Guinea pigs are popular companion pets and veterinary care is becoming increasingly important for these small mammals. This study aimed to evaluate plasma amylase in guinea pigs to determine an association between amylase and mortality, as there are limited studies evaluating prognostic indicators in this species. This retrospective study evaluated medical records from four institutions across the United States from January 1, 2016, to August 31, 2022. A total of 278 records were included, with exclusion of records with unknown survival 30-days post biochemistry panel. Mortality was not linearly associated with amylase concentrations; however, higher mortality was seen with marked elevations [1910-3985 U/L; adjusted odds ratio (aOR): 10.31; 95% confidence interval (CI): 3.55 to 29.89]. Guinea pigs with markedly elevated amylase were seven times more likely to die within 30 days compared to those with a value within the reference range (1339-1573 U/L; AOR: 7.83; 95% CI: 0.67 to 4.16). There was no association between elevated amylase and other factors. Amylase concentrations had overall limited accuracy to predict 30-day mortality, as shown by an area under the curve (AUC) of the receiving operator characteristic curve of 72.7% (95% CI: 65 to 79.9%). For a cut-off value of amylase being 1623 U/L, there was a 70.2% sensitivity and 66.5% specificity. Given the AUC, amylase should not be used as a standalone predictor of mortality in guinea pigs. However, derangements of this biochemical marker could be an indicator of disease in this species.