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CELL DISRUPTION TECHNIQUES
INTRODUCTION: It is the process of breaking the cell membrane in order to extract the intracellular components efficiently like the extraction of genetic material etc. This process is important as many bio-technologically produced compounds are intracellular and must be released from cells before recovery. There are several technologies developed to use at industrial level. The technologies mainly used were bead mill, sonication and French press. enzymes, detergents and osmotic shock. Cell disruption techniques differ for different organism used, based on the composition of the cell membrane, size of the cell and at the scale at which the process needs to be done. Many aspects should be considered while selecting the method of cell disruption for a specific batch. It can be either mechanical or non-mechanical which are further divided to physical and chemical methods. As it is a downstream process, factors that influence the selection of disruption method include the susceptibility of the cells to disruption, product stability, the ease of extraction from the cell debris, the speed of the method and the cost of the method.

MECHANICAL PHYSICAL METHODS: Mechanical methods involve cell disruption by high pressure. In this, additional cooling systems are required as mechanical pressure adds to heat in the system.

FRENCH PRESS AND HOMOGENISER: It is one of the earliest techniques used and is based on the sudden variations in pressure which causes the cell to disrupt. Cell suspension is first passed through the narrow passage called orifice in high pressure and then is released into a chamber of less pressure where it drops to atmospheric pressure level. Due to the drop, the cell explodes allowing extraction of intracellular components. More number of passages are required in order to regulate the pressure variation over organism. Protein release (R) is first order with respect to the number of passes (N)

ln(Rm÷Rm−R) = kNP^a (R is protein released (kg protein/kg biomass), Rm is max protein available, k is rate constant and is a function of temperature, a is the pressure exponent)

BEAD MILL: In this generally beads are used to disrupt the cell based on the location of the required product. The interaction between beads create stress and applies pressure on the cell. Disruption is due to the grinding of beads against the cell suspension. Consists of jacketed grinding chamber with a rotating shaft at the centre. agitators are fitted at the shaft that provides the kinetic energy required for the beads to collide. Based on the type of the cell, bead size is chosen.

ULTRASOUND: In this method, the frequencies produced by the waves are converted into mechanical oscillations by a transducer. It is often used for very small sample volumes, the cell suspension is sonicated with an ultrasonic probe. Ultrasonic vibrators produce high frequency sound with a wave density of about 20kHZ/s which is required for the rupture of the membrane.

OSMOTIC SHOCK: Internal cell conditions needs to stabilized with the outer environment of the cell in order to coexist. Even though cell can regulate the specific conditions required, it fails adjust sudden variations in less time. Hence, by changing the osmotic pressure by giving high or low salt concentration and then changed to the opposite concentration creates a stress making the cell lyse. An isotonic cell under high salt concentration environment loses waster and when is shifted to low salt concentration environment it immediately takes up water and hence explodes because of internal pressure. However, this method requires initial enzyme pre-treatment in order to weaken the cell

NON MECHANICAL ENZYME AND CHEMICAL METHODS: Enzymatic and chemical methods differ with organism based on the cell wall composition. These react with cell membrane components to lyse them. Some detergents break the protein-protein interaction (Sodium dodecyl sulphate(SDS)), few of them solubilise the phospholipase (cationic detergents). They have different mechanisms to lyse the cell. Solvents like alcohol, methyl ethyl ketone etc act on the cell membrane lipids and lyse them helping in extracting the intracellular components. Enzymes are specific in nature and hence different organisms require different enzymes to deal with. For example, lysozyme for gram positive bacteria, zymolase for yeast and fungi etc. some enzymes that are used in common are cellulases, pectinases etc. Even though using different single method of disruption method has its own efficiency, efficiency can be highly scaled up on using combined methods of physical, chemical and enzymatic methods.

REFERENCES [1] 2020. Cell Disruption Methods. Natalia Kakko, Nicoletta Ivanova, Anssi Rantasalo.

[2]Cell disruption technologies. E. D’Hondt, J. Martı´n-Jua´rez, S. Bolado, 2020.

[3]Cell disruption for microalgae biorefineries. E.GünerkenabE.D'HondtaM.H.M.EppinkbL.Garcia-GonzalezaK.ElstaR.H.Wijffelsbc, 2020.

[4]K. McCabe, "Common techniques and technologies for cell disruption", Microfluidics-mpt.com, 2020. [Online]. Available: https://www.microfluidics-mpt.com/blog/common-techniques-and-technologies-for-cell-disruption. [Accessed: 30- Jul- 2020].

[5]New Approaches for the Use of Non-conventional Cell Disruption Technologies to Extract Potential Food Additives and Nutraceuticals from Microalgae. Francisco J. Barba, Nabil Grimi & Eugène Vorobiev, 2020.