Extraction And Modification Of Chitin From The Hericium Erinaceus Residue And The Effect Mechanism Of Multi-scale Structural Changes On Gel Properties

A great amount of residues are produced after the extraction of polysaccharide from Hericium erinaceus,but there is no relevant report on the high-value utilization of such Hericium erinaceus residue(HER).In addition to a small amount of protein,fat and minerals,the main component of the HER is a special insoluble polysaccharide—chitin.As the second abundant biopolymer in nature after cellulose,chitin is renewable,degradable,safe and non-toxic,showing great application value in the fields of food,materials,environmental protection,energy and biomedicine.Therefore,the high-value utilization of chitin from HER is necessary.In this study,chitin was extracted from HER,and its basic structure was characterized.Subsequently,the chitin from the HER was physically and chemically modified to reveal the effect mechanisms of its multi-scale structural changes on gel properties.Finally,the smart hydrogels based on the Hericium erinaceus residue chitin were constructed and applied to drug release.The main findings are as follows:The chitin was extracted from HER by a multi-step extraction procedure.It was found that the concentration of NaOH,reaction temperature and reaction time all have an effect on the yield,purity,molecular weight and degree of acetylation of chitin.When the NaOH concentration is 2%,the reaction temperature is 85^oC and the reaction time is 3 h,the extracted chitin shows the most desirable performance parameters(yield 16.30%,purity97.88%,molecular weight 2.01×10⁵ g/mol,degree of acetylation 77.67%).In addition,the results also found that HER chitin belongs to?-chitin and does not contain glucan components.The chitin extracted from the Hericium erinaceus residue was nano-sized by TEMPO oxidation method,so that it can disperse stably in a heterogeneous system.The average length and width of the nano chitin prepared in this study were detected as 336.6 nm and 6.4 nm,respectively,the Zeta-potential was determined as-27.8 m V under neutral conditions,the basic structure before and after nanoization did not change significantly,and its dispersion was affected by p H and ionic strength.On the other hand,the acid-induced gas phase coagulation method was used to convert the nano-chitin dispersion into a gel successfully in a heterogeneous system without phase transition.NaOH/Urea(11%/4%)solvent system was used to dissolve the HER chitin,so that it can exist stably in a homogeneous system.At the higher than 0.1 mg/m L of chitin concentration,the chitin molecules were found in the form of aggregates,and at the concentration of HER chitin lower than 0.1 mg/m L,the chitin molecules were found as single molecules.The dissolved chitin solution showed favorable stability,but chitin molecules were subject to undergo a phase transition and re-precipitate in the form of suspended particles with heating or reducing the concentration of OH^-ions.In addition,the HER chitin solution could be converted into a hydrogel through a cross-linking reaction under alkaline conditions,forming gelation of HER chitin in a homogeneous system with phase transition.It is worth noting that the minimum concentration of the chitin that formed hydrogels was 5%.The formed hydrogel showed soft physical structure and concentration-dependent,HER chitin solution at higher concentration formed a hydrogel with preferable plasticity.High-field ultrasound treatment were used to physically degrade the HER chitin,and chitin molecules with different molecular weight were prepared by changing the ultrasound time.The influence of chitin molecular weight on gel properties was investigated,and the mechanism of its effect on the performance of the chitin gel was revealed via change of the molecular scale.With ultrasonic treatment time increased from 0 to 15 min,the molecular weight of chitin decreased from 2.01×10⁵ g/mol to 7.65×10⁴ g/mol.Correspondingly,the gel properties of the formed chitin hydrogels also showed significant differences.The longer the ultrasound time,the weaker the plasticity and gel strength of the chitin hydrogel,and greater the swelling degree.Small-angle X-ray scattering result showed that the essence of HER chitin hydrogel formation is by cross-linking its smallest solid domain,which is considered as the basic structural unit,and the size of the basic structural unit in the formed hydrogels was not affected by ultrasonic treatment.However,the determination of gel yield showed that the yield of chitin hydrogels gradually decreased with the increasing ultrasound time.Therefore,the mechanism of the molecular scale of HER chitin molecule on its gel formation can be summarized as:the reduction of HER chitin molecular scale reduces the amount of effective HER chitin that can participate in the cross-linking reaction,thereby reducing the gel yield of chitin hydrogel,and the crosslinking density of the formed chitin hydrogel network structure was also reduced correspondingly,resulting in a weaker gel strength but an enhanced swelling degree of chitin hydrogel.Homogeneous carboxymethylation was used to replace the molecular side chain groups of HER chitin.HER carboxymethyl chitins with different degree of substitution were prepared by changing the amount of sodium chloroacetate.The effect of the degree of substitution of HER carboxymethyl chitin on the gel performance was investigated,and the mechanism of its effect on the performance of the chitin gel was observed based on the change of the functional group scale.As the degree of substitution of carboxymethyl chitin increased from 0 to 0.038,the equilibrium swelling degree of the corresponding hydrogel increased from 29.8 g/g to40.2 g/g.In addition,the introduction of carboxymethyl functional group also endowed HER chitin hydrogels with excellent p H swelling response characteristics.Under low p H environments,the prepared HER carboxymethyl chitin hydrogels exhibit extremely low swelling degree.As p H increased,the swelling degree of the hydrogels gradually increased.Zeta-potential,low-field nuclear magnetic resonance,contact angle and molecular dynamics simulation results showed that the introduction of carboxymethyl functional group increased the swelling degree of the prepared HER chitin hydrogels due to the enhanced negative charge,water mobility,surface hydrophilicity and the formation of hydrogen bonds with water.In-situ co-precipitation technique was used to synthesize Fe₃O₄ nanoparticles inside the prepared HER carboxymethyl chitin hydrogel.A series of p H/magnetic sensitive composite hydrogels were prepared by changing the concentration of precursor Fe²⁺/Fe³⁺,the effect of Fe₃O₄ nanoparticles on the performances of carboxymethyl chitin hydrogels was investigated.The results showed that Fe₃O₄ nanoparticles exhibited excellent compatibility with the HER carboxymethyl chitin hydrogel network structure.Fe₃O₄ nanoparticles were observed uniformly distributed in the three-dimensional network structure of the prepared HER carboxymethyl chitin hydrogels.Higher Fe₃O₄ nanoparticle content in the composite hydrogel was obtained at higher precursor Fe²⁺/Fe³⁺concentration with a stronger magnetic response characteristics,but lower p H response characteristics and equilibrium swelling degree.The Fe₃O₄content dependent behavior of the prepared hydrogels indicates that the hydrogel performance is adjustable.The prepared hydrogels were further used for experiments on 5-Fu sustained release.It was found that the sustained release of 5-Fu from the hydrogel in simulated gastric fluid and intestinal fluid followed the Fick diffusion mechanism and showed different release rates,showing obvious p H-controlled drug release behavior.In addition,the cytotoxicity experiment showed that the composite hydrogels have no cytotoxicity and can be applied in the fields of drug sustained release.