Contract manufacturing organisation RentschlerBiotechnologie will add a 2,000-litre single-use bioreactor to befully operational end of the first quarter of 2015 to its facility inLaupheim, Germany, to match the growing demands forproduction capacity for all clinical phases.

Single-use technologies have gained broad acceptance inbiomanufacturing within the past few years, supporting flexibleand cost-effective clinical production. Beyond this, the firstmanufacturing processes for market products using disposableequipment (Shire, Protalis) have already been approved byEuropean and US authorities. Currently, two thirds of all newbioprocesses are carried out in single-use equipment. Thousand-litre single-use bioreactors (SUBs) represent the commonstandard, although 2,000L SUBs are on the advance. In general,an optimistic growth market in the double-digit range ispredicted.1

Compared with bioreactors made of stainless-steel, single-useproduction plants not only have a low contamination risk, butare also more cost-effective and faster to implement. The initialinvestment costs are approximately 40% lower, and sincesystems for cleaning and sterilisation (CIP/SIP) are obsoleteand time-consuming pipework is not required, the project lead-time for implementation of single-use production plants isreduced by at least eight months compared with stainless-steelreactors, which still have their place in commercial production.

Pharmaceutical customers also benefit from lower energy andmaintenance costs. Of particular note, however, is the highdegree of flexibility. It is possible to design processes modularlyusing single-use production systems and so to scale them easily;

very rapid product changeovers are possible, and as a result anoverall faster time to market can be realised.

Single-use systems with disposable components made ofplastic are also far less damaging to the environment thanwidely believed – particularly when compared with stainlesssteel reactors. Stainless steel reactors need continual cleaningand sterilisation, which results in a high consumption ofchemicals and ultrapure water. Single-use plants have a 46%lower total water consumption and a 35% more favourable CO2balance than stainless steel reactors.2

Due to the heating of large quantities of water for CIP andSIP, the energy consumption of stainless steel reactors isconsiderably higher than the production and disposal of plasticbags, the burning of which can also be used to recover energy.3

Rawlings and Pora have calculated that the total energyconsumption of single-use systems is about half of that ofstainless steel reactors.4 This means that the disadvantages ofdisposable systems – above all the higher costs for consumables– are more than offset by savings in water, energy andchemicals.

Challenges of single-use bio processesFor demanding processes – particularly in the case of high celldensities and product titers – the classical production process instainless steel reactors is still superior, especially during productharvesting. During harvesting, all cells and cell fragments areseparated from the process liquid normally using centrifugationand subsequent filtration. The centrifugation step in single-useprocesses, however, must be mapped with a cascade of deep-bed

October 2014 manufacturing chemist xv

There has been considerable progress in the application of single-use technologies inbiopharmaceutical production. CMO Rentschler Biotechnologie discusses recent advances

Cleanroom plant witha 1000L single-useproduction capacity

Employing new single-usetechnology in bioprocesses

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filtration steps, and low filtration capacities have to beaccepted.5 Single-use centrifuges have only recently becomeavailable, and Rentschler is working on the installation of sucha centrifuge to bypass the bottleneck in harvesting.

The capacities of chromatography systems, which are still toolow, are another challenge. For example, the titer of the 2,000Lsingle-use bioreactor is limited to 3g/L, because otherwiseproblems can occur during purification due to the small columnsizes. Furthermore suppliers are still working on thedevelopment of single-use sensors for measuring pH-value andoxygen. Until these disposable sensors become more robust,optical sensors made of glass or steel will have to be used.

The possible release of so-called leachables and extractablesfrom the plastic bags, which are normally gamma sterilised,represents a major challenge when polymer compounds are usedin single-use systems. Leachables and extractables dissolve outof the bags and can migrate into the cell culture medium.Extractables are defined as substances that are washed outfrom films, bags and tubes under harsh conditions, such as inthe presence of antioxidants, plasticisers or their degradationproducts. In contrast, leachables are washed out in the ongoingprocess. They not only jeopardise patient safety, but they canalso damage the entire manufacturing process.

Cytotoxic leachables are particularly undesirable underprocess conditions, because they adversely affect the growth ofthe cells, their vitality and consequently also the titer. For thisreason, a screening with mammalian cell cultures in addition tothe extractable studies established by the manufacturer makessense. Critical films can be identified at an early stage usingsuch screening, the quality control of single-use bags can beimproved and their implementation can be simplified.Rentschler and other users of single-use bags thereforeregularly carry out appropriate cell culture tests using its owncell lines, culture media and protocols. A standardised cellculture test was recently published from one Dechema workinggroup.6 The further purification progresses the more risk arisesfrom the leachables in terms of patient safety. For depletion ofthe leachables, Renschler follows a risk-based approach whichincorporates both manufacturer’s data and process data. Thisguarantees that depletion is carried out within defined limits sothat patient safety is ensured at all times.

Advances in single-use technologyBy enabling high levels of modularity and flexibility as well assignificant energy savings, single-use bioprocesses represent aparadigm shift in the production of clinical material. Someproducts, however, are better suited to production in stainlesssteel reactors, and commercial scale production is also usuallymore cost-effective in reusable stainless steel reactors. But here,too, a rethink is taking place. Rentschler is one of the first tollmanufacturers worldwide to establish a complete single-useplant for the upstream and downstream area. The company hasa flexible disposable concept consisting of two multi-productsingle-use bioreactor systems with 2 x 1,000L volume of workand an additional 1 x 2,000L bioreactor coming soon. Thisallows an easy scale-up as well as the reduction ofmanufacturing costs and product cycle times.

The disposable plant involves four independent all-purposecleanroom suites for operating the 100% mobile one-wayproduction plant for upstream processing and downstreamprocessing as well as an inoculum suite. All the cleanroomsuites are connected to a plant-wide data logging system whichis preconfigured for ‘plug and play’ of the mobile productionequipment. Incidentally, the company supplies all its plasticwaste to an incineration plant, thereby contributing to therecovery of a significant amount of heat energy.

What makes single-use facilities attractive is mainly the fact

that implementation is cheaper and faster. The initialinvestment is about 40% lower in comparison with a stainlesssteel facility at same scale. Furthermore, the project lead timefor implementation is reduced by at least eight months. In thenear future, the design of single-use production systems will beeven more modular to meet increasing and varying demands.Further technological developments of single-use equipment, inparticular centrifuges, sensors, chromatography and membraneadsorber, will help to resolve operational limitations.

REFERENCES1. E.Langer, R.Rader (2013): Innovation in Stainless-Steel Bioprocessing. LifeScience Leader, 31 October 20132. Ken Davis (2014). Single-Use Systems – Show Me the Green! ISPE SanFrancisco/Bay Area Chapter Newsletter Vol 19/2, 183. Guldager, N. (2010). Cost Advantages of Single-Use Technologies.Pharmaceutical Technology.4. Rawlings, B, and Pora, H. (2009). Environmental Impact of Single-Use andReusable Bioprocess Systems. BioProcess International 7, 18-26.5. Benjamin Minow et al. (2014). High-Cell-Density Clarification by Single-UseDiatomaceous Earth Filtration. BioProcess International 12(4), 36-47.6. Regine Eibl et al. (2014). Standardisierter Zellkulturtest zur frühenIdentifizierung kritischer Filme für CHO-Zelllinien und chemisch definierteKulturmedien. Dechema, January 2014

CONTACTRentschler Biotechnologie GmbH Erwin-Rentschler-Strasse 21D-88471 LaupheimT +49-(0)7392 701-0info@rentschler.de www.rentschler.de

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Virus filtration system for tangential filtration

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