Genotoxic Impurity Detection In Acyclovir And Acyclovir 2/3 Hydrate Preparation Study

Acyclovir is an important antiviral drug, which is possessing antiviral activity and commonly used in the treatment of variety of infections caused by herpes simplex virus. During Acyclovir quality study, it is found that a genotoxic impurity, p-toluene sulphonic acid, is used in Acyclovir production as a catalyst, which should be controlled less than 3pm in Acyclovir. Thus it’s necessary to develop a sensitive and reproducible method to detect p-toluene sulphonic acid content in Acyclovir. Another concern is the Acyclovir polymorph issue. Acylcovir has six polymorphs, in which acyclovir 2/3 hydrate has better bioavailability and clinical effect than others. It is also meaningful to do some research work on Acyclovir 2/3 hydrate preparation process. The research work of this thesis has two parts:The first section of this thesis is the analytical method development to determine p-toluene sulphonic acid content in Acyclovir. P-toluene sulphonic acid in Acyclovir is enriched by SPE column and then detected by HPLC quantitively. The influential factors of solid phase extraction parameters such as SPE column type, sampling flow rate, concentration of eluant and elution flow rate etc are investigated and the optimum process is obtained0.1g of Acyclovir is dissolved into 10 mL hydrochloric acid at the concentration of 10-4M. Acyclovir solution is loaded into Cleanert SAX column at speed of 1.0mL/min. And then wash the column with hydrochloric acid at the concentration of 10-4M until Acyclovir in column is removed completely. Finally, p-toluene sulphonic acid is eluted by 1M of HCl solution from SPE column and diluted to 10 mL. The collected p-tolunene sulphonic acid solution will be detected by HPLC in the following step. HPLC method operational parameters are optimized and a method is developed as follows: Inertsil O DS-SP C18 column is used at the temperature of 30℃, while mobile phase is Acetonitrile-0.1% phosphoric acid solution in the ratio of 15:85(v/v). Flow rate is kept at 1.0mL/min and the elution is monitored at UV 222 nm. The method for p-tolune sulphonic acid enrichment and detection is validated on specificity, repeatability, linearity, sensitivity, accuracy and robustness etc. Its method validation results show this method is reproducible, sensitive and robustness and easy to operate, which could be good for p-toluene sulphonic acid determination in acyclovir.Another concern is on Acyclovir 2/3 hydrate preparation. Acyclovir 2/3 hydrate is prepared by cooling crystallization, anti-solvent crystallization and reactive participation methods in the initial stage. It was found that the TG and PXRD data of Acyclovir obtained by both cooling crystallization and anti-solvent crystallization methods cannot match the reported data of Acyclovir 2/3 hydrate. While the FTIR, TG and PXRD data of Acyclovir prepared by reactive participation matches to that of Acyclovir 2/3 hydrate well. Acyclovir is dissolved in sodium hydroxide solution to get the Acyclovir sodium salt, then the solution is neutralized by hydrochloric acid and Acylcovir 2/3 hydrate is precipitated and crystallized. The parameters of Acyclovir 2/3 hydrate preparation process are optimized such as the concentration of sodium hydroxide and hydrochloric acid, hydrochloric acid feeding speed, crystallization temperature and stirring rate etc and the optimum process is obtained as follows: Acyclovir is dissolved in 1.25 M sodium hydroxide, and then neutralized by 1.0M of Hydrochloric acid with the feeding speed of 2mL/min. The solution temperature is controlled at 10℃ and the stirring rate at 200 rpm during neutralization. After neutralization, keep the solution for 3h without stirring. Acyclovir 2/3 hydrate is precipitated, filtered and dried at 50℃ for 20 h to obtain Acyclovir 2/3 hydrate. The product yield is 93% and this preparation process can be scaled up in production. Acyclovir 2/3 hydrate is kept under 40±2℃ for 6 months and its PXRD a nd HPLC data show it is stable. Finally, crude Acylcovir with high impurities is used for Acylcovir 2/3 hydrate preparation. Its TG and PXRD data confirm the product prepared is Acylcovir 2/3 hydrate, but its related substance is still out of specification. Thus, Acylovir crude needs to be purified firstly when it is used to prepare Acylovir 2/3 Hydrate.