Butternut squash (Cucurbita moschata) is a significant crop in terms of the content of bioactive substances with antioxidant properties – vitamin C, vitamin E, carotenoids, polyphenol compounds – as well as B-complex and potassium vitamins are appreciated by the consumer both raw and heat treated. It is suitable for storage, consumption in the fresh state or heat treatment thanks to its interesting sensory properties. The aim of the thesis was to evaluate the influence of genotype and storage conditions on the content of selected bioactive substances in the fruit of these six varieties: Liscia, Matilda F1, Orange, Serpentine, UG 205 F1 and Waltham F1. From bioactive substances we determined the content of total carotenoids and total polyphenols in the flesh of the fruits before and after storage. The total carotenoid content in the fruit before storage was in the range of 40.62 mg.100 g-1 DS (dry substance) to 76.78 mg.100 g-1 DS, after storage the range transformed into 50.58 mg.100 g -1 DS to up to 109.81 mg.100 g-1 DS. The total polyphenol content before storage ranged from 109.95 mg GAE.100 g-1 DS to 198.96 mg GAE.100 g-1 DS, after storage in the range of 152.51 mg of GAE.100 g-1 DS to 199.83 mg of GAE.100 g-1 DS. The results show that the highest content of total carotenoids in fresh fruits after harvest was determined in the pulp of the varieties UG 205 F1 (76.78 mg.100 g-1 DS), Orange (71.40 mg.100 g-1 DS), and high content of total polyphenols was determined in the Matilda F1 strains (198.96 mg GAE.100 g-1 DS) and Orange (166.09 mg GAE.100 g-1 DS). After two months of the fruit’s storage, the total carotenoid content significantly increased in Liscia by 91%, Orange and Waltham F1 both by 54%. The total polyphenol content considerably rose in Liscia and UG 205 F1 varieties by 39%. The highest content of total polyphenols after storage was found in Serpentine fruit (199.83 mg GAE.100 g-1 DS). It follows that from the point of view of the content of total carotenoids it makes sense to store the fruits of the Orange and Waltham F1 varieties and, in terms of total polyphenols content, the fruits of the Serpentine variety. Taking into account the development of the contents of both groups of bioactive substances, storage of the Liscia variety is recommended. The effect of genotype on total carotenoid and polyphenol content in fruit was statistically significant. Storing these pumpkin varieties resulted in an increment of their total carotenoid and polyphenol content, but it did not have any statistical significance.