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The compound (2S)-eriodictyol is an important flavonoid that can be derived from (2S)-naringenin through flavonoid 3'-hydroxylase (F3'H) catalyzation. F3'H is a cytochrome P450 enzyme that requires a cytochrome P450 reductase (CPR) to function. However, P450s have limited applications in industrial scale biosynthesis, owing to their low activity. Here, an efficient SmF3'H and a matched SmCPR were identified from Silybum marianum. To improve the efficiency of SmF3'H, we established a high-throughput detection method for (2S)-eriodictyol