Plasma protein binding refers to the binding of a drug to plasma proteins after entering the body. The measurement of plasma protein binding is essential during drug development and in clinical practice, as it provides a more detailed understanding of the available free concentration of a drug in the blood, which is in turn critical for pharmacokinetics and pharmacodynamics studies. In addition, the accurate determination of the free concentration of a drug in the blood is also highly important for therapeutic drug monitoring and in personalized medicine. The present study uses C18-coated solid-phase microextraction 96-pin devices to determine the free concentrations of a set of drugs in plasma, as well as the plasma protein binding of drugs with a wide range of physicochemical properties. It should be noted that the extracted amounts used to calculate the binding constants and plasma protein bindings should be measured at respective equilibrium for plasma and phosphate buffer. Therefore, special attention is placed on properly determining the equilibration times required to correctly estimate the free concentrations of drugs in the investigated systems. The plasma protein binding values obtained with the 96-pin devices are consistent with those reported in the literature. The 96-pin device used in this research can be easily coupled with a Concept96 or other automated robotic systems to create an automated plasma protein binding determination protocol that is both more time and labor efficient compared to conventional equilibrium dialysis and ultrafiltration methods.

Immunosuppressive drugs are administered to decrease immune system activity (e.g. of patients undergoing solid organ transplant). Concentrations of immunosuppressive drugs (ISDs) in circulating blood must be closely monitored during the period of immunosuppression therapy due to adverse effects that take place when concentration levels fall outside of the very narrow therapeutic concentration range of these drugs. This study presents the rapid determination of four relevant immunosuppressive drugs (tacrolimus, sirolimus, everolimus, and cyclosporine A) in whole human blood by directly coupling solid-phase microextraction to mass spectrometry via the microfluidic open interface (Bio-SPME-MOI-MS/MS). The BioSPME-MOI-MS/MS method offers ≤ 10% imprecision of in-house prepared quality controls over a 10-day period, ≤ 10% imprecision of ClinCal® Recipe calibrators over a three-day period, and single total turnaround time of ∼ 60 min (4.5 min for high throughput). The limits of quantification were determined to be 0.8 ng mL-1 for tacrolimus, 0.7 ng mL-1 sirolimus, 1.0 ng mL-1 for everolimus, and 0.8 ng mL-1 for cyclosporine. The limits of detection were determined to be 0.3 ng mL-1 for tacrolimus, 0.2 ng mL-1 for sirolimus, 0.3 ng mL-1 for everolimus, and 0.3 ng mL-1 for cyclosporine A. The R2 values for all analytes were above 0.9992 with linear dynamic range from 1.0 mL-1 to 50.0 ng mL-1 for tacrolimus, sirolimus, and everolimus while from 2.5 ng mL-1 to 500.0 ng mL-1 for cyclosporine A. To further evaluate the performance of the present method, 95 residual whole blood samples of tacrolimus and sirolimus from patients undergoing immunosuppression therapy were used to compare the Bio-SPME-MOI-MS/MS method against a clinically validated reference method based on chemiluminescent microparticle immunoassay, showing acceptable results. Our results demonstrated that Bio-SPME-MOI-MS/MS can be considered as a suitable alternative to existing methods for the determination of immunosuppressive drugs in whole blood providing faster analysis, better selectivity and sensitivity, and a wider dynamic range than current existing approaches.

We present a modified microfluidic open interface (MOI) for the direct coupling of Bio-SPME to a liquid electron ionization-tandem mass spectrometry (LEI-MS/MS) system as a sensitive technique that can directly analyze biological samples without the need for sample cleanup or chromatographic separations as well as without measurable matrix effects (ME). We selected fentanyl as test compound. The method uses a C18 Bio-SPME fiber by direct immersion (DI) in urine and plasma and the subsequent quick desorption (1 min) in a flow-isolated volume (2.5 μL) filled with an internal standard–acetonitrile solution. The sample is then transferred to an EI source of a triple-quadrupole mass spectrometer via a LEI interface at a nanoscale flow rate. The desorption and analysis procedure requires less than 10 min. Up to 150 samples can be analyzed without observing a performance decline, with fentanyl quantitation at microgram-per-liter levels. The method workflow is extremely dependable, relatively fast, sustainable, and leads to reproducible results that enable the high-throughput screening of various biological samples.

A semi-automated and sensitive method was developed for simultaneous determination of the six most consumed artificial sweeteners (AS) in surface waters using thin-film solid-phase microextraction (TF-SPME) and high-performance liquid chromatography (HPLC). A triple quadrupole mass spectrometer and an electrospray ionization source (ESI-MS) run in negative ionization and multiple reaction monitoring modes were employed for instrumental analysis. The TF-SPME method was optimized for the extraction phase, sample pH, desorption solvent, extraction time, and desorption time. In-house-synthetized-hydrophilic-lipophilic balance weak anion exchange (HLB-WAX) particles imbedded within a polyacrylonitrile (PAN) binder were selected as the extraction phase for the thin-film coating due to their cost-effectiveness and enhanced sensitivity for artificial sweeteners. Suitable analytical parameters that include linearity (R2 > 0.9914), recovery > 80%, inter, and intra-reproducibility less than 18% were obtained for the AS compounds studied. The developed method estimated limits of detection (LODs) ranging from 0.004 to 0.038 ng mL-1 The SPME method was successfully applied for the determination of ultra-trace levels of AS in water samples collected from Grand River (Ontario, Canada), downstream of three municipal wastewater treatment plants (WWTPs). Concentrations ranging from 0.03 to 20.3 ng mL-1 were found for the AS compounds studied.

In this article, the use of an SPME technique is reported for the first time for direct measurement of free drug concentration in solid tissue. In our investigations, we considered doxorubicin (DOX) spiked in homogenized tissue matrix at transient and equilibrium extraction conditions, with subsequent assessment of obtained experimental results by an in silico approach using mathematical models developed in COMSOL Multyphysics. In silico studies were performed on the basis of transported diluted species (tds) and reaction engineering (re) modules from COMSOL Multiphysics, using the same conditions as those used to attain experimental results. To determine the apparent binding affinity of DOX to the tissue matrix which contains multiple binding species, the experimentally determined binding affinity of DOX with human serum albumin (HSA) was considered to simplify the mathematical calculations. Here, the value of the binding affinity was considered for a single binding site and adjusted by fitting the experimental results with the mathematical model. Bovine lung tissue homogenate was selected as a surrogate matrix, and a biocompatible C-8 commercial SPME fiber was used for extraction of DOX. In total, four mathematical models were herein developed to describe the mass transfer kinetics of solid coatings: in agar gel at static conditions, in PBS solution with agitated conditions, extraction in PBS solution in the presence of an HSA binding matrix, and static extraction in homogenized lung tissue. For all conditions, simulated results were in good agreement with experimental results. The developed mathematical model allows for measurements of free drug concentrations inside the tissue matrix and facilitates calculations of local depletion of DOX by a solid SPME coating. Results of the investigations indicate that local depletion of the free form of DOX, even at the kinetic stage, is negligible for tissue extraction, as the release of the heavily bound analyte (over 99% binding to tissue matrix) is very rapid, thus easily compensating for the loss of the drug to the SPME coating. This indicates that the dissociation rate constant of DOX from lung tissue components is very rapid; therefore, the mass transfer of drug to the fiber coating via free from is very efficient. Our results also indicate that thin coating SPME fibers provide a good way to measure drug distribution after dosing, as extractions via thin coating SPME fibers do not affect the free concentration of the drug, which is responsible for drug distribution in tissue.

An alternative strategy to increase mass transfer entails geometry optimization of the extraction systems including design of solid-phase microextraction (SPME) probes. In this work, a computational model was employed to elucidate practical aspects such as efficiency and kinetics of extraction by employing several new geometries. Extraction of a model analyte at static conditions with the configurations, such as thin-film, fiber, coated tip, and nanoparticles, was numerically simulated to obtain an in-depth understanding of the advantages and limitations of each geometry in microextraction and exhaustive modes. The attained results associated with the equilibration time dependency on shape were in good agreement with previously reported experimental observations. They demonstrate that the mass-transfer is highly dependent on the size and shape of the coatings and increases with a decrease in size of the devices particularly rapidly below 10 μm caused by radial diffusion effect. Nevertheless, extractions performed using octadecyl-functionalized magnetic nanoparticles demonstrated that higher enrichment factors are achievable with the use of a fewer number of particles in comparison to factors achieved via exhaustive extraction, where a larger number of particles must be employed, confirming theoretical predictions. The conclusions reached are valid for any extraction method. The results obtained herein are very useful toward the design and optimization of future extraction technologies and approaches.