Preparation And Characteristics Of Silk Protein Surfactants And Their Application As Anti-drug Delivery Carrier

Silk is a natural protein polymer material of silkworm production,the process of silk floss quilt and textile silk reeling by silk in industry.The silk floos quit and raw silk will be produced in this process,but it also has a large number of industrial waste,such as waste silk,alkaline sericin waste liquid,etc.The waste material will be discarded directly,which not only causes environmental po llution,but also causes a great deal of waste of protein resources.In this experiment,the sericin protein surfactant(SSS)and fibroin protein surfactant(SFS)were preparation through silk sericin,waste silk and other industrial waste of silk floss quilt processing and silk reeling industry.The structures of the twokinds of amino acid-based surfactants were analyzed by different methods,and the various surface activities of the two surfactants were also studied in detail.In addition,two kinds of amino acid-based surfactants were used as raw materials to study their application as drug sustained-release carriers,and the physical and chemical properties were studied in detail in this experiment.At the same time,the cytotoxicity of two kinds of amino acid-based surfactants and catanionic aggregate solution were were investigated and studied.The research contents of this paper are related to the following aspects:(1)In this experiment,the double cocoon of silkworm as raw material for the process of silk floss quilt and fresh cocoon reeling of simulate industry.It would be producted a large number waste,like silk sericin solution,waste silk,cocoon lining and so on in this process.The silk fibroin amino acid solutions and sericin amino acid solutions were obtained by a series of processing and processing of these silk industrial wastes.Then the two amino acid solutions were made into powder by spray drying,and the powder for storage purposes.(2)In order to make full use of these silk industrial wastes,a large amount of silk fibroin and sericin amino acid powder was prepared in the above process into 20% solution,and a silk fibroin and sericin amino acid-based surfactants were prepared by Schotten-Banmann chemical synthesis.Then the structural characterization of the two kinds of amino acid-based surfactants were determined.Two kinds of amino acid-based surfactant and silk fibroin and sericin amino acid solution has certain difference from the apparent morphology and amino acid analysis results.Under the same concentration,the color of amino acid solution is darker than that of its corresponding surfactant,and the proportion of amino acids in solution was also greatly different.The total percentage of glycine and alanine in two kinds of amino acid-based surfactants were 18.86% and 71.05%,respectively.However,the total percentage of glycine and alanine in silk fibroin and sericin amino acid were 66.91% and 91.69%,respectively.In addition,the structures of the two kinds of amino ac id-based surfactants were analyzed by IR and 1H NMR and the results showed that their were the product of the target,namely sodium lauryl silk fibroin sodium salt and sodium lauryl sericin amino acid sodium salt.The results of TG-DTG showed that the synthesis of two kinds of surfactants of the lowest decomposition temperature was 260.62?C and 269.96?C,which indicates that the product has good thermal stability.The sericin and silk fibroin amino acid solutions are considered to have surface activity due to the unique structure of amino acids.In order to prove that the two synthetic amino acid-based surfactants with excellent surface active properties,determination of silk fibroin and sericin solution and two kinds of amino acid-based surfactants of critical micelle concentration(CMC),foaming properties,emulsifying properties,wetting properties and dispersion properties,while the lauryl sodium sulfate(SDS)was used as a control.According to the results,the CMC of SFS,SSS and SDS are 1.19 g/L,1.06 g/L and 1.12 g/L,respectively.The three kind of surfactants were similar,while the CMC of three kind of surfactants were 1/3 of sericin and silk fibroin solution.The emulsifying properties of SFS and SSS were 2~3 times of silk fibroin and sericin amino acid solution,and the emulsification ability of some organic solvents are also stronger than that of SDS.Under the different temperature of SFS,SSS,SDS,silk fibroin and sericin amino acid solution were measured of wetting properties,the results showed that with the increase of temperature of wetting time decrease,at the same temperature,the wetting time were close of SSS and SDS,slightly higher than SFS,but their were less than the wetting time of silk fibroin and sericin amino acid solution.Besides,the foaming properties and dispersing properties of the two amino acid surfactants were obviously superior to silk fibroin and sericin amino acid solutions,and their were close of SDS.These results indicate that silk industrial wastesare excellent raw materials for synthetic amino acid-based surfactants and the synthetic amino acid-based surfactants have superior surface activity properties.(3)In this paper,two kinds of synthesized amino acid-based surfactants were systematically studied as drug release carriers.Cationic anti-drug-cytarabine hydrochloride(CH)was selected as a sustained-release drug and combined with two kinds of anionic surfactants(SFS and SSS)to study the aggregates behavior of catanionic aggregate solution.First,the sample solution were prepared by mixing CH with SFS and SSS,respectively,at different ratios.Then the physical and chemical properties of the mixed solution with different proportions were determined by the morphology of the sample,TEM,conductivity and Zeta.The results showed that both catanionic aggregates solution of SFS-CH and SSS-CH could form catanionic vesicular structures,and the vesicle structure could exist stably.However,as the drug continued to increase,the vesicular structure was trans formed towards the micellar morphology.After that,the in vitro release behavior of different ratios of catanionic aggregates solution was studied by membrane dialysis.The results showed that the ratio of catanionic aggregates had a significant effect on the sustained-release behavior.The drug release rates of SFS-CH and SSS-CH of both catanionic aggregates with X1 = 0.2 were 75.1% and 68.7%,respectively,and the sustained release time were 24 h and 18 h,respectively.Compared with the pure drug contro l group,the kinds of catanionic aggregates has obvious sustained release effect.At the same time,in order to prove the possibility of using these two surfactants as sustained-release carriers,cytotoxicity of two kinds of amino acid-based surfactants of SFS and SSS were studied in two types of cancer cells,CEM and Ramous cells.The results showed that when the concentrations of SFS and SSS were in the range of 100 ?g / m L,the survival rates of cells at 24 h and 48 h were both higher than 80%.It indica ting that two kinds of surfactants had no obvious cytotoxicity to CEM cells and ramous cells at this concentration range.Subsequently,the same with the two kinds of leukemia cells as the research object,two kinds of catanionic aggregates and pure drug CH of anti-tumor effect in0.1,1,10,50 and 100 ? g/m L five concentration was studied.The results showed thatat 24 h and 100 ?g / m L,the inhibitory rates of CH,SFS-CH and SSS-CH on CEM cells were 40.36%,60.41% and 63.28%,respectively;and the inhibition rates on ramous cells were 43.17%,58.09% and 52.82%,respectively.When cultured 48 h,the inhibition rates of pure drugs CH,SFS-CH and SSS-CH on CEM cells were 54.19%,78.75% and 82.01%,respectively;and the inhibition rates on ramous cells were 50.85%,83.40% and 82.09%,respectively.It can be seen from the results that at the same time of incubation,the inhibitory rates of the two catanionic aggregates on CEM cells and Ramous cells were significantly higher than those of pure drug CH group.Besides the inhibitory effect of the three samples on the ce lls increased with increasing concentration,and with a clear time-dependent,with the extension of time,the three samples of the cell inhibition rate also increased.These results indicate that the two kinds of amino acid-based surfactants synthesized from silk industrial waste have anti-drug sustained-release ability and the property was stable.In conclusion,the waste from the silk industrysuch as waste silk,cocoon lining and sericin can be used as raw materials for synthetic two kinds of anionic lauroyl fibroin and sericin amino acid surfactant,and two kinds of amino acid-based surfactants have been tested to have excellent surface activity,which provides a new solution for the waste treatment of the silk industry.At the same time,the two kinds of amino acid-based surfactants were sustained-release carriers,which have stable performance and good sustained-release effect,which provides a new carrier for anti-drug release.In addition,it provides a new application example and experimental basis fo r the research of anticancer drugs / surfactants,and it is a new hope for the high value use of silk industry waste and the sustainable development of silk industry.