Process analytical technology for biopharmaceuticals
13. jul. 2026
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Time and resources are incredibly valuable in research and manufacturing environments. Slow, inefficient manual workflows lead to increased process variability, higher costs, and even product loss. Many professionals are turning to directly integrated Process Analytical Technologies (PAT) to overcome these challenges. PAT is a system that integrates real-time data acquisition and control into manufacturing processes.
Such technology plays a crucial role in ensuring product quality, optimizing process efficiency, and meeting stringent regulatory requirements. Integrating PAT enhances visibility into key parameters and enables real-time process control. This article defines PAT and explains how it is implemented, with specific application examples showing where PAT is used in the biopharmaceutical industry.
This article covers the following topics:
What is PAT?
PAT is a framework introduced by the U.S. Food and Drug Administration (FDA) to enhance process understanding and control by integrating various analytical techniques into production environments [1]. It is a science- and risk-based approach for the design, monitoring, and control of manufacturing processes through real-time or near real-time measurement of critical quality attributes (CQAs) [2,3] and process parameters. PAT is segmented into three categories: design, analysis, and control. This approach ensures consistent product quality and accurate measurement of CQAs [4].
Metrohm Process Analytics provides a variety of solutions that simplify the integration of PAT into operations. This facilitates accurate, nondestructive, real-time analytical results directly from the process.
Learn more about the necessity of using PAT and how we can support your operations by reading our multipart blog series.
Benefits of PAT in biopharmaceuticals
Biopharmaceuticals provide transformative healthcare therapies, yet their complexity poses significant challenges to manufacturing consistency and product quality. These complexities demand in-depth understanding and precise control of production processes, which are increasingly difficult to achieve with conventional approaches. Recognizing these challenges, regulatory bodies like the FDA are emphasizing the advancement of manufacturing sciences, with PAT emerging as a cornerstone of this evolution.
The pharmaceutical sector was among the first to adopt PAT principles. Progress in the biopharmaceutical sector has been gradual, but the potential benefits are significant. PAT gives the biopharma industry a great reason to investigate different analytical tools to develop more robust and efficient manufacturing processes. This approach improves process insight and enables real-time production control, rather than relying only on final product testing [3].
PAT represents a shift from empirical, end-point testing to a science- and risk-based approach centered on real-time process monitoring, control, and optimization. This shift transforms biomanufacturing by forcing the industry to adopt predictive, data-rich process strategies that enhance product quality, reduce variability, and support Quality by Design (QbD) principles. As a result, organizations can now design robust, scalable, and economically viable processes from early development phases—ultimately improving time-to-market while meeting stringent regulatory and quality expectations.
One way to improve production efficiency and product quality is by using specific monitoring methods that provide real-time or near real-time measurements (inline, online, atline; Figure 1). Continuous monitoring of critical and other relevant process parameters is accomplished with suitable process analysis equipment. For instance, process analyzers can continuously monitor important chemical and physical events during bioprocessing. Such real-time information provides manufacturers deeper process insight, building process knowledge [3].
By combining process analysis technologies such as chemometrics (spectrometers), process analyzers, end-point detection methods, and knowledge management systems, manufacturers can implement dynamic and responsive control over their production [3].
Still curious? Read our article about how to choose the right PAT solution for the most accurate and reliable data.
Building a Smart(er) Factory by implementing Process Analytics 4.0 solutions
Key PAT methods for biopharmaceuticals
Real-time monitoring methods are crucial for evaluating cell growth, substrate usage, and product formation. They enable precise bioprocess control and improve yield, productivity, and consistency. It is therefore vital to implement reliable analytical techniques that can monitor key process parameters online [6].
When considering bioprocess monitoring, three primary categories of parameters are typically assessed:
- physical variables (e.g., pressure, temperature, viscosity, stirrer speed)
- chemical variables (e.g., pH, dissolved oxygen (DO), dissolved carbon dioxide (pCO2), nutrients such as glucose, metabolites such as lactate)
- biological variables (e.g., biomass concentration, cell metabolism)
Different analytical techniques are available for each of these variables, which are each designed for specific applications. Some of these techniques are discussed below, including near-infrared spectroscopy, Raman spectroscopy, and chemical sensors.
Regardless of the analysis technique, sensor probes designed for process monitoring in biopharmaceutical industrial facilities must meet specific requirements to operate effectively within bioreactor systems. These probes are typically immersed directly in the bioreactor medium and must be capable of withstanding demanding conditions along with fluctuations in temperature and pressure. For a sensor probe to be effective in an industrial setting, it should provide high selectivity, sensitivity, robustness, repeatability, stability, low detection limits, linearity, short response times, and a long operational lifetime [6]. Implementing analytical techniques online or inline that meet these criteria remains a significant challenge.
Near-infrared spectroscopy (NIRS)
Near-infrared spectroscopy (NIRS) is widely used in PAT for nondestructive, rapid analysis of raw materials, intermediates, and final products. Its key advantages include minimal sample preparation and robust quantification capabilities.
NIR spectroscopy is based on the absorption of near-infrared light by molecules in a sample. The amount of light absorbed provides critical information about the chemical composition and quality. As the near-infrared light interacts with the sample, it is absorbed by chemical bonds. The absorbed light results in specific absorption bands that are characteristic of the chemical components present in the sample. These absorption bands are analyzed to determine the concentration of various components, such as moisture, proteins, sugars, and lipids.
Want to learn more about NIR spectroscopy? Start with our blog series on this topic.
Raman spectroscopy
Raman spectroscopy is a nondestructive, reagentless analytical technique that provides valuable chemical insights about a sample matrix. It works by measuring the interaction between light (photons) and molecules. When photons from an incoming laser interact with a sample, scattering occurs. Some light is scattered elastically (Rayleigh scattering), while a small portion is scattered inelastically (Raman scattering). The resulting Raman shift contains chemical information which is visualized as spectra and analyzed by peak intensity, peak height, or with chemometric models.
Read more about the theory behind Raman spectroscopy and its applications in our blog series.
Frequently Asked Questions (FAQ) about Raman spectroscopy: Theory and usage
A common challenge in biological sample analysis is fluorescence – a disruptive effect that complicates measurements. However, in some cases fluorescence can correlate with biomass, metabolic state, or cell viability, depending on the application. Additionally, there are effective, patented methods (both hardware- and software-based) that minimize the impact of fluorescence through data preprocessing techniques [7]. This allows for accurate measurement of other parameters.
Strict hygiene and sterile working environments are crucial for preventing the growth of unwanted microorganisms. Fiber optics and specially designed Raman probes are the ideal solution to support this. These components play a key role by guiding light to the sample and returning it to the detector. They ensure precise and efficient light transmission—vital for accurate process analysis. The probe acts as an «eye» peering into the process, performing measurements directly within the production environment (Figure 2). Optical probes can be integrated into the process in several ways: inline or online. Their versatility and adaptability are essential for obtaining reproducible and accurate real-time measurements.
These probes are also designed to withstand harsh sample conditions, such as pressure, pH, and temperature variations. Inline probes, when used with Raman optical spectrometers, provide optimal results in terms of laser intensity and the reproducibility of concentration values.
Sometimes it is necessary to avoid using inline probes that come into direct contact with the sample. This is particularly true when the process analyzer needs to be moved or switched between different measuring ports like in lab or pilot plant settings. Another consideration is the time- and resource-intensive process of validating a new PAT sampling point, which many companies cannot afford.
For such cases, a cost-efficient solution is the use of a typical ViewPort™ Ingold connection or PG13.5 connection for bioreactors, ranging from lab-scale (benchtop) to industrial-scale applications. Raman probe shafts are optimized to maximize the Raman signal, while the probe itself remains separate from the sample. Measurements are taken through a window in the ViewPort™. This offers a non-invasive approach to real-time bioprocess monitoring under sterile conditions, without the need to revalidate a PAT measurement point.
Chemical sensors
Many biopharmaceuticals rely on fermentation, where balancing the optimal pH, oxygen, and carbon dioxide levels is crucial. Microorganisms thrive within well-defined ranges for these parameters, which makes precise measurements essential for maintaining their health.
High quality sensors that relay critical information about the fermentation process in a timely manner are crucial. ProTrode sensors from Metrohm Process Analytics fulfill the demanding requirements of biopharmaceutical manufacturing. These sensors are designed to withstand autoclaving and sterilization cycles without issue and are offered in different lengths and with a standard PG 13.5 thread to fit common bioreactors.
The ProTrodes 200 and 300 are ideal for PAT pH measurements. These state-of-the-art sensors deliver reliable, accurate data throughout the entire batch production process.
For dissolved oxygen measurements, the ProTrode DO sensor from Metrohm Process Analytics impresses with its fast response time and unique design that extends the service life of the sensor. The measuring principle of this DO sensor is based on luminescence quenching, and it delivers reliable measurements from 0–250% air saturation (0–50% O2).
For a more detailed introduction to dissolved oxygen measurement (including practical considerations), read our blog article «Dissolved oxygen measurement – easier than ever».
The DO sensor consists of a measuring part (including electronics) and a sensor shaft with a replaceable membrane. The only parts of the sensor in contact with the fermenting solution are the shaft and the membrane. This makes it possible to limit autoclaving and sterilization to these parts, leaving the sensor part with the electronics unaffected.
The ProTrode pCO2 sensor for dissolved carbon dioxide (pCO2) from Metrohm Process Analytics is based on the Severinghaus principle. Its measuring range (10–800 mbar pCO2) is perfectly suited for the requirements of this application.
Where can PAT be applied?
PAT can be applied across various stages of biopharmaceutical production, from early-stage development to full-scale manufacturing. Its primary value lies in providing insights that help optimize critical process steps and ensure product quality. Key application areas include fermentation and cell cultures.
Fermentation
Fermentation plays a vital role in producing a wide array of biotechnological products, ranging from pharmaceuticals to biofuels and food ingredients. The use of biotechnological processes in pharmaceutical production has grown significantly, with more than 59% of newly approved drugs in Europe now being manufactured through biotechnological methods [8].
Fermentation-based biotechnological processes offer sustainable and resource-efficient alternatives to traditional manufacturing. The production of complex bio-based chemicals (e.g., biopolymers, bio-surfactants, proteins, and enzymes through microbial fermentation) reduces reliance on finite resources and minimizes environmental impact through fewer raw materials used. These processes both lead to the development of new products and applications as well as foster a more sustainable world.
Monitoring fermentation processes inline is critical to optimize production and ensure consistent product quality. Traditional analysis methods often involve long sample preparation and delayed results, failing to provide the real-time feedback necessary to adapt quickly to process changes. Inline analysis using advanced techniques, such as NIRS, enables continuous, reagent-free monitoring of key parameters in real time. This allows for quicker adjustments to variables like enzyme and microbial blends, temperature, and fermentation conditions.
The 2060 NIR Process Analyzer is a cutting-edge tool for this application. It allows for the continuous, reagent-free monitoring of key parameters such as ethanol, glucose, maltose, and organic acids directly within the fermentation bioreactor. This provides operators with real-time insights that improve process efficiency, reduce fermentation time, and ultimately increase production throughput and profitability.
Read our Application Note to learn more.
Cell cultures
In biopharmaceutical production, particularly in cell culture processes, maintaining optimal cell health is crucial for successful yields and product quality. The accumulation of lactate as a byproduct of glycolysis is a major challenge in cell culture, as this can induce acidity that causes cellular stress and inhibits growth. Traditionally, glucose and lactate levels are monitored using manual offline laboratory methods. However, these methods are prone to contamination and can fail to provide timely, accurate data.
To address these issues, inline Raman spectroscopy offers a solution for real-time monitoring directly in the bioreactor. The Metrohm 2060 Raman Process Analyzer offers continuous, non-contact analysis of glucose and lactate concentrations, providing immediate feedback for process adjustments. This reagent-free, nondestructive technique ensures that the sample remains unaltered, which reduces the risk of contamination and improves process efficiency.
By using Raman spectroscopy, operators can monitor the state of the cell culture in real time. They can track glucose consumption and lactate accumulation and make timely adjustments to optimize conditions for cell growth. Paired with Metrohm's Vision and IMPACT software, the 2060 Raman Process Analyzer enables the development of a calibration model to compare real-time data against reference methods such as HPLC or titration. This facilitates accurate, efficient process control and enhances the quality and consistency of biopharmaceutical products.
In addition to its non-contact nature, the 2060 Raman Process Analyzer can measure multiple parameters simultaneously. This provides a comprehensive view of the cell culture environment, facilitating early detection of potential batch failures, minimized contamination risks, and improved overall process throughput. By integrating this advanced inline monitoring solution, manufacturers can significantly enhance the control and efficiency of their cell culture processes. This ultimately ensures higher-quality biopharmaceutical products.
Learn more about this application here.
Conclusion
Process Analytical Technology improves manufacturing by enabling real-time monitoring and control of critical process parameters. In biopharmaceutical production, PAT enhances product quality, supports regulatory compliance, and reduces operational costs. By integrating PAT tools such as Raman spectroscopy, near-infrared spectroscopy, and various chemical sensors into production environments, manufacturers gain precise control over critical processes. This enables faster decision-making, minimizes batch failures, and reduces the need for manual intervention or additional costly end-product testing.
References
[1] Guidance for Industry PAT — A Framework for Innovative Pharmaceutical Development, Manufacturing, and Quality Assurance U.S. Department of Health and Human Services Food and Drug Administration Center for Drug Evaluation and Research (CDER) Center for Veterinary Medicine (CVM) Office of Regulatory Affairs (ORA) Pharmaceutical CGMPs September 2004; U.S. Department of Health and Human Services, 2004.
[2] Process Analytical Technology - an overview | ScienceDirect Topics. https://www.sciencedirect.com/topics/chemistry/process-analytical-technology (accessed 2025-04-01).
[3] Undey, C.; Low, D.; Menezes, J. C.; et al. PAT Applied in Biopharmaceutical Process Development And Manufacturing: An Enabling Tool for Quality-by-Design; CRC Press, 2011.
[4] Read, E. K.; Park, J. T.; Shah, R. B.; et al. Process Analytical Technology (PAT) for Biopharmaceutical Products: Part I. Concepts and Applications. Biotech & Bioengineering 2010, 105 (2), 276–284. DOI:10.1002/bit.22528
[5] Owczarek, D. Bioprocessing 4.0 and the Benefits of Introducing AI to Biopharmaceutical Manufacturing. Nexocode. https://nexocode.com/blog/posts/bioprocessing-4-ai-in-biopharmaceutical-manufacturing/ (accessed 2025-04-02).
[6] Veloso, A. C.; Ferreira, E. C. Online Analysis for Industrial Bioprocesses. In Current Developments in Biotechnology and Bioengineering; Elsevier, 2017; pp 679–704. DOI:10.1016/B978-0-444-63663-8.00023-9
[7] Gelwicks, M. J.; Zemtsop, C.; Allen, M. W. Fluorescence Rejection in Raman Spectroscopy: An Algorithmic Approach and Applications. In Photonic Instrumentation Engineering XI; SPIE, 2024; Vol. 12893, pp 84–92. DOI:10.1117/12.3000516
[8] Biopharmazeutika: Marktanteil wächst | vfa. https://www.vfa.de/de/forschung-entwicklung/medizinische-biotechnologie/biotech-deutschland/medizinische-biotechnologie-in-deutschland-2023 (accessed 2025-03-27).
Additional resources
Brochure: Pharmaceutical analysis – Quality control of pharmaceuticals
Flyer: ProTrode 200/300 sensor pharma
Application Note: Inline moisture analysis in fluid bed dryers by near-infrared spectroscopy
Application Note: Inline monitoring of cell cultures with Raman spectroscopy
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