3D Electrode and EIS for Real-Time Microalgae Growth Monitoring

Microalgae are a promising source of biomass and valuable compounds for aquaculture, food, cosmetics, and bioenergy. Compounds like exopolysaccharides (EPS), which microalgae secrete for protection, are of great biotechnological interest.

However, optimizing their production requires precise monitoring of the culture’s growth phases, something current methods do not always facilitate. Periodic sampling, the need for qualified personnel, and laboratory processes (such as centrifugation or flocculation) are often slow, costly, and do not provide real-time data.

What if we could “see” microalgae growth and EPS production continuously, sensitively, and affordably directly in the culture? A recent study led by scientists from the University of Coimbra and the University of Bath, published in the prestigious journal Biosensors and Bioelectronics, presents an innovative solution for precise real-time monitoring of microalgae growth and the metabolites they produce, with various industrial applications. This new technology combines Electrochemical Impedance Spectroscopy (EIS) with three-dimensional (3D) porous electrodes, enabling the development of a probe capable of monitoring microalgae growth and their EPS production in real time.

The Challenge: Monitoring Small, Slow-Growing Organisms

Monitoring microbial growth with electrochemical techniques is not new, especially for bacteria. However, microalgae present particular challenges: they are considerably larger than bacteria (often >20 µm) and their doubling time can be much slower, sometimes taking several days.

This means that electrodes with a much larger surface area than conventional ones are needed, and they must be electrochemically stable for weeks to capture the complete growth dynamics.

The Innovation: Large Surface Area 3D Porous Electrodes

To overcome these challenges, the researchers developed a novel electrode with a three-dimensional, porous structure. They used polyurethane (PU) foams as a base, a common and inexpensive material. These foams were carefully cleaned and hydrophilized (treated to attract water) using oxygen plasma.

Then, they immersed them in a solution of a conductive polymer called PEDOT:PSS (poly(3,4-ethylenedioxythiophene) polystyrene sulfonate), modified to improve its adherence and conductivity. This coating process was repeated twice and finalized with a thermal treatment (annealing) to fix the coating and ensure good electrical conductivity.

The result is an electrode with an internal structure of interconnected pores (similar to a sponge) coated with the conductive polymer. This design offers an enormously larger effective surface area (estimated at 240 cm²) compared to traditional planar electrodes (on the order of 10 mm² in this study).

Crucially, the coating process did not significantly obstruct the pores, maintaining the open structure (porosity only varied by 2% after two coating layers) while increasing electrical conductivity. The stability of these electrodes was confirmed through long-term tests (14 days) in the culture medium (BG11) without cells, where no significant variations in impedance were observed.

How Does Electrochemical Impedance Spectroscopy (EIS) Work?

EIS is a technique that measures a system’s resistance to the flow of an alternating electrical current at different frequencies. When microalgal cells adhere and grow on the surface of the 3D porous electrode submerged in the culture medium, they alter the electrical properties at the electrode-liquid interface.

Imagine that the cells and the EPS they produce act as an insulating layer that slightly hinders the current flow, especially at low frequencies. EIS detects these subtle changes in impedance (a complex measure of resistance and capacitance). By measuring impedance over time, the dynamics of cell growth and the accumulation of products like EPS can be tracked.

Key Results of the Study with Lobochlamys segnis

The researchers used the microalga Lobochlamys segnis (formerly known as Chlamydomonas segnis), a model known for its EPS production, to test the system. They cultured this microalga directly on the 3D porous electrodes for 14 days and performed continuous EIS measurements.

• Real-Time Monitoring and High Sensitivity: The EIS system could continuously track the growth of L. segnis throughout the 14-day experiment. Notably, it worked even with very low initial cell densities (10⁵ cells/mL), corresponding to minimal initial electrode coverage (~0.012%). This demonstrates the high sensitivity of the method.
• Correlation with Traditional Methods: The growth curve obtained from impedance measurements (specifically at low frequency, 0.1 mHz) closely followed the curve obtained through traditional cell counting under a microscope. Applying a logistic growth model, the growth rate estimated by EIS (kz​=0.75 /day) was very similar to that obtained by cell counting (kcells​=0.85 /day). This agreement validates the EIS technique as a reliable indicator of microalgal growth.
• Detection of EPS Production: The study revealed that EPS production by L. segnis significantly impacts impedance measurements, especially in the initial growth phases (days 0-6). The cells and the surrounding EPS layer (bEPS) cover the electrode surface, increasing the low-frequency impedance. The maximum impedance approximately coincided with the time of greatest visible EPS production around the cells (days 8-10).
• Indicator of EPS Release into the Medium: An interesting finding was that the parameter Rsol​ (solution resistance), extracted from the EIS equivalent circuit analysis, proved sensitive to the time when EPS begin to be massively released from the cells into the culture medium (around day 12). The rate of change of Rsol​ also correlated with the initial cell density, being higher in denser cultures. This suggests that EIS could be used not only to monitor cell growth but also to detect peaks in the release of extracellular products.
• Advantage over Planar Electrodes: A direct comparison showed that the 3D porous electrodes, thanks to their larger surface area, allowed tracking of the complete growth curve, including the exponential phase, much better than conventional small-area planar PEDOT:PSS electrodes.

Paulo Rocha, professor at the Department of Life Sciences (DCV) and researcher at the Centre for Functional Ecology (CFE) at the University of Coimbra, explains: “We employ Lobochlamys segnis as a microalgae model system and show that growth can be continuously monitored for at least 14 days. The results indicate that the logistic growth rate obtained through EIS is similar to conventional cell counting. Furthermore, the ohmic resistance proved to be a reliable indicator for detecting the peak point of EPS production.”

Implications for Aquaculture and Biotechnology

The results of this study are very promising for the aquaculture sector and microalgae biotechnology. According to the experts involved, this technology represents a significant advancement for the biotechnology industry. The ability to monitor the growth and production of metabolites like EPS in real time, continuously, sensitively, and potentially affordably, opens up new avenues:

• Bioreactor Optimization: Allows adjustment of culture conditions (light, nutrients, etc.) at the optimal time to maximize the production of biomass or specific compounds like EPS.
• Quality Control: Facilitates more precise monitoring of the culture’s health and physiological state.
• Cost Reduction: Could decrease the need for frequent manual sampling and complex laboratory analyses.
• New Applications: The high sensitivity, even at low densities, suggests potential for more efficient water resource management, such as the early detection of algal blooms.

Conclusion: Towards Smart Microalgae Monitoring

The combination of Electrochemical Impedance Spectroscopy (EIS) with 3D porous electrodes based on PU/PEDOT:PSS emerges as a powerful and sensitive tool for real-time monitoring of the growth of microalgae like Lobochlamys segnis and the dynamics of exopolysaccharide (EPS) production.

By providing continuous data and correlating well with standard methods, this technology offers a promising alternative to conventional monitoring techniques, with the potential to significantly improve efficiency and control in the large-scale production of microalgal biomass and bioproducts in bioreactors and, possibly, in environmental monitoring. Likewise, the developed technology complements the use of artificial intelligence or absorption spectra to determine the growth or pigment content of microalgae.

This advancement is the result of collaboration between researchers from the University of Coimbra (PhD students Francisco Cotta Jr and Felipe Bacellar, CFE researchers Raquel Amaral and Diogo Correia, led by Paulo Rocha) and Professor Kamal Asadi from the University of Bath, UK.

Paulo R.F. Rocha
Centre for Functional Ecology-Science for People & the Planet, Associate Laboratory TERRA, Department of Life Sciences, University of Coimbra
Coimbra, 3000–456, Portugal
Email: [email protected]

Reference (open access)
Cotta, F. C., Amaral, R., Bacellar, F. L., Correia, D., Asadi, K., & Rocha, P. R. (2025). A 3D porous electrode for real-time monitoring of microalgal growth and exopolysaccharides yields using Electrochemical Impedance Spectroscopy. Biosensors and Bioelectronics, 277, 117260. https://doi.org/10.1016/j.bios.2025.117260