This study assesses the metabolic status of rat diabetic cardiomyopathy (DCM) models. Echocardiography is used to detect the diastolic dysfunction in type 2 diabetic rats, and a lower threshold for inducible atrial fibrillation is found in type 2 diabetic rats with diastolic dysfunction compared to the control. Metabolic abnormalities are detected by status changes of reduced nicotinamide adenine dinucleotide (phosphate) (NAD(P)H), which is an essential coenzyme in cells or tissues. Fluorescence lifetime imaging microscopy (FLIM) is used to monitor changes in NAD(P)H in both myocardial tissues and blood. FLIM reveals that the protein-bound proportion of NAD(P)H in rat myocardium in the DCM group is smaller than the control group, which indicates the oxidative phosphorylation rate of the DCM group decreased. Similar results are found for blood plasma of DCM rats by the FLIM study. FLIM exhibits high potential for screening DCM as a label-free, sensitive, and noninvasive method.