Alkylating agents are considered to be archetypal carcinogens.
One suitable technique to evaluate the activity of alkylating compounds
is the NBP assay. This method is based on the formation of a chromophore
in the reaction between the alkylating agent and the nucleophile 4-(p-nitrobenzyl)pyridine (NBP), a trap for alkylating agents
with nucleophilic characteristics similar to those of DNA bases. NBP
is known to react with strong and weak alkylating agents, and much
insight into such alkylation mechanisms in vivo can
be gained from kinetic study of some alkylation reactions in vitro. Since 1925, the NBP assay has evolved from being
a qualitative, analytical tool to becoming a useful physicochemical
method that not only allows the rules of chemical reactivity that
govern electrophilicity and nucleophilicity to be applied to the reaction
of DNA with alkylating agents but also helps to understand some significant
relationships between the structure of many alkylation substrates
(including DNA) and their chemical and biological responses. Given
that advances in this area have the potential to yield both fundamental
and practical advances in chemistry, biology, predictive toxicology,
and anticancer drug development, this review is designed to provide
an overview of the evolution of the NBP method from its early inception
until its recent kinetic–mechanistic approach, which allows
the pros and cons of NBP as a DNA-model to be analyzed. The validity
of NBP as a nucleophilicity model for DNA in general and the position
of guanosine at N7 in particular are discussed.