Hadamard transform CE-UV detection for biological samples

Abstract

A Hadamard transform-capillary electrophoresis-UV (HT-CE-UV) detection technique is described for the analysis of biological samples. Pseudorandom injections of sample and buffer according to a simplex matrix obtained from the corresponding Hadamard matrix is performed with conventional capillaries. Alternating injections are achieved with a novel capillary "T" connector created by drilling conventional capillary dimensions through a 1-cm diameter polycarbonate disc. This connector design coupled with a switching system allows for rapid, electrokinetic injections of solution into alternating sample and buffer capillary arms for UV detection. The standard mixtures of nitric oxide (NO) metabolites, nitrite and nitrate, dissolved in physiological saline solution are injected into the separation capillary according to an 83-element injection sequence to obtain a signal-to-noise ratio (S/N) enhancement of ca. 4.5 over a single injection. Nitrite, being the less concentrated metabolite in NO detection and thereby more difficult to detect, was calibrated with the HT-CE-UV method and a limit of detection (LOD) of 0.56 microM was obtained. Rat blood plasma was analyzed with this detection system and demonstrated to be comparable with NO metabolite concentrations of previously published results. This HT-CE-UV method is described where a unique reservoir tube design that contains 8-microL standard nitrite sample volumes is placed over the end of the capillary arm to explore low volume limits for biological samples.

Bioanalytical profile of the l-arginine/nitric oxide pathway and its evaluation by capillary electrophoresis

“…This sensitivity was sufficient to quantify nitrate and nitrite in the mammalian brain extracts, but was insufficient to detect the nitrite ion in biological fluids with apparently low nitrite content. Other bioanalytical examples and improvements of this technique have been reported subsequently [250][251][252][253][254][255][256][257][258][259][260][261].…”

“…HPCE detection of NO 2 − in biological samples is a good target for such an improvement. For example, McReynolds et al [251] utilized a pseudorandom alternating sample-buffer injection according to a Hadamard matrix, using conventional capillaries with an original "T" connector coupler. By applying an 83-element injection sequence, they were able to improve the SNR by a factor of 4.5 over a single injection mode, leading to an LOQ value of 0.56 μM for nitrite [251].…”

“…For example, McReynolds et al [251] utilized a pseudorandom alternating sample-buffer injection according to a Hadamard matrix, using conventional capillaries with an original "T" connector coupler. By applying an 83-element injection sequence, they were able to improve the SNR by a factor of 4.5 over a single injection mode, leading to an LOQ value of 0.56 μM for nitrite [251]. A detailed description of HPCE/ HT applications with a focus on planar chip CE techniques has been recently published [398].…”

Cross-correlation capillary electrophoresis in unmodified commercial equipment

“…Cross-correlation CE has successfully been performed using UV [10][11][12][13], LIF [14][15][16] or even amperometric detection [17]. It has been used in conventional CE [18,19] and in microchip CE [14,17,21,22].…”

Online photolytic optical gating of caged fluorophores in capillary zone electrophoresis utilizing an ultraviolet light‐emitting diode

“…Here, we demonstrate the first use of an UV-LED array for online photolytic injection of caged fluorophores in CZE. Further, we coupled UV-LED POG to Hadamard transform capillary electrophoresis (HTCE) [17,[28][29][30][31][32][33] to demonstrate enhanced improvements in sensitivity with minimal instrumental modification.…”