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In Sub-Saharan Africa, post-harvest loss (PHL) of fruits and vegetables significantly contributes to food and nutrient insecurity. Although PHL has an economic impact, the nutritional implications of PHL throughout the food supply chain remain largely unexplored. This research aimed to identify the causes of PHL in the Ho West District of Ghana and estimate the associated nutritional losses. The study adopted a cross-sectional mixed-methods design, involving 299 respondents (81 farmers, 81 traders, and 137 household women). Data were collected using structured questionnaires and focus group discussions. Post-harvest losses were categorised as high or low based on the distribution of reported losses and analysed using chi-square tests, ANOVA, and regression models to identify key drivers. Nutritional losses were calculated using adapted protocols from the World Bank and WBCSD Food Loss and Waste Protocol, focusing on key nutrients such as iron, vitamin A, and vitamin C. Key drivers of PHL included age, education, income level, poor storage, transportation delays, and the length of time produce remained unsold. Nutritional loss estimates revealed substantial iron, vitamin A, and vitamin C losses at the retail and household levels; these losses were converted into equivalent daily requirements for women and children based on standard recommended dietary allowances, assuming complete nutrient loss once produce became unfit for consumption. At the household level, vitamin C losses averaged 4.6 mg per 100 g of edible portion of fruits and vegetables lost, based on standard food composition values. Thematic analysis revealed inadequate knowledge of storage, limited preservation tools, and misconceptions about food spoilage. Post-harvest handling practices, socio-demographic factors, and lack of awareness contribute to significant nutrient losses across the fruit and vegetable value chain. Interventions should focus on training, household-level preservation techniques, and improved infrastructure to reduce nutritional losses and support food security.