Chiral analytical method development of liquiritigenin with application to a pharmacokinetic study

Abstract

Pharmacometric characterization studies of liquiritigenin have historically overlooked its chiral nature. To achieve complete characterization, an analytical method enabling the detection and quantification of the individual enantiomers of racemic (±) liquiritigenin is necessary. Resolution of the enantiomers of liquiritigenin was achieved using a simple high-performance liquid chromatographic method. A Chiralpak® ADRH column was employed to perform baseline separation with UV detection at 210 nm. The standard curves were linear ranging from 0.5 to 100 μg/mL for each enantiomer. Limit of quantification was 0.5 μg/mL. The assay was applied successfully to stereoselective serum disposition of liquiritigenin enantiomers in rats. Liquiritigenin enantiomers were detected in serum as both aglycones and glucuronidated conjugates. Both unconjugated enantiomers had a serum half-life of ~15 min in rats. The volume of distribution (Vd) for S- and R-liquiritigenin was 1.49 and 2.21 L/kg, respectively. Total clearance (Cltotal) was 5.12 L/h/kg for S-liquiritigenin and 4.79 L/h/kg for R-liquiritigenin, and area under the curve (AUC0-inf) was 3.95 μg h/mL for S-liquiritigenin and 4.23 μg h/mL for R-liquiritigenin. The large volume of distribution coupled with the short serum half-life suggests extensive distribution of liquiritigenin into tissues.

Pharmacological characterization of liquiritigenin, a chiral flavonoid in licorice

“…Although the isolation of stereoisomers is possible with the validated HPLC method (16), the cost resources of isolating them is often a limiting factor to extensive studies in an academic laboratory. In addition, it is a time consuming process, taking between two to six weeks to collect the amount of individual enantiomers necessary for one single assay.…”

“…The limited commercial availability of pure liquiritigenin enantiomers made necessary the manual separation and collection of pure enantiomers using the analytical methods described earlier (16). The collection of the pure enantiomers was achieved following chiral chromatographic separation of multiple injections of racemic liquiritigenin.…”

“…Separation was carried out isocratically at ambient temperature (25 ± 1°C), with a flow rate of 0.6 mL/min and ultraviolet (UV) detection at 210 nm. The analytical column used was a Chiralpak® AD-RH (150 × 4.6 mm i.d., 5 μm particle size, Chiral Technologies Inc., West Chester, USA) and the mobile phase consisted of acetonitrile, water and acetic acid (50:50:0.05 v:v:v) (16). Enantiomeric peaks were collected in sequence using separate 50 mL tubes and were subsequently dried to completion under nitrogen.…”

“…Liquiritigenin (Fig. 1) belongs to the chiral flavanone family (15) and is an important ingredient present in Glycyrrhiza glabra (liquorice) (16) and some Chinese medicinal herbs. Liquiritigenin has been reported as an important bioactive ingredient of the popular Chinese herb Shao Yao Gan Cao Tang for its analgesic and anti-inflammatory properties (17).…”

“…To more thoroughly understand liquiritigenin’s pharmacological activity, as well as its therapeutic and toxic effects, especially in light of the lipophilicity, a stereospecific HPLC method (16) has been developed and validated to separate and isolate liquiritigenin’s pure enantiomers and examine for the first time the stereoselective liquiritigenin in various biological assays. Previous studies have suggested some pharmacological activity for liquiritigenin including anticancer, anti-inflammatory, antidiabetic, and antioxidant activities (19202122).…”