Straightforward thin-layer chromatography–densitometric method for the determination of phyllanthin in Phyllanthus niruri from different phytogeographical zones

Abstract

Phyllanthus niruri contains various lignan compounds, whose concentrations vary depending on several factors. This study was intended to determine the phyllanthin content of P. niruri obtained from various locations in Indonesia by using thinlayer chromatography (TLC)-densitometry to evaluate the effect of geographical factors on their quality. The TLC system comprised silica gel 60 F 254 for the stationary phase, toluene-ethyl acetate-formic acid (15:10.5:1.5, V/V) for the mobile phase, and documentation under ultraviolet (UV) 254 nm light without chemical reagents. This developed method meets the specificity requirement, as marked by the identical UV spectrum between the phyllanthin sample and the standard (λ max = 279 and 230 nm). Further, it shows good linearity for phyllanthin concentrations in the range of 2.36-11.8 µg/band (r = 0.9924), with LOD 0.532 µg/band and LOQ 1.612 µg/band. It also has good intraday and interday precision, as indicated by RSD of 8.87-9.43 and 6.94%, respectively. Eight of the 15 analyzed samples (collected from Batu, Blitar, Kediri, Nganjuk, Jember, Mojokerto, Banyuwangi, and Surabaya) contained only a trace amount of phyllanthin. In contrast, the other seven had varying levels of phyllanthin (1.376-4.130 mg/g dried herbs). Using the Tawangmangu sample as the reference, these seven samples can be grouped into two: significantly lower phyllanthin contents (Tulungagung) and very significantly lower phyllanthin contents (Lumajang, Bangkalan, Pasuruan, Sidoarjo, and Gresik). It can be concluded that TLC-densitometry designed in this research is a straightforward method that, at the same time, meets the validation parameters. Therefore, it can be repeated to analyze phyllanthin in P. niruri of different phytogeographical origins.