Supervised multivariate data analysis of annotated metabolites in Sorghum bicolor seedlings following UHPLC-MS analysis of the 14 d vs. 29 d group samples. (A) An OPLS-DA scores plot of the predictive component t[1] and the first orthogonal component t0[1], R2(cum): 0.989; Q2(cum): 0.980; R2 permutation: 0.259; Q2 permutation: −0.541; p-value: 0.0000; and components: (1+1+0). The ellipse indicates the 95% limit of the Hotelling T2 distribution for the model. (B) Permutation analysis plotting R2 and Q2 (black and blue dots, respectively) from n = 100 permutation tests in the OPLS-DA model. The y-axis shows R2 and Q2, whereas the x-axis shows the correlation coefficient of permuted and observed data. The cluster of points on the left represents 100 permuted R2s and Q2s, and the two points on the right represent the observed R2(cum) and Q2(cum). Dashed lines denote corresponding fitted regression lines for observed and permutated R2 and Q2. (C) OPLS-DA S plot with discriminant biomarkers at each end of the S-plot. The covariance (variable magnitude) and correlation (reliability) of the variables in the model (indicated with black dots) are represented on the axes as p[1] and p(corr)[1], respectively. The features located at the extreme ends of the plot show a positive association (high magnitude and high reliability) to the respective conditions being compared, while those in the middle can be regarded as shared features. (D) A VIP plot where the metabolites of a VIP score of >1 is significant to metabolomic differences corresponding to developmental changes. (Metabolite abbreviations: ODA-3OH(II) = trihydroxy-octadecadienoic acid; ODA-3OH(IV) = 9,12,13-trihydroxy-10-octadecenoic acid; IAA = indole-3-acrylic acid; coumaroyl-glu II = coumaroyl glucose; SA = salicylic acid; sin-glu = sinapaldehyde glucoside I; sinapald-glu II = sinapaldehyde glucoside II; S-flavanone = sophoraflavanone; trp = tryptophan; coniferyl-Ac = coniferyl acetate; sinapoyl-OH = sinapoyl-alcohol; phe = phenylalanine; nar-glu = naringenin 7-O-beta-D-glucoside; IA-L = indole-3-acetyl-leucine; ODA-2OH (II) = 9,10-dihydroxy-12-octadecenoic acid).