Preparation Of Microbial Enzyme-Responsive Solid Lipid Nanoparticles Loading Budesonide For Colonic Targeted Ulcerative Colitis Treatment

Ulcerative colitis(UC)is an inflammatory bowel disease that affects the mucosa and submucosa layers of the rectum.The latest epidemiological studies have shown that the incidence of UC in China is about 2.22 per 100,000 persons,and the incidence rate has been increasing year by year.UC is characterized by easy recurrence and incurable,making it a chronic disease that seriously affects the health of Chinese residents.The current clinical treatment aims to relieve the clinical symptoms of patients.Steroids are commonly used in the clinical treatment of UC,and studies have shown that topical budesonide(BUD)has a good effect and few side effects through rectal administration.However,rectal administration is difficult for UC patients to accept,and the limited drug exposure of rectal administration restricts the clinical application of topical therapy for BUD.Oral colon-specific drug delivery systems(OCDDSs)have been a hot topic in gastrointestinal drug delivery in recent years.This drug delivery strategy limits the release of drugs in the upper gastrointestinal tract,while the loadecd drug begins to released in the colon,reducing systemic toxicity and side effects,which can be used for topical BUD administration.Compared with the small intestine,the colon flora is more abundant and cellulase activity is higher,which can be used to construct OCDDSs for BUD based on the difference in cellulase gradients between the small intestine and colon.Solid lipid nanoparticles(SLNs)with negatively charged surfaces have high drug loading capacity and are easy to accumulate in inflamed intestinal areas.However,the early burst drug release of SLNs and poor single polysaccharide film-forming ability limit their colon-specific drug delivery ability.In this research,layer-by-layer self-assembly(LBL)technology was applied to form a polyelectrolyte complex(PEC)layer,consisting of negatively charged Sodium cellulose sulfate(NaCS)and positively charged chitosan quaternary ammonium salt(HACC),on the surface of SLNs,and OCDDSs for BUD administration was constructed,which exhibited an excellent antiinflammatory activity in a DSS-induced colitis mouse model.First,refined cotton was used as cellulose raw material to synthesize NaCS using heterogeneous reaction.The effects of sulfonation time on product yield,molecular weight of NaCS and substitution degree of NaCS were investigated.Furthermore,the hydrophobic modification of NaCS was carried out,and the effect of feeding ratio between dodecyl chloride and NaCS on the product characteristics was explored.Afterwards,SLNs were prepared by ultrasound dispersion method,and the preparation process(recrystallization process in nanoparticle preparation),ultrasound time,and parameters(solid lipid/surfactant ratio)were explored.Under the optimized conditions,the drug-loaded solid lipid nanoparticles(SLN-BUD)were spherical morphology with an average particle size of 248.5±1.2 nm,a surface potential of-39.9±0.1 mV,a drug loading capacity of 0.84%,and an encapsulation efficiency of 97.6%.Second,on the basis of the SLNs obtained above,we prepared nanoparticles coated with one,two,and three PEC layers on the surface of nanoparticles by sonication-assisted washless layer-by-layer self-assembly technology,i.e.SLN-BUD-1L,SLN-BUD-2L,and SLN-BUD-3L,respectively,and explored the effect of the number of coating layers on drug release.In the LBL process,the particle sizes of the nanoparticles were increased with the increase of the number of adsorption layers,and the surface potential was alternately inverted with the layer-by-layer adsorption of PEC,and finally stabilized at-30 mV to+20 mV.In the in vitro drug release experiment,with the increase of the number of PEC layers on the surface of nanoparticles,the drug release rate of nanoparticles decreased,and the maximum cumulative drug release also decreased.Moreover,the maximum cumulative release of PEC layers coated nanoparticles in the release medium of PBS solution containing cellulase was much higher than that of PBS without cellulase,indicating that the PEC layer composed of NaCS/HACC had the ability of enzymatic response hydrolysis.In view of the lower drug permeability of SLN-BUD-2L in a cellulase-free environment than SLN-BUD-1L and better enzyme responsiveness than SLN-BUD-3L in a cellulase environment,combining with the preparation time of self-assembly of nanoparticles layer by layer,two PEC layers coated nanoparticles(SLN-BUD-2L)were choosen for later research.Third,the physicochemical properties and drug release characteristics of SLN-BUD-2L were further evaluated.SLN-BUD-2L presented a spherical shell structure with an average particle size of 498.6 ± 14 nm and a surface potential of-28.5±0.4 mV.In the in vitro drug release experiments,75.24%of encapsulated BUD was released from SLN-BUD-2L in PBS release medium containing cellulase at 24 h,while only 29.21%was released in PBS as the control,which indicated great cellulase responsiveness for SLN-BUD-2L.Finally,the therapeutic effect of colon-targeted nanoparticles on UC was investigated,and the therapeutic ability of SLN-BUD-2L on DSS-induced acute ulcerative colitis mouse models was studied.After treatment,the disease activity index(DAI)score of the experimental mice in the SLN-BUD-2L group decreased significantly,the colon length was restored,the epithelial cell structure in the colon tissue section was intact,and the myeloperoxidase(MPO)and colonic tissue inflammatory cytokine levels decreased,indicating that SLN-BUD-2L has a strong therapeutic ability for ulcerative colitis.In summary,BUD loaded NaCS/HACC polyelectrolyte SLNs were prepared by sonicationassisted washless layer-by-layer self-assembly technology,and showed the efficient and safe colonic targeting ability for the treatment of UC.