Biomedical Engineering Reference
In-Depth Information
Raman spectroscopy relies on inelastic scattering involving an energy transfer
between incident light and illuminated light target molecules. Problems in applying
Raman spectroscopy to bioprocesses lie in the strong fluorescence activity of many
biological molecules that often overlay the Raman scattering bonds. In bioprocesses, it
has been used to quantify the production of total intracellular carotenoid astaxanthin
from yeast and alga. Lee et al. [10] have shown quantification of glucose, acetate,
formate, lactate, and phenylalanine in E. coli fermenters.
Optical bio- and chemosensors are most commonly used fluorescent dyes immo-
bilized on the tip of a fiber optic cable. Online measurement of various process
parameters such as pH, CO
2
,orO
2
can be quantified depending on the dye used.
Because the tips are replaceable, this technology offers a broader range of applications
than traditional sensors for pH and off-gas analysis. The change in fluorescence intensity
of ametalorganic dye, caused by quenching of oxygen, is the measuring principle of fiber
optic oxygen sensors. This technology is being taken to the next logical progression for
monitoring bioprocesses by immobilizing enzymes, antibodies, oligonucleotides, and in
some cases even whole cells selective for particular compounds. These affinity sensors
for protein analysis during fermentation and downstream processing have included SPR
and RIF. Most manufacturers of these two kinds of optical biosensors supply instru-
ment-ready surfaces and applicable reagent chemistries for ligand immobilization [31].
The modes of detection are also expanding to include colorimetric, chemiluminescence,
bioluminescence, and electrochemical methods to solve particular process problems.
Pulsed terahertz spectroscopy has become a preferable method for performing
low-frequency spectroscopy over standard FIR (Fourier infrared) techniques. A pulsed
terahertz experiment is similar to conventional pump-probe setups. A terahertz beam
interrogates the properties of a sample (“pump”) and is superimposed on a detector
together with pulses of a second ultrafast beam (“probe”). Varying the time delay of the
beams relative to one another enables the electric field of the terahertz wave to be
reconstructed, and a Fourier transform finally yields the desired spectral information.
Engineers at some of the major optics companies are reportedly working at PTS
modifications to allow determination of the sequence of intermediate tertiary structures
(similar to time-resolved CD) in bioreactors and fermenters. An example is ECOPS
(electrically controlled optical sampling) systems that use two ultrafast lasers that are
phase stabilized to each other. A slight modulation of the length of one of the fiber
oscillators via a piezo element serves to sweep the “probe” pulse through the terahertz
pulse in a precisely controlled manner. PTS is a viable technique for future time-resolved
FIR measurements of protein folding. Thus, it could become a very useful tool for
bioprocess monitoring, since the correct folding of recombinant proteins is one of the
most important factors in the biopharmaceutical industry [31].
Given the high utility of online probes, the most difficult question the bioprocess
engineer has to answer is “Which and how many of these technologies are worth
implementing in my process?” Although NIR is currently viewed as a panacea because it
can measure what most of the other probe types can, it is not always the right tool for the
job. For example, in an industrial process for viral production, an OD meter may best
serve the purpose for measuring cell lysis by monitoring increasing turbidity. A question
that bioengineers need to ask themselves is “Is there truly a need for continuous real time
