Chiral capillary electrophoresis–mass spectrometry of 3,4-dihydroxyphenylalanine: Evidence for its enantioselective metabolism in PC-12 nerve cells

Abstract

A fully automated chiral capillary electrophoresis - tandem mass spectrometric method (CE-MS/MS) was developed for enantiomeric quantification of DOPA and its precursors, phenylalanine (Phe) and tyrosine (Tyr). To avoid MS source contamination, a negatively charged chiral selector, sulfated β-cyclodextrin (sulfated β-CD) that migrated away from the detector was used in combination with the partial filling technique. The six stereoisomers were simultaneously quantified in less than 12 min. Detection limits were 0.48 and 0.51 μM for L- and D-DOPA enantiomers, respectively. Assay reproducibility (RSD, n=6) were 4.43%, 3.15%, 4.91%, 5.16%, 3.96%, and 3.25% for L-/D-DOPA, L-/D-Tyr, and L-/D-Phe at 10.0 μM, respectively. Thanks to the high enantioseparation efficiency, detection of trace D-DOPA in L-/D-DOPA mixtures could be achieved. The assay was employed to study the metabolism of DOPA, a well known therapeutic drug for treating Parkinson’s disease. It was found that L-DOPA was metabolized effectively in PC-12 cells. About 88% of L-DOPA disappeared after incubation at a cell density of 2 × 106 cells/mL for 3 hrs. However, D-DOPA coexisting with L-DOPA in the incubation solution remained intact. The enantiospecific metabolism of DOPA in this neuronal model was demonstrated.

Self-assembly of l-cysteine–gold nanoparticles as chiral probes for visual recognition of 3,4-dihydroxyphenylalanine enantiomers

“…In recent years, chiral selectors were used to modify the electrodes, and some electrochemical methods were developed for recognition of Dopa enantiomers without the need of separation. [9][10][11][12][13] Although progress in chiral discrimination of Dopa has been achieved during the past decades, [4][5][6][7][8][9][10][11][12][13] it is still a challenge to develop a much simple, inexpensive and convenient techniques for chiral recognition and quantication of Dopa. One of the most pressing challenges in the design of chiral assay is to achieve visual discrimination of enantiomers.…”

Evaluation of a microchip electrophoresis-mass spectrometry platform deploying a pressure-driven make-up flow

“…Detection limit (S/N =3) was estimated to be 0.10 μM DOPA. Compared with our previous work where a CE-ESI-MS method was developed for DOPA quantification [20], the present MCE-nanoESI-MS method was more sensitive (LODs: 0.10μM versus 0.50 μM). The increase in assay sensitivity could be attributed to two factors: 1) less dilution of the CE effluent by sheath liquid, and 2) use of nanoESI in the present system [16].…”

“…A review on coupling MCE with ESI-MS was recently given [16]. In an effort to develop MCE-ESI-MS methods with the use of a sheathless interface, referencing to the microchip designs reported in literature and on the basis of our experience with microchip fabrication [17–18] and CE-ESI-MS method development [19–20], we encountered problems /inconveniences such as the difficulty to start electrospray without a pneumatic assistance and easily clogging of the tiny nanoESI emitter. Similar observations were also reported by other research groups [11, 21–22].…”

A microchip electrophoresis-mass spectrometric platform for fast separation and identification of enantiomers employing the partial filling technique

“…Detection limits (S/N =3) were estimated to be 0.043 μM and 0.047 μM for L-DOPA and D-DOPA, respectively. Compared with the CE-MS analytical results [57], the present MCE-nanoESI-MS method was 10-times more sensitive (LODs: 0.04 μM versus 0.50 μM). The increase in assay sensitivity can be attributed to at least two factors: 1) less dilution of the MCE effluent by MUF liquid, and 2) use of nanoESI in the present system.…”