Efficient Ibuprofen Delivery from Anhydrous Semisolid Formulation Based on a Novel Cross-linked Silicone Polymer Network: An In Vitro and In Vivo Study

The use of silicone as a primary polymer in topical semisolid pharmaceutical formulations is infrequent. Recent development of novel silicone materials provides an opportunity to investigate their drug delivery efficiencies. In this study, an anhydrous semisolid formulation was prepared using a novel cross-linked silicone polymer network swollen in isododecane. Similar formulations were prepared using petrolatum, an acrylic, or a cellulose polymer. All formulations contained 5% ibuprofen (IBP). In vitro permeability was evaluated for all formulations and a commercial product using human cadaver epidermis. The silicone formulation delivered IBP more efficiently than all other formulations in terms of flux, cumulative amount, and percent drug release. The silicone formulation showed the maximum flux of 85.9 μg.cm⁻².h⁻¹ and a cumulative IBP release of 261.6 μg in 8 h, whereas the benchmark showed 20.1 μg.cm⁻².h⁻¹ and 30.9 μg, respectively. An in vivo study conducted on rats showed calculated blood AUCs of 59.2 and 17.6 μg.h/g (p < 0.003) for the silicone formulation and the benchmark, respectively. The IBP in excised rat skin was 264 ± 59 μg/g for the silicone formulation and 102 ± 5 μg/g for the benchmark. The results obtained from the in vitro and in vivo studies demonstrate efficient topical IBP delivery by the silicone formulation.     

Continuous versus intermittent infusion of cefepime in neurosurgical patients with post-operative intracranial infections

Cefepime is administered as an intermittent infusion (II); however, continuous infusion (CI) may be advantageous because β-lactam antibiotics exhibit time-dependent antibacterial activity. This retrospective, non-randomised, comparative study included 68 neurosurgical patients with post-operative intracranial infections treated with 4 g/day cefepime over 24 h as a CI (n = 34) or 2 g every 12 h as II (n = 34). CI controlled the intracranial infection more rapidly and effectively than II (6.6 ± 1.9 days vs. 7.8 ± 2.6 days; P = 0.036). By considering the minimum inhibitory concentrations (MICs) to be 4 μg/mL and 8 μg/mL, the percentage of time when the cefepime plasma or CSF concentrations were higher than the MIC (%T_>MIC) was calculated for each patient. For plasma cefepime concentrations, the %T_>MIC in the CI group was higher than in the II group (for MICs of 8 μg/mL, 100% vs. 75%, respectively). The mean calculated area under the curve (AUC) in the CI group was similar to the II group (1197.99 ± 72.15 μg h/mL vs. 890.84 ± 140.78 μg h/mL; P = 0.655). For CSF cefepime concentrations, the %T_>MIC in the CI group was higher than in the II group (for MICs of 4 μg/mL and 8 μg/mL, 83.3% and 75% vs. 25% and 0%, respectively). The mean calculated AUC for the CI group was higher than the II group (220.56 ± 13.59 μg h/mL vs. 86.34 ± 5.69 μg h/mL; P = 0.003). Therefore, CI of cefepime significantly enhanced the antibacterial effect and reduced the treatment duration in neurosurgical patients with post-operative intracranial infections.     

Antidermatitic Perspective of Hydrocortisone as Chitosan Nanocarriers: An Ex Vivo and In Vivo Assessment Using an NC/Nga Mouse Model

The aim of this study to administer hydrocortisone (HC) percutaneously in the form of polymeric nanoparticles (NPs) to alleviate its transcutaneous absorption, and to derive additional wound-healing benefits of chitosan. HC-loaded NPs had varied particle sizes, zeta potentials, and entrapment efficiencies, when drug-to-polymer mass ratios increased from 1:1 to 1:8. Ex vivo permeation analysis showed that the nanoparticulate formulation of HC significantly reduced corresponding flux [~24 μg/(cm² h)] and permeation coefficient (~4.8 × 10^? 3 cm/h) of HC across the full thickness NC/Nga mouse skin. The nanoparticulate formulation also exhibited a higher epidermal (1610 ± 42 μg/g of skin) and dermal (910 ± 46 μg/g of skin) accumulation of HC than those associated with control groups. An in vivo assessment using an NC/Nga mouse model further revealed that mice treated with the nanoparticulate system efficiently controlled transepidermal water loss [15 ± 2 g/(m² h)], erythema intensity (232 ± 12), dermatitis index (mild), and thickness of skin (456 ± 27 μm). Taken together, histopathological examination predicted that the nanoparticulate system showed a proficient anti-inflammatory and antifibrotic activity against atopic dermatitic (AD) lesions. Our results strongly suggest that HC-loaded NPs have promising potential for topical/transdermal delivery of glucocorticoids in the treatment of AD. © 2013 Wiley Periodicals, Inc. and the American Pharmacists Association J Pharm Sci 102:1063–1075, 2013     

Formulation,optimization and evaluation of transferosomal gel for transdermal insulin delivery

The present study deals with the development of transferosomal gel containing insulin by reverse phase evaporation method for painless insulin delivery for use in the treatment of insulin dependent diabetes mellitus. The effect of independent process variables like ratio of lipids (soya lecithin:cholesterol), ratio of lipids and surfactants, and ratio of surfactants (Tween 80:sodium deoxycholate) on the in vitro permeation flux (μg/cm²/h) of formulated transferosomal gels containing insulin through porcine ear skin was optimized using 2³ factorial design. The optimal permeation flux was achieved as 13.50 ± 0.22 μg/cm²/h with drug entrapment efficiency of 56.55 ± 0.37% and average vesicle diameter range, 625–815 nm. The in vitro insulin permeation through porcine ear skin from these transferosomal gel followed zero-order kinetics (R² = 0.9232–0.9989) over a period of 24 h with case-II transport mechanism. The in vitro skin permeation of insulin from optimized transferosomal gel by iontophoretic influence (with 0.5 mA/cm² current supply) also provided further enhancement of permeation flux to 17.60 ± 0.03 μg/cm²/h. The in vivo study of optimized transferosomal gel in alloxan-induced diabetic rat has demonstrated prolonged hypoglycemic effect in diabetic rats over 24 h after transdermal administration.     

Using transdermal iontophoresis to increase granisetron delivery across skin in vitro and in vivo: Effect of experimental conditions and a comparison with other enhancement strategies

The objectives of the study were (i) to investigate the effect of experimental parameters on the iontophoretic transport of granisetron, (ii) to identify the relative contributions of electromigration (EM) and electroosmosis (EO), (iii) to determine the feasibility of delivering therapeutic amounts of drug for the treatment of chemotherapy-induced nausea and vomiting and (iv) to test the in vitro results in a simple animal model in vivo. Preliminary in vitro studies using aqueous granisetron formulations investigating the effect of drug concentration (5, 10, 20 and 40 mM) and current density (0.1, 0.2, 0.3 mA cm^?2) were performed using porcine ear skin. As expected, cumulative delivery in vitro at the 20 and 40 mM concentrations was significantly greater than that at 5 and 10 mM, which were not statistically different (p < 0.05). Increasing the applied current density from 0.1 to 0.3 mA cm^?2 resulted in a ～4.2-fold increase in iontophoretic flux. Furthermore, in the absence of Na⁺ in the formulation, no dependence of iontophoretic flux on drug concentration was reported (at a granisetron concentration of 40 mM, the transport rate was 2.93 ± 0.62 μg cm^?2 min^?1). Co-iontophoresis of acetaminophen was used to show that EM was the predominant transport mechanism accounting for 71–86% of total granisetron delivery. In vivo studies in Wistar rats (40 mM granisetron; application of 0.3 mA cm^?2 for 5 h with Ag/AgCl electrodes and salt bridges) showed an average iontophoretic input rate (k_input) of 0.83 ± 0.26 μg min^?1 and a maximum plasma concentration (C_max) of 0.092 ± 0.004 μg ml^?1. Based on these results and given the known pharmacokinetics, transdermal iontophoresis could achieve therapeutic drug levels for the management of chemotherapy-induced emesis using a reasonably sized (4–6 cm²) patch.     

Bilayered Nail Lacquer of Terbinafine Hydrochloride for Treatment of Onychomycosis

The present study aimed to develop bilayered nail lacquer of terbinafine hydrochloride (TH) for treatment of onychomycosis. The composite nail lacquer formed an underlying drug-loaded hydrophilic layer and overlying hydrophobic vinyl layer. The hydrophilic lacquer made of hydroxylpropyl methylcellulose E-15 contained polyethylene glycol 400 (PEG 400) as a drug permeation enhancer. The vinyl lacquer was composed of poly (4-vinyl phenol) as a water- resistant film former. In vitro permeation studies in Franz diffusion cells indicated that the amount of TH permeated across the human cadaver nail in 6 days was 0.32 ± 0.14, 1.12 ± 0.42, and 1.42 ± 0.53 μg/cm² from control (hydrophilic lacquer devoid of PEG 400), monolayer (hydrophilic lacquer alone), and bilayered nail lacquers, respectively. A higher nail drug load was seen in vitro with the bilayered lacquer (0.59 ± 0.13 μg/mg) as compared to monolayer (0.36 ± 0.09 μg/ mg) and control (0.28 ± 0.07 μg/mg) lacquers. The drug loss despite multiple washing was significantly low (p < 0.001) for the bilayered lacquer owing to the protective vinyl coating. Clinical studies demonstrated the efficacy of bilayered lacquer to achieve better drug load in the nail plate (1.27 ± 0.184 μg/mg) compared to monolayer (0.67 ± 0.18 μg/mg) and control (0.21 ± 0.04 μg/mg) lacquers.     

Clinical Pharmacokinetics of Paclitaxel Liposome with a New Route of Administration in Human Based on the Analysis with Ultra Performance Liquid Chromatography

We investigated the clinical pharmacokinetics of paclitaxel liposome with a new route of administration, which was intrapleural infusion, in nine advanced nonsmall-cell lung cancer (NSCLC) patients with malignant pleural effusions after a single administration. Paclitaxel concentrations were measured in pleural fluid and plasma using a simple and rapid ultra performance liquid chromatography (UPLC) method following intra-and inter-day validations. In subjects, AUC_0–96h values in pleural fluid and plasma were 17831 ± 6439 μgh/mL and 778 ± 328 μgh/mL, respectively, and T_max values were initial time and 6.67 h after administration and the corresponding C_max values were 558 ± 44 μg/mL and 12.89 ± 6.86 μg/mL, respectively. The T_{1/2 IP}, CL_IP and Vd_IP values in pleural fluid were 76 ± 48 h, 0.005 ± 0.002 L/hm² and 0.53 ± 0.23 L/m², respectively. The T_1/2,_pla, CL_pla, and Vd_pla values in plasma were 68.34 ± 56.74 h, 0.184 ± 0.080 L/hm², and 17.53 ± 16.57 L/m², respectively. However, some paclitaxel concentrations from several patients in plasma could not be detected at some designed time-points. Our results might offer new opportunities to design and determine individually appropriate therapeutic dosage regimens based on a pharmacokinetic profile.     

Spray freeze drying for dry powder inhalation of nanoparticles

Formulating nanoparticles for delivery to the deep lung is complex and many techniques fail in terms of nanoparticle stability. Spray freeze drying (SFD) is suggested here for the production of inhalable nanocomposite microcarriers (NCM). Different nanostructures were prepared and characterized including polymeric and lipid nanoparticles. Nanoparticle suspensions were co-sprayed with a suitable cryoprotectant into a cooled, stainless steel spray tower, followed by freeze drying to form a dry powder while equivalent compositions were spray dried (SD) as controls. SFD-NCM possess larger specific surface areas (67–77 m²/g) and lower densities (0.02 g/cm³) than their corresponding SD-NCM. With the exception of NCM of lipid based nanocarriers, SFD produced NCM with a mass median aerodynamic diameter (MMAD) of 3.0 ± 0.5 μm and fine particle fraction (FPF ⩽ 5.2 μm) of 45 ± 1.6% with aerodynamic performances similar to SD-NCM. However, SFD was superior to SD in terms of maintaining the particle size of all the investigated polymeric and lipid nanocarriers following reconstitution (S_f/S_i ratio for SFD ≈ 1 versus >1.5 for SD). The SFD into cooled air proved to be an efficient technique to prepare NCM for pulmonary delivery while maintaining the stability of the nanoparticles.     

Comparative in vitro and ex-vivo myelotoxicity of aflatoxins B1 and M1 on haematopoietic progenitors (BFU-E,CFU-E,and CFU-GM): Species-related susceptibility

Haemato- and myelotoxicity are adverse effects caused by mycotoxins. Due to the relevance of aflatoxins to human health, the present study, employing CFU-GM-, BFU-E- and CFU-E-clonogenic assays, aimed at (i) comparing, in vitro, the sensitivity of human vs. murine haematopoietic progenitors to AFB1 and AFM1 (0.001–50 μg/ml), (ii) assessing whether a single AFB1 in vivo treatment (0.3–3 mg/kg b.w.) alters the ability of murine bone marrow cells to form myeloid and erythroid colonies, and (iii) comparing the in vitro with the in vitro ex-vivo data.We demonstrated (i) species-related sensitivity to AFB1, showing higher susceptibility of human myeloid and erythroid progenitors (IC₅₀ values: about 4 times lower in human than in murine cells), (ii) higher sensitivity of CFU-GM and BFU-E colonies, both more markedly affected, particularly by AFB1 (IC₅₀: 2.45 ± 1.08 and 1.82 ± 0.8 μM for humans, and 11.08 ± 2.92 and 1.81 ± 0.20 μM for mice, respectively), than the mature CFU-E (AFB1 IC₅₀: 12.58 ± 5.4 and 40.27 ± 6.05 μM), irrespectively of animal species, (iii) regarding AFM1, a species- and lineage-related susceptibility similar to that observed for AFB1 and (iv) lack of effects after AFB1 in vivo treatment on the proliferation of haematopoietic colonies.     

Penetration of doripenem into prostatic tissue following intravenous administration in prostatectomy patients

This study examined the prostatic penetration of doripenem in prostatectomy patients. Doripenem 500 mg was administered by a 0.5-h infusion and venous blood and prostatic tissue samples were obtained up to 5 h afterwards. Drug concentrations in plasma and prostatic tissue were measured chromatographically. The observed maximum concentration (C_max) (mean ± standard deviation; n = 9) was 27.5 ± 5.1 μg/mL in plasma and 5.09 ± 1.94 μg/g in prostate tissue and the prostate/plasma C_max ratio was 0.189 ± 0.078. The area under the drug concentration–time curve (AUC) was 49.7 ± 6.9 μg h/mL in plasma and 3.93 ± 1.89 μg h/g in prostate tissue and the prostate/plasma AUC ratio was 0.081 ± 0.047. Based on a three-compartment pharmacokinetic analysis, average drug exposure times above 0.25 μg/mL (the minimum inhibitory concentration for isolates of common pathogens) in the prostate were 23.2% for 500 mg once daily, 46.2% for 500 mg twice daily and 69.9% for 500 mg three times daily. The 500-mg regimens all achieved the drug exposure time target (bacteriostatic 20% or bactericidal 40%) in the prostate, despite the relatively low penetrability of doripenem.     

Levosimendan and its metabolite OR-1896 elicit KATP channel-dependent dilation in resistance arteries in vivo

BackgroundLevosimendan and its long-lived metabolite OR-1896 produce vasodilation in different types of vessels by activating ATP-sensitive (K_ATP) and other potassium channels.MethodsIn the present study we applied intravital videomicroscopy to investigate the in situ effects of levosimendan and OR-1896 on the diameters of real resistance arterioles (rat cremaster muscle arterioles with diameters of ~ 20 μm).ResultsLevosimendan and OR-1896 induced concentration-dependent (1 nM – 100 μM) dilations to similar extents in these arterioles (maximal dilation from 23 ± 2 to 33 ± 2 μm and from 22 ± 1 to 32 ± 1 μm, respectively). The arteriolar dilations induced by the selective K_ATP channel opener pinacidil (1 nM – 10 μM) (maximal dilation from 22 ± 4 μm to 35 ± 3 μm) were diminished in the presence of the selective K_ATP channel blocker – glibenclamide (5 μM) (maximal diameter attained: 22 ± 1 μm). Glibenclamide also counteracted the maximal dilations in response to levosimendan or OR-1896 (to 23 ± 3 μm or 22 ± 5 μm, respectively).ConclusionsIn conclusion, this is the first demonstration that levosimendan and OR-1896 elicit arteriolar dilation in vivo, via activation of K_ATP channels in real resistance vessels in the rat.     

Activation of nicotinamide N-methyltrasferase and increased formation of 1-methylnicotinamide (MNA) in atherosclerosis

Nicotinamide N-methyltrasferase (NMMT) catalyzes the conversion of nicotinamide (NA) to 1-methylnicotinamide (MNA). Recent studies have reported that exogenous MNAexerts anti-thrombotic and anti-inflammatory activity, suggesting that endogenous NMMT-derived MNA may play a biological role in the cardiovascular system.In the present study, we assayed changes in hepatic NNMTactivity and MNAplasma levels along the progression of atherosclerosis in apoE/LDLR^?/? mice, as compared to age-matched wild-type mice. Atherosclerosis progression in apoE/LDLR^?/? mice was quantified in aortic root, while hepatic NNMT activity and MNAplasma concentrations were concomitantly measured in 2-, 3-, 4-, and 6-month-old mice.In apoE/LDLR^?/? mice, atherosclerotic plaques developed in the aortic roots beginning at the age of 3 months and gradually increased in size, macrophage content, and inflammation intensity over time, as detected by Oil-Red O staining, CD68 immunostaining, and in situ zymography (MMP2/MMP9 activity). Hepatic NNMT activity was upregulated approximately two-fold in apoE/LDLR^?/? mice by the age of 2 months, as compared to wild-type mice (1.03 ± 0.14 vs. 0.64 ± 0.23 pmol/min/mg, respectively). MNAplasma concentrations were also elevated approximately two-fold (0.30 ± 0.13 vs. 0.17 ± 0.04 μmol/l, respectively). As atherosclerosis progressed, hepatic NMMTactivity and MNAplasma concentrations increased five-fold in 6-month-old apoE/LDLR^?/? mice at the stage of advanced atherosclerotic plaques (NMMT activity: 2.29 ± 0.34 pmol/min/mg, MNA concentration: 1.083 ± 0.33 μmol/l).In summary, the present study demonstrated that the progression of vascular inflammation and atherosclerosis was associated with the upregulation of hepatic NNMT activity and subsequent increase in endogenous MNAplasma levels. Given the anti-thrombotic and anti-inflammatory properties of exogenous MNA, robust activation of an endogenous NA-MNApathway in atherosclerosis may play an important compensatory role.     

Sulfur mustard causes oxidants/antioxidants imbalance through the overexpression of free radical producing-related genes in human mustard lungs

The aim of this study is to analyze oxidative stress (OS) and changes in expression of reactive oxygen species (ROS) producing-related genes in mustard lungs. Human lung biopsies provided from controls (n = 5) and sulfur mustard (SM)-exposed patients (n = 6). Changes in expression of dual oxidases (DUOXs), aldehyde oxidase 1 (AOX1), thyroid peroxidase (TPO), myeloperoxidase (MPO) and eosinophil Peroxidase (EPO) were measured using RT² Profiler™ PCR Array. OS was evaluated by determining bronchoalveolar lavage fluids (BALF) levels of total antioxidant capacity (TAC) and malondialdehyde (MDA). Higher TAC value was observed in BALF of controls compared with patients (0.138 ± 0.02683 μmol/l vs 0.0942 ± 0.01793 μmol/l), whereas a significant increase in MDA concentration was found in patients (0.486 ± 0.04615 nmol/l vs 0.6467 ± 0.05922 nmol/l). All ROS producing-related genes were overexpressed in the order AOX1> MPO> DUOX2> DUOX1> TPO> EPO. Upregulation of these genes may be a reason for overproduction of ROS, oxidants/antioxidants imbalance, OS and respiratory failures in mustard lungs.     

Genotoxicity and antileishmanial activity evaluation of Physalis angulata concentrated ethanolic extract

Antileishmanial in vitro tests, as well as Ames and micronucleus assays were performed with a concentrated ethanolic extract of Physalis angulata (EEPA)ResultsEEPA did not present mutagenic effect in Salmonella typhimurium strains at concentration reaching 3000 μg/plate and did not induce mutagenic effects after two oral administrations with a 24 h interval at a dose level of 2000 mg/kg. EEPA presented antileishmanial activity and presented an IC₅₀ value of 5.35 ± 2.50 μg/mL and 4.50 ± 1.17 μg/mL against Leishmania amazonensis and Leishmania braziliensis promastigotes, respectively. In the cytotoxicity test against macrophages, the EEPA had a LC₅₀ of 6.14 ± 0.59 μg/mL. Importantly, the IC₅₀ against L. amazonensis intracellular amastigotes was 1.23 ± 0.11 μg/mL.ConclusionEEPA extract is non-mutagenic and presented a promising pharmacological effect against Leishmania parasites.     

Chemical composition,antiproliferative and antioxidant properties of lipid classes in ordinary and dark muscles from chub mackerel (Scomber japonicus)

This study investigated to compare lipid profiles in ordinary and dark muscles from chub mackerel and to examine antiproliferative and antioxidative properties of lipid classes. The average levels of neutral lipids (NL), glycolipids (GL), and phospholipids (PL) in ordinary muscle were 92.32 ± 0.19%, 5.10 ± 0.48%, and 2.58 ± 0.46%; in dark muscle were 96.88 ± 0.15%, 2.59 ± 0.36%, and 0.54 ± 0.29%, respectively. The fatty acid composition indicated that PL had a higher percentage of PUFA (especially 22:6n?3) with lower percentages of SFA and MUFA compared to NL and GL (p < 0.05). The main ion peaks of GL in ordinary and dark muscles showed that monocharged and bischarged molecular ion were presented at m/z 876.9 and 438.8, respectively. In MTT assay, inhibition of AGS and HT-29 cell proliferation was greatest with the 0.5 and 1.0 mg mL^?1 GL treatments. The GL of ordinary muscle with 0.05 mg mL^?1 concentrations markedly decreased the levels of reactive oxygen species (ROS) induced by H₂O₂ compared to the control (p < 0.05). From our results, GL might have antiproliferative and antioxidant properties based on protective effect against the production of intracellular ROS.     

Pulmonary delivery of pyrazinamide-loaded large porous particles

We have improved the aerodynamic properties of pyrazinamide loaded large porous particles (PZA-LPPs) designed for pulmonary delivery. To overcome the segregation of the different components occurring during the spray drying process and to obtain homogeneous LPPs, spray drying parameters were modified to decrease the drying speed. As a result, good aerodynamic properties for lung delivery were obtained with a fine particle fraction (FPF) of 40.1 ± 1.0%, an alveolar fraction (AF) of 29.6 ± 3.1%, a mass median aerodynamic diameter (MMAD_aer) of 4.1 ± 0.2 μm and a geometric standard deviation (GSD) of 2.16 ± 0.16. Plasma and epithelial lining fluid (ELF) concentrations of pyrazinamide were evaluated after intratracheal insufflation of PZA-LPPs (4.22 mg kg⁻¹) into rats and compared to intravenous administration (iv) of a pyrazinamide solution (5.82 mg kg⁻¹). The in vivo pharmacokinetic evaluation of PZA-LPPs in rats reveals that intratracheal insufflation of PZA-LPPs leads to a rapid absorption in plasma with an absolute bioavailability of 66%. This proves that PZA-LPPs dissolve fast upon deposition and that PZA crosses efficiently the lung barrier to reach the systemic circulation. PZA concentrations were 1.28-fold higher in ELF after intratracheal administration than after iv administration and the ratio of ELF concentrations over plasma concentrations was 2-fold greater. Although these improvements are moderate, lung delivery of PZA appears an interesting alternative to oral delivery of the molecule and should now be tested in an infected animal model to evaluate its efficacy against Mycobacterium tuberculosis.     

Quantification of biliary excretion and sinusoidal excretion of 5(6)-carboxy-2′,7′-dichlorofluorescein (CDF) in cultured hepatocytes isolated from Sprague Dawley,Wistar and Mrp2-deficient Wistar (TR−) rats

Hepatic efflux of drug candidates is an important issue in pre-clinical drug development. Here we utilise a method which quantifies and distinguishes efflux of drugs at the canalicular and sinusoidal membranes in rat hepatocyte cultures. Bi-phasic kinetics of transport of 5(6)-carboxydichlorofluorescein (CDF) at the canalicular membrane was demonstrated in Sprague Dawley (SD) and Wistar (W) rat hepatocytes. The high affinity component (Km = 3.2 ± 0.8 μM (SD), 9.0 ± 3.1 μM (W)) was attributed to Mrp2-mediated transport, the low affinity component (Km = 192.1 ± 291.5 μM (SD), 69.2 ± 36.2 μM (W)) may be attributed to transport involving a separate Mrp2 binding site. Data from membranes (Hill coefficient (h) = 2.0 ± 0.5) and vesicles (h = 1.6 ± 0.2) expressing Mrp2 and from SD (h = 1.6 ± 0.4) and Wistar (h = 4.0 ± 0.6) hepatocytes suggests transport involves more than one binding site. In TR⁻ hepatocytes, CDF efflux was predominantly over the sinusoidal membrane (Km = 100.7 ± 36.0 μM), consistent with low abcc2 (Mrp2) expression and compensatory increase in abcc3 (Mrp3) expression. This report shows the potential of using this in vitro method to model changes in biliary excretion due to alterations in transporter expression.     

Plasma protein binding of tetrodotoxin in the marine puffer fish Takifugu rubripes

To elucidate the involvement of plasma protein binding in the disposition of tetrodotoxin (TTX) in puffer fish, we used equilibrium dialysis to measure protein binding of TTX in the plasma of the marine puffer fish Takifugu rubripes and the non-toxic greenling Hexagrammos otakii, and in solutions of bovine serum albumin (BSA) and bovine alpha-1-acid glycoprotein (AGP). TTX (100–1000 μg/mL) bound to protein in T. rubripes plasma with low affinity in a non-saturable manner. The amount of bound TTX increased linearly with the TTX concentration, reaching 3.92 ± 0.42 μg TTX/mg protein at 1000 μg TTX/mL. Approximately 80% of the TTX in the plasma of T. rubripes was unbound in the concentration range of TTX examined, indicating that TTX exists predominantly in the unbound form in the circulating blood of T. rubripes at a wide range of TTX concentrations. TTX also bound non-specifically to H. otakii plasma proteins, BSA, and bovine AGP. The amount of the bound TTX in the plasma of H. otakii and BSA, respectively, was 1.86 ± 0.36 and 4.65 ± 0.70 μg TTX/mg protein at 1000 μg TTX/mL, and that in the bovine AGP was 8.78 ± 0.25 μg TTX/mg protein at 200 μg TTX/mL.     

The chemotherapeutic potential of doxorubicin-loaded PEG-b-PLGA nanopolymersomes in mouse breast cancer model

Vesicles of mPEG-PLGA block copolymer were developed to deliver a therapeutic quantity of doxorubicin (DOX) for breast cancer treatment. The DOX-loaded nanoparticles (NPs) were prepared by the pH-gradient method and then evaluated in terms of morphology, size, DOX encapsulation efficiency and in vitro drug release mechanism.The PEG-PLGA nanopolymersomes were 134 ± 1.2 nm spherical NPs with a narrow size distribution (PDI = 0.121). DOX was entrapped in mPEG-PLGA nanopolymersomes with an encapsulation efficiency and a loading content of 91.25 ± 4.27% and 7.3 ± 0.34%, respectively. The DOX-loaded nanopolymersomes were found to be stable, demonstrating no significant change in particle size and encapsulation efficiency (EE%) during the 6-month storage period of the lyophilized powder at 4 °C. The nanopolymersomes sustained the release of DOX. In cytotoxicity studies of 4T1 cell line samples, free DOX showed a higher cytotoxicity (IC50 = 1.76 μg/mL) than did DOX-loaded nanopolymersomes (15.82 μg/mL) in vitro. In order to evaluate the antitumor efficacy and biodistribution of DOX-loaded nanopolymersomes, murine breast tumors were established on the BALB/c mice, and in vivo studies were performed. The obtained results demonstrated that the prepared drug delivery system was highly effective against a murine breast cancer tumor model and successfully accumulated in the tumor site through an enhanced permeation and retention mechanism.In vivo studies also proved that DOX-loaded nanopolymersomes are stable in blood circulation and could be considered a promising and effective DOX delivery system for breast cancer treatment.     

Resveratrol self-emulsifying system increases the uptake by endothelial cells and improves protection against oxidative stress-mediated death

The aim of the present study was to develop and characterize a resveratrol self-emulsifying drug delivery system (Res-SEDDS), and to compare the uptake of resveratrol by bovine aortic endothelial cells (BAECs), and the protection of these cells against hydrogen peroxide-mediated cell death, versus a control resveratrol ethanolic solution.Three Res-SEDDSs were prepared and evaluated. The in vitro self-emulsification properties of these formulations, the droplet size and the zeta potential of the nanoemulsions formed on adding them to water under mild agitation conditions were studied, together with their toxicity on BAECs. An optimal atoxic formulation (20% Miglyol® 812, 70% Montanox® 80, 10% ethanol 96% v/v) was selected and further studied.Pre-incubation of BAECs for 180 min with 25 μM resveratrol in the nanoemulsion obtained from the selected SEDDS significantly increased the membrane and intracellular concentrations of resveratrol (for example, 0.076 ± 0.015 vs. ethanolic solution 0.041 ± 0.016 nmol/mg of protein after 60 min incubation, p < 0.05). Resveratrol intracellular localization was confirmed by fluorescence confocal microscopy. Resveratrol nanoemulsion significantly improved the endothelial cell protection from H₂O₂-induced injury (750, 1000 and 1500 μM H₂O₂) in comparison with incubation with the control resveratrol ethanolic solution (for example, 55 ± 6% vs. 38 ± 5% viability after 1500 μM H₂O₂ challenge, p < 0.05).In conclusion, formulation of resveratrol as a SEDDS significantly improved its cellular uptake and potentiated its antioxidant properties on BAECs.