Percentage Of Protein Binding Research Articles

Exploring the viability of Zeatin as a prospective therapeutic candidate for investigating the complex interplay between severe acute respiratory syndrome coronavirus (SARS-CoV) and Alzheimer's disease.

The present research aims to explore the intricate link between SARS-CoV infection and susceptibility to Alzheimer's disease, focusing on the role of APOE4, a genetic factor associated with both conditions. Our research aims to uncover shared molecular pathways, considering APOE4's impact on lipid metabolism, immune responses, and neuroinflammation relevant to COVID-19 and AD. The Chyawanprash phytocompounds were subjected to in-silico ADMET profiling and Zeatin a neuroprotective cytokinin emerged as a promising regulator of the ACE2-SPIKE complex as it exhibits favourable pharmacological attributes, presenting as a non-substrate for Permeability glycoprotein, low Protein Binding Percentage, and distinctive toxicity endpoints. Therapeutic candidate. Zeatin's robust binding disrupts the intricate APOE4-ACE2-SPIKE interplay (AAS), offering a potential therapeutic avenue that is further corroborated by Molecular dynamic simulation as the system remained stable without any major fluctuation throughout the 100ns simulation. The AAS binding free energy, determined as -124.849 +/- 15.513 KJ/mol using MMPBSA assay, reveals significant contributions to complex stability from amino acids including, GLN41: 1.211kcal/mol, GLU340: 1.188kcal/mol, ALA344: 1.198kcal/mol, while ARG38: 2.011kcal/mol establishes pivotal strong bonds integral to the interaction between AAS and Zeatin. Rigorous cytotoxicity assessments reveal Zeatin's safety profile, highlighting its inhibitory effect on LN18 cell viability that sharply decreases to 32.47% at 200µg/ml, underscoring its modulatory impact on cellular metabolism. These findings enhance our understanding of the convergent mechanisms linking SARS-CoV and AD, providing valuable insights for potential therapeutic interventions. Further research is warranted to elucidate the specific pathways and molecular mechanisms through which zeatin exerts its protective effects.

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.

Albumin in Tears Modulates Bacterial Susceptibility to Topical Antibiotics in Ophthalmology.

Bacterial keratitis is a serious and vision-threatening condition in veterinary and human patients, one that often requires culture and susceptibility testing to adjust therapy and improve clinical outcomes. The present study challenges the antimicrobial susceptibility testing (AST) paradigm in ophthalmology, enabling more accurate in vitro-to-in vivo translation by incorporating factors normally present during host-pathogen interactions in clinical patients. Thirty bacteria (10 Staphylococcus pseudintermedius, 10 Streptococcus canis, 10 Pseudomonas aeruginosa) were isolated from canine patients with infectious keratitis. For each isolate, commercial plates (Sensititre™ JOEYE2) were used to assess the minimal inhibitory concentration (MIC) of 17 different antibiotics in the absence (0% albumin, control) or presence of canine albumin (0.01–2%). For Staphylococcus pseudintermedius, the experiment was repeated with actual tear fluid collected from canine eyes with ocular surface inflammation. Kruskal-Wallis, Wilcoxon signed rank test and Spearman's correlation tests were used for statistical analysis. Clinical outcomes were unfavorable in selected canine patients with bacterial keratitis (e.g., globe perforation, graft dehiscence) despite standard AST (i.e., 0% albumin in test medium) confirming that most corneal infections (93%) were susceptible to ≥1 topical antibiotics used at the initial visit. Albumin levels ≥0.05% increased MICs in a dose-dependent, bacteria-specific, and antibiotic-specific manner. No significant differences (P = 1.000) were noted in MICs of any antibiotic whether albumin or tear fluid was added to the Mueller-Hinton broth. Percent protein binding inherent to each antibiotic was associated with clinical interpretations (Spearman's rho = −0.53, P = 0.034) but not changes in MICs. Albumin in tears impacted the efficacy of selected ophthalmic antibiotics as only the unbound portion of an antibiotic is microbiologically active. The present findings could improve decision making of clinicians managing bacterial keratitis, reduce development of antimicrobial resistance, influence current guidelines set by the Clinical and Laboratory Standards Institute, and serve as a reference for bacteriological evaluations across medical fields and across species.

Development and characterization of two novel 68 Ga-labelled neuropeptide Y short analogues with potential application in breast cancer imaging.

In vivo receptor targeting with radiolabelled peptide-based probes is an attractive approach for the development of novel radiotracers for molecular imaging. This work presents the development and characterization of two novel neuropeptide Y analogues labelled with a positron emitter 68 Ga, for potential use in breast cancer imaging. Both analogues share the same amino acid sequence and were derivatized with NOTA through either a lysine linker (L1) or an acetylated lysine (L2). In both cases, a single product with radiochemical purity higher than 95% was obtained. The two complexes were hydrophilic, showed remarkable in vitro stability, good cellular uptake, binding affinity in the nanomolar range and high cellular internalization rate. Biodistribution studies revealed low blood uptake and elimination through the urinary tract. The addition of an acetyl group in the spacer increased the lipophilicity of C2 and modified the reactivity of the ε-amino group of the lysine which resulted in lower protein binding and lower percentage of injected dose in bladder and urine. The tumour versus muscle ratio was (3.8±0.4) for 68 Ga-L1 and (4.7±0.4) for 68 Ga-L2. These results encourage performing further studies in order to complete the evaluation of both tracers as potential radiopharmaceutical for breast cancer imaging.

Pharmacokinetics and dialytic clearance of apixaban during in vitro continuous renal replacement therapy

BackgroundTo evaluate the transmembrane clearance (CLTM) of apixaban during modeled in vitro continuous renal replacement therapy (CRRT), assess protein binding and circuit adsorption, and provide initial dosing recommendations.MethodsApixaban was added to the CRRT circuit and serial pre-filter bovine blood samples were collected along with post-filter blood and effluent samples. All experiments were performed in duplicate using continuous veno-venous hemofiltration (CVVH) and hemodialysis (CVVHD) modes, with varying filter types, flow rates, and point of CVVH replacement fluid dilution. Concentrations of apixaban and urea were quantified via liquid chromatography-tandem mass spectrometry. Plasma pharmacokinetic parameters for apixaban were estimated via noncompartmental analysis. CLTM was calculated via the estimated area under the curve (AUC) and by the product of the sieving/saturation coefficient (SC/SA) and flow rate. Two and three-way analysis of variance (ANOVA) models were built to assess the effects of mode, filter type, flow rate, and point of dilution on CLTM by each method. Optimal doses were suggested by matching the AUC observed in vitro to the systemic exposure demonstrated in Phase 2/3 studies of apixaban. Linear regression was utilized to provide dosing estimations for flow rates from 0.5–5 L/h.ResultsMean adsorption to the HF1400 and M150 filters differed significantly at 38 and 13%, respectively, while mean (± standard deviation, SD) percent protein binding was 70.81 ± 0.01%. Effect of CVVH point of dilution did not differ across filter types, although CLTM was consistently significantly higher during CRRT with the HF1400 filter compared to the M150. The three-way ANOVA demonstrated improved fit when CLTM values calculated by AUC were used (adjusted R2 0.87 vs. 0.52), and therefore, these values were used to generate optimal dosing recommendations. Linear regression revealed significant effects of filter type and flow rate on CLTM by AUC, suggesting doses of 2.5–7.5 mg twice daily (BID) may be needed for flow rates ranging from 0.5–5 L/h, respectively.ConclusionFor CRRT flow rates most commonly employed in clinical practice, the standard labeled 5 mg BID dose of apixaban is predicted to achieve target systemic exposure thresholds. The safety and efficacy of these proposed dosing regimens warrants further investigation in clinical studies.

Determining the Drug-Like Properties of Ailanthone, a Novel Chinese Medicine Monomer with Anti-CRPC Activity.

Approximately 40% of compounds with therapeutic potential cannot be successfully developed into drugs owing to their poor pharmaceutical properties, emphasising the need to profile their drug-like properties as early as possible during preclinical development. This study aimed to evaluate the drug-like properties of ailanthone, a novel Chinese medicine monomer that was shown to have activity against castration-resistant prostate cancer tumour growth and metastasis in our previous study. The drug-like properties detected in the present study included effects on permeability, liver microsome stability, plasma protein binding rate, plasma stability, and human ether-à-go-go-related gene inhibition. Additionally, the following results were obtained: the efflux ratio of ailanthone was > 32 during permeability detection; the half-life and intrinsic clearance (Clint) in mouse, rat, and human liver microsomes were > 145 min and < 9.6 µL/min/mg protein, respectively. The Clint(liver) of ailanthone was < 38.0, < 17.3, and < 8.6 mL/min/kg body weight in mice, rats, and humans, respectively. The plasma protein binding percentage of ailanthone was 16.6 ± 4.2% in human plasma, with 62.5% remaining at 120 min after incubation. The IC50 value of ailanthone for the human ether-à-go-go-related gene channels was > 30 µM. Collectively, these results and those from our previous study indicate that the pharmacokinetic properties of ailanthone are suitable for the potential development of this compound as an oral or intravenous drug for the treatment of castration-resistant prostate cancer.

Lipid-PLGA hybrid nanoparticles of paclitaxel: Preparation, characterization, in vitro and in vivo evaluation.

Paclitaxel loaded lipid-polymer nanoparticles (NPs) were successfully synthesized using poly lactide-co-glycolide (PLGA) as polymer and stearyl amine, soya lecithin as lipids via single step nanoprecipitation method. The study was aimed to combine the advantage of structural integrity of hybrid NPs containing PLGA core and lipid in the shell. Surfactants such as polyvinyl alcohol (PVA), tocopheryl polyethylene glycol succinate (TPGS), pluronic 68 (F68) and human serum albumin (HSA) were used as stabilizers. NPs were characterized w.r.t. morphology, particle size, zeta potential, encapsulation efficiency, in vitro drug release, protein binding capability and blood compatibility. NPs were in size range of 150-400nm and the particle size was greatly influenced by type and concentration of surfactants and lipids. TEM analysis confirmed the spherical shape and coating of the lipid on the NPs surface. Highest percentage entrapment efficiency was observed in NPs prepared with HSA as surfactant. The release rate of paclitaxel from modified NPs was much slower as compared to unmodified NPs. The percent protein binding of P-PVA, P-TPGS, P-F68 and P-HSA (unmodified NPs) was found to be 15.11%, 16.27%, 27.90% and 33.72%, respectively demonstrating effect of surface properties of NPs on protein binding. The hemolytic activity of the NPs was found to be dependent on type of surfactant and not on the lipid employed. PVA, TPGS, F68, HSA surfactants showed ~16%, ~10%, ~13%, ~7% hemolysis rate, respectively. The surface nature of NPs had a significant effect on the circulation profile of formulations. The HSA based NPs showed prolonged blood circulation time when compared to NPs without lipid coating. Thus, the synthesized dual lipid coated PLGA NPs with HSA could act as a potential nano-system for controlled delivery of paclitaxel.

Pharmacokinetics of Tylvalosin in Broiler Turkeys (Meleagris Gallopavo) After Single Intravenous and Oral Administration.

Pharmacokinetics of tylvalosin (TVN) were determined in eight broiler turkeys following a single intravenous (IV) and peroral (PO) administration of 25 mg/kg b.w using a crossover design with a 3 weeks washout period. Blood samples were taken between 0.083 and 24 h following TVN administration, plasma was separated and assayed for TVN concentrations by HPLC. The non-compartmental analysis was used to analyze plasma concentration-time curves. After IV administration, the pharmacokinetic profile was best described by a two-compartment model. The mean distribution and elimination half-lives were 0.382 and 5.71 h, respectively. The distribution volume at steady state, total body clearance and mean residence time were 8.30 L/kg, 1.17 L/h, and 7.16 h, respectively. After administering orally, the mean absorption half-life and absorption time of TVN was 0.955 and 2.31 h, respectively. The peak plasma concentration was 1.08 μg/mL and achieved at 2.0 h post-administration and the bioavailability was 53.3%. The plasma protein binding percent was 13%. For a successful clinical efficacy of TVN in broiler turkeys, a dosage regimen of 25 mg/kg b.w, given orally each day is recommended to keep efficient plasma levels above the MIC for most susceptible microorganisms.

Improved dialytic removal of protein-bound uremic toxins by intravenous lipid emulsion in chronic kidney disease rats.

Protein-bound uremic toxins (PBUTs) have received extensive attention, as their accumulation leads to pleiotropic toxic biological effects, while the removal of these solutes by conventional dialysis therapies is severely hampered. This study aimed to examine whether increased removal of PBUTs could be achieved with intravenous lipid emulsion (ILE). PBUTs such as 3-carboxy-4-methyl-5-propyl-2-furanpropionic acid (CMPF), p-cresyl sulfate (PCS) and indoxyl sulfate (IS) were spiked with human serum albumin (HSA) solution and the inhibitory effects of free fatty acid (FFA) on the binding of CMPF, PCS and IS to HSA were examined separately in vitro by ultrafiltration. In vitro dialysis of albumin solution was then performed to investigate the effects of fatty acid (FAs) mixtures infusion on the fractional removal of PBUTs. Finally, the inhibitory effect of FFA on the binding of PBUTs to albumin was examined in uremic rats, and blood purification therapy was conducted to calculate the reduction ratio (RR) and the total solute removal (TSR) of solutes. The percentage protein binding of CMPF, PCS and IS decreased significantly with increasing FFAs concentrations, and the inhibitory effect was more remarkable with the addition of oleic acid or linoleic acid than that of eicosapentaenoic acid and docosahexaenoic acid. In vitro infusion of FAs increased the fractional removal of CMPF to 14.40 ± 2.38%. PCS, IS and indole-3-acetic acid removal increased from 8.00 ± 2.43%, 11.68 ± 1.54% and 15.38 ± 3.97%, respectively, at baseline to 28.21 ± 5.99%, 35.42 ± 5.27% and 40.18 ± 5.05%, respectively, when FAs were present. In vivo, rat serum concentrations of free PBUTs were significantly higher in the ILE group than in the control group, and administration of ILE resulted in higher RRs and more TSR for PBUTs after 3 h of hemodialysis (HD) therapy compared with the control group. Administration of ILE effectively increased the dialytic removal of PBUTs. This method could be applied to current HD therapy.

In-vitro Plasma Protein Binding of Marbofloxacin in Healthy and Disease Condition of Buffalo Calves

Marbofloxacin is a broad-spectrum fluoroquinolone antibiotic developed for use in veterinary medicine for the treatment of skin and soft tissue infections in dogs and cats. Plasma protein binding plays a vital role in distribution, elimination and therapeutic effectiveness of drugs. In the present study we evaluated the plasma protein binding of marbofloxacin in healthy and liver dysfunctioned buffalo calves. In vitro binding of marbofloxacin to plasma proteins was determined by employing the equilibrium dialysis technique and further analyzed by High Performance Liquid Chromatography assay. The plasma protein binding for healthy calves ranges between 25.3±0.34% to 30.4±0.40% with an overall binding of 28.66 ± 0.421%. Kinetic constants (βi) and (Kβ) was 2.6±0.12×10−8 mole/g and 1.9±0.08×10−7 mole, respectively. The percentage of plasma protein binding for liver dysfunctioned buffalo calves extended from 24.5–30.3% with an overall mean of 28.59 ± 0.693%. The binding capacity of the drug to plasma proteins (βi) and dissociation rate constant of protein drug complex (Kβ) were 2.53±0.13 10−5 mole/g and 1.94±0.09×10−6 mole respectively. There was no significant change observed in plasma protein binding and the kinetic constant of liver dysfunctioned buffalo calves when compared to the healthy group.

Roflumilast analogs with improved metabolic stability, plasma protein binding, and pharmacokinetic profile.

With the aim of studying their in vitro and in vivo pharmacokinetics, new chromatographic methods were developed for the determination of three new roflumilast synthetic analogs (I-III) as PDE-4B inhibitors in rat liver S9 fraction, phosphate buffered saline, pH7.4, and human and rat plasma. The developed high performance liquid chromatography-ultra violet (HPLC-UV) methods were performed on a Zorbax Eclipse C8 column and UV detection was carried out at 215nm. The three compounds were tested for their metabolic stability and were found to be metabolically more stable than roflumilast especially the 2-mercaptobenzothiazol-6-yl analog (III) which displayed an in vitro half-life time (247.55minutes) higher than that of roflumilast (12.29minutes) and a low in vitro clearance of 5.67mL/min./kg. Possible phase I metabolites were investigated using ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) showing hydroxylation of the unsubstituted benzothiazol-2-yl (I) and benzothiazole-6-yl (II) analogs and a cleaved benzothiazole metabolite of the 2-mercaptobenzothiazol-6-yl analog (III). Plasma protein binding affinity was tested using equilibrium membrane dialysis method showing a very high percentage (more than 95%) of plasma protein binding of compounds I and II where compound III exhibited lower percentage (53.71%) demonstrating its accessibility for tissue distribution. Also, a UPLC-MS/MS method was developed using an Acquity UPLC BEH shield RP C18 column to be applied to an in vivo pharmacokinetic study in rats following a subcutaneous dose (1mg/kg). Compounds I-III showed improved in vivo pharmacokinetic parameters especially compound III which displayed a half-life 3-fold greater than roflumilast (21hours) and a Cmax value of 113.958ng/mL. Accordingly, this new chemical entity should be subjected to further investigation as it can be a good drug candidate for treating chronic obstructive pulmonary disease.

Development and validation of an ultra-high performance liquid chromatography – high resolution mass spectrometry method for the quantification of total and free teicoplanin in human plasma

ObjectivesThe antibiotic teicoplanin, used for the treatment of infections caused by Gram-positive bacteria, is highly bound to plasma proteins (percentage protein binding, %PB, around 90%) and therapeutic plasma levels of total teicoplanin are 10–100 mg/L. Because of the low free concentrations (i.e. <1–10 mg/L), current immunoassays are not able to quantify free teicoplanin concentrations, although they might be more relevant in therapeutic drug monitoring than total concentrations. Design and methodsIn this study, an ultra-high performance liquid chromatography – high resolution mass spectrometry (UHPLC-HRMS) method for the quantification of total and free teicoplanin in K2EDTA plasma samples was developed and validated. Furthermore, %PB obtained by ultrafiltration was compared with that obtained by equilibrium dialysis using spiked samples from healthy subjects. Analytes were separated using a phenylhexyl column, gradient mobile phase analysis was used, total run time was 4.5 min and teicoplanin was detected by orbitrap MS. ResultsThe precision and accuracy were below 15% and within ±15%, respectively and teicoplanin was found to be stable for at least 14 days in plasma at 4 °C. The %PB of teicoplanin in spiked plasma from healthy subjects as obtained by ultrafiltration (94.1 ± 1.3%) was in good agreement with that obtained by equilibrium dialysis (93.6 ± 0.4%), whereas mean %PB of teicoplanin in samples from infected patients who received the antibiotic was 87.7 ± 4.2% (range: 79.6–95.4%). ConclusionA novel highly sensitive UHPLC-HRMS method was developed and validated for the quantification of total and free teicoplanin in human K2EDTA plasma samples. Amongst others, this method is suitable for therapeutic drug monitoring.

Effect of Ionic Strength, pH and Chemical Displacers on the Percentage Protein Binding of Protein-Bound Uremic Toxins

Background and Objectives: While trying to optimize the dialysis clearances of protein-bound uremic toxins (PBUTs), their percentage protein binding (% PB) is an important parameter. We evaluated the effects of ionic strength, pH change and chemical displacers on the dissociation of PBUTs from albumin in vitro. Methods: PBUTs, such as 3-Carboxy-4-methyl-5-propyl-2-furan-propanoic acid (CMPF), p-cresylsulfate (PCS), indoxyl sulfate (IS) and indole-3-acetic acid (IAA), were spiked with human serum albumin (HSA) solution prepared with different Nacl concentrations and pH values or in the presence of a series of chemical displacers. Ultrafiltration was performed to separate the free and bound fractions, and the % PB of each PBUT was calculated. Results: For all 4 compounds, their % PB decreased with increasing ionic strength, while only slight changes occurred when the pH of the test solution increased from pH 6.0 to pH 8.5; PCS, IS and 3-IAA were relatively easily dissociated from albumin by drug displacement, while CMPF was released from HSA by all studied drugs with difficulty; the PB % for CMPF, PCS, IS and 3-IAA decreased most remarkably in the presence of free fatty acids, such as oleic acid (41.73% for CMPF, 29.9% for PCS, 23.22% for IS, and 20.34% for 3-IAA) and linoleic acid (43.12% for CMPF, 16.65% for PCS, 29.99% for IS, and 16.29% for 3-IAA). Conclusion: The protein binding of PBUTs can be decreased by higher ionic strength, increased pH and the presence of some chemical displacers, including free fatty acids. Effective dialytic removal of PBUTs may be achieved by applying these methods jointly to blood-purification techniques.

Combined molecular modeling and cholinesterase inhibition studies on some natural and semisynthetic O-alkylcoumarin derivatives

Coumarins of synthetic or natural origins are an important chemical class exerting diverse pharmacological activities. In the present study, 26 novel O-alkylcoumarin derivatives were synthesized and have been tested at 100 µM for their in vitro inhibitory potential against acetylcholinesterase (AChE) and butyrlcholinesterase (BChE) targets which are the key enzymes playing role in the pathogenesis of Alzheimer’s disease. Among the tested coumarins, none of them could inhibit AChE, whereas 12 of them exerted a marked and selective inhibition against BChE as compared to the reference (galanthamine, IC50 = 46.58 ± 0.91 µM). In fact, 10 of the active coumarins showed higher inhibition (IC50 = 7.01 ± 0.28 µM – 43.31 ± 3.63 µM) than that of galanthamine. The most active ones were revealed to be 7-styryloxycoumarin (IC50 = 7.01 ± 0.28 µM) and 7-isopentenyloxy-4-methylcoumarin (IC50 = 8.18 ± 0.74 µM). In addition to the in vitro tests, MetaCore/MetaDrug binary QSAR models and docking simulations were applied to evaluate the active compounds by ligand-based and target-driven approaches. The predicted pharmacokinetic profiles of the compounds suggested that the compounds reveal lipophilic character and permeate blood brain barrier (BBB) and the ADME models predict higher human serum protein binding percentages (>50%) for the compounds. The calculated docking scores indicated that the coumarins showing remarkable BChE inhibition possessed favorable free binding energies in interacting with the ligand-binding domain of the target. Therefore, our results disclose that O-alkylcoumarins are promising selective inhibitors of cholinesterase enzymes, particularly BChE in our case, which definitely deserve further studies.

Binding of tylosin, tilmicosin and oxytetracycline to proteins from honeybees, larvae and beehive products

American Foulbrood (AFB) caused by the spore-forming bacterium Paenibacillus larvae is the most serious disease of bacterial origin affecting larvae and pupae of honeybees. Antibiotics are used in many countries for the control of AFB in high incidence areas, but their misuse may lead to antibiotic resistance of bacterial strains and honey contamination. The objective of the present work was to determine, through a biological method, the protein binding of tylosin, tilmicosin and oxytetracycline to worker jelly; honey; pollen; adult bees and larvae in order to propose their kinetic routes. The sensitivity limit of the technique used was 0.05 μg/ml for tylosin and tilmicosin and 0.01 μg/ml for oxytetracycline, respectively. The method had intra and inter-assay correlation coefficients over 0.90, respectively and a coefficient variation of intra-and inter-assay for all antibiotics and processed samples under 5%. Tylosin and oxytetracycline presented lower percentages of protein binding in tissues and hive products (average 15%) in relation to those observed for tilmicosin (29%). In conclusion, tylosin is useful for AFB control in honey bee colonies due to its chemical characteristics, antimicrobial activity and levels of protein binding in bees, larvae, and beehive products.

Pharmacokinetics and Pharmacodynamics of Temocillin.

Temocillin, a 6-α-methoxy derivative of ticarcillin, is a forgotten antibiotic that has recently been rediscovered, and issues about clinical breakpoints and optimal therapeutic regimens are still ongoing. Temocillin spectrum is almost restricted to Enterobacteriaceae. The addition of the α-methoxy moiety on ticarcillin confers resistance to hydrolysis by Ambler classes A and C β-lactamases (extended spectrum β-lactamases, Klebsiella pneumoniae carbapenemase and AmpC hyperproduced enzymes). Temocillin is bactericidal, and the effect of inoculum size on its activity is relatively mild. The proportion of spontaneous resistant mutants in vitro to temocillin is low, as found in vivo. After intravenous infusion, temocillin showed a prolonged elimination half-life of approximately 5h. The percentage of protein binding of temocillin is high (approximately 80%), and is concentration-dependent. Temocillin clearance is mainly renal, and urinary recovery is high, ranging from 72 to 82% after 24h. Furthermore, the penetration of temocillin into bile and peritoneal fluid is high, but poor into cerebrospinal fluid. The cumulative percentage of a 24-h period during which the free drug concentration exceeds the minimum inhibitory concentration (fT>MIC) at steady-state pharmacokinetic conditions seems to be the best pharmacokinetic/pharmacodynamic (PK/PD) index correlating with temocillin efficacy. An fT>MIC of 40-50% is associated with antibacterial effect and survival in vivo. Monte Carlo simulations performed in critically ill patients showed that the 2g every 12h and 2g every 8h regimens provide a 95% probability of target attainment of 40% fT>MIC up to an MIC of 8mg/L. In less severely ill patients or in specific foci of infection, such as urinary tract infection, a 4g daily regimen should be adequate for strains with temocillin MIC up to 16mg/L. Data regarding actual wild-type MIC distribution, clinical efficacy, PK profiling in volunteers or patients, and PD targets are scarce, and further studies are required to support appropriate dosing recommendations and determination of clinical breakpoints.

Exploring binding characteristics and the related competition of different protein-bound uremic toxins

Little is known about potential differences in binding characteristics of protein-bound uremic toxins (PBUTs) in patients with chronic kidney disease (CKD) versus healthy controls. The question arises whether eventual differences are attributed to (i) the elevated levels of competing uremic toxins, and/or (ii) post-translational modifications of albumin.We evaluated the binding characteristics of hippuric acid (HA), indole-3-acetic acid (IAA), indoxyl sulfate (IS), and p-cresylsulfate (pCS) by deriving a binding curve in three distinct conditions: (i) serum from healthy controls (healthy serum), (ii) blank serum from hemodialysis patients (blank HD serum; i.e. cleared from uremic toxins), and (iii) non-treated serum from HD patients (HD serum). Additionally, the mutual binding competition of these uremic toxins was studied in blank HD in pairs. In both experiments, equilibrium dialysis (37 °C, 5 h) was used to separate the free and bound fractions of each PBUT. Free and total PBUT concentrations were quantified by an ultra-high performance liquid chromatography method with tandem mass spectrometer detection and the percentage protein binding (%PB) of each PBUT was calculated.For all four compounds, the binding capacity of healthy serum was higher than blank HD serum, which was comparable to non-treated HD serum, except for HA. The competition experiments revealed that at high uremic concentrations, mutual competition was observed for the strongly bound PBUTs IS and pCS. The %PB of the weakly bound HA and IAA was lower (trend) only for the addition to blank HD serum containing the strongly bound IS or pCS.There is an intrinsic impact on protein binding in uremia, revealing a lower binding capacity, as compared to healthy controls. Competitive binding is only relevant for the strongly bound PBUTs at high uremic concentrations. In addition, at least part of the effect on binding capacity can be attributed to post-translational modifications of albumin.

Effect of sample temperature, pH, and matrix on the percentage protein binding of protein-bound uraemic toxins

While studying and trying to optimise dialysis clearances of protein-bound uraemic toxins (PBUTs), the percentage protein binding (% PB) may be an important parameter and can be calculated from measured free and total concentrations.

In vivo Studies of Protein Binding of Ketorolac in Rat Model by UV-VIS Spectrophotometry and High Performance Liquid Chromatographic Methods

In this study, an attempt has been made to determine the protein binding of ketorolac at different time and concentration in rat model. A total of ten rats were used for this study. The rats were subdivided into two groups and 100?g/ml and 200?g/ml of ketorolac were administered through the intra-peritoneal route to rats of group 1 and group 2, respectively. The serum from the rats were collected and analyzed using UV-VIS spectrophotometry and HPLC. In the UV-VIS Spectrophotometric study, the highest percentage of protein binding for ketorolac was found to be 63.067% in group 1 and 74.63% in group 2. In case of HPLC method, the highest protein binding of ketorolac for group 1 and group 2 were 89.72% and 95.07%, respectively. The observed protein binding percentage of ketorolac by HPLC method was slightly higher than that found in UV-VIS Spectrophotometric method. The result obtained in this study showed that the protein binding of ketorolac changes with the time and drug concentration. The percentage of protein binding was higher for higher drug concentration.Dhaka Univ. J. Pharm. Sci. 15(1): 63-67, 2016 (June)

Interactions Between Sirolimus and Anti-Inflammatory Drugs: Competitive Binding for Human Serum Albumin.

The aim of the present study was investigating the effects of three anti-inflammatory drugs, on Sirolimus protein biding. The binding site of Sirolimus on human serum albumin (HSA) was also determined. Six different concentrations of Sirolimus were separately exposed to HSA at pH 7.4 and 37°C. Ultrafiltration method was used for separating free drug; then free drug concentrations were measured by HPLC. Finally, Sirolimus protein binding parameters was calculated using Scatchard plots. The same processes were conducted in the presence of NSAIDs at lower concentration of albumin and different pH conditions. To characterize the binding site of Sirolimus on albumin, the free concentration of warfarin sodium and Diazepam, site I and II specific probes, bound to albumin were measured upon the addition of increasing Sirolimus concentrations. Based on the obtained results presence of Diclofenac, Piroxicam and Naproxen, could significantly decrease the percentage of Sirolimus protein binding. The Binding reduction was the most in the presence of Piroxicam. Sirolimus-NSAIDs interactions were increased in higher pH values and also in lower albumin concentrations. Probe displacement study showed that Sirolimus may mainly bind to site I on albumin molecule. More considerations in co-administration of NSAIDs and Sirolimus is recommended.