Option G - Modern analytical chemistry

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[edit] G.1 Analytical techniques

[edit] G.1.1

Analytical techniques became more frequently used because of the questions and conflicts regarding the structure of molecules, mechanisms of reaction, the increased research on the molecular level caused an increased need for the understanding of the composition of substances, purity became a factor of greater importance. Therefore the reasons for using analytical techniques are:

• Determination of structure.
• Determination of mechanisms of reaction (by observing the species present).
• Determination of the composition of substances.
• Determination of purity of a substance.
• Separation of substances.
• Identification of substances.

[edit] G.1.2

Information that can be obtained from analytical techniques is the following:

Visible and ultraviolet spectroscopy:

• The determination of the absorption spectrum and the determination of λ_max. e.g. detection of optical whiteners, since they absorb in the UV part of the spectra and emit in the visible part.
• Quantitative analysis, e.g. in drug analysis first determine λ_max for the compound; plot a calibration curve of the absorbance measured against concentration and then the unknown solution can be determined.
• To determine, identify a d-block element, for example a complex of the element is prepared and then analysed, identified.
• Determine the presence of compounds that absorb in visible – UV part of the spectra.
• Used to provide data about the extent of reaction.

Infrared spectroscopy: The IR spectra is very complicated, and is therefore rarely used as the sole analysing technique, it is however used as a supporting technique in the obtaining of the following information:

• Identification of (organic) functional groups, since each of them absorbs in a certain range,
• Determination of the structure of organic compounds – secondary structure of proteins,
• Determination of unsaturation of fats and oils,
• Information on bond strength and length,

Mass spectroscopy: So far the application of mass spectroscopy alone has been limited to:

• Determining relative atomic/isotopic mass,
• Identification of organic compounds.
• Detection and quantification of relative abundance of isotopes in nature. For example the presence of ¹⁸O or ¹³C, dating techniques such as ⁴⁰K/⁴⁰Ar are based upon MS.
• In combination with Gas chromatography (GC) it is used in drug and food testing. e.g. the separated sample is “fed” into the mass spectrograph, where it is then identified.

Chromatography: It is used in the separation of substances, it is often the initial step of many analytical techniques.

Nuclear magnetic resonance (NMR): As many other analytical techniques it is used for structure determination, but along this main function, many uses are becoming increasingly important. It is used in medicine for body scanning as well as monitoring of metabolic processes.

X-ray crystallography: It is used in the determination of structure of crystals, both arrangement and position of atoms can be determined.

[edit] G.2 Principles of spectroscopy

[edit] G.2.1

The electromagnetic spectrum is a spectrum of all electromagnetic radiations. Electromagnetic radiation is the emission and transmission of energy through space in the form of waves, that have an electric field component and a magnetic field component. The sequence of different EM radiations in the EM spectrum is as follows: Gamma rays – X rays – UV light – Visible Light – Infrared – Microwave – Radio waves The energy decreases from Gamma rays to Radio waves as does frequency (v). On the other hand Wavelength (λ) increases from Gamma rays to Radio waves. Visible light is between 400 nm and 700 nm.

[edit] G.2.2

Emission spectra: If substances are provided enough extra energy they, emit light of certain wavelengths. The spectrum that can be detected by such emission is called emission or line spectrum. The entire spectrum is derived from the specimens own radiation.

Absorption spectra: Each substance absorbs certain amounts of light at certain wavelengths. Absorption spectra are obtained by passing light through a specimen and recording the colours that are not absorbed by the specimen, they appear as lines of colour in the spectrum. No “own” radiation is formed

[edit] G.2.3

Electromagnetic radiation in the range from UV to radio waves is absorbed by molecules.

UV to visible light is absorbed by atoms that undergo electronic transition, this means that enough energy is provided for the electrons to change energy level. This type of absorption is typical for complexes of d-block elements.

IR radiation is absorbed by molecules that undergo a change in the dipole moment by vibrational transition. The main forms of vibration are: Symmetrical stretch vibration, asymmetrical stretch vibration, bending vibration.

Radio waves are the ones with the least energy and are absorbed during the rotational motion of particles, the direction is important.

[edit] G.2.4

A double beam Infrared Spectrometer Image:Double beam.gif

A spectrometer has a source of infrared light (hot coil of nichrome wire) that emits radiation over the whole of the frequency range of the detector. The beam is then split into two beams of equal intensity. One beam is passed through the sample, while the other is a reference beam via a rotating disk, the beams are passing alternately. The intensities of the two beams are then compared and the wavelength over which the comparison is made is calibrated and dispersed on the detector via the prism.

[edit] G.3 Visible and ultraviolet spectroscopy

[edit] G.3.1

Absorption of d-block metal complex ions.

For d-block elements to behave as transition metals and form complex ions, the d-orbital has to be partly filled. By experimental data, it is now believed that the d-orbital is divided into five sub-orbitals. Three of them are less energetic and two of them are more energetic. The absorption of visible light causes the electrons to change energy levels within the d-orbitals.

[edit] G.3.2

Absorption of organic molecules.

“Simple” organic molecules are usually colourless since they absorb in the Ultraviolet part of the spectrum, however double bonds increase the wavelength of absorption, so that λ_max increases. This is especially true in highly conjugated molecules (those with many double bonds and a high conjugative effect) where the effect is so high that the compound appears to be coloured. One example is the natural pigment carotene, which is orange.

[edit] G.3.3

The compounds will usually absorb in the UV part of the spectrum (they appear colourless), however, where there are:

• Many double or triple bonds present (high conjugation),
• lone pairs of electrons about the atom,
• The d-orbital is not full, and the metal forms complex ions,

The absorption might extend into the visible part of the spectrum. Increased absorption will occur where there are many de-localised electrons, because it will be more possible for them to undergo electronic transitions. Base all of your predictions upon these facts. Try to think also, whether you know the compound, because it is sometimes the case that they give you a compound that you have used during a practical.

[edit] G.3.4

All calculations can be done using the following formulae, where:

• A is absorbance,
• T is transmittance,
• ε is extinction coefficient
• λ is the wavelength
• c is the concentration

A = -log T
T = ε λ c
T = I/I₀
%T = 100 x T

[edit] G.4 Infrared spectroscopy

[edit] G.4.1

Infrared radiation does not provide enough energy to promote an electron to a higher energy level, but does cause the electron to vibrate within its equilibrium position. Each bond can absorb radiation of the same frequency as the natural frequency with which an atom vibrate – the IR region of the electromagnetic spectrum. Two conditions are needed for absorption to occur:

• The radiation, wavelength of light has to be the same as that of the vibration of the atom
• The molecule has to have a change in dipole moment

Different molecules and functional groups are affected differently by IR and therefore their absorption of the latter changes:

H₂O: There are three kinds of vibration that cause a change in the dipole moment of water. They are: Asymmetric stretch vibration, Symmetric stretch vibration, bending vibration.

-CH₂-: CH₂ experiences many vibrations that cause its change in dipole moment, some concern only specific bonds – Symmetric stretching, Asymmetric stretching, In-plane rocking, In-plane scissoring, Out-of-plane wagging, Out-of-plane twisting. Other vibrations are that of the entire molecule.

SO₂: The kinds of vibration that cause the absorption of IR light of a sulphur dioxide molecule is the same as that of water.

CO₂: Unlike H₂O and SO₂, carbon dioxide does not have any lone pairs of electrons, therefore the structure is not bent but linear. Absorption will be therefore be caused only by asymmetric stretching and bending, a symmetric stretch will not cause a change in the dipole moment, and it therefore does not cause absorption.

[edit] G.4.2

The relationship between wavelength and wave number is inverse. The wave number is calculated per centimetre, that is λ^-1 the units for is are cm^-1. The higher the wave number, the higher the energy involved.

[edit] G.4.3

Interpretation

Just keep in mind that every functional group has a specific range of wave numbers not a single value of the wave number.

[edit] G.5 Nuclear magnetic resonance (NMR) spectroscopy

[edit] G.5.1

1H, 13C, 19F, 31P have magnetic moments (spin) of ½.

[edit] G.5.2

The case of 1H nucleus will be considered. As stated before its magnetic moment (spin) is ½, which means that in a magnetic field (N-S) it will act as a little magnet, aligning itself either in the same direction as the field (N-S) or in the opposite direction (S-N). These two states involve different energy levels. The one which is oriented according to the field has a lower energy than the one aligned against. However if ΔE (difference between the two states) is provided, the latter is in the radio wave range, the orientation of the proton will change (the energy is transmitted via rotation). Each proton will absorb in a specific range, which is dependent upon ΔE. The detector records whether a specific frequency is absorbed or not. Since the frequency is proportional to the strength of the field, it is the latter that is varied.

The frequency at which a proton absorbs is dependent upon the amount of shielding (by electrons) that a proton has. In covalent bonding electrons are shared, therefore there is a varying electron density around the proton (shielding) according to the environment the proton is in. As the magnetic field is applied to a molecule the electrons around a proton start circulating faster, this is called shielding, the higher the density of electrons, the higher the shielding, the greater the absorbance, because ΔE is greater. However in molecules, the electron density and with it shielding changes with the presence of different atoms, for example O usually attracts more electrons, therefore hydrogen in the vicinity of O has less shielding, and it absorbs less. All of the absorbance is relative to TMS (tetra methyl silane), chemical shift is calculated according to the amount of shielding around TMS. The lower the value of the chemical shift the higher the level of shielding (because it is closer to that of TMS). In high resolution NMR, multiple peaks can be observed, this is due to probability. The higher the probability that a certain arrangement of spins will occur the higher the peak will be, compared to the peaks of the same group. From multiple peaks the number of neighbouring hydrogen atoms can be determined. If the number of peaks is n the number of hydrogen atoms is n-1.

[edit] G.5.3

For interpretation, keep the following in mind:

• The peaks are in a range
• The relative heights (areas under the peaks) of peaks give you the ratios of the numbers of different hydrogen atoms, not the actual value
• The number of peaks in high resolution NMR give you the number of neighbouring hydrogen atoms as explained at the end of G.5.2

[edit] G.5.4

NMR in medicine

Since protons in water, lipids, carbohydrates and proteins give different signals (effectively only those of water and lipids are used), they are used to determine the arrangement of the latter, making a map of each plane of the section of the body that was scanned. It is used to determine the abnormalities of the tissues, for example to detect tumours. Since there are no known side effects or damages to the body the technique is used regularly.

[edit] G.6 Mass spectrometry & 12.1 The mass spectrometer

[edit] G.6.1

When a specimen is bombarded with electrons in a mass spectrometer, it forms positive ions. The biggest molecular ion is M+, and can then be further be split into smaller ions and neutral radicals. The detector in the mass spectrometer, detects only ions of positive charge (it has to be calibrated beforehand), the array of all the readings is called the fragmentation pattern. In a fragmentation pattern, the peak of highest abundance is assigned the name of base peak and the arbitrary abundance of 100. All the other fragments are then represented as peaks with heights that give their relative abundance (relative to the base peak).

[edit] G.6.2

Nowadays the relative atomic masses are very accurately determined. Mass spectrometers are also becoming very accurate instruments. The combination of the two can give us the molecular formula of the specimen very accurately. If we take the m/e ratio it is usually given to 4 decimal places, the number of combinations of relative atomic masses (which are also given to 4 decimal places, and beyond) that will give us the obtained m/e value is very limited, therefore the molecular formula of the compound can be determined. In some cases, however, the molecular ion does not form, as it is too unstable. In these cases the structure is more difficult to determine, though it is still possible.

[edit] G.6.3

For interpretation, keep in mind that:

• The peak with the highest abundance is due to the ion which is most easily formed,
• M+ does not necessarily make part of the fragmentation pattern,
• Peaks which have m/e (M+1), (M+2) or (M+4) are due to isotopes. (M+1) is usually due to 13C, (M+2) is due to 37Cl or 81Br, if the ratio M+/(M+2) is one, the peak is most certainly due to 81Br, if it is 3:1 the isotope involved is 37Cl. (M+4) is due to the presence of two 37Cl or 81Br.

12.1.1

The mass spectrometer: Image:Mass spectrometer.gif

The specimen is subject to the following stages during the process of mass spectroscopy. At first it is vaporised (1) then it is ionised (2), bombarded with electrons. The sample is then accelerated (3) by an electric field, deflected by a big magnet (4) and finally detected at the detector (5). See figure above.

12.1.2

The mass spectrometer gives us m/e ratios that can give us the mass of the fragment, if multiplied by the charge. Since the heights of the peaks give us the abundance of an isotope, we can calculate the relative atomic mass of an element, by comparing the heights of peaks at certain m/e, and calculating the average mass, relative mass can then be obtained from the charge.

[edit] G.7 Chromatography

[edit] G.7.1

Chromatography is a separating technique, it is usually the initial step of many analytical techniques. Substances separated by chromatography are then analysed and identified by mass spectroscopy (GC-MS is extensively used in drug and food testing). It can also be used as a technique to determine the purity of substances.

[edit] G.7.2

Components in a mixture have a different tendency to adsorb (interaction with particles on the surface of a solid, the greater the adsorption the smaller the movement) on to a surface or dissolve in a solvent. This provides a powerful means of separating the components, therefore all chromatographic techniques require a mobile phase (the solvent in the case of paper chromatography - PC) and a stationary phase (the paper in the case of PC). The mobile phase in a chromatographic technique passes the sample over a stationary phase, which causes different species of molecules in a sample to separate.

[edit] G.7.3

Adsorption: It is defined as the interaction between molecules in the mobile phase and particles in the stationary phase. It is very dependent upon the structure of molecules, different molecules interact differently with the stationary phase, this is used as a means of separation. The higher the adsorption the smaller the movement. This principle is the base of Paper chromatography, Thin-layer chromatography, column chromatography, liquid chromatography, gas chromatography gel permeation chromatography, affinity chromatography.

Partition: It occurs when molecules have a different solubility in different solvents, for example an organic molecule is scarcely soluble in water, but very soluble in an ether. The ether and water are immiscible, it is possible to separate the organic molecule and water, by passing it through ether. Another case of partition has to do with the size of molecules, by putting a “grid” only molecules of specific size will pass, thus being separated form the rest. Examples of techniques based on this are: high pressure liquid chromatography, liquid chromatography, column chromatography.

Ion exchange: This principle can be found in water softeners. A surface – resin is covered with ion acceptors, bound with Na⁺ (in the example of water softeners), when Ca²⁺ is passed through the resin, the two species of ions are exchanged. Example of a technique using this principle is ion exchange chromatography.

Molecular exclusion: In biochemistry it is very important to separate macromolecules. In affinity chromatography specific binding sites, selectively react, bind with specific substrates, and so remove the molecules from the mixture.

[edit] G.7.4

Paper chromatography: Stationary phase is paper, the mobile phase is a solvent. The sample is placed on paper, the solvent then separates the solvent by diffusion and adsorption of the sample.

Thin-layer chromatography: Stationary phase is a thin layer of gel on a hard surface, the mobile phase is a solvent. Similar functioning to paper chromatography.

Column chromatography: Stationary phase is an inert solid soaked with a solvent, mobile phase is a second solvent. The solvent is poured on the solid, and left to soak. The second solvent with sample is poured over and left to separate. Bands of different solutions form.

Ion exchange chromatography: Stationary phase is an ionic resin, mobile phase is a solution of ions. Once the resin and solution come in contact ion exchange occurs.

Gas-liquid chromatography: Stationary phase is a liquid, mobile phase is a gas carrying the sample. The gas is bubbled through the liquid, different substance separate, are detected and condensed.

High-performance liquid chromatography: Stationary phase is a grating, the mobile phase is a solvent, it is poured over the grid, and then high pressures are applied to speed up the process.

[edit] G.7.5

If food or drugs are involved, GC should be used. In the case of pigments, paper chromatography can be applied, in a mixture of ions ion exchange chromatography, in the case of molecules of different size are present HPLC should be applied. In the case of separation of Amino acids TLC should be used.

[edit] G.8 X-ray crystallography

[edit] G.8.1 & G.8.2

Image:X-ray.gif

The arrowed lines (V1 and V2) represent x-rays with an angle of incidence θ. It can be observed from the diagram that the rays are diffracted (but are assumed to be reflected) by the particles in the lattice, at the same angle θ, following the law of reflection. It can also be said that V2 travels a longer distance than V1. The difference is exactly WY+YZ. Since x-rays are waves the combination of two, can either reinforce the signal or diminish it. The waves are initially in phase, if WY+YZ can be expressed as nλ (an integer multiple of λ, the wavelength of the x-rays) the waves will remain in phase, however if this is not the case (i.e. the expression is (n + ½) λ) the waves shift out of phase. When the waves are in phase they can be summed and the signal is reinforced (constructive interference), but if the signal is reduced the waves are out of phase (destructive interference).

When we have the reading regarding the intensity of radiation (whether it is increased or decreased) we are able to calculate the distance between crystal planes – d. This can be done using Bragg equation.

Derivation of the equation:

Path difference of V1 and V2 = WY + YZ

Applying trigonometry, we can say WY+YZ = XY sin θ + XY sin θ = 2XY sin θ

XY is equal to the distance between the crystal planes, therefore 2XY sin θ = 2d sin θ, because the path difference can be expressed as a relation to the λ, so the whole equation is:

n λ = 2d sin θ

[edit] G.8.3

X-ray diffractometer:

All diffractometers are composed of 4 basic components. A source of x-rays, that are passed through a slit, to localise the waves, which are then diffracted by a crystal, which directs the light to photographic paper. The sample can either be a single crystal or a fine crystal powder, the signal is however different, in the case of a single crystal the reading will be made up of dots, while the fine powder will give a line, continuous reading. Very complex molecules, for example DNA give a combination of the two, because there are plenty superimposed atoms in the structure of DNA.

[edit] G.8.4

The unit cell is the simplest arrangements of atoms which when repeated will reproduce the whole structure.

[edit] G.8.5

Unit cells are usually cubes, therefore the calculations about the size can all be performed by using Bragg’s equation.

[edit] G.8.2

Just pay attention to the layers of the unit cell and Bragg’s equation gives you the distance between 2 layers.

[edit] G.8.6

Remember that hydrogen does not show on the electron-density diagrams/contour maps. The larger the ion, and the more negative it is and the more electrons there are around, the bigger it appears.

[edit] G.8.7

X-ray crystallography is used for the determination of the structure of molecules, crystals, even as complex as proteins. Even though it is very accurate it has some basic limitations. It can be applied only to solids and hydrogen is usually not detected. X-ray diffraction can give us the information about the arrangements of ions and atoms, however it can rarely be used for the identification of the species to which the atom belongs.