Immobilization of coacervate microcapsules in multilayer sodium alginate beads for efficient oral anticancer drug delivery.

We have designed and evaluated coacervate microcapsules-immobilized multilayer sodium alginate beads (CMs-M-ALG-Beads) for oral drug delivery. The CMs-M-ALG-Beads were prepared by immobilization of doxorubicin hydrochloride (DOX) loaded chitosan/carboxymethyl coacervate microcapsules (DOX:CS/CMCS-CMs) in the core and layers of the multilayer sodium alginate beads. The obtained CMs-M-ALG-beads exhibited layer-by-layer structure and rough surface with many nanoscale particles. The swelling characteristic and drug release results indicated that 4-layer CMs-M-ALG-Beads possessed favorable gastric acid tolerance (the swelling rate <5%, the cumulative drug release rate <3.8%). In small intestine, the intact DOX:CS/CMCS-CMs were able to rapidly release from CMs-M-ALG-Beads with the dissolution of ALG matrix. Ex vivo intestinal mucoadhesive and permeation showed that CMs-M-ALG-Beads exhibited continued growth for P(app) values of DOX, which was 1.07-1.15 folds and 1.28-1.38 folds higher than DOX:CS:CMCS-CMs in rat jejunum and ileum, respectively, demonstrating that CMs-M-ALG-Beads were able to enhance the absorption of DOX by controlled releasing DOX:CS/CMCS-CMs and prolonging the contact time between the DOX:CS/CMCS-CMs and small intestinal mucosa.

Preparation and characterization of antibacterial, antioxidant, and biocompatible p-coumaric acid modified quaternized chitosan nanoparticles.

To fabricate multifunctional nanoparticles (NPs) based on chitosan (CS) derivative, we first prepared quaternized CS (2-hydroxypropyltrimethyl ammonium chloride CS, HTCC) via a one-step approach, then synthesized p-coumaric acid (p-CA) modified HTCC (HTCC-CA) for the first time through amide reaction, and finally fabricated a series of NPs (HTCC-CA NPs) using HTCC-CAs with different substitution degrees and sodium tripolyphosphate (TPP) by ionic gelation. Newly-prepared HTCC and HTCC-CAs were characterized by FT-IR, 1H NMR, elemental analysis (EA), full-wavelength UV scanning, silver nitrate titration, and Folin-Ciocalteu methods. DLS and TEM results demonstrated that three selected HTCC-CA NPs had moderate size (< 350 nm), good dispersion (PDI < 0.4), and positive zeta potential (11-20 mV). The HTCC-CA NPs had high antibacterial activity against six bacterial strains, and the minimum inhibitory concentration (MIC) values were almost the same as the minimum bactericidal concentration (MBC) values (250-1000 µg/mL). Also, the HTCC-CA NPs had good antioxidation (radical scavenging ratio > 65 %) and low cytotoxicity (relative cell viability >80 %) to the tested cells. Totally, HTCC-CA NPs with high antibacterial activity, great antioxidation, and low cytotoxicity might serve as new biomedical materials for promoting skin wound healing.

Carboxymethyl chitosan and carboxymethyl cellulose based self-healing hydrogel for accelerating diabetic wound healing.

In this study, a new type of biodegradable, injectable, self-healing, and low-toxic CMCSH, formed by N, O-carboxymethyl chitosan-heparin (CMCS-Hep) and carboxymethyl cellulose-aldehyde (CMC-A), was designed to deliver drug for promoting the progress of the diabetic wound healing. CMCS was modified with Hep for the first time to synthesize CMCS-Hep, and CMC-A was synthesized by the periodate oxidation method. First, SOD encapsulated in the CMCSH was applied to the diabetic wound bed to moderate the microenvironment, then rhEGF retained in the CMCSH was sustainedly released to the wound area. These results indicated that the dual-drug delivery system had the ability to improve drug availability, promote cell migration and proliferation, reduce DNA damage, shorten the inflammatory period, and accelerate the deposition of collagen fibers and the formation of blood vessels in the model with diabetic skin injury, suggesting that CMCSH as drug carriers had positive effects on diabetic wound healing.

Antibacterial porous sponge fabricated with capric acid-grafted chitosan and oxidized dextran as a novel hemostatic dressing.

This work aims to fabricate multifunctional hemostatic sponges (C-ODs). Porous C-ODs were first constructed by using capric acid-modified chitosan (CSCA) and oxidized dextrans (ODs) with different oxidation degrees. Batches of experiments showed that (i) CSCA (33.39% of grafting degree), ODs, and C-ODs (100-200 µm in pore size) were synthesized, evidenced by FT-IR, 1H NMR, elemental analysis, hydroxylamine hydrochloride titration, and SEM results; (ii) among C-ODs, C-OD2 had appropriate porosity (85.0%), swelling (20 times its dry weight), absorption, water retention, water vapor transmission, and mechanical properties; (iii) C-OD2 possessed low toxicity (relative cell viability > 86%), low hemolysis rate (0.65%), suitable tissue adhesion (4.74 kPa), and strong antibacterial efficacy (five strains); and (iv) C-OD2's dynamic blood clotting was within 30 s. In three animal injury models, C-OD2's hemostasis time and blood loss were fairly lower than commercial gelatin sponge. Totally, C-OD2 might serve as an ideal hemostatic dressing.

Novel amidinothiourea-modified chitosan microparticles for selective removal of Hg(II) in solution.

Glutaraldehyde-crosslinked chitosan microparticles (CGP) prepared via the inversed-phase emulsification were successively modified by epichlorohydrin (ECH) and amidinothiourea (AT) as novel adsorbent (CGPET) for selective removal of Hg(II) in solution. FTIR, EA, XPS, SEM-EDX, TG, DTG, and XRD results indicated that CGPET had ample -NH2 and CS, relative rough surface, mean diameter of ~40 µm, great thermal stability, and crystalline degree of 2.4%, beneficial to the uptake of Hg(II). The optimum parameters (pH 5, dosage 1 g/L, contact time 4 h, and initial concentration 150 mg/L) were acquired via batches of adsorption experiments. Adsorption behavior was well described by the Liu isothermal and pseudo-second-order kinetics models, and the maximum adsorption capacity was 322.51 mg/g, surpassing many reported adsorbents. Regeneration and coexisting-ion tests demonstrated that CGPET had outstanding reusability (Rr > 86.89% at the fifth cycle) and selectivity (Rs > 93%). Besides, its potential adsorption sites and mechanisms were proposed.

Fabrication of methyl acrylate and tetraethylenepentamine grafted magnetic chitosan microparticles for capture of Cd(II) from aqueous solutions.

MCS-MA-TEPA microparticles, with 251.22 mg g-1 of adsorption capacity for Cd(II), higher than most of the counterparts, were first fabricated by chemical coprecipitation, spray drying, and Michael addition reaction, without any cross-linker participation. These Fe3O4-nanoparticle-embedded microparticles of 5.95 µm in size, derived from modifications by methyl acrylate (MA) and tetraethylenepentamine (TEPA) on magnetic chitosan (MCS) microparticles, were of plum-pudding-like and wrinkle-like topography portrayed by TEM and SEM. Such features were beneficial to adsorbent recycling and Cd(II) capture. BET examinations illustrated 6.084 m2 g-1 of specific surface area, 0.015 mL g-1 of pore volume, and 6.536 nm of pore diameter. FTIR, VSM, XRD, TEM-SAED, TG, and DTG characterizations were indicative of successful synthesis, satisfactory magnetism, well-defined architecture, and good thermostability. Optimal adsorption parameters for Cd(II) were determined via batch experiments. Thermodynamic parameters and adsorption data fitting implied an exothermic, spontaneous, monolayer, and chemisorption process. XPS analyses confirmed a potential adsorption mechanism that N and O atoms on microparticles chelated with Cd(II) ions in solutions. Additionally, MCS-MA-TEPA-Cd(II) microparticles were magnetically separated easily and had outstanding reusability even after five-time recycling, with a slight adsorption capability loss (< 12%). Altogether, MCS-MA-TEPA microparticles might serve as a promising adsorbent for contaminated water scavenging.

Cr(VI) and Pb(II) capture on pH-responsive polyethyleneimine and chloroacetic acid functionalized chitosan microspheres.

PEI-ECH-CMCS microspheres (MPs) were first constructed via elaborately programmed procedures. Fourier transform infrared spectroscopy, conductometric titration, Brunauer-Emmett-Teller, X-ray diffraction, pH at zero point of charge (pHzpc), scanning electron microscopy, X-ray photoelectron spectroscopy, and swelling results demonstrated that chitosan-based adsorbent had ample -NH2 and -COOH, specific surface area of 29.040 m2/g, porous 3D architectures, pHzpc of 4.2, uniform spherical surfaces, narrow size distribution (19-33 µm), and pH-responsive swelling features, advantageous to Cr(VI) and Pb(II) capture. Adsorption parameters were obtained from batch experiments and pH 3 and 5 were chosen for Cr(VI) and Pb(II) capture. Pseudo-second-order kinetic and Liu isotherm models well interpreted adsorption behavior, and thermodynamic, isotherm, and kinetic studies revealed an exothermic, spontaneous, monolayer, and chemical adsorption process. Maximum adsorption capacity for Cr(VI) or Pb(II) was 331.32 or 302.56 mg/g, exceeding CS-based adsorbents reported. Excellent reusability and feasibility were evidenced by adsorption capacity loss < 12.10% and high removal efficiency for Cr(VI) (95.79%) and Pb(II) (91.40%) in synthetic effluents. Finally, potential adsorption mechanisms were proposed.

Optical diagnosis of cervical intraepithelial neoplasm (CIN) using polarization-sensitive optical coherence tomography.

We present a polarization-sensitive optical coherence tomography (PS-OCT) technique that can quantify the polarization changes (the degrees of circular polarization, DOCP) caused by the scattering changes induced by cervical intraepithelial neoplasia (CIN). The axial and lateral resolutions of our PS-OCT system are 13 microm and 15 microm, respectively. Uterine cervical conization tissue samples from 18 patients were examined, and 71 areas were imaged for in vitro studies; about 2-4 areas per sample were imaged and processed for diagnosis. The scanned areas had a size of 2 mm (axial) X 2 mm (lateral) X 4 mm (transversal). We quantified the slope of the axial decay of the DOCP signal near the cervical epithelium by a linear fitting procedure. The excised samples were then investigated by two pathologists, and their histological findings were later compared with the PS-OCT results. Our results show that the sensitivity and specificity are 94.7% and 71.2%, respectively.

Selective Adsorption toward Hg(II) and Inhibitory Effect on Bacterial Growth Occurring on Thiosemicarbazide-Functionalized Chitosan Microsphere Surface.

The work presented here aims to fabricate dual-purpose adsorbent with adsorption selectivity for Hg(II) and antibacterial activity. TSC-PGMA-MACS microspheres were first constructed via esterification of malic acid (MA) with chitosan (CS) and through successively grafting glycidyl methacrylate (GMA) and thiosemicarbazide (TSC) onto MACS microsphere surfaces. Fourier transform infrared spectroscopy, elemental analysis, energy-dispersive X-ray spectrometry, X-ray diffraction, differential scanning calorimetry, thermogravimetry, differential thermogravimetry, scanning electron microscopy, and Brunauer-Emmett-Teller results provided ample evidence that new mesoporous adsorbent, with 35.340 m2 g-1 of specific surface area and abundant -NH2 and C═S, was successfully fabricated and had loose crystalline, thermodynamically stable, and well-defined architectures, beneficial for Hg(II) adsorption and bacterial cell killing. Optimal adsorption parameters were determined via varying pH, time, concentrations, and temperatures, and pH 6.0 was chosen as an optimal pH for Hg(II) adsorption. Adsorption behavior, described well by pseudo-second-order kinetic and Langmuir isotherm models, and thermodynamic parameters implied a chemical, monolayer, endothermic, and spontaneous adsorption process, and the maximum adsorption capacity for Hg(II) was 242.7 mg g-1, higher than most of the available adsorbents. Competitive adsorption exhibited excellent adsorption selectivity for Hg(II) in binary-metal solutions. Besides, TSC-PGMA-MACS microspheres had outstanding reusability even after five times recycling, with adsorption capability loss <14%. Several potential adsorption sites and bonding modes were proposed. Notably, TSC-PGMA-MACS microspheres before and after adsorption were of high antibacterial activity against Escherichia coli and Staphylococcus aureus (MICs, 2 and 0.25 mg mL-1), superior to CS powders, and possible antibacterial mechanisms were also summarized. Altogether, dual-purpose TSC-PGMA-MACS microspheres might be promising adsorbent for contaminated water scavenging.

Preparation, characterization, and evaluation of 3,6-O-N-acetylethylenediamine modified chitosan as potential antimicrobial wound dressing material.

This work aims to prepare 3,6-O-N-acetylethylenediamine modified chitosan (AEDMCS) and evaluate its potential use as an antimicrobial wound dressing material. UV, FTIR, and 1H NMR results demonstrated N-acetylethylenediamine groups were successfully grafted to C3OH and C6OH on polysaccharide skeletons. TGA, XRD, and solubility tests indicated that as compared with chitosan, AEDMCS had diminished thermostability, decreased crystallinity, and greatly improved solubility. AEDMCS, with degrees of deacetylation and substitution being respectively 90.3% and 0.72, exhibited higher antibacterial activity than chitosan against six bacteria generally causing wound infections. Meanwhile, AEDMCS had permissible hemolysis and cytotoxicity and low BSA adsorption even at a AEDMCS concentration of 25mg/mL. Acute toxicity tests showed AEDMCS was nontoxic. Moreover, the wound healing property was preliminarily evaluated, illustrating that AEDMCS enhanced wound healing rates as expected and had no significant differences as compared with chitosan. These results suggested AEDMCS might be a potential material used as antibacterial wound dressings.

Characterization and antibacterial mechanism of poly(aminoethyl) modified chitin synthesized via a facile one-step pathway.

This work aims to synthesize poly(aminoethyl) modified chitin (PAEMC) and ascertain its antibacterial activity and mechanism. FTIR and 1H NMR results proved aminoethyl moieties were grafted to C6OH and C3OH on chitin backbone in the form of polymerization. XRD and TG/DTG analyses manifested its well-defined crystallinity and thermostability. PAEMC, with average molecular weight (MW) of 851.0 kDa, degree of deacetylation (DD) of 27.95%, and degree of substitution (DS) of 1.77, had good solubility in aqueous solutions over the pH range of 3-12, and also possessed high antimicrobial activity against Staphylococcus epidermidis, Staphylococcus aureus, Pseudomonas aeruginosa, Escherichia coli, Bacillus proteus, and Klebsiella pneumoniae, commonly causing chronic wound infections. Nucleic acid release, protein leakage, increased inner membrane permeability, and decreased cell surface hydrophobicity implied bacterial cytomembranes were substantially compromised in the presence of PAEMC. Microscopically, PAEMC visually perturbed bacteria, illustrating deformed and even collapsed morphologies. Overall, PAEMC possessed good solubility, effectively destroyed bacteria via a membrane damage mechanism, and might serve as an antibacterial agent for treatments of chronic wound infections.

Uptake of Pb(II) and Cd(II) on Chitosan Microsphere Surface Successively Grafted by Methyl Acrylate and Diethylenetriamine.

A novel adsorbent, CS-MA-DETA microspheres, for uptake of heavy metal ions from aqueous solutions was first fabricated via two-step grafting methyl acrylate (MA) and diethylenetriamine (DETA) onto chitosan (CS) microsphere surface in the absence of cross-linkers. CS-MA-DETA microspheres of 3.04 µm in mean diameter were of uniformly wrinkle-like topography sketched out by SEM, whose surface after decoration by MA and DETA was stable and beneficial to metal ion capture. Its chemical composition, microstructure, and thermal property were characterized by elemental analysis, FTIR, XRD, BET, and TGA techniques, and the achieved quantitative results mainly included C/N ratio (4.76), crystallinity (31.20%, 19.75% of CS), specific surface area (27.806 m2 g-1), pore diameter (3.452 nm), and mass loss at the first stage (3%, around 10% of CS), which indicated a successful synthesis, well-defined structure, and good thermostability. Adsorption tests of CS-MA-DETA microspheres were performed in Pb(II) and/or Cd(II) solution(s) at various pH values, contact time, and initial concentrations, exhibiting an excellent adsorption capability. Its maximum adsorption capacity calculated by Langmuir model was 239.2 mg Pb(II)/g, or 201.6 mg Cd(II)/g, which was higher than those of most available CS-based adsorbents. Furthermore, several adsorption kinetic and isotherm models were employed to investigate its uptake behavior, implying that it was mainly a monolayer adsorption and chemisorption process. Five-cycle reusability tests demonstrated CS-MA-DETA microspheres could be repeatedly used without significant capacity loss (<10%). Additionally, several potential bonding modes and adsorption sites for both metal ions were also proposed. Overall, CS-MA-DETA microspheres with outstanding adsorption performance toward Pb(II) and/or Cd(II) might serve as a new absorbent for wastewater purification.

Preparation and characterization of poly(maleic acid)-grafted cross-linked chitosan microspheres for Cd(II) adsorption.

A novel adsorbent, composed of poly(maleic acid)-grafted cross-linked chitosan microspheres (PMACCMs), was prepared via cross-linking with glutaraldehyde and modification by grafting maleic acid. FTIR, zeta potential, elemental analysis, 13C NMR, DTG, laser particle size analysis, SEM, and BET methods were applied to characterize PMACCMs, exhibiting a successful fabrication, good thermostability, and well-defined surface microstructure beneficial to Cd(II) adsorption. The effects of pH, contact time, and initial concentration on Cd(II) adsorption were also investigated, and the maximum adsorption capacity was 39.2mgg-1, indicating a great improvement as compared with that (14.5mgg-1) of cross-linked chitosan microspheres. The experimental data were well fitted with pseudo-second-order kinetic and Langmuir isotherm models. Five-cycle reusability tests demonstrated PMACCMs could be repeatedly used with a small adsorption capacity loss (<15%). Additionally, the adsorption mechanism was proposed. All the results confirmed that PMACCMs, which presented outstanding adsorption capability and reusability, could be a good candidate for wastewater purification.

Itaconic acid grafted carboxymethyl chitosan and its nanoparticles: Preparation, characterization and evaluation.

This work aims to synthesize a novel itaconic acid (IA) grafted carboxymethyl chitosan (PICMCS), and further fabricate its nanoparticles for potential biomedical applications. First, PICMCS was prepared via free-radical polymerization of IA monomer, in the presence of ammonium persulfate as an initiator and nitrogen as a protector. Its chemical structure was confirmed by FTIR and 1H NMR. The IA substitution degree calculated by elemental analysis data was 1.85, implying that IA was successfully grafted to carboxymethyl chitosan (CMCS). XRD and TGA patterns illustrated its well-defined crystallinity and thermostability. Second, PICMCS nanoparticles were fabricated by electrostatic attraction between carboxyl and amino groups in the absence of any additional agent, which were of obvious core-shell structures with an average particle size of 144nm and a polydispersity index of 0.11. PICMCS nanoparticles exhibited excellent physical stability after storage at 25°C for 30days, without any aggregation. PICMCS nanoparticles with high negative surface charge also indicated the good stability, especially in neutral or alkaline media. Additionally, the cytotoxicity experiments showed that either PICMCS or its nanoparticles had better cytocompatibility toward L929 cells than CMCS. These findings above suggested that PICMCS was a kind of promising material for preparing nanoparticles used in biomedical field.

Synthesis, characterization, and evaluation of poly(aminoethyl) modified chitosan and its hydrogel used as antibacterial wound dressing.

This study aims to develop new antibacterial hydrogel wound dressings composed of poly(aminoethyl) modified chitosan (PAEMCS). FTIR, 1H NMR, and elemental analysis demonstrated that PAEMCS was successfully synthesized via grafting poly(aminoethyl) groups onto hydroxyl groups on chitin first, and removing acetyl groups from the grafted polymer afterward. XRD and TGA implied its well-defined crystallinity and thermostability. Furthermore, a series of hydrogels were fabricated under the participation of dipotassium hydrogen phosphate (DHP). The gelation tests suggested that the higher concentration of PAEMCS or DHP was beneficial to the formation of hydrogels. The pH values of hydrogels at 37°C were all in the range of 7.12-7.50. The rheological tests indicated that PAEMCS-based hydrogels were of lower DHP addition and higher elasticity than CS-based hydrogels to achieve the same gelation temperature under the same polymer's concentration. Additionally, the swelling, anti-bacteria, and cytotoxicity experiments showed that PAEMCS-based hydrogels possessed excellent hygroscopicity, high antibacterial activity against E. coli, S. aureus, or S. epidermidis, and good cytocompatibility toward L929 cells or HUVECs, respectively. All the results implied that PAEMCS-based hydrogels not only maintained inherent multiple properties of chitosan but also possessed excellent antibacterial activity, and might be promising antibacterial hydrogel dressings used in wound therapy.

Fabrication and evaluation of thermosensitive chitosan/collagen/α, ß-glycerophosphate hydrogels for tissue regeneration.

Thermosensitive hydrogels whose physiological properties are similar to extracellular matrix have been extensively used for tissue regeneration. Polysaccharides and proteins, as biocompatible substrates similar to bio-macromolecules that could be recognized by human body, are two preferred polymers for fabrication of such hydrogels. A series of novel thermosensitive hydrogels (CS-ASC-HGs) containing chitosan (CS) and acid-soluble collagen (ASC) were thus prepared, in the presence of α, ß-glycerophosphate, to mimic extracellular microenvironment for tissue regeneration. Rheological measurements demonstrated excellent thermosensitivity. FT-IR and SEM indicated CS-ASC-HGs possessed 3D porous architectures with fibrous ASC, and the molecular structure of ASC was well-maintained in hydrogels. Hemolysis, acute toxicity, and cytotoxicity tests suggested CS-ASC-HGs were of good biocompatibility. CS-ASC-HGs were able to support the survival and proliferation of L929 cells encapsulated in them. Moreover, CS-ASC-HGs had better pH stability and biocompatibility than pure CS hydrogel. These results suggested that CS-ASC-HGs could serve as promising scaffolds for tissue regeneration.

Heterotrophic nitrogen removal by a newly-isolated alkalitolerant microorganism, Serratia marcescens W5.

A new microbe, Serratia marcescens W5 was successfully isolated. Its feasibility in purification of excessively nitrogen-containing wastewater was evaluated using inorganic nitrogen media. Single factor tests showed that W5 exhibited high ammonium removal rates (above 80%) under different culture conditions (pH 7-10, C/N ratios of 6-20, 15-35°C, 0-2.5% of salinity, respectively). Besides various organic carbon sources, W5 was able to utilize calcium carbonate with 28.05% of ammonium removed. Further experiments indicated that W5 was capable of resisting high-strength ammonium (1200mg/L) with the maximum removal rate of 514.13mgL(-1)d(-1). The nitrogen removal pathway of W5 was also tested, showing that both nitrite and nitrate were efficiently removed only in the presence of ammonium, with hydroxylamine as intermediate, which was different from the conventional nitrogen removal pathway. All the results verified that W5 was a good candidate for the purification of excessively nitrogenous wastewater.

3,6-O-[N-(2-Aminoethyl)-acetamide-yl]-chitosan exerts antibacterial activity by a membrane damage mechanism.

A novel chitosan derivative, 3,6-O-[N-(2-aminoethyl)-acetamide-yl]-chitosan (AACS), was successfully prepared to improve water solubility and antibacterial activity of chitosan. AACS had good antibacterial activity, with minimum inhibitory concentrations of 0.25mg/mL, against Escherichia coli and Staphylococcus aureus. Cell membrane integrity, electric conductivity and NPN uptake tests showed that AACS caused quickly increasing the release of intracellular nucleic acids, the uptake of NPN, and the electric conductivity by damaging membrane integrity. On the other hand, hydrophobicity, cell viability and SDS-PAGE experiments indicated that AACS was able to reduce the surface hydrophobicity, the cell viability and the intracellular proteins through increasing membrane permeability. SEM observation further confirmed that AACS could kill bacteria via disrupting their membranes. All results above verified that AACS mainly exerted antibacterial activity by a membrane damage mechanism, and it was expected to be a new food preservative.

Transport mechanism of doxorubicin loaded chitosan based nanogels across intestinal epithelium.

Chitosan/carboxymethyl chitosan nanogels (CS/CMCS-NGs) could enhance the oral bioavailability of doxorubicin hydrochloride (DOX). To identify the mechanisms that support this recent observation, different transport pathways of CS/CMCS-NGs through the small intestine were studied in this work. Transcellular mechanisms were investigated in the presence of different inhibitors of protein-mediated endocytosis. A reduction of 52.32±18% of drug transport was found when clathrin-mediated endocytosis was inhibited, which demonstrated that clathrin-mediated endocytosis played an important role in the transcellular transport of DOX:CS/CMCS-NGs. The paracellular transport results showed that CMCS in NGs could produce a transient and reversible enhancement of paracellular permeability by depriving Ca(2+) from adherens junctions, whose efficacy as an absorption enhancer was about 1.7-3.3 folds higher than CS in NGs in GI tract. Finally, in vivo experiment showed that the transport capacity of DOX:CS/CMCS-NGs was significantly inhibited by extra added Ca(2+), which confirmed that the higher capacity to binding Ca(2+) of CS/CMCS-NGs was beneficial for transport of DOX.