1. Dnyanasadhana College, Thane. Department of Chemistry T. Y. B. Sc
Dnyanasadhana College, Thane. Department of Chemistry T.Y.B.Sc. Analytical Chemistry TURBIDIMETRY AND NEPHELOMETRY Dr.G.R.Bhagure

2. Theory, Instrumentation, Application

3. Theory
Origin of Scattering
A focused, monochromatic beam of radiation of wavelength l, passing through a medium containing particles whose largest dimensions are less than l is observed to scatter in all directions. For example, visible radiation of 500 nm is scattered by particles as large as 750 nm in the longest dimension.

4. Rayleigh, or small-particle scattering large-particle scattering
Elastic scattering
Rayleigh, or small-particle scattering
large-particle scattering
For larger particles, the distribution of scattered light increases in the forward direction and decreases in the backward direction as the result of constructive and destructive interferences.
Rayleigh scattering occurs when the scattering particles largest dimension is less than 5% of the radiation’s wavelength.

5. Interaction of Light with Particles  Small molecules  Light scattered symmetrically  

6. Interaction of Light with Particles Big molecules  Light scattered unsymmetrically  

7. Turbidimetry and Nephelometry
Turbidimetry and Nephelometry are two related techniques in which an incident source of radiation is elastically scattered by a suspension of colloidal particles.
In turbidimetry, the detector is placed in line with the radiation source, and the

8. Decrease in the radiation’s transmitted power is measured.
In Nephelometry, scattered radiation is measured at an angle of 90° to the radiation source.
The similarity of the measurement of Turbidimetry to absorbance, and of Nephelometry to fluorescence, is evident in the block instrumental designs shown in Figure In fact, turbidity can be measured using a UV/Vis spectrophotometer, such as a Spectronic- 20, whereas a spectrofluorometer is suitable for Nephelometry.

9. Turbidimetry A method in which the decrease in transmitted radiation due to scattering is
measured.
Nephelometry :A method in which the intensity of scattered radiation is measured at an angle of 90° to the source.

10. Applications Turbidimetry and nephelometry
Applications Turbidimetry and nephelometry are widely used to determine the clarity
of water, beverages, and food products. For example, the turbidity of water is determined
using nephelometry by comparing the sample’s scattering to that of a set of
standards. The primary standard for measuring turbidity is formazin (Figure 10.55),
which is an easily prepared, stable polymer suspension.26 Formazin prepared by mixing
a 1 g/100 mL solution of hydrazine sulfate, N2H4•H2SO4, with a 10-g/100 mL solution
of hexamethylenetetramine produces a suspension that is defined as 4000 nephelometric
turbidity units (NTU). A set of standards with NTUs between 0 and 40 is
prepared and used to construct a calibration curve. This method is readily adapted to
the analysis of the clarity of orange juice, beer, and maple syrup.

11. A number of inorganic cations and anions can be determined by precipitating
them under well-defined conditions and measuring the transmittance or scattering
of radiation from the precipitated particles. The transmittance or scattering, as
given by equation or is proportional to the concentration of the scattering
particles, which, in turn, is related by the stoichiometry of the precipitation reaction
to the analyte’s concentration. Examples of analytes that have been determined
in this way are listed in Table The turbidimetric determination of
SO4
2– in water following its precipitation as BaSO4 is described in Method

14. TURBIDIMETRY & NEPHELOMETRY 
Light interact with particle cause light scattering 
 Wavelength of scattered light is the same as of the incident light Detect by turbidimeter
 (detector is parallel to light source) or nephelometer
(detector is aligned at an angle °)

15. commonly used in immunoassay
detect AgAb complex particles (diameter = nm)
•Rayleigh-Debye light scatter 
use spectrophotometer in turbidimetry 
Light scatter in Nephelometry may be measured by end point or kinetic method

16. APPLICATIONS:
• Immunoglobulins,
• Specific proteins: Haptoglobin, Transferrin, Alpha 1 AT, Lipoproteins, Albumin
• Coagulation factors: Antithrombin III
• Theraputic drugs
Formation of immune complexes

17. Interaction of Light with Particles  Small molecules  Light scattered symmetrically  

18. Turbidimeter visible light Filter Photocell Detector Read Out Device
Sample Cell
visible light
Filter

19. Nephelometer visible light Light Trap Photocell Detector Sample Cell
Graduated Disc
Collimating Lens
Light Trap
visible light
Photocell Detector
Sample Cell
Read Out Device