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Modeling Yeast Cell Viability in Continuous-Flow Pulsed Electric Field Treatment

This study demonstrates that in continuous-flow pulsed electric field treatment of yeast, the repetition rate and residence time act as independent factors influencing cell viability beyond their combined pulse count, necessitating their separate specification for accurate modeling and process control.

Original authors: Jânio Anselmo, Afrânio de Castro Antônio Júnior, Jéssica Rodrigues, Guilherme Brasil Pintarelli, Daniela Ota Hisayasu Suzuki

Published 2026-10-26

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CC BY 4.0

Original authors: Jânio Anselmo, Afrânio de Castro Antônio Júnior, Jéssica Rodrigues, Guilherme Brasil Pintarelli, Daniela Ota Hisayasu Suzuki

Original paper licensed under CC BY 4.0 (https://creativecommons.org/licenses/by/4.0/). ⚕️ This is an AI-generated explanation of a preprint that has not been peer-reviewed. It is not medical advice. Do not make health decisions based on this content. Read full disclaimer

In the world of food processing, keeping liquids like juice or wine fresh without cooking them is a delicate balancing act. Traditional heat treatment kills harmful microbes but often ruins the flavor, color, and nutritional value of the product. To solve this, scientists have developed a method called pulsed electric field treatment. Imagine sending a rapid series of short, sharp electrical shocks through a liquid. These shocks create tiny, temporary holes in the membranes of bacteria and yeast cells. If the shocks are strong enough and frequent enough, these holes become permanent, causing the cells to die without heating the liquid to a damaging degree. This technique is widely used in laboratories, but moving it from a static container to a flowing pipe for industrial use introduces a new layer of complexity. In a flowing system, the liquid is constantly moving, meaning every drop spends a different amount of time in the electric field depending on how fast the pump is pushing it.

Researchers at the Federal University of Santa Catarina in Brazil set out to understand exactly how this flow affects the death of yeast cells, a common microorganism used in fermentation and a frequent cause of spoilage in beverages. They built a specialized chamber where a yeast suspension could flow continuously between two metal plates while being hit with electrical pulses. The team tested a wide range of conditions, changing the strength of the electric field, how often the pulses fired, and how fast the liquid moved. They wanted to see if the total number of pulses a cell received was the only thing that mattered, or if the timing of those pulses and the speed of the flow created different outcomes.

The study revealed a surprising truth: the total count of pulses is not enough to predict whether a cell will survive. The researchers found that two different setups could deliver the exact same number of pulses to a cell, yet produce very different results. In one specific comparison, they adjusted the speed of the flow and the frequency of the pulses so that a cell received the same number of shocks in both cases. However, when they counted the surviving yeast, one setup left three times as many living cells as the other. This proved that the history of the exposure matters. It is not just about how many times a cell is hit, but how quickly those hits come and how long the cell is exposed to the electric field overall.

To explain this, the team developed a new way of thinking about the treatment. Instead of just counting pulses, they treated the speed of the pulse repetition and the time the liquid spent in the chamber as two separate, independent factors. They also carefully measured the temperature rise caused by the electricity. While the electric shocks generated heat, the researchers found that the heat itself was not the main cause of cell death in their experiments. The electrical effect was far more powerful, killing cells even when the temperature rise was relatively small. By separating the electrical effects from the thermal ones, they created a model that could accurately predict how many cells would survive under different operating conditions.

This work is significant because it challenges the simple idea that more pulses always equal more killing. The researchers showed that in a flowing system, the timing of the pulses interacts with the movement of the liquid in a complex way. If the pulses come too fast or the liquid moves too slowly, the cells experience a different kind of stress than if the pulses are spaced out differently, even if the total number of shocks is identical. This finding suggests that engineers designing industrial machines for pasteurizing liquids cannot just set a total pulse count. They must carefully tune the flow rate and the pulse frequency separately to ensure the treatment is effective without wasting energy or overheating the product. The study provides a clearer roadmap for making this technology reliable for real-world use, ensuring that beverages can be treated safely while keeping their fresh, natural qualities intact.

Technical Summary: Modeling Yeast Cell Viability in Continuous-Flow Pulsed Electric Field Treatment

Problem Statement
Optimizing Pulsed Electric Field (PEF) processes for continuous-flow systems presents a significant challenge because the biological response depends on coupled factors that are often conflated in batch processing. In continuous-flow chambers, the number of pulses a cell receives is the product of the pulse repetition rate (RR) and the mean residence time ( $t_{res}$ ). A critical gap in current modeling is the assumption that conditions yielding the same effective pulse number ( $N_p = RR \cdot t_{res}$ ) produce identical biological outcomes. However, in flow-through systems, different combinations of flow rate and repetition rate create distinct thermal signatures, local exposure histories, and pulse spacings, which may lead to different cell inactivation rates despite identical nominal pulse counts. Existing models based solely on total pulse number, treatment time, or delivered energy often fail to distinguish these operating conditions.

Methodology
The study investigated Saccharomyces cerevisiae (strain TB10) in a parallel-plate continuous-flow chamber with a 1 mm electrode gap. The experimental matrix varied three primary parameters while holding others constant:

Applied Voltage: 300–500 V (300–500 kV/m).
Repetition Rate (RR): 150–250 Hz.
Volumetric Flow Rate (Q): 10–20 mL/min.
Pulse Characteristics: 4 µs monopolar rectangular pulses.

The system utilized a peristaltic pump to drive the suspension through the chamber, with active cooling via thermoelectric modules to manage temperature rise. Researchers recorded voltage and current waveforms, measured temperature rise ( $\Delta T$ ) between inlet and outlet, and assessed cell viability using methylene-blue staining and Neubauer chamber counting (7 replicates per condition).

A novel effective-dose mathematical model was developed to describe cell viability ( $C_v$ ) as:
$C_v = C_{v0} \cdot e^{-(\Omega_T + \Omega_E)}$
Where:

$\Omega_T$ (Thermal dose) was calculated using literature kinetic constants for S. cerevisiae (Török & King, 1991) and integrated over the residence time, rather than being fitted to the data.
$\Omega_E$ (Electrical dose) treated repetition rate and residence time as separate, independent factors rather than collapsing them into a single pulse count. The model incorporated a threshold field ( $E_c$ ) and sensitivity exponents for repetition rate ( $\eta_R$ ) and residence time ( $\eta_t$ ).

Key Results

Decoupling of Pulse Number and Viability: The study demonstrated that conditions with identical effective pulse numbers ( $N_p$ ) produced significantly different viable-cell densities. Specifically, a condition with 150 Hz at 15 mL/min and a condition with 200 Hz at 20 mL/min (both $N_p \approx 156$ ) resulted in a 3.12-fold difference in viable-cell density ( $9.6 \times 10^4$ vs. $3.0 \times 10^5$ cells/mL), with no overlap between replicates ( $p = 5.8 \times 10^{-4}$ ).
Model Performance: The proposed model, which separates RR and $t_{res}$ , reproduced the experimental medians with high accuracy ( $R^2 = 0.9979$ ). Leave-one-out cross-validation yielded $R^2_{LOO}$ values between 0.61 and 0.84.
Thermal vs. Electrical Contribution: The computed thermal dose was more than four orders of magnitude lower than the electrical dose, indicating that inactivation was primarily driven by electroporation rather than thermal effects, despite measurable temperature rises (up to 39.2 °C).
Parameter Sensitivity: The model identified a residence-time sensitivity exponent ( $\eta_t \approx 4.89$ ), suggesting a steeper-than-proportional sensitivity to transit time, likely due to cumulative damage mechanisms and membrane resealing dynamics.

Significance and Claims
The primary contribution of this work is the integration of experimental data with a mathematical framework that explicitly decouples repetition rate and flow rate in continuous-flow PEF modeling. The authors claim that:

Repetition rate and residence time must be specified separately. Relying solely on the product $N_p$ is insufficient for predicting cell viability in continuous systems because pulse spacing affects membrane recovery and thermal history.
The effective-dose model provides a consistent framework for comparing operating conditions that share the same nominal exposure but differ in their dynamic history.
Thermal contributions can be quantified independently using literature constants, preventing the conflation of thermal and electrical effects in the model fitting process.

The study concludes that continuous-flow PEF viability modeling must account for coupled electrical, thermal, and hydrodynamic variables. The developed model serves as a tool for selecting operating conditions that balance cell inactivation, temperature rise, and treatment capacity, though the authors note that the model is currently validated only within the specific experimental matrix (300–500 kV/m, 150–250 Hz, 10–20 mL/min) and requires further testing for extrapolation to other matrices or geometries.

Technical Summary: Modeling Yeast Cell Viability in Continuous-Flow Pulsed Electric Field Treatment

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