Acid-base reaction-based dispersive solid phase extraction of favipiravir using biotin from biological samples prior to capillary electrophoresis analysis.

In this study, an acid-base reaction-based dispersive solid-phase extraction method was developed for the extraction of favipiravir from deionized water, plasma and urine samples prior to its determination using a capillary electrophoresis-diode array detector. The target analyte was extracted from the samples using biotin as a green adsorbent. To reach this goal, the pH of the solution was first adjusted to 9.0 (using borate buffer), and the ionic strength of the solution was enhanced by adding sodium chloride (2.5%, w/v). Thereafter, an appropriate amount of biotin was dissolved in the solution and a homogenous phase was obtained. By adding hydrochloric acid to the solution, an acid-base reaction occurs via protonation of biotin, which decreases its solubility. During this procedure, the analyte was adsorbed onto the tiny particles of the produced adsorbent dispersed into the solution. The resulting mixture was sonicated to facilitate the adsorption of the analyte onto the adsorbent surface. After the collection of biotin particles through centrifugation, the analyte was eluted using acetonitrile and then used in the determination stage. Under the optimal extraction conditions, the calibration curve was linear from 250 to 3000 ng mL-1 with a coefficient of determination of 0.9968. Low limit of detection, and quantification, good repeatability on the same day and different days (relative standard deviation ≤ 8.2%), and acceptable extraction recovery were accessed. The applicability of the method was examined by performing it on spiked plasma and urine samples, and its performance was verified.

Spermidine Exerts Protective Effects in Random-Pattern Skin Flap Survival in Rats: Possible Involvement of Inflammatory Cytokines, Nitric Oxide, and VEGF.

BACKGROUND: Distal necrosis and inflammation are two of the most common health consequences of random-pattern skin flaps survival (SFS). Anti-inflammatory effects of spermidine have been identified in various studies. On the other hand, considering the involvement of the nitric oxide molecule in the spermidine mode of action and also its role in skin tissue function, we analyzed the possible effects of spermidine on the SFS and also, potential involvement of nitrergic pathway and inflammatory cytokine in these phenomena. METHODS: Each rat was pretreated with either a vehicle (control) or various doses of spermidine (0.5, 1, 3, 5, 10 and 30 mg/kg) and then was executed a random-pattern skin flap paradigm. Also, spermidine at the dose of 5 mg/kg was selected and one group rats received spermidine 20 min prior to surgery and one additional dose 1 day after operation. Then, 7 days after operations, interleukin (IL)-6, tumor necrosis factor (TNF)-α, interferon-gamma (IFN-γ), and nitrite levels were inquired in the tissue samples by ELIZA kit. Vascular endothelial growth factor expression was assessed by DAPI staining and fluorescent microscopes. The concentrations of three polyamines, including spermidine, spermine, and cadaverine, were analyzed using HPLC. RESULTS: Pretreatment with spermidine 5 mg/kg improved SFS considerably in microscopic skin H&E staining analysis and decreased the percentage of necrotic area. Moreover, spermidine exerted promising anti-inflammatory effects via the modulation of nitric oxide and reducing inflammatory cytokines. CONCLUSIONS: Spermidine could improve skin flaps survival, probably through the nitrergic system and inflammation pathways. This preclinical study provides level III evidence for the potential therapeutic effects of spermidine on SFS in rats, based on the analysis of animal models. Further studies are needed to confirm these findings in clinical settings. LEVEL OF EVIDENCE III: This journal requires that authors assign a level of evidence to each article. For a full description of these Evidence-Based Medicine ratings, please refer to the Table of Contents or the online Instructions to Authors www.springer.com/00266 .

Online preconcentration and chiral separation of ofloxacin in exhaled breath condensate by capillary electrophoresis.

Breath analysis is an effective method of monitoring systemic or respiratory ailments. A simple chiral capillary electrophoresis method coupled with an online field-amplified sample injection stacking method is presented for ultratrace quantification of the enantiomers of ofloxacin in exhaled breath condensate (EBC). The study is focused on the use of EBC as an easily available biological sample to monitor ofloxacin's enantiomers levels with good patient compliance. The proposed method was validated in accordance with FDA guidelines over the concentration range of 0.004-1.0 µg mL-1 of racemic ofloxacin. Inter- and intra-day precision and accuracy were within the acceptable limit (below 8.50 %). The method was specific for routine analysis of ofloxacin's enantiomers. A small volume of EBC samples from seven patients under ofloxacin therapy was analyzed using the proposed method in which the concentrations of "R" and "S" enantiomers were between 0.0026 and 0.056 µg mL-1.

Biomarkers in exhaled breath condensate as fingerprints of asthma, chronic obstructive pulmonary disease and asthma-chronic obstructive pulmonary disease overlap: a critical review.

Asthma, chronic obstructive pulmonary disease (COPD) and asthma-COPD overlap are the third leading cause of mortality around the world. They share some common features, which can lead to misdiagnosis. To properly manage these conditions, reliable markers for early and accurate diagnosis are needed. Over the past 20 years, many molecules have been investigated in the exhaled breath condensate to better understand inflammation pathways and mechanisms related to these disorders. Recently, more advanced techniques, such as sensitive metabolomic and proteomic profiling, have been used to obtain a more comprehensive understanding. This article reviews the use of targeted and untargeted metabolomic methodology to study asthma, COPD and asthma-COPD overlap.

Capillary electrophoresis-mass spectrometry in pharmaceutical and biomedical analyses.

Acquiring accurate and reliable results are crucial in pharmaceutical and biomedical analyses. There is a demand for the ongoing development and validation of advanced analytical and bioanalytical methods. Mass spectrometry (MS) has become an extremely powerful tool for the identification, quantification, and characterization of small and macro-(bio)molecules. Capillary electrophoresis (CE) presents fast and high-resolution separation and in combination with MS allows sensitive and selective identification and detailed characterization. To date, CE-MS has been used to analyze a wide range of molecules, including pharmaceuticals, biopharmaceuticals, metabolites, peptides, and proteins. This review provides an update on recent applications and approaches of CE-MS relevant to biomedical and (bio)pharmaceuticals between January 2018 and May 2022. Furthermore, the latest developments on the hyphenation of CE with MS, as well as different CE modes including capillary isotachophoresis, capillary zone electrophoresis, and micellar electrokinetic capillary chromatography along with on-capillary on-line analyte stacking methods such as field-amplified sample injection, transient isotachophoresis, dynamic pH junction, and solid-phase extraction are discussed.

Ion-pair hollow fiber liquid-phase microextraction combined with capillary electrophoresis for the determination of biogenic amines in rat tissues.

Herein, the development of an ion-pair hollow fiber liquid-phase microextraction (IP-HF-LPME) procedure followed by capillary electrophoresis (CE) with indirect UV detection for the simultaneous extraction and determination of aliphatic biogenic amines including spermidine, spermine, and cadaverine in tissue samples of rat is described. In this study, tissue samples were firstly homogenized with a cold trichloroacetic acid solution then the developed method was applied to the supernatant resulting from tissue homogenization. Different CE separation and indirect UV detection conditions, as well as IP-HF-LPME extraction conditions were studied in detail and optimized. A 11 mmol L-1 imidazole solution containing 13 % (v/v) ethanol adjusted at pH 3.3 (by acetic acid) was the best running buffer for indirect UV detection of non-UV-absorbing amines. 1-Octanol in combination with salicylic acid, respectively as the membrane solvent and ion-pair reagent provided the highest extraction recovery for the studied amines. Method performances were established and good results of linearity, recovery, sensitivity, and repeatability were achieved for all the studied amines. Limits of detection were found to be between 2.8 and 4.5 ng mL-1 and limits of quantification were 15 ng mL-1 and the dynamic range was 15-2000 ng mL-1 for all three analytes.

Ultrasound-assisted electromembrane extraction of clonazepam from plasma and determination using capillary electrophoresis.

In this work, ultrasound-assisted electromembrane extraction (UA-EME) coupled with capillary electrophoresis (CE) and diode array detection (DAD) was developed for the determination of clonazepam from plasma samples. A comparative study was carried out between conventional EME and UA-EME methods to investigate the influence of the ultrasound waves on the extraction efficiency. The central composite design was used for the optimization of the variables affecting these methods to achieve the best extraction efficiency. Under optimal extraction conditions, the UA-EME provided better extraction recovery in a shorter time (58% in 13 min) than the EME method (42% in 30 min). Ultrasound reduces the extraction time and increased recovery by reducing the thickness of the barrier layer. In addition, this method provided a higher pre-concentration factor (203) and a lower limit of detection (3 ng mL-1) with good repeatability (RSDs were less than 10.11%).

Overview of therapeutic drug monitoring of immunosuppressive drugs: Analytical and clinical practices.

Immunosuppressant drugs (ISDs) play a key role in short-term patient survival together with very low acute allograft rejection rates in transplant recipients. Due to the narrow therapeutic index and large inter-patient pharmacokinetic variability of ISDs, therapeutic drug monitoring (TDM) is needed to dose adjustment for each patient (personalized medicine approach) to avoid treatment failure or side effects of the therapy. To achieve this, TDM needs to be done effectively. However, it would not be possible without the proper clinical practice and analytical tools. The purpose of this review is to provide a guide to establish reliable TDM, followed by a critical overview of the current analytical methods and clinical practices for the TDM of ISDs, and to discuss some of the main practical aspects of the TDM.

Recent advances in the determination of unbound concentration and plasma protein binding of drugs: Analytical methods.

Drugs in the bloodstream are available in both free and bound forms in which the free drug is responsible for pharmacological activities. Since protein binding determines the amount of free concentration of the drug in the blood, determining the protein binding in the early stages of drug discovery and development is of great importance. Besides, it is most beneficial to measure the free concentration of a drug in personalized medicine and therapeutic drug monitoring. For this reason, the need for sensitive, selective, and fast analytical methods to measure the free concentration of drugs and their protein binding has increased. This review aims to summarize recent advancements in analytical approaches used for the determination of free drug concentration and plasma protein binding and will focus on the most important approaches used to determine plasma protein binding. Furthermore, the concepts of each method will be described and discussed, along with their inherent advantages and disadvantages.

Electromembrane extraction as a new approach for determination of free concentration of phenytoin in plasma using capillary electrophoresis.

PURPOSE: Electromembrane extraction is a new membrane-based extraction method in which charged compounds are extracted by an electric field. So far, this method has been used to extract and isolate a variety of acidic and basic drugs from various samples, including blood and plasma. However, in this procedure, it is not yet clear whether only unbound fraction of a drug is extracted or the total drug. The aim of this study is to reveal the nature of drug extraction in the presence of plasma proteins. METHODS: To determine the nature of the extraction, the electromembrane extraction was performed from plasma solutions of phenytoin with concentrations 0.03 and 1.0 µg/mL, then the result was compared with the values obtained from the electromembrane extraction of ultrafiltrate of the same solutions (free concentration) and protein-free ultrafiltrate of plasma with final concentration of 0.03 and 1.0 µg/mL (total concentration). For this purpose, EME followed by capillary electrophoresis coupled with diode array detection was optimized and validated. RESULTS: The results showed that the electromembrane extraction method was only able to extract the unbound fraction of phenytoin from plasma samples. The method was validated over a concentration range of 0.03-4 µg/mL. The inter and intra-assay precisions were less than 6.7%. The phenytoin protein binding was also determined to be in agreement with the literature data and confirms the validity of this method. CONCLUSION: This sensitive and quick EME approach for determining the free concentration of a phenytoin, can be a good alternative to classic methods for therapeutic drug monitoring and pharmacokinetic studies.

Determination of phenytoin in exhaled breath condensate using electromembrane extraction followed by capillary electrophoresis.

Application of hollow fiber-based electromembrane extraction was studied for extraction and quantification of phenytoin from exhaled breath condensate (EBC). Phenytoin is extracted from EBC through a supported liquid membrane consisting of 1-octanol impregnated in the walls of a hollow fiber, and into an alkaline aqueous acceptor solution inside the lumen of the fiber. Under the obtained conditions of electromembrane extraction, that is, the extraction time of 15 min, stirring speed of 750 rpm, donor phase pH at 11.0, acceptor pH at 13.0, and an applied voltage of 15 V across the supported liquid membrane, an enrichment factor of 102-fold correspond to extraction percent of 25.5% was achieved. Good linearity was obtained over the concentration range of 0.001-0.10 µg/mL (r2 = 0.9992). Limits of detection and quantitation were 0.001 and 0.003 µg/mL, respectively. The proposed method was successfully applied to determine phenytoin from EBC samples of patients receiving the drug. No interfering peaks were detected that indicating excellent selectivity of the method. The intra- and interday precisions (RSDs) were less than 14%.