# Development and Validation of an HPLC-UV Method for the Quantification of Acyclovir and Ganciclovir in the Plasma of Pediatric Immunocompromised Patients

## TL;DR

A new HPLC-UV method was developed and validated to measure acyclovir and ganciclovir in the blood of immunocompromised children.

## Contribution

A low-cost, efficient HPLC-UV method for quantifying antiviral drugs in small plasma samples is introduced.

## Key findings

- The HPLC-UV method showed linearity in the range of 0.5–40 mg/L for both drugs.
- Results from the HPLC-UV method correlated significantly with those from LC-MS/MS.
- The method uses minimal sample volume and is suitable for routine diagnostics when LC-MS/MS is unavailable.

## Abstract

Acyclovir and ganciclovir comprise the prophylaxis and treatment of herpesvirus and cytomegalovirus infections occurring in immunocompromised patients. Their therapeutic drug monitoring is fundamental because of interindividual variability leading to side effects and drug resistance and is performed through several techniques, such as liquid chromatography coupled with UV spectrophotometry (HPLC-UV) or mass spectrometry (LC-MS/MS). Therefore, we developed and validated a low-cost, non-time-consuming, and low-sample-consuming HPLC-UV method. Briefly, 100 µL of sample was used for sample preparation, mainly consisting of precipitation through organic solvent. In total, 20 µL was injected into the instrument. Chromatographic separation was obtained eluting mobile phases A (10 mM ammonium formiate 0.01% formic acid) and B (acetonitrile) on a Poroshell 120 SB-C8 2.1 × 150 mm, 2.7 µm for 12 min isocratically (97:3; A:B) at a flow rate of 0.2 mL/min. The linearity range (0.5–40 mg/L) of the method allowed us to quantify both the Cmin and Cmax of acyclovir and ganciclovir. Plasma concentrations measured on a small cohort of patients undergoing acyclovir (31) and ganciclovir (9) treatment by the proposed method and the LC-MS/MS methods, already in use, were significantly correlated. The proposed HPLC-UV method may be implemented in diagnostics as an alternative method in case of the unavailability of the LC-MS/MS system.