Parijoto fruit (Medinilla speciosa Blume) of Family Melastomataceae is a plants grow on the slopes of the Muria Mountains, Colo Village, Kudus District, Indonesia. The methanol extract of Parijoto fruit was able to reduce blood sugar levels, improve sexual function, exhibit moderate cytotoxicity in T47D cancer cells, potential for chemoprevention, ability to lower glucose levels, antihyperlipidemic, antiobesity, stimulating effect on the quantity spermatozoa, increase the number of Leydig cells, Sertoli cells, primary spermatocytes and seminiferous tubule diameter. Parijoto fruit contains flavonoid compounds, saponins, terpenoids, glycosides and tannins, beta-carotene and antioxidants, and alkaloids. So that these plants can be developed and preserved, it is necessary to carry out continuous research, so that the types of compounds can be known. To determine the content of active compounds from plants, and identification, so that pure compounds are obtained. This study aims to provide information on the results of the isolation and identification of phytoconstituents from the methanol extract of parijoto fruit. The methanol extract of parijoto fruit was diffracted using n-hexane. Fraction then was continued with the isolation process. The process of isolation and identification of methanol extract of parijoto fruit using Preparative Thin Layer Chromatography method. Structure these compounds were determined based on the Gas chromatography-mass spectrometry (GC-MS), Fourier Transform Infrared Spectroscopy (FT-IR) and NMR spectra. This study succeeded in isolating three compounds from the methanol extract of parijoto fruit. Namely stigmasterol (1) of n-hexane fraction, stigmasterol (2) and 1-hexanol (3) of methanol fraction.