With the development of the technology of fruit juice adulteration, ordinary methods are difficult to effectively identify whether fruit juice adulteration. The carbon isotope in the biological body is influenced by the external factors, such as climate, environment and so on. Materials from different sources have their own isotope abundance. Although the difference of the isotope abundance is very small, the stable isotope ratio mass spectrometry can be determined accurately. Now stable isotope ratio mass spectrometry has been developed for detection adulteration and also can trace the origin of the materials. Doing some pretreatment to the freshly squeezed orange juice and homemade adulteration orange juice, using isotope mass spectrometer detected the main carbohydrates 'delta 13 c. HPLC device using a Xtimate Sugar-Ca column , the individual components (fructose, glucose, and sucrose) are isolated on a preparative. According to the above requirements to do a series of experiments, through statistical analysis found that the biggest difference of carbohydrates ' delta 13 c is less than 2.1 ‰ in the freshly squeezed orange juice, and in the homemade adulterated orange juice is far greater than 2.1 ‰. Finally, experiments were performed on commercial orange juice, by comparing the difference of the main carbohydrates ' delta 13 c, preliminary identification of orange juice adulteration.