With the optimization of transfusional and chelation regimens, beta-thalassemia has changed from a pediatric disease with poor life expectancy into a chronic disease. Bone demineralization is an important cause of morbidity in older patients; the etiology is multifactorial and partially unknown. We examined, cross-sectionally, 111 adult patients with beta-thalassemia major (66 females and 45 males, 32.6 ± 6 years) who were regularly transfused, sufficiently chelated and replaced for endocrine defects. Bone demineralization was detected in 92.7% of patients with different severity according to gender and site: osteopenia was the prominent finding at the femur, osteoporosis at the lumbar spine (p < 0.001), more evident in males. The femoral site was more influenced by biochemical and clinical factors; despite adequate replacement, the femoral T-score was lower in the hypogonadic group than in the eugonadic group (p = 0.047). A significant correlation was found between the bone mass and body mass index (BMI), alkaline phosphatase (ALP), and pre-transfusional Hb levels. The multivariate analysis indicated as significant regressors ALP, BMI and hypoparathyroidism (T-score, p = 0.005, 0.035, and 0.002; Z-score, 0.002, 0.009, and 0.003, respectively) at the femoral site; whereas, only ALP at the lumbar spine (p = 0.008 and 0.045 for T-and Z-scores, respectively). The statistical significance was reached more frequently by the T-score, while the Z-score seemed to be a less sensitive parameter. Despite best care facilities, bone demineralization in thalassemic patients remains a challenge. Further exploration of the relationships between bone loss and endocrine, biochemical and hematologic parameters is warranted to find effective measures to reduce the risk of fracture in this disease.