Fully soluble polymeric foams for in-vial dried blood spot collection and analysis of acidic drugs by capillary electrophoresis.

Polymeric foams tailor-made of polyvinylpyrrolidone (PVP) and carboxymethylcellulose/oxidized 6-carboxycellulose (CMC07/OC) composite were proposed as suitable sorbents for the collection and analysis of dried blood spots (DBSs). The PVP and CMC07/OC foams were easy to prepare, enabled collection of minute volumes of capillary blood, and blood drying at ambient temperature. The resulting foams were prepared as small porous discs with uniform dimensions (approx. 6 × 3 mm) and were fully soluble in aqueous solutions. The DBSs were formed in standard capillary electrophoresis (CE) vials fitted with the soluble foam discs and enabled the direct in-vial DBS processing and at-line analysis by CE. The DBSs were pretreated with a simple process, which involved a complete dissolution of the foam disc in an acidic solution and a simultaneous hollow fiber liquid-phase microextraction (HF-LPME) in one step. The complete solubility of the foam disc with the DBS served for a quantitative transfer of all blood components into the eluate and a nearly exhaustive HF-LPME of target analytes, whereas the blood matrix and the polymeric foam components were efficiently retained by the organic solvent impregnated in the walls of the HF. The suitability of the PVP and CMC07/OC foams for the collection and the direct analysis of DBSs was demonstrated by the HF-LPME/CE determination of model acidic drugs (warfarin, ibuprofen, naproxen, ketoprofen, and diclofenac) at therapeutically relevant concentrations. Repeatability of the analytical method was better than 8.1% (RSD), extraction recoveries ranged from 70 to 99% (for PVP foam), calibration curves were linear over two orders of magnitude (R2 higher than 0.9991), and limits of detection were less than 44 µg/L (for concentrations in undiluted capillary blood). The soluble polymeric foams exhibited non-significant variations in analyte concentrations for DBSs prepared from blood samples with different hematocrit levels and for aged DBSs (less than 9.2%), moreover, they outperformed standard DBS sampling devices in terms of sample pretreatment time and extraction recovery.

At-line coupling of hollow fiber liquid-phase microextraction to capillary electrophoresis for trace determination of acidic drugs in complex samples.

Direct analysis of complex samples is demonstrated by the at-line coupling of hollow fiber liquid-phase microextraction (HF-LPME) to capillary electrophoresis (CE). The hyphenation of the preparative and the analytical technique is achieved through a 3D-printed microextraction device with an HF located in a sample vial of a commercial CE instrument. The internal geometry of the device guides the CE separation capillary into the HF and the CE injection of the HF-LPME extract is performed directly from the HF lumen. The 3D-printing process ensures uniform dimensions of the devices, their constant position inside the sample vial, and excellent repeatability of the HF-LPME as well as the CE injection. The devices are cheap (∼0.01 €) and disposable, thus eliminating any possible sample-carryover, moreover, the at-line CE analysis of the extract is performed fully autonomously with no need for operator's intervention. The developed HF-LPME/CE-UV method is applied to the determination of acidic drugs in dried blood spot and wastewater samples and is characterized by excellent repeatability (RSD, 0.6-9.6%), linearity (r2, 0.9991-0.9999), enrichment (EF, 29-97), sensitivity (LOD, 0.2-3.4 µg/L), and sample throughput (7 samples/h). A further improvement of selected characteristics of the analytical method is achieved by the at-line coupling of HF-LPME to capillary isotachophoresis (ITP) with electrospray ionization-mass spectrometry (ESI-MS). The HF-LPME/ITP-ESI-MS system facilitates enhanced selectivity, matrix-free analytical signals, and up to 34-fold better sensitivity due to the use of ESI-MS detection and additional on-capillary ITP preconcentration of the HF-LPME extracts.

Dried Blood Spot Self-Sampling with Automated Capillary Electrophoresis Processing for Clinical Analysis.

A simple and convenient concept of blood sampling followed by a fully automated analysis is presented. A disposable sampling kit is used for accurate self-sampling of capillary blood from a finger prick. A high-throughput blood sampling is thus enabled, which is essential in many clinical assays and considerably improves life quality and comfort of involved subjects. The collected blood samples are mailed to a laboratory for a fully automated dried blood spot (DBS) processing and analysis, which are performed with a commercial capillary electrophoresis instrument. Quantitative results are obtained within 20 min from the DBS delivery to the laboratory. The presented concept is exemplified by the determination of warfarin blood concentrations and demonstrates excellent analytical performance. Moreover, this concept is generally applicable to a wide range of endogenous and exogenous blood compounds and represents a novel and attractive analytical tool for personalized health monitoring.

Hollow Fiber Liquid-Phase Microextraction At-Line Coupled to Capillary Electrophoresis for Direct Analysis of Human Body Fluids.

A simple and cheap all-in-one concept for at-line coupling of hollow fiber liquid-phase microextraction (HF-LPME) to commercial capillary electrophoresis (CE) is demonstrated, which enables the direct analysis of complex samples. A disposable microextraction device compatible with injection systems of Agilent CE instruments is proposed, which consists of a short segment of a porous HF attached to a tapered polypropylene holder. The holder maintains a constant position of the HF in a CE vial during extraction and simultaneously guides the injection end of a separation capillary into the HF lumen for automated CE injection and analysis. In a typical analytical procedure, the HF is impregnated with a water-immiscible solvent, its lumen is filled with 5 µL of an aqueous acceptor solution, and the microextraction device is placed in a 2 mL glass CE vial containing 550 µL of a donor solution. The vial is agitated at 750 rpm for 10 min, and the resulting acceptor solution is injected directly from the HF lumen into the commercial CE. No additional manual handling is required, except for the transfer of the CE vial to the CE autosampler. Multiple complex samples can be simultaneously pretreated in a multiple-well plate format, thus significantly reducing the total analysis time. Suitability of the analytical method is demonstrated by the direct determination of model basic drugs (nortriptyline, haloperidol, loperamide, and papaverine) in physiological solutions, urine, and dried blood spot (DBS) samples. Repeatability of the method is better than 12.8% (%RSD), extraction recoveries range between 34 and 76%, and enrichment factors are 37-84. The method is linear in a range of 2 orders of magnitude (R2 ≥ 0.9977) with limits of detection of 0.7-1.55 µg/L. The method has a high potential for the direct analysis of DBS samples since DBS elution and HF-LPME are performed simultaneously during the 10 min agitation. The manual DBS handling is thus reduced to inserting the DBS punch into the CE vial only. Moreover, the universal character of the HF-LPME might extend the applicability of the method to a wide range of analytes/matrices, and combination with other commercial detectors might improve the selectivity/sensitivity of the CE analysis.

Capillary Electrophoresis with Capacitively Coupled Contactless Conductivity Detection for Quantitative Analysis of Dried Blood Spots with Unknown Blood Volume.

Blood volume in dried blood spot (DBS) analysis is assumed to be constant for DBS punches with a fixed area. However, blood volume in the punch is dependent on several factors associated with the blood composition and is preferentially normalized by off-line analysis for quantitative purposes. Instead of using external instrumentation, we present an all-in-one approach for the simultaneous determination of exact blood volume in the DBS punch and the quantitation of target analytes. A DBS is eluted with 500 µL of elution solvent in a sample vial, and the eluate is directly subjected to an automated analysis by capillary electrophoresis with capacitively coupled contactless conductivity detection (CE-C4D). The capillary blood volume in the eluate is calculated from the concentrations of the inorganic blood constituents (K+, Na+, or Cl-) determined by CE-C4D, which are linearly proportional to the blood volume originally sampled onto the DBS card. Alternatively, conductivity of the DBS eluate can be used for the blood volume determination by using C4D in a nonseparation flow-through mode. The methods are suitable for the determination of blood volume in unknown DBS samples by punching out the entire DBS or by subpunching a small section of a large DBS with variations of the true vs the determined volume ≤5.5%. Practical suitability was demonstrated by the simultaneous CE-C4D determination of K+ and Na+ (for DBS volume calculation) and amino acids (target analytes) in unknown DBS samples. Quantitative analysis of selected amino acids (related to inborn metabolic disorders) in the unknown DBS was compared with a standard analytical procedure using wet-blood chemistry, and an excellent fit was obtained. The use of CE-C4D represents an important milestone in quantitative DBS analysis since the detection technique is universal, and the separation technique enables the determination of cations and/or anions and the use of multiple detectors, which further enhance selectivity/sensitivity of the analysis and the range of detectable analytes.

The effect of membrane thickness on supported liquid membrane extractions in-line coupled to capillary electrophoresis for analyses of complex samples.

The effect of membrane thickness on extraction performance was systematically examined in extractions through supported liquid membranes (SLM), which were in-line coupled to capillary electrophoresis (CE). Three porous polypropylene membranes with different thickness (25, 100 and 170 µm) were used as supports for SLM extractions of model basic drugs (nortriptyline, papaverine, haloperidol and loperamide) from complex samples. The analytes were transferred through the SLMs by a pH gradient and were in-line injected, separated and quantified using a commercial CE instrument with ultraviolet (UV) detection. Transfers of the model drugs through SLM decreased with the increased membrane thickness (in the order: 25 > 100 > 170 µm) and highest transfers were achieved for the thinnest membrane. Interferences from complex sample matrices were efficiently eliminated, moreover, impregnation of the 25 µm membrane required significantly reduced volume of organic solvent. Mechanical stability of the impregnated 25 µm membrane was excellent during in-line injections, which necessitated direct contact of CE separation capillary with the SLM. Repeatability of the hyphenated SLM-CE-UV method (using the 25 µm membrane) was lower than 11% (RSD values of peak areas) and calibration curves were strictly linear in 0.5-30 µg/mL concentration range (coefficients of determination ≥ 0.997). Transfers of the basic drugs from donor solutions (standard and undiluted human urine/plasma) through the SLMs ranged from 45 to 231% and limits of detection were between 0.02 and 0.15 µg/mL.