A simple RP-HPLC method for determination of nitisinone in pharmaceuticals

Zana Mustafa,Zorica Arsova-Sarafinovska,Maya Shishovska,Aleksandra Grozdanova

doi:10.33320/10.33320/maced.pharm.bull.2019.65.02.007

Abstract

Nitisinone is a reversible inhibitor of 4-hydroxyphenylpyruvate dioxygenase and an active substance in the orphan drug used for the treatment of hereditary tyrosinemia type I, a rare genetic disease caused by mutations in the gene for the enzyme fumarylacetoacetase. The aim of our study was to develop a simple and accurate RP–HPLC method with UV detection for routine determination of nitisinone in commercially available pharmaceutical forms. The chromatographic separation was achieved on a reversed-phase column Purospher STAR® RP–8 end–capped (150 x 4.6 mm i.d., particle size 5 μm), with a mobile phase consisted of a mixture of acetonitrile and water acidified with o-phosphoric acid with pH adjusted to 3.0, 65:35 (V/V), filtered through 0.45μm nylon filter. The flow rate was kept at 1 mL/min. A diode array detector measured the UV absorbance at 272 nm. The injection volume was 10 μL. The method was validated by a determination of system suitability, specificity, linearity, range, accuracy, precision, detection limit and quantitation limit. Then, the method was applied for determination of nitisinon in commercially available capsules. The proposed RP-HPLC method allows a simple, accurate, precise and rapid determination of nitisinone in pharmaceuticals. The advantages of the method include simple sample treatment, good precision (RSD less than 2%) and high recovery (greater than 99%). The method could be recommended for routine analysis in quality control laboratories, in stability studies as well as for the evaluation of potentially counterfeit capsules containing nitisinone. Key words: Nitisinone; High performance liquid chromatography; Method validation; Hereditary tyrosinemia type I

Highlights

Hereditary tyrosinaemia type 1 (HT-1) is a rare genetic disease caused by mutations in the gene for the enzyme fumarylacetoacetase
Nitisinone (NTBC) is a reversible inhibitor of 4hydroxyphenylpyruvate dioxygenase and an active substance in the orphan drug used for the treatment of hereditary tyrosinemia type I (De Laetet al., 2013)
The mobile phase consisted of a mixture of acetonitrile and water acidified with o-phosphoric acid with pH adjusted to 3.0, 65:35 (V/V) and the reversedphase column Purospher STAR® RP–8 end–capped (150 x 4.6 mm i.d., particle size 5 μm), was found optimal for isocratic determination of NTBC in pharmaceuticals

Summary

Introduction

Hereditary tyrosinaemia type 1 (HT-1) is a rare genetic disease caused by mutations in the gene for the enzyme fumarylacetoacetase. It usually presents with liver failure it can be manifest as chronic liver disease. It is considered that there is a significant lifetime risk of developing hepatocellular carcinoma. Before the Nitisinone (NTBC) is a reversible inhibitor of 4hydroxyphenylpyruvate dioxygenase and an active substance in the orphan drug used for the treatment of hereditary tyrosinemia type I (De Laetet al., 2013). The chemical name of nitisinone is 2-(2-nitro-4trifluoromethyl benzoyl)-1,3-cyclohexanedione (Fig. 1), with empirical formula C14H10F3NO5, and molecular weight is 329.228 g/mol.

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