he cultivation of Calendula officinalis has gained importance not only for its medicinal properties but also for its ornamental value. C. officinalis flowers contain carotenoids and oleanolic acid, valuable phytoconstituents with various medicinal properties. However, to meet the increasing demand for this plant, it is necessary to adopt sustainable cultivation practices that optimize growth, enhance flowering, and maximize the content of bioactive compounds. An experiment was conducted under greenhouse conditions during the 2021/2022 and 2022/2023 seasons on C. officinalis using chemical fertilizer (NPK-RD: recommended dose of NPK), organic fertilizer (poultry manure), and biofertilizers. The biofertilizers included different combinations of plant growth-promoting microbes: N-fixing bacteria (Az), P-solubilizing bacteria, K-solubilizing bacteria (Bc), and vesicular-arbuscular mycorrhizal (VAM) fungi. The results showed that the application of poultry manure was more pronounced compared to biofertilizers. The principal component analysis (PCA) revealed that NPK-RD, moderate and higher levels of organic fertilizer, and Az+Bc+VAM treatments achieved significant superiority in all tested parameters. A higher dosage of poultry manure (95.20 m3 ha−1) was significantly superior to chemical fertilizer in plant height, branch number, plant dry weight (DW), chlorophyll, carbohydrates, flower number, flower yield fresh weight (FW), and carotenoids content. However, the flowering start, flowering period, flower yield DW, and oleanolic acid content obtained by poultry manure at 95.20 m3 ha−1 were significantly equal to NPK-RD treatment. The inoculation with Az+Bc+VAM microbes positively impacted carotenoids content in the C. officinalis flower. These results underline the potential of organic fertilizer and plant growth-promoting microbes in improving growth, flowering and phytochemical quality of the C. officinalis plant with the possibility of avoiding chemical fertilization.