Objective To investigate the clinical features, muscle pathological changes, blood and urine screening by mass spectrometry, and gene mutations of riboflavin responsive multiple acyl-CoA dehydrogenation deficiency (RR-MADD) caused by electron transfer flavoprotein dehydrogenase (ETFDH) mutation, and to provide help for early diagnosis and treatment. Methods A retrospective analysis was performed for related data of 15 patients with RR-MADD caused by ETFDH mutation who were diagnosed in our hospital from 2009 to 2019. Results The 15 patients had a mean age of onset of 32.1±13.6 years and had the initial symptom of limb weakness, among whom 53.3% had weakness in all extremities and 46.7% had weakness in both lower limbs. All patients had increases in muscle enzymes, and electromyography showed myogenic damage in 80% of the patients, myogenic and neurogenic damage in 13.3% of the patients, and normal results in 6.7% of the patients. The positive rates of blood and urine mass spectrometry were 66.7% and 22.2%, respectively. Gene detection showed that all patients had mutations at different loci of the ETFDH gene, among whom 40% had single heterozygous mutations, 40% had compound heterozygous mutations, and 20% had homozygous mutations. Conclusions This disease mainly manifests as fluctuating muscle weakness and elevated muscle enzymes. Muscle pathological examination should be performed for such patients as soon as possible, and a combination of blood and urine metabolic screening and gene detection may help with early diagnosis and timely treatment of such patients.