Cross-linked hyaluronic acid hydrogel films: new biomaterials for drug delivery

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Abstract

A new hyaluronic acid (HA)-based hydrogel film was prepared and evaluated for use in drug delivery. This biocompatible material crosslinks and gels in minutes, and the dried film swells and rehydrates to a flexible hydrogel in seconds. HA was first converted to the adipic dihydrazide derivative and then crosslinked with the macromolecular homobifunctional reagent poly(ethylene glycol)–propiondialdehyde to give a polymer network. After gelation, a solvent casting method was used to obtain a HA hydrogel film. The dried film swelled sevenfold in volume in buffer, reaching equilibrium in less than 100 s. Scanning electron microscopy (SEM) of the hydrogel films showed a condensed and featureless structure before swelling, but a porous microstructure when hydrated. The thermal behavior of the hydrogel films was characterized by differential scanning calorimetry. The enzymatic degradation of the HA hydrogel films by hyaluronidase was studied using both SEM and a spectrophotometric assay. Drug release from the hydrogel film was evaluated in vitro using selected anti-bacterial and anti-inflammatory drugs. This novel biomaterial can be employed for controlled release of therapeutic agents at wound sites.

Introduction

Hyaluronic acid (HA) is a naturally-occurring linear polysaccharide comprised of β-1,4-linked d-glucuronic acid (β-1,3) N-acetyl-d-glucosamine disaccharide units, and it is the only non-sulfated glycosaminoglycan (GAG) in the extracellular matrix (ECM) of all higher animals. This polyanionic polymer has a range of naturally-occurring molecular sizes from 1000 to 10,000,000 Da, and has unique physicochemical properties and distinctive biological functions [1]. HA has been implicated in water homeostasis of tissues, in the regulation of the permeability of other substances by steric exclusion phenomena, and in the lubrication of joints [2]. HA also binds specifically to proteins in the ECM, on the cell surface, and within the cell cytosol, thereby having a role in cartilage matrix stabilization [2], [3], cell motility [4], [5], growth factor action [6], morphogenesis and embryonic development [7], and inflammation [8].

Unmodified HA has many important applications in drug delivery and surgery. For example, HA is used as an adjuvant for ophthalmic drug delivery [9], and was found to enhance the absorption of drugs and proteins via mucosal tissues [10], [11], [12], [13]. In addition, HA has important applications in the fields of viscosurgery, viscosupplementation, and wound healing [14], [15], [16].

HA is also an attractive building block for new biocompatible and biodegradable polymers with applications in drug delivery, tissue engineering, and viscosupplementation [17], [18]. Chemical modification [19] allows the physicochemical properties and in vivo residence time of HA to be tailored to specific applications while retaining its natural biocompatibility, biodegradability, and lack of immunogenicity [18]. Composite materials containing HA [20], [21], [22], functional modification of carboxylic groups on HA, polymer–drug conjugates for controlled drug release [23], crosslinking into hydrogels and surface coating [18], and uses of HA-derived biochemical probes [24] have been recently summarized. Many HA-derived biomaterials have been evaluated in vitro and in vivo as potential biomedical devices [25]. For example, a new internally-esterified material, the HA autocrosslinked polymer hydrogel (ACP gel), was found to be effective in reducing postsurgical adhesions in laparoscopic surgery [26]. In addition, esterification of HA with various therapeutically-active and/or inactive alcohols produced mucoadhesive biopolymers with new physicochemical properties [27], [28] for use in controlled drug release and tissue engineering [29].

Obtaining HA-derived biomaterials with sufficient strength to withstand biomechanically stressful applications has been problematic. HA derivatives with the best mechanical properties have generally been prepared with highly alkaline crosslinking conditions and at elevated temperatures [30], [31], conditions that preclude inclusion of sensitive molecules or living cells during preparation of the polymer network hydrogel. Additionally, such reactions employed small molecular crosslinking reagents, often in large excess, which required considerable purification in order to obtain materials for physiological use.

We describe herein a new HA hydrogel well-suited to drug delivery, prepared using a macromolecular crosslinker under neutral aqueous conditions. This new biocompatible material crosslinks and gels in minutes and swells from a flexible dry film to a flexible porous hydrogel in seconds. The hydrogel can be used directly in any biological system without purification after gelling, since it only contains two biocompatible polymers. The swelling properties of the hydrogels, including equilibrium and swelling kinetics, were evaluated. Enzymatic degradation of the hydrogels by hyaluronidase (HAse), and in vitro release of anti-bacterial and anti-inflammatory drugs were examined.

Section snippets

Materials

Fermentation-derived hyaluronan (HA, sodium salt, Mr=1.5×106) was provided by Clear Solutions Biotechnology, Inc. (Stony Brook, NY). 1-Ethyl-3-[3-(dimethylamino)-propyl]carbodiimide (EDCI), adipic dihydrazide (ADH), and acridine orange were purchased from Aldrich Chemical Co. (Milwaukee, WI). Bovine testicular hyaluronidase (HAse, 880 U/mg), indomethacin, pilocarpine, diclofenac sodium, hydrocortisone, 6α-methyl-prednisolone, prednisolone, cortisone, corticosterone, dexamethasone, prednisone,

Results and discussion

Tissue-compatible hydrogels provide a natural, hydrophilic environment for controlled drug delivery [36], and the rate of drug release can be regulated by controlling gel-swelling and crosslinking density. Crosslinked HA hydrogels [37] have been prepared with diepoxybutane, ethyleneglycol diglycidylether (EGDGE), or polyglycerol polyglycidylether (PGPGE) in strongly alkaline conditions with ethanol as a co-solvent [30], [31]. These crosslinking methods produce HA hydrogel films with water

Conclusion

In conclusion, a novel fast-gelling and fast-swelling HA hydrogel was developed as a potential drug delivery system. This polymer network hydrogel was prepared from HA–ADH and a macromolecular crosslinker, PEG–diald. SEM images showed that the network structure of the hydrogel film is condensed in the dried state, and becomes porous after swelling. The thermal behavior of the HA hydrogel and its components were studied by DSC, which revealed that the hydrogel had a microstructure that differed

Acknowledgements

Financial support for this work was provided by Center for Biopolymers at Interfaces (The University of Utah (UUtah)), and by the start-up funds provided by UUtah to GDP. We thank C. Wang, Professor J. Kopecek, and Professor S.-W. Kim for valuable discussions and experimental assistance.

References (51)

  • M. Rochira et al.

    Novel vaginal delivery systems for calcitonin. 2. Preparation and characterization of HYAFF(R) microspheres containing calcitonin

    Int. J. Pharm.

    (1996)
  • P.T. Prisell et al.

    Evaluation of hyaluronan as a vehicle for peptide growth factors

    Int. J. Pharm.

    (1992)
  • S.P. Zhong et al.

    Biodegradation of hyaluronic acid derivatives by hyaluronidase

    Biomaterials

    (1994)
  • D. Papini et al.

    Diffusion of macromolecules in membranes of hyaluronic acid esters

    J. Control. Release

    (1993)
  • L.M. Benedetti et al.

    Microspheres of hyaluronic acid esters — fabrication methods and in vitro hydrocortisone release

    J. Control. Release

    (1990)
  • T.C. Laurent et al.

    Functions of hyaluronan

    Ann. Rheum. Dis.

    (1995)
  • J.R.E. Fraser et al.

    Hyaluronan: its nature, distribution, functions and turnover

    J. Intern. Med.

    (1997)
  • G.P. Dowthwaite et al.

    An essential role for the interaction between hyaluronan and hyaluronan binding proteins during joint development

    J. Histochem. Cytochem.

    (1998)
  • C. Hardwick et al.

    Molecular cloning of a novel hyaluronan receptor that mediates tumor cell motility

    J. Cell Biol.

    (1992)
  • W.F. Cheung et al.

    Receptor for hyaluronan-mediated motility (RHAMM), a hyaladherin that regulates cell responses to growth factors

    Biochem. Soc. Trans.

    (1999)
  • B.P. Toole

    Hyaluronan in morphogenesis

    J. Intern. Med.

    (1997)
  • B. Gerdin et al.

    Dynamic role of hyaluronan (HYA) in connective tissue activation and inflammation

    J. Intern. Med.

    (1997)
  • M.F. Saettone et al.

    Mucoadhesive ophthalmic vehicles: evaluation of polymeric low-viscosity formulations

    J. Ocular Pharm.

    (1994)
  • A.R. Moore et al.

    Hyaluronan as a drug delivery system for diclofenac: a hypothesis for mode of action

    Int. J. Tissue React.

    (1995)
  • K. Morimoto et al.

    Effects of viscous hyaluronate-sodium solutions on the nasal absorption of vasopressin and an analogue

    Pharm. Res.

    (1991)
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