This study delves into the intricacies of aquaporin water channels (AQP) in cartilage formation, emphasising their expression and functionality in chondrogenic cultures. Cartilage, a vital element in joint function and mobility, is a robust avascular tissue. It is composed of chondrocytes and an extracellular matrix (ECM) rich in glycosaminoglycans and type II collagen. This unique composition provides cartilage with the ability to withstand considerable mechanical stress while maintaining its structural integrity. During the process of chondrogenesis, chondroblasts are responsible for synthesising and depositing ECM components. In this context, AQPs are hypothesised to play an indispensable role in regulating cell volume and water homeostasis, critical for proper ECM formation and cartilage functionality. Our research employed micromass cultures from limb buds of 4-day-old chicken embryos. We conducted RNA sequencing to analyse AQP transcript levels, complemented by western blotting for protein expression. The impact of the AQP inhibitor Phloretin on chondrogenic differentiation and ECM formation was assessed through MTT assays and metachromatic staining. Finally, RT-qPCR was utilised to determine the expression of cartilage-specific genes. The findings revealed transcription-level expressions of AQP1, AQP3, AQP4, AQP9, AQP10, and AQP11. Intriguingly, Phloretin markedly diminished ECM production but paradoxically upregulated chondrogenic markers and boosted mitochondrial activity in chondrogenic cells. These observations underscore a complex and nuanced role of AQPs in cartilage formation. This investigation offers insights into the role of water transport mechanisms in cartilage development and highlights the potential for targeting these channels in enhancing the quality of cartilage grafts for tissue engineering. The effects of Phloretin on chondrogenic cultures pave the way for further exploration into the molecular mechanisms of chondrogenesis, offering potential avenues for future therapeutic applications.