Capillary electrophoresis coupled with chloroform‐acetonitrile extraction for rapid and highly selective determination of cysteine and homocysteine levels in human blood plasma and urine

Abstract

A rapid and selective method has been developed for highly sensitive determination of total cysteine and homocysteine levels in human blood plasma and urine by capillary electrophoresis (CE) coupled with liquid–liquid extraction. Analytes were first derivatized with 1,1′‐thiocarbonyldiimidazole and then samples were purified by chloroform–ACN extraction. Electrophoretic separation was performed using 0.1 M phosphate with 30 mM triethanolamine, pH 2, containing 25 μM CTAB, 2.5 μM SDS, and 2.5% polyethylene glycol 600. Samples were injected into the capillary (with total length 32 cm and 50 μm id) at 2250 mbar*s and subsequent injection was performed for 30 s with 0.5 M KОН. The total analysis time was less than 9 min, accuracy was 98%, and precision was <2.6%. The LOD was 0.2 μM for homocysteine and 0.5 μM for cysteine. The use of liquid–liquid extraction allowed the precision and sensitivity of the CE method to be significantly increased. The validated method was applied to determine total cysteine and homocysteine content in human blood plasma and urine samples obtained from healthy volunteers and patients with kidney disorders.

Advances in the Determination of Nonprotein Amino Acids in Foods and Biological Samples by Capillary Electrophoresis

“…In this work, the use of an electrokinetic injection with pH mediated stacking enabled to reach a LOD for homocysteine of 0.8 M. Subsequently, the authors improved the methodology introducing several modifications which enabled to achieve a LOD of 0.2 M [36]. These modifications included the use of a liquid-liquid extraction with chloroform-ACN to purify the sample and determine homocysteine and cysteine levels in urine (the previous approach was not suitable for determining both analytes in matrices like urine in which salt levels vary considerable), a different composition of the running buffer (see Table 2), and an in-capillary preconcentration step based on the use of field amplified sample stacking and pH mediated stacking.…”

“…These modifications included the use of a liquid-liquid extraction with chloroform-ACN to purify the sample and determine homocysteine and cysteine levels in urine (the previous approach was not suitable for determining both analytes in matrices like urine in which salt levels vary considerable), a different composition of the running buffer (see Table 2), and an in-capillary preconcentration step based on the use of field amplified sample stacking and pH mediated stacking. In this way, homocysteine and cysteine levels were determined in human plasma and urine samples from healthy subjects and patients with kidney disorders (see Figure 5), observing a decrease in the homocysteine levels in urine from patients with kidney disorders [36]. Other detection modes different from UV were also hyphenated with CE 21 to perform the determination of homocysteine in biological samples.…”

Recent progress of sample stacking in capillary electrophoresis (2016–2018)

“…Ivanov et al. presented a method for sensitive determination of total cysteine and homocysteine levels in human blood plasma and urine by applying FASS and pH mediated stacking. The analytes were first derivatized with 1,1‐thiocarbonyldiimidazole and purified by LLE.…”

Red-Emissive Carbon Dots for “Switch-On” Dual Function Sensing Platform Rapid Detection of Ferric Ions and l-Cysteine in Living Cells

“…These methods mainly include inductively coupled plasma mass spectrometry (ICP-MS), 5 inductively coupled plasma-atomic emission spectrometry (ICP-AES), 6 atomic absorption spectrometry, membrane separation, 7 chromatographic separation, 8 and capillary electrophoresis. 9 Although these methods are widely used, the disadvantages such as time consuming, cumbersome operation, expensive equipment, poor selectivity, and low detection efficiency are in urgent need of improvement. Therefore, the development of a fast, accurate, environment-friendly, and rapid sensing platform for Fe 3+ ions and l -Cys is of great significance.…”