Upstream processing

The upstream processing element of the manufacture of a batch of biopharmaceutical product begins with the removal of a single ampoule of the working cell bank. This vial is used to inoculate a small volume of sterile growth medium, with subsequent incubation under 

Over half of all biopharmacuticals thus far approved are produced in recombinant E. coli or S. cerevisiae. Industrial-scale bacterial and yeast fermentation systems share many common features, an overview of which is provided below. Most remaining biopharmaceuticals are produced using animal cell culture, mainly by recombinant BFIK or CFiO cells (or hybridoma cells in the case of some monoclonal antibodies Appendix 1). While industrial-scale animal cell culture shares many common principles with microbial fermentation systems, it also differs in several respects, as subsequently described. Microbial fermentation/animal cell culture is a vast speciality area in its own right. As such, only a summary overview can be provided below and the interested reader is referred to the Further Reading section. 

Simple organic molecules Ethanol Butanol Acetone Acetic acid Lactic acid Saccharomyces cerevisiae Pachysolen tamiophilus, some Clostridium spp. Clostridium acetobutylicum, C. saccharoacetobutylicum Clostridium acetobutylicum, C. saccharoacetobutylicum Various acetic acid bacteria Lactobacillus spp. 

Amino acids Lysine Glutamic acid Corynebacterium glutamicum Corynebacterium glutamicum 

Enz5mies Proteases Amylases Cellulases Various Bacilli, e.g. Bacillus licheniformis Bacillus subtilis, Aspergillus oryzae Trichoderma viride, Penicillium pinophilum 

Main purification (chromatography) r Concentration (if necessary) and initial purification Cell harvesting and recovery of crude product 

Final product formulation Product filling, freeze drying (if required) and sealing - Labelling and packaging 

---

Improved feedstock pretreatment is important to minimize catalyst consumption and reduce subsequent spent-catalyst handling requirements. Selective hydrogenation of dienes can be used to reduce acid consumption, both in HF and H2SO4 alkylation (29). More effective adsorptive treating systems have been appHed to remove oxygen-containing contaminants that are frequently introduced in upstream processing steps. 

Rupture Disks A rupture disk is a device designed to function by the bursting of a pressure-retaining disk (Fig. 26-15). This assembly consists of a thin, circular membrane usually made of metal, plastic, or graphite that is firmly clamped in a disk holder. When the process reaches the bursting pressure of the disk, the disk ruptures and releases the pressure. Rupture disks can be installed alone or in combination with other types of devices. Once blown, rupture disks do not reseat thus, the entire contents of the upstream process equipment will be vented. Rupture disks are commonly used in series (upstream) with a relief valve to prevent corrosive fluids from contacting the metal parts of the valve. In addition, this combination is a reclosing system. 

Running different products through mill, or change in upstream process, resulting in different feed characteristics and unsuitable milling conditions, e.g. overheating due to blocked outlet... 

The delayed coking feed stream of residual oils from various upstream processes is first introduced to a fractionating tower where residual lighter materials are drawn off and the heavy ends are condensed. The heavy ends are removed and heated in a furnace to about 900 to 1,000 F and then fed to an insulated vessel called a coke drum where the coke is formed. When the coke drum is filled with product, the feed is switched to an empty parallel drum. Hot vapors from the coke drums, containing cracked lighter hydrocarbon products, hydrogen sulfide, and ammonia, are fed back to the fractionator where they can be treated in the sour gas treatment system or drawn off as intermediate products. 

Iron is usually present in FCC feed as tramp iron and is not cata-lytically active. Tramp iron refers to various corrosion by-products from upstream processing and handling. 

Product extraction Effluent and waste disposal Medium preparation Seed vessel Purification Cell free supernatant Cell biomass Production bioreactor Downstream processing Medium sterilisation Primary culture Upstream processing... 

Though the total degrees of freedom is seen to be (C + 4) some of the variables will normally be fixed by general process considerations, and will not be free for the designer to select as design variables . The flash distillation unit will normally be one unit in a process system and the feed composition and feed conditions will be fixed by the upstream processes the feed will arise as an outlet stream from some other unit. Defining the feed fixes (C + 2) variables, so the designer is left with ... 

This chapter aims to overview the manufacturing process of therapeutic proteins. It concerns itself with two major themes (1) sources of biopharmaceuticals and (2) upstream processing. The additional elements of biopharmaceutical manufacturing, i.e. downstream processing and product analysis, are discussed in Chapters 6 and 7 respectively. 

Biopharm production can be divided into upstream and downstream processing (Figure 5.5). Upstream processing refers to the initial fermentation process that results in the initial generation of product, i.e. the product biosynthesis phase. Downstream processing refers to the actual purification of the protein product and generation of finished product format (i.e. filling into its final product containers,... 

Upstream processing is deemed to commence when a single vial of the working cell bank system (see later) is taken from storage and the cells therein cultured in order to initiate the biosynthesis of a batch of product. The production process is deemed complete only when the final product is filled in its final containers and those containers have been labelled and placed in their final product packaging. 

The upstream processing element of the manufacture of a batch of biopharmaceutical product begins with the removal of a single ampoule of the working cell bank. This vial is used to inoculate a small volume of sterile media, with subsequent incubation under appropriate conditions. This describes the growth of laboratory-scale starter cultures of the producer cell line. This starter culture is, in turn, used to inoculate a production-scale starter culture that is used to inoculate the production-scale bioreactor (Figure 5.7). The media composition and fermentation conditions required to... 

Figure 5.7 Outline of the upstream processing stages involved in the production of a single batch of product. Initially, the contents of a single ampoule of the working cell bank (a) are used to inoculate a few hundred millilitres of media (b). After growth, this laboratory-scale starter culture is used to inoculate several litres/tens of litres of media present in a small bioreactor (c). This production-scale starter culture is used to inoculate the production-scale bioreactor (d), which often contains several thousands/tens of thousands litres of media. This process is equally applicable to prokaryotic or eukaryotic-based producer cell lines, although the bioreactor design, conditions of growth, etc., will differ in these two instances...

Overall, therefore, the routine manufacture of a biopharmaceutical product is initiated by large-scale culture of its producing cell line (upstream processing). Subsequent to this, the product is recovered, purified and formulated into final product format. These latter operations are collectively termed downstream processing and are described in Chapter 6. 

Downstream processing serves to (a) recover the therapeutic protein from its producer cell source upon completion of the upstream processing phase, (b) purify the protein and (c) formulate the protein into final product format. 

Upstream processing (fermentation) and downstream processing (purification and formulation) are physically separated, by being undertaken in separate buildings. Fermentation is generally undertaken in specially designed 42 000 1 stainless steel vessels. After recovery of the product from the cells, a number of chromatographic purification steps are undertaken, essentially within... 

---