Single-use manufacturing equipment for the production of certain biological compounds (e.g., recombinant proteins from mammalian cell cultures) makes good sense. Such equipment reduces water and energy use, decreases the need for equipment sterilization and waste treatment, and optimizes space in a manufacturing facility. But consider the plastic resins used to construct the disposable parts of such equipment. In BPI’s April 2014 issue, Tony Kingsbury discussed the fundamentals of how plastics are made. In this second installment of BPI’s series on bioprocess plastics, I explain why certain resins are used in single-use manufacturing equipment, how they are made, how their quality is ensured, and whether alternative resins are available readily.

Resins: Natural and Synthetic Polymers
By narrow definition, resins are hydrocarbon secretions of coniferous plants. They are naturally occurring polymers. Examples include amber; pine rosins used on violin strings; certain mastics; and two of the gifts brought by the Magi, frankincense and myrrh (1). Plant-made resins are built up from smaller units derived from isoprene, a five-carbon molecule with two double bonds.

Resins found in plastic compounds are synthetic polymers (although monomer units from which they are made can come from plants). Like plant-derived resins, synthetic resins are polymers derived from smaller molecules then hardened into solid, lacquer-like substances. The resin is the polymer itself, without the additives used to make a plastic compound more processable or to enhance its properties during its service life.

Types of Synthetic Polymers
Plastic resins are broadly classified as thermoplastic (heat softening) or theromoset. Thermoplastic resins such as polyethylene, polypropylene, polyvinyl chloride (PVC), and some urethanes soften and melt when heated. The different types of polyethylene include high density, low density, and linear low density. Different types of catalysts — notably Ziegler-Natta and metallocene — are used to polymerize polyethylene and polypropylene. Polymers produced with different reaction conditions and with different types of catalysts have significantly different properties (2).

During processing, synthetic polymers often are mixed with other compounding ingredients (e.g., antioxidants, pigments, and/or lubricants). The compounded polymer is heated until it softens and melts, then it is shaped (e.g., molded or extruded) to a desired form. Once a shaped polymer has cooled and hardened, it maintains its form throughout its service life. Each polymer has a characteristic melting temperature range that determines the limits of the temperatures under which a finished product can operate. Each polymer also has a characteristic melting temperature, range, chemical and water resistances, gas permeability, hardness, set of mechanical properties (e.g., tensile strength, elongation, and modulus), and other properties (3).

At the end of its service life, a part derived from thermoplastic resins can be recycled, and the resin is isolated and reused. Some thermoplastic resins also can be blended or alloyed with other thermoplastics to have properties intermediate between component polymers. Blended polymers must be fairly miscible with each other on a molecular level, but many polymers cannot be alloyed with others.

Thermosets such as phenol formaldehyde resins, epoxy resins, and others have chemical bonds that crosslink the polymer chains. They do not soften when heated and cannot be as easily recycled as thermoplastics. Like all plastics, thermosets can be incinerated, and the energy contained in their chemical bonds can be captured. Alternatively, thermoset plastics — like any large organic molecule — can be pyrolyzed (heated to high temperatures in the absence of oxygen), producing hydrogen and low–molecular-weight (MW) hydrocarbons, which can be used for some other purpose.

The thermoplastic/thermoset classification also includes some hybrids. Some thermoplastic polymers can be crosslinked after they are processed to enhance their mechanical properties. Some thermoplastics have reversible bonds between polymer chains so that they act somewhat like thermoset polymers but can be remelted. Thermoplastic elastomers (modified thermoplastics which behave similarly to synthetic rubbers) fall into this latter category.

Manufacture of Synthetic Polymers
Polymers (either synthetic or natural) are made up of small repeating (monomer) units that have been linked together. The monomer units may all be the same (homopolymer), or there may be two (copolymer) or three (terpolymer) different types of monomers in a single polymer. The polymerization or linking process typically involves either a catalyst-enabled polymer growth process (polyolefins), free radical chain reaction (PVC, polystyrene, and others), an acid–base chain reaction (polyurethanes, phenolic resins, polycarbonates, and others), or a condensation or water-loss reaction (polyesters, polyamides, and others). When the monomers used are gases at room temperature, the polymerization reactions are carried out in high-pressure reactors. Production of commercial quantities of almost any thermoplastic polymer is an expensive proposition, so there are not typically any small-scale polymer manufacturing operations.

For thermoplastic resins, the polymerization reaction is carried out in batch processes at high temperatures, room temperature, or temperatures in between. The finished product has a certain density and a certain average MW and MW distribution. It has a primary structure determined by the way adjacent monomer units are bound together. It also can have a secondary structure depending on attractive forces among components of the polymer chains. Polymers also have certain amounts of chain branching and certain random defects in the polymer structure, both factors that affect performance in the finished product. Finished resin particles as they come out of the reactor have a characteristic morphology (size and shape). As they come out of the polymerization reactor, thermoplastic resin particles have all of their properties fully formed. Further processing into finished plastic products involves melting and reshaping the polymer without changing its molecular weight or chemical properties. The newly made polymer contains tiny amounts of residual monomer; manufacturing by-products; polymerization catalysts; chain terminators; and possibly emulsifiers, surfactants, antioxidants, and water. Those substances can affect a polymer’s performance in certain critical end uses.

Thermoset resins are often polymerized in some sort of mold. Monomer units or prepolymers (liquids) react to form the cross-linked solid finished product. Because they cannot be remelted, thermoset polymers are not sold in particle form. Resins used in bioreactors are mostly thermoplastics of one type or another.

Quality Assurance for Thermoplastic Resins
Thermoplastic resin manufacturers routinely track certain properties of the resins they produce. The type of properties they measure depends on the type of polymer; examples include color; density; solvent extractables; melt index (a measure of melt viscosity); particle-size distribution; solution viscosity (a measure of molecular weight and molecular weight distribution); molecular weight distribution by gel permeation chromatography; and mechanical properties such as modulus, tensile strength, and elongation. Some measured properties are sales specifications (properties that a manufacturer certifies that a resin meets). Other properties are typical values (those that the manufacturer says are typical of the product but are not guaranteed to customers).

Statistical analysis of resin production data over time allows the manufacturer to determine the probability of each of the measured properties (particularly the sales specifications) being within the guaranteed limits on any batch of resin made. This process is known as statistical quality control, and it is one component of best manufacturing practices (4). To ensure that their product suppliers routinely follow best manufacturing practices, many customers require that their suppliers have ISO (or similar) certifications for their manufacturing units (5).

Selecting the Right Polymer for End Use
General Considerations: Selecting the right polymer for a finished product’s intended end use is important, regardless of what that end use is. Factors typically taken into consideration include

• service temperature range of the finished product
• compatibility with contacting chemicals
• compatibility with contacting plastics
• required mechanical properties (tensile strength, modulus, ultimate enlongation) of finished product
• impact resistance
• clarity and opacity
• permeability to gases
• ease of connecting system components. Additional specific requirements for certain end uses can include
• electrical resistivity (e.g., for wire insulation or antistatic flooring)
• stain resistance (e.g., for household goods)
• flame resistance (e.g., for children’s apparel)
• UV resistance (e.g., for roofing membranes)
• microbial resistance (e.g., for products susceptible to mold or mildew attack)
• water resistance (e.g., for submersible cables).

Selecting Resins for Single-Use Bioreactors: Plastic resins typically are used for several functions in bioreactors. A cell culture vessel is a multilayered disposable plastic bag. The inner (contact) layer is often made of plastics with low extractables, low cytotoxicity, and low cell-binding capacity; good resistance to acids and bases; and good water resistance. Good low-temperature flexibility also is required if cell cultures are to be stored at very low temperatures in a reaction vessel. Low-density polyethylene (LDPE) or polypropylene (PP) is sometimes used. A second layer of polyvinyl alcohol (PVA) or flexible PVC acts as a gas barrier. The outer layer of LDPE or polyethylene terephthalate (PET) provides mechanical strength. Flexible tubing is often made of C-Flex, a thermoplastic elastomer. The material has low extractables, low cytotoxicity, good flexibility, and good shatter resistance.

An impeller or mixing blade in a bioreactor is sometimes made of polysulfone, a high-strength, dimensionally stable, chemically resistant thermoplastic with the highest operating temperature range of any thermoplastic. Gas spargers (for introducing gas bubbles into a reaction vessel) are sometimes made of high density polyethylene (HDPE), a plastic with high strength, good dimensional stability, and low extractables. Ports (for introducing materials and removing samples) on a biocontainer are often made from LDPE for the same reasons that LDPE is used in the inner layer of a bioreactor vessel. Housing seals are often made from HDPE, ultra–high- density PE, or polysulfone because of these materials have exceptional dimensional stability, low cytotoxicity, and low extractables (6–8).

Bioreactors used for medical purposes (such as to manufacture drugs or vaccines) must meet certain FDA requirements (9). Often manufacturers use medical-grade resins (compliant with US Pharmacopeia, class VI standards) in these applications. On the other hand, bioreactors used for nonmedical purposes are not subject to particular standards or guidelines. However, general good manufacturing procedures dictate that bioreactors for nonmedical products have the same properties as those mandated for bioreactors for drug products. Low extractabilty and low cell toxicity are two such concerns. Likewise, the plastic’s chemical resistance, strength, dimensional stability, and other properties are largely dictated by the end use for successful cultivation of cells in suspension.

Sourcing Resins for Bioreactors: the Bottom Line
If a bioreactor is used for medical purposes then the manufacturer of its thermoplastic components has less latitude is substituting one resin for another. In any case, the manufacturer of a bioreactor used for drug products should use medical-grade ingredients (e.g., medical-grade resins) if available. Most manufacturers of thermoplastic resins have a limited number (if any) of medical grades available. They are extra-pure resins that have been through additional testing to ensure low levels of volatiles, extractables, and other impurities. To ensure reliable product quality from a thermoplastic resin manufacturer, a bioreactor manufacturer should always buy high-grade polymer from a supplier that uses best manufacturing practices as evidenced by ISO (or similar) certifications.

References

1 Frankincense & Myrrh: A Gift of Tree History. University of Georgia, Warnell School of Forestry and Natural Resources, December 2011; www.warnell.uga.edu/outreach/pubs/pdf/forestry/Frank%20&%20 Myrrh%20pub%202011.pdf.

2 Tullo AH. Metallocenes Rise Again. Chem. Eng. News 88, 2010: 10–18.

3 Van de Velde K, Kiekens P. Thermoplastic Polymers, Overview of Properties. Polymer Testing 20, 2001: 885–893.

4 ReVelle JB. Quality Essentials: A Reference Guide from A to Z. ASQ Quality Press: Milwaukee, WI, 2004: 185–186.

5 International Standards Organization (ISO); www.iso.org/iso/ home/standards/certification.htm.

6 Weber A, et al. Development and Qualification a Scalable, Disposable Bioreactor for GMP-Compliant Cell Culture. BioProcess Int. 11(4) 2003: S6–S17.

7 Barbaroux M, Sette A. Properties of Materials Used in Single- Use Flexible Containers: Requirements and Analysis. BioPharm Int. 2 November 2006.

8 Allegro STR 200 Single-Use Stirred Tank Bioreactor; www.pall.com/ pdfs/Biopharmaceuticals/253981-PALL-Allegro-STR-200-datasheet. pdf.

9 Guidance for Industry, Q7A Good Manufacturing Practice for Active Pharmaceutical Ingredients. US Food and Drug Administration: Silver Spring, MD; ww.fda.gov/ICECI/ComplianceManuals/ CompliancePolicyGuidanceManual/ucm200364.htm.

Paul Daniels is a retired senior staff chemist for Exxon Mobil Chemical; p.daniels1051@gmail.com.