Isolation,Purification And Antihypertensive Test Of Laminine Synthesis

Laminine is a widely-existing natural active substance in seaweed kelp-non-protein amino acid.As an antihypertensive drug with little side effects and good antihypertensive effect,it has extremely broad market and research application prospects.Because the content of laminin in seaweed kelp is minimal,and the current separation and purification process is not yet mature,it is impossible to achieve industrial production to meet the market demand.In response to this problem,this subject is based on chemical synthesis and explores a complete and efficient production process of laminine,laying the foundation for industrialization.The research content is as follows:1.Optimization of the synthesis process of laminine:First,using lysine as a substrate,nickel ions are connected to?-NH₂ of lysine to produce nickel lysine.Using the response surface to optimize the synthesis method of nickel lysine,the study determined the best synthesis conditions of nickel lysine:pH=10.15 of the reaction solution,temperature 47?,and reaction 4.2h.A standard curve was established using an atomic absorption spectrophotometer,and the nickel lysine yield was calculated to be 89.23%,and the complex ratio(Ni²⁺:lysine)was 2.The yield is increased by about10%relative to the unoptimized synthetic route.The results of infrared spectroscopy prove that Ni²⁺is connected to the C=O bond and N bond of lysine.Second,the methylation reaction of nickel lysine with dimethyl carbonate?DMC?produces laminine.The silica gel column was separated and purified and analyzed by means of nuclear magnetic resonance?NMR?,mass spectrometry?MS?,ultraviolet and high-performance liquid.The data of NMR and MS confirmed that the synthesized methylated compound was laminine.2.Research on the post-processing technology of laminine synthesis:the ion exchange resin D001x7 is used to separate and purify laminine,the response surface method is used to optimize the adsorption conditions,the pre-column derivation of o-phthalaldehyde?OPA?,and the HPLC quantitative analysis to calculate the resin adsorption the amount.Finally,it was determined that the aqueous solution of crude laminic acid was adsorbed in a dynamic and static manner:shaking at a constant temperature of 30°C for 40 min,and then transferred to a chromatography column with a constant flow rate of 0.7 ml/min.Desorption:pH 6.6 citric acid-disodium hydrogen phosphate buffer?pH 7.0 citric acid-disodium hydrogen phosphate buffer?pH 10.0 ammonia-ammonium chloride buffer.Analysis by nuclear magnetic resonance?NMR?,liquid-mass spectrometry?LC-MS?and melting point analyzer confirmed that the compound isolated and purified was laminine.According to calculations,this purification process will increase the separation yield to 64%,which is about 14%higher than that of silica gel columns.3.Evaluation of the antihypertensive activity of pure laminic acid product:Spontaneously hypertensive rats were selected,and randomized trials were conducted to explore the effects of different doses,cycles of action,and administration time on blood pressure.The test showed that regardless of the dose size,laminin exhibited antihypertensive activity.Among them,the high-dose?4.0mg/?kg·d??administration has the most obvious antihypertensive effect.The single-dose experiment shows that laminin can play an immediate antihypertensive effect within 0.5h after administration.With the prolonged medication cycle,the effect of reducing blood pressure becomes more and more significant.This study significantly improves the synthesis process of laminine.The atomic absorption spectrophotometry used makes the calculation of nickel lysine yield more accurate.In this project,the newly established crude product pretreatment method-water extraction and alcohol precipitation method combined with sodium type D001×7 strong acid cation exchange resin,obtained cuminine with higher separation yield.The pre-column derivatization method of phthalaldehyde?OPA?provides a specific analytical method for the study of laminine.The antihypertensive test of rats has proved the antihypertensive effect of pure cumbrine,eliminating the influence of impurities.In summary,this study established a complete and efficient production process of laminine,which provides ideas for the study of laminine production technology and provides a basis for the clinical study of laminine.This is of great significance to the industrialization of laminine.