Stress-Induced Analysis of Piperine Variability in Different Marketed Black Pepper Powders and Seeds by a Validated High-Performance Thin-Layer Chromatography Method

Summary

In this article, a simple and sensitive stress-induced high-performance thin-layer chromatography (HPTLC) method is proposed for the quantitative analysis of piperine (a bioactive marker) in the alcoholic extracts of five different marketed powders (P1—P5) and dried seeds (P6) of black pepper (Piper nigrum). HPTLC was performed on glass-backed silica gel 60 F₂₅₄ HPTLC plates with toluene, ethyl acetate, and glacial acetic acid (7:3:0.5; v/v) as the mobile phase. Scanning and quantification of the developed HPTLC plate (λ = 280 nm) furnished compact and intense spots of piperine at R _F = 0.40 ± 0.01. For the proposed method, the estimated regression equation (Y) and r ² were 9.473X + 2262.23 and 0.997, respectively, in the range of 100–900 ng, the limit of detection (LOD) and limit of quantification (LOQ) were 19.63 and 59.47 ng, respectively. The quantified piperine values (% w/w) in P1, P2, P3, P4, P5, and P6 were 1.75, 2.71, 1.21, 1.67, 2.45, and 1.80, respectively. In the stress study, the treatment of piperine with acid, alkali, and hydrogen peroxide showed its complete degradation. On the other hand, there was no effect of dry or moist heat, and photochemical and ultraviolet light (λ = 254 nm) on the stability of piperine. The present maiden report on a validated HPTLC method for the stress induced analysis of piperine, therefore, would help in the selection of good-quality marketed products. Furthermore, the developed method may help in selecting or rejecting the in-process procedural conditions for making the products and formulations more efficacious and safe.