At-line Measurements Research Articles

A portable dry film FTIR instrument for industrial food and bioprocess applications.

The main objective of this study was to design, build, and test a compact, multi-well, portable dry film FTIR system for industrial food and bioprocess applications. The system features dry film sampling on a circular rotating disc comprising 31 wells, a design that was chosen to simplify potential automation and robotic sample handling at a later stage. Calibration models for average molecular weight (AMW, 200 samples) and collagen content (68 samples) were developed from the measurements of industrially produced protein hydrolysate samples in a controlled laboratory environment. Similarly, calibration models for the prediction of lactate content in samples from cultivation media (59 samples) were also developed. The portable dry film FTIR system showed reliable model characteristics which were benchmarked with a benchtop FTIR system. Subsequently, the portable dry film FTIR system was deployed in a bioprocessing plant, and protein hydrolysate samples were measured at-line in an industrial environment. This industrial testing involved building a calibration model for predicting AMW using 60 protein hydrolysate samples measured at-line using the portable dry film FTIR system and subsequent model validation using a test set of 26 samples. The industrial calibration in terms of coefficient of determination (R2 = 0.94), root mean square of cross-validation (RMSECV = 194 g mol-1), and root mean square of prediction (RMSEP = 162 g mol-1) demonstrated low prediction errors as compared to benchtop FTIR measurements, with no statistical difference between the calibration models of the two FTIR systems. This is to the authors' knowledge the first study for developing and employing a portable dry film FTIR system in the enzymatic protein hydrolysis industry for successful at-line measurements of protein hydrolysate samples. The study therefore suggests that the portable dry film FTIR instrument has huge potential for in/at-line applications in the food and bioprocessing industries.

Real Time AMC Monitoring with Novel Chemical Ionization Mass Spectrometry at Single-Digit pptv Concentrations

In this study, emissions of photoresist resin deposited on wafters in a FOUP are compared using impinger/GCMS at-line measurements and on-line Vocus AMC measurements based on chemical ionization mass spectrometry. While GCMS measurements are routinely used for VOC measurements, the Vocus AMC monitor allows simultaneous, real-time detection of acids, bases, and condensable organics, providing unique insights into the cleanroom and FOUP microenvironment Photoresist resin outgassing measurements show comparable absolute concentrations between the two methods (GCMS and CI-TOF) for targeted compounds in real world sampling scenarios. The rate of outgassing was tracked in real time with the Vocus and also allowed identification of unknown compounds emitted from the resin. Continuous monitoring of the cleanroom air around the FOUP highlights the importance of the cleanroom environment which can directly affect the FOUP microenvironment, and additionally we show that Entegris Barrier Material EBM can guarantee a good wafer protection.

Optical glucose sensor for microfluidic cell culture systems

Glucose is the primary energy source of human cells. Therefore, monitoring glucose inside microphysiological systems (MPS) provides valuable information on the viability and metabolic state of the cultured cells. However, continuous glucose monitoring inside MPS is challenging due to a lack of suitable miniaturized sensors. Here we present an enzymatic, optical glucose sensor element for measurement inside microfluidic systems. The miniaturized glucose sensor (Ø 1 mm) is fabricated together with a reference oxygen sensor onto biocompatible, pressure-sensitive adhesive tape for easy integration inside microfluidic systems. Furthermore, the proposed microfluidic system can be used as plug and play sensor system with existing MPS. It was characterized under cell culture conditions (37 °C and pH 7.4) for five days, exhibiting minor drift (3% day−1). The influence of further cell culture parameters like oxygen concentration, pH, flow rate, and sterilization methods was investigated. The plug-and-play system was used for at-line measurements of glucose levels in (static) cell culture and achieved good agreement with a commercially available glucose sensor. In conclusion, we developed an optical glucose sensor element that can be easily integrated in microfluidic systems and is able to perform stable glucose measurements under cell culture conditions.

Combining on‐line and at‐line measurements for monitoring of cell distributions in bioreactors

Chemie Ingenieur TechnikVolume 94, Issue 9 p. 1237-1237 Vortrag Combining on-line and at-line measurements for monitoring of cell distributions in bioreactors A. Schaum, A. SchaumSearch for more papers by this authorP. Jerono, P. JeronoSearch for more papers by this authorT. Meurer, T. MeurerSearch for more papers by this author A. Schaum, A. SchaumSearch for more papers by this authorP. Jerono, P. JeronoSearch for more papers by this authorT. Meurer, T. MeurerSearch for more papers by this author First published: 25 August 2022 https://doi.org/10.1002/cite.202255393AboutPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Share a linkShare onFacebookTwitterLinkedInRedditWechat No abstract is available for this article. Volume94, Issue9Special Issue: (Bio)Process Engineering – a Key to Sustainable Development: ProcessNet and DECHEMA-BioTechNet Jahrestagungen 2022 together with 13th ESBES SymposiumSeptember 2022Pages 1237-1237 RelatedInformation

Applicability of Raman and near-infrared spectroscopy in the monitoring of freeze-drying injectable ibuprofen

The freeze-drying process is an expensive, time-consuming and rather complex process. Therefore, process analytical technology (PAT) tools have been introduced to develop an optimized process and control critical process parameters, which affect the final product quality. The aim of the present work was to study the applicability of at-line near-infrared (NIR) and Raman spectroscopy approach in the monitoring of the freeze-drying process. Freeze-dried powders, which were developed previously, were manufactured as a multi-component system, containing ibuprofen (IBP). The NIR proved to be a useful tool for the monitoring of the freeze-drying process, since it was able to determine residual moisture content (RMC) and hence predict its values by using the partial least square (PLS) model. In addition, the evaluation of the correlation between the NIR and off-line HPLC IBP content results showed that NIR spectra were consistent with the HPLC measurements, even though overlapping absorption bands in multi-component system were observed. This research also studied the ability of using the at-line Raman measurements for the evaluation of the crystallinity and polymorphic transformations during the process, such as IBP ionization and mannitol polymorphism. The results were in correlation with XRPD results, but parameters of PLS models were not optimal. Nevertheless, this approach still assured better process understanding. To conclude, high applicability of the at-line NIR in the monitoring of the freeze-dried powder production was successfully demonstrated, suggesting that it can be used as a single tool to monitor RMC and IBP content as well as process deviations during the freeze-drying process.

Multiplexed Microfluidic Cartridge for At-Line Protein Monitoring in Mammalian Cell Culture Processes for Biopharmaceutical Production.

The biopharmaceutical market has been rapidly growing in recent years, creating a highly competitive arena where R&D is critical to strike a balance between clinical safety and profitability. Toward process optimization, the recent development and adoption of new process analytical technologies (PAT) highlight the dynamic complexity of mammalian/human cell culture processes, as well as the importance of fine-tuning and modeling key metabolites and proteins. In this context, simple, rapid, and cost-effective devices allowing routine at-line monitoring of specific proteins during process development and production are currently lacking. Here, we report the development of a versatile microfluidic protein analysis cartridge allowing the multiplexed bead-based immunodetection of specific proteins directly from complex mixtures with minimal hands-on time. Colorimetric quantification of Chinese hamster ovary (CHO) host cell proteins as key impurities, monoclonal antibodies as target biopharmaceuticals, and lactate dehydrogenase as a marker of cell viability was achieved with limits of detection in the 1–10 ng/mL range and analysis times as short as 30 min. The device was further demonstrated for the monitoring of a Rituximab-producing CHO cell bioreactor over the course of 8 days, providing comparable recoveries to standard enzyme-linked immunosorbent assay (ELISA) kits. The high sensitivity combined with robustness to matrix interference highlights the potential of the device to perform at-line measurements spanning from the bioreactor to the downstream processing.

Real-time monitoring and control for high-efficiency autonomous laser fabrication of silicon nanoparticle colloids

Nanotechnology is a significant research tool for biological and medical research with major advancements achieved from nanoparticle (Np) applications in biosensing and biotherapeutics. For laser ablation synthesis in solution (LASiS) to be chosen by researchers for Np colloid production, the process must effectively compete with chemical synthesis in terms of produced colloid quality and productivity while taking advantage of LASiS benefits in terms of its ‘green-synthesis’ and single-step functionalisation abilities. In this work, a newly developed integrated LASiS Np manufacturing system is presented including a Np flow reactor design, an at-line Np size monitoring via 180° dynamic light scattering, and a UV-Vis spectroscopy system used to estimate colloid concentration and stability. The experimental outcomes are discussed in terms of Np productivity and quality via these at-line measurements from the UV-Vis and DLS systems. The developed instrument was validated via off-line SiNps DLS, UV-Vis and morphology tests via TEM. Ultra-high quality and nanoparticle fabrication rate efficiency was achieved and is reported here.

Characterization of lysozyme PEGylation products using polarized excitation-emission matrix spectroscopy.

The growing use of therapeutic proteins requires accurate analytical techniques for measuring biophysical and structural changes during manufacturing. This is particularly true for the PEGylation of proteins, because characterization of PEGylation reactions and products can often be difficult due to the relatively small impact on protein structure, the lack of an accessible polyethylene glycol (PEG) chromophore, and the heterogeneous final product mixtures. Intrinsic fluorescence spectroscopy is one potential solution due to its relatively high sensitivity to small changes in protein structure and its suitability for online or atline measurements. In this study, we use the PEGylation of lysozyme as a model system to determine the efficacy of polarized excitation-emission matrix (pEEM) spectroscopy as a rapid tool for characterizing the structural variability of the lysozyme (LZ) starting materials and PEGylated products with varying PEG-to-protein ratios (PPR). Dynamic light scattering showed that as PPR increased from 0 to 2.8, the hydrodynamic radius increased from ∼2.2 to 4.8 nm. pEEM measurements provided several sources of information: Rayleigh scattering to identify size changes and aggregate/particle formation, and fluorescence emission to assess chemical and structural changes. PEGylation induced sufficient physicochemical changes in LZ, which produced changes in the pEEM spectra, largely due to variations in the hydrophobic environments of tryptophan residues close to a PEG attachment site. These significant spectral changes when modeled using conventional multivariate analysis methods were able to easily discriminate the raw product solutions according to the degree of PEGylation and were also able to predict PPR with reasonable accuracy (root mean square error for calibration ∼10%, relative error of prediction < 20%), considering the reference size exclusion chromatography method error of ∼7.2%. The variable selection of the pEEM data suggests that equivalent predictions could be obtained with faster and simpler two-dimensional spectra, making the method a more viable online measurement method.

Comprehensive analysis of the start-up period of a full-scale drinking water biofilter provides guidance for optimization

Abstract. The use of biofilters to produce drinking water from anaerobic groundwater is widespread in some European countries. A major disadvantage of biofilters is the long start-up period required for virgin filter medium to become fully functional. Although individual aspects of biofilter start-up have previously been investigated, no comprehensive study in full scale using inherent inoculation has previously been documented. A thorough investigation of a full-scale drinking water biofilter was carried out over 10 weeks of start-up. The many spatial and temporal changes taking place during start-up were documented using a holistic approach. In addition to collection of many samples over time (frequency) and space (filter depth), this study entailed the use of multiple sample media (water, backwash water and filter media) and multiple types of analyses (physical, chemical and microbiological). The decrease in filter effluent concentrations of individual substances to compliance levels followed a specific order that was shown to coincide with the spatiotemporal development of bacteria on the filter media. Due to the abiotic nature of the iron removal process, iron disappears at the earliest in the start-up period followed by substances that require growth of microorganisms. Ammonium disappears next, with nitrite appearing briefly near the end of ammonium removal, followed by manganese. The thorough overall picture obtained by these efforts provides guidance for optimization and monitoring of the start-up. Guidance for optimization includes shortening the start-up by focusing on kick-starting the ammonium removal; limiting the monitoring burden (at-line measurements of ammonium in finished water supplemented with manual manganese measurements when ammonium removal is complete); and improving filter design by isolating the removal processes in separate, smaller filters.

Process analytical technologies in food industry - challenges and benefits: A status report and recommendations.

Process analytical technologies in food industry - challenges and benefits: A status report and recommendations.

Development of at‐line assay to monitor charge variants of MAbs during production

One major challenge currently facing the biopharmaceutical industry is to understand how MAb microheterogeneity affects therapeutic efficacy, potency, immunogenicity, and clearance. MAb micro-heterogeneity can result from post-translational modifications such as sialylation, galactosylation, C-terminal lysine cleavage, glycine amidation, and tryptophan oxidation, each of which can generate MAb charge variants; such heterogeneity can affect pharmacokinetics (PK) considerably. Implementation of appropriate on-line quality control strategies may help to regulate bioprocesses, thus enabling more homogenous material with desired post-translational modifications and PK behavior. However, one major restriction to implementation of quality control strategies is the availability of techniques for obtaining on-line or at-line measurements of these attributes. In this work, we describe the development of an at-line assay to separate MAb charge variants in near real-time, which could ultimately be used to implement on-line quality control strategies for MAb production. The assay consists of a 2D-HPLC method with sequential in-line Protein A and WCX-10 HPLC column steps. To perform the 2D-HPLC assay at-line, the two columns steps were integrated into a single method using a novel system configuration that allowed parallel flow over column 1 or column 2 or sequential flow from column 1 to column 2. A bioreactor system was also developed such that media samples could be removed automatically from bioreactor vessels during production and delivered to the 2D-HPLC for analysis. With this at-line HPLC assay, we have demonstrated that MAb microheterogeneity occurs throughout the cell cycle whether the host cell line is grown under different or the same nominal culture conditions.

PAT-tools for process control in pharmaceutical film coating applications

Recent development of analytical techniques to monitor the coating process of pharmaceutical solid dosage forms such as pellets and tablets are described. The progress from off- or at-line measurements to on- or in-line applications is shown for the spectroscopic methods near infrared (NIR) and Raman spectroscopy as well as for terahertz pulsed imaging (TPI) and image analysis. The common goal of all these methods is to control or at least to monitor the coating process and/or to estimate the coating end point through timely measurements.

An integrated scale‐down plant for optimal recombinant enzyme production by Pichia pastoris

An integrated bioprocess was created in a scale-down production plant by developing a two-stage enzyme production process with Pichia pastoris, containing a cell-breeding reactor and a production reactor in combination with a three-stage downstream process. To harvest the secreted enzymes, a disc separator and a cross-flow microfiltration clear the broth from the cells. Purification with hydrophobic interaction chromatography removes other proteins, concentrates the product, and prepares the enzyme solution for lyophilization. Fully automated and broad observable multi-stage parallel process courses have been developed using industrial process control systems and at-line measurements for enzyme concentration and enzyme activity. Optimal process conditions were found by application of Design of Experiments (DoE) for the production process.

Optimization of Recombinant Enzyme Production with Pichia pastoris in an Integrated Industrial Scale-down Production Plant

A scale down version of a multi component production plant, including a cell breeding reactor, a protein production reactor and a disc clarifier for enzyme separation is combined to an Integrated Bioprocess. Fully automated and global observable multistage parallel process courses are developed using industrial process control systems and atline measurements for enzyme concentration and enzyme activity. Optimal production conditions were found by application of DoE - Design of Experiment.

PAT – PROCESS ANALYTICAL TECHNOLOGY IN CULTIVATION PROCESSES WITH RECOMBINANT ESCHERICHIA COLI

Abstract An extensive monitoring system is build up to fully automate and scan production processes of recombinant Escherichia coli for optimal process performance. It is based on in-line, on-line and at-line measurements for substrates and products. MATLAB r is connected via OPC OPC Object linked and embedded (OLE) for process control to the SCADA SCADA Supervisory control and data acquisition system MFCS/win r to realise on-line calculations of cell specific and volumetric reaction rates, based on the applied analytical systems. For on-line determination of intracellular target protein recombinant GFP is expressed either as single or fusion protein.

Correlation between Inertial Impaction and Laser Diffraction Sizing Data for Aerosolized Carrier-based Dry Powder Formulations

The purpose of the study was to determine whether the drug fine particle fraction (FPF) from different dry powder aerosol formulations measured by laser diffraction at a range of flow rates correlated with that measured by inertial impaction. Ten binary formulations were prepared containing 1.5% w/w salbutamol base or sulphate, blended with the sieved (63-90 microm) fraction of different sugars (regular lactose, spray-dried lactose, sorbitol, dextrose or maltose). A further six ternary formulations were prepared containing 1.5% w/w salbutamol sulphate, 97% coarse lactose (63-90 microm) and 1.5% micronised or intermediate-sized lactose (1-50 microm). The FPF particles (< 5 microm) of these formulations were measured by laser diffraction and inertial impaction at flow rates between 28.3 and 100 l min(-1). When only the particles with diameter < 60 microm obtained by laser diffraction were considered the FPF (< 5 microm) could be determined and this enabled the aerosolisation of all 16 blends to be feasibly compared at flow rates ranging from 28.3 to 100 l min(-1). A significant linear correlation was found between the fine fractions measured by laser diffraction and the salbutamol fine fractions determined by inertial impaction (r2 = 0.934). Such correlation was also confirmed for formulations containing added fine lactose. Particle size measured by laser diffraction under the employed conditions reflected the aerodynamic properties of the drug. Laser diffraction can be used as on-, in- and/or at-line measurements and controls for dry powder aerosol formulations.

At‐line Determination of Glucose, Ammonia, and Acetate in High Cell Density Cultivations of Escherichia coli

AbstractA sophisticated measurement system for at‐line determination of the main C‐source glucose, the by‐product acetate, and the N‐source ammonium for high cell density cultivations (HCDC) of Escherichia coli K12 TG1 is presented. One flow diffusion technique (FDA) system is used for glucose measurement in the range of 0.5 up to 40 gL–1 in the cultivation broth. Another FDA system detects the amount of the undesired by‐product acetate. The ammonium concentration in the range of 0.2 to 2.5 gL–1 is determined on‐line by a flow injection analysis (FIA) system. For verification purposes, an HPLC system which is also connected to the bioreactor for at‐line measurements is utilized. Several HCDC with cell densities of more than 100 gL–1 have been carried out. The courses of growth‐determining substrates have been detected at‐line. All used systems have shown an excellent compliance with off‐line measurements.

On-Line Monitoring of Physiological Parameters in High Cell Density Cultivations with Recombinant E. coli

An extensive monitoring system of high cell density cultivations (HCDC) with Escherichia coli was developed. It is based on at-line measurements of the main carbon source glucose, the nitrogen source ammonia and the undesired by-product acetate using flow analysis techniques. Besides, MATLAB® was connected to the SCADA system MFCS/win via OPC. Experiments were carried out with two different strains, wild type strain E. coli K12 TG1 (DSMZ 6056) and E. coli JM 105 with green fluorescence protein (GFP). Final cell densities up to 140 gl -1 were reached

Measuring Nylon Carpet Yarn Heat History by Remote NIR Spectroscopy

Heat setting of nylon carpet yarns is one of the most critical processes in carpet manufacturing. The temperature at which the yarn is heat set—the heat-set temperature (HST)—is one of the most important operational variables in the process. Previous investigations with laboratory, bench-top, near infrared (NIR) reflectance instrumentation had shown NIR to be a simple, rapid, and accurate laboratory technique for measuring the HST of nylon carpet yams. The potential of NIR for measuring the HST/heat history of nylon carpet yarns directly on the carpet yarn bobbin and in the manufacturing area is evaluated in a two-phase program. In this paper, Phase One—the evaluation of remote fiber optic sampling systems and the development of a laboratory measurement of HST/heat history directly on the bobbin—is presented and discussed. Nylon 66 yams are heat set by both the Suessen and Superba methods over wide HST ranges. NIR calibrations for HST are developed and validated with two remote fiber optic sampling systems—an interactance probe ("Probe") and a remote reflectance head ("Head"). Excellent agreement is obtained between the stated Suessen and Superba HSTs and the NIR results for both fiber optic sampling systems, with high R2 values (normally &gt;0.9) and low residuals (standard deviation of differences, SDD) of normally &lt;3° for both systems on both heat-set methods. The Probe sampling system is the best overall system for at-line measurements, with low sampling system weight, simpler operation, faster analysis time, and increased flexibility. The Head sampling system is the best overall system for on-line measurements, where its superior accuracy can be optimized.

The Benefits of near Infrared Analysis for Food Product Quality

This is a poster presentation. Raw materials have a decisive influence on the quality of a product. Therefore it is important to check raw materials not only for quality and composition but also for identity. It is also essential to check material during the production process to prevent defective products and to save costs resulting from any out of specification batch. Such in-process measurements are also used for process control. The power of NIR spectroscopy includes both qualitative and quantitative analysis. Results can be obtained within a few seconds. Since no chemicals or time consuming sample preparation is necessary NIR spectroscopy is an ideal analytical method for at-line and in-line measurements. NIR spectroscopy is well established to measure moisture, protein, and fat – the three major constituents in food processing. For these three and a few other constituents a selection of 19 specific wavelengh filters can give all necessary information. But NIR spectroscopy is not only an appropriate tool to analyze these “standard parameters” it can also be very helpful when determining less conventional or minor parameters. Therefore often a higher flexibility regarding the wavelength selection is required which makes the collection of a full spectrum necessary. Some of these less common applications are the determination of caffeine in coffee, theobromine in chocolate or trans fatty acids in margarine. Other product specific parameters which have a strong influence on processing behaviour like the tenderness of peas or the degree of substitution in modified starch can also be measured by NIR.