Chemical exchange saturation transfer (CEST) imaging has been demonstrated to discuss the concentration changes of amide proton, glutamate, creatine, or glucose measured at 3.5, 3.0, 2.0, and 1.0-1.2 ppm. However, these peaks in z-spectra are quite broad and overlap with each other, and thus, the independence of a CEST signal on any specific metabolite is still open to question. Here, we described whether there was interference among the CEST signals and how these CEST signals behave when the power of the presaturation pulse was changed. Based on these results, further experiments were designed to investigate a method to increase the independence of the CEST signal in both phantoms and animals. The result illustrates a clear interference among CEST signals. A presaturation power adjusted pulsed- (PPAP-) CEST method which was designed based on the exchange rates of the metabolites can be used to remove contributions from other exchanging species in the same sample. Further, the method was shown to improve the independence of the glutamate signal in vivo in the renal medulla in mice. The PPAP-CEST method has the potential to increase the independence of any target CEST signals in vivo by choosing the appropriate combination of pulse amplitudes and durations.