Degree Of Plasma Protein Binding Research Articles

Distribution of quetiapine between serum and whole blood in therapeutic drug-monitoring specimens.

Quetiapine use is on the rise, leading to a corresponding increase in acute intoxications, some of which have fatal outcomes. When assessing whole-blood quetiapine concentrations during forensic autopsies, interpretations are primarily based on toxicity data from studies of serum concentrations. To our knowledge, there are only two previous studies that have attempted to establish the ratio between whole blood and serum quetiapine concentrations with limited populations and high variability of results. Paired specimens of whole blood and serum from 16 quetiapine users recruited from the Psychiatric Clinic, St. Olav University Hospital were analyzed using LC-MS-MS. Quetiapine concentrations in both matrices were determined and compared. The mean blood:serum ratio of quetiapine was 0.74 (standard deviation (SD) = 0.05, 95% confidence interval (CI) 0.71-0.76, P < 0.001), range 0.66-0.85. Simple linear regression showed strong linear correlation between quetiapine concentrations in the two matrices (B = 0.774, P > 0.001, r = 0.999). Our results imply that quetiapine occurs at lower concentrations within erythrocytes than in plasma. This is most likely due to a high degree of plasma protein binding. Other factors which may influence the distribution of quetiapine between these compartments are solubility, metabolism and passive or active efflux mechanisms. We did not observe any covariation between blood:serum ratios and serum concentrations. Quetiapine was consistently present at lower concentrations in whole blood than in serum. If so inclined to, a conversion factor of ∼0.7 may be considered for extrapolation of concentrations from serum to whole blood, at least in cases with therapeutic quetiapine concentration levels.

Improving In Vitro-In Vivo Extrapolation of Clearance Using Rat Liver Microsomes for Highly Plasma Protein-Bound Molecules.

It is common practice in drug discovery and development to predict in vivo hepatic clearance from in vitro incubations with liver microsomes or hepatocytes using the well-stirred model (WSM). When applying the WSM to a set of approximately 3000 Novartis research compounds, 73% of neutral and basic compounds (extended clearance classification system [ECCS] class 2) were well-predicted within 3-fold. In contrast, only 44% (ECCS class 1A) or 34% (ECCS class 1B) of acids were predicted within 3-fold. To explore the hypothesis whether the higher degree of plasma protein binding for acids contributes to the in vitro-in vivo correlation (IVIVC) disconnect, 68 proprietary compounds were incubated with rat liver microsomes in the presence and absence of 5% plasma. A minor impact of plasma on clearance IVIVC was found for moderately bound compounds (fraction unbound in plasma [fup] ≥1%). However, addition of plasma significantly improved the IVIVC for highly bound compounds (fup <1%) as indicated by an increase of the average fold error from 0.10 to 0.36. Correlating fup with the scaled unbound intrinsic clearance ratio in the presence or absence of plasma allowed the establishment of an empirical, nonlinear correction equation that depends on fup Taken together, estimation of the metabolic clearance of highly bound compounds was enhanced by the addition of plasma to microsomal incubations. For standard incubations in buffer only, application of an empirical correction provided improved clearance predictions. SIGNIFICANCE STATEMENT: Application of the well-stirred liver model for clearance in vitro-in vivo extrapolation (IVIVE) in rat generally underpredicts the clearance of acids and the strong protein binding of acids is suspected to be one responsible factor. Unbound intrinsic in vitro clearance (CLint,u) determinations using rat liver microsomes supplemented with 5% plasma resulted in an improved IVIVE. An empirical equation was derived that can be applied to correct CLint,u-values in dependance of fraction unbound in plasma (fup) and measured CLint in buffer.

CEFOVECIN PROTEIN BINDING AS A PREDICTOR FOR EXTENDED DURATION OF ACTION: A REVIEW OF CURRENT LITERATURE AND IN VITRO ANALYSIS IN MULTIPLE ZOOLOGICAL SPECIES.

Cefovecin is a third-generation cephalosporin antibiotic with an efficacy of 2 wk following a single injection in domestic dogs and cats. A high degree of plasma protein binding to cefovecin has been proposed as one of the mechanisms responsible for the long elimination half-life, but protein binding has not been evaluated extensively in nondomestic species. In this study, a review of the current literature was conducted, and pharmacokinetic data were compiled for species in which cefovecin has been evaluated thus far. Additionally, in vitro cefovecin protein binding was evaluated in plasma from 22 nondomestic species representing a broad range of taxa. Animals of the order Carnivora demonstrated protein-binding levels of >98%, which is supportive of the long elimination half-life seen in related species. Protein binding was highly variable in Artiodactyl and Perissodactyl species, with dolphins (Tursiops truncatus) and barasingha (Rucervus duvaucelii) displaying high protein binding (99.12% to >99%), but not gazelles (Eudorcas thomsonii) or equids (91.76-92.70%). Cefovecin was not highly bound in any reptiles or birds, corresponding to short half-lives reported for these taxa. These results suggest that a high percentage of plasma protein binding in vitro may predict in which species cefovecin may exhibit a long half-life and duration of action in vivo. These findings may aid in selecting species for cefovecin pharmacokinetic research and for empirical treatment of infections caused by susceptible bacteria.

Lack of Correlation between In Vitro and In Vivo Studies on the Inhibitory Effects of (‒)-Sophoranone on CYP2C9 is Attributable to Low Oral Absorption and Extensive Plasma Protein Binding of (‒)-Sophoranone.

(‒)-Sophoranone (SPN) is a bioactive component of Sophora tonkinensis with various pharmacological activities. This study aims to evaluate its in vitro and in vivo inhibitory potential against the nine major CYP enzymes. Of the nine tested CYPs, it exerted the strongest inhibitory effect on CYP2C9-mediated tolbutamide 4-hydroxylation with the lowest IC50 (Ki) value of 0.966 ± 0.149 μM (0.503 ± 0.0383 μM), in a competitive manner. Additionally, it strongly inhibited other CYP2C9-catalyzed diclofenac 4′-hydroxylation and losartan oxidation activities. Upon 30 min pre-incubation of human liver microsomes with SPN in the presence of NADPH, no obvious shift in IC50 was observed, suggesting that SPN is not a time-dependent inactivator of the nine CYPs. However, oral co-administration of SPN had no significant effect on the pharmacokinetics of diclofenac and 4′-hydroxydiclofenac in rats. Overall, SPN is a potent inhibitor of CYP2C9 in vitro but not in vivo. The very low permeability of SPN in Caco-2 cells (Papp value of 0.115 × 10−6 cm/s), which suggests poor absorption in vivo, and its high degree of plasma protein binding (>99.9%) may lead to the lack of in vitro–in vivo correlation. These findings will be helpful for the safe and effective clinical use of SPN.

The correlation of plasma protein binding and molecular properties of selected antifungal drugs

Antifungal agents are the group of drugs commonly prescribed in the treatment of fungal infections, which are widely spread among the global population. Their properties, such as absorption, distribution, metabolism, route of elimination or plasma protein binding (PPB), considerably influence their therapeutic success, while a number of the molecular physicochemical properties of the drug notably influence all these processes. Lipophilicity (log P), molecular weight (Mw), volume (Vol), polar surface area (PSA) and solubility (log S) play important roles in drug absorption, penetration into tissues, distribution and route of elimination or the degree of plasma protein binding. In this study, the relationships between these five molecular properties of eight antifungal drugs and their plasma protein binding data obtained from relevant literature were investigated. The Selected physicochemical molecular descriptors of the drug were calculated using software packages. The established relationships between PPB and PSA; Mw; Vol and log S were showed relatively poor correlation (r &lt; 0.35). The best correlation was obtained for the relationship between PPB data and the lipophilicity descriptor X log P3 (correlation coefficient r = 0.55). In further investigation, multiple linear regression analysis was applied. The best correlation was obtained with application of lipophilicity with polar surface area (r = 0.918) and volume (r = 0.916) or molecular weight (r = 0.896) as independent variables.

Improved Ultrafiltration Method to Measure Drug Release from Nanomedicines Utilizing a Stable Isotope Tracer.

An important step in the early development of a nanomedicine formulation is the evaluation of stability and drug release in biological matrices. Additionally, the measurement of encapsulated and unencapsulated nanomedicine drug fractions is important for the determination of bioequivalence (pharmacokinetic equivalence) of generic nanomedicines. Unfortunately, current methods to measure drug release in plasma are limited, and all have fundamental disadvantages including non-equilibrium conditions and process-induced artifacts. The primary limitation of current ultrafiltration (and equilibrium dialysis) methods for separation of encapsulated and unencapsulated drug and determination of drug release is the difficulty in accurately differentiating protein bound and encapsulated drug. Since the protein binding of most drugs is high (>70%) and can change in a concentration- and time-dependent manner, it is very difficult to accurately account for the fraction of non-filterable drug that is encapsulated within the nanomedicine and how much is bound to protein. The method in this chapter is an improvement of existing ultrafiltration protocols for nanomedicine fractionation in plasma, in which a stable isotope tracer is spiked into a nanomedicine containing plasma sample in order to precisely measure the degree of plasma protein binding. Determination of protein binding then allows for accurate calculation of encapsulated and unencapsulated nanomedicine drug fractions, as well as free and protein-bound fractions.

Antibiotic Distribution into Cerebrospinal Fluid: Can Dosing Safely Account for Drug and Disease Factors in the Treatment of Ventriculostomy-Associated Infections?

Ventriculostomy-associated infections, or ventriculitis, in critically ill patients are associated with considerable morbidity. Efficacious antibiotic dosing for the treatment of these infections may be complicated by altered antibiotic concentrations in the cerebrospinal fluid due to variable meningeal inflammation and antibiotic properties. Therefore, doses used to treat infections with a higher degree of meningeal inflammation (such as meningitis) may often fail to achieve equivalent exposures in patients with ventriculostomy-associated infections such as ventriculitis. This paper aims to review the disease burden, infection rates, and common pathogens associated with ventriculostomy-associated infections. This review also seeks to describe the disease- and drug-related factors that influence antibiotic distribution into cerebrospinal fluid and provide a critical appraisal of current dosing of antibiotics commonly used to treat these types of infections. A Medline search of relevant articles was conducted and used to support a review of cerebrospinal fluid penetration of vancomycin, including critical appraisal of the recent paper by Beach et al. recently published in this journal. We found that in the intensive care unit, ventriculostomy-associated infections are the most common and serious complication of external ventricular drain insertion and often result in prolonged patient stay and increased healthcare costs. Reported infection rates are extremely variable (between 0 and 45%), hindered by the inherent diagnostic difficulty. Both Gram-positive and Gram-negative organisms are associated with such infections and the rise of multi-drug-resistant pathogens means that effective treatment is an ongoing challenge. Disease factors that may need to be considered are reduced meningeal inflammation and the presence of critical illness; drug factors include physiochemical properties, degree of plasma-protein binding, and affinity to active transporter proteins present in the blood-cerebrospinal fluid barrier. The relationship between cerebrospinal fluid antibiotic exposures in the setting of ventriculostomy-associated infection and clinical response has not been fully elucidated for many of the antibiotics commonly used in its treatment. More thorough and clinically relevant investigations are needed to better define blood pharmacokinetic/pharmacodynamics targets and optimal therapeutic exposures for treatment of ventriculostomy-associated infections. It is hoped that this future research will be able to provide clearer recommendations for clinicians frequently faced with dosing-related dilemmas when treating patients with these challenging infections.

Pharmacokinetics of long-acting cefovecin in copper rockfish (Sebastes caurinus).

To assess the pharmacokinetic properties of cefovecin in a cold-water teleost species. 10 healthy adult copper rockfish (Sebastes caurinus), sex unknown. Cefovecin (16 mg/kg) was administered SC to the rockfish. Blood samples were collected at predetermined points for measurement of plasma cefovecin concentrations (3 samples/fish). Plasma cefovecin concentrations were measured via liquid chromatography with mass spectrometry. Pharmacokinetic analysis was performed by means of naïve pooled analysis and compartmental modeling. Plasma protein binding of cefovecin was determined by ultrafiltration. Cefovecin administration appeared to be well tolerated by the rockfish. Pharmacokinetic analysis resulted in a maximum plasma concentration of 104.8 μg/mL at 2.07 hours after administration. Plasma terminal half-life was 32.5 hours, and area under the curve was 5,132 h·g/mL. Plasma protein binding was low (< 10%) for plasma concentrations of 10 and 100 μg of cefovecin/mL when assessed at 7.8° and 20°C. Plasma concentrations > 1 μg/mL persisted for the full 7-day follow-up period. SC administration of cefovecin to copper rockfish at a dose of 16 mg/kg yielded plasma concentrations > 1 μg/mL that persisted to 7 days, but some interindividual variability was observed. The low degree of plasma protein binding but high circulating concentration of free drug may allow an extended administration interval in rockfish. Studies are needed to assess the efficacy and safety of this dose in rockfish.

Assays for determination of ertapenem for applications in therapeutic drug monitoring, pharmacokinetics and sample stability.

Carbapanems are a class of β-lactam antibiotics with broad-spectrum potency and high β-lactamase resistance. Ertapenem, a member of this class, sold under the trade name Invanz™, has been of interest in the world of antibiotic therapeutic drug monitoring owing to its highly standardized 1 g dose and its high degree of plasma protein binding. Owing to the relative newness of this drug, fewer than 30 methods for ertapenem quantification have been published. Among these about half utilize biological matrices at the sample type. Liquid-liquid extraction and protein precipitation prevail as the most frequently used sample preparation techniques, despite their low recoveries compared with solid-phase extraction. Additionally, high-performance liquid chromatography with ultraviolet detection (HPLC-UV) is the instrumentation choice for most ertapenem assays. While these approaches may not achieve the highest possible sensitivity for ertapenem quantification, they provide clinically relevant tools for monitoring ertapenem in real patients. Sample stability is an ongoing concern for laboratories that handle ertapenem analysis, with buffering being of paramount importance, as well as low temperature (<-70°C) storage, to ensure minimal drug degradation in situ.

Effect of protein binding on unbound atazanavir and darunavir cerebrospinal fluid concentrations

HIV-1 protease inhibitors (PIs) exhibit different protein binding affinities and achieve variable plasma and tissue concentrations. Degree of plasma protein binding may impact central nervous system penetration. This cross-sectional study assessed cerebrospinal fluid (CSF) unbound PI concentrations, HIV-1 RNA, and neopterin levels in subjects receiving either ritonavir-boosted darunavir (DRV), 95% plasma protein bound, or atazanavir (ATV), 86% bound. Unbound PI trough concentrations were measured using rapid equilibrium dialysis and liquid chromatography/tandem mass spectrometry. Plasma and CSF HIV-1 RNA and neopterin were measured by Ampliprep/COBAS® Taqman® 2.0 assay (Roche) and enzyme-linked immunosorbent assay (ALPCO), respectively. CSF/plasma unbound drug concentration ratio was higher for ATV, 0.09 [95% confidence interval (CI) 0.06-0.12] than DRV, 0.04 (95%CI 0.03-0.06). Unbound CSF concentrations were lower than protein adjusted wild-type inhibitory concentration-50 (IC50 ) in all ATV and 1 DRV-treated subjects (P < 0.001). CSF HIV-1 RNA was detected in 2/15 ATV and 4/15 DRV subjects (P = 0.65). CSF neopterin levels were low and similar between arms. ATV relative to DRV had higher CSF/plasma unbound drug ratio. Low CSF HIV-1 RNA and neopterin suggest that both regimens resulted in CSF virologic suppression and controlled inflammation.

Abstract 3660: Crenolanib: A next generation FLT3 inhibitor

Abstract Every year in the US there are approximately 12,000 new cases of acute myeloid leukemia (AML). Amongst these patients, 25% have internal tandem duplications (ITDs) of the type III receptor tyrosine kinase, FLT3, causing constitutive activation of the receptor. This mutation has been associated with a poor survival rate, thus making the FLT3/ITD an important target in the treatment of AML. Treatment with small molecule tyrosine kinase inhibitors (TKIs) has shown high response rates in AML patients.1 However, this response is often temporary due to the development of resistance-conferring point mutations.2 Furthermore, QTc prolongation has proven to be a dose-limiting toxicity with some FLT3 TKIs.1 Crenolanib is a TKI with activity against PDGFR and FLT3 and is currently being tested in trials involving patients with a variety of solid tumors. We have determined that crenolanib has potent activity in vitro against FLT3. Crenolanib is cytotoxic to the FLT3/ITD-expressing leukemia cell lines Molm14 and MV411, with an IC50 of 7 nM and 8 nM, respectively. In immunoblots, crenolanib inhibited phosphorylation of both the wild type FLT3 receptor (in SEMK2 cells) and the FLT3/ITD receptor (in Molm14 cells) in culture medium with an IC50 of 1-3 nM. Importantly, the IC50 of crenolanib against the D835Y mutated form of FLT3 was 8.8 nM in culture medium. Using primary AML patient samples, we determined that crenolanib was cytotoxic to FLT3/ITD-expressing samples in a manner comparable to agents such as sorafenib and AC220. Furthermore, crenolanib had cytotoxic activity against primary samples that were obtained from patients who had developed D835 resistance mutations while receiving FLT3 TKIs. In vitro, the IC50 of crenolanib for inhibition of FLT3/ITD in plasma was found to be 34 nM, indicating a relatively low degree of plasma protein binding. From pharmacokinetic studies of crenolanib in solid tumor patients, steady state trough plasma levels of roughly 500 nM were found to be safe and tolerable, suggesting that crenolanib could potentially inhibit the target in vivo as effectively as AC220. Unlike AC220, crenolanib has no significant activity against c-KIT, which may be an advantage in that myelosuppression can be avoided. Furthermore, there was no evidence of QTc prolongation in patients treated with crenolanib. In summary, crenolanib offers a number of advantages over other FLT3 TKIs. Clinical trials of crenolanib in AML patients with FLT3 activating mutations are planned.

The Involvement of a Na+- and Cl–-Dependent Transporter in the Brain Uptake of Amantadine and Rimantadine

Despite their structural similarity, the two anti-influenza adamantane compounds amantadine (AMA) and rimantadine (RIM) exhibit strikingly different rates of blood-brain barrier (BBB) transport. However, the molecular mechanisms facilitating the higher rate of in situ BBB transport of RIM, relative to AMA, remain unclear. The aim of this study, therefore, was to determine whether differences in the extent of brain uptake between these two adamantanes also occurred in vivo, and elucidate the potential carrier protein facilitating their BBB transport using immortalized human brain endothelial cells (hCMEC/D3). Following oral administration to Swiss Outbred mice, RIM exhibited 2.4-3.0-fold higher brain-to-plasma exposure compared to AMA, which was not attributable to differences in the degree of plasma protein binding. At concentrations representative of those obtained in vivo, the hCMEC/D3 cell uptake of RIM was 4.5-15.7-fold higher than that of AMA, with Michaelis-Menten constants 6.3 and 238.4 μM, respectively. The hCMEC/D3 cellular uptake of both AMA and RIM was inhibited by various cationic transporter inhibitors (cimetidine, choline, quinine, and tetraethylammonium) and was dependent on extracellular pH, membrane depolarization and Na⁺ and Cl⁻ ions. Such findings indicated the involvement of the neutral and cationic amino acid transporter B⁰,⁺ (ATB⁰,⁺) in the uptake of AMA and RIM, which was demonstrated to be expressed (at the protein level) in the hCMEC/D3 cells. Indeed, AMA and RIM appeared to interact with this transporter, as shown by a 53-70% reduction in the hCMEC/D3 uptake of the specific ATB⁰,⁺ substrate ³H-glycine in their presence. These studies suggest the involvement of ATB⁰,⁺ in the disposition of these cationic drugs across the BBB, a transporter with the potential to be exploited for targeted drug delivery to the brain.

The degree of plasma protein binding of sodium cromoglycate.

Journal Article The degree of plasma protein binding of sodium cromoglycate Get access B Clark, B Clark Fisons Limited, Pharmaceutical Division, Research & Development Laboratories, Bakewell Road, Loughborough, Leicestershire, LE11 0QY, UK Correspondence: Fisons Limited, Pharmaceutical Division, Research & Development Laboratories, Bakewell Road, Loughborough, Leicestershire, LE11 0QY, U.K. Search for other works by this author on: Oxford Academic Google Scholar G F Moss, G F Moss Fisons Limited, Pharmaceutical Division, Research & Development Laboratories, Bakewell Road, Loughborough, Leicestershire, LE11 0QY, UK Search for other works by this author on: Oxford Academic Google Scholar M G Neale M G Neale Fisons Limited, Pharmaceutical Division, Research & Development Laboratories, Bakewell Road, Loughborough, Leicestershire, LE11 0QY, UK Search for other works by this author on: Oxford Academic Google Scholar Journal of Pharmacy and Pharmacology, Volume 30, Issue 1, September 1978, Pages 386–387, https://doi.org/10.1111/j.2042-7158.1978.tb13266.x Published: 12 April 2011 Article history Received: 24 January 1978 Published: 12 April 2011

Effect of Obesity on the Pharmacokinetics of Drugs in Humans

The prevalence of obesity has dramatically increased in recent years and now includes a significant proportion of the world's children, adolescents and adults. Obesity is linked to a number of co-morbidities, the most prominent being type 2 diabetes mellitus. While many agents are available to treat these conditions, the current knowledge regarding their disposition in the obese remains limited. Over the years, both direct and indirect methodologies have been utilized to assess body composition. Commonly used direct measures include underwater weighing, skinfold measurement, bioelectrical impedance analysis and dual-energy x-ray absorptiometry. Unfortunately, these methods are not readily available to the majority of clinicians. As a result, a number of indirect measures to assess body composition have been developed. Indirect measures rely on patient attributes such as height, bodyweight and sex. These size metrics are often utilized clinically and include body mass index (BMI), body surface area (BSA), ideal bodyweight (IBW), percent IBW, adjusted bodyweight, lean bodyweight (LBW) and predicted normal weight (PNWT). An understanding of how the volume of distribution (V(d)) of a drug changes in the obese is critical, as this parameter determines loading-dose selection. The V(d) of a drug is dependent upon its physiochemical properties, the degree of plasma protein binding and tissue blood flow. Obesity does not appear to have an impact on drug binding to albumin; however, data regarding alpha(1)-acid glycoprotein binding have been contradictory. A reduction in tissue blood flow and alterations in cardiac structure and function have been noted in obese individuals. At the present time, a universal size descriptor to describe the V(d) of all drugs in obese and lean individuals does not exist. Drug clearance (CL) is the primary determinant to consider when designing a maintenance dose regimen. CL is largely controlled by hepatic and renal physiology. In the obese, increases in cytochrome P450 2E1 activity and phase II conjugation activity have been observed. The effects of obesity on renal tubular secretion, tubular reabsorption, and glomerular filtration have not been fully elucidated. As with the V(d), a single, well validated size metric to characterize drug CL in the obese does not currently exist. Therefore, clinicians should apply a weight-normalized maintenance dose, using a size descriptor that corrects for differences in absolute CL between obese and non-obese individuals. The elimination half-life (t((1/2))) of a drug depends on both the V(d) and CL. Since the V(d) and CL are biologically independent entities, changes in the t((1/2)) of a drug in obese individuals can reflect changes in the V(d), the CL, or both. This review also examines recent publications that investigated the disposition of several classes of drugs in the obese--antibacterials, anticoagulants, antidiabetics, anticancer agents and neuromuscular blockers. In conclusion, pharmacokinetic data in obese patients do not exist for the majority of drugs. In situations where such information is available, clinicians should design treatment regimens that account for any significant differences in the CL and V(d) in the obese.

Multiple‐Dose Pharmacokinetics and Safety of Desloratadine in Subjects With Moderate Hepatic Impairment

Desloratadine, a nonsedating histamine H(1)-receptor antagonist, is metabolized to 3-hydroxy (3-OH) desloratadine. Impaired hepatic function could result in increased exposure to desloratadine. This study assessed possible differences in the pharmacokinetics and safety of desloratadine and 3-OH desloratadine in subjects (N = 21) with moderate hepatic dysfunction or normal liver function. Subjects were given desloratadine 5 mg once daily for 10 days and were assessed in several pharmacokinetic parameters. A similar degree of plasma protein binding to desloratadine and 3-OH desloratadine was observed in healthy volunteers and subjects with moderate hepatic impairment. All subjects with hepatic impairment were normal metabolizers. Three subjects with normal liver function, all African American, were identified as poor metabolizers. Exposure to desloratadine in the poor metabolizers was 2.6- to 6.5-fold greater than in other subjects with normal liver function. Eleven treatment-related adverse events, all mild to moderate in severity, were reported. Results suggest that subjects with moderate hepatic impairment experienced a greater increase in desloratadine exposure than subjects with normal liver function. Poor metabolizers had more exposure to desloratadine than normal metabolizers with or without hepatic impairment. Desloratadine administered at a daily dose of 5 mg was well tolerated.

Antimicrobial therapy for bacterial meningitis

Bacterial meningitis is an uncommon condition in mature horses (Newton 1988). It is more prevalent in neonates, where it is reported to occur in 8–10% of septicaemic foals (Moore 1995), where reported survival rates are poor (Newton 1988; Santschi and Foreman 1989; Moore 1995). Survival rate for adult horses was reported as 60% in a case series of 5 horses with bacterial meningitis (Mitchell et al. 2006). There are no published comprehensive case series of bacterial meningitis in foals. Early diagnosis is essential to provide accurate prognosis and to institute proper treatment (Santschi and Foreman 1989). The primary goal of treatment is rapid sterilisation of the cerebrospinal fluid (CSF) using appropriate antimicrobial therapy based on bacterial culture and sensitivity testing of the CSF (Moore 1995). However, due to the specialist nature of the central nervous system (CNS) and presence of the blood brain barrier (BBB), CNS concentrations of antimicrobial agents may be substantially below those obtained in serum, and below the minimal inhibitory concentration or minimal bactericidal concentration for the bacterial isolates involved in the disease process. This paper reviews the nature of poor antimicrobial penetration into the CNS and discusses the antimicrobial drugs that may be appropriate to use in treatment of bacterial meningitis in horses. Meningitis is defined as inflammation of the 3 membranes, or meninges, surrounding the brain and spinal cord. These consist of the dura mater, the arachnoid and pia mater (Fig 1). Inflammation of the meninges due to bacterial infection is termed bacterial meningitis. The space between the arachnoid and pia mater is filled with CSF. This protects the brain and spinal cord against impact from the surrounding bony wall, provides nutrition to the brain and spinal cord and permits variations in blood volume within the cranial cavity. The existence of a barrier between the brain or CSF and the peripheral circulation was first recognised by Ehrlich in 1885 (Saunders et al. 1999), and allows the adult brain to function in a well-controlled internal environment that is separate from that of the internal environment of the rest of the body as a whole. The integrity of this barrier is due to the presence of tight junctions between the cerebral endothelial cells forming the BBB and between the blood and choroid plexus epithelial cells (blood-cerebrospinal fluid barrier) (Saunders et al. 1999). The presence of endothelial cell tight junctions and basement membrane also restricts entry of certain therapeutic agents into the CSF and CNS. The presence of the BBB, combined with the lack of a traditional lymphatic system, are important factors in mediating the immune response within the CNS. As such, the CNS is often classified as an ‘immune privileged’ site (Wekerle et al.1986). In health, the key factors that determine penetration of a molecule into the CNS and CSF in adults are lipid solubility and molecular size (Saunders et al. 1999). Highly lipid soluble molecules are considered to dissolve in the lipid structure of cell membranes and can thus transverse cell membranes. For molecules of low lipid solubility there is a clear relationship between molecular size and penetration into the brain and CSF. In healthy human subjects, molecules with size greater than 5000 Daltons are unable to penetrate from blood to CSF (Saunders et al. 1999). In addition, a high degree of plasma protein binding of therapeutic agents limits their permeability to the CNS, since plasma proteins (principally albumin) are too large to cross the BBB. This situation is altered in acute CNS inflammation, where penetration of molecules of larger molecular weight

Unexpected relationship between plasma protein binding and the pharmacodynamics of 2‐NAP, a CCK1‐receptor antagonist

WHAT IS ALREADY KNOWN ABOUT THIS SUBJECT?: * Two chemically diverse CCK1 receptor antagonists have been shown clinically to inhibit CCK-evoked contraction of human gallbladder [2, 3]. These studies have not examined the relationship between plasma concentration and effect, the latter usually considered to be predictive from the free drug concentration [8]. * We wanted to examine our novel CCK1 receptor antagonist in this validated model and also to explore its PK-PD relationship. * 2-NAP inhibited CCK-evoked human gallbladder contraction in vivo but at a plasma free concentration that was, in theory, too low to have achieved adequate CCK1 receptor occupancy. * The study serves as a caveat to the assumption that free plasma concentration can be used to predict pharmacological effect. To study the pharmacokinetics and pharmacodynamics of 2-NAP (2-naphthalenesulfonyl-L-aspartyl-(2-phenethyl)amide), a selective CCK1 receptor antagonist in healthy volunteers. 2-NAP was given to 12 healthy male volunteers in an ascending dose, safety and PK phase 1a study by 1 h i.v. infusion (0.6-9.6 mg kg(-1) h(-1)). A further 12 healthy male volunteers received i.v. CCK-8S (6.25 pmol kg(-1) h(-1)) to produce gallbladder contraction, measured by ultrasound recordings of gallbladder volume, and the effect of concurrent i.v. 2-NAP administration was studied. Plasma protein binding in vitro and ex vivo was measured by ultrafiltration and by equilibrium dialysis. 2-NAP was generally well tolerated, displayed linear pharmacokinetics and a very high degree of plasma protein binding (99.9%). A 105 min i.v. CCK-8S infusion induced a reduction in gallbladder volume of 14.9 (+/-7.0) ml during placebo co-infusion and this was reduced to 2.4 (+/-5.9) ml when 2-NAP was co-infused with CCK-8S (P = 0.00024, paired t-test, mean change 12.5 ml; 95% CI For mean 7.4, 18.3 ml). This extent of inhibition was consistent with a 2-NAP total plasma concentration of 36 microm, but when protein binding corrections were made, the 'free concentration' of 2-NAP was only 0.04 microm, a value much less than the average equilibrium dissociation constant of 2-NAP for human CCK1 receptors ( approximately 0.7 microm). The pharmacological effect of a drug is usually considered to be determined by its free concentration. However, the complete inhibition of CCK-8S-evoked gallbladder contraction by a free plasma concentration of 0.04 microm 2-NAP was much greater than would have been predicted from simple drug-receptor occupancy theory and cautions against the general use of free concentration of drug for predicting pharmacological effect.

Population pharmacokinetics of lorazepam and midazolam and their metabolites in intensive care patients on continuous venovenous hemofiltration

The objective is to study the population pharmacokinetics of lorazepam and midazolam in critically ill patients with acute renal failure who are treated with continuous venovenous hemofiltration (CVVH). Twenty critically ill patients with acute renal failure on CVVH therapy were administered either lorazepam (n = 10) or midazolam (n = 10) by continuous infusion. CVVH was performed with an ultrafiltrate flow of 2 L/h with filtrate substitution in the predilution or postdilution mode. Blood flow through the 1.9-m 2 cellulose triacetate membrane filter was 180 mL/min. For 48 hours, multiple blood and ultrafiltrate samples were obtained for determination of concentrations of the drug and its metabolites. The pharmacokinetics of lorazepam is described best by a 1-compartment model. No significant covariates were identified. Total-body clearance was 6.4 L/h, and volume of distribution was 376 L. Ultrafiltration clearance was 0.31 L/h, equivalent to approximately 5% of total clearance. Average degree of plasma protein binding was 82.9% for lorazepam, with a sieving coefficient of 0.16 +/- 0.03. For lorazepamglucuronide, degree of plasma protein binding was 39.5%, and sieving coefficient was 0.48 +/- 0.07. The pharmacokinetics of midazolam is described best by a 1-compartment model. No significant covariates were identified. Total-body clearance was 8.5 L/h, and volume of distribution was 157 L. Clearance by ultrafiltration was 0.055 L/h, equivalent to approximately 0.7% of total clearance. Average degree of plasma protein binding was 95.8%, with a sieving coefficient of 0.04 +/- 0.03. For the metabolite 1-hydroxymidazolamglucuronide, average degree of plasma protein binding was 43.4%, with a sieving coefficient of 0.45 +/- 0.06. Neither lorazepam nor midazolam is removed efficiently by CVVH. CVVH contributes significantly to the removal of the glucuronide metabolites lorazepamglucuronide and 1-hydroxymidazolamglucuronide.

Serum albumin-adjusted phenytoin levels: an approach for predicting drug efficacy in patients with epilepsy, suitable for developing countries

The antiepileptic drug phenytoin has a high degree of plasma protein binding. Therefore, total phenytoin levels in plasma are misleading indicators of clinical efficacy. This study was designed to investigate whether serum albumin-adjusted phenytoin levels in Indian patients with epilepsy predict clinical outcome better than total phenytoin levels. Fifty patients with epilepsy were included in the study and were followed-up for a period of 6 months. Serum albumin levels were estimated spectrophotometrically using the bromocresol green dye method, and serum phenytoin levels were estimated using high pressure liquid chromatography. Values were expressed as mean +/- SEM. Corrected phenytoin levels were calculated using the Sheiner-Tozer equation. Corrected phenytoin levels = Measured total phenytoin(micromol/l) [(albumin g/1 x 0.9)+ 0.1] 40. At Visit 1, mean serum albumin levels were 44.1 +/- 1.1 micromol/l and mean serum phenytoin levels were 33.9 +/- 2.8 g/l. After correction of the total phenytoin levels using the Sheiner-Tozer equation, 30% of the patients shifted to a different category. The follow-up visits showed similar results. Throughout the study, the corrected phenytoin levels were better indicators of clinical outcome than the total levels. In 23% of patients there was a significant difference between total and corrected phenytoin levels. In patients with serum albumin levels in the hyper- and hypoalbuminemic range, corrected phenytoin levels were better indicators of clinical outcome. In developing countries like India, where estimation of free drug levels is expensive and suitable equipment is not available in most centers, serum albumin-adjusted levels can be used by pharmacologists to predict response and thus assist in clinical decision-making.

PKQuest: capillary permeability limitation and plasma protein binding - application to human inulin, dicloxacillin and ceftriaxone pharmacokinetics.

BackgroundIt is generally assumed that the tissue exchange of antibiotics is flow limited (complete equilibration between the capillary and the tissue water). This assumption may not be valid if there is a large amount of plasma protein binding because the effective capillary permeability depends on the product of the intrinsic capillary permeability (PS) and the fraction of solute that is free in the blood (fwB). PKQuest, a new generic physiologically based pharmacokinetic software routine (PBPK), provides a novel approach to modeling capillary permeability in which the only adjustable parameter is the PS of muscle.MethodsAll the results were obtained by applying PKQuest to previously published human pharmacokinetic data.ResultsThe PKQuest analysis suggests that the highly protein bound antibiotics dicloxacillin and ceftriaxone have a significant capillary permeability limitation. The human muscle capillary PS of inulin, dicloxacillin and ceftriaxone was 0.6, 13 and 6 ml/min/100 gm, respectively. The ceftriaxone protein binding is non-linear, saturating at high plasma concentrations. The experimental ceftriaxone data over a wide range of intravenous inputs (0.15 to 3 gms) was well described by PKQuest. PKQuest is the first PBPK that includes both permeability limitation and non-linear binding.ConclusionsBecause of their high degree of plasma protein binding, dicloxacillin and ceftriaxone appear to have a diffusion limited exchange rate between the blood and tissue and are not flow limited as had been previously assumed. PKQuest and all the examples are freely available at .