Two-dimensional temperature-responsive chromatography using a poly(N-isopropylacrylamide) brush-modified stationary phase for effective therapeutic drug monitoring

Kenichi Nagase,Masakazu Inoue,Hideko Kanazawa,So Inoue

doi:10.1038/s41598-022-06638-1

Abstract

Therapeutic drug monitoring (TDM) is an effective pharmacological approach for controlling drug concentration in a patient’s serum. Herein, a new two-dimensional chromatography system was developed using two poly(N-isopropylacrylamide) (PNIPAAm)-modified bead-packed columns for effective and safe drug monitoring. PNIPAAm-modified silica beads were prepared as packing materials using atom transfer radical polymerization of NIPAAm. The increase in the retention times of the drugs requiring TDM with increasing temperature, was attributed to enhanced hydrophobic interactions at elevated temperatures. The drugs and serum proteins were separated on the prepared column at 40 °C using an all-aqueous mobile phase. Differences in the hydrophobic interactions accounted for the elution of the serum proteins and drugs at short and long retention times, respectively, and a primary column was employed to separate the serum proteins and drugs. After eluting the serum proteins from the column, the drug was introduced into the secondary column, leading to a peak of its purified form and enabling determination of the drug concentration. Two-dimensional temperature-responsive chromatography can benefit TDM by allowing the drug concentration in the serum to be measured in all-aqueous mobile phases without sample preparation.

Highlights

Therapeutic drug monitoring (TDM), that is, the monitoring of drug concentrations in patients, is essential for effective pharmacological therapy, as the dosages of specific types of drugs differ among patients because of their varying absorption, distribution, and elimination levels[1]
A PNIPAAm brush with a low polymer density was prepared on silica beads by diluting CPTMS (ATRP initiator) on the beads, as the highly dense PNIPAAm brush-modified bead-packed column exhibited an excessively long retention time for analytes in previous studies[40,43]; this long retention time was not suitable for the short analysis time required for TDM
The PNIPAAm brush-modified beads (PN-1000 and PN-1500) exhibited larger carbon compositions compared to the silane layer-modified silica beads, indicating successful modification of PNIPAAm on the silica beads via atom transfer radical polymerization (ATRP) under the conditions employed in this study

Summary

Introduction

Therapeutic drug monitoring (TDM), that is, the monitoring of drug concentrations in patients, is essential for effective pharmacological therapy, as the dosages of specific types of drugs differ among patients because of their varying absorption, distribution, and elimination levels[1]. Ordinary chromatography requires an organic solvent in the mobile phase to adjust drug retention in the column. Ordinary chromatography requires sample preparation (pretreatment) to remove serum proteins This time-consuming sample preparation process, which generally requires organic solvents for serum protein denaturation, delays the monitoring of drug concentrations in a patient. To overcome these issues, we developed a method based on temperature-responsive chromatography using poly(N-isopropylacrylamide) (PNIPAAm)-modified bead-packed columns and two-dimensional chromatography. Temperature-responsive chromatography employing a PNIPAAm-modified stationary phase can modulate analyte retention by changing the column temperature through the intrinsic properties of P NIPAAm6–8. Chromatography does not require addition of an organic solvent to the mobile phase to control analyte retention.

Methods

Results

Conclusion