Adeline Siew is science editor for BioPharm International.

Adeline Siew, PhD

The packaging or container closure system that is meant to protect a pharmaceutical product can be a source of contamination. Comprehensive extractables and leachables (E&L) studies are, therefore, required to identify and quantify harmful impurities that could affect the quality and safety of drug products.

spoke with Lester Taylor, Pharma marketing manager, Agilent Technologies; Andrew Blakinger, manager, Extractables and Leachables Testing, Eurofins Lancaster Laboratories; and Fran DeGrazio, vice-president, Global Scientific Affairs and Technical Services, West Pharmaceutical Services, about the ins and outs of extractables and leachables assessments in biologic drug products.  

 What are the E&L challenges for biologics compared to small-molecule drugs?

 Compared to small-molecule drugs, biologics face additional challenges. For example, the efficacy of a biologic drug may potentially be reduced through undesirable interactions of leachables with drug molecules through post-translational modification (PTM) biochemical reactions (e.g., oxidation, aggregation, clipped variants, unfolding, adducts formation, and glycosylation). Alternatively, a leachable arising from single-use systems (SUS) or components used for bioprocessing may adversely affect the manufacturing process through cellular toxicity and Chinese hamster ovary (CHO) cell death thereby reducing the productivity of the bioprocess. There are several examples where leachables have been associated with these undesirable effects on biologic manufacturing and drug efficacy, leading to major manufacturing losses and even worse, dangerous side-effects and loss of drug efficacy. 

 The evaluation of biologics for leachables presents many unique challenges. The protein itself can interfere with testing, so removal prior to analysis may be warranted. But if care is not taken, this process can unintentionally remove potential leachables, resulting in false negatives, or it may lead to contamination of the sample that may result in the generation of false positives. 

	Other ingredients in large-molecule formulations, such as polysorbate 80 and other surfactants/stabilizers, can also cause issues. These compounds often interfere with chromatographic analyses in the form of multiple large peaks that display numerous ions by mass spectrometry throughout the retention time window. These large surfactant peaks can easily mask leachables. Furthermore, proteins, surfactants/stabilizers, and other ingredients in large-molecule formulations are difficult to clean from mass spectrometers and, therefore, may carry over from one analytical run to the next if not dealt with properly. 

 The likelihood of leachables in any drug product will depend on the packaging materials, type of formulation ingredients, and conditions of use. The occurrence and impact of leachables in biologic products can present greater challenges compared to that of small synthetic molecules due to several factors. Biologics are living molecules that can be difficult to solubilize and stabilize, and quality attributes are not easily characterized compared to small molecules. The formulation ingredients for biologics often contain co-solvents or surfactants and will have more propensity to extract chemicals from packaging materials compared to typical small-molecule formulations. 

	Biologic products are complex and very sensitive to their environments. Extractables or potential leachables that may migrate into a drug product have the potential to interact, and therefore, affect the product quality, safety, or stability. In general, biologic products are formulated to solubilize, stabilize, and optimize pharmacokinetic properties consistent with the route of administration. Anything that migrates from the packaging that could interfere with this optimized environment is of concern. This includes interactions with active or excipients in a drug product formulation that lead to said quality, safety, or stability issues.

	Additionally, large molecules have greater surface areas with sites that have a propensity for interactivity based on polarity and charge. This can lead to conformational modifications and other interactions that may impact product quality.

	Primary packaging and container closure systems

 What are the key considerations when selecting primary packaging material for biologic drug products?

With every drug product and especially biologics, the most inert primary package possible must be chosen to minimize the potential for interactions to occur. Potential leachables are not the only interaction of which to be wary. Because of their reactive nature, biologic drug products can adhere to surfaces or absorb into materials. An understanding of possible interfacial interactions must be a consideration. In addition, there are other packaging considerations that must be addressed, such as container closure integrity, particle generation, and other performance concerns.

 For any drug product, it is crucial to ensure the packaging does not adulterate the drug product. Any compounds that leach from the packaging could affect the product in a variety of ways, including impacting patient safety if compounds are toxic or interfering with other analytical assays during release testing. 

	There are a number of other potential E&L risks that are unique to large molecules. Leachables may cause conformational changes in the protein or may cause the protein to aggregate. Large-molecule drug products may also chelate inorganic leachables. These types of interactions can increase the toxicity of the drug product, reduce the product’s efficacy, or affect the product’s stability. It is, therefore, important to fully evaluate the E&L risks to avoid costly delays in getting a product to market. 

What components in a container closure system can pose E&L risks to a biologic drug product?

 Typically, the container and closure components that come into direct contact with the drug product usually have the highest impact in terms of leachables observed. However, there have been many examples of leachables arising from package labels such as the inks or adhesives, as well as from secondary packaging components. These risks should, therefore, be assessed during bioprocess development.

Nearly any component in a container closure system may pose E&L risk to a biologic. Because many biologics are packaged in prefilled syringes, some of the most common components of concern are rubber stoppers. Rubber stoppers are notorious for containing nitrosamines and polynuclear aromatic hydrocarbons (PAHs), both of which are carcinogenic. Glass prefilled syringes are another common example of a component type posing a special risk to biologics. During manufacturing, tungsten pins are used to hold open the fluid path in the syringe barrel. Because manufacturing occurs at extremely high temperatures, the formation of tungsten oxides is possible. The residual tungsten oxide on the glass syringe can then leach into the final biologic drug product and cause protein aggregation or degradation. 

The most common primary packaging system for a biologic drug product is a vial system. This system is composed typically of a glass vial with an elastomeric rubber stopper and an aluminum seal with a plastic flip-off button. The other common primary package is a prefilled syringe system, which is typically a glass syringe with an elastomeric plunger and a tip cap or needle shield. Each of these components has the potential to leach substances into a drug product with contact over time. Of course, the extractables of most significant concern from glass materials are metal ions. It is well known that some biologics drugs are sensitive to various metal ions. Although these reactions are drug-product specific, these reactions are a consideration when evaluating packaging components.

	Other types of extractables are expected from elastomeric components. Elastomeric components are composed of much more than just the base polymer. Elastomer formulations typically have six to 12 added ingredients that are mixed with the base polymer under heat and pressure. This process causes chemical crosslinking to occur, which result in the formation of reaction products. These reaction products, along with residual compounds of the original raw materials, may interact with the active drug product or environment. Many of these compounds are organic; some may be inorganic, and, therefore, provide an additional source of metal ions.

	In the case of a prefilled syringe system, there is the potential for even more extractables. A glass syringe may be formed with the use of a tungsten pin. This can result in tungsten residuals that are known to interact with proteins. Another issue is that syringes typically use silicone oil as a lubricant for easier plunger movement. Silicone oil can migrate into the drug product and silicone oil droplets can act as a nucleus for particle formation/growth and protein aggregation.

	Newer packaging components are now being introduced to the industry; for example, engineered polymers are replacing glass. These polymers, such as cyclic polyolefins, are much lower in extractables and have lower surface tension characteristics that make them suitable for biologic drug products.

Why is it important to fully characterize contact materials and understand the material of construction for the container closure system and their associated E&L?

 Fully characterizing contact materials is crucial to ensure the materials chosen do not negatively affect the safety or efficacy of the drug as a result of leachables. Ideally, multiple options for container closure systems should be evaluated during the initial extractables screening. Then the packaging with the lowest risk can be selected. Establishing an extractable compound profile helps to ensure that the observed compounds are not overlooked during subsequent leachables evaluations. The constituents of large-molecule drug products often interfere with the analytical tests used to evaluate E&Ls. By establishing a material’s extractable profile, leachable analysis by mass spectrometry, using extracted ion analysis, can specifically target those compounds to evaluate their presence in the drug product. This technique effectively eliminates any matrix interferences and ensures leachables are not overlooked.

 What assessments should be performed to evaluate the potential risks of E&Ls from primary packaging that meets the biologic drug product?

 It is important to take a risk-based approach to choosing and evaluating the packaging components. It should start with supplier information on the components or system, addressing questions such as:

		What are the basic material characteristics of the components?

		Are there special needs associated with the biologic drug product application, such as the environmental conditions of storage?

	Once this information is gathered, basic evaluation by standard compendial methods is needed for compliance and allows one to begin to ‘qualify’ a component for use. But this is only the first step in proving suitability. Once compendia requirements are passed, material characterization is essential to better understand what may be extracted from the material (at levels critical to the drug product).

	The following highlights the best practice recommendation for addressing E&L for a primary package:

		Material characterization: Each individual component should be assessed to assure it has broad applicability for the application.

		Controlled extractables study: This study is a comprehensive program to understand what could be extracted from the components under a broader series of solvents, if the material characterization information is found not to be sufficient. It is crucial to perform a risk assessment to decide if this step is needed, and to determine the appropriate next step in the process based on the application. The solvents used should be aqueous-based, with considerations for organic solvents (if needed), pH, extraction conditions (such as time), extraction methods, material-to-solvent extraction ratios, etc.

		Simulation study: Depending on the drug product application, it may be appropriate to complete a simulation study, instead of going directly into a leachables study. This study is highly probable when it is especially challenging to reach the analytical evaluation threshold (AET). This may occur in a circumstance such as when evaluating a large-volume parenteral (LVP) application where there is a significant volume of drug solution. If many extractables are found from the controlled extractables study, the simulation study is a way to help identify the probable leachables to target in a formal leachables study.

		Data assessment: To determine the targets for a leachables study, it is important to evaluate the risk in the specific drug application.

		Leachables study: Method development and validation for specific leachables in the drug product should occur. Leachables testing should be conducted over drug product shelf life, at both room temperature, and accelerated conditions. The leachables should be identified based on the safety concern threshold (SCT). The SCT is the threshold dose below which a leachable would present negligible safety concerns for carcinogenic and noncarcinogenic effects. The recommended SCT for parenteral drug products, per the Product Quality Research Institute (PQRI) Extractables & Leachables Working Group for parenteral and ophthalmic drug product (PODP), is 1.5 ug/day (as described in an April 2018 workshop). 

		Special considerations for biologics include: biologic activity, efficacy, degradation, oxidation, chemical modification, immune adjuvant activity. 

 Typically, the first step is to perform an extractables profiling study on the packaging component of interest to identify the potential list of leachables in the drug formulation. The profiling study results may be used to perform a risk assessment with two goals: 

		To identify potential ‘bad actors’ from the list of extractables through predicting toxicity or performing toxicology experiments 

		To select components that have more desirable extractables profiles for the final process and eliminate components found to likely contribute to an undesirable leachable.

 How do you identify and quantify potential E&L from container closure systems?

 The first step is to expose the components of the container closure system to several model extraction solvents at exaggerated conditions of time and/or temperature. The resulting solutions are then screened by headspace and direct injection gas chromatography–mass spectrometry (GC/MS) for volatile and semi-volatile organic compounds, liquid chromatography–mass spectrometry time of flight (LC/MS–TOF) for non-volatile organic compounds, and inductively coupled plasma-mass spectrometry (ICP/MS) for elemental impurities. Additional testing methods may be used if appropriate, such as those specific for halide ions, nitrosamines, or PAHs. At Eurofins, we use the Wiley/National Institute of Standards and Technology (NIST) databases to identify compounds detected by GC/MS. For those compounds detected by LC/MS, we have a propriety database, the Eurofins Extractables Index, containing more than 1500 non-volatile organic compounds. If a compound cannot be identified via the database, additional testing may be necessary. Not only does this additional testing require advanced instrumentation (e.g., quadrupole time of flight [Q-ToF]), but it also requires the expertise of experienced and highly educated analysts.â©

 Establishing a holistic extractables profile for an article of interest is a complex and intensive process involving the use of a variety of analytical technologies. Gravimetric studies and total organic and inorganic carbon analysis are often performed to gain an understanding of the total extractable content. Fingerprinting of extracts using spectroscopic methods such as ultraviolet–visible spectroscopy (UV–VIS) and Fourier transform infrared spectroscopy (FTIR) resulting in generic information about constituent chemical classes is also common. 

	These methodologies are typically followed by more specific qualitative studies to identify volatile, semi-volatile, and non-volatile extractables using GC/MS and LC/MS techniques respectively (including high resolution accurate mass [HRAM] determination). Compounds are usually identified above the (AET) that has been determined for the material or article of interest. The AET for an article of interest depends on the target dose and number of doses expected to be stored in the container closure system or component. Analytically, the AET is used to estimate a detector response threshold using a set of reference standards carefully selected to represent the chemicals expected to be extracted. Once a list of extractable peaks above AET is identified, relative quantitation is also performed to better inform risk assessment.

	In parallel, it is also important to identify any elemental impurities that result from the extractions. This [assessment] is usually performed through either inductively coupled plasma optical emission spectrometry (ICP–OES) and inductively coupled plasma mass spectrometry (ICP–MS) methods depending on required specificity and sensitivity.

 Which analytical techniques are robust enough to identify potential E&Ls?

There is no one method that will identify all potential E&Ls. Multiple analytical techniques are needed for comprehensive assessment of extractables and leachables. For inorganic species, ICP/MS or OES are typically employed. GC/MS and LC/MS are the most common techniques for detection and identification of organic compounds. There are various LC/MS configurations for robust non-volatile organic analysis. Various additional features/techniques can improve sensitivity. One such example is ion mobility and Q-ToF to enable more precise analyses and identification of unknowns by combining ion mobility and mass-to-charge ratio.

Articles

To ensure the safety and efficacy of biologic drug products, the materials that come in contact with the drug must be fully characterized to determine their impact.

Assessing the E&L Risk in Biologic Drugs

When FDA’s process analytical technology (PAT) initiative was first unveiled, the aim was to have the industry move away from a mindset of “testing quality” at the end of production to instead develop methods that monitor, assess, predict, and control quality from the start of the manufacturing process. PAT tools have facilitated ongoing monitoring and adjustment of bioprocesses to ensure that biologic drug products are produced with high quality, meeting the required specifications all the time. 

spoke to Jim Mills, senior vice-president of Technical Operations at Abzena; Karl Rix, vice-president, Business Unit Bioprocess, Eppendorf; Tommy Smith, lead scientist, Cell Culture, GE Healthcare Life Sciences; Richard Moseley, chief technologist, Microsaic Systems; and Christian Grimm, R&D director, Process Analytical Technologies, Sartorius Stedim Biotech, about the advances in PAT tools for bioprocessing. 

In your opinion, what are the three most significant advances in analytical tools for monitoring processes in biopharmaceutical manufacturing? 

The three significant advances are on-line biomass measurement, at-line substrate and catabolite measurement, and the recent promise of at-line product measurement.

In my opinion, the important advances are in peripherals, such as sensors, pumps, valves, and motors, being designed with additional feedback and intelligence, allowing more precise monitoring of the entire process. This goes hand in hand with advances in bioprocess software that allow customized use of the gathered data, for example, in tailored feedback control loops. 

	Significant advances are also caused by the application of multivariate data analysis tools to interpret bioprocess data, develop a thorough process understanding, and use this knowledge for process design and ideally in-process corrective actions during manufacturing. 

	Finally, I consider scale-down models as important analytical tools; although they are not directly used during manufacturing, they are still of high relevance for it. I think efforts in the creation of scalable bioprocess systems have advanced and will further advance the possibilities for bioprocess analysis, both in development and for troubleshooting.  

The three most significant advances in analytical tools for monitoring upstream processes in biopharmaceutical manufacturing are near-infrared (NIR) spectroscopy, Raman, and radio-frequency (RF) impedance. In upstream processing, NIR and Raman are useful for real-time monitoring of cell culture parameters including pH, dissolved oxygen, and various metabolites. After calibration for a given process and media, these two tools provide constant information about the culture without the need for manual sampling. The same is true of RF impedance in relation to cell population density within a culture. 

	These tools are important for keeping the entire cell culture process under close surveillance. An additional advantage to using these three in-process analytical instruments is a reduced risk for contamination due to manual intervention and off-line analyses.

The analytical tools commonly used to monitor upstream and downstream bioprocessing have been traditional, simple instruments. For upstream process monitoring, these include pH monitors, glucose measurements, and gas pressure gauges. For downstream purification, the tools are slightly more sophisticated, such as enzyme-linked immunosorbent assay (ELISA) and high-performance liquid chromatography with ultraviolet detection (HPLC–UV). However, these tools are not suitable for the current challenges faced by the biopharmaceutical industry, particularly those driven by the stricter regulations surrounding biologics, or biopharmaceuticals. Efficient quality-by-design (QbD) strategies are now needed, giving bioprocessing operators and managers access to more detailed product and process information at the point-of-need, in other words, where the reaction and process is taking place. 

	The first significant advancement in analytical tools is the development of better multivariate data analysis (MVA). The biopharmaceutical industry, previously familiar in making simpler small-molecule drugs, has found that MVA is suitable for the challenge of safely controlling a bioprocess. MVA is used to link the critical quality attributes (CQAs) of the target product (as an example, glycosolyation is a CQA for monoclonal antibody safety and potency) to a set of critical process parameters (CPPs). CPPs can include temperature, pressures, and flow rates. 

	Adopted industry-wide as the preferred choice of analysis, mass spectrometry (MS) can provide the information needed to monitor and control the processing of biologics using MVA, as it measures CQAs and CCPs directly. Until recently, MS instruments were large, power hungry, and capital-intensive equipment to purchase and maintain. These instruments still form much of the centralized-laboratory infrastructure, where samples from the bioprocessing workflow are analyzed. However, turning a sample analysis around can often take days or even weeks. A significant advancement is the emergence of compact, miniaturized MS instruments containing powerful analytical software tools, more suited to point-of-need applications. Such compact instruments may be deployed at almost any step in the bioprocess (at the ‘point-of-need’) and can be used to analyze the contents of the cell media, or buffer solution. As a result, analysis times become instantaneous, speeding up time for manufacture and also the time to market for new biologic drug candidates.

From my perspective, the most significant developments of analytical tools for use in biopharmaceutical processes are the full integration of ready-to-use single-use inline sensors such as in-line capacitance for biomass monitoring in bioreactor bags, the use of optical spectroscopic equipment and methods for inline integration into bioprocess solutions (e.g., NIR, Raman, UV–VIS or fluorescence), and the use of data analytics such as multivariate statistical and other chemometric methods to transfer data into knowledge.

How widespread, would you say, is the use of PAT tools in biopharmaceutical manufacturing? 

Obviously, the implementation of PAT tools is not as widespread in biopharmaceutical production as in chemical or pharmaceutical manufacturing. In biopharmaceutical processes, we have to distinguish between the status within upstream and downstream processes. Interestingly, the implementation of relatively new tools such as spectroscopy is more mature in upstream than in downstream, even if the matrix under test is far more complex in a cell culture for example than in a later polishing step. Compared to classical pharmaceutical processes, however, we are far behind in the implementation of PAT tools. This may change significantly in the coming period especially with the increased use of continuous and intensified manufacturing approaches within the industry.

Simple, low cost, and robust sensors have been used for PAT in the chemical industry for many years, for example, Fourier-transform infrared (FTIR) spectroscopy is used to monitor and control small-molecule API synthesis. However, the data produced from these types of simple sensors has very limited value for MVA in a bioprocessing context. Instruments suited to analyzing complex biologics such as MS have been proposed and explored in the past by biopharmaceutical companies, but the lack of deployability of the large MS instruments, and the limited software analysis that were available at the time did not lend themselves well to being PAT tools. The advent of miniaturized mass spectrometers has presented an important step forward for the adoption of point-of-need MS by biopharmaceutical manufacturers. 

I think the majority of the biopharmaceutical companies are interested in implementing PAT tools in manufacturing even if they are not using them now. I expect it will take a couple of years more until PAT is widespread in bioprocessing. Furthermore, PAT tools will get increasingly important at earlier development stages, I believe, because gaining sufficient process and product knowledge during the research and development phase helps implementing PAT at later stages. Regulatory agencies encourage PAT implementation, and we might see even more incentives from their side in the future.  

I agree that the concept of PAT is not new in pharmaceutical manufacturing. However, the ability to consistently and robustly measure, monitor, and control such complex processes as biologics manufacturing are still largely outside of the state-of-the art. It has much to do with the difficulty in directly measuring macromolecular characteristics in-process. Many developmental PAT tools have been proposed using indirect measurement methods relying on complex statistical correlations between traditional physical on-line measurements and the parameter being inferred. Because of the difficulty in direct measurement, trust of such inferred measurements has been hard to gain. In a highly regulated industry such as the biopharmaceutical sector, which also has high costs and long cycle times for product development, these challenges have acted as a significant barrier to PAT adoption for most companies, especially small and medium enterprises (SMEs) and contract manufacturing organizations (CMOs).

	I think PAT tools are beginning to gain a foot-hold in bioprocess development to save time and cost during the early transition from research into development. One aspect that has helped with this is more vendors, such as Sartorius, providing already integrated solution for PAT in their product offerings (e.g., single-use biomass measurement), which makes adoption simpler for companies. However, even though PAT is starting to be used during process development, generally for GMP manufacturing, traditional quality control testing is still relied upon.

What positive changes have PAT tools contributed to in biopharmaceutical manufacturing over the past 10 years?

In upstream processes, periodic sampling (daily or twice daily) is the traditional norm. However, with the advent of PAT, constant pH, dissolved oxygen, and metabolite monitoring are now possible. Such monitoring allows for proactive management of the system instead of a reactive response. Now the operator can have an insight into what the status of the system is at any time as well as an opportunity to add feeds, bases, and acids when needed instead of at or after the traditional sampling time during the day. The gain in this instance is a more consistent quality of product due to the ability to keep parameters that affect quality attributes steady or within desired ranges over the course of a production run.

	The increased use of PAT has led to a reduction of contamination incidences in manufacturing due to the ability to monitor processes with minimal manual intervention or sampling. In upstream work, this includes non-invasive pH and dissolved oxygen monitoring through optical devices that do not need to be inserted into the bioreactor post steam-in-place or irradiation. Another example is the monitoring of cell density via an RF impedance probe that is inserted into the system before steam, autoclave, or irradiation. Overall, this monitoring inevitably results in a cost savings to the company during the life of the production cycle. Tighter controls over the process are also a result of PAT tools. 

Working with complex biological systems producing the target molecule or drug substance makes the implementation of PAT tools in biopharmaceutical production processes far more difficult than in chemical synthesis. However, the recent implementation of several PAT tools in terms of design of experiment (DoE), inline sensors, spectroscopy probes, and the use of multivariate statistical methods have increased fundamental process-knowledge and given insight into the mechanisms of CPPs on the CQAs. This enabled more reliable process automation and was a major step forward in the direction of QbD implementation in bioprocesses.

PAT is a system for designing, analyzing, and controlling manufacturing through timely measurements of critical quality and performance attributes, with the goal of ensuring final product quality. I think this exactly describes the most important positive changes PAT tools contribute to. They allow achieving better process understanding and therefore predictability. Associated with process control, they can help to minimize process variations. As a result, a more consistent product quality can be achieved. I assume PAT tools also help to save costs, as they can help to reduce the number of failed batches.

There is broad recognition that PAT tools are essential for helping to simplify manufacturing, and this work has already led to the adoption of relatively simple instruments being integrated into process steps where PAT is relatively straightforward. However, the complexity of bioprocessing means important decision points in the process are not all routinely covered by PAT. Important improvements to MS hardware and software means the remaining difficult bioprocessing steps can be monitored and controlled using miniaturized MS-based PAT systems. 

What challenges still exist in PAT implementation for bioprocessing?

One important challenge I see is transferability. PAT already plays a key role during research and development, where it helps in increasing process knowledge and developing a process control strategy, but it usually happens at small scales. To be applied in manufacturing, PAT tools and control strategies need to be validated and transferred across scales and facilities. 

	Secondly, PAT implementation requires specialized expertise of how to integrate sensors, how to validate models, and how to program feedback control loops. It will often require the combined efforts of vendors and users to tackle this challenge.

The understanding of PAT and its relevance to complex bioprocessing has improved enormously. There are, however, many challenges that need to be tackled before there can be full implementation of PAT tools into the whole of bioprocessing. For the implementation of a miniaturized MS PAT tool, the key challenges are: 

		To integrate the mass spectrometer in environments normally hostile to the instrument without damaging the instrument or compromising the safety of the product 

		Acquire reliable data over an acceptable time 

		Provide accurate and useful information to the end-user. 

I think the main challenge is still the implementation of robust, reliable direct measurement tools that can be validated and applied in the GMP manufacturing environment.

The remaining hurdles for PAT implementation for bioprocessing could be split into three categories. There are, of course, residual technical and application challenges due to the fact that we have to monitor and control complex biological systems, whose responses to changes of macroscopic environmental factors cannot be predicted completely. In addition, the number and complexity of important parameters are quite high, hence, requiring analytical methods that are selective and sensitive enough to measure these parameters in an online way. Furthermore, the current trend in the industry to use single-use equipment-not only in process development but going further to manufacturing scale-requires an implementation of PAT tools in single-use systems as well, which is a technical challenge per se. 

	Secondly, the implementation of PAT in bioprocess requires mindset changes of the acting people. There is still, in some cases, the perception of the operators, that they are faced with a black box batch process, which they better not touch if they were able to run it in a semi-optimal way. If our industry is moving forward to continuous or intensified processes, there is a clear demand for implementing PAT tools so that the processes can be run in a stable and robust regime. 

	The third important aspect for the hesitation of PAT implementation in bioprocessing is the lack of a strong economical driving force. We still have legacy production processes running in the industry that are far away from optimal in terms of efficiency and quality. The recent increase of more biosimilars coming to market, higher consolidation in the industry, and changes in local markets will give the demand of more cost competitive manufacturing. These drivers will lead to a higher degree of implementation of PAT in the near future.

): The vast number of potential analytical instruments that could be integrated into a biopharmaceutical process presents a logistical challenge. A single unit that implements NIR or Raman for metabolite monitoring is typically not an issue. However, when instrumentation such as HPLC/MS, metabolite/cell density analyzers, and sample purification modules are all individually attached to a sterile sampling mechanism, the footprint of the equipment and distance of travel of the sample becomes a burden.

What advances in analytical tools for biopharmaceutical manufacturing would you like to see within the next five years?

We believe that any advances should be aimed at bringing cheaper and safer biologics to all the people who require them. For analytics, this means the tools must: 

		Add important value to bioprocessing by maximizing product yields, ensure product safety, and minimize process downtime

		Be reliable and robust to suit the point-of-need applications needed for PAT. 

	In the next five years, we would like to see miniaturized mass spectrometers routinely integrated into upstream and downstream bioprocessing environments, where they can both monitor and control the production of biologics, through their unique ability to detect and analyse proteins, and small molecules.

Next PAT advances should be on further implementation of inline sensors for monitoring and controlling the CPPs, including more multi attribute methods. For example, HPLC or MS could be used to get online access to CQAs and mechanistic or hybrid process models could improve advanced process control. In addition, the industry has concentrated mainly on implementation of PAT tools in the upstream area, but we would likely see more development in the downstream part to set the basis for the vision of real-time release.

It would be optimal to see more all-in-one modular offerings allowing the targeted industry to implement a single instrument for all, or most, desired analysis parameters.

In my opinion, biopharmaceutical manufacturing would benefit from additional sensors for online monitoring of metabolites and CQAs of the product. These parameters of course can be analyzed offline and online-integration of some sensor types is feasible; however, I hope to see, for example, metabolite sensors in the future, which can be as easily used as today’s pH and dissolved oxygen sensors. Often single-use technology is applied in manufacturing; it would be also desirable for new sensor technology to simplify handling and avoid sterility issues.

I’d like to see actual real-time measurement of product concentration and product quality. 


	Vol. 31, Number 7
	July 2018
	Pages: 39–43

	When referring to this article, please cite as A. Siew, “Advances in Analytics for Bioprocessing,” 

Advances in process analytical technology tools are needed to address the challenges of process monitoring and quality control.