Formulation and release of alaptide from cellulose-based hydrogels

The modern drug alaptide, synthetic dipeptide, shows regenerative effects and effects on the epitelisation process. A commercial product consisting of 1% alaptide hydrophilic cream is authorised for use in veterinary practice. This study focuses on the formulation of alaptide into semi-synthetic polymer-based hydrogels. The aim of the present study is to prepare hydrogels and to evaluate the liberation of alaptide from hydrogels. The hydrogels were prepared on the basis of three gel-producing substances: methylcellulose, hydroxyethylcellulose and hydroxypropylcellulose. To enhance the drug release from hydrogel humectants, glycerol, propylene glycol and ethanol in various concentrations were evaluated. The permeation of the alaptide from gels into the acceptor solution was evaluated with the use of the permeable membrane neprophane. The amount of drug released from prepared hydrogels was determined spectrophotometrically. Hydrogels with optimal alaptide liberation properties were subjected to the study of rheological properties in the next phase. The optimal composition of hydrogel as established in this study was 1% alaptide + 3% hydroxyethylcellulose with the addition of 10% glycerol as humectant. Due to the advantageous properties of hydrogels in wounds, alaptide could be incorporated into a hydrogel base for use in veterinary medicine. Epitelisation, gel-producing substance, humectant, methylcellulose, hydroxyethylcellulose, hydroxypropylcellulose Hydrogel research increased significantly in the last decades; due to their properties, hydrogels are widely used as suitable bases of preparations in human and veterinary medicine. The properties of hydrophilic gels, or hydrogels, are significantly different from those of ointments and creams. Hydrogels are usually formed by hydrophilic polymers which under certain conditions and polymer concentrations create a gel with a high percentage of water. They can additionally contain glycerol or propylene glycol as humectants and ethanol as an antimicrobial agent. Hydrogels create a smooth film that adheres well, can be removed easily and exhibits a cooling effect. The active substances have been dispersed in hydrogels (dissolved, suspended or emulsified). The base with the incorporated active ingredient must have suitable properties, especially biocompatibility and chemical, microbiological and physical stability (Bronaugh and Maibach 2001). Peptide drugs are becoming an essential part of modern therapy. Beside their therapeutic effect they can perform the function of another non-peptide drug carrier. Their reactions on changes of the pH milieu, biocompatibility, thermosensitivity and good mechanical properties have been reported (Galaev and Mattiasson 2008). Alaptide is spirocyclic synthetic dipeptide, chemically prolyl-leucyl-glycine amide derivative (Nedvidková et al. 1994). Due to its positive influence on regenerative processes in various skin diseases it can be used in veterinary medicine. For a long time it has been used as a commercial 1% hydrophilic cream that stimulates granulation tissue ACTA VET. BRNO 2012, 81: 301–306; doi:10.2754/avb201281030301 Address for correspondence: PharmDr. Zbyněk Sklenář, Ph.D., MBA Institute of Pharmacology 1st Faculty of Medicine, Charles University in Prague Albertov 4, 128 00 Prague 2, Czech Republic Phone: +420 224 968 104 E-mail: [email protected] http://actavet.vfu.cz/ growth and accelerates epithelisation and wound healing processes. It has been used for skin and mucosal injury treatment (lacerations, abrasions, anal fissures, small bite injuries), burns, frostbite. It may also be used for bedsores, ulcers, sutures, claw bed injuries, chronic relapsing injuries, post-operative wounds, dog paw burns by salt in winter, etc. The usual healing time is 3−10 days; in chronic cases the healing process takes longer. The mechanism of action of alaptide is probably due to its multiple water-binding ability and capability to improve regeneration. It is also possible to use alaptide in pregnant and nursing animals (Alaptide veterinary ointment 2011). Practice has shown that in wounds treated with alaptide, granulation tissue with marked epithelisation line rises and granulation progresses equally without hypergranulation. Thanks to the base, the wound surface remains moist, and traumatisation during rebandaging does not appear. Before application of a local topical preparation containing alaptide, it is necessary to clean the wound carefully, because the hydrophilic cream can worsen the drainage of wound secretions (Lexmaulová 2002). Using alaptide in a hydrogel base, it is expected that drug release and absorption by the wound will be improved. A hydrogel base is more physiologically appropriate for use in wounds; it is better tolerated and more easily washed out. The aim of this study was the incorporation of the drug alaptide into a hydrogel formulation based on semi-synthetic polymers – cellulose derivatives, and its liberation through the permeable membrane nephrophan. The composition of this membrane is very similar to that of human and animal membranes. In particular, it is commonly used in clinical practice as a dialysance membrane. The release is enhanced by glycerol, propylene glycol and ethanol, where glycerol and propylene glycol also act as humectants. Based on a previous study of alaptide-liberation from dermatological semi-solid chitosan-based preparations, it has been found that glycerol and propylene glycol can have an influence on alaptide release in dermatological semi-solid preparations (Šimunková et al. 2008). Polymer concentrations have been chosen on the basis of previous experiments with drug liberation and rheologic properties of hydrogels. In previous works the influence of various membranes on alaptide permeation from hydrogels (Vitková et al. 2011) has been evaluated. Four membranes – cellophane, chicken skin, snake skin and intestinal wall were studied. It was found that the best permeation occurred through the intestinal wall, less through chicken skin and the least through snake skin. Materials and Methods Chemicals Alaptide (ALA) – micronised form (Výzkumný ústav pro farmacii a biochemii –VÚFB, Czech Republic), ethanol 96% (V/V) (ETH) (Ethanolum 96% SL 1 – Liehovary, Leopoldov, Slovakia), glycerol 85% (GLY) (Galvex spol. s.r.o., Banská Bystrica, Slovakia), hydroxyethylcellulose – Natrosol 250 HR (HEC) (Aqualon, USA), hydroxypropylcellulose – Klucel MCS (HPC) (Aqualon, USA), lactic acid (Lachema, o. p., Brno, Czech Republic), methylcellulose (MC) (Aqualon, USA), preservative agent – Sepicide HBR (Sep. HBR) (SEPPIC, Paris, France; composition: butyl p-hydroxybenzoate, ethyl p-hydroxybenzoate, sodium methyl p-hydroxybenzoate and propyl p-hydroxybenzoate in phenoxyethanol), propylene glycol (PPG) (Merck, Darmstadt, Germany), purified water (Aqua purificata SL 1– FaF UK, Bratislava, Slovakia), sodium chloride (Centrachem, Switzerland). Equipment Analytic scales – Kern & Sohn GmbH (Germany), electric mixer – VEB ML W Prűfgeräte – Werk (Germany), Viskotester VT 500 – Haake Mess-Technik GmbH (Germany), pH meter – WTW Weilheim (Switzerland), spectrophotometer – Philips Pye Unicam (Great Britain), permeation equipment, permeation membrane (Nephrophan) – VEB Filmfabrik Wolfen (Germany). Hydrogel composition Gels based on three semi-synthetic cellulose polymers were prepared containing 2% methylcellulose, 3% hydroxyethylcellulose or 2.5% hydroxypropylcellulose with suspended alaptide at a 1% concentration. Humectant addition in single hydrogels was 5% PPG, 10% GLY and 15% ethanol. Drug liberation and content determination Drug liberation from hydrogel was evaluated 7 and 14 days after preparation in 6 parallel evaluations. 302