UV-Visible Spectroscopy for Colorimetric Applications

*Sonia Karuppaiah and Manikandan Krishnan*

#### **Abstract**

UV-visible spectroscopy is an interpretive skill that amplitude the variety of different wavelengths of UV or visible light, which are captivated by or transferred via a pattern new assessment to an implication or blank constituent. This asset is encouraged by way of the pattern combination, doubtlessly subject to network on what is within the representative and at what attention. Because this spectroscopy execution confides on the control of mild. Therefore, illuminate can be described by its wavelength, which can be useful in UV-visible spectroscopy to analyse or identify different substances.

**Keywords:** detectors, filters, monochromators, sources, UV-visible spectroscopy

#### **1. Introduction**

The analytical chemistry is based on the quality of colour in coloured solution, we observe the colour, the colour's depth, or intensity. These observations led to the technique called colorimetry, the colour of a solution identify species while the intensity of the colour depends on identifying the concentration of the species present. The important and sensitive colour tests have been developed for the detection and determination of a wide range of chemical species, both inorganic and organic in nature, this used the development of visible and ultraviolet spectrometer [1].

The wavelength range of UV radiation starts at 400 nm, the blue end of visible light, and ends at 200 nm. The radiation has sufficient energy to excite electrons. When light passes through the solution and emerges as red light, then the solution is red. Because the solution has allowed the red component of white light to pass through, whereas if the solution has led the red component of white light to pass through because it has absorbed the complementary colours, yellow and blue [2].

If the solution has more concentration, more yellow and blue light is absorbed, and more intensely red solution appears to the eye. There is a difficulty in comparing the intensity of the two colours. The wavelength range of UV radiation starts at the end of visible light of 400 nm and ends at 800 nm [3]. The atoms or molecules have sufficient energy to excite valence electrons. Visible light starts the wavelength from 800 to 400 nm.

## **2. Theory**

#### **2.1 Electronic excitation in molecules**

The atoms are held strongly by sharing electrons in a molecule. The electron in a molecule moves in molecular orbitals at discrete energy levels. When the energy of the electrons is at a minimum, the molecules are in the lowest energy state or ground state. The molecules can absorb radiation and move to a higher energy state or excited state. The movement of electrons from a higher energy state is called electronic excitation [4]. The frequency captivates or effuse by a molecule and the power is related by, ΔE=hγ. The amount of energy required is based upon the variation in energy linking the ground state E0 and the excited state E1 of the electron. It is stated as ΔE ¼ E1 � E0 ¼ hγ

where, E1 is the energy of the excited state.

E0 is the energy of the ground state.

The full strength of a molecule is the same as the sum of electronic, vibrational, and rotational electricity. The importance of the energies decreases inside the following order: Eelec, Evib, and Erot. Ultraviolet energy is computed, the assimilation spectrum arising from a single electronic transition must contain a single discrete line. However, an awesome line is not obtained because digital absorption is superimposed upon rotational and vibrational sublevels. Suppose of complex molecules in conjugation with an excess of two atoms, discrete bands merge to bring about broad absorption bands or "band envelops" [5]. Three distinct types of electrons are involved in organic molecules. They are as follows:


#### **2.2 Electronic transition in organic molecule**

A rule to predict how molecules undergo a transition is given by Quantum mechanics. Some transitions are "allowed" while others are "Forbidden."


which undergo these transitions are saturated halides, alcohols, ethers, aldehydes, ketones, amines, etc. This transition requires less energy. In saturated alkyl halides, the energy required for such a transition decreases with the increase in the size of the halogen atom.

In methyl chloride and methyl iodide due to the electronegativity of chlorine atom, the n electrons on chlorine atom are comparatively difficult to excite, whereas the methyl iodide is 258 nm as n electrons on iodide atom are loosely bound.


*2.2.1 Beer's and Lambert's law*

There are two laws related to the absorption of radiation [7].

I ¼ Ia þ It

I = Intensity of incident light.

Ia = Intensity of absorbed light.

It = Intensity of transmitted light.

#### *2.2.2 Beer's law*

The intensity of a beam of monochromatic light drops exponentially with expanding in the concentration of absorbing species arithmetically.

**Figure 1.** *The electronic transition.*

� *dI dc* <sup>∝</sup>I the decline in the intensity of incident light*;* with concentration, C is proportional to the strength of incident light, IÞ � *dI dc* <sup>¼</sup> K I eliminate and introducing constant proportionality K � *dI dc* <sup>¼</sup> K dc rearranging terms ð Þ � ln I ¼ Kc þ b on integration ð Þ

(1)

When concentration, C = 0, there is no absorbance I = I0. Exchange in Eq. (1).

$$\begin{aligned} -\ln \mathbf{I}\_0 &= \mathbf{K} \times \mathbf{0} + \mathbf{b} \\ -\ln \mathbf{I}\_0 &= \mathbf{b} \end{aligned}$$

Substitute the value of -ln I0 = b in Eq. (1).

$$\begin{aligned} \text{\$-\$\ln \$I = Kc -\$\ln I\_0\$}\\ \text{\$-\$\ln \$I\_0 -\$\ln I = Kc\$}\\ \text{\$\ln \$I\_0/I = Kc\$ (Since \$\log A - \log B = \log A/B)\$}\\ \text{\$I\_0/I = e^{Kc}\$ (separate natural logarithm)}\\ \text{\$I/I\_0 = e^{-Kc}\$ (reversed on both sides)} \end{aligned} \tag{2}$$

#### *2.2.3 Lambert's law*

The rate of decrease of intensity (monochromatic light) with the thickness of the medium is directly proportional to the intensity of incident light.

� *dI dt* <sup>∝</sup>I the decline in the intensity of incident light*;* with concentration, C is proportional to the intensity of incident light, I � *dI dt* <sup>¼</sup> K I separate and introducing constant proportionality K � *dI <sup>I</sup>* <sup>¼</sup> K dt reposition terms ð Þ � ln I ¼ Kt þ b on integration ð Þ (3)

When concentration, t = 0, existent is never absorbance I = I0. Substituting in Eq. (3).

$$\begin{aligned} -\ln \mathbf{I}\_0 &= \mathbf{K} \times \mathbf{0} + \mathbf{b} \\ -\ln \mathbf{I}\_0 &= \mathbf{b} \end{aligned}$$

Substitute the rate of -ln I0 = b in Eq. (1).

*UV-Visible Spectroscopy for Colorimetric Applications DOI: http://dx.doi.org/10.5772/intechopen.101165*

$$\begin{aligned} & -\ln \text{ I} = \text{Kt} - \ln \text{ I}\_0\\ & -\ln \text{ I}\_0 - \ln \text{ I} = \text{Kt} \\ & \ln \text{ I}\_0/\text{I} = \text{Kt} \left( \text{Since } \log \text{A} - \log \text{ B} = \log \text{ A/B} \right) \\ & \ln \text{I}\_0/\text{I} = e^{\text{Kt}} \left( \text{removing natural logarithm} \right) \\ & \text{I}/\text{I}\_0 = e^{-\text{Kt}} \left( \text{Inverse on bilateral} \right) \end{aligned} \tag{4}$$

Combine and equate Eqs. (3) and (4)

$$\mathbf{I}/\mathbf{I}\_0 = e^{-Kct}$$

$$\mathbf{I} = Ioe^{-Kct}$$

<sup>I</sup> <sup>¼</sup> *Io*10�*Kct* <sup>ð</sup>Converting natural logarithm to base 10&K <sup>¼</sup> <sup>K</sup> � ð Þ <sup>0</sup>*:*<sup>4343</sup>

<sup>I</sup>*=Io* <sup>¼</sup> <sup>10</sup>�*Kct* ð Þ reposition terms Io*=*<sup>I</sup> <sup>¼</sup> <sup>10</sup>*Kct* ð Þ reverse on both sides

Log Io*=*I ¼ Kct Taking log on both sides ð Þ

Transmittance (T) = I/Io and Absorbance (A) = log 1/T. Hence A <sup>¼</sup> log <sup>1</sup> *I=Io:*

$$\mathbf{A} = \log \text{Io/I} \tag{6}$$

(5)

Substitutes Eq. (6) in Eq. (5)

$$\begin{aligned} \mathbf{A} &= \mathbf{K} \mathbf{C}t \text{ (Instead of } \mathbf{K} \text{, we write } \mathbf{e})\\ \mathbf{A} &= \mathbf{e} \mathbf{c}t \end{aligned}$$

Where, A = Absorbance or optical density or extinction coefficient ε = molecular extinction coefficient C = Concentration of drug (mmol/lit) T = pathlength (1 cm) ε can also expressed as

$$e = E \sum\_{1cm}^{1@} \text{\textit{\chi}} \text{Molecule weight} / 100$$

Where *E* P1% <sup>1</sup>*cm* means the absorbance of 1% W/V solution using a path length of 1 cm.

#### **2.3 Deviation from Beer's and Lambert's law**

Beer and Lambert's law is found to be obeyed by the system if a straight line passes through the origin and a graph is plotted between absorbance and concentration.

But there is always a deviation from the linear relationship between the absorbance and concentration particularly at higher concentration, and hence the absorption curve changes with the change in concentration of the solution. The deviation may be positive or negative, if the resulting curve is concave upwards it is called positive deviation. If the resulting curve is concave downwards it is called negative deviation, which is depicted in **Figure 2** [8].

#### *Colorimetry*

**Figure 2.** *Deviation from Beers & Lamberts law.*

### **2.4 The reason for changing deviation from Beer's law**

#### *2.4.1 Instrumental deviation*

Factors like stray radiation improper slit width, fluctuations in single and when monochromatic light is not used.

#### *2.4.2 Physiochemical changes in solution*

i. The law does not hold if the substance ionises, dissociates, or associates in solution. Since the nature of the ionised species in solution varies with the concentration.

Example: Benzoic acid in benzene is associated to form dimer and hence deviation occurs.


#### **3. Effects of solvents UV spectra**

Chromophore: the term chromophore is used to denote a functional group or presence of some structural feature that gives colour to a compound [9].

Example: Nitro group is a chromophore because its presence in a compound gives the yellow colour to the compound. It can be defined as any group which exhibits absorption of electromagnetic radiation in the visible or ultraviolet region. It may or may not impart any colour to the compound. Some of the important chromophores are ethylenic, acetylenic, carbonyls, acids, esters, nitrile group, etc.

There are two types of chromophores. The chromophore in which the group contains π electrons and they undergo n ! π\* transitions, the compounds like ethylene, acetylene, etc.

The other type of chromophore contains both π electrons and n (non-bonding) electrons. This type of chromophore undergoes two types of transitions, π ! π\* and n ! π\* and examples include carbonyls, nitriles, azo compounds, and nitro compounds.

#### **3.1 Changes in position and intensity of absorption**

For isolated chromophore groups such as >C=C < and -C � C-, absorption takes place in the far ultraviolet region which cannot be easily studied.

But the role of absorption is maximum and the intensity of absorption can be edited in exceptional approaches by some structural adjustments or change of solvent.

#### *3.1.1 Bathochromic shift or redshift*

It involves the shift of absorption most in the direction of longer wavelength because of the presence of certain groups such as OH and NH2 called auxochromes or by change of solvent. A Bathochromic shift is also produced when two or more chromophores are present in conjugation in the molecule.

Example: Ethylene shows π ! π\* transition at 170 nm, whereas 1,3 -butadiene (where two double bonds are in conjugation) shows λmax at 217 nm.

#### *3.1.2 Hypsochromic or blue shift*

The shift of absorption maximum towards shorter wavelength and may be by the removal of conjugation or by change of solvent. The absorption shift towards a shorter wavelength is also called the blue shift.

Example: Aniline shows maximum absorption at 280 nm, because the pair of electrons on the nitrogen atom is in conjugation with the π bond system of the benzene ring. In acidic solution, a blue shift is caused and absorption takes place at a shorter wavelength 200 nm. The electron pair is no longer present and hence conjugation is removed.

#### *Colorimetry*

**Hyperchromic effect:** The effect is due to an increase in the intensity of absorption and it is brought about by the introduction of an auxochrome.

**Hypochromic effect:** It involves a decrease in the intensity of absorption and is brought by groups that are able to distort the geometry of the molecule.

**Auxochrome:** It is a group that itself does not act as a chromophore but when attached to a chromophore it shifts the adsorption maximum towards a longer wavelength along with an increase in the intensity of absorption.

#### **3.2 Instrumentation**

The various components of a UV-VIS spectrophotometer are as follows [3]:

