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Influence of Organic Solvents and β-cyclodextrins on Capillary Zone Electrophoresis Separation of Five Biogenic Amines and Two B Vitamins

M. Matuskova
M. Matuskova
, 
I. Cizmarova
I. Cizmarova
, 
P. Chalova
P. Chalova
, 
O. Stefanik
O. Stefanik
, 
A. Horniakova
A. Horniakova
, 
P. Mikus
P. Mikus
 e 
J. Piestansky
J. Piestansky
   | 06 ago 2022
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Article Category: Original Paper
Pubblicato online: 06 ago 2022
Pagine: 43 - 53
Ricevuto: 14 gen 2022
Accettato: 30 mag 2022
DOI: https://doi.org/10.2478/afpuc-2022-0012
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© 2022 M. Matuskova et al., published by Sciendo
This work is licensed under the Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International License.

Figure 1

Optimisation of the background electrolyte (BGE) composition. 1 = thiamine hydrochloride (THI), 2 = tyramine hydrochloride (TYR), 3 = dopamine hydrochloride (DOP), 4 = 5-hydroxytryptamine hydrochloride (5-HT), 5 = noradrenaline hydrochloride (NOR), 6 = adrenaline hydrochloride (ADR), 7 = pyridoxine hydrochloride (PYR). Concentration of the analytes in the sample was 10 μg/mL−1. Applied separation current was 50 μA. For more operation conditions see the part Instrumentation.
Optimisation of the background electrolyte (BGE) composition. 1 = thiamine hydrochloride (THI), 2 = tyramine hydrochloride (TYR), 3 = dopamine hydrochloride (DOP), 4 = 5-hydroxytryptamine hydrochloride (5-HT), 5 = noradrenaline hydrochloride (NOR), 6 = adrenaline hydrochloride (ADR), 7 = pyridoxine hydrochloride (PYR). Concentration of the analytes in the sample was 10 μg/mL−1. Applied separation current was 50 μA. For more operation conditions see the part Instrumentation.

Figure 2

Effect of organic solvent addition into the background electrolyte (BGE) on the simultaneous separation of five biogenic amines and two B vitamins. (a) Addition of methanol (MeOH); (b) addition of acetonitrile (ACN); (c) addition of isopropanol (IP); (d) addition of tetrahydrofuran (THF). BGE = 25 mM GABA + 50 mM HAc + 0.1% mHEC. Concentration of the analytes in the sample was 10 μg/mL−1. Applied separation current was 50 μA. 1 = thiamine hydrochloride (THI), 2 = tyramine hydrochloride (TYR), 3 = dopamine hydrochloride (DOP), 4 = 5-hydroxytryptamine hydrochloride (5-HT), 5 = noradrenaline hydrochloride (NOR), 6 = adrenaline hydrochloride (ADR), 7 = pyridoxine hydrochloride (PYR). For more operation conditions, see the “Instrumentation” section.
Effect of organic solvent addition into the background electrolyte (BGE) on the simultaneous separation of five biogenic amines and two B vitamins. (a) Addition of methanol (MeOH); (b) addition of acetonitrile (ACN); (c) addition of isopropanol (IP); (d) addition of tetrahydrofuran (THF). BGE = 25 mM GABA + 50 mM HAc + 0.1% mHEC. Concentration of the analytes in the sample was 10 μg/mL−1. Applied separation current was 50 μA. 1 = thiamine hydrochloride (THI), 2 = tyramine hydrochloride (TYR), 3 = dopamine hydrochloride (DOP), 4 = 5-hydroxytryptamine hydrochloride (5-HT), 5 = noradrenaline hydrochloride (NOR), 6 = adrenaline hydrochloride (ADR), 7 = pyridoxine hydrochloride (PYR). For more operation conditions, see the “Instrumentation” section.

Figure 3

Effect of simultaneous addition of tetrahydrofuran (THF) and isopropanol (IP) into the background electrolyte (BGE) on the separation of five biogenic amines and two B vitamins. BGE = 25 mM GABA + 50 mM HAc + 0.1% m-HEC. Concentration of the analytes in the sample was 10 μg/mL−1. Applied separation current was 50 μA. 1 = thiamine hydrochloride (THI), 2 = tyramine hydrochloride (TYR), 3 = dopamine hydrochloride (DOP), 4 = 5-hydroxytryptamine hydrochloride (5-HT), 5 = noradrenaline hydrochloride (NOR), 6 = adrenaline hydrochloride (ADR), 7 = pyridoxine hydrochloride (PYR). For other operation conditions, see the “Instrumentation” section.
Effect of simultaneous addition of tetrahydrofuran (THF) and isopropanol (IP) into the background electrolyte (BGE) on the separation of five biogenic amines and two B vitamins. BGE = 25 mM GABA + 50 mM HAc + 0.1% m-HEC. Concentration of the analytes in the sample was 10 μg/mL−1. Applied separation current was 50 μA. 1 = thiamine hydrochloride (THI), 2 = tyramine hydrochloride (TYR), 3 = dopamine hydrochloride (DOP), 4 = 5-hydroxytryptamine hydrochloride (5-HT), 5 = noradrenaline hydrochloride (NOR), 6 = adrenaline hydrochloride (ADR), 7 = pyridoxine hydrochloride (PYR). For other operation conditions, see the “Instrumentation” section.

Figure 4

Effect of CE-β-CD addition into the background electrolyte (BGE) on the simultaneous separation of five biogenic amines and two B vitamins. (a) Addition of CE-β-CD at various concentration levels; (b) addition of CE-β-CD at 1 mg/mL−1 concentration level and isopropanol; (c) addition of CE-β-CD at 2 mg/mL−1 concentration level and isopropanol; (d) addition of CE-β-CD at 2.5 mg/mL−1 concentration level and isopropanol. BGE = 25 mM GABA + 50 mM HAc + 0.1% mHEC. Concentration of the analytes in the sample was 10 μg/mL−1. Applied separation current was 50 μA. 1 = thiamine hydrochloride (THI), 2 = tyramine hydrochloride (TYR), 3 = dopamine hydrochloride (DOP), 4 = 5-hydroxytryptamine hydrochloride (5-HT), 5 = noradrenaline hydrochloride (NOR), 6 = adrenaline hydrochloride (ADR), 7 = pyridoxine hydrochloride (PYR). For more operation conditions, see the “Instrumentation” section.
Effect of CE-β-CD addition into the background electrolyte (BGE) on the simultaneous separation of five biogenic amines and two B vitamins. (a) Addition of CE-β-CD at various concentration levels; (b) addition of CE-β-CD at 1 mg/mL−1 concentration level and isopropanol; (c) addition of CE-β-CD at 2 mg/mL−1 concentration level and isopropanol; (d) addition of CE-β-CD at 2.5 mg/mL−1 concentration level and isopropanol. BGE = 25 mM GABA + 50 mM HAc + 0.1% mHEC. Concentration of the analytes in the sample was 10 μg/mL−1. Applied separation current was 50 μA. 1 = thiamine hydrochloride (THI), 2 = tyramine hydrochloride (TYR), 3 = dopamine hydrochloride (DOP), 4 = 5-hydroxytryptamine hydrochloride (5-HT), 5 = noradrenaline hydrochloride (NOR), 6 = adrenaline hydrochloride (ADR), 7 = pyridoxine hydrochloride (PYR). For more operation conditions, see the “Instrumentation” section.

Figure 5

Effect of HP-β-CD addition into the background electrolyte (BGE) on the simultaneous separation of five biogenic amines and two B vitamins. (a) Addition of HP-β-CD at various concentration levels; (b) addition of HP-β-CD at 5 mg/mL−1 concentration level and isopropanol; (c) addition of HP-β-CD at 10 mg/mL−1 concentration level and isopropanol; (d) addition of HP-β-CD at 15 mg/mL−1 concentration level and isopropanol. BGE = 25 mM GABA + 50 mM HAc + 0.1% mHEC. Concentration of the analytes in the sample was 10 μg/mL−1. Applied separation current was 50 μA. 1 = thiamine hydrochloride (THI), 2 = tyramine hydrochloride (TYR), 3 = dopamine hydrochloride (DOP), 4 = 5-hydroxytryptamine hydrochloride (5-HT), 5 = noradrenaline hydrochloride (NOR), 6 = adrenaline hydrochloride (ADR), 7 = pyridoxine hydrochloride (PYR). For more operation conditions, see the “Instrumentation” section.
Effect of HP-β-CD addition into the background electrolyte (BGE) on the simultaneous separation of five biogenic amines and two B vitamins. (a) Addition of HP-β-CD at various concentration levels; (b) addition of HP-β-CD at 5 mg/mL−1 concentration level and isopropanol; (c) addition of HP-β-CD at 10 mg/mL−1 concentration level and isopropanol; (d) addition of HP-β-CD at 15 mg/mL−1 concentration level and isopropanol. BGE = 25 mM GABA + 50 mM HAc + 0.1% mHEC. Concentration of the analytes in the sample was 10 μg/mL−1. Applied separation current was 50 μA. 1 = thiamine hydrochloride (THI), 2 = tyramine hydrochloride (TYR), 3 = dopamine hydrochloride (DOP), 4 = 5-hydroxytryptamine hydrochloride (5-HT), 5 = noradrenaline hydrochloride (NOR), 6 = adrenaline hydrochloride (ADR), 7 = pyridoxine hydrochloride (PYR). For more operation conditions, see the “Instrumentation” section.

Accuracy and precision data obtained by the CZE-UV method.

Intra-day, n = 5
Found (μg.mL−1) RSD (%) % Nom.
Nominal (μg.mL−1) THI TYR DOP 5-HT NOR ADR PYR THI TYR DOP 5-HT NOR ADR PYR THI TYR DOP 5-HT NOR ADR PYR
0.5 0.54 - - 0.43 - - 0.56 8.2 - - 10.7 - - 15.5 108.1 - - 86.8 - - 112.1
1 1.09 - - 1.02 - - 0.99 13.1 - - 8.2 - - 10.6 108.5 - - 101.6 - - 99.4
2.5 2.77 2.64 2.07 2.42 2.95 2.84 2.59 11.0 8.6 15.1 4.2 8.9 4.4 5.4 110.7 105.5 82.9 97.0 117.8 113.5 103.7
5 4.97 5.00 4.95 5.06 4.97 4.99 5.10 3.5 8.2 7.7 1.9 13.4 3.3 3.8 99.4 100.1 98.9 101.2 99.5 99.8 101.9
10 9.23 9.21 10.21 9.77 9.71 9.54 9.22 1.2 1.5 8.0 2.7 2.3 4.8 1.7 92.3 92.1 102.1 97.7 97.1 95.4 92.2
25 25.47 25.99 26.15 25.51 24.08 25.11 25.01 1.0 1.6 6.5 0.4 1.2 0.4 1.7 101.9 104.0 104.6 102.0 96.3 100.5 100.0
50 49.91 49.65 49.42 49.79 50.48 50.02 50.10 0.4 2.1 3.4 1.2 2.6 4.0 0.5 99.8 99.3 98.9 99.6 101.0 100.1 100.2

Selected operation and validation parameters of the proposed CZE-UV method.

THI TYR DOP 5-HT NOR ADR PYR
tm (min) 12.09 12.95 14.09 14.60 15.06 15.73 18.44
RSDtm (%), n=6 0.24 0.02 0.11 0.12 0.10 0.07 0.21
RSDarea (%), n=6 1.82 4.30 8.21 5.47 4.76 4.27 1.67
Calibration equation y=104.03x + 77.112 y=15.15x − 3.721 y=9.64x + 8.563 y=44.70x + 11.626 y=7.27x − 8.347 y=12.43x − 5.641 y=38.48x − 3.648
r2 0.9994 0.9991 0.9957 0.9996 0.9924 0.9979 0.9995
Linear range 0.5 – 50 2.5 – 50 2.5 – 50 0.5 – 50 2.5 – 50 2.5 – 50 0.5 – 50
LOD (μg.mL−1) 0.15 1.25 1.25 0.25 1.25 1.25 0.25
LOQ (μg.mL−1) 0.5 2.5 2.5 0.5 2.5 2.5 0.5
N 31729 28753 27813 28162 29880 22327 27182
R 2.99 3.57 1.47 1.32 1.76 6.27

Stability study of the analytes performed under various storage conditions.

% difference from the initial concentration, n = 5
20°C 4°C −20°C −80°C
c (μg.mL−1) 2.5 15 30 2.5 15 30 2.5 15 30 2.5 15 30
THI −8.6 0.7 0.6 −0.9 −4.4 −1.3 0.26 −2.4 3.8 0.5 0.2 0.2
TYR −13.8 6.5 −2.4 9.8 8.1 −4.2 3.1 14.5 2.5 1.0 1.4 −4.6
DOP −9.4 −0.6 0.4 2.4 −2.6 0.3 3.3 −2.4 4.2 −8.6 −0.7 1.0
5−HT 2.4 0.1 0.7 4.2 −1.2 1.2 0.5 0.7 2.6 0.4 3.3 1.6
NOR 2.0 2.7 6.3 −2.8 −0.4 9.1 −5.0 1.5 2.7 −14.6 6.5 5.0
ARD −1.1 1.4 2.4 −1.0 −7.1 −1.6 3.3 2.7 2.0 −7.0 7.5 2.3
PYR 1.8 −1.2 −3.4 3.1 −9.0 −1.0 −2.8 −10.7 −1.4 0.7 −8.1 −0.6
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