以绿茶和绿茶与白参菌共发酵茶为研究对象,建立快速同时检测绿茶与白参菌共发酵前后茶叶中的6种活性成分含量变化的高效液相色谱检测方法。将白参菌与绿茶共发酵,制备白参菌发酵茶。采用WondaSil C18柱(4.6 mm×250 mm,5 μm),乙腈(A)- 0.2%H3PO4溶液(B)为流动相,梯度洗脱(0～15 min,5%A→15%A; 15～40 min,15%A→25%A;40～45 min,25%A→5%A),流速1 mL/min,柱温30 ℃,检测波长为280 nm检测茶叶发酵前后的活性成分含量。结果表明,绿茶经过与白参菌共发酵后,咖啡碱和可可碱含量几乎没有变化,表没食子儿茶素、表儿茶素、表没食子儿茶素没食子酸酯、表儿茶素没食子酸酯的含量均大幅下降。6种活性成分在6.25~2 000 μg/mL下线性关系良好,相关系数(R)在0.995 5～0.999 9,检出限在0.312 5～1.870 μg/mL,绿茶的加标回收率在95.06%~111.06%,白参菌茶的加标回收率在85.06%~118.88%。白参菌与绿茶共发酵前后化学成分的含量发生变化,该液相检测方法快速、准确、重现性好,能够满足白参菌茶中多种活性成分同时定性、定量检测需求,具有一定的应用价值。
Using green tea and green tea co-fermented with Schizophyllum communeh (S. communeh fermented tea) as raw materials, we established a high-performance liquid chromatography (HPLC)-based method for rapid and simultaneous determination of changes in the contents of six active ingredients in green tea leaves before and after co-fermentation with S. communeh. Using a WondaSil C18 column (4.6 mm × 250 mm × 5 μm) with acetonitrile (A) and 0.2% phosphoric acid (B) as the mobile phase, we performed separation by gradient elution (0-15 min, 5% A→15% A; 15-40 min, 15% A→25% A; 40-45 min, 25% A→5% A) with a flow rate of 1 mL/min and column temperature of 30 ℃. The contents of active ingredients in the tea leaves before and after fermentation were measured using a detection wavelength of 280 nm. The caffeine and theobromine contents in green tea barely changed after co-fermentation, whereas the epicatechin, epigallocatechin, epicatechin gallate and epigallocatechin gallate contents significantly decreased. All six active ingredients exhibited good linear relationships in the range of 6.25-2 000 μg/mL, with correlation coefficients in the range of 0.995 5-0.999 9. Detection limits were in the range of 0.3125-1.870 μg/mL and the recovery rates obtained using the standard addition method were in the range of 95.06%-111.06% for green tea and 85.06%-118.88% for S. communeh fermented tea. The HPLC-based method developed in this study has potential applications in the testing of active components of tea products, as it is rapid and accurate, with good reproducibility, and has been proven suitable for simultaneous qualitative and quantitative determination of multiple active ingredients in S. communeh fermented tea.
[1] BARBOSA D S. Green tea polyphenolic compounds and human health[J]. Journal für Verbraucherschutz und Lebensmittelsicherheit, 2007, 2(4): 407-413.
[2] 卢明. 茶叶的研究与应用[J].中国社区医师, 2005,7(22):16.
[3] 陈睿. 茶叶功能性成分的化学组成及应用[J]. 安徽农业科学, 2004,32(5):1 031-1 033;1 036.
[4] 宛晓春,李大祥,张正竹,等. 茶叶生物化学研究进展[J]. 茶叶科学, 2015(1):1-10.
[5] 龚智宏,陈思,高江涛,等.半制备型液相色谱法分离纯化茶叶鲜叶中7种儿茶素类化合物[J].色谱, 2017,35(11): 1 192-1 197.
[6] 王向阳,周政子,刘绘景. 茶多酚对萝卜干3种致病菌的抑菌机理研究[J].中国食品学报, 2013,13(8):96-101.
[7] 王丽丽,黄建安,刘仲华. 茶叶降脂功效及其作用机理研究进展[J].中国茶叶, 2011(9):12-15.
[8] LI Qin, LI Yongdi, LUO Yu, et al. Shifts in diversity and function of the bacterial community during the manufacture of Fu brick tea[J]. Food microbiology,2019,80:70-76.
[9] 夏丽飞,陈玫,蔡丽. 白参菌发酵茶品质及安全性研究[J]. 中国农学通报, 2019,35(4):143-146.
[10] WANG Q P,ŠARKANJ B,JURASOVIC J. Evaluation of microbial toxins, trace elements and sensory properties of a high-theabrownins instant Pu-erh tea produced using Aspergillus tubingensis via submerged fermentation[J]. International Journal of Food Science & Technology, 2019, 54(5):1 541-1 549.
[11] 刘佳奇,熊涛,李军波,等.乳酸菌发酵茶饮料的工艺优化及其发酵前后香气成分分析[J].食品与发酵工业,2016,42(8):109-114.
[12] 侯冬岩,刁全平,李铁纯.不同发酵类型的茶叶茶多酚含量的比较分析[J]. 鞍山师范学院学报, 2015,17(6):38-40.
[13] 王峰.超高效液相色谱法(UPLC)快速测定茶饮料中的咖啡因[J].食品与发酵工业,2011,37(11):213-215.
[14] 朱叶梅,张雯,杨少杰,等.HPLC法测定茶叶中的7种多酚类化合物的含量[J].云南化工,2018,45(9):66-68.
[15] 孔谦,陈栋,陈庄,等.柱前衍生-反相高效液相色谱测定茶叶中19种氨基酸含量[J].食品科技,2018,43(10):320-325.
[16] 王富花.HPLC分析测定不同茶叶中的游离氨基酸[J].食品研发与开发,2018,39(1):141-146.
[17] GE Yahui, BIAN Xiong, SUN Baoqing. Dynamic profiling of phenolic acids during Pu-erh tea fermentation using derivatization LC-MS approach[J]. Journal of agricultural and food chemistry, 2019, 67(16): 4 568-4 577.
[18] 王玉娇,刘通,邓亚美,等.QuEChERS-气相色谱-串联质谱法快速检测茶叶中6种禁用香精成分[J].色谱,2019,37(7):766-772.
[19] 李志,焦彦朝,王兴宁.气相色谱-质谱联用法测定茶叶中21种农药残留[J].中国酿造,2019,38(3):181-186.
[20] 邝柏贤,陈曼,周慈敏,等.茶多酚检测方法综述[J]. 广东茶业, 2016(2):14-16.
[21] ZHANG M H, LUYPAERT J,FERNǍNDEZ P J A, et al. Determination of total antioxidant capacity in green tea by near-infrared spectroscopy and multivariate calibration[J]. Talanta, 2004, 62(1):25-35.
[22] 孙延春,张英.紫外光谱法测定饮料中的咖啡因含量[J]. 化学研究, 2011,22(1):77-79.
