Free serum concentrations of antibiotics determined by ultrafiltration: extensive evaluation of experimental variables.

Aim: To assess the impact of experimental conditions on free serum concentrations as determined by ultrafiltration and HPLC-DAD analysis in a wide range of antibiotics. Materials & methods: Relative centrifugation force (RCF), temperature, pH and buffer were varied and the results compared with the standard protocol (phosphate buffer pH 7.4, 37°C, 1000 × g). Results: Generally, at 10,000 × g the unbound fraction (fu) decreased with increasing molecular weight, and was lower at 22°C. In unbuffered serum, the fu of flucloxacillin or valproic acid was increased, that of basic or amphoteric drugs considerably decreased. Comparable results were obtained using phosphate or HEPES buffer except for drugs which form metal chelate complexes. Conclusion: Maintaining a physiological pH is more important than strictly maintaining body temperature.

[Box: see text].

Interprofessional Therapeutic Drug Monitoring of Carbapenems Improves ICU Care and Guideline Adherence in Acute-on-Chronic Liver Failure.

(1) Background: Acute-on-chronic liver failure (ACLF) is a severe, rapidly progressing disease in patients with liver cirrhosis. Meropenem is crucial for treating severe infections. Therapeutic drug monitoring (TDM) offers an effective means to control drug dosages, especially vital for bactericidal antibiotics like meropenem. We aimed to assess the outcomes of implementing TDM for meropenem using an innovative interprofessional approach in ACLF patients on a medical intensive care unit (ICU). (2) Methods: The retrospective study was conducted on a medical ICU. The outcomes of an interprofessional approach comprising physicians, hospital pharmacists, and staff nurses to TDM for meropenem in critically ill patients with ACLF were examined in 25 patients. Meropenem was administered continuously via an infusion pump after the application of an initial loading dose. TDM was performed weekly using high-performance liquid chromatography (HPLC). Meropenem serum levels, implementation of the recommendations of the interprofessional team, and meropenem consumption were analyzed. (3) Results: Initial TDM for meropenem showed a mean meropenem serum concentration of 20.9 ± 9.6 mg/L in the 25 analyzed patients. Of note, in the initial TDM, only 16.0% of the patients had meropenem serum concentrations within the respective target range, while 84.0% exceeded this range. Follow-up TDM showed serum concentrations of 15.2 ± 5.7 mg/L (9.0-24.6) in Week 2 and 11.9 ± 2.3 mg/L (10.2-13.5) in Week 3. In Week 2, 41.7% of the patients had meropenem serum concentrations that were within the respective target range, while 58.3% of the patients were above this range. In Week 3, 50% of the analyzed serum concentrations of meropenem were within the targeted range, and 50% were above the range. In total, 100% of the advice given by the interprofessional team regarding meropenem dosing or a change in antibiotic therapy was implemented. During the intervention period, the meropenem application density was 37.9 recommended daily doses (RDD)/100 patient days (PD), compared to 42.1 RDD/100 PD in the control period, representing a 10.0% decrease. (4) Conclusions: Our interprofessional approach to TDM significantly reduced meropenem dosing, with all the team's recommendations being implemented. This method not only improved patient safety but also considerably decreased the application density of meropenem.

Tigecycline Soft Tissue Penetration in Obese and Non-obese Surgical Patients Determined by Using In Vivo Microdialysis.

BACKGROUND AND OBJECTIVE: Tigecycline, a broad-spectrum glycylcycline antibiotic, is approved for use at a fixed dose irrespective of body weight. However, its pharmacokinetics may be altered in obesity, which would impact on the antibiotic's effectiveness. The objective of this study was to investigate the plasma and subcutaneous tissue concentrations of tigecycline in obese patients compared with those in a non-obese control group. METHODS: Fifteen obese patients (one class II and 14 class III) undergoing bariatric surgery and 15 non-obese patients undergoing intra-abdominal surgery (mainly tumour resection) received a single dose of 50 or 100 mg tigecycline as an intravenous short infusion. Tigecycline concentrations were measured up to 8 h after dosing in plasma (total concentration), in ultrafiltrate of plasma (free concentration), and in microdialysate from subcutaneous tissue, respectively. RESULTS: In obese patients, total peak plasma concentration (1.31 ± 0.50 vs 2.27 ± 1.40 mg/L) and the area under the concentration-time curve from 0 to 8 h (AUC8h,plasma: 2.15 ± 0.42 vs 2.74 ± 0.73 h⋅mg/L), as normalized to a 100 mg dose, were significantly lower compared with those of non-obese patients. No significant differences were observed regarding the free plasma concentration, as determined by ultrafiltration, or the corresponding AUC8h (fAUC8h,plasma). Concentrations in interstitial fluid (ISF) of subcutaneous tissue were lower than the free plasma concentrations in both groups, and they were lower in obese compared to non-obese patients: the AUC8h in ISF (AUC8h,ISF) was 0.51 ± 0.22 h⋅mg/L in obese and 0.79 ± 0.23 h⋅mg/L in non-obese patients, resulting in a relative tissue drug exposure (AUC8h,ISF/fAUC8h,plasma) of 0.38 ± 0.19 and 0.63 ± 0.24, respectively. CONCLUSION: Following a single dose of tigecycline, concentrations in the ISF of subcutaneous adipose tissue are decreased in heavily obese subjects, calling for an increased loading dose. EU CLINICAL TRIALS REGISTRATION NUMBER: EudraCT No. 2012-004383-22.

Interprofessional Collaboration between ICU Physicians, Staff Nurses, and Hospital Pharmacists Optimizes Antimicrobial Treatment and Improves Quality of Care and Economic Outcome.

(1) Background: Antibiotic resistance is a worldwide health threat. The WHO published a global strategic plan in 2001 to contain antimicrobial resistance. In the following year, a workshop identified crucial barriers to the implementation of the strategy, e.g., underdeveloped health infrastructures and the scarcity of valid data as well as a lack of implementation of antibiotic stewardship (ABS) programs in medical curricula. Here, we show that interprofessional learning and education can contribute to the optimization of antibiotic use and preserving antibiotic effectiveness. We have initiated interprofessional rounds on a medical intensive care unit (MICU) with a focus on gastroenterology, hepatology, infectious diseases, endocrinology, and liver transplantation. We integrated ICU physicians, hospital pharmacists, nursing staff, and medical students as well as students of pharmacy to broaden the rather technical concept of ABS with an interprofessional approach to conceptualize awareness and behavioral change in antibiotic prescription and use. Methods: Clinical performance data and consumption figures for antibiotics were analyzed over a 10-year period from 2012 to 2021. The control period covered the years 2012-2014. The intervention period comprised the years 2015-2021, following the implementation of an interprofessional approach to ABS at a MICU of a German university hospital. Data from the hospital pharmacy, hospital administration, and hospital information system were included in the analyses. A specific electronic platform was developed for the optimization of documentation, interprofessional learning, education, and sustainability. The years 2020 and 2021 were analyzed independently due to the SARS-CoV-2 pandemic and the care of numerous COVID-19 patients at the MICU. Results: Implementation of an interprofessional ABS program resulted in the optimization of antibiotic management at the MICU. The suggestions of the hospital pharmacist for optimization can be divided into the following categories (i) indication for and selection of therapy (43.6%), (ii) optimization of dosing (27.6%), (iii) drug interactions (9.4%), (iv) side effects (4.1%), and (v) other pharmacokinetic, pharmacodynamic, and pharmacoeconomic topics (15.3%). These suggestions were discussed among the interprofessional team at the MICU; 86.1% were consequently implemented and the prescription of antibiotics was changed. In addition, further analysis of the intensive care German Diagnosis Related Groups (G-DRGs) showed that the case mix points increased significantly by 31.6% during the period under review. Accordingly, the severity of illness of the patients treated at the ICU as measured by the Simplified Acute Physiology Score (SAPS) II increased by 21.4% and the proportion of mechanically ventilated patients exceeded 50%. Antibiotic spending per case mix point was calculated. While spending was EUR 60.22 per case mix point in 2015, this was reduced by 42.9% to EUR 34.37 per case mix point by 2019, following the implementation of the interprofessional ABS program on the MICU. Through close interprofessional collaboration between physicians, hospital pharmacists, and staff nurses, the consumption of broad-spectrum antibiotics, e.g., carbapenems, was significantly reduced, thus improving patient care. In parallel, the case mix and case mix index increased. Thus, the responsible use of resources and high-performance medicine are not contradictory. In our view, close interprofessional and interdisciplinary collaboration between physicians, pharmacists, and nursing staff will be of outstanding importance in the future to prepare health care professionals for global health care to ensure that the effectiveness of our antibiotics is preserved.

Similar Piperacillin/Tazobactam Target Attainment in Obese versus Nonobese Patients despite Differences in Interstitial Tissue Fluid Pharmacokinetics.

Precision dosing of piperacillin/tazobactam in obese patients is compromised by sparse information on target-site exposure. We aimed to evaluate the appropriateness of current and alternative piperacillin/tazobactam dosages in obese and nonobese patients. Based on a prospective, controlled clinical trial in 30 surgery patients (15 obese/15 nonobese; 0.5-h infusion of 4 g/0.5 g piperacillin/tazobactam), piperacillin pharmacokinetics were characterized in plasma and at target-site (interstitial fluid of subcutaneous adipose tissue) via population analysis. Thereafter, multiple 3-4-times daily piperacillin/tazobactam short-term/prolonged (recommended by EUCAST) and continuous infusions were evaluated by simulation. Adequacy of therapy was assessed by probability of pharmacokinetic/pharmacodynamic target-attainment (PTA ≥ 90%) based on time unbound piperacillin concentrations exceed the minimum inhibitory concentration (MIC) during 24 h (%fT>MIC). Lower piperacillin target-site maximum concentrations in obese versus nonobese patients were explained by the impact of lean (approximately two thirds) and fat body mass (approximately one third) on volume of distribution. Simulated steady-state concentrations were 1.43-times, 95%CI = (1.27; 1.61), higher in plasma versus target-site, supporting targets of %fT>2×MIC instead of %fT>4×MIC during continuous infusion to avoid target-site concentrations constantly below MIC. In all obesity and renally impairment/hyperfiltration stages, at MIC = 16 mg/L, adequate PTA required prolonged (thrice-daily 4 g/0.5 g over 3.0 h at %fT>MIC = 50) or continuous infusions (24 g/3 g over 24 h following loading dose at %fT>MIC = 98) of piperacillin/tazobactam.

Perioperative administration of cefazolin and metronidazole in obese and non-obese patients: a pharmacokinetic study in plasma and interstitial fluid.

OBJECTIVES: To assess plasma and tissue pharmacokinetics of cefazolin and metronidazole in obese patients undergoing bariatric surgery and non-obese patients undergoing intra-abdominal surgery. PATIENTS AND METHODS: Fifteen obese and 15 non-obese patients received an IV short infusion of 2 g cefazolin and 0.5 g metronidazole for perioperative prophylaxis. Plasma and microdialysate from subcutaneous tissue were sampled until 8 h after dosing. Drug concentrations were determined by HPLC-UV. Pharmacokinetic parameters were calculated non-compartmentally. RESULTS: In obese patients (BMI 39.5-69.3 kg/m2) compared with non-obese patients (BMI 18.7-29.8 kg/m2), mean Cmax of total cefazolin in plasma was lower (115 versus 174 mg/L) and Vss was higher (19.4 versus 14.2 L). The mean differences in t½ (2.7 versus 2.4 h), CL (5.14 versus 4.63 L/h) and AUC∞ (402 versus 450 mg·h/L) were not significant. The influence of obesity on the pharmacokinetics of metronidazole was similar (Cmax 8.99 versus 14.7 mg/L, Vss 73.9 versus 51.8 L, t½ 11.9 versus 9.1 h, CL 4.62 versus 4.13 L/h, AUC∞ 116 versus 127 mg·h/L). Regarding interstitial fluid (ISF), mean concentrations of cefazolin remained >4 mg/L until 6 h in both groups, and those of metronidazole up to 8 h in the non-obese group. In obese patients, the mean ISF concentrations of metronidazole were between 3 and 3.5 mg/L throughout the measuring interval. CONCLUSIONS: During the time of surgery, cefazolin concentrations in plasma and ISF of subcutaneous tissue were lower in obese patients, but not clinically relevant. Regarding metronidazole, the respective differences were higher, and may influence dosing of metronidazole for perioperative prophylaxis in obese patients.

Measurement of Free Plasma Concentrations of Beta-Lactam Antibiotics: An Applicability Study in Intensive Care Unit Patients.

BACKGROUND: The antibacterial effect of antibiotics is linked to the free drug concentration. This study investigated the applicability of an ultrafiltration method to determine free plasma concentrations of beta-lactam antibiotics in ICU patients. METHODS: Eligible patients included adult ICU patients treated with ceftazidime (CAZ), meropenem (MEM), piperacillin (PIP)/tazobactam (TAZ), or flucloxacillin (FXN) by continuous infusion. Up to 2 arterial blood samples were drawn at steady state. Patients could be included more than once if they received another antibiotic. Free drug concentrations were determined by high-performance liquid chromatography with ultraviolet detection after ultrafiltration, using a method that maintained physiological conditions (pH 7.4/37°C). Total drug concentrations were determined to calculate the unbound fraction. In a post-hoc analysis, free concentrations were compared with the target value of 4× the epidemiological cut-off value (ECOFF) for Pseudomonas aeruginosa as a worst-case scenario for empirical therapy with CAZ, MEM or PIP/tazobactam and against methicillin-sensitive Staphylococcus aureus for targeted therapy with FXN. RESULTS: Fifty different antibiotic treatment periods in 38 patients were evaluated. The concentrations of the antibiotics showed a wide range because of the fixed dosing regimen in a mixed population with variable kidney function. The mean unbound fractions (fu) of CAZ, MEM, and PIP were 102.5%, 98.4%, and 95.7%, with interpatient variability of <6%. The mean fu of FXN was 11.6%, with interpatient variability of 39%. It was observed that 2 of 12 free concentrations of CAZ, 1 of 40 concentrations of MEM, and 11 of 23 concentrations of PIP were below the applied target concentration of 4 × ECOFF for P. aeruginosa. All concentrations of FXN (9 samples from 6 patients) were >8 × ECOFF for methicillin-sensitive Staphylococcus aureus. CONCLUSIONS: For therapeutic drug monitoring purposes, measuring total or free concentrations of CAZ, MEM, or PIP is seemingly adequate. For highly protein-bound beta-lactams such as FXN, free concentrations should be favored in ICU patients with prevalent hypoalbuminemia.

Quantification of microdialysis related variability in humans: Clinical trial design recommendations.

OBJECTIVE: Target-site concentrations obtained via the catheter-based minimally invasive microdialysis technique often exhibit high variability. Catheter calibration is commonly performed via retrodialysis, in which a transformation factor, termed relative recovery (RR), is determined. Leveraging RR values from a rich data set of a very large clinical microdialysis study, promised to contribute critical insight into the origin of the reportedly high target-site variability. The present work aimed (i) to quantify and explain variability in RR associated with the patient (including non-obese vs. obese) and the catheter, and (ii) to derive recommendations on the design of future clinical microdialysis studies. METHODS: A prospective, age- and sex-matched parallel group, single-centre trial in non-obese and obese patients (BMI=18.7-86.9 kg/m2) was performed. 1-3 RR values were obtained in the interstitial fluid of the subcutaneous fat tissue in one catheter per upper arm of 120 patients via the retrodialysis method (nRR=1008) for a panel of drugs (linezolid, meropenem, tigecycline, cefazolin, fosfomycin, piperacillin and acetaminophen). A linear mixed-effects model was developed to quantify the different types of variability in RR and to explore the association between RR and patient body size descriptors. RESULTS: Estimated RR was highest for acetaminophen (69.7%, 95%CI=65.0% to 74.3%) and lowest for piperacillin (40.4%, 95%CI=34.6% to 46.0%). The linear mixed-effects modelling analysis showed that variability associated with the patient (σ=15.9%) was the largest contributor (46.7%) to overall variability, whereas the contribution of variability linked to the catheter (σ=5.55%) was ~1/6 (16.8%). The relative contribution of residual unexplained variability (σ=12.0%, including intracatheter variability) was ~1/3 (36.4%). The limits of agreement of repeated RR determinations in a single catheter ranged from 0.694-1.64-fold (linezolid) to 0.510-3.02-fold (cefazolin). Calculated fat mass affected RR, explaining the observed lower RR in obese (ΔRRmean= -29.7% relative reduction) versus non-obese patients (p<0.001); yet only 15.8% of interindividual variability was explained by this effect. No difference in RR was found between catheters implanted into the left or right arm (p=0.732). CONCLUSIONS: Three recommendations for clinical microdialysis trial design were derived: 1) High interindividual variability underscored the necessity of measuring individual RR per patient. 2) The low relative contribution of intercatheter variability to overall variability indicated that measuring RR with a single catheter per patient is sufficient for reliable catheter calibration. 3) The wide limits of agreement from multiple RR in the same catheter implied an uncertainty of a factor of two in target-site drug concentration estimation necessitating to perform catheter calibration (retrodialysis sampling) multiple times per patient. To allow routine clinical use of microdialysis, research efforts should aim at further understanding and minimising the method-related variability. Optimised study designs in clinical trials will ultimately yield more informative microdialysis data and increase our understanding of this valuable sampling technique to derive target-site drug exposure.

Meropenem Plasma and Interstitial Soft Tissue Concentrations in Obese and Nonobese Patients-A Controlled Clinical Trial.

BACKGROUND: This controlled clinical study aimed to investigate the impact of obesity on plasma and tissue pharmacokinetics of meropenem. METHODS: Obese (body mass index (BMI) ≥ 35 kg/m2) and age-/sex-matched nonobese (18.5 kg/m2 ≥ BMI ≤ 30 kg/m2) surgical patients received a short-term infusion of 1000-mg meropenem. Concentrations were determined via high performance liquid chromatography-ultraviolet (HPLC-UV) in the plasma and microdialysate from the interstitial fluid (ISF) of subcutaneous tissue up to eight h after dosing. An analysis was performed in the plasma and ISF by noncompartmental methods. RESULTS: The maximum plasma concentrations in 15 obese (BMI 49 ± 11 kg/m2) and 15 nonobese (BMI 24 ± 2 kg/m2) patients were 54.0 vs. 63.9 mg/L (95% CI for difference: -18.3 to -3.5). The volume of distribution was 22.4 vs. 17.6 L, (2.6-9.1), but the clearance was comparable (12.5 vs. 11.1 L/h, -1.4 to 3.1), leading to a longer half-life (1.52 vs. 1.31 h, 0.05-0.37) and fairly similar area under the curve (AUC)8h (78.7 vs. 89.2 mg*h/L, -21.4 to 8.6). In the ISF, the maximum concentrations differed significantly (12.6 vs. 18.6 L, -16.8 to -0.8) but not the AUC8h (28.5 vs. 42.0 mg*h/L, -33.9 to 5.4). Time above the MIC (T > MIC) in the plasma and ISF did not differ significantly for MICs of 0.25-8 mg/L. CONCLUSIONS: In morbidly obese patients, meropenem has lower maximum concentrations and higher volumes of distribution. However, due to the slightly longer half-life, obesity has no influence on the T > MIC, so dose adjustments for obesity seem unnecessary.

Brain Exposure to Piperacillin in Acute Hemorrhagic Stroke Patients Assessed by Cerebral Microdialysis and Population Pharmacokinetics.

BACKGROUND: The broad antibacterial spectrum of piperacillin/tazobactam makes the combination suitable for the treatment of nosocomial bacterial central nervous system (CNS) infections. As limited data are available regarding piperacillin CNS exposure in patients without or with low-grade inflammation, a clinical study was conducted (1) to quantify CNS exposure of piperacillin by cerebral microdialysis and (2) to evaluate different dosing regimens in order to improve probability of target attainment (PTA) in brain. METHODS: Ten acute hemorrhagic stroke patients (subarachnoid hemorrhage, n = 6; intracerebral hemorrhage, n = 4) undergoing multimodality neuromonitoring received 4 g piperacillin/0.5 g tazobactam every 8 h by 30-min infusions for the management of healthcare-associated pneumonia. Cerebral microdialysis was performed as part of the clinical neuromonitoring routine, and brain interstitial fluid samples were retrospectively analyzed for piperacillin concentrations after the first and after multiple doses for at least 5 days and quantified by high-performance liquid chromatography. Population pharmacokinetic modeling and Monte Carlo simulations with various doses and types of infusions were performed to predict exposure. A T>MIC of 50% was selected as pharmacokinetic/pharmacodynamic target parameter. RESULTS: Median peak concentrations of unbound piperacillin in brain interstitial space fluid were 1.16 (range 0.08-3.59) and 2.78 (range 0.47-7.53) mg/L after the first dose and multiple doses, respectively. A one-compartment model with a transit compartment and a lag time (for the first dose) between systemic and brain exposure was appropriate to describe the brain concentrations. Bootstrap median estimates of the parameters were: transfer rate from plasma to brain (0.32 h-1), transfer rate from brain to plasma (7.31 h-1), and lag time [2.70 h (coefficient of variation 19.7%)]. The simulations suggested that PTA would exceed 90% for minimum inhibitory concentrations (MICs) up to 0.5 mg/L and 1 mg/L at a dose of 12-16 and 24 g/day, respectively, regardless of type of infusion. For higher MICs, PTA dropped significantly. CONCLUSION: Limited CNS exposure of piperacillin might be an obstacle in treating patients without general meningeal inflammation except for infections with highly susceptible pathogens. Brain exposure of piperacillin did not improve significantly with a prolongation of infusions.

Plasma and tissue pharmacokinetics of fosfomycin in morbidly obese and non-obese surgical patients: a controlled clinical trial.

OBJECTIVES: To assess the pharmacokinetics and tissue penetration of fosfomycin in obese and non-obese surgical patients. METHODS: Fifteen obese patients undergoing bariatric surgery and 15 non-obese patients undergoing major intra-abdominal surgery received an intravenous single short infusion of 8 g of fosfomycin. Fosfomycin concentrations were determined by LC-MS/MS in plasma and microdialysate from subcutaneous tissue up to 8 h after dosing. The pharmacokinetic analysis was performed in plasma and interstitial fluid (ISF) by non-compartmental methods. RESULTS: Thirteen obese patients (BMI 38-50 kg/m2) and 14 non-obese patients (BMI 0-29 kg/m2) were evaluable. The pharmacokinetics of fosfomycin in obese versus non-obese patients were characterized by lower peak plasma concentrations (468 ±âŸ139 versus 594 ±âŸ149 mg/L, P = 0.040) and higher V (24.4 ±âŸ6.4 versus 19.0 ±âŸ3.1 L, P = 0.010). The differences in AUC∞ were not significant (1275 ±âŸ477 versus 1515 ±âŸ352 mg·h/L, P = 0.16). The peak concentrations in subcutaneous tissue were reached rapidly and declined in parallel with the plasma concentrations. The drug exposure in tissue was nearly halved in obese compared with non-obese patients (AUC∞ 1052 ±âŸ394 versus 1929 ±âŸ725 mg·h/L, P = 0.0010). The tissue/plasma ratio (AUCISF/AUCplasma) was 0.86 ±âŸ0.32 versus 1.27 ±âŸ0.34 (P = 0.0047). CONCLUSIONS: Whereas the pharmacokinetics of fosfomycin in plasma of surgical patients were only marginally different between obese and non-obese patients, the drug exposure in subcutaneous tissue was significantly lower in the obese patients.

Determination of total or free cefazolin and metronidazole in human plasma or interstitial fluid by HPLC-UV for pharmacokinetic studies in man.

Cefazolin (CFZ) plus metronidazole (MTZ) is commonly used for perioperative antibiotic prophylaxis. An HPLC-UV method is described for the simultaneous determination of total or free cefazolin and metronidazole in human plasma or in microdialysate of subcutaneous tissue. Separation was performed isocratically using a reversed phase column and phosphate buffer/acetonitrile as mobile phase. The validation characteristics were similar for both drugs. Linearity has been shown down to 0.1 mg/L (R > 0.9990). Intra- and inter-assay precision (CV) and in-accuracy were < 5%. The method was applied to the determination of cefazolin and metronidazole in plasma and microdialysate of surgical patients following 30-min intravenous infusion of cefazolin/metronidazole 2.0/0.5 g.

Determination of total and free ceftolozane and tazobactam in human plasma and interstitial fluid by HPLC-UV.

Ceftolozane/tazobactam is a new cephalosporin/beta-lactamase inhibitor combination. An HPLC-UV method is described for the determination of total and free ceftolozane and tazobactam in human plasma and in microdialysate of subcutaneous tissue, respectively. Separation was performed using a reversed-phase column with phosphate buffer/acetonitrile as eluent and photometric detection at 260 nm (ceftolozane) or 220 nm (tazobactam). Linearity has been shown down to ceftolozane/tazobactam 0.1/0.05 mg/L in plasma and 0.03/0.015 mg/L in saline, respectively. The plasma protein binding of both drugs as determined by ultrafiltration was less than 10%. Temperature, pH or relative centrifugation force (up to 3000 x g) had no significant impact on the protein binding. The method was applied to the determination of ceftolozane and tazobactam in plasma and interstitial fluid of healthy volunteers following intravenous infusion of ceftolozane/tazobactam 1.0/0.5 g.

Drug combinations and impact of experimental conditions on relative recovery in in vitro microdialysis investigations.

The need for pharmacokinetic knowledge about antibiotics directly at the site of infection, typically the interstitial space fluid (ISF) of tissues, is gaining acceptance for effective and safe treatment. One option to acquire such data is the microdialysis technique employing a catheter with a semipermeable membrane inserted directly in the ISF. A prerequisite is catheter calibration, e.g. via retrodialysis, yielding a conversion factor from measured to true ISF concentrations, termed relative recovery. This value can be influenced by various factors. The present investigation assessed the impact of three of them on relative recovery using seven drugs: (I) drug combinations/order, (II) air in the microdialysis system, (III) flow rate changes inherent when using common in vivo microdialysis pumps. All experiments were performed in a standardised in vitro microdialysis system. (I) Relative recovery of single antibiotics (linezolid, meropenem, cefazolin, metronidazole, tigecycline) was determined in microdialysis and retrodialysis settings and compared with values using either antibiotic or antibiotic+analgesic (acetaminophen and metamizole) combinations or single drugs with reversed microdialysis order. For assessing these factors for lower relative recovery values (as in in vivo), these were mimicked by increasing the flow rate for linezolid. (II) For the impact of air, linezolid relative recovery of freshly carbonated solutions was compared to degassed ones in microdialysis and retrodialysis settings. For each condition in (I) and (II), summary statistics of relative recovery were calculated and for the impact of the factors a linear mixed-effect model developed. (III) From samples taken during an automatic flush sequence (15 µL/min) of an in vivo pump and afterwards switching to the flow rate of 1 and 2 µL/min for 120 min, the time necessary for relative recovery to reach equilibrium was determined. (I) High relative recovery values (flow rate 2 µL/min: ≥84%; flow rate 5 µL/min: ≥65%) were observed for all investigated single drugs. Intra- and intercatheter variability ranged from 0.3%-11% and 3%-25%, respectively. Based on these values and on the statistical model, the impact of drug combination versus single drug as well as of reversed order was small with changes in relative recovery of smaller equal 9%. (II) Compared to degassed solutions, relative recovery in carbonated solutions was 23% and 19% lower (relative reduction) in the microdialysis and retrodialysis setting, respectively, with increased intercatheter variability (up to 37%). (III) As expected, relative recovery increased after the flush sequence and was constant 10-15 min after the switch to the typical 1 and 2 µL/min flow rate. Given the intercatheter variability, combinations and the order of drugs showed minor but clinically negligible impact on relative recovery. In contrast, air in the microdialysis catheter/system caused falsely low and inconsistent relative recovery values and must be avoided when performing a trial. Also changes in flow rate at the end of pump flush sequence impacted relative recovery. Hence, a sufficient equilibration time of 10-15 min prior to sampling should be implemented in sampling protocols. In vitro microdialysis presents a highly valuable complementary platform to clinical microdialysis studies impacting the design, sampling schedule and data analysis of such trials to gain knowledge of target-site pharmacokinetics for contributing to better informed decisions in the individual patient/special populations in future.

Impact of Experimental Variables on the Protein Binding of Tigecycline in Human Plasma as Determined by Ultrafiltration.

Tigecycline, a tetracycline derivative, shows atypical plasma protein binding behavior. The unbound fraction decreases with increasing concentration at therapeutic concentrations. Moreover, uncertainty exists about the magnitude of tigecyline's protein binding in man. Unbound fractions between 2.5% and 35% have been reported in plasma from healthy volunteers, and between 25% and 100% in patients, respectively. In the present study, the protein binding of tigecycline has been investigated by ultrafiltration using different experimental conditions. Whereas temperature had only a marginal influence, the unbound fraction at 0.3/3.0 mg/L was low at pH 8.2 (9.4%/1.9%) or in unbuffered pooled plasma (6.3%/1.2%), compared with plasma buffered with HEPES to pH 7.4 (65.9%/39.7%). In experiments with phosphate buffer and EDTA, the concentration dependency was markedly attenuated or abolished, which is compatible with a cooperative binding mechanism involving divalent cations such as calcium. The unbound fraction in clinical plasma samples from patients treated with tigecycline was determined to 66.3 ± 13.7% at concentrations <0.3 mg/L compared with 41.3 ± 16.0% at >1 to <5 mg/L. To summarize, tigecycline appears to be only moderately bound to plasma proteins as determined by ultrafiltration, when a physiological pH is maintained.

Unbound fraction of fluconazole and linezolid in human plasma as determined by ultrafiltration: Impact of membrane type.

Ultrafiltration is a rapid and convenient method to determine the free concentrations of drugs in plasma. Several ultrafiltration devices based on Eppendorf cups are commercially available, but are not validated for such use by the manufacturer. Plasma pH, temperature and relative centrifugal force as well as membrane type can influence the results. In the present work, we developed an ultrafiltration method in order to determine the free concentrations of linezolid or fluconazole, both neutral and moderately lipophilic antiinfective drugs for parenteral as well as oral administration, in plasma of patients. Whereas both substances behaved relatively insensitive in human plasma regarding variations in pH (7.0-8.5), temperature (5-37°C) or relative centrifugal force (1000-10.000xg), losses of linezolid were observed with the Nanosep Omega device due to adsorption onto the polyethersulfone membrane (unbound fraction 75% at 100mg/L and 45% at 0.1mg/L, respectively). No losses were observed with Vivacon which is equipped with a membrane of regenerated cellulose. With fluconazole no differences between Nanosep and Vivacon were observed. Applying standard conditions (pH 7.4/37°C/1000xg/20min), the mean unbound fraction of linezolid in pooled plasma from healthy volunteers was 81.5±2.8% using Vivacon, that of fluconazole was 87.9±3.5% using Nanosep or 89.4±3.3% using Vivacon. The unbound fraction of linezolid was 85.4±3.7% in plasma samples from surgical patients and 92.1±6.2% in ICU patients, respectively. The unbound fraction of fluconazole was 93.9±3.3% in plasma samples from ICU patients.

Variable Linezolid Exposure in Intensive Care Unit Patients-Possible Role of Drug-Drug Interactions.

BACKGROUND: Standard doses of linezolid may not be suitable for all patient groups. Intensive care unit (ICU) patients in particular may be at risk of inadequate concentrations. This study investigated variability of drug exposure and its potential sources in this population. METHODS: Plasma concentrations of linezolid were determined by high-performance liquid chromatography in a convenience sample of 20 ICU patients treated with intravenous linezolid 600 mg twice daily. Ultrafiltration applying physiological conditions (pH 7.4/37°C) was used to determine the unbound fraction. Individual pharmacokinetic (PK) parameters were estimated by population PK modeling. As measures of exposure to linezolid, area under the concentration-time curve (AUC) and trough concentrations (Cmin) were calculated and compared with published therapeutic ranges (AUC 200-400 mg*h/L, Cmin 2-10 mg/L). Coadministered inhibitors or inducers of cytochrome P450 and/or P-glycoprotein were noted. RESULTS: Data from 18 patients were included into the PK evaluation. Drug exposure was highly variable (median, range: AUC 185, 48-618 mg*h/L, calculated Cmin 2.92, 0.0062-18.9 mg/L), and only a minority of patients had values within the target ranges (6 and 7, respectively). AUC and Cmin were linearly correlated (R = 0.98), and classification of patients (underexposed/within therapeutic range/overexposed) according to AUC or Cmin was concordant in 15 cases. Coadministration of inhibitors was associated with a trend to higher drug exposure, whereas 3 patients treated with levothyroxine showed exceedingly low drug exposure (AUC ∼60 mg*h/L, Cmin <0.4 mg/L). The median unbound fraction in all 20 patients was 90.9%. CONCLUSIONS: Drug exposure after standard doses of linezolid is highly variable and difficult to predict in ICU patients, and therapeutic drug monitoring seems advisable. PK drug-drug interactions might partly be responsible and should be further investigated; protein binding appears to be stable and irrelevant.

Protein binding characteristics and pharmacokinetics of ceftriaxone in intensive care unit patients.

AIMS: The aim of the present study was to assess the pharmacokinetics of total and unbound ceftriaxone in intensive care unit (ICU) patients and its protein binding characteristics. METHODS: Twenty patients (m/f 15/5, age 25-86 years, body weight 60-121 kg, APACHE II 7-40, estimated glomerular filtration rate 19-157 ml min(-1) , albumin 11.7-30.1 g l(-1) , total bilirubin <0.1-36.1 mg dl(-1) ) treated with intravenous ceftriaxone were recruited from two ICUs. Timed plasma samples were obtained using an opportunistic study protocol. Ceftriaxone concentrations were determined by high-performance liquid chromatography; unbound concentrations were determined after ultrafiltration using a new method which maintains physiological pH and temperature. The pharmacokinetics was described by a one-compartment model, the protein-binding characteristics by Michaelis-Menten kinetics. RESULTS: For total drug, the volume of distribution was 20.2 l (median; interquartile range 15.6-24.5 l), the half-life 14.5 h (10.0-25.5 h) and the clearance 0.96 l h(-1) (0.55-1.28 l h(-1) ). The clearance of unbound drug was 1.91 l h(-1) (1.46-6.20 l h(-1) ) and linearly correlated with estimated glomerular filtration rate (slope 0.85, y-intercept 0.24 l h(-1) , r(2) = 0.70). The unbound fraction was higher in ICU patients (33.0%; 20.2-44.5%) than reported in healthy volunteers, particularly when renal impairment or severe hyperbilirubinaemia was present. In all patients, unbound concentrations during treatment with ceftriaxone 2 g once daily remained above the EUCAST susceptibility breakpoint (≤1 mg l(-1) ) throughout the whole dosing interval. CONCLUSIONS: Protein binding of ceftriaxone is reduced and variable in ICU patients due to hypoalbuminaemia, but also to altered binding characteristics. Despite these changes, the pharmacokinetics of unbound ceftriaxone is governed by renal function. For patients with normal or reduced renal function, standard doses are sufficient.