The purpose of this paper is explore if the use of a high-performance work system (HPWS) facilitates the development of organizational ambidexterity directly (OA) or through a mediating variable such as human resource flexibility (HRF). We propose a quantitative approach. The theoretical model and the hypotheses proposed were tested using a sample of 100 Spanish hotels. The data analysis method used was that of Partial Least Squares (PLS). This paper represents a valuable contribution to the literature in several ways. Firstly, this research advances in the understanding of the ambidexterity antecedent’s factors. Especially in the role that human capital plays and in ambidextrous learning. Thus, this research expands the scarce empirical evidence about the role of human resources in the achievement of simultaneity in exploration and exploitation learning. Secondly, the present paper has incorporated a mediator variable (human resource flexibility) between HPWS and organizational ambidexterity which so far never been treated in studies linking human resources and organizational ambidexterity. And finally, our findings demonstrate that the relation between human resources and ambidexterity is partially mediated by human resource flexibility. The present research is likely to provide managers with some evidence on how and what can be achieved with a suitable human resource management in hotel firms. Firstly, the utilization of HPWS (i.e. comprehensive staffing, extensive training, development performance appraisal, and an equitable reward system) can facilitate organizational ambidexterity. This is so because human resource management system can improve employees’ ability to efficiently exploit existing knowledge and effectively explore new knowledge. Secondly, the utilization of HPWS can facilitate human resource flexibility. This capability plays a very important role in organizations, since it allows them to quickly adapt to the changes operated in the environment, and the present research work has served to verify that this capability is equally an antecedent factor in organizational ambidexterity development. Considering the context in which the present study has taken place ‒Spanish hotels‒ this outcome presents highly interesting practical implications due to the fact that the Spanish model for flexibility in the tourism sector has almost exclusively focused on temporary employment which, despite the possibility for this type of work to generate improvements in productivity via labor cost reductions in the short term. Therefore, the present research can actually help managers to look for alternative labor flexibility formats –i.e. behavioral flexibility (develop employee’s ability to adapt their responses and actions to any new circumstances which eventually arise in the workplace), skill flexibility (develop employee’s versatility in order to perform different tasks and/or functions), and human resource practice flexibility (implement people management practices which can easily adapt to any organizational contingencies– .