Separation and quantification of lignans in Phyllanthus species by a simple chiral densitometric method

Abstract

A sensitive, selective, and robust high-performance TLC (HPTLC) method using chiral TLC plates for qualitative and quantitative analysis of phyllanthin (A), hypophyllanthin (B), niranthin (C), and nirtetralin (D), the active lignans of Phyllanthus species, was developed and validated. The effectiveness and role of various stationary phases viz TLC silica gel 60F(254), HPTLC silica gel 60F(254), and chiral TLC plates in the quantitation were evaluated. A precoated chiral TLC plate was found suitable for the simultaneous analysis of four pharmacologically active lignans. For achieving good separation, the optimized mobile phase of n-hexane/acetone/1,4-dioxane (9:1:0.5 by volume) was used (R(f) = 0.30, 0.36, 0.41, and 0.48 for compounds A, B, C, and D, respectively). A densitometric determination of the above compounds was carried out in reflection/absorption mode at 620 nm. Optimized chromatographic conditions provide well-separated compact bands for the tested lignans. The calibration curves were found linear in the concentration range of 100-500 ng/band. Recoveries of A-D were 99.98, 100.51, 99.22, and 98.74%, respectively. The method was validated according to ICH guidelines. The method reported here is reproducible and applied for the quantitative analysis of the above lignans in the leaves of four Phyllanthus species, i. e., P. amarus, P. maderaspatensis, P. urinaria, and P. virgatus.

Phyllanthus amarus: Ethnomedicinal uses, phytochemistry and pharmacology: A review

“…The main active constituents of P. amarus are lignans (phyllanthin, hypophyllanthin, nirurin niranthin, phyltetralin, niranthine, nirtetralin etc. (Morton, 1981;Chevallier, 2000;Srivastava et al, 2008;Kassuya et al, 2006;Huang et al, 2003;Maciel et al, 2007;Singh et al, 2009), flavonoids (quercetin, quercetrin, rutin, gallocatechin, phyllanthusiin, kaempferol etc. ), (Foo and Wong, 1992;Foo, 1993a;Londhe et al, 2008;Morton, 1981), Ellagitannins viz.…”

“…Secondary metabolites Phyto-constituents References 1 Lignans Phyllanthin, hypophyllanthin, niranthin, phyltetralin, nirtetralin, isonirtetralin, hinokinin Morton (1981), Sharma et al (1993), Chevallier (2000), Srivastava et al (2008), Kassuya et al (2006), Huang et al (2003), Singh et al (2009) Lintetralin, isolintetralin, demethylenedioxy-niranthin, 5-demethoxy-niranthin Maciel et al (2007) (3-(3,4-dimethoxy-benzyl)-4-(7-methoxybenzo[1,3]dioxol-5-yl-methyl)-dihydrofuran-2-one, 4-(3,4-dimethoxy-phenyl)-1-(7-methoxybenzo[1,3]dioxol-5-yl)-2,3-bismethoxymethyl-butan-1-ol Singh et al (2009) 2…”

“…Optimized chromatographic conditions provided well separated compact bands for the tested lignans. The calibration curves were found linear in the concentration range of 100-500 ng/band (Srivastava et al, 2008). Sharma et al (1993) have developed a reversed phase HPLC procedure for standardizing P. amarus on the basis of its two bioactive lignans, phyllanthin and hypophyllanthin.…”

Development of a UHPLC–MS/MS method for the quantitation of bioactive compounds in Phyllanthus species and its herbal formulations

“…The quantitative analysis of bioactive lignans has been carried out in Phyllanthus species by traditional methods such as high-performance thin-layer chromatography (HPTLC) and HPLC [13][14][15][16][17][18][19][20][21][22][23][24][25]. Several researchers have reported fingerprinting, identification and characterization of bioactive compounds in crude extracts of Phyllanthus species using LC-MS [24][25][26][27][28][29][30][31][32][33][34].…”

Hplc–MS/MS Method for The Determination of Four Lignans from Phyllanthus Urinaria L. in Rat Plasma and its Application

“…High-performance thin layer chromatography (HPTLC) [14], chiral TLC [15], HPLC-UV [16], HPLC-fluorescence [17] and HPLC-MS [18] methods have been developed for the quantitation of lignans in the genus Phyllanthus (P. niruri, P. urinaria, P. amarus and so on). A HPLC-fluorescence method has been used for determination of four lignans (hypophyllanthin, phyllanthin, phyltertralin and niranthin) from Phyllanthus niruri L. in vivo for a PK study [19].…”