Figure 9: Comparison of biological and technical variance. Citrate, creatinine, PAG, and urea were measured in urine samples from natrosol-, furosemide-, or HCBD-treated rats by 1H NMR metabolite quantification. Glucose and lactate values were close to or below the LLOQ and thus excluded from evaluation. To depict biological variance, samples from individual animals ( per treatment group, one dot represents one individual animal), including 4 male (filled symbol) and 4 female (open symbol) animals, were measured. Data from natrosol-treated animals are available from 4 different treatment days ((a), (b), (c), (d); different animals were used on different treatment days). Technical variance is determined from pooled rat urine samples, which are additionally modified concerning pH (depicted values for pH 3 and 9 except for urea) and 6-fold increases in salt-concentration, that is, changes in the accepted range (see before). Additionally, each sample is measured 9 times up to day 14. Values are given as relative concentrations normalized to the mean of the respective treatment and period (biological variance) or the value from the first measurement of the original sample (technical variance). The horizontal grey range (15%) and dotted horizontal lines (20%) show the range of variation stated as acceptable by the “Guidance for Industry” [12].