Fig. 1.

Experimental condition. (A) The concept of body-time detection with conventional and molecular timetable methods illustrated by Linné’s flower clock. In the conventional method a single indicator monitored over a few days detects internal body time; in the molecular timetable method, multiple metabolic “flowers” are simultaneously measured at a few time points, which reduce efforts in sampling. (B) Metabolite timetable construction. We collected time-course blood samples, isolated plasma, measured with LC–MS, and selected circadian-oscillating metabolites as indicators. In body-time detection, we collected blood samples at a few time points, isolated plasma, measured with LC–MS, and estimated body time. (C) Image of the constant routine (CR) experiments. During CR, subjects stayed in chairs while various measurements were performed. Note: the man in this picture is demonstrating the set-up and is not an actual subject in this study. (D) Sampling schedule for each subject. Black circles indicate the time points when blood samples were taken and when subjects ate during CR. White boxes indicate the time when subjects were awake, and black boxes indicate when they were asleep. The blood samples of three subjects during CR1 were used for timetable construction, and other samples were used for body-time estimation. (E) Measured melatonin (Upper) and cortisol (Lower) rhythms in the collected blood samples during CR1 (Left) and CR2 (Right). The cortisol and melatonin levels show that some subjects (e.g., subject E in CR2) have shifted internal body time against sampled time. CR, constant routine; SCR, semiconstant routine; BT, body time; LC–MS, liquid chromatography mass spectrometry.