RESUMO
ObjectiveTo investigate the transformation mechanism and content variation of saponins from Polygalae Radix before and after being boiled with licorice juice and water. MethodSimulated licorice juice boiled products and simulated water boiled products of onjisaponin B， onjisaponin Z， onjisaponin F， polygalasaponin ⅩⅩⅧ were prepared by simulated processing technology， and analyzed by ultra-performance liquid chromatography-quadrupole-electrostatic field orbitrap high resolution mass spectrometry（UPLC-Q-Exactive Orbitrap/MS）. Then the contents of onjisaponin B， onjisaponin Z， onjisaponin F， polygalasaponin ⅩⅩⅧ and tenuifolin in Polygalae Radix， licorice-boiled Polygalae Radix and water-boiled Polygalae Radix were determined by UPLC-triple quadrupole tandem mass spectrometry（UPLC-QQQ-MS/MS）. ResultDuring the boiling process with licorice juice and water， onjisaponin B could be hydrolyzed to produce 4-methoxycinnamic acid， desacylsenegin Ⅲ， polygalasaponin ⅩⅩⅧ and tenuifolin， onjisaponin Z could be hydrolyzed to produce 3，4，5-trimethoxycinnamic acid， onjisaponin TF， polygalasaponin ⅩⅩⅧ and tenuifolin， onjisaponin F could be hydrolyzed to produce 3，4，5-trimethoxycinnamic acid， onjisaponin G， polygalasaponin ⅩⅩⅧ and tenuifolin， and polygalasaponin ⅩⅩⅧ was hydrolyzed to produce tenuifolin. After being boiled with licorice juice or water， the content of onjisaponin B decreased significantly（P<0.05， P<0.01）， but the contents of onjisaponin Z， onjisaponin F， polygalasaponin ⅩⅩⅧ and tenuifolin increased significantly（P<0.05， P<0.01） in Polygalae Radix. Compared with the water-boiled products， the contents of onjisaponin Z and tenuifolin increased significantly（P<0.05， P<0.01）， and the change of tenuifolin content was the most significant in the licorice-boiled products.However， there was no significant difference in the content of onjisaponin B， onjisaponin F and polygalasaponin ⅩⅩⅧ between the water-boiled products and the licorice-boiled products. ConclusionBeing boiled with licorice juice or water can hydrolyze onjisaponin B， onjisaponin Z， onjisaponin F and polygalasaponin ⅩⅩⅧ， and generate secondary glycosides and aglycones（organic acids） through deglycosylation， which leads to obvious changes in the contents of onjisaponins after Polygalae Radix being processed.It is inferred that licorice juice can promote the hydrolysis of some onjisaponins in Polygalae Radix to onjisaponin Z and tenuifolin.This study provides an experimental basis for revealing processing mechanism of Polygalae Radix.
RESUMO
Objective: To optimize and establish the best hydrolysis method of diethyl ester 4-amino-N5-formyl-N8,N10-dideazatetrahydrofolate through the optimization of simple compound of diethyl N-(4-aminobenzoyl)-L-glutamate.Methods: To increase the low yield of hydrolysis reaction of diethyl ester 4-amino-N5-formyl-N8,N10-dideazatetrahydrofolate due to the by-products and difficult purification, we studied the effect of NaOH and KOH, two kinds of alkalis, three concentrations between 0.175-1 mol/L and five types of reaction time involved in 20, 30, 60, 120 and 180 min on the common side chain diethyl N-(4-aminobenzoyl)-L-glutamate.A high performance liquid chromatography was established for measuring the target product and the by-products in reaction liquid in different reaction conditions.Finally, on the basis of the best hydrolysis method of diethyl ester 4-amino-N5-formyl-N8,N10-dideazatetrahydrofolate, we completed the optimization of the hydrolysis reaction conditions of diethyl ester 4-amino-N5-formyl-N8,N10-dideazatetrahydrofolate.Results: We developed the best reaction condition for the hydrolysis of diethyl ester 4-amino-N5-formyl-N8,N10-dideazatetrahydrofolate, which could be carried out easily and efficiently.The results indicated that treated with the optimized condition of 0.3 mol/L KOH in 60 min at the room temperature, diethyl ester 4-amino-N5-formyl-N8,N10-dideazatetrahydrofolate was converted into its diacid derivative in 95.6 % yield, which turned to be a better reaction condition compared with the previous reaction condition.The structures of those compounds were identified to be correct by 1H nuclear magnetic resonance(1H NMR), 13C nuclear magnetic resonance(13C NMR) and electrospray ionization time of flight mass spectrometry (ESI-MS).The purity of the diacid derivative of the compound was determined to be 96% by high performance liquid chromatography(HPLC).The new hydrolysis reaction condition could not only avoid the formation of single ester hydrolysis product and amide bond hydrolysis product, but also improve the yield of the hydrolysis reaction.Conclusion: We have developed an efficient reaction for the hydrolysis of diethyl ester 4-amino-N5-formyl-N8,N10-dideazatetrahydro.Since the final step of the synthesis of classical folic acid antagonists is always the catalyzed hydrolysis of the side chain glutamate, the reaction also has great significance for anti-folic acid anti-tumor inhibitors synthesis.