Introduction: The growing popularity of dietary supplements in Poland raises the need to verify the knowledge and practices of healthcare workers who have a key role in recommending their use. Understanding these issues is important to ensure patient safety and the correct use of dietary supplements. Therefore, the aim of the study was to examine the level of knowledge, practices, and sources of information on dietary supplements among various professional groups of healthcare workers. Material and Methods: The study conducted from September 2023 to June 2024 involved 743 healthcare professionals, including doctors, pharmacists, and nurses. Data were collected online using an original questionnaire. The questionnaire was placed in portals associated with target groups. Results: A total of 73.49% of respondents declared knowledge of dietary supplements. The most common source of information was scientific articles (42.8%), and only 7.00% of respondents used official sources such as the Chief Sanitary Inspectorate. Advertisements had a smaller impact on doctors (p < 0.001), with 46.97% of respondents stating that they did not pay attention to them. In addition, 84.52% of respondents considered the composition of the dietary supplement to be a key factor in their recommendation. Conclusions: Knowledge of dietary supplements among healthcare professionals varies, particularly between different professional groups. The results indicate a need for further education and standardization of information to ensure safe and responsible recommendation of supplements to consumers.