We propose the separation algorithm to simultaneously measure two-dimensional refractive index distribution and thickness profile of transparent samples using three wavelengths. The optical system was based on the Mach-zehnder interferometer with LD (Laser Diode)-based multi-wavelength sources. A LCR (Liquid Crystal Retarder) was used to obtain interference images at four phase states and then the optical phase of the object is calculated by four-bucket algorithm. Experimental results with a glass rod are provided at the different wavelengths of 635nm, 660nm and 675nm. The refractive indices of the sample are distributed with accuracy of less than 0.0005 and the thickness profile of sample was cylindrical type. This result demonstrates that it is possible to separate refractive index distribution and thickness profile of samples in two dimensions using the proposed algorithm.