Objective To investigate the effect of Betulinic acid (BA) on myocardial hypertrophy and fibrosis induced by isoproterenol (ISO) in rats and its possible mechanism. Methods 70 SD rats were divided into control group, model group, BA low-dose group, BA high-dose group and Pro group (propranolol positive treatment), with 10 rats in the control group and 15 rats in each other group. The myocardial hypertrophy model rats were established by ISO induction. The rats in BA low-dose group and BA high-dose group were given BA at the dose of 20 mg/kg and 100 mg/kg, respectively, and the rats in Pro group were given propranolol at the dose of 40 mg/kg, once a day, for a total of 21 days. Left ventricular ejection fraction (LVEF), left ventricular fraction shortening (LVFS), left ventricular systolic pressure (LVSP) and left ventricular posterior wall thickness at end diastole (LVPWd) were measured by echocardiography. Heart mass index (HMI) and left ventricular mass index (LVMI) were statistically analyzed. The histopathological changes of rat myocardium were detected by HE staining, the morphological changes of rat cardiomyocytes were detected by WGA fluorescence staining, and the degree of myocardial fibrosis of rat was detected by Masson staining. The expression level of inflammatory factors in rat myocardium was detected by RT-qPCR. The protein expression levels of myocardial hypertrophic markers (ANP, BNP and β-MHC), fibrosis markers (Collagen Ⅰ, Collagen Ⅲ and α-SMA) and the related proteins of β2-AR/ERK1/2 pathway were detected by Western blot. Results Compared with control group, LVEF, LVFS and LVSP in model group were decreased (P<0.05), while LVPWd, HMI and LVMI index were increased (P<0.05). The arrangement of myocardium fibers were disordered, a large number of inflammatory cells infiltrated, the cross-sectional area of myocardium increased (P<0.05), and the collagen volume fraction increased (P<0.05). The protein levels of ANP, BNP, β-MHC, Collagen Ⅰ, Collagen Ⅲ, α-SMA, β2-AR and p-ERK1/2 in myocardial tissue of rat were increased (P<0.05), and the mRNA expression levels of IL-1β, IL-6 and TNF-α were increased (P<0.05). Compared with model group, LVEF, LVFS and LVSP in BA high-dose group and Pro group were increased (P<0.05), while LVPWd, HMI and LVMI index were decreased (P<0.05). The degree of myocardial lesion was significantly decreased, the cross-sectional area of cardiomyocytes was decreased (P<0.05), and the collagen volume fraction was decreased (P<0.05). The protein levels of ANP, BNP, β-MHC, Collagen Ⅰ, Collagen Ⅲ, α-SMA, β2-AR and P-ERK1/2 in myocardial tissue were decreased (P<0.05), and the mRNA expression levels of IL-1β, IL-6 and TNF-α were decreased (P<0.05). There was no significant difference in all indexes between BA high-dose group and Pro group (P>0.05).Conclusion BA can ameliorate the myocardial hypertrophy and fibrosis induced by ISO in rats, and the mechanism may be related to inhibition of β2-AR/ERK1/2 pathway