Simultaneous Estimation of Protein and Nucleic Acid Using Derivative Spectrophotometric Method

Purpose: Simple and precise first derivative zero crossing spectrophotometric method has been developed for the simultaneous estimation of protein and nucleic acid without prior separation. Approach: The concentration ranges for protein and nucleic acid were taken in the range of 60-140 m g mL -1 and 20-100 m g mL -1 respectively in 0.1 M NaOH. Absorption spectra of the samples were recorded between 200 nm to 400 nm against a reagent blank. Zero-order spectra of protein and nucleic acid were stored individually within the above concentration ranges and were derivatized in first order using scaling factor 5 for both the substances. First derivative amplitudes were recorded at 261 nm and 289 nm for estimation of protein and RNA respectively for pure substances and binary mixture. Finding: Limit of detection was found to be 2.90 m g mL -1 and 0.36 m g mL -1 and limit of quantitation was 8.80 m g mL -1 and 1.02 m g mL -1 respectively for protein and nucleic acid respectively. Precision was found to be 1.38 % for protein and 1.25 % for RNA. Reproducibility was found to be 1.54 % for protein and 1.69 % for RNA. Conclusions: Thus, proposed method can be adapted for simultaneous determination of protein and nucleic acid and better alternate technique for immunoassays and electrophoretic methods.