Photometry: Principle, applications and types

Photometry

Fig. Principle of colorimetry

When light is passed through a coloured solution, certain wavelengths are selectively absorbed giving a plot of the absorption spectrum of the compound in solution. The wavelength at which maximum absorption is called the absorption maximum (λ_max) of that compound. The light that is not absorbed is transmitted through the solution and gives the solution its colour.

Photometric instruments measure transmittance, which is defined as follows:

                               Intensity of the emergent (or transmitted) light         Ie

Transmittance (T)= ------------------------------------------------------------- = ------

                                               Intensity of the incident light                    Io

Transmittance is usually expressed on a range of 0 to 100%.

If the concentration of the substance in solution is increased linearly, or if the path length that the light beam has to traverse is increased, transmittance falls exponentially. So a term absorbance is defined so that it is directly proportional to the concentration of the substance.

                                                Ie

Absorbance (A)= log1/T = log  ----

                                                Io

Absorbance has no units. Photometric instruments electronically convert the measured transmittance to absorbance values.

Cerebrospinal fluid (CSF) : Formation, composition, function, pathological state and biochemical analysis

A. Formation

Cerebrospinal fluid (CSF) is a clear, colourless fluid filling the ventricles and subarachnoid space. CSF production is a result of the combined processes of diffusion, pinocytosis and active transfer. The majority is produced by selective dialysis of blood plasma by a specialized sponge-like structure called the "choroid plexus" of third, lateral and fourth ventricles.

Fig. CSF collection

The anatomy of the ventricular system allows for movement of CSF in and around all the major structures of the brain. From the lateral ventricles located within the cerebral hemisphere, it circulates through the foramina of Monro into the third ventricle. At its caudal end, the third ventricle is connected by aqueduct of Sylvius to the fourth ventricle. CSF then flows into the basal cisterns and subarachnoid space by two lateral foramina of Lusckha and median foramina of Magendie. From the cisterns the CSF flows / throughout the subarachnoid space and over the hemispheric convexities and around the spinal cord.

Protein separation techniques: Protein fractionation

Fig. Protein purification steps

Proteins differ in their molecular size and charge. They can be separated on the basis of their following properties:

1.       molecular size

2.       solubility

3.       electrical charge

4.       adsorption properties

5.       specific bioaffinity.

Many techniques for protein purification exist, but the emphasis here is on some of the most popular procedures and the principles involved in their use.  Protein fractionation is required to separate and characterize a protein in detail.

Qualitative and qualitative estimation of Proteins

Proteins are the most abundant biomolecules, constituting more than 50% of the dry weight of the cell. They are known to perform many different biological functions.  Qualitative and quantitative estimation of proteins in biological fluids is based on the physico-chemical properties of proteins.

Denaturation of proteins

In each type of protein, the three dimensional structure is essential for its biological function.  A protein in the biologically active form is termed as the native protein and the native state is called “optimum” state. The disruption of the three dimensional structure is called denaturation.  On denaturation, the primary structure remains intact.  It is an all or none phenomenon and does not cause any alteration in the quantity of protein present.