Synthesis And Properties Of Biodegradable Carboxymethyl Chitosan And Grafted Polylactide Hydrogels

1-ethyl-3-(3-dimethyl-aminopropyl)-1-carbodiimide (EDC) andN-hydroxysuccinimide (NHS) are common protein coupling agents. They can beeffectively used to couple amino and carboxyl groups to form amide bonds.Carboxymethyl chitosan (CMCS) is a natural polysaccharide with good biocompatibility,biodegradability. It is widely used in tissue engineering, drug delivery, health food, andwound dressing. CMCS has a lot of amino and carboxyl groups which can be crosslinkedto yield a chemical hydrogel by EDC/NHS catalysis. Poly(lactic acid)(PLA) is asynthetic biodegradable material. PLA can be grafted onto the main chain of CMCS byEDC/NHS catalysis, yielding a biodegradable CMCS-PLA hydrogel. Using EDC/MHScatalytic system, this work focuses on the two types of synthetic hydrogels.CMCS hydrogels were prepared by crosslinking CMCS with EDC/NHS at roomtemperature. The amount of EDC and EDC/NHS ratio revealed to be influencing factorson the reaction. Vial-inverting observation, solid-state13C nuclear magnetic resonance(NMR), differential scanning calorimetry (DSC), thermogravimetric analysis (TGA)confirmed the effective crosslinking of CMCS by using EDC/NHS. With increasing theamount of EDC/NHS, the crosslink density of CMCS gel is improved. The hydrogelsexhibit typical pH-responsive character. A minimum swelling ratio is obtained at theisoelectric point in the pH range of3to5. The swelling ratio remains almost constantwith increasing crosslink destiny of CMCS gel. Rheological tests show that as thecrosslink density increases, the elastic modulus increases. In frequency sweep, it is confirmed that as crosslink density increases, the rigidity of carboxymethyl chitosanhydrogel is enhanced. In strain sweep, the elastic modulus of the low crosslink densityCMCS hydrogels is basically constant, whereas the elastic modulus of high crosslinkdensity CMCS hydrogels exhibits a downward trend, thus showing good resistance toyield but also brittleness. Degradation of hydrogels was carried out at37℃in phosphatebuffered saline (PBS) or in PBS containing0.2mg/ml lysozyme. The hydrogels appearrather stable in PBS for10days. The initial mass loss of15to45%is assigned to thedissolution of uncrosslinked CMCS. The hydrogel with low crosslink density is degradedafter10days in the presence of lysozyme, while the hydrogel with high crosslink densityis hardly degraded. Two model drugs, bovine serum albumin (BSA) and thymopentin(TP-5) ae loaded in CMCS hydrogels. Preliminary drug release studies show that thehydrogels are promising carrier of hydrophilic drugs. BSA loaded high crosslink densityCMCS hydrogels present faster release rate as compared to low crosslink densityhydrogels. In contrast, TP-5loaded high crosslink density CMCS hydrogels presentslower release rate as compared to low crosslink density hydrogels. These findings areattributed the difference of molar masses between BSA and TP5. BSA with much highermolar mass cannot penetrate the center of highly crosslinked hydrogels.Novel carboxymethyl chitosan-polylactic acid hydrogels (CMCS-PLA gel) wereprepared by adjusting the EDC/NHS. Solid-state13C-NMR, SEM and FT-IRmeasurements confirmed PLA blocks are successfully grafted onto the CMCS mainchains. Vial-inverting observation, DSC measurements confirmed the effectivecrosslinking of carboxymethyl chitosan by using EDC/NHS. With increasing the amountof EDC/NHS, the crosslink destiny of CMCS-PLA gel can be improved. Similarly, thehydrogels exhibit typical pH-responsive behavior. A minimum swelling ratio is alsoobtained at the isoelectric point in the pH range of3to5. Compared to CMCS-Gel,CMCS-PLA gel exhibits relatively low swelling ratio. Rheological studies allowed toelucidate the visco-elactic behaviors of the hydrogels. Frequency sweep shows that as thecrosslink density increases, the rigidity of CMCS-PLA gel is enhanced. In the strainsweep, the elastic modulus of CMCS hydrogels remains basically constant. Compared tohigh EDC/NHS CMCS-Gel, CMCS-PLA gel presents higher toughness since the graftingof PLA decreases the crosslink density of hydrogels. Degradation of hydrogels wascarried out at37℃in phosphate buffered saline (PBS) or in PBS containing0.2mg/mllysozyme. The hydrogels appear rather stable in PBS for10days. The initial mass loss of15to30%is assigned to the dissolution of uncrosslinked CMCS (similar to CMCS-Gel). The hydrogel with low crosslink density is degraded after10days in the presence oflysozyme, while the hydrogel with high crosslink density is hardly degraded. A modeldrug, thymopentin (TP-5) was loaded in CMCS-PLA gel. Preliminary drug releasestudies show that the hydrogels are promising carrier of hydrophilic drugs.Cytotoxicity is of major importance for the applications biomaterials. Preliminarystudies were performed to evaluate the cytotoxicity of CMCS-Gel and CMCS-PLA gelby using MTT assay. The results confirmed that all hygrogels present non-toxicity. Themorphology of L929cells during culture exhibits good growth state, thus proving thatCMCS-Gel and CMCS-PLA gel present no toxicity. Therefore, both the CMCS-Gel andCMCS-PLA gel present outstanding biocompatibility, and are promising for applicationsin the fields of tissue engineering and controlled drug delivery.