Rapid Determination of Anthocyanins in Bilberries

Bilberry (Vaccinium myrtillus L.) extracts are high in anthocyanin content and are widely used in nutritional supplements for improving visual acuity and treating circulatory disorders. Bilberries cannot be cultivated and are hard to harvest and process, which makes them one of the most expensive botanical ingredients resulting in a high susceptibility to adulteration. A reliable analytical method is needed to determine anthocyanins in nutraceuticals to ensure their efficacy. This paper describes a sensitive, fast, and accurate UHPLC method to determine 15 anthocyanins and 5 anthocyanidins in a bilberry-based nutritional supplement with a run time of < 30 min.

Experimental

A Thermo Scientific Dionex UltiMate™ 3000 Rapid Separation LC system was used for this study. For individual components of the system and preparation of solutions, refer to Thermo Scientific Dionex Application Note 281 (1).

Figure 1: Column: Acclaim™ 120, C18, 2.2 μm, Analytical 2.1 × 150 mm. Eluent: A: 10% Formic Acid, B: 10% Formic Acid, 22.5% CH3CN, 22.5% MeOH; gradient: 0.0–12.0 min 9% B, 12.0–25.00 min 35% B, 25.00–30.00 min 50% B, 30.00–35 min 9% B; temp: 35 °C; flow-rate: 0.475 mL/min; detection: Vis 520 nm; injection volume: 2.0 μL.

Results and Discussion

Figure 1 shows the separation of anthocyanins in a bilberry-based nutritional supplement. All 15 anthocyanins and 5 anthocyanidins present in bilberries were detected in this product (see Table I for a list of analytes separated).

Table I: Concentrations of quantified analytes

The reported LODs ranged from 0.20–1.56 μg/mL. Sample recoveries ranged from ranged from 84%–108% suggesting good method accuracy.

Reference

(1) Dionex Corporation, "Fast Determination of Anthocyanins in Bilberries" Application Note 281, LPN 2848: Sunnyvale, CA.

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