Bioavailability enhancement of glucosamine hydrochloride by chitosan
Graphical abstract
Introduction
Osteoarthritis is a degenerative disease leading to chronic joint pain and limitation of movement. Typical features of osteoarthritis are the degeneration or progressive loss of the structure and functionality of articular cartilage (Lorenz and Richter, 2006). Cartilage, a flexible connective tissue, is composed of chondroblasts that produce a large amount of extracellular matrix composed of glycosaminoglycans and proteoglycans (Murray and Keeley, 2009). Glucosamine, also known as 2-amino-2-deoxyglucose (CAS 3416-24-8) (Fig. 1), is an amino sugar biosynthesized endogenously in animals and human. Glucosamine functions as a precursor for glycosaminoglycans and is structurally incorporated in mucopolysaccharides, glycoproteins and proteoglycans. As a food supplement, glucosamine may benefit the generation of cartilage structure, and has been applied to facilitate the alleviation of arthritis (Matheson and Perry, 2003, Rovati et al., 2012, Towheed et al., 2005). Although glucosamine has not been approved by US FDA as a drug, it is regarded as a prescription drug for osteoarthritis in Europe (Russell et al., 2002). However, the oral bioavailability of glucosamine is relatively poor in the vertebrate (6.1–26% in human (Meulyzer et al., 2008, Setnikar et al., 1993), ∼19% in rats (Aghazadeh-Habashi et al., 2002) and ∼12% in dogs (Adebowale et al., 2002)), which may be due to its transport-facilitated limited absorption and presystemic loss in gut and liver (Ibrahim et al., 2012, Setnikar et al., 1993). As a dietary supplement, glucosamine is commercially available in hydrochloride and sulphate salt forms with high water solubility, which demonstrate bioequivalence in vivo (Aghazadeh-Habashi and Jamali, 2011) and linear pharmacokinetics in the dose range of 750–1500 mg in human (Persiani et al., 2005). However, following a conventional dose (1500 mg/day), the plasma concentration achieved (2.7–17.4 μM) is far below the required level for response (Ibrahim et al., 2012). The low and variable bioavailability of glucosamine could have led to inconsistency of clinical trial outcomes, which further casts doubt on its use for clinical treatment of osteoarthritis (Aghazadeh-Habashi and Jamali, 2011, Towheed and Anastassiades, 2007). Therefore, improvement in the oral bioavailability of glucosamine is highly desirable.
Glucosamine is a small hydrophilic molecule like glucose, which cannot easily cross the biological membrane via transcellular pathway. It has reported to be a substrate of glucose facilitative transporters (especially GLUT2) (Uldry et al., 2002), which could be inhibited by flavonoid quercetin (Ibrahim et al., 2012) and the transport process can be saturable (Klip and Paquet, 1990, Persiani et al., 2005). Paracellular transport can also play an important role for the transport of small hydrophilic drugs (especially at the upper small intestine with leaky barriers) (Lafforgue et al., 2008, Sugano et al., 2010). Increasing the paracellular transport of glucosamine is expected to lead to an enhancement in its absorption.
The primary aim of our study was to find an approach to improve the oral bioavailability of glucosamine. Although several classes of compounds, including surfactants, fatty acids, cyclodextrins, cellulose ethers, chelators, and positive charged polymers (Junginger, 2007), are known to be able to enhance absorption of low permeable hydrophilic drugs by acting on the mucous layer, the membrane components or the tight junctions of the intestinal epithelium, it is uncertain whether any of them is suitable for glucosamine absorption enhancement or not. In fact, during our previous formulation development of glucosamine, it was found that: (1) addition of cellulose ethers (including CMC-Na and HPMC) even lead to a lower absorption of glucosamine in rats (data not shown); (2) surfactants like SLS (high toxicity to the intestinal membrane (Uchiyama et al., 1999)) and Tween 80 (odoriferous smell and bitter taste) have safety or taste concerns. In our preliminary study in rats, addition of 1% chitosan (positive charged polymer) exhibited a significant enhancement of glucosamine oral bioavailability, which prompted us to investigate further on the mechanism of such absorption enhancement as well as to optimize this chitosan containing formulation.
Chitosan is a natural and cationic polysaccharide obtained by deacetylation of chitin, a polymer which is abundant in the outer skeleton of crustaceous shells such as crabs and shrimps. Chitosan has been approved as food supplement in many countries and is commonly regarded as non-toxic, biological compatible and biodegradable substance (Kean and Thanou, 2010). With a pKa value of ∼6.5, chitosan exhibits a pH-dependent solubility with high solubility at low pH but insoluble at higher pH ranges. It is soluble in dilute HCl, HNO3, and HClO4. However, chitosan is sparing soluble in H2SO4 at room temperature and incompatible with drugs in sulphate form (Tu et al., 2010). An important property is its ability to reversibly interact with components at the epithelial tight junction when chitosan is protonated in its uncoiled configuration at pH below 6.5, leading to widening of the paracellular route temporarily and increasing the paracellular permeability of drugs across mucosal epithelia (Thanou et al., 2001). A number of studies have confirmed the improvement of permeability of poorly absorbed hydrophilic compounds such as atenolol (Schipper et al., 1999), mannitol (Artursson et al., 1994), dextran (Vllasaliu et al., 2012), insulin (Fernandez-Urrusuno et al., 1999) and salmon calcitonin (Sinswat and Tengamnuay, 2003) across the intestinal or nasal epithelia.
In the present study, we first test its ability by a well-established in vitro model, Caco-2 cell model (Artursson et al., 2001, Fong et al., 2012, Zhang et al., 2006, Zhang et al., 2007), followed by animal studies (rats and beagle dogs) to verify the absorption enhancement.
Section snippets
Materials
Considering the compatibility with chitosan, glucosamine hydrochloride has been chosen for the current study. d-(+)-glucosamine hydrochloride (referred as “glucosamine” in this paper) was purchased from Qingdao Highsun Biochemical Products Co., Ltd., China. Chitosan hydrochloride (referred as “chitosan”) with average molecular weight of ∼200 kDa and deacetylation degree of 83% was purchased from Zhejiang Golden-Shell Biomedical Co., Ltd., China. 9-Fluorenylmethyl chloroformate (FMOC-Cl), sodium
Cytotoxicity of glucosamine and chitosan to Caco-2 cells
The cytotoxicity profile of glucosamine and chitosan on Caco-2 cells was shown in Fig. 2. For glucosamine (Fig. 2A), the results showed that the relative viability of Caco-2 cells maintained to be greater than 80% after 4 h incubation with glucosamine concentrations up to 3 mg/ml in PBS+ 6.0 and 1 mg/ml in PBS+ 7.4.
For chitosan (Fig. 2B), the cell viabilities did not remarkably change over 1–100 μg/ml in PBS+ 7.0 and 1–50 μg/ml in PBS+ 7.4, whereas a concentration-dependent cytotoxicity behaviors
Discussion
The present in vitro and in vivo studies showed a significant enhancement of oral absorption of glucosamine by chitosan. Our results suggest that a glucosamine formulation with chitosan could potentially provide enhanced therapeutic effect at conventional dose. Further clinical studies, however, are needed to confirm such results.
In general, chitosans with high degree of deacetylation (DDA > 80%) and high molecular weight were preferred for absorption enhancement of hydrophilic compounds due to
Conclusion
Our study showed that chitosan could significantly enhance the intestinal permeability of glucosamine by reversibly opening the tight junction of intestinal epithelium cells. Further in vivo studies in rats and beagle dogs demonstrated that the presence of chitosan could increase the plasma concentration and bioavailability of glucosamine without altering its elimination. Chitosan can serve as an effective absorption enhancer for glucosamine.
Acknowledgments
Financial support from General Research Fund CUHK 480809 and Comprehensive Drug Enterprises Limited; Technical support from Dr. Zhao, Lizi, Mr. Zeng, Guixiong in Sun Yat-sen University.
References (67)
- et al.
Epithelial transport of drugs in cell-culture. VIII: effects of sodium dodecyl-sulfate on cell-membrane and tight junction permeability in human intestinal epithelial (Caco-2) cells
J. Pharm. Sci.
(1993) - et al.
Caco-2 monolayers in experimental and theoretical predictions of drug transport
Adv. Drug Deliv. Rev.
(2001) - et al.
Influence of molecular weight on oral absorption of water soluble chitosans
J. Control. Release
(2005) - et al.
Effect of chitosan on epithelial permeability and structure
Int. J. Pharm.
(1999) - et al.
In vitro and in situ evaluation of herb–drug interactions during intestinal metabolism and absorption of baicalein
J. Ethnopharmacol.
(2012) Introduction: claudins, tight junctions, and the paracellular barrier
- et al.
Improvement of intestinal absorption of insulin and water-soluble macromolecular compounds by chitosan oligomers in rats
Int. J. Pharm.
(2008) - et al.
Absorption and bioavailability of glucosamine in the rat
J. Pharm. Sci.
(2012) - et al.
Trimethylated chitosans as non-viral gene delivery vectors: cytotoxicity and transfection efficiency
J. Control. Release
(2005) - et al.
Biodegradation, biodistribution and toxicity of chitosan
Adv. Drug Deliv. Rev.
(2010)
In vitro evaluation of mucoadhesive properties of chitosan and some other natural polymers
Int. J. Pharm.
Osteoarthritis: cellular and molecular changes in degenerating cartilage
Prog. Histochem. Cytochem.
Comparison of pharmacokinetics of glucosamine and synovial fluid levels following administration of glucosamine sulphate or glucosamine hydrochloride
Osteoarthr. Cartil.
Rapid colorimetric assay for cellular growth and survival: application to proliferation and cytotoxicity assays
J. Immunol. Methods
pH-Dependent passive and active transport of acidic drugs across Caco-2 cell monolayers
Eur. J. Pharm. Sci.
Glucosamine oral bioavailability and plasma pharmacokinetics after increasing doses of crystalline glucosamine sulfate in man
Osteoarthr. Cartil.
Characterization of chitosan hydrochloride-mucin interaction by means of viscosimetric and turbidimetric measurements
Eur. J. Pharm. Sci.
Chitosans as absorption enhancers of poorly absorbable drugs. 3: influence of mucus on absorption enhancement
Eur. J. Pharm. Sci.
Enhancing effect of chitosan on nasal absorption of salmon calcitonin in rats: comparison with hydroxypropyl- and dimethyl-beta-cyclodextrins
Int. J. Pharm.
Oral drug absorption enhancement by chitosan and its derivatives
Adv. Drug Deliv. Rev.
GLUT2 is a high affinity glucosamine transporter
FEBS Lett.
Absorption-promoting effects of chitosan in airway and intestinal cell lines: a comparative study
Int. J. Pharm.
Mechanistic study on the intestinal absorption and disposition of baicalein
Eur. J. Pharm. Sci.
Improvement of intestinal absorption of forsythoside A in weeping forsythia extract by various absorption enhancers based on tight junctions
Phytomedicine
The bioavailability and pharmacokinetics of glucosamine hydrochloride and low molecular weight chondroitin sulfate after single and multiple doses to beagle dogs
Biopharm. Drug Dispos.
The glucosamine controversy; a pharmacokinetic issue
J. Pharm. Pharm. Sci.
Single dose pharmacokinetics and bioavailability of glucosamine in the rat
J. Pharm. Pharm. Sci.
Effect of chitosan on the permeability of monolayers of intestinal epithelial cells (Caco-2)
Pharm. Res.
Absorption enhancers: applications and advances
AAPS J.
Upper gastrointestinal (GI) pH in young, healthy men and women
Pharm. Res.
Enhancement of nasal absorption of insulin using chitosan nanoparticles
Pharm. Res.
Probit Analysis: A Statistical Treatment of the Sigmoid Response Curve
Influence of chitosans on permeability of human intestinal epithelial (Caco-2) cells: the effect of molecular weight, degree of deacetylation and exposure time
Cited by (31)
Nanoconfined ReS<inf>2</inf> in biomass-derived 3D porous N-doped carbon architecture as anode for stable lithium-ion storage
2021, Electrochimica ActaCitation Excerpt :More detailed information about the chemical components and local structures of the ReS2NC2 were provided by XPS. As depicted from Fig. 6a, the spectra of Re 4f are inclusive of two peaks ascribed to Re 4f7/2 and Re 4f5/2, which are situated at 41.4 and 43.8 eV, respectively [36–39]. The S spectra (Fig. 6b) can be devided into a couple of peaks located at a binding energy of 162.0 and 163.4 eV, which is attributed to S 2p3/2 and S 2p1/2 mode, respectively [36–39].
Preparation and drug release property of tanshinone IIA loaded chitosan-montmorillonite microspheres
2019, International Journal of Biological MacromoleculesCitation Excerpt :Enlightened from the above points, it is expected that the TA molecules dissolved in ethyl alcohol should be effectively loaded on the CS matrix during the dehydration process. CS, a natural cationic polysaccharide derived from chitin, has been widely used for the matrix of drug delivery system due to its high biocompatibility, biodegradability, and non-toxicity [16,17]. Because of the abundant NH2 and OH groups in CS backbone and its positively charged surface, CS can be easily prepared into different carrier materials, such as microspheres [18], nanoparticles [19], hydrogels [20], microgels [21], films [22], and other materials [23].
Hierarchical flower-like SnS grafted with glucosamine-derived nitrogen-doped carbon with enhanced reversible Li-storage performance
2018, Applied Surface ScienceCitation Excerpt :Recently, biomass derived nanostructured nitrogen doped carbon has been widely investigated for their prominent performance for applications in energy conversion and storage [32,33]. Glucosamine can be easily extracted from naturally abundant chitin and commercially available in bulk quantity and can be employed as the precursor of nitrogen doped carbons [34,35]. This paper presents a simple hydrothermal reaction followed by an annealing procedure to prepare flower-like SnS/nitrogen doped carbon (SnS/NC) composites with glucosamine hydrochloride as carbon and nitrogen sources.
Enhanced oral bioavailability of docetaxel in rats combined with myricetin: In situ and in vivo evidences
2017, European Journal of Pharmaceutical Sciences
- 1
These authors contributed equally in the experimental works.