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This paper proposes an extremum-seeking control approach based on sliding mode to achieve the dynamic optimization of methane outflow rate in anaerobic digestion processes. Open-loop analysis for a two-population model shown that the system becomes unstable due at the accumulation of volatile fatty acids (VFA). Then the controller is designed to achieve the regulation of VFA concentration close to the optimal set-point while maximizing the methane production. The control law is based on a variable-structure feedback to iteratively extremize the methane outflow rate and converges to the neighborhood of the optimum with sliding mode motions. In contrast with previous works on extremum-seeking control with sliding mode, the control scheme includes an observer-based uncertain estimator which computes the unknown terms related to the growth kinetics and the inlet composition. Practical stabilizability for …