Mechanical Cell-Disruption Methods

Liquid shearing, such as high pressure homogenization, solid shear, such as bead milling and the extrusion of frozen cells are the most common mechanical methods for cell disruption (D souza and Killedar, 2008 Hatti-Kaul and Mattiasson, 2003). These methods are mainly based on the use of mechanical force to disrupt the cell. Some of these techniques, such as bead milling, have already found applications on a large scale (Prasad, 2010). 

In addition to these conventional methods, ultrasonication has also been used, where shock waves are created due to the generated high frequency vibrations that disrupt the cells. It is a simple method, but consumes a lot of power, uses probes that have short working times, and generates heat during the process. Thus, most ultrasonication systems are contained in a cooling jacket. 

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PHA is accumulated intracellularly in Gram-negative bacterial strains and normally its recovery after the fermentation include several steps - briefly these are (i) the separation of cells from the fermentation broth by centrifugation (ii) after that, the bacterial cells are pre-treated by heat, freeze dried, or salted, before extraction to avoid polymer degradation (iii) the PHA is therefore extracted, normally by using chlorinated solvents or other methods such as enzymatic digestion or mechanical cell disruption and (iv) PHA purification. The process of PHA recoveiy is shown in Figure 2.9. 

Cell disruption methods can be roughly divided into two categories mechanical ones, e.g., bead mill, homogenizers, and ultrasound and nonmechanicals like organic solvents, osmotic shock, supersonic fluid feed (SFF) and pulsed electric field (PEF). For extracting microalgal components, several methods were applied (Table 2.6). 

In general, the physical structure of the tissue must be broken down mechanically followed by an extraction procedure, before the sample can be analyzed. Homogenization using blenders, probe homogenizers, cell disrupters, sonicators, or pestle grinders is particularly useful for muscle, liver, and kidney samples. Regardless of the method used for tissue disruption, the pulse, volume of extraction solvent added, and temperature should be validated and standardized in order to ensure reproducible analytical results. During cell disruption, care should be taken to avoid heat build-up in the sample, because the analyte may be heat labile. 

The dates of these references tell us about the current interest in this topic. However, despite the promising results obtained, and the great interest paid to this new alternative method, the mechanism of cell disruption is still unclear and the quantification of the technological process is not yet precise. Furthermore, it would be essential to understand the relationship between the degree of inactivation and the dimensions and the structure of cell membranes, and the shape of the micro-organism. 

Physical Methods Physical methods include mechanical disruption by milling, homogenization, or ultrasonication. Typical high-speed bead mills are composed of a grinding chamber filled with glass or steel beads which are agitated with disks or impellers mounted on a motor-driven shaft. The efficiency of cell disruption in a bead mill depends on the concentration of the cells, the amount and size of beads, and the type and rotation speed of the agitator. The optimum wet solid content for the cell suspension for a bead mill is typically somewhere between 30 percent to 60 percent by volume. The amount of beads in the chamber is 70 percent to 90 percent by... 

There are a range of physical and chemical methods available at laboratory scale for cell disruption which involve the use of reagents or temperature and pressure changes to break the cell wall to release the desired products. However, at an industrial scale it is more common to use a mechanical disruption technique, and a number of companies have developed efficient... 

The choice of cell lysis method depends to a large extent on sample type. Mammahan cells, bacteria, and yeast all have different requirements for lysis depending on the presence or absence of a cell wall. In some cases, a combination of chemical and mechanical disruption may yield best results. Another important factor in the choice of lysis method is the sample size of cells to be dismpted. If only a very small volume of sample is available, care must be taken to reduce loss and avoid cross-contamination. Consideration should also be given to the compatibility of the chosen method with downstream applications. 

Physical or mechanical methods of cell disruption are the most widely researched in terms of containment. The underlying principle is either by breakage of the cell wall by mechanical contact, the application of liquid or hydrodynamic shear forces, or the application of solid shear forces. Cell disruption by non-physical methods generally involve simple operations which may be carried out in large tanks or vessels, which may or may not require agitation. 

The advantage of chemical treatment for disruption purpose compared to mechanical cell decomposition is its low energy consumption. But chemical methods are not very... 

Physical and mechanical methods are more suitable for cell disruption in general, because these methods cost less and do not affect intracellular biopolymer chemical integrity. Depending on the chemical method, temperature and material, it may affect chemical integrity if no judicious choice of solvents and operating conditions is made. Physical methods can be ultrasound, hydrocyclone, mill balls, Hughes press or osmotic pressure. 

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