Sample Digestion

 The effect of boiling on UDPGA stability was investigated both with regards to

heating time as well as the chemical nature of the solution in which the UDPGA was

dissolved in. Boiling in water for 3 minutes resulted in a 12.7% loss of UDPGA (present

as standard solution), compared to boiling in 0.25M H2PO4 for the same amount of time

(7.7% loss). A final boiling time of 1 min was chosen since measurement of the

temperature inside the glass tube showed that, within 15 seconds, the temperature of the

water within the tube rose to 980C when the tube was plunged into briskly boiling water.

In addition, UDPGA loss when boiling in buffer for this period of time was minimal

(Figure 6-1), and did not lead to appreciable decomposition of UDPGA to UDP and UMP

(Bedford et al., 2003; Figure 6-2). Boiling of the liver sample for 1 minute in 0.30 M

buffer, pH 4.3 resulted in less sample loss than boiling for 1 min or 3 min in 0.25 M

buffer, pH 3.4 (Figure 6-3). Thus the final sample treatment conditions chosen were

boiling for 1 minute in 0.30 M H2PO4 in H20, pH 4.3.

 While liver samples were boiled in buffer as a 1 in 5 dilution (0.1 g liver with 0.4

mL buffer), intestinal samples were boiled as 2 in 5 (0.2 g with 0.3 mL), since UDPGA

concentrations in intestine were expected to be significantly less than in liver.