Resolution and isolation of enantiomers of (±)‐isoxsuprine using thin silica gel layers impregnated with l‐glutamic acid, comparison of separation of its diastereomers prepared with chiral derivatizing reagents having l‐amino acids as chiral auxiliaries

Abstract

Thin silica gel layers impregnated with optically pure l-glutamic acid were used for direct resolution of enantiomers of (±)-isoxsuprine in their native form. Three chiral derivatizing reagents, based on DFDNB moiety, were synthesized having l-alanine, l-valine and S-benzyl-l-cysteine as chiral auxiliaries. These were used to prepare diastereomers under microwave irradiation and conventional heating. The diastereomers were separated by reversed-phase high-performance liquid chromatography on a C18 column with detection at 340 nm using gradient elution with mobile phase containing aqueous trifluoroacetic acid and acetonitrile in different compositions and by thin-layer chromatography (TLC) on reversed phase (RP) C18 plates. Diastereomers prepared with enantiomerically pure (+)-isoxsuprine were used as standards for the determination of the elution order of diastereomers of (±)-isoxsuprine. The elution order in the experimental study of RP-TLC and RP-HPLC supported the developed optimized structures of diastereomers based on density functional theory. The limit of detection was 0.1-0.09 µg/mL in TLC while it was in the range of 22-23 pg/mL in HPLC and 11-13 ng/mL in RP-TLC for each enantiomer. The conditions of derivatization and chromatographic separation were optimized. The method was validated for accuracy, precision, limit of detection and limit of quantification.

Chirality of β2-agonists. An overview of pharmacological activity, stereoselective analysis, and synthesis

“…Chiral derivatizing reagents prepared by substitution of the fluorine atoms in 1,5-difluoro-2,4-dinitrobenzene (DFDNB) with three amino acids: L-alanine, L-valine, and (S)-benzyl-L-cysteine cysteine were used for enantioresolution of racemic β 2 -agonist isoxsuprine on reversedphase HPLC and reversed-phase thin-layer chromatography [117].…”

“…TLC served for the resolution of enantiomers of isoxsuprine using thin silica gel layers impregnated with L-glutamic acid. The limit of detection of each enantiomer was 11-13 ng/mL [117].…”

Assessment and application of Marfey’s reagent and analogs in enantioseparation: a decade’s perspective

“…ii. The structures optimized for the lowest energy, developed by the same software as mentioned above, of the two diastereomeric derivatives of (±)-isoxsuprine prepared with the FDNP reagent having S-benzyl-L-cysteine (Bhushan & Nagar, 2015) iii. On the similar lines it was interpreted that the L-L diastereomer was retained for lesser time in comparison with the L-D diastereomer of SeMet prepared with FDNP-L-Met .…”

“…The lowest energy optimized structures (Figure 7) of the two diastereomeric derivatives (of orciprenaline, for example) showed that in the [( S )‐ l ]‐diastereomer, the hydrophobic groups ‐CH 2 SCH 3 on the stereogenic center of CDR (having S ‐methyl‐ l‐ cysteine) and ‐C 6 H 3 (OH) 2 on stereogenic center of orciprenaline considered to be cis to each other and in the [( R )‐ l ]‐diastereomer, the same two groups have trans ‐type arrangement. The diastereomeric derivative with a cis ‐type arrangement (which is more hydrophobic) interacts more strongly with ODS material of the C 18 column than the trans ‐type arrangement, hence it has a longer retention time. The structures optimized for the lowest energy, developed by the same software as mentioned above, of the two diastereomeric derivatives of (±)‐isoxsuprine prepared with the FDNP reagent having S ‐benzyl‐ l‐ cysteine (Bhushan & Nagar, 2015) supported the cis ‐type arrangement of the hydrophobic groups, i.e. ‐CH 2 SC 6 H 5 on the stereogenic center of CDR and ‐CH 2 OC 6 H 5 present on the stereogenic center of isoxsuprine, respectively, in [(−)‐ l ]‐diastereomer, and trans ‐type arrangement in the [(+)‐ l ]‐diastereomer. On the similar lines it was interpreted that the l ‐ l diastereomer was retained for lesser time in comparison with the l‐ D diastereomer of SeMet prepared with FDNP– l‐ Met (Nagar & Bhushan, 2014).…”

“…Bhushan and Nagar (2015) synthesized FDNP reagents having S-benzyl-L-cysteine (SBLC), L-Val and L-Ala, as chiral auxiliaries by stirring the reaction mixture (10 ml of 0.1 M solution of SBLC, for example) and 10 ml of 0.2 M solution of DFDNB in acetone-water (6:4, v/v) at 40 C in a round-bottom flask for 1 h. It was kept at room temperature for 5 min and then 20 ml of water was added followed by addition of HCl (10 ml, 1 M) to induce crystallization. The FDNP reagents were characterized and were used to prepare diastereomeric derivatives of (±)-isoxsuprine (Figure 1j) under MWI (for 80 s at 75% of 800 W) and conventional heating (at 45 C for 1 h with constant stirring).…”