米糠酚类物质的大孔树脂分离纯化工艺

【目的】建立米糠中酚类物质的大孔树脂分离纯化工艺,比较米糠提取物纯化前后总酚、总黄酮含量及抗氧化能力,为米糠的深加工利用提供参考。【方法】比较9种不同类型大孔吸附树脂(HPD-100、HPD-200A、HPD-300、HPD-700、D101、HPD-722、AB-8、ADS17和HPD-826)对米糠提取物中总酚的静态吸附和解吸性能,筛选出对脱脂米糠酚类物质纯化效果最佳的大孔吸附树脂类型;通过考察其静态吸附动力学曲线,比较不同上样液浓度、上样流速和上样体积下的动态吸附曲线以及采用不同浓度乙醇洗脱的解吸效果,建立米糠多酚的大孔吸附树脂吸附和解吸工艺条件;比较HPD-300型大孔吸附树脂纯化前后总酚、总黄酮含量,采用高效液相色谱法结合应用酚类标准品分析米糠提取物经大孔吸附树脂纯化前后其各单体酚类物质种类与含量,并通过氧自由基吸收能力(oxygen radical absorption capacity,ORAC)和细胞抗氧化分析法(cellular antioxidant activity,CAA)比较其纯化前后的抗氧化活性。【结果】9种类型大孔吸附树脂对脱脂米糠提取物酚类物质均有一定的吸附能力,其中HPD-300型大孔吸附树脂对脱脂米糠总酚静态吸附和解吸效果明显优于其他类型树脂,其饱和吸附量为9.04 mg GAE·g~(-1),解吸率为82.62%,同时米糠总酚静态吸附在6 h时达到饱和值。HPD-300型大孔吸附树脂最佳吸附和解吸工艺条件为:米糠酚类提取液上样浓度为1.0 mg GAE·m L-1,上样流速为3.0 BV·h~(-1),上样体积为3.5 BV,洗脱剂为70%乙醇溶液,洗脱流速为3.0 BV·h~(-1)。经HPD-300型大孔吸附树脂纯化后,米糠酚类提取物的总酚和总黄酮含量分别从纯化前的88.07μg GAE·mg~(-1)和30.04μg CE·mg~(-1)提高到320.72μg GAE·mg~(-1)和133.67μg CE·mg~(-1),分别提高了2.6和3.4倍。经高效液相色谱分析,米糠酚类提取物经HPD-300型大孔吸附树脂纯化后,其中的没食子酸、原儿茶酸、香草酸、对羟基苯甲酸、咖啡酸、丁香酸、表儿茶素、香草醛、对香豆酸和阿魏酸等10种酚类单体组成未见明显变化和损失,且上述酚类单体在提取物中的含量较纯化前分别提高0.4—2.4倍。纯化后提取物的ORAC和CAA值分别为3 248.21μmol TE·g~(-1)和95.24μmol QE·g~(-1),较纯化前的1 327.51μmol TE·g~(-1)和29.19μmol QE·g~(-1),分别提高了1.4和2.2倍。【结论】HPD-300型大孔吸附树脂适合米糠酚类物质的分离纯化,经其纯化后的米糠提取物中总酚、总黄酮含量及抗氧化活性提高1—3倍,且不会造成单体酚的明显损失。

Separation and Purification of Polyphenols in Rice Bran by Macroporous Resins

【Objective】The objective of this study is to screen a resin with a good adsorption and desorption performance to rice bran polyphenols (RBP) and optimize the purification technique, to compare the total phenols, total flavonoids content and antioxidant activity before and after purification, thus providing a theoretical basis for the deep processing and utilization of rice bran.【Method】The behavior of static adsorption and desorption, dynamic adsorption and desorption of 9 macroporous resins (HPD-100, HPD-200A, HPD-300, HPD-700, D101, HPD-722, AB-8, ADS17, and HPD-826) with different polarities to RBP was determined. Kinetic characteristics of static adsorption of the preferred resin were analyzed. The purification technique of macroporous resin to RBP was developed through the dynamic adsorption and desorption curves under different loading and elution conditions, including phenolic concentrations in loading samples, sampling rates, sampling volumes and ethanol concentrations in eluent. The phenolic profiles in RBP before and after resin purification were analyzed by HPLC analysis. Their total phenolic and flavonoid contents, oxygen radical absorption capacity (ORAC) and cellular antioxidant activity (CAA) were also estimated. 【Result】HPD-300 exhibited the best capability of adsorption and desorption to RBP among the 9 selected macroporous resins. The saturated absorption quantity of HPD-300 resin to RBP was 9.04 mg GAE·g^-1, and the desorption rate was 82.62%. It took 6 h for HPD-300 resin to reach absorption equilibrium. The optimized purification parameters were as follows: the phenolic concentration of loading sample is 1 mg GAE·mL^-1, sampling rate is 3.0 BV·h^-1, sampling volume is 3.5 BV, eluent is 70% ethanol, flow velocity is 3.0 B V·h^-1. After HPD-300 resin purification, the total phenolic content increased by 2.6 times from 88.07 µg GAE·mg^-1 to 320.72 µg GAE·mg^-1, and the total flavonoids content increased by 3.4 times from 30.04 µg CE·mg^-1 to 133.67 µg CE·mg^-1. Ten phenolic compounds were identified from rice bran phenolic extract, which were gallic acid, protocatechuic acid, vanillic acid, p-hydroxy benzoic acid, caffeic acid, clove acid, epicatechin, catechins, vanillic aldehyde, p-coumaric acid and ferulic acid by HPLC analysis. The phenolic profiles of the extract were not changed after HPD-300 resin purification, but the concentrations of 10 phenolic compounds increased by 0.4-2.4 times. The ORAC and CAA antioxidant activity of the extract changed from 1 327.51 µmol TE·g^-1 and 29.19 µmol QE·g^-1 before purification to 3 248.21 µmol TE·g^-1 and 95.24 µmol QE·g^-1 after purification, and increased by 1.4 and 2.2 times, respectively.【Conclusion】The total phenolic and flavonoid contents and antioxidant activity increased by 1-3 times after HPD-300 resin purification. The individual phenolic compounds in the extract were not lost significantly during purification. Therefore, HPD-300 macroporous resin was suitable to purify rice bran phenolic extract.