Controllable Preparation And Structure Analysis Of Hydroxypropyl Chitosan And Its Application In Drug-loaded Microspheres

Hydroxypropylation is one of the most effective methods to improve the water solubility of chitosan,which expands its application in medicine,textile,paper making,wast-water treatment,etc.However,there has been no conclusive evidence to show the specific substitution sites in the structure of hydroxypropyl chitosan(HPCS),which bring difficulties to the synthesis design of HPCS derivatives and the quality control of the molar degree of substitution(MS).Therefore,hydroxypropyl chitosan(HPCS)samples with different MSs were prepared,as well as the amino-protected HPCS sample,in order to determine the substitution sites and the MS and acetylation degree(DA)of HPCS,and thus lay the foundation for their application in drug-loaded microspheres.The results of the study are as follows.Firstly,the sample preparation and ¹H NMR methods for HPCS were established and optimized.With the increase of DCl concentration,the HOD peak moves linearly to the low field,and when its concentration is greater than 1.91 M,the HOD peak can be moved to the position that it does not interfere with the characteristic peak of HPCS.While increasing the temperature can make the HOD peak move linearly to the high field,when moving to 60?70?,the HOD peak can be moved to the position that does not interfere with the HPCS characteristic peak.Secondly,the amino-protected and amino-unprotected HPCS samples were prepared,respectively,and the two ¹H NMR spectra were compared.The results show that 5.12 ppm(H_1-N),4.16 ppm(H₂')and 1.27 ppm(H_9-N)are related to amino substitution.Moreover,¹H NMR,¹³C NMR and ¹H-¹³C HSQC were used to analyze the structure of hydroxypropyl chitosan.With no DCl added in the HPCS sample as a control,the differences in the HPCS spectra were investigated to fully explore the substitution of hydroxypropylation site.The results show that 3.88/70.0 ppm and 3.89/76.4 ppm,3.93,3.78/60.0 and 4.05?3.30/66.2 ppm,4.13/63.0 ppm are 3,6,and 2 substitution sites,respectively.Once again,the ¹H NMR method to determine the total degree of substitution,NH₂-substitution degree,OH-substitution degree and acetylation degree were established.It was found that water presaturation is not suitable for the quantitative calculation of MS.Finally,hydroxypropyl chitosan with amino-protection was prepared and the drug-loaded microsphere with about 550 nm was obtained.The encapsulation efficiency and drug loading were 71.77%and 6.52%,respectively.The drug release within 24 h was78.85%.In summary,¹H NMR method we developed is successfully used for the structural analysis and MS determination of HPCS.The ¹H NMR method can be widely used to characterize the structure of amino-containing sugars(especially chitosan and its derivatives)and determine the degree of substitution.and chitosan microspheres were successfully obtained.