The Preparation And Characteristics Of Genipin Crosslinked Soy Protein/chitosan-based Chili Alkaline Hydrogel And Transdermal Absorption

A novel hydrogel system composed of soybean protein and chitosan blended with naturally non-toxic genipin was developed for controlling capsaicin delivery. The impact of various factors on the preparation of the hydrogel was examined. Based on single factor experiment, the response surface methodology was used to optimize the hydrogel preparation process. Genipin soybean protein chitosan hydrogel was prepared under optimal conditions as a controlled-release carrier of capsaicin. The skin irritation studies of rattus norvegicus in vitro transdermal study investigated hydrogel toxicity, and capsaicin release. Additionally, the release profile of a model drug (capsaicin) from test hydrogel was investigated in pH7.4buffer. The structures of soybean protein hydrogel, chitosan hydrogel, and hydrogel were characterized by infrared spectrophotometer. These results showed obvious crosslinking systems for soybean protein and chitosan by genipin in the hydrogels. The hydrogel swelled in the pH7.4buffer. The controlled release of capsicine was realized in26h. Therefore, the genipin-corsslinked hydrogel could be a suitable carrier for site-specific drug delivery in the skin. The main contents were as follows:1、Set up HPLC and LC-MS/MS method which was for determining the capsaicin. HPLC method:Chromatographic separation was carried out by a Kromasil-C18(5μm,4.6mm x250mm) reversed-phase column; the mobile phase is methanol-water (91:9, v/v). Capsaicin was quantified by a UVD following RP-HPLC separation at the wavelength of281nm. The flow rate was1ml/min, the injection volume was10μand the column temperature was maintained at25℃. The standard curve of capsaicin and regression equation was drawn.LC-MS/MS method:Chromatographic separation was carried out by a Kromasil-C18(5μm,4.6mm×250mm) reversed-phase column; the mobile phase is methanol-water (9:1, v/v). Scanning mode used cation and reaction monitoring mode (MRM); the voltage of ion source spray was-4500V; the temperature of ion source atomization was300℃; atomization air was82.7kPa; the molecular weight of capsaicin debris was137.3; the DP voltage was+70V; the CE voltage was+66V; the CXP voltage was+10V; the peak time was2.46min; Standard curve equation was y=27.484x+27800(R2=0.9995), The linear range is50ng～0.1mg. Then inter-day precision, intra-day precision, recovery and stability were measured and met the test requirements.2、Genipin crosslinked soy protein/chitosan hydrogel was selected as a controlled-release carrier of capsaicin by the comparison of chitosan hydrogel, soy protein hydrogel and hydrogel. The effect of single factors on the preparation of hydrogel was studied. Base on the Single-factor experiments, a three-factor-three-level experiment design had been developed by Central experiment design method. The optimized conditions were as follows:1%soybean protein solution with1.5%chitosan solution volume ratio was2:1, cross linker concentration was70mmol/mL, and cross linking time was5min. The condition can be used to prepare for hydrogel.3、The surface morphology and structure of chitosan-based hydrogels were characterized by infrared spectrophotometry and scanning electron microscopy. The results showed that there was amide bond or ester bond in the hydrogels. The results showed obvious crosslinking systems for soybean protein and chitosan by genipin in the hydrogels. But there were some large voids without rules on the surface. Then the drug loading, release, tack, the degradation rate and other parameters were tested, and the results were213μg/g,26.1h,0.7kg,90%respectively. The results of stability test and skin irritation test shown the chitosan-based hydrogels was stable and non-toxic.4、In vitro percutaneous studies were performed by Genipin soybean protein chitosan hydrogel that was prepared under optimal conditions. Hydrogels and capsaicin ointment Qn-t curve equation obtained by transdermal experiment were y=6.8828x+3.3191(R2=0.9321), y=74.375x+20.123(R-=0.9258). cumulative permeation amount Qn, steady-state permeation rate Js, residence time Tlag of hydrogels and capsaicin ointment were141.15μg-cm-2and153.06μg·cm-2,5.8812μg·cm-2·h-1and51.02μg·cm-2·h-1,0.48h and0.27h. As cumulative permeation amount of the two consistent with the circumstances, the permeation rate of capsaicin ointment was greater than hydrogel. So transdermal effect of hydrogel were longer working time, longer residence time and more pronounced pharmacodynamic.