绿茶中阻断亚硝胺合成的有效成分的分离鉴定

Separation and identification of active constituents in green tea for blocking N-nitrosamine formation

茶叶用沸水(1:10)提取10 min,过滤并冷冻干燥(Ⅰ);另将茶水用氯仿、乙酸乙酯先后提取浓缩,并与残水一起分别冷冻干燥,得组分Ⅱ、Ⅲ、Ⅳ。用Sephadex LH20对组分Ⅱ分离,以2 ml/min流速用氯仿:甲醇:石油醚(1:2:1)洗脱,监测280 nm处紫外吸收以收集馏分。对馏分F_(7～10)浓缩、纯化与重结晶,经UV和MS分析并与茶儿茶素标准相对照,F_7为(一)-表儿茶素(EC)和(一)-儿茶素(C), F_8为(一)-表没食子儿茶素(EGC), F_9为(一)-表儿茶素没食子酸酯ECG,F_10为(一)-表没食子儿茶素没食子酸酯(EGCG);HPLC测定其纯度,F_7为17％C和83％EC,F_8为99％以上EGC,F_9为92.5％ECG,F_10为93.6％EGCG。茶叶中阻断N-亚硝化的主要有效成分为茶多酚(组分Ⅲ),尤为茶儿茶素混合物;茶儿茶素的阻断能力与其与NO_2^-摩尔比及本身的抗氧化能力有关。

Green tea was extracted in boiling water(1:10)for to min, and the filtrate was freozen-dried(crude water extract, Ⅰ);or loaded with liqu- id-liquid partitioning from water to organic solvent of chloroform(CHCl_3) and ethyl acetate(EtoAc)respectively(CHCl_3 extract, Ⅱ,EtoAc extract, Ⅲ;lyophilizing aqueous residue, Ⅳ).EtoAc extract was loaded onto Sepadex LH-20, eluted with CHCl_3-methanol-petrol (1:2:1)at a flow rate of 2ml/min and fractions were collected based on monitoring UV absorbance at 280nm.Fractions(F7-F10)were concentrated, purified and recrystallized. The analytical data were obtained by UV and MS as well as HPLC. It was confirmed that data of F7 was identical with that of catechin(C)and epicatechin(EC)standard; F8, epigallocatechin(EGC); F9, epicatechin gallate(ECG);F10, epigallocatechin gallate(EGCG). The purity in HPLC of F7 was 17％C and 80% EC; F8, 99% EGC: F9, 92.5% ECG: F10, 93.6% EGCG.The result in blocking N-nitrosation in- dicated that the active constituent was mainly due to the polyphenols (EtoAc extract)in tea,especially tea catechines(flavanols).The effect of tea catechines on N-nitrosation depended on their molar ratio with nitrite as well as their antioxidative activity.