158 / 172
D). However, even with less free, unreacted urea in the sample, recovery by these urease sources was
not improved. From the data in Table 5, the lower activity urease sources failed substantially to
hydrolyze all the urea in these UF fertilizers, while the high activity sources did hydrolyze all the urea.
The fact that high activity sources (No. 1, 7, and 20 ml extracts of No.2) show the same percent urea
hydrolyses as the amount found by HPLC analyses, support that such percentages urea are indeed
present in these UF fertilizers samples. The combined performances of HPLC and the high activity urease
sources indicate that the AOAC Method 959.03 is very much dependent on the type of urease.
The above data show that analyses of Urea-Formaldehyde fertilizer solutions using the urease method
are not consistent. In most cases, the amount of free and unreacted urea analyzed by the urease
method showed a low biased (Table 6). However, in some occasions the results showed a high bias
(Table 3).
These data support the discussion that AOAC official Method 959.03 (Urease Method) is not suitable for
the determination of free-unreacted urea in the Urea-Formaldehyde fertilizer solutions. The HPLC
Method consistently provides more accurate analyses of unreacted urea in this class of fertilizers.
As mentioned above, there are two AOAC Official Methods by HPLC for the determination of unreacted
urea in these fertilizers, namely AOAC Official Method 983.01, and AOAC Official Method 2003.14.
The advantage with AOAC 983.01 is the use of water as the mobile phase, however due to the use of
water, the peak separations in the complex compositions of these fertilizers (Figure 1) is not efficient
and some of the peaks might overlap with that of urea resulting in erroneous data. A collaborative
study
3
by the International Standards organization (ISO-CD 18643) involving 13 laboratories for the
biuret content of several fertilizers, including the water-soluble Urea Triazone fertilizers,has showed that
AOAC 983.01 HPLC method results in erroneous analyses of some of these UF fertilizer solutions. The
same study showed that the AOAC 2003.14 method accurately predicts the amounts ofunreacted urea
in these fertilizers.
Table 8 shows the inconsistency of the AOAC Official Method 983.01 (HPLC) for the determination of
biuret in the Urea-Triazone fertilizers (Grade 26-0-0). The biuret co-elutes with the free and unreacted
urea in these fertilizers and their peaks overlap resulting in erroneous analytical data.