黑茶是六大茶类之一，是西北边疆地区人们日不可少的必需饮品。人们长期的消费实践和我们前期的研究均表明，黑茶具有明显的降脂减肥作用。本研究从黑茶化学物质组学研究入手，通过对65个中国黑茶中22个常量化学成分指标分析，采用主成分分析探寻了不同黑茶的常规化学物质组学表征属性；采用UPLC- Q/TOF等分析方法，探讨了不同黑茶的化学指纹图谱特征差异，表征了6个不同黑茶类型中19种已知活性成分组成特征差异，并从不同黑茶中共分离鉴定出了92种化学成分，主要包括儿茶素类及其衍生物、黄酮类、花青素与原花色素类、有机酸类、生物碱类、茶黄素及其衍生物，且多数化学成分是其他茶类中含量低微或不存在的活性物质。因此，基本探明了黑茶的化学物质组学特征。选择工艺与品质特征典型的茯砖茶和六堡茶，通过建立营养性肥胖大鼠模型和高脂血症大鼠模型、3T3-L1前脂肪细胞模型和HepG2细胞模型，结合脂质代谢生理生化指标分析、细胞增殖分化与代谢分析、基因表达及蛋白质组学分析，确证了黑茶具有显著的减肥和降血脂功效，且有一定的量效关系；动物实验表明，与脂质代谢相关基因的qPCR分析发现，黑茶可促使与脂肪生成相关的FAS和SREBP-1c基因及与脂肪细胞分化相关的C/EBP-α基因显著下调表达，抑制肝脏中脂肪酸的合成，减少了脂质的聚集。相反，促使与脂肪分解(脂肪酸β-氧化)相关的PPAR-α和CPT-1a基因及调降血液中胆固醇含量的LDLR基因显著上调表达，促进脂肪酸β-氧化，增加能量消耗，减轻体重，同时催化TG、TC、LDL-C分解，降低血液中血脂水平。细胞学研究表明，黑茶可以有效抑制3T3-L1前脂肪细胞的增殖和分化，调控肝细胞HepG2中与脂肪代谢相关酶系的基因表达，有效控制肝细胞HepG2中的脂质积累。双向凝胶电泳与LC-MS/MS联用的定量蛋白质组学分析进一步确证，黑茶(茯砖茶)是通过下调与脂肪酸的生物合成起重要作用的相关酶ACACB和FAS及与脂肪生成密切相关的前体蛋白3-磷酸甘油生成所需的关键酶GPD1的表达，上调与脂肪酸β-氧化相关的酶，如肉碱-O-酶棕榈酰转移酶(CPT1和CPT2)、烯酰辅酶A水合酶(ECHS1)、羟酰辅酶A脱氢酶(HADH)的表达，从而抑制脂肪酸、脂肪的生成以及增强脂肪酸β-氧化来调节脂质代谢。COX2(细胞色素C氧化酶亚基2)、ACACB(乙酰辅酶A羧化酶2)、FAS(脂肪酸合成酶)、ECHS1(烯酰辅酶A水合酶)等蛋白质可能是黑茶改善脂质代谢紊乱的潜在药物靶标。因此，从生化代谢、细胞学、基因表达和蛋白质组学水平上基本探明了黑茶的降脂减肥作用机理。
Dark tea, one of the six major types of Chinese tea, which has been a necessary beverage for the ethnic groups living in border regions of northern and western China. Despite the rough raw material and less water soluble components in dark tea, the significantly anti-obesity and hypolipidemic effect of dark tea were observed by our previous studies and the long-time consumption. For exploring the chenomics characteristic, 22 common chemical components in 65 Chinese dark teas were analyzed and the differences in the composition of different dark tea was explored by PCA method. UPLC-Q/TOF was used for comprehensive analysis of the water extract from six types of dark tea. The difference of chemical fingerprint chromatography and characteristic component were also compared in different types of dark tea. Through MRM analysis, 19 known bio-active ingredient showed a significantly different level in different dark teas. The full chemical profile showed significantly difference in different dark teas. 92 phytochemical compounds were identified in dark teas, which include catechins and its derivatives, phenolic acid and its derivatives, flavones and flavoneglycoside, organic acids and procyanidins, etc. Fuzhuan brick tea and Liubao tea were selected for evaluating the anti-obesity and hypolipidemic effect of dark tea. Through establishing the model of obesity and hyperlipidemia rats, 3T3-L1 preadipocyte cell and HepG2 cell model, physiological and biochemical analysis, cell proliferation, differentiation and metabolism analysis, gene expression and proteomic analysis, the results showed that dark tea could significantly lose weight and reduce blood fat in a dose-depended manner. Animal experiment showed that dark tea significantly suppressed the increase of body weight and accumulation of adipose tissue, and reduced the level of serum TG, TC and LDL-C in obese rats fed a high-fat diet. Moreover, dark tea attenuated the gene expression of FAS, SREBP-1c and C/EBP-α, which is related to lipogenic metabolism and adipose differentiation. In contrast, the gene expressions of enzymes involved in energy expenditure and lipodieresis including hepatic PPAR-α, CPT-1a and LDLR gene expression were increased by dark tea treatment. Cell experiment showed that dark tea inhibited proliferation and differentiation 3T3-Ll, and also lowered TG and TC contents in HepG2 in a dose-depended manner. Dark tea also regulated the genes expression that related to lipid metabolism. Quantitative proteomics analysis using 2-DE-LC-MS/MS technology also showed that dark tea significantly down-regulated the protein expression of ACACB, FAS and GPD195, which related fatty acid synthesis and lipogenesis. And the protein expression of CPT1, CPT2, ECHS1 and HADH that related to β-oxidation were up-regulated by dark tea treatment, which lead to inhibition of fatty acid synthesis and enhance the β-oxidation of fatty acid. Additionally, some putative drug targets were also revealed that could attenuate deleterious effects of metabolic diseases on liver such as COX2, ACACB, FAS, ECHS1. In conclusion, this research preliminarily revealed the chemical characteristics of dark tea, explored the effect and mechanisms of dark tea on regulating hyperlipidemia and reducing obesity on physiological and biochemical, cytology, genomic and proteomic levels.
