Study On The Production Quality Standard Of Lonicerae Japonicae Flos,the Raw Material Of Reduning Injection

The Lonicerae Japonicae Flos is derived from the dried buds or early blooming flowers of Lonicera japonica Thunb.It is harvested and dried before blooming in early summer.It is sweet and cold in nature,attach to the lung,heart and stomach meridians.It has the effects of clearing away heat,detoxifying and evacuating wind and heat.It is used for swelling and furuncle,throat palsy,erysipelas,heat toxin,hemorrhagic dysentery,wind-heat cold and febrile fever.Lonicerae Japonicae Flos is mainly distributed in Henan,Shandong,Hebei provinces in China,and is mostly cultivated.Lonicerae Japonicae Flos has many origins,abundant varieties and different harvesting and processing methods,which make the quality of Lonicerae Japonicae Flos different.In the market,Lonicerae Flos and sulfur fumigated Lonicerae Japonicae Flos are also sold,which has an impact on the quality,safety,effectiveness and stability of the preparation of Reduning injection with Lonicerae Japonicae Flos as the raw material.We intend to study different origins,different varieties,different harvesting and processing,different packaging and storage by establishing a quality evaluation method for Lonicerae Japonicae Flos.We studied the quality standard of Lonicerae Japonicae Flos seeds and seedlings,and standardized the production of medicinal materials from the source.The contents of glandular hairs in different harvesting periods of Lonicerae Japonicae Flos were qualitatively studied to find the differences of metabolites in different harvesting periods.The relationship between glandular hairs and their quality can be further revealed to provide some basic research for the quality differences of Lonicerae Japonicae Flos at different harvesting periods.The research contents of this paper are as follows:1.Literature ResearchThe chemical constituents of Lonicerae Japonicae Flos were summarized.It was found that Lonicerae Japonicae Flos contains many components,such as organic acids,flavonoids,iridoid glycosides,triterpene saponins and trace elements.After sorting out the quality evaluation of Lonicerae Japonicae Flos,it was found that the evaluation of Lonicerae Japonicae Flos medicinal materials mainly focused on chlorogenic acid and luteolin,and gradually developed towards multi-index component determination.The research progress of Lonicerae Japonicae Flos,Lonicerae Flos and sulfur fumigated Lonicerae Japonicae Flos was summarized,and the chemical constituents of the three species were found to be different and different,and the three could be distinguished by fingerprint,character identification and spectral imaging.2.Study on standardization of Lonicerae Japonicae Flos germplasmDifferent batches of Lonicerae Japonicae Flos seeds on the market were measured for their seed quality and morphological indexes,and their purity,clarity,1000-grain weight,water content,germination rate and viability were determined.Principal component analysis,cluster analysis and other methods were used to analyze them.According to water content,germination rate and 1000-grain weight,honeysuckle seeds were divided into three gradesThe seedling quality and morphological indexes of different batches of Lonicerae Japonicae Flos seedlings on the market were measured,and the ground diameter,plant height,main root length,number of branches and branches were measured.The annual,two-year and three-year Lonicerae Japonicae Flos cuttings were divided into three grades by plant height,ground diameter and branches number.3.Quality evaluation of Lonicerae Japonicae FlosCharacter identification and microscopic identification of Lonicerae Japonicae Flos were studied.The microscopic characteristics of Lonicerae Japonicae Flos were observed by powder identification and scanning electron microscopy.The structure of glandular hair,non glandular hair,pollen grain and calcium oxalate cluster crystal of Lonicerae Japonicae Flos can be clearly seenThe conditions of different deployment systems,different extraction methods,different Pharmacopoeia methods,different temperature and humidity were compared,On the basis of the original Pharmacopoeia,the TLC identification method of honeysuckle was optimized,and AICl3 was added as the chromogenic agent.and the comparison of luteolin and rutin reference spots was increased.The light yellow strip at RF value 0.34 and the light blue strip at RF value 0.17 can be used to distinguish Lonicerae Japonicae Flos and Lonicerae Flos,and the light blue white strip at RF value 0.3 and the light blue strip at RF value 0.71 can be used to distinguish Lonicerae Japonicae Flos and sulfur fumigated Lonicerae Japonicae Flos.The test items of Lonicerae Japonicae Flos samples were determined,and the moisture,ash and acid-insoluble ash all met the requirements of Chinese Pharmacopoeia.At the same time,impurities,flower opening rate,leachate determination and SO2 residue detection were added.In 46 batches of Lonicerae Japonicae Flos samples,the content of leachate was more than 30%,and the residual SO2 was less than 50 mg·kg-1.Eleven inorganic elements in Lonicerae Japonicae Flos were determined by ICP-MS.In 46 batches of Lonicerae Japonicae Flos,some elements such as Pb,Cd and Cu exceeded the standard.The contents of Sb,Ga,Ni,Cr,V and Ti elements were higher and distributed in the range of 10-30 mg·kg-1.The difference of Ga elements among different batches was the greatest and distributed in the range of 1-600 mg·kg-1.GC-MS method was used to determine 74 pesticide residues in 30 batches of Lonicera japonica Flos.It was found that the main pesticide residues in Lonicera japonica Flos were organic chlorine,including alpha-BHC,beta-BHC,-BHC,-BHC and penttnnitrobenzene(PCNB),and their contents were very low.The fingerprints of 30 batches of Lonicera japonica Flos were established by HPLC method,and 33 common peaks were identified.The common peak area was analyzed by principal component analysis(PCA)and partial least squares(OPLS-DA).Three regions of Shandong,Jiangsu,Henan and Hebei were clearly distinguished,and 11 different components were screened out.The method can distinguish Lonicerae Japonicae Flos,Lonicerae Flos and sulfur fumigated Lonicerae Japonicae Flos by increasing or decreasing characteristic peaks.Ten components,including seven organic acids,two iridoid glycosides and one flavonoid,were determined in 48 batches of Lonicerae Japonicae Flos from different origins by UPLC.Using OPLS-DA analysis,it was found that there was a dispersion trend in samples from different origins.The quality of medicinal materials from different origins(Shandong,Henan,Hebei,Shaanxi,Jiangsu)was evaluated by GRA and TOPSIS methods,and the samples from Jiangsu and Henan were better.In addition,a NIR quantitative model and a one-test multi-evaluation method were established for the quality evaluation of Lonicerae Japonicae Flos.UFLC-Q-TRAP-MS/MS was used to determine 29 kinds of nucleoside amino acids in 51 batches of Lonicerae Japonicae Flos,including 21 kinds of amino acids and 8 kinds of nucleosides.The results showed that the contents of alanine,phenylalanine,glutamine,aspartic acid and lysine were higher in honeysuckle.Using GRA and TOPSIS to evaluate LoniceraeJaponicae Flos,S10 is the best,S25 is the worst.4.Study on the collection,processing and packaging storage of Lonicerae Japonicae FlosThe content of 10 components in samples collected and processed by UPLC method was determined,and the quality of flowers was compared from different harvest time(rice grain,Sanqing,Erbai,Dabai,Yinhua,Jinhua),different stubble(one stubble flower,two stubble flower,three stubble flower,four stubble flower),different processing methods(air drying,infrared,microwave,sun drying,shade drying,green killing),different temperature(40?,50?,60?,70?).Combined with the biomass and the content of effective components,it was found that Lonicerae Japonicae Flos was better to be harvested in the period from Er Bai to Da Bai,and combined with the actual processing situation,appearance character and the content of effective components in the production area,Lonicerae Japonicae Flos was better to be processed by low-temperature inactivation and variable temperature drying.Through the established quality evaluation method of Lonicerae Japonicae Flos,the quality of Lonicerae Japonicae Flos such as different packaging materials(nylon bags,plastic bags,vacuum bags),different storage temperatures(low temperature 0-10?,normal temperature 25?)were investigated,and the optimal packaging storage conditions were explored.It was found that the moisture and chlorogenic acid content of Lonicerae Japonicae Flos did not change significantly within 6 months.Combined with the cost and actual situation,Lonicerae Japonicae Flos were packed in nylon bags and stored in a dry and cool place.5.Study on the changes of glandular hair morphology and composition of Lonicerae Japonicae Flos in different harvest timeMorphology(density and length of glandular hairs)of Lonicerae Japonicae Flos at different harvest time were studied by stereomicroscope and scanning electron microscope.It was found that the density of glandular hair and non glandular hair decreased gradually with the growth of flower bud,and the density was the highest at grain stage and the lowest at golden flower stage.There was no significant change in the length of glandular hair and non glandular hair.The glandular hairs of honeysuckle were obtained by laser microdissection,UFLC-QTOF-MS/MS was used to analyze the non-volatile substances in the glandular hairs of Lonicerae Japonicae Flos.It was found that there were 28 compounds in the glandular hairs of Lonicerae Japonicae Flos,including 5 organic acids,2 sugars,5 iridoids and 16 flavonoids.UFLC-QTOF-MS/MS technology was used to analyze the changes of glandular hair components in different harvest periods of Lonicerae Japonicae Flos.Combined with PC A and OPLS-DA analysis,it was found that different harvest time could be distinguished obviously.4 components were screened and identified in different harvest time,including 2 iridoid glycosides and 2 organic acids.6.SummaryThe quality standard(draft)of Lonicerae Japonicae Flos,the seed and seedling standard of Lonicerae Japonicae Flos,and the technical specifications for the harvesting,processing and storage of Lonicerae Japonicae Flos for Reduling injection were established,and the identification method with Lonicerae Flos and sulfur fumigated Lonicerae Japonicae Flos was constructed to provide scientific support for ensuring the effective,safe and stable quality of Lonicerae Japonicae Flos.