Studies On Processing Technology And Quality Standard Of Semen Sojae Preparatrm

Objective:Semen Sojae Preparatrm (SSP) was fermented by soybean, Folium Mori and Herba Artemisiae Annuae together to process finished product. It has the effects of inducing diaphoresis, relieving restlessness,and dispelling emission heat. It is used to treat flu, headache, fidgeting due to deficiency and insomnia, etc. Present clinical used SSP is process as 2005 edition pharmacopoeia of people republic of china, the processing control mainly depend on experiences and lack of quantizing control system, technological parameters was not defined clearly, for this reason make the quality of SSP unsteadily, and directly affect the security and validity of medication in clinical use. In order to guarantee the stable and steerable quality of SSP and create the scientific and normative processing technology, we studied and quantized the processing technological parameters. Meanwhile, we qualitative identify the soybean, Folium Mori and Herba Artemisiae Annuae in SSP, and establish the assaying methods of multi-index ingredients in SSP, enacted and consummated the scientific and advisable quality standard for providing the scientific evidence and method to control and evaluate the quality of SSP objectively.Methods:1 Studies on processing technologyUse single factor test, with the contents of Isoflavonids, Daidzein and Genistein of SSP as indexes, to investigate technological parameters of processing technology, such as dose percentage of Folium Mori and Herba Artemisiae Annuae, relative density of physic liquor in soybean, cooking time, fermentation temperature, fermentation time, repeated fermentation time and appreciably steaming time, etc; use 4 factors, 3 levels orthogonal design, with the content of total isoflavones in Folium Mori and Herba Artemisiae nnuae as control index, select the decoction time, decoction frequency and water dose of Folium Mori and Herba Artemisiae Annuae as the influencing factors of the content of total flavones, to optimize the best decoction technology of Folium Mori and Herba Artemisiae Annuae, furthermore, to testify the decoction technology with practice, through the verification test, determine the decoction technology of Folium Mori and Herba Artemisiae Annuae. Finally, optimize the processing technology of SSP, identify and quantify the technological parameters. In order to investigate the feasibility of the processing technology, take the above-mentioned research result to proceed three semi-works production and check out whether all indexes are accord with the requirement, supply evidence for normalization manufacture and quality control of SSP.2 Studies on quality standardUse the optimal processing technology to process the SSP, to research and establish the quality standard of SSP from aspects of thin-layer chromatography and assaying etc.2.1 thin layer chromatography (TLC) methodUse TLC method to optimum the preparation method of test solution, developing solvent system, visual inspection method, and establish the TLC methods of soybean, Folium Mori and Herba Artemisiae Annuae in SSP.2.2 assaying methodHPLC and UV methods were established to determine the content of Daidzein, Genistein and isoflavones, as well as the methodology. Via detecting the content of sample and regulating homologue content limitation, make considerable evidence to evaluate the quality of Semen Sojae Praeparatum.Results:1 The studies of the processing technologyBy investigating the processing technology parameters, the optimal processing technology is: take 90 g Folium Mori and 100 g Herba Artemisiae Annuae, add in water about 18 times, decoct for 3 times, 1 hour per time, filter and liquid medicine concentrate to relative density for 1.10～1.12 g/cm3, then mix in 1000 g soybean, absorb completely and steam 1.5 hour, take out and cool a little, cover soybean with dregs of decoction of Folium Mori and Herba Artemisiae Annuae, put them in the incubator at (30±2)℃, ferment for 6～8 days until"yellow clothes"overgrow, take out and remove dregs of a decoction, wash cleanly and dry. Via determining all indexes of three semi-works production, the results showed that the quality of product accord requirement, it was demonstrated that the processing technology is reasonable and feasible.2 The studies of the quality standard2.1 The studies of TLC identification2.1.1 the TLC identification of soybean in SSPAccording to TLC method, used the Daidzein and Genistein as the comparison, the same color spot appeared in test chromatogram and reference standard chromatogram, it should show positive result to prove the existence of soy bean in SSP.2.1.2 the TLC identification of Folium Mori and Herba Artemisiae Annuae in SSPAccording to TLC method, use Folium Mori and Herba Artemisiae Annuae as the the comparison, due to use developing solvent system and visual inspection method identically, the the same color spot appeared in test chromatogram and reference standard chromatogram, it should show positive result to prove the existence of Folium Mori and Herba Artemisiae Annuae in SSP.2.2 The studies of the assaying2.2.1 HPLC method for the determination of Daidzein and GenisteinChromatographic conditions and system suitability test: 18 alkyl silane bonded silica was used as filler and methanol-0.1% glacial acetic acid(50:50) as the mobile phase, the detection wavelength is 261 nm, and the flow rate is 1 ml/min. Theoretical plate number calculated by Genistein should not be lower than 3000.Reference standard solution preparation: precisely take appropriate amounts of Daidzein and Genistein reference substance and put them into MeOH to make containing 10.3μg and 9.92μg Daidzein and Genistein per milliliter solution respectively.Test solution preparation: precisely take the 0.5 g power of SSP and put them into erlenmeyer flask, precisely add up to 20 mL 50% alcohol and weighting, deal with ultrasound 50 min and take out, then repeated weighting, use the same solution to make up for the missing solution, shake up evenly and filtration.Determination: draw the test solution and control solution 10μl respectively, inject into HPLC to determine the content.According to the dry SSP to calculate, the content of Daidzein (C15H10O4) and Genistein (C15H10O5) should not be lower than 0.063%and 0.040% respectively.2.2.2 UV method for the determination of IsoflavonesReference standard solution preparation: precisely take appropriate amounts of Genistein and put it into alcohol to make it containing 72.0μg Genistein per milliliter solution.Standard curve preparation: precisely take 1.0ml, 2.0ml, 3.0ml, 4.0ml, 5.0ml Genistein reference standard solution and put them into 50ml erlenmeyer flask, add up alcohol to solve the residue and dilute to scale, and shake up evenly, use UV method to determine the absorption value at 261nm, abscissa was concentration and ordinate was absorption, draw the standard curve.Determination: precisely draw 5 ml extracting solution, put it into a evaporating dish, and make it dry, add up 15 ml water to solve the residue, extract with acetidin from water solution 6 times, and toties quoties 25 ml. then combine acetidin layer, and make it dry, add up alcohol to solve the residue into 25 ml volumetric flask and dilute to scale, and shake up evenly, then precisely draw 3 ml into 10ml volumetric flask and dilute to scale, and shake up evenly, to determine the absorption value with UV method, standby.According to the dry SSP to caculate, the content of Isoflavones should not be lower than 0.223% by the content of Genistein (C15H10O5). Conclusions:1 via the quantify study all procedure of processing technology, formulate the feasible processing technology parameter and establish quantization indexes of processing technology, accomplish the normalization processing technology, it was steady and feasible to process SSP.2 the study established TLC methods of identifying Folium Mori and Herba Artemisiae Annuae in SSP in the same thin layer plate, developing solvent, visual inspection method at the first time, and TLC methods of identifying soybean, it was easy, special and accurate, and provide new the method for identifying the real and false of SSP.3 HPLC and UV methods were established for the determination of the content of Daidzein, Genistein and Isoflavones, they were simple, stable and reliable, and use to multi-index composition assaying, further to control the quality of SSP.The study formulated and perfected feasible and normative processing technology and quality standard to provide the evidence for pharmacopoeias standardization of SSP, control the stable quality efficiently and certify the security and reliability for clinical use.