Figure 4. The metabolic process of fatty acids and incorporation of label into the TCA cycle from [1-¹³C₈] octanoate.

(A) Schematic diagram of fatty-acid metabolic process. Long-chain fatty acids are catalyzed by ACSL1, 3–6 (acy-CoA synthetase long-chain 1, 3–6) and CPT1 (carnitine O-palmitoyltransferase 1). Then, long-chain fatty acids are transferred from the cytosol to the mitochondria by CACT (carnitine acylcarnitine translocase) for subsequent β-oxidation. On the other hand, medium-chain fatty acids enter directly into the mitochondrial matrix, and subsequently acyl-CoAs are synthesized. Acyl-CoA-fatty acid products are dehydrogenased by ACADM and ACADL, and the products are subjected to the β-oxidation spiral. (B) Western blotting analysis of ACADM, ACADL, and HADHA in oocytes, 8-cell embryos, and blastocysts (U: unfertilized oocyte, 8: 8-cell embryo, and B: blastocyst). Actin was used as a loading control. An amount of extracted protein corresponding to 100 oocytes or embryos was loaded per lane. The representative result is shown from three independent experiments. (C) Incorporation of ¹³C label into TCA-cycle metabolites in mitochondria from [1-¹³C₈] octanoate-containing culture media. Box plot showing the amount of metabolites derived from octanoate (shaded boxes) and [1-¹³C₈] octanoate (open boxes) cultures. Error bars represent SD. N.D. means that the metabolite concentration was below the detection limit of the analysis. P-value was calculated by Student's t-test (*P < 0.05 and ** P < 0.01).