http://informahealthcare.com/phdISSN: 1083-7450 (print), 1097-9867 (electronic)

Pharm Dev Technol, Early Online: 1–4! 2014 Informa Healthcare USA, Inc. DOI: 10.3109/10837450.2014.965327

TECHNICAL NOTE

Novel pectin-based carriers for colonic drug delivery

Wujie Zhang1, Kirsten Mary Mahuta1*, Brandon Anthony Mikulski1*, Jenna Nicole Harvestine1*, James ZacharyCrouse1*, Jung Chull Lee1, Matey Georgiev Kaltchev1, and Charles Samuel Tritt2

1Biomolecular Engineering Program, Department of Physics & Chemistry, 2Biomedical Engineering Program, Department of Electrical Engineering

and Computer Science, Milwaukee School of Engineering, Milwaukee, WI, USA

Abstract

Pectin-based hydrogel carriers have been studied and shown to have promising applicationsfor drug delivery to the lower GI tract, especially to the colonic region. However, making surethese hydrogel carriers can pass through the upper GI tract and reach the targeted regions,after oral administration, still remains a challenge to overcome. A solution to this problem is topromote stronger cross-linking interactions within the pectin-based hydrogel network. Thecombined usage of a divalent cation (Ca2+) and the cationic biopolymer oligochitosan hasshown to improve the stability of pectin-based hydrogel systems – suggesting that these twocross-linkers may be used to eventually help improve pectin-based hydrogel systems forcolonic drug delivery methods.

Keywords

Colonic drug-delivery, hydrogel,microcapsule, microsphere, pectin

History

Received 20 August 2014Revised 28 August 2014Accepted 8 September 2014Published online 25 September 2014

Introduction

There is an increasing need for colonic drug delivery systems fortherapies such as colon cancer treatment. Pectin-based hydrogelmicrosphere systems have shown great potential for drug deliveryto the lower GI-tract, especially to the colon1–4. Pectin is anaturally occurring polysaccharide found in citrus fruits and applepeels, composed of galacturonic acid with carboxyl groups, and ischaracterized by the degree of methyl esterification. In general,the low methyl (LM) pectin has been shown to have excellentcapability for hydrogel formation5–8.

The advantages for using a pectin-based drug delivery systeminclude its dual-responsiveness to pH (releasing the drug at a higherpH) as well as to its microbial enzymatic degradation at the colonsite3,4,9–11. Recently, pectin-based systems have been tested forcolonic drug delivery/release. For example, the chitosan/pectinpolyelectrolyte complex systems were investigated for the colon-specific delivery of vancomycin (abrupt release in the presence ofbeta-glucosidase)12 and theophylline (98.58 ± 1.92% drug releasein presence of rat fecal content)13. Moreover, Newton et al.investigated the in vitro dissolution profile of colon targeted HPMCE15 LV/Pectin/Chitosan matrix tablets, confirming that theprepared matrices could achieve targeted colon drug release14.Jung et al. showed that the enzymatically charged modified pectin-based hydrogel could be used as a colon-targeted drug deliverycarrier with indomethacin as a model drug1. Ramteke et al. alsoshowed its potential to deliver glipizide to the colonic region byusing the Pectin-Bora rice beads, especially with the in vivo gammascintigraphy evidence, revealing the beads showed degradation

upon reaching the colon15. Aside from chitosan-coating, speciallycoated calcium pectinate beads have been designed to reduceswelling of the beads and delay the drug release for colonic drugdelivery of theophylline (silica-coated; organic-inorganic hybridbeads)16 and curcumin (Eudragit� S100 coated, Rohm Pharma,Darmstadt, Germany)7. On the contrary, pectin-coating has alsobeen demonstrated as a potential route of developing colon-targeted drug delivery systems, e.g. pectin-coated chitosan-LDHbionanocomposite beads17. Interestingly, p53 polyplex-loadedenteric-coated calcium pectinate microbeads were evaluated fororal gene delivery for colorectal cancer therapy. Less than 10% ofrelease was observed within the upper GI tract, their in vivo studiesconfirmed that this type of novel microbeads could be used todeliver pDNA specifically to rat colons18.

The pectin-based delivery system’s pH responsiveness ismainly due to the pH-dependent ionization (at a lower pH) ofthe carboxyl groups of pectin19. This pH-dependent responsive-ness of the pectin-based carrier is desirable, considering there isa gradual pH increase in the body fluid from the upper to thelower GI tract. However, the distinct challenge is to keep the drugcarrier intact while retaining the drug within its carrier, since theintestinal fluid pH could be at a level that is high enough to causethe disintegration of the pectin-based hydrogel systems beforethey reach the colonic region. Microbial enzymes can also play amajor role in the hydrogel disintegration process. For example,trypsin, found in the intestinal fluid, captures divalent calciumcations (Ca2+) from the hydrogels, which in turn act as cofactorsto increase the enzyme activity20,21. This subsequently facilitatesthe dissociation of the pectin-based hydrogel systems.

Recently, it has been reported that the oligochitosan, a naturalsaccharide derived from crab and other crustacean shells, couldalso serve as a cross-linker to form the hydrogel micro-spheres22–24. These microspheres exhibit a red blood cell-likeshape – possibly because of the lower diffusivity of oligochitosanwithin the pectin solution as compared to Ca2+, and this may

*These authors contributed equally to this work.

Address for correspondence: Wujie Zhang, Biomolecular EngineeringProgram, Department of Physics & Chemistry, Milwaukee School ofEngineering, Milwaukee, WI 53202, USA. E-mail: zhang@msoe.edu

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occur, depending on the orientations of the pectin molecules (i.e.crosslinking of pectin molecules), within the hydrogel. It is ofgreat interest to investigate whether the change of the cross-linkeror the combination of different cross-linkers could lead to a morestable hydrogel microsphere at a higher pH (�6.8), eventuallyachieving a successful colonic drug delivery. This method isadvantageous because it is simpler and less time-consuming thanthe approaches based on chemically modified pectin or coatedsystems3,4,7,12,13,16,25.

Methods

Pectin-based microspheres were prepared by using a BUCHI�

encapsulator device (electrostatic spray/vibration method)22.Briefly, 3% (w/v) LM pectin (20.4% esterification; Willpowder;Miami Beach, FL) was sprayed into a cross-linker-containinggelling solution (pH: �6) to form hydrogel microspheres. The150 mM CaCl2, 5% (w/v) oligochitosan (MW: 2–3 KDa; 95%deacetylation; Zhejiang Golden-Shell Pharmaceutical Co. Ltd;Zhejiang, China), and 75 mM CaCl2/2.5% (w/v) oligochitosansolution was used as the gelation solution to prepare the Calcium-Pectin (Ca-Pectin; i.e. Calcium Pectinate), Oligochitosan-Pectin,and Ca-Oligochitosan-Pectin microspheres, respectively. The

microspheres were formed either by intermolecular chelatebinding of cations (Ca2+) (formation of macromolecular aggre-gates; egg-box structure) and/or the formation of pectin-oligochitosan electrolyte complexes (pectin and chitosan havecomplementary charges at a pH around 6), depending on thetype of the gelation solution used (Figure 1). The differencesin morphology of these three types of hydrogel microspheres(Table 1) are most likely due to the diffusion/gelation speed of thecross-linkers. The morphology of a carrier, which determines thesurface area-to-volume ratio, could potentially influence its drugdelivery/release performance22.

In vitro responsiveness of different types of pectin-basedhydrogel microspheres were investigated using simulated gastric(containing 0.32 g pepsin, 0.2 g NaCl and 0.7 mL hydrochloricacid per 100 mL solution; pH: 2) and intestinal (containing 1 gtrypsin and 0.68 g KH2PO4 per 100 mL solution; pH: 6.8) fluidsas well as using a solution of a pectinase enzyme (5.7 U/mL(1.5 mL enzyme solution/1 mL microsphere solution); mimickingcolonic enzyme)7,26,27. These microspheres were then placed ina shaker and checked for their morphology and integrity every10 min under an optical microscope. Images were processed byusing the NIH ImageJ software (Bethesda, MD) to determine themicrospheres’ average size.

Table 1. Comparison of different pectin-based hydrogel systems.

Stability in simulated fluids

System Shape Typical image Gastric Intestinal

Ca-pectin Spherical High Low

Oligochitosan-pectin Red blood cell-like High Medium

Ca-oligochitosan-pectin Oblate spheroid High High

Figure 1. Schematic showing the possiblemolecular structure of the Ca-Oligochitosan-Pectin hydrogel.

2 W. Zhang et al. Pharm Dev Technol, Early Online: 1–4

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Results and discussion

All three types of microspheres remained stable and intact withinthe simulated gastric fluid, with no significant swelling(Figure 2), which means they could be used to shield theencapsulated drug(s) from stomach degradation and/or releaseafter oral administration. When placed in the simulated intestinalfluid, Ca-Pectin and Oligochitosan-Pectin microspheres started toswell and then immediately disintegrated (Figure 2). However, theCa-Oligochitosan-Pectin microspheres stayed intact with onlyslight swelling (3.4%) during the entire 2-hour-long testingperiod, showing an improved stability when using both Ca2+ andoligochitosan as cross-linkers during the hydrogel formationprocess. Improved stability suggests the Ca-Oligochitosan-Pectinmicrospheres have potential as a drug carrier to pass through thesmall intestine and reach the colonic region. In addition, all thesethree types of microspheres are responsive to the pectinase,decreasing in size over time. The microscopic images, demon-strating the responsiveness of the Ca-Oligochitosan-Pectin micro-spheres to the pectinase, are shown in Figure 3. The averagediameter of the Ca-Oligochitosan-Pectin microspheres was

decreased by 34.8% within 2 h; in the same period of time, theaverage sizes of Ca-Pectin and Oligochitosan-Pectin microsphereswere reduced by 43.1% and 56.0% respectively. Based on thesein vitro results, the Ca-Oligochitosan-Pectin microsphere systemcould be a good carrier candidate for colonic drug delivery; whileremaining stable in the stomach and small intestine, it disinte-grates and releases the drug under colonic enzymatic degradation.These three hydrogel microsphere systems are summarized intoTable 1 based on their response to the simulated body fluids.

Conclusions

For drug delivery to reach the colon site, after oral administration,the drug carrier should be stable, remain intact with limitedswelling, and also retain the drug as it passes through the upperGI tract. The Ca-Pectin system is the most commonly usedpectin-based hydrogel drug delivery system. However, this systemcould not pass through the small intestine region, before reachingthe colon, because of its high pH and enzymatic responsiveness.To further improve this system, oligochitosan could be used as acoupled cross-linker (i.e. Ca-Oligochitosan-Pectin microspheres)

Figure 2. Typical images of different types of microspheres placed in simulated gastric and intestinal fluid at different times. (A), D) and (G): Ca-Pectin microspheres; (B), (E) and (H): Oligochitosan-Pectin microspheres; (C), (F) and (I): Ca-Oligochitosan-Pectin microspheres. Top panel: 120 minafter incubation in simulated gastric fluid, middle and bottom panel: 0 and �40 min after incubation in simulated intestinal fluid. Scale bar: 500 mm.

DOI: 10.3109/10837450.2014.965327 Novel pectin-based carriers for colonic drug delivery 3

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not only to reinforce the hydrogel network but also to improve thestability of the hydrogel system. This modification could be easilyachieved by mixing a calcium chloride solution with a premadeoligochitosan solution (e.g. 1 to 1 volume ratio); this mixture canthus be used as the gelling solution for hydrogel formation. This isadvantageous when compared to other routes such as chemicallymodifying pectin or by coating the delivery system. Furtherin vitro and in vivo studies that use this Ca-Oligochitosan-Pectinhydrogel system for colonic drug delivery should be conducted,e.g. for colon cancer treatment. In addition, the delivery systemoptimization (e.g. choice of pectin type and gelling solution),based on the drug properties, drug-carrier interactions, as well asthe desired responsiveness of the drug carrier, is necessary.

Declaration of interest

This work is supported by the Seed Money Grant from the Rader Schoolof Business and the Faculty Summer Development Grant at theMilwaukee School of Engineering.

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Figure 3. Typical images of the Ca-Oligochitosan-Pectin microspheres in pectinase solution at 0 (A), 30 (B), and 120 (C) min. The size of themicrospheres decreased with time: from 288.7 ± 14.8mm (0 min) to 225.0 ± 12.6mm (30 min) and 188.2 ± 20.3mm (120 min). Scale bar: 400 mm.

4 W. Zhang et al. Pharm Dev Technol, Early Online: 1–4

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