• Robustness of the method

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• Analysis can also be automated

• Higher degree of precision

• Inexpensive

• Doesn’t require sophisticated instrumentation.

Over the last 50 years or so there has been a growing tendency to make use of certain instruments to achieve quantitative analysis. For example, instrumental techniques such as potentiometric, conductometric, photometric, amperometric etc., have been applied to locate the end point in a titration or to follow the course of a chemical reaction. It should be noted that in such cases the titrimetric methods are not basically altered from their standard procedures, the instrument simply act as a substitute for an indicator.

Recently such methods commonly known as the instrumental methods of analysis have been increasingly used especially in the field of industrial and commercial quantitative analysis due to their rapidity and sensitivity.

Traditionally, instrumental analyses are divided into five categories

The electroanalytical methods apply an electrical signal to the sample and/or monitor an electrical property of the sample. The separative methods rely upon separation of the components of a sample prior to measuring a property of the components.

1. Spectral Methods

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Classification of analytical methods and advantages of instrumental methods These types of method analysis use an instrument to calculate the amount of radiation that is

absorbed, emitted or scattered by the sample. If the amount of absorbed radiation is measured, the technique is absorptiometry or absorption spectrophotometry.

If the absorbed energy is electromagnetic radiation in the X-ray, ultraviolet or visible region of the spectrum, the subsequently emitted electromagnetic radiation is a form of luminescence termed either fluorescence or phosphorescence. The spectral methods of analysis are listed in Table 1.

Table 1: List of the major spectral methods of chemical analysis

Some of the spectral methods are explained in brief here:

A. UV-VIS spectroscopy

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Classification of analytical methods and advantages of instrumental methods B. IR spectroscopy

C. Atomic absorption spectroscopy D. Flame emission spectroscopy E. Atomic emission spectroscopy

A. UV-VIS spectroscopy

Ultraviolet–visible spectroscopy refers to absorption in the ultraviolet-visible region of electromagnetic radiation. The absorption or reflectance in the visible range directly affects the perceived colour of the chemicals involved. Ultraviolet and visible radiation absorption promotes electronic transitions σ→σ*, n→σ*, n→π* and π→π*.

Interpretation and uses of ultraviolet spectra

UV-spectrum of particular compound is recorded in conjunction with other spectral data such as infrared to deduce its molecular structure.

Ultraviolet spectra, however, do not furnish prime information as such. It acts as complimentary or even supplementary evidence to infrared.

B. IR spectroscopy

Infrared Spectroscopy is the analysis of infrared light interacting with a molecule. Molecular vibrations can occur by two different mechanisms. Quanta of infrared radiation can excite atoms to vibrate directly. Most organic molecules are fairly large and their resultant vibrational spectra are complex.

Infrared absorption spectra

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Classification of analytical methods and advantages of instrumental methods Two spectral regions are distinguished, region of group frequencies and fingerprint region. In

group frequency region, principal absorption bands may be assigned to vibration units consisting of only two atoms. Such influences reveal themselves on careful study and offer valuable evidence as to the nature of neighboring atoms. Fingerprint region extends from 1400 cm^-1 to 400 cm^-1. Absorption bands found here are related to vibrations of molecule as a whole. Resulting bands are unique to a particular molecule.

Advantages of IR-Spectroscopy:

 Non-Destructive

 Provides insights about functional groups present in compound

 Cheap and fast

C. Atomic absorption spectroscopy

Modern atomic absorption spectroscopy was introduced in 1955 as a result of independent work of A. Walsh and C. T. J. Alkemade. Developing a quantitative atomic absorption method requires several considerations, including:

 choosing a method of atomization,

 selecting wavelength and slit width,

 preparing sample for analysis,

 minimizing spectral and chemical interferences, and

 selecting a method of standardization.

Advantages of atomic absorption spectroscopy

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 High sample throughput

 Easy to use

 High precision

 Inexpensive technique

D. Flame emission spectroscopy

Flame photometry is based on measurement of intensity of light emitted when a metal is introduced into a flame. The wavelength of the colour tells us what the element is, and the colour's intensity tells us how much of the element is present.

In early experiments, visible colour of the flame was used to confirm the presence of certain elements in the sample, particularly alkali metals and alkaline-earth metals. Later the whole ultraviolet and visible range was utilized using a spectrophotometer. This instrument permitted us to select the wavelengths of the radiation and measure its intensity with considerable accuracy.

The spectrophotometric technique has proven to be one of the most reliable and easily used techniques for the determination of concentrations of sodium, potassium, calcium, and magnesium. Flame photometry is also named as flame emission spectroscopy because of the use of a flame.

Advantages of Flame emission spectroscopy:

 High sensitivity and high reliability for determination of elements in first two columns of periodic table.

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 In addition to the determination of metals, it can be applied to non-metal analysis by utilizing the infrared region of the spectrum

 Flame photometry is a simple, rapid method for the routine determination of elements that can be easily excited.

E. Atomic emission spectroscopy

In emission spectroscopy, a sample is excited by absorbing thermal or electric energy and the radiation emitted by the excited sample is studied for both qualitative and quantitative analysis. Most of the spectroscopic techniques are related to molecules but emission spectroscopy is related to atoms. Therefore, this technique is used as a method of elemental- metal analysis. With it, all metallic elements can be identified and quantitatively determined in very low concentrations, as can be metalloids, such as arsenic, silicon and selenium.

Solids, liquids or gases can be analyzed quite easily by this method.

Advantages of Atomic emission spectroscopy:

 Regarded as most reliable method for elemental quantitative analysis available at present.

 The method can be used for quantitative analysis of about seventy elements at concentration level as low as 1 ppm.

 High sensitivity.

2. Electroanalytical Methods

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Classification of analytical methods and advantages of instrumental methods The electroanalytical methods are divided into categories, as shown in Fig.1. During analysis

with most of the electroanalytical methods, electrical contact with the sample is completed by dipping at least two electrodes into the solution.

Amperometry is based on the control potential between the two electrodes and the current is measured.

Potentiometry is based on the controlled current between two electrodes and the potential is measured.

Fig. 1. Various electroanalytical methods

During electroanalytical studies using coulometry and electrogravimetry, either a potential or a current can be applied to the electrodes in the solution. The measurement of quantity Q of electricity that is consumed during an electrochemical reaction is coulometry. When weight of a reaction product after an exhaustive electrolysis is calculated, the method is electrogravimetry. In Conductometry, electrical conductance of solution is calculated.

Conductance is defined as the inverse of the electrical resistance. Normally an alternating electrical potential is applied to electrodes during the measurement. In voltammetry potential is applied to one of the electrodes while the current flowing through the electrode is measured. During the current measurement the potential is varied in some predetermined manner. Polarography is the series of voltammetric methods in which the electrode to which

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Classification of analytical methods and advantages of instrumental methods the potential is applied has a constantly renewable surface. The most common electrode used

for polarographic measurements is dropping mercury electrode.

3. Separative Methods

In the process of separation, the components can be individually assayed either qualitatively or quantitatively. Among the non-instrumental separative methods are distillation, extraction, precipitation, filtration, osmosis and reverse osmosis.

Chromatography is a technique in which a mixture is separated into its basic components.

The sample is placed on the edge of the stationary phase (a solid or liquid) and a mobile phase (a liquid or gas) is admitting to flow over the stationary phase. Strong components sticks to the stationary phase are swept less rapidly than weakly adhere. Result causes separation of components

Chromatography is easily categorized into following:

1. liquid chromatography 2. gas chromatography

Depending upon the state or nature of the mobile phase. Some of the chromatographic methods are explained in brief here:

 Paper chromatography

 Thin-layer chromatography

 Gas chromatography

 High performance liquid chromatography

A. Paper chromatography

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Classification of analytical methods and advantages of instrumental methods Paper chromatography is used to separate colored chemicals or substances. It is primarily

used as a teaching tool, having been replaced by other chromatography methods.

Apparatus

Apparatus required for paper chromatography consists of a support for paper, a solvent trough, and an air-tight chamber. Size of chamber may vary from an ordinary test-tube to large aquarium depending on size of paper. Sample is applied prior to dipping into eluting solvent as a small spot.

Fig. 2 Basic apparatus for paper chromatography

Methods of detection commonly used are:

 inherent visible colors of components,

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 reactions with color-producing reagents,

 ultraviolet absorbance,

 infrared absorbance,

 fluorescence,

 radioactivity,

 bioautography, or

 extraction and further chemical or physical tests.

Test reagents are applied by spraying, immersing, or by exposing to vapors. Bioautography involves placing paper in contact with culture medium followed by examination of growth of bacteria along paper strip.

Advantages of paper chromatography

 Requires very less quantitative material.

 Cheaper compared to other chromatographic methods.

 Both unknown inorganic as well as organic compounds can be identified by paper chromatography method.

 Doesn’t occupy much space compared to other analytical methods or equipments.

B. Thin – layer chromatography

Operations performed in TLC are essentially same as in paper chromatography. Instead of paper, thin layer adsorbent supported on a glass or plastic plate is used.

Nature of layer: Commonly used adsorbents are silica gel, alumina, diatomaceous earth, and powdered cellulose. These materials can be combined with binder to make a cohesive layer.

Preparation of layer:

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Classification of analytical methods and advantages of instrumental methods Layers of 0.15 to 2.0 mm thickness are satisfactory. Layers are produced by spreading film of

an aqueous slurry of adsorbent over entire surface. Slurry must be neither too thick (viscous) nor too thin, or it will not spread properly. In commercial spreading machines a slotted trough travels over glass and deposits a uniform layer. Binder requires about 30 minutes to "set".

The edges should be cleaned and plates stored in a cabinet. Pre-coated TLC plates (glass or plastic) are available from many laboratory supply firms.

Methods of Development and detection:

Procedure must be conducted in closed chamber. In order to detect spots, iodine vapor is used. Another detecting reagent is a spray of sulfuric acid. There are a host of reagents available, and, of course, spot can be scraped off, eluted, and investigated by any available method.

Advantages:

Compared to paper chromatography, thin-layer is more versatile, faster, and reproducible.

C. Gas chromatography

Basic GLC apparatus

A gas chromatograph requires a completely closed system. Components of GC are shown in Fig.3.

Carrier gas passes through pressure regulators which control flow rate through apparatus.

Sample is introduced into heated chamber either through silicone rubber septum or by sampling valve. Carrier gas carries sample components through column where they are separated. A thermostated oven is provided for column, injector, and detector. There are innumerable variations of basic components of gas chromatograph.

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Classification of analytical methods and advantages of instrumental methods Fig. 3. Essential features of gas chromatograph

Carrier gas

Common carrier gases are helium, nitrogen, hydrogen, and argon. These gases are relatively inexpensive, and are not hazardous to handle. Choice of carrier gas is usually based on availability in high purity grade. Thermal conductivity detectors work best with hydrogen or helium. Helium is popular choice for analysis. Flow through column is caused by difference in pressure between inlet and outlet. Pressure regulating valves maintain an inlet pressure, Pi outlet pressure, Po, is normally atmospheric pressure. On some instruments, flow controllers automatically vary Pi to maintain constant flow rate.

As gases are compressible, complications arise in determining flow rate and volume of gas passing through column.

Sample introduction

It is important to introduce sample in shortest time and in smallest volume possible. Sample chamber may be heated for rapid vaporization of liquid samples. In some instruments, sample is injected directly into column at inlet. Size of sample is dictated by several factors:

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 amount available,

 capacity of the column, and

 sensitivity of detector

Ordinary chromatograph can handle liquid samples in range of 0.1 to 10 µl and gaseous samples in range from 1 to 10 ml.

Columns

There are two types of columns, packed and open tubular. Packed columns are easier to fabricate, less expensive, last longer, have higher capacity. Open tubular columns have less pressure drop. Packed columns are usually 1 to 20 m long. Open tubular columns are usually 10 to 50 m long. Columns are bent in U- or W-shape or coiled to fit oven. Short columns are often made of glass. Longer columns are made of copper, aluminum, or stainless steel.

Detectors

Separations performed in column must be sensed and recorded. Detector must ignore large amount of carrier gas and find trace amounts of sample components contained therein.

Requirements for detector are:

 low limit of detection,

 linear response over an extreme range of concentrations,

 uniform response to all possible substances,

 simple calibration,

 short response time,

 small internal volume,

 low noise,

 long term stability;

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 it must be simple,

 inexpensive,

 robust, and

 safe to operate

Advantages of gas chromatography:

 High resolution power compared to other methods.

 High sensitivity when used with thermal detectors.

 Relatively good accuracy and precision.

 Separation and analysis of sample very quickly.

 Sample with less quantity is also separated.

D. High performance liquid chromatography

This apparently simple expedient, combined with a number of other improvements in apparatus and technique, has evolved into a new practice of liquid chromatography which is competitive with gas chromatography in speed and resolution of complex mixtures. HPLC column consists of packing materials.

Packing Materials:

Various types of materials such as pellicular beads (covered with thin layer of porous material), silica beads are used for packing the column. Another type of chemically bonded packing utilizes silicone polymers which are more stable because of their three-dimensional cross-linked structure.

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Classification of analytical methods and advantages of instrumental methods High Pressures: Columns packed with small particles require high inlet pressures in order to

give a reasonable flow rate. Pressures up to 10,000 psi are not difficult to handle in the small columns used (2 to 3 mm diam.). Improved, pulse-free pumping systems are incorporated in modern liquid chromatographs.

Detectors:

The smaller columns and faster flow rates place rigid requirements on the detection system.

Flow-through detectors with low dead volumes and high sensitivity are a necessity. Two types of detectors are currently popular. One is based on a flow-through micro-cell placed in an ultraviolet spectrophotometer.

A second popular detector, the differential refractometer, continuously monitors the difference in refractive index between the pure mobile phase (reference stream) and the column effluent.

Advantages of high performance liquid chromatography:

 Widespread applicability

 Greater reproducibility

 High resolution

 Columns can be re-used

Electrophoresis is the separative method that takes advantage of the relative mobility of ions toward an electrode of opposite charge (to the ion) and away from an electrode of similar charge.

The buffered solution through which the ions normally travel is either supported by porous paper or is in a gel.

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Classification of analytical methods and advantages of instrumental methods Mass spectrometry can be used either alone or in combination with some other analytical

technique, such as gas or liquid chromatography.

4. Radioanalytical methods

Radioanalytical chemistry focuses on the analysis of sample for their radionuclide content.

Various methods are employed to purify and identify the radio-element of interest through chemical methods and sample measurement techniques.

The importance of radio-analytical chemistry spans many fields including chemistry, physics, medicine,

pharmacology, biology, ecology, hydrology, geology, forensics, atmospheric sciences, health protection, archeology, and engineering.

Advantages of radio-analytical methods:

 Good accuracy

 Adaptability to wide variety of applications

They minimize or even eliminate the need for separations that are required in other analytical methods.

5. Thermoanalytical methods

Thermal analysis is a branch of materials science where the properties of materials are studied as they change with temperature. Several methods are commonly used:

 Differential thermal analysis (DTA)

 Differential scanning calorimetry (DSC)

 Thermogravimetric analysis (TGA)

 Advantages of Thermo-analytical methods:

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 High resolution

 Rapid response time

 Easy measurement on samples of different configurations

I. Do you know

1. Analytical chemistry can be divided into areas called qualitative analysis and quantitative analysis.

2. Thermal analysis is a branch of materials science where the properties of materials are studied as they change with temperature.

3. Radio-analytical methods are employed to purify and identify the radio- element of interest.

II. True or false

1. Qualitative analysis deals with the identification of substances. (True)

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Classification of analytical methods and advantages of instrumental methods 2. In a gravimetric determination, a known volume of sample solution is treated

with a suitable reagent which quantitatively precipitates the desired constituent present in the sample solution. (True)

3. Infrared Spectroscopy is not the analysis of infrared light interacting with a molecule. (False)

4. In group frequency region, principal absorption bands may be assigned to vibration units consisting of only two atoms. (True)

III. Interesting facts.

1. Separations using mass spectrometry are based upon the relative motion in an electrical or magnetic field of the components of a gaseous mixture of sample ions.

2. The smaller columns and faster flow rates place rigid requirements on the detection system.

3. Various types of materials such as pellicular beads (covered with thin layer of porous material), silica beads are used for packing the column.

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