Dissolution/diffusion-based Injection for Fast Separation of Epinephrine and Norepinephrine by Short Capillary Electrophoresis†

doi:10.7503/cjcu20150156

Abstract

Abstract:

Shortening capillary is an effective strategy to speed up capillary electrophoresis(CE), but a very fine injection approach must be availableto obtain an ultra-short initial zone which is remains a big challenge. To conquer it, an easy injection approach called dissolution/diffusion-based injection(DDI) was explored simply based on the properties of an analyte able to dissolve and diffuse into a solution. DDI was easily operated by letting the capillary inlet touch either a sample solution or liquid-dried film, giving initial sample zones <15 μm. The resulted DDI-CE is reproducible, with relative standard deviation of peak time below 1.6%, and was shown to be applicable to the fast(40 s) separation of epinephrine and norepinephrine mixtures through only 3 cm effective length under 400 V/cm. Its applicability was further validated by quantification of epinephrine injection.

Key words: Micrometer-zonal injection, Fast short capillary electrophoresis, Epinephrine, Amino acid, Separation

CLC Number:

O657

TrendMD:

HU Can, CHEN Yi. Dissolution/diffusion-based Injection for Fast Separation of Epinephrine and Norepinephrine by Short Capillary Electrophoresis^†[J]. Chem. J. Chinese Universities, 2015, 36(9): 1681.

Figures/Tables 6

Fig.1 Setup of DDI-CE1. Separation capillary; 2. detection window; 3. grounded electrode reservoir; 4. platinum electrodes; 5. DC power supply; 6. high voltage electrode reservoir; 7. cleaning solution vial; 8. sample vial; 9. slit opening(ca.1 mm); 10. longitudinal slider; 11. cross slider; 12. sample plate and holder; 13. sample solution membrane or dried trace; 14. micro-objective of inverted fluorescence microscope.

Fig.2 Photos of sample solution(A1, A2) and dried film(B1, B2), and electropherograms via DDI for about 0.2 s(C)(A1) shows a solution film formed in a 1 cm2 well with 1 μL of 0.1 mmol/L FITC,(B1) its dried trace,(A2) and(B2) their corresponding DDI zones, respectively. (C) a. dried film; b. solution film; c. spontaneous injection; d. electrokinetic injection. The electropherograms were obtained after separation at 400 V/cm, through 1 cm capillary filled with 20 mmol/L borax(pH=9.5).

Fig.3 Resolution decreases with initial sample zone length and increases with separation lengthInjection time: a. 0.2 s; b and d. 1 s; c and e. 3 s; capillary: a—c. 50 μm i.d.×3 cm; d. 4 cm; e. 6 cm; sample: 50 μmol/L FITC-labeled E and NE; running buffer: 60 mmol/L borax at pH=10. Other conditions are the same as in Fig.2.

Fig.4 Successive DDI-CE of FITC dryness(A1—A3) and solution(B1—B3) membranes, respectivelyOther conditions are the same as in Fig.2. c(FITC)/( μmol·L-1): (A1), (B1) 20; (A2), (B2) 50; (A3), (B3) 100.

Fig.5 DDI-CE of standard and real samples(A) together with their working curves(B)(A) Sample state: a. solution film, b. film dried for 1 h, c. film dried for >1 d, d. real sample, all analytes were prepared at 50 μmol/L; (B) injection diluted for 100 times, and labeled by FITC; DDI injection time: 0.2 s; separation distance: 3 cm. Other conditions are the same as in Fig.2.

Fig.6 DDI-CE of amino acid mixturesSample: 50 μmol/L FITC-labled Arg, Leu, Phe, Ala, Gly, Glu and Asp. Capillary: 50 μm(i.d.)×6 cm;buffer: 20 mmol/L borax at pH=9.2. Other conditions are the same as in Fig.2.

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