Separation and determination of peptide hormones by capillary electrophoresis with laser-induced fluorescence coupled with transient pseudo-isotachophoresis preconcentration.
A new method for the determination of the peptide hormones and their fragments by capillary electrophoresis (CE) with laser-induced fluorescence (LIF) detection and transient pseudo-isotachophoresis (pseudo-tITP) preconcentration was established in this study. The LIF detector used an argon ion laser with excitation wavelength at 488 nm and emission wavelength at 535 nm. Fluorescein isothiocyanate (FITC) was used as precolumn derivatization reagent to label cholecystokinin tetrapeptide (CCK-4), neurotensin (NT), neurotensin hexapeptide (NT(8-13)), and neurokinin B (NKB). Borate (10 mmol/L, pH 9.0) was selected as derivatization medium to get the high efficiency. When the addition of 70% (v/v) methanol and 1% (m/v) sodium chloride (NaCl) to the sample matrix, and with borate buffer (110 mM, pH 9.5) and 20% (v/v) methanol as running buffer, a preconcentration based on the pseudo-tITP afforded 100-fold improvement in peak heights compared with the traditional hydrodynamic injection (2.3% capillary volume). The detection limits (signal/noise=3) based on peak height were found to be 0.04, 0.1, 0.2, and 0.08 nmol/L for NT(8-13), NT, NKB, and CCK-4, respectively. The method was validated and applied to qualitative analysis of NT and NT(8-13) in human cerebrospinal fluid sample.
Chen
Ying
Y
Ministry of Education, Key Laboratory of Analysis and Detection Technology for Food Safety, Fuzhou University, Fuzhou, Fujian 350002, China.
Xu
Liangjun
L
Zhang
Lan
L
Chen
Guonan
G
D015815:Cell Adhesion Molecules / Q000032:analysis / Q000134:cerebrospinal fluid / Q000302:isolation & purification; D019075:Electrophoresis, Capillary / Q000379:methods*; D016650:Fluorescein-5-isothiocyanate / Q000737:chemistry; D005453:Fluorescence; D006801:Humans; D054020:Lasers, Gas; D015287:Neurokinin B / Q000032:analysis / Q000134:cerebrospinal fluid / Q000302:isolation & purification; D009496:Neurotensin / Q000032:analysis / Q000134:cerebrospinal fluid / Q000302:isolation & purification; D010446:Peptide Fragments / Q000032:analysis / Q000134:cerebrospinal fluid / Q000302:isolation & purification; D036361:Peptide Hormones / Q000134:cerebrospinal fluid / Q000737:chemistry / Q000302:isolation & purification*; D020794:Receptor Protein-Tyrosine Kinases / Q000032:analysis / Q000134:cerebrospinal fluid / Q000302:isolation & purification; D013345:Subarachnoid Hemorrhage / Q000134:cerebrospinal fluid
D015815:Cell Adhesion Molecules; D010446:Peptide Fragments; D036361:Peptide Hormones; D009496:Neurotensin; C054381:neurotensin (8-13); D015287:Neurokinin B; C096905:PTK7 protein, human; D020794:Receptor Protein-Tyrosine Kinases; D016650:Fluorescein-5-isothiocyanate