Chemical exchange saturation transfer (CEST) magnetic resonance imaging (MRI) is capable of measuring dilute labile protons and microenvironmental properties. However, the CEST contrast is dependent upon experimental conditions—particularly, the radiofrequency (RF) irradiation scheme. Although continuous‐wave RF irradiation has been used conventionally, the limited RF pulse duration or duty cycle of most clinical systems requires the use of pulsed RF irradiation. Here, the conventional numerical simulation is extended to describe pulsed‐CEST MRI contrast as a function of RF pulse parameters (i.e., RF pulse duration and flip angle) and labile proton properties (i.e., exchange rate and chemical shift). For diamagnetic CEST agents undergoing slow or intermediate chemical exchange, simulation shows a linear regression relationship between the optimal mean RF power of pulsed‐CEST MRI and continuous‐wave‐CEST MRI. The optimized pulsed‐CEST contrast is approximately equal to that of continuous‐wave‐CEST MRI for exchange rates less than 50 s−1, as confirmed experimentally using a multicompartment pH phantom. In the acute stroke animals, we showed that pulsed‐ and continuous‐wave‐amide proton CEST MRI demonstrated similar contrast. In summary, our study elucidated the RF irradiation dependence of pulsed‐CEST MRI contrast, providing useful insights to guide its experimental optimization and quantification. Magn Reson Med, 2011. © 2011 Wiley‐Liss, Inc.