There has been rapidly accelerating interest in the utilization of volatile organic compounds (VOCs) as non-invasive methods for rapid point-of-care medical diagnostics. There is widespread variation in analytical methods and protocols, with little understanding of the effects of sample storage on VOC profiles. This study aimed to determine the effects on VOC profiles of different storage times, at room temperature, prior to freezing, of sealed urine samples from healthy individuals. Analysis using Field Asymmetric Ion Motility Spectrometry (FAIMS) determined the alterations in VOC and total ion count profiles as a result of increasing room temperature storage times. Results indicated that increasing exposure time to room temperature prior to freezing had a threefold effect. Firstly, increased urinary VOC profile variability, with a plateau phase between 12 and 48 hours, before further degradation. Secondly, an increase in total ion count with time exposed to room temperature. Finally, a deterioration in VOCs with each sample run during the analysis process. This provides new insight into the effect of storage of urine samples for VOC analysis using FAIMS technology. Results of this study provide a recommendation for a 12-hour maximum duration at room temperature prior to storage.