In this industrial revolution era, university-level education emphasises higher-order thinking skills. This research aims to analyse the effect of implementing the PjB-HOTS learning model on cognitive learning, creative thinking skills, analytical thinking skills, and metacognitive skills of the students studying osmoregulation concepts in Animal Physiology courses. The sample consisted of 61 fifth-semester students divided into two classes. An open-ended test was used to measure students' cognitive learning levels, creative thinking skills, analytical thinking skills, creative thinking skills, and metacognitive skills. The research data were analysed using the Analysis of Covariance (ANCOVA). The results of this research show that the learning model had a significant effect on the cognitive learning gains (0.000< [alpha]= 0.05), creative thinking skill (0.001< [alpha]= 0.05), analytical thinking skill (0.000< [alpha]= 0.05), and metacognitive skill (0.000< [alpha]= 0.05). These results indicate that the combined syntax of PjBL and HOTS can foster students' higher-order thinking skills and improve their cognitive learning gains. We recommend the PjB-HOTS learning model be used for other biological concepts at the university-level.