This study investigated the time-related changes in the alanine aminotransferase (ALT), aspartate aminotransferase (AST) and alkaline phosphatase (ALP) values of serum and plasma samples stored at refrigerator (2 - 8oC, average of 40C) and room temperature (30-340C, average of 320C) for a period of 120 hours (5 days). Blood samples were obtained from a total of 20 patients that presented with cases of liver malfunction at the Ebonyi State University Teaching Hospital, Abakaliki, Nigeria. The enzyme assays were carried out immediately upon sample collection and separation to obtain the baseline value (BV), and thereafter at specified time intervals across the 120 hours. Results showed that values not significantly different (p > 0.05) from the BV can be obtained from serum and plasma samples within the specified storage durations at the different temperatures as follows: For serum ALT, 40C – 48 hours, 320C – 16 hours; plasma ALT, 40C – 36 hours, 320C –10 hours; serum AST, 40C – 36 hours, 320C – 10hours; plasma AST, 40C – 36 hours, 320C – 8 hours; serum and plasma ALP, 40C – 30 hours, 32oC – 10 hours. All the enzymes were found to be more stable in refrigerated samples than in those kept at room temperature; also the enzymes were more stable in serum than in plasma for all the storage temperatures. It was concluded that in general, reliable values for serum and plasma ALT, AST and ALT (values not significantly different from the BV) can be obtained from refrigerated samples if analysed within 30 hours of blood collection and separation, while for samples kept at room temperatures (320C) analysis should be carried out within 8 hours. Beyond these time points (30 hours for refrigerated samples and 8 hours for samples kept at room temperature) values obtained for the serum and plasma enzymes studied were significantly different (P