Development of a Simple, Rapid, and Robust Isocratic Liquid Chromatographic Method for the Determination of Pyrimethamine and its Synthetic Impurities in Bulk Drugs and Pharmaceutical Formulations
Pyrimethamine is an important antiparasitic drug in the treatment of malaria and toxoplasmosis and is often used in combination with either sulfadoxine, sulfalene, or sulfadiazine. Determining the content of pyrimethamine and investigating the related substances is currently possible applying either a compendial monograph utilizing thin layer chromatography as well as liquid chromatographic methods used by the respective manufacturers. To provide a simple method which is capable of determining the content of pyrimethamine and of resolving four of its potential synthetic impurities a very simple, cheap, precise, and accurate isocratic RP-HPLC method was developed. All analytes can be separated within a total runtime of 30 min and the method was linear within the concentration ranges of 0.12–0.740, 0.104–0.621, 0.120–0.710, 2.0–11.8, and 1.01–5.80 µg mL−1 for pyrimethamine, impurity A, impurity B, impurity C, and impurity D, respectively. These substances were separated by employing a Eurospher-II C18H column (250 × 4.6 mm, 5 µm particle size), a mobile phase being a mixture of a 0.05 M KH2PO4 buffer solution (pH 2.6) and methanol in the ratio 40:60 (v/v). The analysis was carried out at 30 °C, applying a flow rate of 1.2 mL min−1, and a detection wavelength of λ = 215 nm. The coefficients of determinations (R 2) for the five analytes were greater than 0.994 for pyrimethamine and all impurities. Results of recovery studies were within the range of 89.1–105.1% for all substances. In all tested genuine batches of pyrimethamine raw material impurities within the specified limits were present which is concurrent with results obtained from using the present manufacturer’s method.
KeywordsPyrimethamine RP-HPLC Quality control Bulk drug and tablet analysis Malaria Impurity profiling
- B. P.
The European Directorate for the Quality of Medicines and HealthCare
High-performance thin layer chromatography
Catholic Academic Exchange Service
- Ph. Eur.
- Ph. Int.
Reversed-phase high-performance liquid chromatography
Tanzania Food and Drugs Authority
Thin layer chromatography
United States Pharmacopoeia
Thanks to the KAAD (Catholic Academic Exchange Services, Bonn, Germany) for awarding scholarship to Yonah Hebron Mwalwisi, to the European Directorate for the Quality of Medicines and HealthCare (EDQM) for the provision of samples and reference substances, the Tanzania Food and Drugs Authority (Dar es salaam, Tanzania) for facilitating the research stay of Yonah H. Mwalwisi at the Institute of Pharmacy and Food Chemistry at Würzburg, and Christine Erk (Institute for Pharmacy and Food Chemistry, Würzburg) for LC/MS support.
YHM, LH, and UH designed the study, YHM performed sample preparation and analysis, data collection, data analysis and interpretation. YHM and LH drafted the manuscript. UH and LH supervised the study and contributed in data interpretation. UH, LH, and EK reviewed of the manuscript. All authors have read and approved the final manuscript.
Compliance with Ethical Standards
Conflict of interest
The authors declare that they have no competing interests.
This work was funded by the Catholic Academic Exchange Services, Bonn, Germany (KAAD) by providing a scholarship to YHM.
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