On-line monitoring of the methanogenic fermentation by measurement of culture fluorescence

Summary Real-time on-line fluorescence measurements of the coenzymes NAD(P)H and F₄₂₀ were evaluated as indicators of stability in a glucose-fed anaerobic methanogenic digester. A probe designed forin situ fluorimetric measurement of NAD(P)H provided an assessment of activity of the total microbial community, while the response of a fluorescence probe designed to measure coenzyme F₄₂₀ correlated well with methanogenic activity. The two fluorescence-monitoring probes responded directly to fermentation imbalance during periods of substrate overloading and corresponded to traditional offline measurements, suggesting that the probes may be suitable candidates for inclusion in an on-line process control system for anaerobic digestion.Florida Agricultural Experimental Station, Journal Series no. R-00326     

On-line monitoring of the denitrification process by measurement of NADH fluorescence

On-line monitoring method of the denitrification process was developed by NADH fluorescence measurements using the facultative microorganism, Ochrobactrum anthropi SY509. The NADH fluorescence signals showed a rapid drop and a rise at the initiation and termination points of the denitrification, respectively. This NADH fluorescence method could be applied successfully to the monitoring of the denitrification process in sequential batch and continuous operations while these rapid changes of fluorescence were not observed in a batch operation due to accumulation of some metabolites secreted from the microorganism.     

On-line biomass monitoring of CHO perfusion culture with scanning dielectric spectroscopy

In this work, dielectric spectroscopy was used to monitor two CHO perfusion culture experiments (B14 and B16). The capacitance of the cell suspension was recorded every 20 minutes over an excitation frequency range of 0.2 MHz to 10.0 MHz. A phase plot of the capacitance at a low excitation frequency vs. the value at a higher frequency proved to be an accurate indicator of the major transition points of the culture, i.e., maximum cell viability, end of lactate consumption, point of zero viability. For both experiments, the capacitance signal correlated very well (R(2) >0.98) with viable cell number up to concentrations of 1 x 10(7) cells/mL. Visual observation of the capacitance spectra indicated that changes in the capacitance relative to frequency were related to the cellular morphology. A multivariate model was developed using off-line data that could predict the median cell diameter within a single experiment (B14) with an error of 0.34 microm (2%). Upon extension to a subsequent experiment (B16), the predicted error was 1.18 microm (9%).     

基于傅里叶变换近红外光谱实时分析1,3-丙二醇发酵过程生物量的在线监测方法

生物量是反映生物发酵过程进展的重要参数,对生物量进行实时监测可用于对发酵过程的调控优化。为克服目前主要采用的离线方法检测生物量时间滞后和人工测量误差较大等缺点,本研究针对1,3-丙二醇发酵过程设计了一个基于傅里叶变换近红外光谱实时分析技术的生物量在线监测实验平台,通过对实时采集光谱预处理以及敏感光谱段分析,应用偏最小二乘算法,建立了1,3-丙二醇发酵过程生物量变化的动态预测模型。以底物甘油浓度为60 g/L和40 g/L的发酵过程作为外部验证实验,分析得到模型的预测均方根误差分别为0.341 6和0.274 3,结果表明所建立的模型具有较好的实时预测能力,能够实现对1,3-丙二醇发酵过程中生物量的有效在线监测。     

Evaluation of a new annular capacitance probe for biomass monitoring in industrial pilot-scale fermentations

The four-pin electrode capacitance probe has already shown to be a valuable tool for on-line monitoring viable biomass concentration in industrial-type fermentations. A new prototype annular probe was developed and its performance in real-time monitoring the concentration of viable cells during industrial pilot-scale fermentation for the production of an Active Pharmaceutical Ingredient (API) was investigated and compared to the four-pin probe. A set of 14 fermentations was monitored on-line: four of them with the four-pin probe, the remaining with the annular probe. The performance of both the annular and the four-pin electrode probe were compared against each other and against off-line measurements (viscosity and packed mycelial volume). The prototype annular probe showed to have higher signal intensity and sensitivity than the standard four-pin probe, with higher signal-to-noise ratio. Furthermore, its new design and construction proved to be easier to handle in an industrial environment.     

Hypothetical model for monitoring microbial growth by using capacitance measurements--a minireview.

Microbiological impedance devices are used routinely by food and manufacturing industries, and public health agencies to measure microbial growth and metabolism. In this paper a hypothetical model explaining the effects of microbial growth and metabolism on capacitance at electrode-medium interfaces, that can be supported by fundamental theories and principles of electrochemistry, is presented. This model provides a framework to interpret changes in capacitance during microbial growth and metabolism and can be used to generate and test hypotheses on factors (i.e., temperature, microbial cell density, microbial growth and medium conductivity) contributing to increases or decreases in capacitance.     

On-line biomass monitoring in bench-scale stirred bioreactors using parts of a liquid chromatography system

A cell density monitor, comprising parts of a liquid chromatography system, was adapted for on-line real-time measurement of bacterial cell density up to 75 g l^–1. The device dilutes the sample containing cells and measures the absorbancy at 280 nm. This detector gives an excellent correlation with off-line measurements, good reproducibility, a low limit of detection (3.5 mg l^–1), and becomes a low cost alternative for fermentation monitoring in small stirred bioreactors.     

On-line measurement in biotechnology: techniques.

Bioprocesses are generally ill controlled. This is due to the fact that the measurement of relevant variables is difficult. Therefore, fundamental knowledge of metabolic interrelations is, at least in vivo, limited. In this article, some of the most important measurement techniques are reviewed in order to provide an evaluation of their current state. Emphasis is given to the underlying principles and on-line capability which allow to judge their importance and potential for exploitation resulting in well (maybe entirely) controlled bioprocesses in the future.     

Real time monitoring biomass concentration in Streptomyces clavuligerus cultivations with industrial media using a capacitance probe

On-line monitoring biomass concentration in mycelial fed-batch cultivations of Streptomyces clavuligerus grown with soluble and partially insoluble complex media, was investigated with an in-situ capacitance probe fitted to an industrial pilot-plant tank. Standard off-line and on-line biomass determinations, including cell dry weight, packed mycelial volume, viscosity, DNA concentration and total CO(2) evolution in the exhaust gases, were performed throughout the experiments and compared to on-line capacitance measurements. Linear relations between capacitance and all other measurements were developed for both media that hold only in defined process phases, depending on the biomass state and the amount of insoluble matter present. For the industrial complex culture media good linear relations were obtained in the fast growth phase between capacitance and DNA concentration and total CO(2) evolution, while in the subsequent transition and stationary phases only with apparent viscosity was a reasonable correlation found. The capacitance probe was shown to be a valuable tool for real-time monitoring biomass concentration in industrial-like cultivation of mycelial streptomycetes.     

Studies of on-line viable yeast biomass with a capacitance biomass monitor

A commercially available biomass monitor has been employed in a number of applications. For capacitance monitors, a relationship between capacitance measurement and cell counts or colony forming units has been reported in the literature. However, for use as an online instrument, a more practical correlation with the biomass concentration is needed. In this study, we followed the batch growth of brewer's yeast and a correlation with viable biomass concentration (g DW/L) was demonstrated. This correlation was utilized with the capacitance biomass monitor in a control loop to maintain setpoint biomass levels in a cyclic reactor under perturbations. Not only did the system demonstrate the capability of the biomass monitor to control biomass in such a system, but it also confirmed the correlation reported in our earlier work. (c) 1994 John Wiley & Sons, Inc.     

On-line measurement of hybridoma growth by culture fluorescence

Summary Fluorescence measurement of viable hybridoma cell cultures provides a convenient method for monitoring the progress of a batch culture. It is shown that cell concentration changes as low as 35,000 cells/ml during initial stages of growth can be measured reliably. This sensitivity, however, decreases to 2 × 10⁶ cells/ml at cell concentration greater than 2 × 10⁶ cells/ml. The culture fluorescence of hybridoma culture is a characteristic property of the cell and the medium used. Consequently, processes in which the medium composition and cell lines are invariant, a direct on-line estimate of viable cell count can be made using the method investigated in this paper.     

On-line monitoring of lipolytic activity by sequential injection analysis

An automated sequential injection analysis using stop-flow technique for the on-line determination of lipolytic activity has been developed. It is based on a colorimetric method using a chromogenic substrate, 1,2-O-dilauryl-rac-glycero-3-glutaric acid-(6-methylresorufin)-ester. The system permits a linear range analysis between 5–100 lipolytic activity units ml^–1, without external dilution of the sample, a sampling frequency of 5 samples per hour and a relative standard deviation (RSD) of 5%. The analyser has been used for the on-line monitoring of Candida rugosa fed-batch fermentation with excellent performance, regarding its reliability and reproducibility.     

On-line monitoring of yeast cell growth by impedance spectroscopy

The application of impedance spectroscopy to estimate on-line cell concentration was studied. The estimation was based on the relative variation between electrical impedance measured at low (10 kHz) and high frequencies (10 MHz). Studies were carried out to characterise the influence of changes in physical and chemical parameters on the impedance measurement. Two different possibilities to perform on-line measurements were tested: a simple set-up, based on an in situ probe, gave good results but was not suitable for high agitation and aeration rates. An ex situ flow-through on-line measuring cell was used to overcome these problems, showing a better performance. The use of this set-up for the growth monitorisation of a Saccharomyces cerevisiae culture showed an efficient performance, having the correlation between estimated and measured S. cerevisiae a Pearson coefficient of 0.999.     

On-line monitoring of intrinsic PEEP in ventilator-dependent patients.

Measurement of the intrinsic positive end-expiratory pressure (PEEP(i)) is important in planning the management of ventilated patients. Here, a new recursive least squares method for on-line monitoring of PEEP(i) is proposed for mechanically ventilated patients. The procedure is based on the first-order model of respiratory mechanics applied to experimental measurements obtained from eight ventilator-dependent patients ventilated with four different ventilatory modes. The model PEEP(i) (PEEP(i,mod)) was recursively constructed on an inspiration-by-inspiration basis. The results were compared with two well-established techniques to assess PEEP(i): end-expiratory occlusion to measure static PEEP(i) (PEEP(i, st)) and change in airway pressure preceding the onset of inspiratory airflow to measure dynamic PEEP(i) (PEEP(i,dyn)). PEEP(i, mod) was significantly correlated with both PEEP(i,dyn) (r = 0.77) and PEEP(i,st) (r = 0.90). PEEP(i,mod) (5.6 +/- 3.4 cmH(2)O) was systematically >PEEP(i,dyn) and PEEP(i,st) (2.7 +/- 1.9 and 8.1 +/- 5.5 cmH(2)O, respectively), in all the models without external PEEP. Focusing on the five patients with chronic obstructive pulmonary disease, PEEP(i,mod) was significantly correlated with PEEP(i,st) (r = 0.71), whereas PEEP(i,dyn) (r = 0.22) was not. When PEEP was set 5 cmH(2)O above PEEP(i,st), all the methods correctly estimated total PEEP, i.e., 11.8 +/- 5.3, 12.5 +/- 5.0, and 12.0 +/- 4.7 cmH(2)O for PEEP(i,mod), PEEP(i,st), and PEEP(i,dyn), respectively, and were highly correlated (0.97-0.99). We interpreted PEEP(i,mod) as the lower bound of PEEP(i,st) and concluded that our method is suitable for on-line monitoring of PEEP(i) in mechanically ventilated patients.     

On-line measurement of catecholamine secretion

A method for the on-line measurement of catecholamine secretion from chromaffin cells is described. Isolated chromaffin cells are placed on a membrane filter and superfused. The perfusate is passed directly over an electrochemical detector, where secreted catecholamines are oxidized. The catecholamine content in the perfusate is measured and recorded as a continuous current output from the detector. This on-line monitor has been used to study the time course of catecholamine secretion, the desensitization of the secretory process, and the recovery of the secretory response to acetylcholine.     

On-line and in-situ monitoring technology for cell density measurement in microbial and animal cell cultures

Commercially available on-line and in-situ devices for monitoring cell density are reviewed in this article. Principles of operation are described as well as capabilities of these probes in specific measurement applications based on literature reports. Pilot-scale experimental observations from three optical density probes, the Cerex, Monitek and Wedgewood designs, have been included for Escherichia coli fermentations. Requirements for future on-line and in-situ instruments are discussed as well as the impact of current limitations on widespread application.     

On-line estimation of biomass using dynamic oxygen balancing

Summary A method for the on-line estimation of biomass is proposed. The method is based on a liquid-phase dynamic mass balance for oxygen during a sequence of no aeration transients. The method has the advantage that no samples need be withdrawn and the biomass need not be homogeneously distributed in the vessel. The technique thus has applications in the estimation of immobilised biomass and plant organ culture, e.g. hairy roots.