Therapeutic drug monitoring for valproate: deriving a novel formula for calculation of free concentration.

BACKGROUND: Monitoring free valproate concentrations, as with other highly protein-bound anticonvulsants, is essential in clinical situations where protein binding may be disrupted. Conversion of measured total concentrations to approximate free concentrations offers a cost-effective alternative. This study evaluated the relationship between total and free valproate concentrations for discordance and the impact of key determinants. A novel formula was devised that incorporates significant variables. METHODS: A multicentre, cross-sectional observational analytical study included 101 subjects 18 years and older using valproate for 6 months or longer. Participants were recruited from private and public sector healthcare settings from primary to tertiary level in, South Africa, during 2017-2019. RESULTS: Free valproate concentrations could be measured for 84 subjects. Discordance for concomitant total and free valproate concentrations was 79.1%. Among 19 participants with elevated free concentrations, 15 (78.9%) had total valproate concentrations within the recommended reference range. Calculations based on the study-derived formula were more accurate in predicting free valproate concentration than previously proposed methods. CONCLUSION: This study proposes that the novel formula for calculating free valproate enables more accurate prediction.

The use of concomitant medications on nephrotoxicity associated with teicoplanin: A retrospective observational study.

BACKGROUND: Teicoplanin (TEIC) is a nephrotoxic agent. However, little is known about the effects of concomitant medications on nephrotoxicity. In this study, we investigated the effects of concomitant drugs on nephrotoxicity. METHODS: A retrospective observational case-control study was conducted on patients (≥18 years) who started TEIC at the Tokyo Dental College, Ichikawa General Hospital, between January 2013 and April 2023. The primary outcome was nephrotoxicity, defined as an increase in serum creatinine levels of ≥50 % or ≥0.5 mg/dL from baseline. Logistic regression analysis was used to determine the risk factors for nephrotoxicity associated with TEIC. In addition, we investigated the relationship between nephrotoxicity and predicted free TEIC concentrations. RESULTS: Of 305 patients, 43 (14.1 %) developed nephrotoxicity. The multivariate logistic regression analysis identified that serum albumin (odds ratio [OR] = 0.50, 95 % confidence interval [CI] 0.27-0.89, p = 0.02), concomitant use of loop diuretics (OR = 2.22, 95 % CI 1.10-4.59, p = 0.03), antivirals (OR = 3.24, 95 % CI 1.32-7.62, p < 0.01), and vasopressors (OR = 2.57, 95 % CI 1.10-5.78, p = 0.03) were the associated risk factors for nephrotoxicity in patients administered with TEIC. In 216 patients, predicted TEIC concentrations were 3.6 [interquartile range (IQR), 2.6-4.9] µg/mL in the nephrotoxicity group versus 3.6 [IQR, 2.5-4.7] µg/mL in the non-nephrotoxicity group, with no significant difference (p = 0.69). CONCLUSION: Our results indicate the importance of modifying the concomitant use of loop diuretics, antivirals, and vasopressors.

Therapeutic drug monitoring of free vancomycin concentration in practice: A new analytical technique based on the HFCF-UF sample separation method.

The aim of this study was to establish a method for free vancomycin concentration determination in human plasma and apply it to clinical therapeutic drug monitoring (TDM). The unbound vancomycin in plasma was separated by the hollow fiber centrifugal ultrafiltration (HFCF-UF) technique and analyzed by HPLC. Chromatographic conditions were optimized, the specificity, linearity, precision, recovery and stability of the method were examined, and plasma samples of patients were measured. The standard curve for free vancomycin is y = 0.0277x - 0.0080 with good linearity within 0.25-50 µg·mL-1 . The relative and absolute recovery rates for vancomycin were 98.63-101.0% and 88.41-101.2%, respectively. The intraday and interday precision RSDs were <10%. Plasma was stable under several conditions. The TDM value of the free vancomycin concentration of 20 patients was 0.99-38.51 µg·mL-1 , and the correlation between the free and total concentrations was not significant. The unbound fraction of vancomycin ranged from 25.5 to 84.8%, with large variation. The operation of free vancomycin separation by HFCF-UF was simple and suitable for TDM in practice. The unbound fraction of vancomycin in clinical samples varied significantly between individuals. It is recommended to perform free concentration TDM in critically ill patients.

Human plasma protein levels alter the in vitro antifungal activity of caspofungin: An explanation to the effect in critically ill?

BACKGROUND: Recent studies have shown low caspofungin concentrations in critically ill patients. In some patients, the therapeutic target, area under the total plasma concentration curve in relation to the minimal inhibition concentration (AUCtot /MIC), seems not to be achieved and therapeutic drug monitoring (TDM) has been proposed. Caspofungin is highly protein-bound and the effect of reduced plasma protein levels on pharmacodynamics has not been investigated. OBJECTIVES: Fungal killing activity of caspofungin in vitro was investigated under varying levels of human plasma protein. METHODS: Time-kill studies were performed with clinically relevant caspofungin concentrations of 1-9 mg/L on four blood isolates of C. glabrata, three susceptible and one strain with reduced susceptibility, in human plasma and plasma diluted to 50% and 25% using Ringer's acetate. RESULTS: Enhanced fungal killing of the three susceptible strains was observed in plasma with lower protein content (p < .001). AUCtot /MIC required for a 1 log10 CFU/ml kill at 24 h in 50% and 25% plasma was reduced with 36 + 12 and 80 + 9%, respectively. The maximum effect was seen at total caspofungin concentrations of 4-9 × MIC. For the strain with reduced susceptibility, growth was significantly decreased at lower protein levels. CONCLUSIONS: Reduced human plasma protein levels increase the antifungal activity of caspofungin in vitro, most likely by increasing the free concentration. Low plasma protein levels in critically ill patients with candidemia might explain a better response to caspofungin than expected from generally accepted target attainment and should be taken into consideration when assessing TDM based on total plasma concentrations.

Digestion of Lipid-Based Formulations Not Only Mediates Changes to Absorption of Poorly Soluble Drugs Due to Differences in Solubilization But Also Reflects Changes to Thermodynamic Activity and Permeability.

The aim of this study was to evaluate the effect of lipid digestion on the permeability and absorption of orally administered saquinavir (SQV), a biopharmaceutics classification system (BCS) class IV drug, in different lipid-based formulations. Three LBFs were prepared: a mixed short- and medium-chain lipid-based formulation (SMCF), a medium-chain lipid-based formulation (MCF), and a long-chain lipid-based formulation (LCF). SQV was loaded into these LBFs at 26.7 mg/g. To evaluate the pharmacokinetics of SQV in vivo, drug-loaded formulations were predispersed in purified water at 3% w/w and orally administered to rats. A low dose (0.8 mg/rat) was employed to limit confounding effects on drug solubilization, and consistent with this design, presolubilization of SQV in the LBFs did not increase in vivo exposure compared to a control suspension formulation. The areas under the plasma concentration-time curve were, however, significantly lower after administration of SQV as MCF and LCF compared to SMCF. To evaluate the key mechanisms underpinning absorption, each LBF containing SQV was digested, and the flux of SQV from the digests across a dialysis membrane was evaluated in in vitro permeation experiments. This study revealed that the absorption profiles were driven by the free concentration of SQV and that this varied due to differences in SQV solubilization in the digestion products generated by LBF digestion. The apparent first-order permeation rate constants of SQV (kapp,total) were estimated by dividing the flux of SQV in the dialysis membrane experiments by the concentration of total SQV on the donor side. kapp,total values strongly correlated with in vivo AUC. The data provide one of the first studies of the effect of digestion products on the free concentration of a drug in the GI fluid and oral absorption. This simple permeation model may be a useful tool for the evaluation of the impact of lipid digestion on apparent drug permeability from lipid-based formulations. These effects should be assessed alongside, and in addition to, the more well-known effects of lipids on enhancing intestinal solubilization of poorly water-soluble drugs.

Factors to influence the accuracy of albumin adjusted free valproic acid concentration.

BACKGROUND: Therapeutic drug monitoring (TDM) is recommended during treatment with valproic acid (VPA), as is the measurement of free VPA concentration (MfVPA). However, MfVPA is unavailable in many institutions. Based on the highly protein-bound characteristics of VPA, an albumin-adjusted formula has been proposed to predict free VPA concentration (PfVPA). Nevertheless, the factors affecting the accuracy of this formula remain unknown, as does the concordance between MfVPA and PfVPA. METHODS: Adult patients receiving VPA and undergoing TDM were enrolled. Free and total serum concentration (TVPA) were categorized as subtherapeutic, therapeutic, or supratherapeutic based on the reference range of 5-15 and 50-100 µg/mL, respectively. Concordance was defined as MfVPA and PfVPA, or MfVPA and TVPA, falling within the same category. Multivariate logistic regression with generalized estimating equation was adopted to identify factors affecting concordance, and the receiver operating characteristic curve was applied to determine the cutoff values of predictors. RESULTS: A total of 98 data points from 51 participants were included for analysis. The concordance of MfVPA and PfVPA, and MfVPA and TVPA, was 72% and 44%, respectively. Blood urea nitrogen (BUN) (0.97 [0.95-0.99], P = 0.01) and TVPA (0.97 [0.95-0.99], P = 0.02) had a significant influence on the concordance of MfVPA and PfVPA. The cutoff values of TVPA and BUN for the accuracy of the albumin-adjusted formula were 56.4 µg/mL and 51.05 mg/dL, respectively. CONCLUSION: If MfVPA is not available, the albumin-adjusted formula should be applied before VPA dosage adjustment when TVPA is < 56.4 µg/mL and BUN is < 51.05 mg/dL.

Comparative study on quantitation of human myoglobin by both isotope dilution mass spectrometry and surface plasmon resonance based on calibration-free analysis.

The activity of proteins rather than the concentration of proteins in biopharmaceutical and in vitro diagnostics are often the primary focus. Nonetheless, development of a calibration-free concentration analysis (CFCA) approach that accurately quantifies the concentration of proteins based on molecular interactions with specific monoclonal antibodies and without the requirement of external calibrators would be beneficial to diagnostics. Generally, only analytes that interact with the antibody (Ab) are quantified by CFCA. Moreover, protein concentrations measured by CFCA usually vary when different Abs are used, and are lower than those obtained by amino acid analysis because any non-native state population of the target protein is not captured by the Ab. To achieve comparable results between CFCA and traditional amino acid analysis (AAA), an Ab that recognizes the target protein irrespective of its conformation should be used. In this report, three different monoclonal antibodies were used to quantify purified human myoglobin in solution by CFCA. The concentrations obtain by the Abs (i.e., 2.985, 2.912, 3.032 mg mL-1) were comparable with that obtained by AAA. Moreover, isotope dilution mass spectrometry (IDMS) gave a human myoglobin concentration of 2.851 mg mL-1, which is also in agreement with the results from CFCA. The performance of CFCA was evaluated by measuring various parameters, including within-day and between-day precision. The results demonstrated that the active concentration measured by CFCA is comparable with that of IDMS when the appropriate Ab is used. Recommended procedures for performing the new CFCA approach are provided. This study shows that CFCA represents a primary method for accurate protein concentration determination, which should aid the development of certified reference materials. Graphical abstract.

Calibration-free concentration analysis for an analyte prone to self-association.

Calibration-free concentration analysis (CFCA) based on surface plasmon resonance uses the diffusion coefficient of an analyte to determine the concentration of that analyte in a bulk solution. In general, CFCA is avoided when investigating analytes prone to self-association, as the heterogeneous diffusion coefficient results in a loss of precision. The derivation for self-association of the analyte was presented here. By using the diffusion coefficient for the monomeric state, CFCA provides the lowest possible concentration even though the analyte is self-associated.

Analysis of the dose-dependent stage-specific in vitro efficacy of a multi-stage malaria vaccine candidate cocktail.

BACKGROUND: The high incidence and mortality rate of malaria remains a serious burden for many developing countries, and a vaccine that induces durable and highly effective immune responses is, therefore, desirable. An earlier analysis of the stage-specific in vitro efficacy of a malaria vaccine candidate cocktail (VAMAX) considered the general properties of complex multi-component, multi-stage combination vaccines in rabbit immunization experiments using a hyper-immunization protocol featuring six consecutive boosts and a strong, lipopolysaccharide-based adjuvant. This follow-up study investigates the effect of antigen dose on the in vitro efficacy of the malaria vaccine cocktail using a conventional vaccination scheme (one prime and two boosts) and a human-compatible adjuvant (Alhydrogel(®)). RESULTS: IgG purified from rabbits immunized with 0.1, 1, 10 or 50 µg doses of the VAMAX vaccine candidate cocktail was analysed for total IgG and antigen-cocktail-specific titers. An increase in cocktail-specific titers was observed between 0.1 and 1 µg and between 10 and 50 µg, whereas no significant difference in titers was observed between 1 and 10 µg. Antigen component-specific antibody titers and stage-specific in vitro efficacy assays were performed with pooled IgG from animals immunized with 1 and 50 µg of the VAMAX cocktail. Here, the component-specific antibody levels showed clear dose dependency whereas the determined stage-specific in vitro IC50 values (as a correlate of efficacy) were only dependent on the titer amounts of stage-specific antibodies. CONCLUSIONS: The stage-specific in vitro efficacy of the VAMAX cocktail strongly correlates with the corresponding antigen-specific titers, which for their part depend on the antigen dose, but there is no indication that the dose has an effect on the in vitro efficacy of the induced antibodies. A comparison of these results with those obtained in the previous hyper-immunization study (where higher levels of antigen-specific IgG were observed) suggests that there is a significant need to induce an immune response matching efficacy requirements, especially for a PfAMA1-based blood stage vaccine, by using higher doses, better adjuvants and/or better formulations.

Temocillin (6 g daily) in critically ill patients: continuous infusion versus three times daily administration.

OBJECTIVES: The growing incidence of infections caused by Enterobacteriaceae producing ESBLs has led to increased use of carbapenems. Temocillin, which resists most ß-lactamases, may be a useful alternative. The aim of this study was to assess the pharmacokinetics and target attainment rates of 6 g of temocillin daily divided into three administrations every 8 h (three times daily) or administered by continuous infusion in critically ill patients. PATIENTS AND METHODS: This was a prospective, two-centre, randomized, controlled study in patients with intra-abdominal or lower respiratory tract infections caused by Enterobacteriaceae. RESULTS: Thirty-two patients were included and analysed for clinical efficacy, and pharmacokinetics were measured in 29 of them. Four patients undergoing continuous veno-venous haemofiltration (CVVH) were analysed separately. Mean, median and range of percentages of the dosing interval during which the free drug concentration remained >16 mg/L were 76.4, 98 and 18.7-98.9 in patients treated three times daily and 98.9, 89.7 and 36.4-99.9 in patients with continuous infusion, respectively. Clinical cure rates were 79% and 93% in each of these groups, respectively (not significant). Patients with CVVH received a daily dose of 750 mg given by continuous infusion and had a mean free drug concentration of only 13.8 ±â€Š1.9 mg/L. No adverse event attributable to temocillin was observed. CONCLUSIONS: Temocillin (6 g daily) given by continuous infusion allows a larger proportion of critically ill patients to have free drug serum concentrations covering infections caused by Enterobacteriaceae with an MIC of 16 mg/L compared with administration three times daily. Clinical efficacy compared with carbapenems in documented severe infections needs to be further studied.

Prediction of ROS1 and TRKA/B/C occupancy in plasma and cerebrospinal fluid for entrectinib alone and in DDIs using physiologically based pharmacokinetic (PBPK) modeling approach.

PURPOSE: Entrectinib (ENT) is a potent c-ros oncogene 1(ROS1) and neurotrophic tyrosine receptor kinase (NTRKA/B/C) inhibitor. To determine the optimum dosage of ENT using ROS1 and NTRKA/B/C occupancy in plasma and cerebrospinal fluid (CSF) in drug-drug interactions (DDIs), physiologically-based pharmacokinetic (PBPK) models for healthy subjects and cancer population were developed for ENT and M5 (active metabolite). METHODS: The PBPK models were built using the modeling parameters of ENT and M5 that were mainly derived from the published paper on the ENT PBPK model, and then validated by the observed pharmacokinetics (PK) in plasma and CSF from healthy subjects and patients. RESULTS: The PBPK model showed that AUC, Cmax, and Ctrough ratios between predictions and observations are within the range of 0.5-2.0, except that the M5 AUC ratio is slightly above 2.0 (2.34). Based on the efficacy (> 75% occupancy for ROS1 and NTRKA/B/C) and safety (AUC < 160 µM·h and Cmax < 8.9 µM), the appropriate dosing regimens were identified. The appropriate dosage is 600 mg once daily (OD) when administered alone, reduced to 200 mg and 400 mg OD with itraconazole and fluconazole, respectively. ENT is not recommended for co-administration with rifampicin or efavirenz, but is permitted with fluvoxamine or dexamethasone. CONCLUSION: The PBPK models can serve as a powerful approach to predict ENT concentration as well as ROS1 and NTRKA/B/C occupancy in plasma and CSF.

Determination of free concentration of endocannabinoids in brain tissue.

The release of metabolites from their bound to free forms is the main regulatory path in living species. Therefore, the ability to determine the free concentrations of small molecules is highly critical in many biological samples. The main challenges in achieving this task are the interferences inherent to complex matrices and the ability to distinguish between the free and total concentrations. This paper presents a non-invasive microextraction method that enables the determination of endocannabinoids in brain tissue. The proposed method is based on two key principles: the availability of the free concentration of endocannabinoids for partitioning to the solid-phase microextraction (SPME) fiber; and negligible depletion enabled by the small volume of extraction phase on the fiber. These features allow the presented SPME method to provide information about the free concentration of analytes without disturbing the binding equilibrium between the analytes and the matrix. The determination of spiked samples with known concentrations enables the percentage of analyte bound to the tissue to be calculated, which can then be applied to calculate the total concentration from the determined free concentration. This manuscript focuses on the determination of the free concentration and tissue binding percentages of endocannabinoids in brain tissue. Significantly, SPME's small size and potential for non-invasive sampling enable its application in live animal subjects with minimal tissue damage.

Protein binding investigation of first-line and second-line antituberculosis drugs.

Data on protein binding are incomplete for first-line antituberculosis drugs, and lacking for second-line antituberculosis drugs that are used extensively for multi-drug-resistant tuberculosis (levofloxacin, linezolid and moxifloxacin). Thus, the main purposes of this study were to investigate: (i) the relationship between carrier protein concentration and drug binding; and (ii) the feasibility of predicting free drug concentration using in-vitro and in-vivo results. In-vitro experiments were performed on spiked plasma mimicking real-case samples (drug combinations from clinical practice). Median in-vivo protein binding was 1.5% for ethambutol, 9.7% for isoniazid, 0.7% for pyrazinamide and 88.2% for rifampicin; and median in-vitro protein binding was 26.2% for levofloxacin, 12.8% for linezolid and 46.3% for moxifloxacin. Albumin concentration (<30 g/L) had a moderate impact on moxifloxacin binding and a strong impact on levofloxacin, linezolid and rifampicin binding. Determination of the free drug concentration seems to be of little value for ethambutol, isoniazid, moxifloxacin and pyrazinamide; limited value for linezolid because of its low binding; and major value for rifampicin in hypoalbuminaemic patients with tuberculosis, and levofloxacin because total concentration was an inaccurate reflection of free concentration. The free concentration predicted by the mathematical model was suitable for levofloxacin and linezolid, whereas the real free concentration should be measured for rifampicin. Further investigations should be carried out to investigate the benefit of using free concentration for levofloxacin, linezolid and rifampicin, particularly in the critical period of active tuberculosis associated with hypoalbuminaemia.

Hypoalbuminemia and Pharmacokinetics: When the Misunderstanding of a Fundamental Concept Leads to Repeated Errors over Decades.

Surprisingly, misinterpretation of the influence of hypoalbuminemia on pharmacokinetics and the clinical effects of drugs seems to be a current problem, even though hypoalbuminemia has no impact on the pharmacologically active exposure. Exceptions to this fact are highly protein-bound anaesthetics with high elimination capacity (i.e., <5 drugs on the market). To assess the frequency of misinterpretation of the influence of hypoalbuminemia on pharmacokinetics and the clinical effects of drugs between 1975 and 2021, a PubMed literature review was conducted. Each paragraph on albumin binding was classified as correct, ambiguous or incorrect, creating two acceptable categories: (1) content without any errors, and (2) content containing some incorrect and/or ambiguous statements. The analyses of these articles showed that fewer than 11% of articles contained no interpretation errors. In order to contain this misinterpretation, several measures are proposed: (1) Make the message accessible to a wide audience by offering a simplified and didactic video representation of the lack of impact of albumin binding to drugs. (2) Precise terminology (unbound/free form/concentration) should be used for highly bound drugs. (3) Unbound/free forms should be systematically quantified for highly plasma protein bound drugs for clinical trials as well as for therapeutic drug monitoring.

Measurement of free fraction, total concentration and protein binding for testosterone, triiodothyronine and thyroxine.

Aims: Measuring the total and free concentrations of hormones is useful, but the technology to do this simultaneously is lacking. Methods: A new method offers the ability to measure these parameters concurrently for testosterone, thyroxine and triiodothyronine. Results: The free concentrations showed significant correlations with patients' vital statistics. Overall, 67% of correlations for total concentration showed that the new and classical methods had equal accuracy, or that comprehensive ultrafiltration was more accurate. The protein binding term was found to correlate significantly with the patients' luteinizing hormone, prostate-specific antigen and height. Conclusion: Comprehensive ultrafiltration for measuring the total concentration, free concentration and protein binding term uses less sample and is much faster than measuring these parameters with three separate methods.

Quantitative determination of plasma free and total concentrations of antivitamin K drugs using a new sensitive and rapid LC-MS/MS method: Application in hemodialysis patients.

BACKGROUND AND AIMS: Vitamin K antagonists (VKAs) are the first-line anticoagulants used in end stage renal disease. This population experiences a significant variability in their International Normalized Ratio (INR) over time. There is a need for methods allowing the study of the pharmacokinetics of free and total concentrations of VKAs to explain INR variability. MATERIALS AND METHODS: We developed and validated a high-performance liquid chromatography-tandem mass spectrometry method allowing the quantification of warfarin and fluindione free and total plasma concentrations. Chromatographic separation was achieved in a raptor biphenyl column and the spectrometry acquisition was set in multiple reaction monitoring mode after negative electrospray ionization. We then applied it in describing the plasma free and total concentrations of VKAs in samples from 50 hemodialysis patients. RESULTS: The developed method is rapid, sensitive and specific. Our cohort results showed a correlation between free and total VKA concentrations. The free VKA concentrations tended to be higher in patients with higher INR. Although VKAs are highly albumin-bound drugs, albumin concentration did not totally explain the high inter-individual total VKA concentrations variability. CONCLUSION: This opens the door to further studies to understand the factors involved in their variability.

[Elucidation and Prediction of Absorption Behavior After Oral Administration of Poorly Water-Soluble Drugs as Different Lipid-Based Formulations].

Lipid-based formulations (LBFs) are isotropic mixtures typically comprising lipids, surfactants, and/or co-solvents, in which drugs are pre-solubilized. After oral administration, LBFs are piggybacked into endogenous lipid digestion pathways. This triggers drug super-saturation and improves absorption. However, super-saturation poses a risk of drug precipitation, which generally leads to poor drug absorption. Furthermore, a series of aqueous colloidal species including digestion products (typically fatty acids and monoglycerides) and endogenous molecules (bile acids and phospholipids) increase the drug solubilization capacity of the intestinal fluid (compared with that of the normal intestinal fluid). However, the solubilization/precipitation behavior may change according to the LBF composition (e.g., the drug loading amount and type of formulation excipients), which may ultimately lead to differences in oral absorption. This review summarizes the results of the evaluation and prediction of the effect of LBFs composition on oral absorption and provides an in-depth understanding of the drug absorption mechanisms when using LBFs.

The impact of quantity of lipid based formulations with different compositions on the oral absorption of ritonavir: A trade-off between apparent solubility and permeability.

In this study, the effect of the quantity of lipid-based formulations (LBFs) on the oral absorption of ritonavir (RTV), a model for poorly water-soluble drugs, was investigated. Two types of LBFs, comprising short- and medium-chain lipids (LBF-SMC) and long-chain lipids (LBF-LC) loaded with different masses of RTV, were prepared. Then, the respective LBFs were dispersed in distilled water at concentrations of 1.0, 2.0, and 3.0% w/w, which provided the same drug concentration for all formulations. When 1.0% LBF-SMC and LBF-LC were orally administered to rats, the oral absorption was significantly improved compared with that of the suspension (a reference formulation) because of enhanced solubilization of RTV in the gastrointestinal tract; however, this improvement was lower for LBF-LC than for LBF-SMC. The oral absorption decreased with increasing LBF concentration for both LBF-SMC and LBF-LC. The in vitro permeation in sequence with in vitro digestion revealed that this phenomenon was caused by a reduction in the free drug concentration in the gastrointestinal tract. Moreover, the effect of decreasing the free concentration was more remarkable for LBF-LC than for LBF-SMC because of the greater solubilization capacity of LC digestion products. These findings may be useful for designing improved drug delivery systems.

Comparison of liquid-liquid extraction, microextraction and ultrafiltration for measuring free concentrations of testosterone and phenytoin.

Aim: The purpose of the study was to find methods suitable for measuring the free concentrations of testosterone and phenytoin. Materials & methods: Sample solutions of the compounds in buffer and human albumin were processed using liquid-liquid extraction, microextraction and ultrafiltration and analyzed by LC-MS/MS. Results: Liquid-liquid extraction with dibutyl phthalate provided complete extraction from buffer solutions and partial extraction from albumin samples. Spintip C18 devices provided exhaustive extraction from buffer and albumin samples. Spintip C8 devices offered complete extraction from buffer and approximately 50% recovery from albumin samples. Centrifree ultrafiltration devices showed high recovery of free concentrations from all the samples, while Amicon and Nanosep devices provided partial recovery. Conclusion: Spintip C8 and Centrifree devices proved useful for measuring free concentrations.

Does Monitoring Total and Free Polymyxin B1 Plasma Concentrations Predict Polymyxin B-Induced Nephrotoxicity? A Retrospective Study in Critically Ill Patients.

INTRODUCTION: The correlation between total and free polymyxin B (PMB including PMB1 and PMB2) exposure in vivo and acute kidney injury (AKI) remains obscure. This study explores the relationships between plasma exposure of PMB1 and PMB2 and nephrotoxicity, and investigates the risk factors for PMB-induced acute kidney injury (AKI) in critically ill patients. METHODS: Critically ill patients who used PMB and met the criteria were enrolled. The total plasma concentration and plasma binding of PMB1 and PMB2 were analysed by liquid chromatography-tandem mass spectrometry and equilibrium dialysis. RESULTS: A total of 89 patients were finally included, and AKI developed in 28.1% of them. The peak concentration of PMB1 (Cmax (B1)) (adjusted odds ratio (AOR) = 1.68, 95% CI 1.08-2.62, p = 0.023), baseline BUN level (AOR = 1.08, 95% CI 1.01-1.16, p = 0.039) and hypertension (AOR = 3.73, 95% CI 1.21-11.54, p = 0.022) were independent risk factors for PMB-induced AKI. The area under the ROC curve of the model was 0.799. When Cmax (B1) was 5.23 µg/ml or more, the probability of AKI was higher than 50%. The ratio of PMB1/PMB2 decreased after PMB preparation entered into the body. The protein binding rate in critically ill patients indicated significant individual differences. Free Cmax (B) and free Cmax (B1) levels in the AKI group were significantly (p < 0.05) higher than those in the non-AKI group. Total and free concentrations of PMB in patients showed a positive correlation. CONCLUSIONS: Both the ROC curve and logistic regression model showed that Cmax (B1) was a good predictor for the probability of PMB-induced AKI. Early therapeutic drug monitoring (TDM) of PMB should be considered in critically ill patients. Compared with Cmin (B), Cmax (B) and Cmax (B1) may be helpful for the early prediction of PMB-induced AKI in critically ill patients.