Bone Mineral Content Analysis Protocol

After removing the attachment on the right tibia, bone mineral content was measured. The tibia was dried at 105 °C for 48 h then defatted with ether for seven days and dried to constant weight at 105 °C. The dried bones were crushed and put in the crucible then carbonized on an electric furnace until smokeless and burned in a muffle furnace at 550–600 °C for 24 h (RC-MF12-30, Beijing Ruicheng Yongchuang Technology Co., Ltd., Beijing, China). After obtaining the tibia ash, it was made into sample decomposition liquid. The calcium (Ca) was analyzed by the Ethylene Diamine Tetraacetie Acid (EDTA) method according to the GB/T6436–2018. The phosphorus (P) was determined via the molybdate-yellow colorimetry method based on the GB/T6437–2018.