Aas800

We collected diet samples twice per week and stored them at −20 °C. At the end of the experiment, they were used to analyze the composition and nutrient levels of the basal diet. We determined the DM content using oven drying at 105 °C to constant mass (method 930.15, AOAC) [22 ] and calculated the dry matter intake (DMI) using DM. We used Kjeldahl nitrogen analysis to determine the CP (method 945.16, AOAC) [22 ] and used a Soxhlet extractor to determine the EE content (method 945.16, AOAC) [22 ]. NDF and ADF were analyzed using heat-stable amylase (A3306, Sigma Chemical Co., St. Louis, MO, USA) and sodium sulfite according to the procedure of Van Soest [23 (link)]. In addition, we used a muffle furnace to measure the ash content by combustion (method 942.05, AOAC) [22 ]. We used the colorimetric method and atomic absorption spectrometry to analyze phosphorus (Spectrophotometer UV755N, Yoke Instrument Co., Ltd., Shanghai, China) and calcium (PerkinElmer AAS800, Waltham, MA, USA), respectively [24 (link)].

Samples of each diet were collected twice per week, stored at −20°C, and composited weekly for the analysis of DM, CP, ether extract (EE), neutral detergent fiber (NDF), acid detergent fiber (ADF), ash, calcium (Ca), and phosphorus (P). We calculated dry matter intake (DMI) according to the feed intake recorded by Automatic feeding equipment (Insentec Co., The Netherlands) and DM obtained through dietary analysis. The DM content was determined by oven drying at 105°C to constant mass (method 930.15, AOAC) [13 ], the CP content was determined using Kjeldahl nitrogen analysis (method 945.16, AOAC) [13 ], and the EE content was determined using a Soxhlet extractor (method 945.16, AOAC) [13 ]. NDF and ADF were analyzed using heat-stable amylase (A3306, Sigma Chemical Co., St. Louis, MO, USA) and sodium sulfite according to the procedure of Van Soest et al [14 (link)]. The ash content was measured by combustion using a muffle furnace (method 942.05, AOAC) [13 ]. A colorimetric method was used for the analysis of phosphorus (Spectrophotometer UV752N, Yoke Instrument Co. Ltd., Shanghai, China) and calcium was measured using atomic absorption spectrometry (PerkinElmer AAS800, Waltham, MA, USA).

Automatic feeding equipment (Institute of Animal Sciences, Chinese Academy of Agricultural Sciences, Beijing, China, and NanShang Husbandry Science and Technology Co. Ltd., Henan, China) was used to record DMI after calving. Samples of each diet were collected twice per week, stored at −20°C and composited weekly for analysis of DM, CP, ether extract (EE), neutral detergent fiber (NDF), acid detergent fiber (ADF), ash, calcium (Ca), and phosphorus (P). The DM content was determined by oven drying at 105°C until the weight is constant (method 930.15, AOAC) [17 ]. The CP content was determined using Kjeldahl nitrogen analysis (method 945.16, AOAC) [17 ]. The EE content was determined using a Soxhlet extractor (method 945.16, AOAC) [17 ]. The NDF and ADF were analyzed using heat-stable amylase (Sigma no. A3306, Sigma Chemical Co., St. Louis, MO, USA) and sodium sulfite according to the procedure of Van Soest et al [18 (link)]. Ash was measured by combustion using a muffle furnace (method 942.05, AOAC) [17 ]. The colorimetric method was used for analysis of phosphorus (Spectrophotometer UV752N, Yoke Instrument Co. Ltd., Shanghai, China) and calcium was measured using atomic absorption spectrometry (PerkinElmer AAS800, Waltham, MA, USA).