Methods for Cell disruption : The methods of cell disruption used to release the product depends on the composition of plasma membrane, presence or absence of cell wall, choice of location and the kind of product (enzymatic/non-enzymatic or simply bioactive substance). The different methods utilizes a unique mechanism to disturb the integrity of plasma membrane. These methods can be classify into 3 major section:
1. Physical Methods- These methods play with the physical parameters to damage the cell to release the product.
A. Thermolysis- This method is easy, economical and require no additional specialized equipment. It can be used only if the product is thermostable.This method gives a heat shock to kill the organism and as a result it disturb the cellular integrity without affecting the product. The effect of heat shock depends on ionic strength, presence of chelating agents such as EDTA and presence of other proteolytic enzymes.
B. Osmotic Shock- Most of the mammalian cell have a plasma membrane with active transporter to maintain the osmotic balance. Maintaining an osmotic balance is an active process with expenditure of energy. Prolonged exposure to the cell with hypotonic liquid such as water causes osmotic imbalance and ultimately causes lysis of cell. In this process due to inflow of water, cell swell and ultimately burst to release the products. According to the Hoff's equation, osmotic pressure Π is directly propotional to the concentration of solute and temperature.
R= Gas constant, T=absolute temp (K), Ci-Co =difference between total solute concentration inside and outside the cell (moles per litre)
Each mammalian cell has differential suspectibility towards osmotic shock. Red blood cells as shown in Figure 28.2, will be lysed with the addition of a tiny drop of water. Plant or bacterial cells are more resistant towards osmotic lysis due to presence of thick cell wall.
C. Sonication- A typical laboratory sonicator is given in Figure 28.3. A sonicator generates the ultrasound waves of frequency more than 20kHz to cause cell disruption by cavitation. The interaction of ultrasound with liquid causes compression and decompression very rapidly. The bubble formed in liquid, compresses several thousand atmospheres and gives shock waves to the cell wall or plasma membrane to cause cell lysis (Figure 28.3). Generation of ultrasonic waves in liquid causes rapid change in temperature and may cause thermal denaturation. Hence, ultrasonication medium needs to be cool and a long duration should be avoided.