Nanoparticles Prepared From N,N-Dimethyl-N-Octyl Chitosan as the Novel Approach for Oral Delivery of Insulin: Preparation, Statistical Optimization and Characterization

Document Type : Research article

Authors

1 Department of Pharmaceutics, School of Pharmacy, Hamadan University of Medical Sciences, Hamadan, Iran.

2 Department of Pharmaceutics, Faculty of Pharmacy, Tehran University of Medical Sciences, Tehran, Iran.

Abstract

In this study, N,N-Dimethyl-N-Octyl chitosan was synthesized. Nanoparticles containing insulin were prepared using PEC method and were statistically optimized using the Box-Behnken response surface methodology. The independent factors were considered to be the insulin concentration, concentration and pH of the polymer solution, while the dependent factors were characterized as the size, zeta potential, PdI and entrapment efficiency. The optimized nanoparticles were morphologically studied using SEM. The cytotoxicity of the nanoparticles on the Caco-2 cell culture was studied using the MTT cytotoxicity assay method, while the permeation of the insulin nanoparticles across the Caco-2 cell monolayer was also determined. The optimized nanoparticles posed appropriate physicochemical properties. The SEM morphological studies showed spherical to sub-spherical nanoparticles with no sign of aggregation. The in-vitro release study showed that 95.5 ± 1.40% of the loaded insulin was released in 400 min. The permeability studies revealed significant enhancement in the insulin permeability using nanoparticles prepared from octyl chitosan at 240 min (11.3 ± 0.78%). The obtained data revealed that insulin nanoparticles prepared from N,N-Dimethyl-N-Octyl chitosan can be considered as the good candidate for oral delivery of insulin compared to nanoparticles prepared from N,N,N-trimethyl chitosan.

Keywords

Main Subjects


Patel V. A short review on the association between
depression and diabetes.
Int. J. Med. Sci. Public Health
(2014) 3: 3-5.

Li CL and Deng YJ. Oil-based formulations for oral

delivery of insulin.
J. Pharm. Pharmacol . (2004) 56:
1101-7.

Pillai O and Panchagnula R. Insulin therapies - past,

present and future.
Drug Discov. Today (2001) 6:
1056-61.

Ubaidulla U, Khar RK, Ahmed FJ and Panda AK.

Development and
in-vivo evaluation of insulin-loaded
chitosan phthalate microspheres for oral delivery.
J.
Pharm. Pharmacol.
(2007) 59: 1345-51.
Bayat A, Larijani B, Ahmadian S, Junginger HE and

Rafiee-Tehrani M. Preparation and characterization of

(1)

(2)

(3)

(4)

(5)
 
Shamsa ES et al. / IJPR (2018), 17 (2): 442-459458
insulin nanoparticles using chitosan and its quaternized

derivatives.
Nanomedicine (2008) 4: 115-20.
Sadeghi AM, Dorkoosh FA, Avadi MR, Weinhold M,

Bayat A, Delie F, Gurny R, Larijani B, Rafiee-Tehrani

M and Junginger HE. Permeation enhancer effect

of chitosan and chitosan derivatives: Comparison of

formulations as soluble polymers and nanoparticulate

systems on insulin absorption in Caco-2 cells.
Eur. J.
Pharm. Biopharm
. (2008) 70: 270-8.
Radwant MA and Aboul-Enein HY. The effect of oral

absorption enhancers on the
in-vivo performance of
insulin-loaded poly(ethylcyanoacrylate) nanospheres

in diabetic rats.
J. Microencapsul. (2002) 19: 225-35.
Mesiha M, Plakogiannis F and Vejosoth S. Enhanced

oral absorption of insulin from desolvated fatty acid-

sodium glycocholate emulsions.
Int. J. Pharm. (1994)
111: 213-6.

Morishita I, Morishita M, Takayama K, Machida

Y and Nagai T. Hypoglycemic effect of novel oral

microspheres of insulin with protease inhibitor in

normal and diabetic rats.
Int. J. Pharm . (1992) 78:
9-16.

Bernkop-Schnürch A. The use of inhibitory agents

to overcome the enzymatic barrier to perorally

administered therapeutic peptides and proteins.
J.
Control. Release
(1998) 52: 1-16.
Takeuchi H, Yamamoto H, Niwa T, Hino T and

Kawashima Y. Enteral absorption of insulin in rats from

mucoadhesive chitosan-coated liposomes.
Pharm. Res.
(1996) 13: 896-901.

Asada H, Douen T, Waki M, Adachi S, Fujita T and

Yamamoto A. Absorption characteristics of chemically

modified-insulin derivatives with various fatty acids in

the small and large intestine.
J. Pharm. Sci. (1995) 84:
682-7.

Chen Y, Siddalingappa B, Chan PH and Benson

HA. Development of a chitosan-based nanoparticle

formulation for delivery of a hydrophilic hexapeptide,

dalargin.
Biopolymers (2008) 90: 663-70.
Ito M, Ban A and Ishihara M. Anti-ulcer effects of

chitin and chitosan, healthy foods, in rats.
Jpn. J.
Pharmacol
. (2000) 82: 218-25.
Sieval AB, Thanoual M, Kotzkb AF, Verhoefa JC,

Brussee J and Junginger HE. Preparation and NMR

characterization of highly substituted
N-trimethyl
chitosan chloride.
Carbohydr. Polym. (1998) 36: 157-
65.

Thanou M, Verhoef JC and Junginger HE. Oral

drug absorption enhancement by chitosan and its

derivatives.
Adv. Drug Deliv. Rev. (2001) 52: 117-26.
Mao S, Bakowsky U, Jintapattanakit A and Kissel

T. Self-assembled polyelectrolyte nanocomplexes

between chitosan derivatives and insulin.
J. Pharm.
Sci
. (2006) 95: 1035-48.
Mao HQ, Roy K, Troung-Le VL, Janes KA, Lin

KY, Wang Y, August JT and Leong KW. Chitosan-

DNA nanoparticles as gene carriers: Synthesis,

characterization and transfection efficiency.
J. Control.
Release
(2001) 70: 399-421.
Mahjub R, Dorkoosh F, Amini M, Khoshayand M and

Rafiee-Tehrani M. Preparation, statistical optimization,

and
in-vitro characterization of insulin nanoparticles
composed of quaternized aromatic derivatives of

chitosan.
AAPS PharmSciTech. (2011) 12: 1407-19.
Kadam RS, Bourne DW and Kompella UB.

Nano-advantage in enhanced drug delivery with

biodegradable nanoparticles: Contribution of reduced

clearance.
Drug Metab. Dispos. (2012) 40: 1380-8.
Roller S and Covill N. The antifungal properties of

chitosan in laboratory media and apple juice.
Int. J.
Food Microbiol.
(1999) 47: 67-77.
Rhoades J and Roller S. Antimicrobial actions

of degraded and native chitosan against spoilage

organisms in laboratory media and foods.
Appl.
Environ. Microbiol.
(2000) 66: 80-6.
Julienne MC, Alonso MJ, GÓMez Amoza JL and

Benoit JP. Preparation of poly(D,L-Lactide/Glycolide)

nanoparticles of controlled particle size distribution:

Application of experimental designs.
Drug Dev. Ind.
Pharm
. (1992) 18: 1063-77.
Ibrahim H, Bindschaedler C, Doelker E, Buri P and

Gurny R. Aqueous nanodispersions prepared by a

salting-out process.
Int. J. Pharm. (1992) 87: 239-46.
Leorux JC, Allemann E, Doelker E and Gurny R.

New approach for the preparation of nanoparticles by

an emulsification-diffusion method.
Eur. J. Pharm.
Biopharm
. (1995) 41: 14-8.
Elsayed A, Al-Remawi M, Qinna N, Farouk A, Al-

Sou’od K and Badwan A. Chitosan–sodium lauryl

sulfate nanoparticles as a carrier system for the
in-vivo
delivery of oral insulin.
AAPS PharmSciTech. (2011)
12: 958-64.

Rojanarata T, Petchsangsai M, Opanasopit P,

Ngawhirunpat T, Ruktanonchai U, Sajomsang W

and Tantayanon S. Methylated N-(4-N ,N -dimet

hyl aminob en zyl) chitosan for novel effective gene

carriers.
Eur. J. Pharm. Biopharm. (2008) 70: 207-14.
Yoo HS, Lee JE, Chung H, Kwon IC and Jeong SY. Self-

assembled nanoparticles containing hydrophobically

modified glycol chitosan for gene delivery.
J. Control.
Release
(2005) 103: 235-43.
Sandri G, Rossi S, Bonferoni MC, Ferrari F, Zambito

Y, Di Colo G and Caramella C. Buccal penetration

enhancement properties of N-trimethyl chitosan:

Influence of quaternization degree on absorption of a

high molecular weight molecule.
Int. J. Pharm. (2005)
297: 146-55.

Sandri G, Bonferoni MC, Rossi S, Ferrari F, Gibin S,

Zambito Y, Di Colo G and Caramella C. Nanoparticles

based on N-trimethylchitosan: Evaluation of

absorption properties using
in-vitro (Caco-2 cells) and
ex-vivo
(excised rat jejunum) models. Eur. J. Pharm.
Biopharm
. (2007) 65: 68-77.
Helgason T, Awad TS, Kristbergsson K, McClements

DJ and Weiss J. Effect of surfactant surface coverage

on formation of solid lipid nanoparticles (SLN).
J.
Colloid Interface Sci.
(2009) 334: 75-81.
Mohanraj VJ and Chen Y. Nanoparticles – A Review.

(6)

(7)

(8)

(9)

(10)

(11)

(12)

(13)

(14)

(15)

(16)

(17)

(18)

(19)

(20)

(21)

(22)

(23)

(24)

(25)

(26)

(27)

(28)

(29)

(30)

(31)

(32)
 
New chitosan derivative for insulin oral delivery459
Trop. J. Pharm. Res
. (2006) 5: 561-73.
Wada R, Hyon SH and Ikada Y. Lactic acid oligomer

microspheres containing hydrophilic drugs.
J. Pharm.
Sci.
(1990) 79: 919-24.
Fresta M, Puglisi G, Giammona G, Cavallaro G,

Micali N and Furneri PM. Pefloxacine mesilate-

and ofloxacin-loaded polyethylcyanoacrylate

nanoparticles: Characterization of the colloidal drug

carrier formulation.
J. Pharm. Sci. (1995) 84: 895-902.
Uchida T, Yagi A, Oda Y, Nakada Y and Goto S.

Instability of bovine insulin in poly(lactide-co-

glycolide) (PLGA) microspheres.
Chem. Pharm. Bull.
(1996) 44: 235-6.

Werle M, Takeuchi H and Bernkop-Schnurch A.

Modified chitosans for oral drug delivery.
J. Pharm.
Sci
. (2009) 98: 1643-56.
Di Colo G, Zambito Y and Zaino C. Polymeric

enhancers of mucosal epithelia permeability: Synthesis,

transepithelial penetration-enhancing properties,

mechanism of action, safety issues.
J. Pharm. Sci .
(2008) 97: 1652-80.

Reis CP, Veiga FJ, Ribeiro AJ, Neufeld RJ and Damge

C. Nanoparticulate biopolymers deliver insulin orally

eliciting pharmacological response.
J. Pharm. Sci .
(2008) 97: 5290-305.

(33)

(34)

(35)

(36)

(37)

(38)

Sadeghi AM, Dorkoosh FA, Avadi MR, Saadat P,

Rafiee-Tehrani M and Junginger HE. Preparation,

characterization and antibacterial activities of chitosan,

N-trimethyl chitosan (TMC) and N-diethylmethyl

chitosan (DEMC) nanoparticles loaded with insulin

using both the ionotropic gelation and polyelectrolyte

complexation methods.
Int. J. Pharm. (2008) 355: 299-
306.

Mahobia S, Bajpai J and Bajpai AK. An
in-vitro
investigation of swelling controlled delivery of insulin

from egg albumin nanocarrier.
Iran. J. Pharm. Res .
(2016) 15: 695-711.

Mahjub R, Heidari Shayesteh T, Radmehr M, Vafei SY,

Amini M, Dinarvand R and Dorkoosh FA. Preparation

and optimization of N-trimethyl-O-carboxymethyl

chitosan nanoparticles for delivery of low-molecular-

weight heparin.
Pharm. Dev. Technol. (2016) 21: 14-
25.

Mahjub R, Radmehr M, Dorkoosh FA, Ostad SN and

Rafiee-Tehrani M. Lyophilized insulin nanoparticles

prepared from quaternized N-aryl derivatives of

chitosan as a new strategy for oral delivery of insulin:

in-vitro
, ex-vivo and in-vivo characterizations. Drug
Dev. Ind. Pharm
. (2014) 40: 1645-59