Delivery Of Low Molecular Weight Heparin Research Articles

Development and in vivo evaluation of nanoemulsions for oral delivery of low molecular weight heparin

AimThe aim of this study was the development and in vivo evaluation of different types of nanoemulsions for oral delivery of low molecular weight heparin (LMWH). MethodsNanoemulsions (NE) were developed consisting of mainly non-digestible components and variations of polyglycerol-, PEGylated or zwitterionic surfactants. To incorporate LMWH in the oily droplets, hydrophobic ion pairs were formed with several cationic surfactants. Loaded and unloaded NE were evaluated for their size, zeta potential, stability towards pancreatin, hemolytic and toxic properties. Finally, an in vivo study on oral LMWH delivery was performed. ResultsDroplet size of all formulations was below 150 nm with a PDI <0.4. Highest logK1-butanol/water was obtained for the complex of LMWH and didecyldimethylammonium bromide (HIP). This complex was incorporated into NE containing polyglycerol- and PEGylated surfactants up to a concentration of 5% (m/v) and in zwitterionic NE up to 10% (m/m). NE containing polyglycerol-surfactants were digested to a higher extent than NE containing PEGylated surfactants. The zwitterionic NE without glycerides did not show any lipolysis but the zwitterionic NE which contained high amounts of glycerides was rapidly degradable. NE with polyglycerol-surfactants were less toxic and hemolytic than NE with PEGylated surfactants and zwitterionic NE. Toxic potential increased with HIP-loaded formulations. When performing in vivo studies, oral bioavailability of LMWH was increased up to 2.6% with non-digestible NE. ConclusionAccording to these results, non-digestible nanoemulsions improve oral bioavailability of LMWH more efficiently than biodegradable formulations.

NOVEL NANOPARTICLES FOR THE ORAL DELIVERY OF LOW MOLECULAR WEIGHT HEPARIN: IN VITRO AND IN VIVO ASSESSMENT

Objective: The objective of the present study was to prepare and evaluate a novel oral formulation of nanoparticles for the systemic delivery of low molecular weight heparin (LMWH). Methods: Nanoparticles were prepared by polyelectrolyte complexation (PEC) method using polymers sodium alginate and chitosan. Entrapment efficiency of LMWH in nanoparticles was found to be ̴88%. Fourier transform infrared spectroscopy (FTIR), differential scanning calorimetry (DSC), X‑ray diffraction (XRD), Scanning electron microscopy (SEM) studies carried for nanoparticles. In vitro release studies were performed for the formulations. Ex vivo permeation studies were performed optimized formulation by using small intestine of rat and in vivo studies were conducted on rat model.Results: In vitro release studies demonstrated that the release of LMWH was negligible in the stomach and high in the small intestine. FTIR has indicated that there is no interaction between the ingredients in nanoparticle. DSC and XRD studies confirmed that the amino groups of chitosan interacted with the carboxylic groups of alginate. Invitro % drug release of 95% was shown by formulation AC5. Ex vivo permeation studies have elucidated that ̴ 73% of LMWH was transported across the epithelium. Nanoparticles have shown enhanced oral bioavailability of LMWH as revealed by 4.5 fold increase in AUC of plasma drug concentration time curve.Conclusion: The results suggest that the nanoparticles prepared can result in targeted delivery of LMWH into systemic circulation via intestinal and colon routes. Novel nanoparticles thus prepared in this study can be considered as a promising delivery system.Keywords: Antifactor Xa activity, Chitosan, Differential scanning calorimetry, Sodium alginate, Low-molecular-weight heparin, Oral bioavailability.

Development and characterization of polymer lipid hybrid nanoparticles for oral delivery of LMWH

The present study aimed to develop an improved oral delivery system for low-molecular-weight heparin (LMWH), novel polymer lipid hybrid nanoparticles were developed. LMWH loaded chitosan polymer lipid hybrid nanoparticles (LMWH-CS-PLNs) were developed using double emulsification and solvent evaporation method. The performance of developed formulations was evaluated by using in vitro and in vivo behavior, such as drug release studies, in vitro permeation study, in vivo venous thrombolytic study, in vitro uptake studies by using intestinal epithelium resembling Caco-2 cell lines. The new CS-PLNs might provide an effective strategy for oral delivery of LMWH with improved encapsulation efficiency as compared to CS-NPs and SA-LNPs.

Preparation and evaluation of a novel oral delivery system for low molecular weight heparin.

Objective:The objective of the present work was to prepare and evaluate a novel oral formulation for systemic delivery of low molecular weight heparin (LMWH). The formulation consisted of Eudragit S 100-coated positively charged liposomes encapsulating LMWH and a penetration enhancer.Materials and Methods:Positively charged liposomes were first prepared by the thin film hydration method using lipid (soy phosphotidylcholine and cholesterol) and stearyl amine (SA) in the optimum ratio of 16:1, along with cetylpyridinium chloride (CPC) as a penetration enhancer. Prepared liposomes were coated with negatively charged Eudragit S 100 (0.3% w/v). The formulations were studied for various in vitro and in vivo properties. Differential scanning calorimetry (DSC), x-ray diffraction (XRD), scanning electron microscopy (SEM), Fourier transform infrared spectroscopy (FTIR) studies, and in vitro drug release were used for in vitro characterization of the formulations. Ex vivo permeation studies were performed by using distal small intestine of rat. Oral absorption studies were conducted with the rat model.Results:Coating of the liposomes was confirmed by SEM and particle size determination studies. In vitro release studies of coated liposomes have demonstrated that the release of LMWH was in the following order: Stomach < small intestine < distal small intestine < colon. Ex vivo permeation studies have shown a fivefold increase in permeation of LMWH with Eudragit S 100-coated liposomes compared to uncoated, uncharged liposomes. Oral absorption studies have showed that with Eudragit-coated liposomes, the oral bioavailability of LMWH was improved, compared to plain LMWH solution. This is revealed by a threefold increase in the area under the curve (AUC) of the plasma concentration time curve.Conclusion:A novel formulation for oral delivery of LMWH was thus successfully prepared and evaluated.

Development of subcutaneous sustained release nanoparticles encapsulating low molecular weight heparin.

The objective of the present research work was to prepare and evaluate sustained release subcutaneous (s.c.) nanoparticles of low molecular weight heparin (LMWH). The nanoparticles were prepared by water–in-oil in-water (w/o/w) emulsion and evaporation method using different grades of polylactide co-glycolide (50:50, 85:15), and different concentrations of polyvinyl alcohol (0.1%, 0.5%, 1%) aqueous solution as surfactant. The fabricated nanoparticles were evaluated for size, shape, zeta potential, encapsulation efficiency, in vitro drug release, and in vivo biological activity (anti-factor Xa activity) using the standard kit. The drug and excipient compatibility was analyzed by Fourier transform infrared spectroscopy (FTIR), differential scanning calorimetry (DSC) and X-ray diffraction (XRD) studies. The formation of nanoparticles was confirmed by scanning electron microscopy; nanoparticles were spherical in shape. The size of prepared nanoparticles was found between 195 nm and 251 nm. The encapsulation efficiency of the nanoparticles was found between 46% and 70%. In vitro drug, release was about 16–38% for 10 days. In vivo drug, release shows the sustained release of drug for 10 days in rats. FTIR studies indicated that there was no loss in chemical integrity of the drug upon fabrication into nanoparticles. DSC and XRD results demonstrated that the drug was changed from the crystalline form to the amorphous form in the formulation during the fabrication process. The results of this study revealed that the s.c. nanoparticles were suitable candidates for sustained delivery of LMWH.

PH-responsive thiolated chitosan nanoparticles for oral low-molecular weight heparin delivery: in vitro and in vivo evaluation

The aim of present study was to investigate a pH-responsive and mucoadhesive nanoparticle system for oral bioavailability enhancement of low-molecular weight heparin (LMWH). The thioglycolic acid (TGA) was first covalently attached to chitosan (CS) with 396.97 ± 54.54 μmol thiol groups per gram of polymer and then the nanoparticles were prepared with thiolated chitosan (TCS) and pH-sensitive polymer hydroxypropyl methylcellulose phthalate (HPMCP) by ionic cross-linking method. The obtained nanoparticles were characterized for the shape, particle size, zeta potential, drug entrapment efficiency and loading capacity. In vitro results revealed the acid stability of pH-responsive nanoparticles, which had a significant control over LMWH release and could effectively protect entrapped drugs in simulated gastric conditions. By the attachment of the thiol ligand, an improvement of permeation-enhancing effect on freshly excised carp intestine (1.86-fold improvement) could be found. The mucoadhesive properties were evaluated using fluorescently labeled TCS or CS nanoparticles. As compared with the controls, a significant improvement of mucoadhesion on rat intestinal mucosa was observed in TCS/HPMCP nanoparticles via confocal laser scanning microscopy. The activated partial thromboplastin time (APTT) was significantly prolonged and an increase in the oral bioavailability of LMWH was turned out to be pronounced after oral delivered LMWH-loaded TCS/HPMCP nanoparticles in rats, which suggested enhanced anticoagulant effects and improved absorption of LMWH. In conclusion, pH-responsive TCS/HPMCP nanoparticles hold promise for oral delivery of LMWH.

Oligomeric bile acid-mediated oral delivery of low molecular weight heparin

Intestinal transporters are limited to the transport of small molecular substrates. Here, we describe the development of apical sodium-dependent bile acid transporter (ASBT)-targeted high-affinity oligomeric bile acid substrates that mediate the transmembrane transport of low molecular weight heparin (LMWH). Several oligomers of deoxycholic acid (oligoDOCA) were synthesized to investigate the substrate specificity of ASBT. To see the binding of oligoDOCA on the substrate-binding pocket of ASBT, molecular docking was used and the dissociation rate constants (KD) were measured using surface plasmon resonance. The KD for tetrameric DOCA (tetraDOCA) was 50-fold lower than that for monomeric DOCA, because tetraDOCA interacted with several hydrophobic grooves in the substrate-binding pocket of ASBT. The synthesized oligoDOCA compounds were subsequently chemically conjugated to macromolecular LMWH. In vitro, tetraDOCA-conjugated LMWH (LHe-tetraD) had highest selectivity for ASBT during its transport. Orally administered LHe-tetraD showed remarkable systemic anticoagulation activity and high oral bioavailability of 33.5±3.2% and 19.9±2.5% in rats and monkeys, respectively. Notably, LHe-tetraD successfully prevented thrombosis in a rat model of deep vein thrombosis. These results represent a major advancement in ASBT-mediated LMWH delivery and may facilitate administration of many important therapeutic macromolecules through a non-invasive oral route.

Oral Delivery of Low Molecular Weight Heparin by Polyaminomethacrylate Coacervates

Oral bioavailability of low molecular weight heparin (LMWH) can be achieved by several advanced drug delivery approaches. Here, a new preparation method for coacervates (CAs) using non-toxic polyethylene glycol derivates was developed. LMWH were coacervated with polyaminomethacrylates (Eudragit® RL or RS) using polyethylene glycol (PEG) derivatives as non-toxic solvents. CAs were analyzed for their physicochemical properties and pharmacokinetic parameters were determined for different formulations in rabbits. CAs from both polymer types using various PEGs were of irregular shape and had particle sizes of around 40 μm, encapsulation efficiencies of >90%, and complete LMWH in vitro release was obtained within 2 h. In vivo, oral Absorption at doses of 300 IU/kg was rather low (F < 2.5%) while dose increase resulted in a maximum at 600 IU/kg (FRL: 6.0 ± 1.2%; FRS: 5.8 ± 2.5%) and 1,200 IU/kg did not result in higher bioavailability (FRL: 4.6 ± 0.4%; FRS: 4.1 ± 0.8%). CAs were applicable to various LMWH types where the oral availability decreased in the order fondaparinux>enoxaparin>nadroparin>certoparin depending mainly on the molecular weight. CAs prepared by an organic solvent-free method allowed the oral delivery of LMWHs. The therapeutic efficiency and the simple and solvent-free manufacturing process underlines the high potential of this new preparation method.

Systemic heparin delivery by the pulmonary route using chitosan and glycol chitosan nanoparticles

The aim of this study was to evaluate the performance of chitosan (CS) and glycol chitosan (GCS) nanoparticles containing the surfactant Lipoid S100 for the systemic delivery of low molecular weight heparin (LMWH) upon pulmonary administration. These nanoparticles were prepared in acidic and neutral conditions using the ionotropic gelation technique. The size and zeta potential of the NPs were affected by the pH and also the type of polysaccharide (CS or GCS). The size (between 156 and 385 nm) was smaller and the zeta potential (from +11 mV to +30 mV) higher for CS nanoparticles prepared in acidic conditions. The encapsulation efficiency of LMWH varied between 100% and 43% for the nanoparticles obtained in acidic and neutral conditions, respectively. X-ray photoelectron spectroscopy studies indicated that the surfactant Lipoid S100 was localized on the nanoparticle's surface irrespective of the formulation conditions. In vivo studies showed that systems prepared in acidic conditions did not increase coagulation times when administered to mice by the pulmonary route. In contrast, Lipoid S100-LMWH GCS NPs prepared in neutral conditions showed a pharmacological efficacy. Overall, these results illustrate some promising features of CS-based nanocarriers for pulmonary delivery of LMWH.

Comparative Studies on Chitosan and Polylactic-co-glycolic Acid Incorporated Nanoparticles of Low Molecular Weight Heparin

This study was performed to test the feasibility of chitosan and polylactic-co-glycolic acid (PLGA) incorporated nanoparticles as sustained-release carriers for the delivery of negatively charged low molecular weight heparin (LMWH). Fourier transform infrared (FTIR) spectrometry was used to evaluate the interactions between chitosan and LMWH. The shifts, intensity, and broadening of the characteristic peaks for the functional groups in the FTIR spectra indicated that strong interactions occur between the positively charged chitosans and the negatively charged LMWHs. Three types of LMWH nanoparticles (NP-1, NP-2, and NP-3) were prepared using chitosan with or without PLGA: NP-1 nanoparticles were formed by polyelectrolyte complexation after single mixing, NP-2 nanoparticles were prepared by polyelectrolyte complexation after single emulsion-diffusion-evaporation, and NP-3 nanoparticles were optimized by double emulsion-diffusion-evaporation. NP-3 nanoparticles of LMWH prepared by the emulsion-diffusion-evaporation method showed significant differences in particle morphology, size, zeta potential, and drug release profile compared to NP-1 nanoparticles formed by polyelectrolyte complexation. Another ionic complex of LMWH with chitosan-incorporated PLGA nanoparticles (NP-2) showed lower drug entrapment efficiency than that of NP-1 and NP-3. The drug release rate of NP-3 was slower than the release rates of NP-1 and NP-2, although particle morphology of NP-3 was similar to that of NP-2. Cell viability was not adversely affected when cells were treated with all three types of nanoparticles. The data presented in this study demonstrate that nanoparticles formulated with chitosan-PLGA could be a safe sustained-release carrier for the delivery of LMWH.

Laser-engineered dissolving microneedles for active transdermal delivery of nadroparin calcium

There is an urgent need to replace the injection currently used for low molecular weight heparin (LMWH) multidose therapy with a non- or minimally invasive delivery approach. In this study, laser-engineered dissolving microneedle (DMN) arrays fabricated from aqueous blends of 15% w/w poly(methylvinylether-co-maleic anhydride) were used for the first time in active transdermal delivery of the LMWH nadroparin calcium (NC). Importantly, an array loading of 630IU of NC was achieved without compromising the array mechanical strength or drug bioactivity. Application of NC-DMNs to dermatomed human skin (DHS) using the single-step ‘poke and release’ approach allowed permeation of approximately 10.6% of the total NC load over a 48-h study period. The cumulative amount of NC that permeated DHS at 24h and 48h attained 12.28±4.23IU/cm2 and 164.84±8.47IU/cm2, respectively. Skin permeation of NC could be modulated by controlling the DMN array variables, such as MN length and array density as well as application force to meet various clinical requirements including adjustment for body mass and renal function. NC-loaded DMN offers great potential as a relatively low-cost functional delivery system for enhanced transdermal delivery of LMWH and other macromolecules.

Chitosan nanoconstructs for improved oral delivery of low molecular weight heparin: In vitro and in vivo evaluation

The aim of present study was to investigate the potential of mucoadhesive polymer chitosan (CS) and N-trimethyl chitosan (TMC) based nanoparticulate systems for oral bioavailability enhancement of low molecular weight heparin (LMWH). The TMC was synthesized by methylation of chitosan followed by characterization using infrared spectroscopy and 1H-NMR spectroscopy. The IR and NMR spectra of TMC confirmed the presence of trimethyl groups and estimated the degree of quaternization for TMC about 46%. TMC nanoparticles were then prepared by ionic gelation method. The developed CS-NPs and TMC-NPs were characterized for various parameters including morphology, particle size, zeta potential, entrapment efficiency, in vitro release behavior and storage stability at different temperature and simulated gastrointestinal tract conditions. The fluorescent microscopy study confirmed the higher particle uptake of TMC-NPs by gastrointestinal epithelium in comparison to the CS-NPs. The concentration of LMWH in the systemic circulation followed by oral administration of formulations was estimated using FXa chromogenic assay. A significant increase ( p < 0.05) in the oral bioavailability of LMWH was observed with TMC-NPs than both CS-NPs as well as plain LMWH solution. These findings suggested that TMC nanoparicles hold promise for oral delivery of LMWH and clinical applicability for the treatment of vascular disorders like deep vein thrombosis and pulmonary embolism, etc.

Cytotoxicity assessment of heparin nanoparticles in NR8383 macrophages

The bioavailability of low molecular weight heparin (LMWH) has been increased by encapsulation in nanoparticles. As a complement to these results, the cytotoxicity and apoptosis induced by LMWH nanoparticles prepared by two methods [nanoprecipitation (NP) and double emulsion (DE)] using Eudragit ® RS (RS) and poly-ɛ-caprolactone (PCL) have been analysed. Particle sizes varied from 54 to 400 nm with zeta potential values between −65 and +63 mV. Our results showed that the method of nanoparticle preparation affects their properties, especially in terms of drug incorporation and cell tolerance. Cell viability ranged from 6% to 100% depending on the preparation method and physicochemical properties of the particles and the type of toxicity assay. Particle diameter and zeta potential seemed to be the most valuable cytotoxicity markers when cell viability was measured by Trypan blue exclusion and MTT respectively. Nanoparticles prepared by DE were better tolerated than those of NP. LMWH encapsulation into the cationic nanoparticles reduces remarkably their toxicity. Apoptosis evaluation showed activated caspases in exposed cells. However, no nuclear fragmentation was detected in NR8383 cells whatever the tested nanoparticles. DE nanoparticles of RS and PCL can be proposed as a good LMWH delivery system due to their low toxicity (IC 50 ∼ 2.33 and 0.96 mg/mL, respectively).

Inhalable Lactose-Based Dry Powder Formulations of Low Molecular Weight Heparin

Currently low molecular weight heparin (LMWH) is administered as subcutaneous injection. This study sought to investigate the feasibility of LMWH as an inhalable dry powder (DPI) formulation and evaluate the interaction of the drug with lactose when used as a carrier. The study also compares the extent of pulmonary absorption of LMWH administered as a dry powder with that administered as an aerosolized aqueous solution. The formulations were prepared by mixing LMWH in an aqueous solution of lactose followed by lyophilization of the resulting solution. The lyophilized preparation was then ground and sieved. Physical characterization of the formulations was performed by Fourier transform infrared spectroscopy (FTIR), X-ray powder diffraction (XRD), scanning electron microscopy (SEM), particle size analysis, and determination of aerodynamic diameter. For in vivo studies, formulations were administered to anesthetized rats, and drug absorption was monitored by measuring plasma antifactor Xa activity. In the FTIR scan, all characteristic peaks of lactose and LMWH were observed, suggesting that there was no strong interaction between lactose and LMWH. Although the aerodynamic diameter of the formulation (DPI-2) that was sieved through 170- and 230-mesh screens was similar to that of the formulation (DPI-1) sieved through 120- and 170-mesh screens, the particle sizes of the two formulations were significantly different. Dry powder formulations of LMWH were better absorbed compared to an inhalable solution of LMWH. One of the dry powder formulations (DPI-2) produced an almost 1.5-fold increase in the relative bioavailability (41.6%) compared to the liquid formulation of LMWH (32.5%). Overall, the data presented here suggest that lactose does not adversely affect the physical-chemical characteristics of the drug, and that lactose can be used as a carrier for pulmonary delivery of LMWH.

Enhancement of Transdermal Delivery of Heparin by Various Physical and Chemical Enhancement Techniques

Conventional anticoagulants such as unfractionated heparin and warfarin have numerous limitations compared with low-molecular-weight heparin (LMWH). However, the need for repetitive parenteral administration is still a major disadvantage of LMWH, and the absorption of macromolecules such as LMWH across the gastrointestinal tract is very poor. Due to these problems with oral delivery of LMWH, transdermal delivery can be considered as an alternate route of administration. However, overcoming the skin barrier is necessary for the transport of larger molecules across the stratum corneum. This review focuses on the transdermal delivery of LMWH, providing a brief overview of heparin delivery via invasive and oral routes and discusses the advantages of using LMWH rather than heparin for transdermal delivery, and the primary reasons for poor permeability of LMWH. Various strategies employed for transdermal delivery of heparin are summarized, and chemical and physical enhancement techniques or suitable formulations that can be used to improve transcutaneous penetration and various chemical enhancers that act on the skin by different modes of action are discussed. We also consider physical approaches such as iontophoresis, electroporation, and ultrasound, as well as combination strategies to deliver heparin. The developments in physical and chemical enhancement strategies over the past decade are summarized. In addition, recent novel approaches such as microneedles employed for the transdermal delivery of LMWH are also discussed.

Synthesis and Evaluation of Pegylated Dendrimeric Nanocarrier for Pulmonary Delivery of Low Molecular Weight Heparin

This study tests the hypothesis that pegylated dendrimeric micelles prolong the half-life of low molecular weight heparin (LMWH) and increase the drug's pulmonary absorption, thereby efficacious in preventing deep vein thrombosis (DVT) in a rodent model. Pegylated PAMAM dendrimer was synthesized by conjugating G3 PAMAM dendrimer with methyl ester of polyethylene glycol 2000 (PEG-2000). Fourier transform infrared (FTIR), nuclear magnetic resonance (NMR) spectra and thin layer chromatography (TLC) were used to evaluate the identity and purity of pegylated dendrimer. The particle size distributions of the formulations were measured by using a Nicomp Zeta meter, and drug entrapment efficiency was studied by azure A assay. The efficacy of pegylated dendrimers in enhancing pulmonary absorption, prolonging drug half-life, and preventing DVT was studied in a rodent model. FTIR, NMR and TLC data confirmed that PAMAM dendrimer was conjugated to PEG-2000. The entrapment efficiency of LMWH in PEG-dendrimer micelles was about 40%. Upon encapsulation of LMWH, the particle size of PEG-dendrimer micelles increased from 11.7 to 17.1 nm. LMWH entrapped in PEG-dendrimer produced a significant increase in pulmonary absorption and the relative bioavailability of the formulation was 60.6% compared to subcutaneous LMWH. The half-life of the PEG-dendrimer-based formulation was 11.9 h, which is 2.4-fold greater than the half-life of LMWH in a saline control formulation. When the formulation was administered at 48-h intervals, the efficacy of LMWH encapsulated in pegylated dendrimers in reducing thrombus weight in a rodent model was very similar to that of subcutaneous LMWH administered at 24-h intervals. Pegylated PAMAM dendrimer could potentially be used as a carrier for pulmonary delivery of LMWH for the long-term management of DVT.

Enhanced transdermal delivery of low molecular weight heparin by barrier perturbation

The purpose of this work was to investigate the in vitro transdermal delivery of low molecular weight heparin (LMWH). Hairless rat skin was mounted on Franz diffusion cells and treated with various enhancement strategies. Passive flux was essentially zero and remained low even after iontophoresis (0.065 U cm −2 h −1) or application of ultrasound (0.058 U cm −2 h −1). A significant increase in flux across tape stripped skin (4.0 U cm −2 h −1) suggests the interaction of stratum corneum (SC) with LMWH which was confirmed using Differential Scanning Calorimetry and Fourier Transform-Infrared spectrophotometry. Maltose microneedles were then employed as a means to locally disrupt and bypass the SC. Transepidermal water loss (TEWL) and transcutaneous electrical resistance (TER) were measured to confirm the barrier disruption. Microneedles breached the SC resulting in increased TEWL, decreased TER and enhanced LMWH permeability (0.175 U cm −2 h −1). Microneedles when used in conjunction with iontophoresis had a synergistic effect on LMWH delivery resulting in enhancement of flux by 14.7-fold as compared to iontophoresis used alone. Confocal laser scanning microscopy substantiated the evidence about LMWH interaction with SC. In conclusion, LMWH was shown to interact with SC and therefore tape stripping or microneedles dramatically increased its delivery due to disruption of the SC skin barrier.

Complexation of a Poly-l-Arginine with Low Molecular Weight Heparin Enhances Pulmonary Absorption of the Drug

This study tests the hypothesis that complexation between a cationic polymer, poly-L-arginine (PLA), and an anionic drug, low molecular weight heparin (LMWH), enhances pulmonary absorption and reduces the epithelial toxicity. Enoxaparin, a LMWH, was complexed with PLAs of different molecular weights at varying concentrations. The resulting complexes were characterized by measuring particle size and zeta potential, and by quantitating the interactions between PLA and enoxaparin using an azure A assay. Changes in transepithelial electrical resistance (TEER) and cytotoxicity induced by enoxaparin-PLA complex were investigated in Calu-3 cells. Pulmonary absorption of LMWH was determined by measuring plasma anti-factor Xa levels. A bronchoalveolar lavage (BAL) study was performed to investigate if the PLA-based formulations caused any cellular or biochemical changes in the lungs. The particle size of enoxaparin-PLA complexes decreased and the zeta potential values of the complex became less negative as the concentration of positively charged PLA in the complex increased. In vitro experiments suggest that addition of enoxaparin-PLA complex to the apical side of a polarized cell monolayer results in a significant increase in permeability to 14C-mannitol and a decrease in TEER. Pulmonary formulations of enoxaparin plus 0.0125% or 0.0625% PLA of molecular weight 93 kDa led to a twofold increase in the relative bioavailability of LMWH compared to the control (enoxaparin plus normal saline). The BAL study showed that the enoxaparin-PLA complex formulation did not elicit any significant increases in marker enzyme activities compared to the normal saline-treated or untreated control groups. PLA could be used as a carrier for the pulmonary delivery of LMWH.

Oral low molecular weight heparin delivery by microparticles from complex coacervation

As low molecular weight heparins exhibit limited oral absorption they usually have to be administered parenterally. Their strong negative charge appears to be one of the biggest hurdles to overcome in order to increase oral absorption. Complex coacervation has been proposed as a microencapsulation technique for increased oral drug absorption on the basis of charge compensation. Optimized tinzaparin/acacia gum mixture were coacervated with either gelatin A or B leading to microparticles with monodispersed size distribution, good fluidity and high encapsulation rates (>90%), while mean particle size varied between 5 and 20 μm, respectively, depending on the gelatin type. Tinzaparin was homogeneously distributed throughout the particle matrix and anti-Xa activity was maintained during preparation and storage. Drug release occurred in dependency of the pH triggering the dissociation between tinzaparin/acacia and gelatin. Cell binding experiments on Caco-2 led to slightly increased adhesion of gelatin A microparticles compared to gelatin B (A: 3.5 ± 0.3%; B: 2.5 ± 0.3%; solution: 1.9 ± 0.1%), while drug transport did not differ from free tinzaparin solution. In-vivo results demonstrated an oral bioavailability of about 4.2 ± 2.9% with gelatin B particles while gelatin A led to no absorption of tinzaparin. In conclusion, tinzaparin microparticles exhibited excellent particle properties in vitro and demonstrate potential for a formulation increasing the oral bioavailability of low molecular weight heparins.

Evaluation of the oral bioavailability of low molecular weight heparin formulated with glycyrrhetinic acid as permeation enhancer

Low molecular weight heparin (LMWH) is the agent of choice for anticoagulant therapy and prophylaxis of thrombosis and coronary syndromes. However, its therapeutic use is limited due to poor oral bioavailability. The aim of this study was to investigate the oral delivery of LMWH, ardeparin formulated with 18-beta glycyrrhetinic acid (GA), as an alternative to currently used subcutaneous (sc) delivery. Drug transport through Caco-2 cell monolayers was monitored in the presence and absence of GA by scintillation counting and transepithelial electrical resistance. Regional permeability studies using rat intestine were performed using a modified Ussing chamber. Cell viability in the presence of various concentrations of enhancer was determined by MTT assay. The absorption of ardeparin after oral administration in rats was measured by an anti-factor Xa assay. Furthermore, the eventual mucosal epithelial damage was histologically evaluated. Higher ardeparin permeability (~7-fold) compared to control was observed in the presence of 0.02 % GA. Regional permeability studies indicated predominant absorption in the duodenal segment. Cell viability studies showed no significant cytotoxicity below 0.01 % GA. Ardeparin oral bioavailability was significantly increased (F(relative)/(S.C). = 13.3%) without causing any damage to the intestinal tissues. GA enhanced the oral absorption of ardeparin both in vitro and in vivo. The oral formulation of ardeparin with GA could be absorbed in the intestine. These results suggest that GA may be used as an absorption enhancer for the oral delivery of LMWH.