Purpose: To employ restriction inhibition assay (RIA) to screen phytochemical-rich fractions (PRFs)
with high affinity for Eco
RI and Hin
dIII restriction sequences and correlate their interaction to an
anticancer activity.
Methods: pBR322 linear plasmid DNA was used as a template to screen the sequence-selective
inhibition of aqueous extracts of
Cinnamomum zeylanicum
and
Picrorhiza kurroa
, respectively. The
template was further incubated with different concentrations of PRFs prior to digestion with restriction
endonucleases Hin
dIII and Eco
RI. The Expressed Sequence Tags (ESTs) and Sequence Tag Sites
(STS) of oncogenes were screened for the presence of Eco
RI and Hin
dIII restriction sequences to
associate an anticancer property to PRF.
Results: The inhibitory concentrations of
Cinnamomum zeylanicum aqueous extract against Hin
dIII and
Eco
RI endonucleases were approximately 2.5 and 5 μg/μl, respectively. No binding was observed for
Picrorhiza kurroa at both Hin
dIII and Eco
RI restriction sites. The saponin-rich fractions of
Cinnamomum
zeylanicum showed significant (p < 0.001) inhibition as compared to control at concentrations of
0.28±1.45 μg/μl for Eco
RI and 0.11±2.68 μg/μl for Hin
dIII endonucleases. Both Eco
RI and Hin
dIII
restriction sites were found repeatedly in the STS and ESTs of BRCA2, the early onset oncogene.
Conclusion: The inhibition of endonucleases by phytochemical-rich fractions provides direct evidence
of the use of RIA for screening as well as demonstrating the binding specificity of these PRFs. The
presence of 5’-AAGCTT-3’ & 5’-GAATTC-3’ in the ESTs of BRAC2 provides an insight into the use of
screened components as leads in the search for novel anticancer compounds.