|Abstract or Summary
- This study was undertaken to standardize and evaluate the
tyrosinase test as a variety identification procedure for wheat. The
potential value of the tyrosinase test lies in its ability to quantify
the colored products, rather than relying on visual estimation as
in the phenol test.
The specific objectives of this study were to: (a) standardize
the methodology of the tyrosinase test, (b) determine the effect of
various non-genetic characteristics on the results of the test,
(c) determine the ability of the tyrosinase test to distinguish Pacific
Northwest wheat varieties, and (d) compare the relative merits of
the tyrosinase and phenol tests as variety identification methods for
The test parameters evaluated were soaking period, temperature,
pH of tyrosine solution, concentration of tyrosine solution, and
shaking effect on enzyme reaction.
The following procedures are suggested for conducting the
tyrosinase test on wheat seed:
For the bulk-seed method soak 2 g of seeds in 10 ml of . 1%
tyrosine solution in sodium citrate buffer .11 M, pH 8, for 2 hours
at 35 C. For the single-seed method, soak the seed in 5 ml of . 1%
tyrosine solution in sodium citrate buffer .11 M, pH 8, in a shaking
water bath for 17 hours at 35 C. Then read the absorbance of the
resulting solution at 470 nm against the substrate solution.
Tyrosinase activity of WS-1 and Yamhill wheat seeds declined
with increasing degrees of deterioration induced by artificial aging.
Vitavax treatment caused a reduction in color development or enzyme
activity PCNB (Pentachloronitrobenzene) depressed color development
in WS-1 but not in Yamhill. HCB (Hexachlorobenzene) had no
effect on either variety.
Yamhill seeds produced in Pendleton developed a darker colored
product or had more enzyme or higher enzyme activity than seeds
produced in Moro, while Hyslop seeds from the two locations produced
similar colored product or were comparable in enzyme activity.
Tyrosinase activity of seeds of Hyslop variety was not affected
by seed protein contents of 9, 11, and 13%, indicating that total seed
protein does not affect pericarp enzyme (tyrosinase) quantity or
Larger seeds possessed greater tyrosinase activity than small
seeds because of more pericarp tissue. Variation in enzyme activity
also occurred in seeds of similar size within a variety.
Comparisons with the phenol test showed that the bulk-seed
tyrosinase test probably will differentiate more varieties than the
phenol test and it is less affected by fungicide seed treatments.
Both tests are affected by seed deterioration and production area.
Both tests are relatively quick and can be completed in approximately
the same amount of time.
The phenol test is superior to individual-seed tyrosinase test
for detecting varietal mixtures. Seeds within a variety differ in
tyrosinase activity because of variations in seed size and other
The bulk-seed test may be useful in differentiating between
varieties belonging to the same phenol color group. Such a quantitative
measurement may also be useful in describing varieties being
entered under the Plant Variety Protection Act. The tyrosinase
test may also provide more accurate results on fungicide treated