Free radicals are associated with degenerative diseases such as cholesterol, the pathogenesis of diabetes, liver damage, nephrotoxicity, inflammation, cancer, and premature aging. Long-term use of synthetic antioxidants has a toxic effect compared to natural antioxidants, so it is necessary to explore natural antioxidants to replace synthetic antioxidants, such as using plants containing flavonoids and phenols. It is known that the extract and methanol fraction of Parijoto fruit contained flavonoids, saponins, tannins, and glycosides. This study aimed to determine whether extracts, fractions and isolates of Parijoto fruit (Medinilla speciosa Blume) have antioxidant and anticholesterol activity in vitro. This study was carried out in several stages, namely extraction, fractionation, and isolation of parijoto fruit. The parijoto fruit underwent antioxidant activity test in vitro using the DPPH method and anticholesterol activity in vitro testing using the Lieberman-Burchard method. The collected data underwent normality and homogeneity tests. If the results are normal and homogeneous, One Way Anova parametric test is carried out, followed by post-hoc LSD. If the results obtained are not normal and not homogeneous, then a non-parametric test analysis is carried out, namely Kruskal Wallis and continued by Mann Whitney. The antioxidant and anticholesterol tests showed significant differences between each group. The 40.64 μg/ml methanol fraction group had the highest antioxidant activity while the lower methanol isolate had the highest anticholesterol atcivity of 94.27%. Methanol extract, methanol fraction, n-hexane fraction, upper methanol isolate, lower methanol isolate from Parijoto fruit had potential as antioxidants and anticholesterol in vitro.