| Objective: At present,commercially available medicinal gelatin is extracted from connective tissues such as bone,tendons,skin,and tendons of mammals such as pigs,cattle,and sheep.Its resources are limited,its production process is not conducive to environmental protection,and its production cost is relatively high,which cannot fully meet the diversified needs of domestic and international markets.In recent years,due to the frequent outbreak of some zoonotic diseases such as bovine spongiform encephalopathy,foot-and-mouth disease,and some religious factors,the application of gelatin extracted from cattle,pigs and other mammals has been greatly restricted.Using grass carp fish as raw material,this study adopts the control of grass carp fish gelatin and enzyme solution method,and the current medical gelatin is widely used in medical field characteristics and structure of comparison,to prove that grass carp fish gelatin as the possibility of a new source of medicinal gelatin,for the industrialization of grass carp fish gelatin and provide theoretical basis and its application in medical field.Treatment under the guidance of TCM theory,adopting the traditional fire burns of the active ingredients of Chinese medicine knotweed glycoside as the model drug,grass carp fish gelatin as the main material,the preparation of giant knotweed nucleoside scale gelatin film,and to study the efficacy and mechanism of action,for the application of grass carp fish gelatin as a supplementary material of TCM and provide the basis.Methods:(1)The appearance,morphology,amino acid composition,relative molecular weight distribution,and characteristic viscosity of the two gelatins were determined by scanning electron microscopy,high performance liquid chromatograph,SDS gel electrophoresis,dynamic rheometer and Ostwald viscosimeter,and the similarities and differences between grass carp scale gelatins and medicinal gelatins were compared.(2)Using ultraviolet spectrometer,Fourier transform infrared spectrometer,X-ray diffractometer,round two color spectrometer,differential thermal scanner,atomic force microscope,and other instruments on grass carp fish gelatin and secondary structure of medical gelatin were characterized structurally comparison between the two,exploring new sources of grass carp fish gelatin as a medicinal gelatin.(3)Using polydatin,the active component of polydatin as the model drug,and grass carp gelatin as the main excipient,polydatin gelatin film was prepared and its preparation process was optimized by orthogonal test to investigate the application performance of grass carp gelatin as a pharmaceutical excipient.(4)Using the rat model of deep second-degree burn on the back,wet-burn ointment was used as the positive control.The wound healing rate,Masson staining section,IL-6,Mc P-1,SOD,CD31,CD68,GRP78,and NF-?B indexes were detected and analyzed to investigate the therapeutic effect of polydatin fish scale gelatin film on deep second-degree burn.Results:(1)The gelatins of grass carp scales were different from those of medicine gelatin.Under scanning electron microscopy,the images showed that their surfaces were very smooth,and no obvious difference was observed between the microstructure of them.Amino acid analysis results showed that the glycine content of medicinal gelatin was higher than that of grass carp scale gelatin,and the hydroxyproline content of grass carp scale gelatin was higher than that of medicinal gelatin.The total content of glycine,proline,and hydroxyproline in grass carp scale gelatin was 40.9%,while that of medicinal gelatin was 38.8%.SDS gel electrophoresis results showed that there were two obvious ? bands in both fish scale gelatin and medicinal gelatin of grass carp.The relative molecular weight of ?1 chain in fish scale gelatin of grass carp was about 124000,and the relative molecular weight of ?2 chain was about107000,which was smaller than that of medicinal gelatin.The proportions of? and ? in the gelatin of grass carp scales were 41.2% and 48.3%,respectively,and the freezing strength was 310 Bloom g.The gelatins were 33.1% and19.4%,respectively,and the freezing strength was 200 Bloom g.The results of dynamic rheometer showed that the gelation temperatures of fish scale gelatin and medicinal gelatin were 20.2? and 24.9?,and the melting temperatures were 25.4? and 29.0?,respectively.The viscosities of fish scale gelatin and medicinal gelatin were 7.24 m Pa.s and 6.69 m Pa.s,respectively,at 60?.(2)UV spectrometric results showed that both fish scale gelatin and medicinal gelatin had absorption peaks at 235 nm and 280 nm,and the absorption intensity of fish scale gelatin at 280 nm was slightly higher than that of medicinal gelatin.Compared with the gelatin of grass carp,the amide A band became wider and the amide I band moved to lower wave numbers.X-ray diffraction analysis showed that the two samples were consistent with the characteristics of gelatin,and the diffraction peak intensity of fish scale gelatin of grass carp was stronger than that of medicinal gelatin at the angle of 7.89°2?.The results of circular dichroism spectrometry showed that the two gelatins had obvious absorption peaks at 198 nm,while the absorption peaks at 220 nm were not obvious.Compared with the two gelatins,the absorption intensity of fish scale gelatin was slightly higher than that of medicinal gelatin at 220 nm.Differential calorimetry analysis showed that the first absorption peak appeared in both samples when heated up to 30?,and the second main absorption peak was located at about 65?.The melting temperature of grass carp gelatin was slightly lower than that of medicinal gelatin.Atomic force microscopy(AFM)showed that the highest point of fish scale gelatin of grass carp was 3.12 nm,and that of medicinal gelatin was 2.24 nm.Both samples had good uniformity and smoothness,and did not contain a complete three-strand spiral structure.(3)The technological research results of polydatin fish scale gelatin film showed that the optimal production conditions were as follows:PVC plastic plate was used as template,the percentage of gelatin of grass carp scale was 8%,the percentage of glycerin was 0.3%,the percentage of polydatin extract was 1%,and the coating temperature was 40?.(4)Polydatin fish-scale gelatin membrane can significantly promote collagen fiber synthesis and angiogenesis,increase the number of macrophages and the ability of scavenging oxygen free radicals,and downregulate the expression of IL-6,Mc P-1 and NF-?B in deep second-degree scald wounds of rats.Conclusion:(1)The gelatin from grass carp scales prepared by controlled enzymatic hydrolysis is similar to the commercially available gelatin in physical properties and structure,and can be used as a new source of gelatin in the medical field.(2)The formulation of polydatin fish scale gelatin film preparation is reasonable,the preparation process is stable and feasible,and the established content determination method is accurate and reliable,which can provide qualified samples for the efficacy and mechanism of action.Fish scale gelatin has good film forming ability and drug loading ability,so it is suitable to be used as an excipient of this preparation.(3)Polydatin fish-scale gelatin membrane can significantly promote the wound healing of deep second-degree scald in SD rats.The mechanism of action is to promote the synthesis and angiogenesis of collagen fibers,increase the number of macrophages and the ability of scavenging oxygen free radicals in scald wounds,and reduce inflammation by downregulating the expression of IL-6,Mc P-1,and NF-?B.Effect of promoting scald wound healing. |
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