The Processes - - BioPharm International


The Processes

BioPharm International

Fermentation Processes

Process developers must consider a complex set of conditions that affect cell propagation, product yield, and concentration of nutrients, waste, and products. The performance of a fermentor or bioreactor is governed by thermodynamics (such as the solubility of oxygen in the medium), microkinetics (such as cell growth and product formation), and transport of materials (moving nutrients into the cells, removing waste products, and so on). Optimal mixing ensures effective oxygen transfer, heat transfer, and dispersal of materials. Minor deficiencies in circulation of the medium can have major effects on growth and protein production. System designers therefore have to consider the fluid viscosity and momentum and the sizes of the cells involved.

A key decision in designing a process has to do with the timing of how nutrients and other ingredients are added to the reactor. Batch and fed-batch processes are commonly used for microbial fermentations.

In batch fermentation , medium and an inoculum of cells are added to the fermentor at the beginning of fermentation, the system is closed, and nothing but sterile air is added for the rest of the process.

Batch processes are simpler to scale up than other processes. They offer the flexibility often required, especially in multiproduct facilities, where several different cell lines are grown to manufacture different products. Because each batch is processed separately, there are periods of downtime, which are useful for cleaning and sterilizing the reactor. In a batch process, the environment changes as nutrients are depleted and product and metabolites accumulate. In a fed-batch process, fresh nutrients are added periodically. When nutrients approach depletion, the hungry cells are "fed." Frequent addition of fresh culture medium replenishes the nutrient supply. Various methods are used for controlling the rate at which the feed medium is delivered.

In some cases, the feed is delivered at a fixed rate, which is matched to the growth rate of the culture. Tighter feed control is often accomplished by linking feed rate to the culture's demand for nutrients as measured by the pH, or the amount of dissolved oxygen being consumed by the culture. High cell densities can be achieved in fed-batch fermentations, which, in turn, result in high levels of product formation.

Continuous culture is often used for assessing scale-up issues. A continuous feed of fresh medium is supplied, and fluid containing cells and cell products is removed at the same rate. Continuous culture offers several advantages over batch processes. Fermentor use is more efficient, with less downtime. High cell density and product output can be maintained for longer periods. Labor costs may be reduced because of less frequent cleaning. This method is appropriate for proteins that are continuously produced in large quantities.

In perfusion propagation , animal cells are held at high concentrations inside a growth chamber, and fresh medium is circulated around them. That provides continuous addition of nutrients and removal of waste products. Perfusion systems are commonly used in antibody production.

Suspension vs. Anchorage

Another key distinction is between suspended and anchored cells.

In suspension cell culture , the cells being grown float freely in the culture medium. Suspension cell culture is simple and easy to manage, and it works well for bacteria and yeasts, types of organisms in which each cell is a separate, independent entity.

Animal cells are a different matter. Most animal cells evolved to live not independently but as parts of tissues or organs. They are not naturally well adapted to growing in suspension.

Scientists have taken two general approaches to working with animal cells. On the one hand, cells can be engineered to make them adapt better to growing in suspension. Today, most of the mammalian cells commonly used in bioprocessing can be grown in suspension, if desired.

A suspension cell culture system must be delicately balanced. The mix of cells in suspension needs a lot of oxygen — but too much will kill them.

Mechanisms such as agitation and air sparging are used, but both can cause hydrodynamic shear stress leading to cell damage or death. Decreased oxygen concentrations slow or stop cell growth, but keeping dissolved oxygen at a high level causes formation of precipitates after growth has leveled off. Detergents can be used to lower surface tension and increase viscosity, thus protecting against shear and foaming.

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