TOPIC : Physical - Mechanical Methods of Cell Disruption

Cell Disruption Microorganisms are protected by extremely tough cell walls. Any potential method of disruption must that labile materials are not denatured by the process or hydrolyzed by the enzymes present in the cell. Methods suitable for large scale production can be divided into two major categories. Physico – Mechanical Methods Liquid Shear Solid Shear Agitation with abrasives Freeze- thawing Ultrasonication 2. Chemical Methods

Physical Mechanical Methods Liquid Shear : Most widely used method in large scale enzyme production procedures. Method is based on high pressure homogenization. Most commonly used homogenizer is APV – Manton Gaulin-homogenizer, which is a high pressure positive displacement pump with an adjustable valve and a restricted orifice. The microbial cell slurry is passed through a narrow channel between the valve and an impact ring followed by a sudden pressure drop at the exit to the narrow orifice. Pressure drop across valve is believed to cause cavitaion in the slurry and the shock waves are produced.

Working pressures are extremely high. 60 – 90% cell disruption can be carried out. Slurry can be cooled down to 0 to 4° C to minimize the loss in enzymatic activity due to heat generation.

Solid shear : Frozen microorganisms ( - 25° C) are passed through a narrow orifice by applying pressure. At laboratory scale Hughes press or an X press is used to obtain small samples of enzymes or microbial cell walls. Disruption is due to combination of liquid shear through a narrow orifice and the presence of ice crystals. The operating temperature can be further lowered down to - 35° C. 90 % of the cell disruption can be achieved in a single pass.

Agitation with abrasives : Carried out in a disintegrator containing a series of rotating discs and small beads. The beads are made up of mechanically resistant materials like glass , alumina , ceramics , and some titanium compounds. Up to 85 % of cell disruption can be achieved by this method. Problem associated with this method is heat generation which demands the use of cooling jackets.

Freezing – Thawing : Freezing and thawing of microbial cell paste will cause ice crystals to form and their expansion. Thawing leads to disruption of cells. It is a slow method with a limited release of cellular materials ,therefore, not a whole technique in its own. It is used in combination with other techniques.

Ultrasonication : High frequency vibration at the tip of an ultrasonication probe leads to cavitation . The shock waves thus produced cause cell disruption. Method is very effective on small scale . Power requirements are high. Large amount of heat is generated , therefore, cooling is needed. The probes has a short working life and are only effective over short range.