The initial stage of derivatization method development
embraced testing of the impact of reaction conditions on the yield
of binding of purified ITCs to NAC. In order to shift the reaction
equilibrium towards the formation of ITC-NAC conjugates, we
decided to apply the large excess of NAC (0.2 M) in relation to
the typical content of ITCs (0.02 mM) that can be expected in water
extracts obtained from Brassicaceae plants (Piekarska et al., 2014).
In addition, we included sodium bicarbonate (0.2 M) into the reaction
mixture, so as NAC occurs in the form of sodium salt, which is
easily soluble in water. Such an approach was previously reported
to increase the rate of dithiocarbamate formation