Use of at‐line and in‐situ near‐infrared spectroscopy to monitor biomass in an industrial fed‐batch Escherichia coli process

Abstract

One of the key goals in bioprocess monitoring is to achieve real-time knowledge of conditions within the bioreactor, i.e., in-situ. Near-infrared spectroscopy (NIRS), with its ability to carry out multi-analyte quantification rapidly with little sample presentation, is potentially applicable in this role. In the present study, the application of NIRS to a complex, fed-batch industrial E. coli (RV308/PHKY531) process was investigated. This process undergoes a series of temperature changes and is vigorously agitated and aerated. These conditions can pose added challenges to in-situ NIRS. Using the measurement of a key analyte (biomass) as an illustration, the details of the relationship between the at-line and in-situ use of NIRS are considered from the viewpoint of both theory and practical application. This study shows that NIRS can be used both at-line and in-situ in order to achieve good predictive models for biomass. There are particular challenges imposed by in-situ operation (loss of wavelength regions and noise) which meant the need for signal optimisation studies. This showed that whilst the at-line modelling process may provide some useful information for the in-situ process, there were distinct differences. This study shows that the in-situ use of NIRS in a highly challenging matrix (similar to those encountered in current industrial practice) is possible, and thus extends previous works in the area.

Application of near‐infrared spectroscopy for monitoring and control of cell culture and fermentation

“…In many production processes, conventional off-line analysis is performed daily or at the end of the fermentation, which results in a rather unreliable and inefficient analysis procedure. 10 At-line measurements offer an improvement to this situation: the analyzer is within the vicinity of the bioreactor and samples taken from the vessel are analyzed quickly, typically within minutes. 10 However, the ideal approach is monitoring on-line, preferably in situ where the analyzer is in direct contact with the process stream, providing real-time determination of the analytes.…”

Cole–Cole, linear and multivariate modeling of capacitance data for on-line monitoring of biomass

“…Unfortunately, its use in in-situ applications is limited because the measurement is very sensitive to bubbles, cell aggregation and non-cellular scattering particles present in the suspension [8]. Success in on-line, in-situ monitoring of biomass with near-infrared probes has been shown by several authors [9][10][11]. Optical monitoring of cell concentration and average cell volume can also be achieved with in-situ microscopy (ISM) [5,12].…”

Real‐time update of calibration model for better monitoring of batch processes using spectroscopy

“…Nearinfrared spectroscopy (NIR) has found the most widespread acceptance. Recent studies include the monitoring of biomass (Arnold et al, 2002;Zhang et al, 2002), glucose (Arnold et al, 2000;Jung et al, 2002;Rhiel et al, 2002a;Sivakesava et al, 2001a), lactose (Macedo et al, 2002), glutamine (Rhiel et al, 2002a), glutamate (Arnold et al, 2000) exopolysaccharides (Macedo et al, 2002) ammonia (Arnold et al, 2000;Rhiel et al, 2002a), lactate (Rhiel et al, 2002a;Sivakesava et al, 2001a), and acetate (Zhang et al, 2002). However, interest is shifting toward mid-infrared spectroscopy (MIR) because it allows higher selectivity and signal-to-noise ratio than NIR.…”

“…This is to ensure that all the spectral variations in the bioreactor are taken into account in the calibration model. Culture media are composed of many different molecules resulting in calibration sets that can easily approach 100 standards (Arnold et al, 2002;Cannizzaro, 2002;Kornmann et al, 2003c;Macedo et al, 2002;Pollard et al, 2001;Rhiel et al, 2002b). Preparing and collecting close to 100 standards is very timeconsuming.…”