| Purpose: 1.The light transmittance and color of antler commercial herbs of different specifications and grades were measured.The fuzzy concept and color of pilose antler traditional character identification were quantified,and the identification and grading basis of different sizes and grades of velvet antler commercial herbs were provided,and a new and quantitative method to describe Chinese herbal medicine pieces was established.2.By the determination of water-soluble components,such as amino acids and polysaccharides and nucleosides of antler commercial herbs of different specifications and grades.The results were statistically analyzed,and the results were compared with the traditional identification and grading of specifications and grades,providing quality control basis for the classification of specifications and grades.3.Through quantitative identification index of transmittance and chroma,main component index of total hydrolysate amino acids,total essential amino acids,total free amino acids,total polysaccharide,total monosaccharide,total nucleoside of different specifications and grade antler commercial herbs,carry on tprincipal component analysis,and cluster analysis is carried out on principal components screened out by principal component analysis,and quality comprehensive evaluation method is provided from statistical point of view,and its specification and classification basis are simplified.Materials and methods: 1 Materials:Different specifications and grades deer antler commodity medicinal materials are detailed in Table 1-1.2 Methods: 2.1 Quantitative identification research 2.1.1 Transmittance measurement: The transmittance of the medicinal herbs of different specifications and grades of the deer antler was measured by the transmittance tester,make a statistical analysis of the results of the measurement,quantitative description of the fuzzy concepts of "translucent","waxy","eyed",and so on in the identification of characters.It will integrate the specifications and grades with no significant difference,and redivide the specifications and grades,in order to provide a new and quantifiable method for the identification of antler and similar Chinese medicinal materials.2.1.2 Chromaticity measurement : Through L*,a*,b* 3D chromaticity space,color difference meter was used to determine the color characteristic parameters of different specifications and grade deer antler medicinal herbs(external)and powder(internal).The results of the measurement are analyzed statistically to quantitatively describe the characteristic color of the traditional character identification,and provide the basis for quantitative identification and classification.2.2 Component analysis 2.2.1 Amino acids:With DNFB(2,4-dinitrofluorobenzene)as a derivatization reagent,the type and content of free and hydrolytic amino acids were determined by amino acid automatic analyzer before column derivatization,and analyzed by cluster analysis.The difference between different specifications and grades was compared.2.2.2 Polysaccharide ingredients:The samples were degreased,and crude polysaccharides were extracted by water extraction and alcohol precipitation.Sevage was used to remove protein and H2O2(hydrogen peroxide)was decolorized,and refined polysaccharides were got by washing with ethanol,petroleum ether and acetone.The content of the total polysaccharide in each sample was determined by phenol sulfuric acid colorimetry,and the polysaccharides were hydrolyzed to monosaccharide by TFA(Trifluoroacetic acid).Using PMP(1-phenyl-3-methyl-5 pyrazolone)as the derivatization reagent,the content of 6 kinds of monosaccharides and the composition of polysaccharides were determined by pre column derivation HPLC method,and the results were analyzed by cluster analysis to compare the differences between different specifications and grades.2.2.3 Nucleoside components:The contents of 5 nucleosides were determined by HPLC method,and the results were analyzed by cluster analysis.The differences between different specifications and grades were compared to provide quality control for identification and grading.2.3 Research on quality evaluation method: Indicators were composed of transmittance,chroma,total amino acids,total essential amino acids hydrolyzed,total free amino acids,total polysaccharides,total 6 monosaccharide and total 5 nucleosides.These indicators are analyzed by SPSS19.0 software for principal component analysis,and cluster analysis is carried out on principal components screened out by principal component analysis,and quality comprehensive evaluation method is provided from statistical point of view,and its specification and classification basis are simplified.Results: 1 Quantitative identification research results 1.1 Transmittance measurement results:Different specifications plun welwet antler yinpian of light transmittance different is statistically significant;Except for the second and third wax scale,white piece of first and second,third and fourth,the rest of the various specifications and grades of light transmittance are statistically significant.1.2 Chromaticity measurement results: Different specifications and grades of Cornu Cervi Pantotrichum color,at L*,a*,b* 3D color space has difference(P<0.05),And the The difference of L value of different grades of decoction pieces was the most significant.2 Component analysis results 2.1 Results of amino acid analysis:The correlation coefficients of 18 kinds of amino acids were all greater than 0.9986.In all samples,Cys and Trp were not detected in the "white slices" of the free samples,and all the hydrolysate samples were not detected in Trp.The rest samples contained 18 kinds of amino acids and 8 kinds of essential amino acids.The results of cluster analysis show that the angle of amino acids is basically the same as that of the division of the various specifications of the deer antler with the traditional experience.2.2 Results of polysaccharide analysis:Different specifications and grades of plum blossom deer antler pieces contain polysaccharides,the total polysaccharide content was 2.58~13.65 mg.g-1,and the correlation coefficient of linear regression equation is r=0.9994.The rate of recovery was 98.2%~103.9%,and RSD% was 2.2%.The results of cluster analysis showed that the division of "wax slices" and other specifications was basically consistent with the traditional experience from the angle of total polysaccharide components.In the analysis of polysaccharides,except for “blood slices” and “white slices”,other samples contain 6 kinds of monosaccharides,such as glucosamine,glucuronic acid,galactosidase,glucose,galactose and fucose.And 6 kinds of monosaccharide content determination,correlation coefficients of 6 kinds of monosaccharide were all greater than 0.9982,the average recovery were all between 96.4%-102.1%,with RSD% less than 3.0%.The content of monosaccharide in each sample is different,except for the variety of monosaccharide.2.3 Results of nucleoside analysis:The correlation coefficients of 5 kinds of nucleoside were all greater than 0.9993,the average recovery were all between 98.5%-103.8%,with RSD% less than 2.5%.There are different types of nucleosides in different specifications and grades of plum blossom deer antler slices.In addition to no thymidine found in "white slices",the 5 nucleosides were contained in all the other samples,and the contents of the nucleosides were also different except for the types of nucleosides.The results of cluster analysis showed that the classification of antler’s specifications from the angle of nucleoside was basically consistent with the traditional experience.3 Research results of quality evaluation method:By the principal component analysis,2 principal components(F1,F2)are selected.By the principal component analysis,2 principal components(F1,F2)are selected.According to the variance contribution rate of the 2 principal components,the comprehensive evaluation index was set up,and the quality ranking of the different specifications and grades deer antler medicinal materials was obtained;By cluster analysis,the specifications and grades of tdeer antler medicinal materials were reclassified.In addition to the “wax slices”,“blood slices”,the original 3 grades,4 grades are reduced to 2 grades,and the classification of other specifications and grades is consistent with the traditional division of experience.Conclusion:This study provides the basis for identification and classification of velvet antler commercial herbs by means of transmittance and chromaticity,and establishes a new and quantitative identification method for pilose antler and similar Chinese medicine;Through amino acids,polysaccharides,nucleosides and other components and content analysis,establish quality evaluation methods respectively,and these methods are simple,feasible,accurate,sensitive and reproducible.Through the establishment of principal component analysis,comprehensive quality evaluation is carried out,and the method of clustering analysis to provide differential quantization and simplified basis is feasible. |
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