Nephelometry | Turbidometry-Instrumental Methods of Analysis

Nephelometry is the method of measurement of the amount of light that is scattered by the particles which are suspended in the liquid medium. Nephelometry is used for the measurement of the size as well as the concentration of tiny microscopic particles like viruses, bacteria, and some other solids that are suspended in the water. Nephelometry may also be used for the measurement of the turbidity of water. With the help of the nephelometric method, one can get information about the quality as well as the safety of the sources of drinking water and other aquatic environments. So, in this article, I will talk about the complete basic information of nephelometry and turbidometry.

Nephelometry

Nephelometry is a scientific technique that is used to measure the amount of light scattered by particles suspended in a liquid medium. Generally, it is used in clinical laboratories for the measurement of the concentration of proteins like immunoglobulins as well as in blood samples. The basic principle of nephelometry is based on the scattering of light by the particles present in the sample.

In nephelometry, a beam of light is directed through the sample, and the amount of light scattered by the particles in the sample is measured at a specific angle. The scattered light intensity is directly proportional to the particles concentration in the sample. The scattered light is detected with the help of a photo-detector, and the signal is amplified and processed to calculate the concentration of the particles in the sample.

For the measurement of water pollution and monitoring the air continuously, nephelometry and turbidimetry are used. Turbidity is monitored in water whereas smoke and dust are monitored in the air. This technique is also used in the determination of the molecular weight of high polymers that are settled down on the earth from the many factories.

Turbidimetry is used for evaluating the weakening of the intensity of a light flux as it passes through a solution that is containing the suspended particles. Due to the scattering of light and absorption, the intensity decreases.

When light passes through comparatively stable suspensions, some of the incident radiant energy is lost through absorption, refraction, and reflection, while the remaining is transmitted. It is understood that the optical properties of each suspension will change depending on the concentration of the dispersed phase. This is because the turbidimetric analysis is based on comparing the intensity of the transmitted light through such suspensions to the concentration of the dispersed phase.

In another case, when the earlier mentioned suspension is seen at 90° (i.e., right angles) to the direction of the incident light, the system looks opalescent due to light reflection from the suspension's particles. The Tyndall effect indicates the dispersion of light. The visible opalescence or cloudiness is the net outcome of the suspension's irregularly and widely reflected light. As a result, the final measurement of scattered light intensity as an accurate representation of dispersed phase concentration provides the foundation of nephelometric analysis. (The word nephele is derived from the Greek which means cloud).

It has already been proven to be the most sensitive and effective, particularly in the case of very dilute suspensions with a concentration of less than 100 mg L. Although it is important to note that turbidimetric analysis is similar to flame photometry, and nephelometric analysis is similar to fluorimetry.

Nephelometry Principle and Turbidimetry

The scattering of light absorption caused by solid or colloidal particles suspended in solution is the basis for nephelometry and turbidimetry. When light passes through the suspension, some of the incident radiant energy is lost by absorption, reflection, and reaction, although the remaining passes through it.

 

The two types of instrumental analysis or techniques that are based on the phenomenon of light scattering (SOL) by particulate matter in a solution are nephelometry and turbidimetry. It is generally accepted that when a beam of light passes through two different mediums, the intensity of incident and transmitted light varies that is, in a transparent medium, the intensity of incident and the transmitted light is the same, whereas, in a medium containing suspended particles, the intensity of transmitted light is less than that of the incident beam due to interaction between particulate matter and the beam of light.

It is worth noting that there is no influence on the radiating strength of the light beam in either situation, if a sensitive photo-detector is put on the opposite side of the medium, the net change in intensity of incidence to the transmitted light beam is thus recorded.

However, if other parameters remain constant, the amount of fall in beam intensity that reaches the photo-detector is proportional to the concentration of particles that is responsible for scattering. The different angles with respect to incident and scattered beams are selected for measurement based on the dimension and extent of particulate matter present in the medium, for example, In nephelometric analysis, the power of a dispersed beam of light is measured at a 90-degree angle to the incidence beam, whereas in turbidimetry, the power of a scattered beam of light is measured at a 180-degree angle to the incidence beam.

Choice Between Both the Techniques

The choice between nephelometric and turbidimetric methods is determined by the two parameters that are given below;

Scattered Radiation Intensity vs Incidence Radiation Intensity

Turbidimetry is preferred over nephelometry when the medium that is to be analyzed has a high concentration of particle matter. However, when extremely dilute solutions have a less particulate matter, which results in less scattering, nephelometry is preferred over turbidimetry.

Size of Particulate Matters

Nephelometry is preferred when particles suspended in the medium are of smaller dimension because they scatter light at a right angle to the incidence beam, which is why nephelometric measurements are usually done at an angle of 90-degree to a collimated beam, whereas turbidimetry is preferred when particles of larger dimension scatter light to such an extent that measurements are done at an angle of 180-degree to collimated beam.

Factors Affecting Scattering of Light in Solution

1) The order and rare of material mixing within the solution.

2) Agitation of solutions that contain particulate matter.

3) The effect of solution temperature on component solubility

4) Medium viscosity.

5) Electrolyte or protective colloid presence or absence.

6) Concentration of solution.

7) Number of particulate matter that is suspended.

8) Particulate matter dimension.

9) Radiating beam wavelength.

10) The medium's refractive index.

The turbidimetric analysis is dependent based on measuring the intensity of transmitted light as a function of dispersed phase concentration.

log I0/It=k’lc            (1)

Nephelometry is not the same as turbidimetry. In the nephelometric analysis, through the sample solution, the light passes directly means through the particles that are suspended, and at an angle of 90°C, then the amount of scattered radiation is recorded and noted down. The analysis of nephelometry is based on the measurement of scattered light intensity as a function of dispersed phase concentration.

It is necessary to recognize that incident and scattered light have the same wavelength in nephelometry, however, scattered light has a longer wavelength than incoming light in fluorimetry.

Theory

The equation may be used to calculate the transmitted intensity I for turbidimetric readings.Where,

I0 = incident intensity

It = transmitted intensity

C = absorbing particles concentration in the solution

l = thickness of the solutions absorbing layer

The basic equation of turbidimetry is equation (1) and is very close to the equation of Bouguer - Lambert-Beer that is given below;

It = I0 x 10-k’lc

Where k' is the molar turbidity coefficient of the solution.

Turbidance is defined by the International Pharmacopeia as "a measure of the light-scattering effect of suspended particles", and Turbidity (meaning in Hindi) is defined as "a measurement of the reduction in incident beam intensity per unit length of a given suspension."

The process of measuring the amount of light scattered by suspended particles at right angles (perpendicular) to the incoming beam is known as nephelometry.

Turbidimetry and nephelometry can be used to quantify precipitates formed by the interaction of extremely dilute reagent solutions or other particular materials, such as colloidal dispersion of chemical and inorganic compounds and bacterial cell suspensions. (microbial assays).

However, it is important to note that to achieve the primary goal of obtaining equally accurate analytical results and consistent results, the following experimental parameters must be strictly followed when producing suspensions with reasonably uniform characteristic features, namely:

i) To solve the issues presented by birefringence, highly dilute solutions of bacterial cells may be utilized,

ii) The concentrations of the two ions that combine to form the proper precipitate, in addition to the ratio of concentrations in the mixed solutions,

iii) The procedure specifics, such as the sequence and rate of mixing,

iv) The concentrations of other salts and compounds present, such as protective colloids such dextrin, gelatin, and gum arabic; and

v) The temperature

Operating Conditions of Nephelometry and Turbidimetry

A variety of specifications must be met for nephelometric and turbidimetric analysis to be successful.

Because the amount of light scattered or absorbed depends on the size of the particles in the solution, accurate findings will be determined by the technique of creating the suspensions and the repeatability of their optical characteristics.

The optical characteristics of suspension and particle size are influenced by the following factors:

a) Concentration ratio of mixed solutions,

b) The order in which the solutions are mixed,

c) Ion concentration in the precipitate,

d) The temperature,

e) Mixing rate,

f) The presence of foreign electrolytes,

g) Dispersion stability,

h) Non-electrolytes present,

i) The presence of protective colloids, as well as

j) The amount of time necessary to achieve maximum turbidity.

Nephelometry has several advantages over other methods of protein quantification, including its high sensitivity, specificity, and accuracy. It is also a rapid and simple technique, making it ideal for use in clinical laboratories. Overall, nephelometry is a valuable tool for diagnosing and monitoring a wide range of medical conditions. I hope you have liked the article on nephelometry and turbidometry. In this article, I have provided complete basic information on nephelometry. If you have any points ask in the comment.