Study On The Attenuation Of Arsenic Trioxide By Total Phenolic Acid From Nandina Domestica

Arsenic,the main component of which is arsenic trioxide.Arsenic trioxide is a novel anti-tumor drug capable of targeting natural products that induce apoptosis in tumor stem cells.Although arsenic trioxide can be targeted to induce apoptosis of tumor stem cells,it has obvious toxic side effects on heart,liver and kidney.The literature and previous studies showed that the extract of Nandina domestica are the main components of reducing the hepatorenal toxicity caused by arsenic trioxide.The phenolic acids in Nandina domestica were the main attenuating active components,and the attenuation of single phenolic acids was not as good as that of total phenolic acids.The total phenolic acids have obvious multi-target and multi-pass traditional Chinese medicine characteristics for the attenuation of arsenic trioxide.However,the separation and purification process of the total phenolic acids from Nandina domestica needs to be optimized,and the chemical constituents of the total phenolic acids from Nandina domestica need to be further clarified.Based on this,the related research of total phenolic acid on arsenic trioxide was carried out.This topic was divided into four parts,the specific research content is as follows: 一、Study on extraction and purification process of total phenolic acid from Nandina domesticaBy using Fulin phenolic acid colorimetry and Gallic acid as reference substance,the optimum extraction process was 20 times 70% ethanol reflux 3 times and 1.5 h.each time.Purification by Polyamide Resin Column Chromatography and the optimum purification conditions were as follows: the concentration of raw drug in the extract liquid was 5 mg/ml,the volume of the sample loading was 15 ml,the rate of flow was 2 ml/min,the impurity was first removed use water with 3 bv volume,then eluted with 3 bv volume of 30% ethanol at a flow rate of 1 ml/min and the total phenolic acid content was 72.2%.二、Study on the detoxification of arsenic trioxide by total phenolic acidThe effects of purified total phenolic acid on arsenic trioxide toxicity from Nandina domestica were compared by using the acute injury model induced by arsenic trioxide.Biochemical indicator effects showed that the compared with the model group,the levels of ALT and AST,BUN were significantly decreased(P<0.01),MDA were decreased(P<0.05),Cr 和 SOD,CAT were increased(P<0.05).And the biochemical indexes of total phenolic acids were better than those of total extracts.The histomorphology showed that Some hepatocytes in the model group displayed hepatocyte edema,punctate necrosis,typical hepatic sinusoid dilatation,steatosis,and a large number of inflammatory cell infiltration in the tissue.A small number of liver cells appeared edema and a small number of inflammatory cells infiltrated in the total extraction group.In the middle and low dose group of total phenolic acids show that a few hepatocyte edema was normal.In the model group,some glomerular atrophy and glomerular dilatation,inflammatory cells were seen in the renal capsule cavity,the renal tubular epithelial cells were swollen,some of them appeared to fall off,type and cell tube type were seen in the lumen,and hyperemia and swelling were seen in the interstitial and vascular tissues.Some of the renal tubules in Nandina domestica total extraction group were fused and expanded to form a large cystic cavity,the tissue could be seen blood vessel congestion,the glomerular capillary gadolinium could be clearly seen,and a small amount of inflammatory cell infiltration could be seen in the tissue.There were no obvious lesions in the glomerulus and renal tubules in the low,middle and high dose group of total phenolic acid in Nandina domestica.There was no tube type in the lumen of the tube,the renal interstitial was slightly widened and a few parts were swollen.But did not show significant dose dependence.The results indicated that the total phenolic acid in Nandina domestica has the effect of reducing liver and kidney toxicity to arsenic trioxide,and the effect of reducing toxicity is better than that of total extract.三、Component Separation and Identification of total phenolic acid in Nandina domesticaThe total phenolic acids from Nandina domestica were separated and purified by column chromatography and 24 compounds were identified.There are 10 simple phen olic acids(1-10),8 lignans(11-18),6 flavonoids(19-24).Including gallic acid(1),protocatechualdehyde(2),3,4-dihydroxyacetophenone(3),isovanillic acid(4),vanillic acid(5),vanillin(6),methyl dihydrocaffeate(7),methyl hydroxyphenylacetate(8),methyl gallate(9),progallin A(10)rel-(αR,βR)-4-Hydroxy-β-(4-hydroxy-3-methoxyphenyl)-γ,3-dimethoxybenzenepropano(11),isolariciresinol(12)(1S,2 R,3 R)-1,2,3,4-tetrahydro7-hydroxy-1-(4-hydroxy-3-methoxyphenyl)-6-methoxy3-acetate-2,3-naphthalenedimethanol(13),sesquipinsapol B(14),2,3-Dihydro-2-(4-hydroxy-3-methoxyphenyl)-3-(hydroxymethyl)-7-methoxy-5-benzofuranpropanol(15)glochidiobioside(16),β-D-Glucopyranoside,[1,2,3,4-tetrahydro-7-hydroxy-1-(4-hydroxy-3-methoxyphenyl)-3-(hydroxymethyl)-6-methoxy-2-naphthalenyl]methyl(17),syringaresinol-4’,4’-O-bis-β-D-glucoside(18),amentoflavone(19),(2R,3R)-2-(3,4-Dihydroxyphenyl)-3,4-dihydro-2H-1-benzopyr an-3,5,7-triol(20),5,7,4’-trihydroxyflavone(21),kaempferol-3-rhamnoside(22),apigenin-4’-O-glucoside(23),cinchonain Ib(24),among them compounds4～9,11,13～17,20～24 were separated from the plant for the first time.四、Qualitative and Quantitative Analysis of total phenolic acids in Nandina domesticaKempferol-3-rhamnoside was selected as the index component for TLC Identification,and 25℃,30%～70% humidity conditions,dichloromethane-methanol-acetic acid(10:1:0.5)was used as the unfolding agent 254 under ultraviolet lamp observation.The total phenolic acid solution from Nandina domestica fruit on the same thin plate was in the same position as kaempferol-3-rhamnoside,showing the same spots and has better durability.The content of syringaresinol-4’,4’-O-bis-β-D-glucoside was determined as the index component.The chromatographic column of SP ODS-A(4.6 mm ×250 mm,5μ m)was selected by HPLC.Acetonitrile-0.1% phosphoric acid solution was used as mobile phase and elution gradient :0 min,11%A;60min,25%A,elution time 60 min,flow rate 1 ml/min,injection volume 10 column greenhouse temperature,detection wavelength 210 nm chromatographic conditions,the content of 6.82mg/g.