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.
Bhushan
Ravi
R
Department of Chemistry, Indian Institute of Technology Roorkee, Roorkee, India. rbushfcy@iitr.ernet.in
Kumar
Virender
V
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