Indirect resolution of baclofen enantiomers from pharmaceutical dosage form by reversed-phase liquid chromatography after derivatization with Marfey's reagent and its structural variants.
A high-performance liquid chromatographic (HPLC) method was developed for chiral assay of baclofen enantiomers in pharmaceutical formulations using an indirect approach. Baclofen enantiomers were derivatized with Marfey's reagent (FDNP-L-Ala-NH2) and its structural variants FDNP-L-Phe-NH2, FDNP-L-Val-NH2, FDNP-L-Leu-NH2 and FDNP-L-Pro-NH2. The resultant diastereomers were separated on RP-TLC [triethylammonium phosphate buffer (pH 4.0, 50 mm)-acetonitrile, 50:50] and on a C18 column using a linear gradient (45 min) of acetonitrile and 0.01% aqueous trifluoroacetic acid (TFA) with UV detection at 340 nm. The differences in the retention times (Delta t R) of diastereomers due to the five chiral reagents were compared. The maximum and minimum difference in retention times between separated diastereomers was for FDNP-L-Leu-NH2 and FDNP-L-Pro-NH2, respectively. The effect of flow rate, acetonitrile content and TFA concentration on resolution was studied. The method was validated for linearity, repeatability, limit of detection and limit of quantification.
Department of Chemistry, Indian Institute of Technology Roorkee, Roorkee, India. rbushfcy@iitr.ernet.in
Ravi
R
Bhushan
Virender
V
Kumar
D000409:Alanine / Q000031:analogs & derivatives* / Q000737:chemistry; D001418:Baclofen / Q000737:chemistry / Q000302:isolation & purification*; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D002855:Chromatography, Thin Layer; D004136:Dinitrobenzenes / Q000737:chemistry*; D004304:Dosage Forms; D018755:GABA Agonists / Q000737:chemistry / Q000302:isolation & purification*; D012015:Reference Standards; D015203:Reproducibility of Results; D013237:Stereoisomerism
D004136:Dinitrobenzenes; D004304:Dosage Forms; D018755:GABA Agonists; C047134:Marfey's reagent; D001418:Baclofen; D000409:Alanine