Determination of Oseltamivir Quality by Colorimetric and Liquid Chromatographic Methods

Abstract

We developed a colorimetric and chromatographic assay for oseltamivir to assess the authenticity of Tamiflu (F. Hoffmann-La Roche Ltd., Basel, Switzerland) because of a growing concern about counterfeit oseltamivir. The colorimetric assay is quantitative and relies on an extractable colored ion-pair complex of oseltamivir with Congo red or bromochlorophenol blue. The reverse-phase chromatographic assay uses an alkaline mobile phase with UV detection. Both methods were evaluated for variability and selectivity and subsequently applied to batches of oseltamivir products acquired through the Internet. The Congo red test showed greater assay sensitivity, linearity, and accuracy. Colorimetric and chromatographic analysis showed all batches of oseltamivir product were within ±15% of the stated amount of active ingredient. The antiviral drug oseltamivir phosphate has been recommended by Centers for Disease Control and Prevention as an adjunct in the effective treatment and prevention of influenza. Oseltamivir and zanamivir are both approved by the Food and Drug Administration (FDA) for use in controlling both influenza A and B viruses (1). Oseltamivir phosphate is the active ingredient in Tamiflu (F. Hoffmann-La Roche Ltd., Basel, Switzerland) and is available in capsule and powder form. The specter of an avian influenza pandemic has given Tamiflu much notoriety, and anticipation of the potential public health threat has prompted a demand for the product. Consequently, criminal elements have already begun to produce counterfeit Tamiflu. Recently, US Customs agents seized counterfeit Tamiflu entering the United States (2). The bogus products, which contained vitamin C and lacked the active ingredient of oseltamivir phosphate, had been purchased through the Internet. Although these shipments were quickly detected by a joint effort of the FDA and US Customs and Border Protection, these products would easily have gone unnoticed in developing countries, where insufficient resources and infrastructure hamper the ability to monitor and preserve drug quality. WHO estimates that up to 25% of the medicines consumed in developing countries are counterfeit or substandard (3). Simple and affordable colorimetric assays provide a practical means to rapidly monitor drug quality in resource-poor areas. Because oseltamivir phosphate (Figure 1) possesses amine groups, the protonated form may act as a cationic site for anionic dyes such as Congo red and bromochlorophenol blue to produce colored ion-pairing complexes. Congo red has been used in colorimetric determinations of chitosan and poly (N-vinyl-2-pyrrolidone) while bromophenol blue has been used i