Based on oil samples and rock samples in several petroliferous basins, this paper studied that secondary alterations influence on fluorescence color and spectral parameters of crude oil and oil inclusions by using microscopic fluorescent spectral and geochemical parameters. The results demonstrated that secondary alterations influence on spectral parameters by changing the ratio of saturated hydrocarbon and aromatic hydrocarbon(saturation/aromatics). Biodegradation and water washing lead to redshift of fluorescence parameters while evaporative fractionation and oil cracking lead to blueshift. Beside, mixed oil lead to redshift and blueshift. The secondary alterations and main period of accumulation in research area were recognized by frequency histogram of crude oil and oil inclusion. The relationship between QF-535 and the parameters measuring the secondary alterations could the extent of secondary alterations from which crude oil suffered. In the microscopic fluorescence test, oil inclusions suffered from stretch or leakage, thermal cracking and photooxidation should be rejected.