Low-carbohydrate, high-protein (LCHP) diets have been shown to induce weight loss and beneficial changes in blood lipids that suggest cardiovascular disease risk reduction; however, LCHP diets have not been adequately investigated for health effects on the skeleton. A randomized trial to determine the effects of a LCHP diet on bone mineral status, biomarkers of bone turnover, indicators of acid-base balance, calcium homeostasis and fasting lipids in healthy pre-menopausal women was conducted. Women, aged 32 - 45 y, with a body mass index between 25-41 kg/m2 were randomized into one of two diet groups: LCHP (n = 13) or high-carbohydrate, low-fat (HCLF) (n = 12). Anthropometric (body weight, lean mass, fat mass) and bone mineral density (BMD) and content (BMC) measures and markers of lipid metabolism were taken at weeks 0, 6, and 12. Measures of acid-base balance, protein metabolism, and calcium homeostasis were conducted at weeks 0, 1-4, 6, and 12. Serum osteocalcin was analyzed at weeks 0, 1, 2, 6, and 12, while urinary NTx was analyzed at weeks 0, 1 and 2. Weight loss was significant at the end of 12 weeks in both diet groups (P < 0.05) but there was no Diet x Time interaction. Total proximal femur BMD was lower in the LCHP group (P < 0.05) compared to the HCLF group by week 12. Femoral neck BMC decreased in the LCHP diet group (P < 0.05), whereas total forearm BMC increased (P < 0.05) in the HCLF diet group by week 12 of the study. Serum osteocalcin showed significant main effects of diet (P < 0.05) and time (P < 0.0001), but a Diet x Time interaction was not observed. Urinary NTx exhibited no main diet effect, time effect or Diet x Time interaction at weeks 1 or 2. Urinary pH was lower in the LCHP group compared to the HCLF group throughout the study (P < 0.0001). Urinary calcium excretion was higher in the LCHP group and lower in the HCLF group (P < 0.0001) compared to baseline values at all intervals of the study. Urinary phosphorus excretion exhibited a significant diet effect (P < 0.001) and time effect (P < 0.002), while no Diet x Time interaction was observed. Total cholesterol, high-density and low-density lipoprotein cholesterol, and triacylglycerol concentrations did not differ between diets during the study. In conclusion, a LCHP diet appears to stimulate bone loss, while a HCLF diet appears to attenuate bone loss in healthy pre-menopausal women undergoing 12 weeks of weight loss.