Dietary Phosphorus Rhythms in Laying Hens

At the age of 70 weeks, a total of 120 hens were randomly selected to evaluate dietary interventions of body phosphorus rhythms. The hens were fed with 4 phosphorus regimens: (1) RR, provided with regular phosphorus diet at both 09:00 and 17:00 (conventional feeding without considering daily rhythms of body phosphorus metabolism); (2) RL, provided with regular phosphorus diet at 09:00 and low phosphorus diet at 17:00 (dynamic feeding converse to the body phosphorus rhythms found in Exp. 1); (3) LR, provided with low phosphorus diet at 09:00 and regular phosphorus diet at 17:00 (dynamic feeding consistent with the body phosphorus rhythms found in Exp. 1); (4) LL, provided with low phosphorus diet at both 09:00 and 17:00 (direct restriction without considering daily rhythms of body phosphorus metabolism). Each feeding regimens included 6 replicates, and each replicate contained 5 hens. The regular and the low phosphorus diet contained 0.32% and 0.14% NPP, respectively. The feeding trial lasted for 12 weeks (according to the literature, changes in eggshell and bone mineralization status could be observed in 8 to 12 weeks after dietary phosphorus interventions in laying hens) [26 (link), 27 (link)]. On the last 3 d of the feeding trial, all the eggs were collected for egg quality analysis. On the last day of the feeding trial, two egg-laying hens were randomly selected from each replicate (sampled at 6 and 18 h post-oviposition, respectively). The following samples were collected: blood (for serum), uterine (stored at −80 ℃, for Western-blotting analysis ), femur (left side, stored in 4% paraformaldehyde for histological analysis; right side, stored at −80 ℃ for the determination of mineralization status and gene expressions).