Application of porous layer open tubular capillary columns with narrow inner diameter in nanoflow-high performance liquid chromatography for N-derivatized amino acid enantiomers

doi:10.3724/SP.J.1123.2019.09002

Abstract

Abstract:

Porous layer open tubular capillary columns with a narrow inner diameter (NPLOT) have potential in life science, especially in single-cell analysis. In this work, quinidine-based chiral NPLOT columns were prepared by in-situ thermal-initiated polymerization. The porous layer of the organic polymer was prepared with a 6 μm i. d capillary by in-situ thermal-initiated polymerization, following which the thickness and morphology of the NPLOT column were observed for different durations (3 h, 6 h, and 9 h) of polymerization. The thicknesses of the porous layer were 103±51 nm and 210±51 nm when polymerization was conducted for 3 h and 6 h, respectively. Subsequently, the columns prepared under 3 h of polymerization were applied in nanoflow-high performance liquid chromatography for the enantioseparation of N-derivatized amino acid enantiomers. Baseline separation was achieved in 2 min. The sample volume consumed was of the picoliter level, which would provide a means of research for single-cell analysis.

Key words: porous layer open tubular columns with narrow inner diameter (NPLOT), in-situ thermal-initiated polymerization, nanoflow-high performance liquid chromatography (nano-HPLC), enantioseparation

LI Ruonan, WANG Lijuan, ZHANG Dongtang, WANG Yanan, GUO Guangsheng, WANG Xiayan. Application of porous layer open tubular capillary columns with narrow inner diameter in nanoflow-high performance liquid chromatography for N-derivatized amino acid enantiomers[J]. Chinese Journal of Chromatography, 2020, 38(3): 350-355.

Figures/Tables 6

Fig. 1 Scheme of nano-HPLC-UV detection system PLOT: porous layer open tubular; DAQ: data acquisition.

Fig. 2 SEM images of poly(MQD-co-HEMA-co-EDMA) PLOT column prepared by in-situ thermal-initiated polymerization for different durations a. bare capillary; b. 3 h; c. 6 h; d. 9 h.

Fig. 3 SEM image of porous layer in poly(MQD-co-HEMA-co-EDMA) PLOT columns with narrow inner diameter by heating for 3 h

Fig. 4 Chromatograms of N-derivatized amino acid enantiomers Chromatographic column: poly(MQD-co-HEMA-co-EDMA) PLOT column, 6 μm i.d., total length 44.60 cm, effective length 40.70 cm; polymerization time: 3 h; sampling condition: 1.38 MPa, 20 s; sample concentration: 1.0 g/L; separation condition: 6.89 MPa; mobile phase flow rate: 32.24 nL/min.

Fig. 5 Chromatograms of N-derivatized amino acid enantiomers Sample: a. 10.0 g/L 3, 5-DClB-DL-Val; b. 10.0 g/L 3, 5-DClB-DL-Trp; c. complex of 5.0 g/L 3, 5-DClB-DL-Trp and 5.0 g/L 3, 5-DClB-DL-Val. Chromatographic column: poly(MQD-co-HEMA-co-EDMA) PLOT column, 6 μm i.d., total length 59.40 cm, effective length 54.20 cm; polymerization time: 3 h; sampling condition: 1.38 MPa, 20 s; separation condition: 1.38 MPa; mobile phase flow rate: 5.00 nL/min.1. methanol; 2.3, 5-DClB-L-Val; 3. 3, 5-DClB-L-Trp; 4. 3, 5-DClB-D-Val; 5. 3, 5-DClB-D-Trp.

Table 1 Resolutions, plate numbers, retention factors, and enantioselectivity of N-derivatized amino acid enantiomers

Sample	R	N_L/m	N_D/m	k_L	k_D	α
m-ClB-DL-Met	1.55	601	404	1.17	1.89	1.61
m-ClB-DL-Ala	1.52	925	1185	1.26	1.73	1.38
3, 5-DMB-DL-Leu	1.25	618	302	0.77	1.27	1.65

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