Bioprocess instrumentation

Process variables measured in bioprocess instrumentation can be categorized into one ofthree groups physical, chemical, and biochemical variables. Table 13.1 summarizes these three types of variables. 

Table 13.1 Process variables measured and estimated in bioprocess instrumentation.

Temperature Ihe temperature in a bioreactor is an important parameter in any bioprocess, because all microorganisms and enzymes have an optimal temperature at which they function most efficiently. For example, optimal temperature for cell growth is 37 °C for Escherichia coli and 30 °C for Saccharomyces sp, respectively. Although there are many types of devices for temperature measurements, metal-resistance thermometers or thermistor thermometers are used most often for bioprocess instrumentation. The data of temperature is sufficiently reliable and mainly used for the temperature control of bioreactors and for the estimation of the heat generation in a large-scale aerobic fermentor such as in yeast production or in industrial beer fermentation. 

Add to these new instruments the breakthroughs in computers (more power, speed, and the ability to go wireless) and software (e.g., chemometrics packages), and the capacity to monitor and affect bioprocesses in real time is more than possible it is almost commonplace. 

A trivial yet important application is following ethanol production via a bioprocess. Sivakesava et al.1 simultaneously measured glucose, ethanol, and the optical cell density of Saccharomyces cerevisiae during ethanol fermentation, using an off-line approach. Samples were brought to an instrument located near the fermentation tanks and the measurements made in short order. While they eventually used MIR due to the interfering scatter of the media, they proved that Raman could be used for this application. 

Similarly, in a paper by Sonnleitner,29 many of the instruments available for monitoring bioprocesses are described. He discusses various conventional and nonconventional monitoring instruments and evaluates them for usefulness and benefits, also discussing their pitfalls. 

Having seen the number of papers devoted to bioprocess analyses utilizing vibrational spectroscopy, it cannot be considered an experimental tool any longer. Manufacturers are responding to pressure to make their instruments smaller, faster, explosion-proof, lighter, less expensive, and, in many cases, wireless. Processes may be followed in-line, at-line, or near-line by a variety of instruments, ranging from inexpensive filter-based to robust FT instruments. Raman, IR, and NIR are no longer just subjects of feasibility studies they are ready to be used in full-scale production. 

D. Dochain and M. Perrier. Advanced Instrumentation, Data Interpretation, and Control of Biotechnological Processes, chapter Monitoring and Adaptive Control of Bioprocesses, pages 347-400. Kluwer Academic Publishers, 1998. 

Instrumentation. A Pharmacia BioPilot Column Chromatography system was used to perform large-scale size exclusion chromatography (SEC) with an 11.3 x 90 cm BioProcess column packed with Sephacryl S-200 HR gel. High performance size exclusion (HPSEC) and ion exchange chromatography (HPIEC) were conducted with Pharmacia Superose 6 and 12 (HR 10/30) and Mono-Q (HR 5/5) columns respectively, equipped with Beckman model 520 system controller and Beckman model HOB HPLC pumps. 

The BioView sensor (DELTA Light Optics, Denmark) was developed especially for industrial applications. It is capable of completely automatic optical measurement for monitoring and control of different bioprocesses. The instrument is conceived to withstand harsh industrial environments (e.g., high temperature, moisture) and electromagnetic interference. For data transfer a single-fiber asynchronous modem is used, which allows a distance between the computer and spectrometer of up to several hundred meters. 

The BioView sensor includes a software package (CAMO ASA, Norway) for data analysis and on-Une estimation of different bioprocess variables simultaneously. Thus, the instrument is able to predict the trends of the concentration courses of different variables during a cultivation and is used to give information about important process steps (e.g., feeding time, harvesting time, etc.). The instrument is able to monitor on-line several fluorophores in situ and non-invasively during cultivation processes and permits an estimation of different bioprocess variables simultaneously. The increasing of cell mass concentration and the product formation as well as the actual metabolic state of the cells is simultaneously detectable by this fluorescence technique. 

As with refining and petrochemical processes, bioprocesses must be operated automatically so as to achieve a consistent production of various bioproducts in a cost-effective way. In particular, there is a strong demand to optimize bioprocesses by controlling them automatically to promote labor-saving operations. To achieve this, it is necessary to understand what is happening in a bioreactor (instrumentation) and to properly manipulate the control variables that affect the performance of a bioreactor operation (control). 

To overcome this difficulty, many challenges have been met in the fields of bioprocess control. In this chapter, the basic principles of the instrumentation and control of bioprocess are explained. 

In this section, we describe the instrumentation for monitoring the current status of bioprocess plants, focusing on the measurement of process variables as the basis of bioprocess control. 

Biosensors Enzymatic analysis of components and products from bioprocesses is widely utilized because this type of analysis is both selective and sensitive. To use enzymes for the automatic analysis and instrumentation of bioprocesses, various sensors using enzymatic reactions, the so-called "biosensors, have been... 

The goal of bioprocess control is to maintain important process variables in a bioreactor at a desired level regardless of time-dependent environmental changes. Process control will be performed by the following two steps based on the information obtained through the instrumentation. 

Analysis of 02 as well as C02 in exhaust gas is becoming generally accepted and is likely to be applied as a standard measuring technique in bioprocessing. It is possible to multiplex the exhaust gas lines from several reactors in order to reduce costs. However, it should be taken into account that the time delay of measurements with classical instruments is in the order of several minutes, depending on the efforts for gas transport (active, passive) and sample pretreatment (drying, filtering of the gas aliquot). 

The principles, sampling systems, control of the measuring device and application of MS for bioprocesses have been summarized by Heinzle [157,158] and Heinzle and Reuss [162]. Samples are introduced into a vacuum (< 10 5 bar) via a capillary (heated, stainless steel or fused silica, 0.3 x 1000 mm or longer) or a direct membrane inlet, for example, silicon or Teflon [72,412]. Electron impact ionization with high energy (approx. 70 eV) causes (undesired) extensive fragmentation but is commonly applied. Mass separation can be obtained either by quadrupole or magnetic instruments and the detection should be performed by (fast and sensitive) secondary electron multipliers rather than (slower and less sensitive) Faraday cups (Fig. 21). 

Liquid samples can be collected from the bioreactor sampling port and introduced into the electronic nose instrument manually. More appealing in bioprocessing is to sample on-line. An electronic nose system monitors non-in-vasively by sampling from the off-gas port of the bioreactor [23]. The humidity... 

Modern NIR equipment is generally robust and precise and can be operated easily by unskilled personnel [51]. Commercial instruments which have been used for bioprocess analyses include the Nicolet 740 Fourier transform infrared spectrometer [52, 53] and NIRSystems, Inc. Biotech System [54, 55]. Off-line bioprocess analysis most often involves manually placing the sample in a cuvette with optical pathlengths of 0.5 mm to 2.0 mm, although automatic sampling and transport to the spectrometer by means of tubing pump has been used (Yano and Harata, 1994). A number of different spectral acquisition methods have been successfully applied, including reflectance [55], absorbance [56], and diffuse transmittance [51]. 

Transferability of spectral data and models in NIR spectroscopy. This subject is an issue that is pertinent to the future use of NIR for bioprocess monitoring. Pre-processing to remove baseline shifts and noise in spectra from individual machines or direct standardisation by data transformation with a representative subset can be used to calibrate across instruments [61]. 

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