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Fig. 2 DLD-dependent enzyme metabolite analysis and mitochondrial function in dldh–/– zebrafish larvae. (A–H) Biochemical analysis by GC/MS of DLD-dependent enzyme metabolite and aa, relative to undecanoic acid standard and normalized to protein α-ketoglutarate, valine, isoleucine, leucine, lysine, glutarate, α-ketoisovalerate, and lactate in 7 dpf WT (dldh+/+ and dldh+/–)and dldh–/– zebrafish larvae. n = 5 biological replicates/condition. ***P < 0.001; ****P < 0.0001; Welch’s unpaired t test. Data are shown as mean ± SD. (I–K) Lactate and pyruvate, analyzed using lactate and pyruvate oxidase-based colorimetric assays, in 7 dpf dldh–/– zebrafish larvae. Data were normalized to WT sibling controls. *P < 0.05. Student’s t test. (L and M) Analysis of NADH and GSH and GSSG levels (percentage of control) showed no differences between dldh–/– and WT larvae at 7 dpf. All colorimetric analyses were conducted on 4 WT and 3 dldh–/– larval biological replicate samples. Data are shown as mean ± SD.