- 學位論文
探討彰化和美、高雄甲仙與屏東內埔地區生產的蜂膠產品之酚類、類黃酮含量與其抗氧化力的差異
A study on the content of phenols, flavonoids, and antioxidant activity of propolis from Changhua (Hemei), Kaohsiung (Jiaxian) and Pingtung (Neipu), Taiwan
摘要
蜂膠是一種樹脂黏性混合物，蜜蜂從樹芽、花、葉子和其他植物來源收集，蜜蜂用唾液和蜂蠟分泌物和蜂膠混合。蜂膠含有樹脂、蠟質、多醣、多酚和次級代謝產物（如黃酮類化合物和蜂膠素）。蜂膠的成分因地理位置、季節以及蜜蜂採集的樹種和花卉而異。因此本研究旨在確認來自台灣三個地理來源的蜂膠其酚類、類黃酮及抗氧化能力。使用不同的分析方法進行其化學表徵以確定地理來源對蜂膠特性的影響。 蜂膠樣品來自三個不同地理位置，其樣品採集的海拔高度(above mean sea level; amsl) 在彰化和美 (15 m amsl) 的沿海地區、高雄甲仙 (450 m amsl) 的丘陵地區以及屏東內埔 (80 m amsl) 的平原地區 ，蜂膠於2021年11月初至2022年2月初期間採集。本研究使用蒸餾水和不同濃度的乙醇 (55、65、75、85及95%) 和不同萃取時間 (30, 60, 90, 120, 150和180分鐘)，利用超音波輔助來萃取。在萃取條件中，85%的乙醇溶劑和90分鐘萃取時間為最佳的萃取組合。為分析蜂膠的化學成分，運用超高效液相層析儀 (UPLC)，使用280 nm紫外線波長分析蜂膠中的化合物。為了測定蜂膠樣品的總酚含量 total phenolic content (TPC) 和總黃酮含量 total flavonoid content (TFC)，使用福林-西奧卡特比色法 (Folin-Ciocalteu method; FCM) 比色法和氯化鋁比色法 (Aluminum chloride colorimetric assay; ACCA)。於FCM分析顯示，丘陵山區樣品的TPC明顯高於沿海地區及平原地區樣品。當使用ACCA分析蜂膠樣品的TFC時，發現了和TPC相似的趨勢。使用鐵降低抗氧化能力 (Ferric-reducing ability of plasmas; FRAP)，其抗氧化能力測定法顯示，蜂膠的抗氧化能力與TPC和TFC含量有直接相關性。丘陵山區的蜂膠樣品具有最高的抗氧化能力，其次是沿海地區，最後是平原地區。主成分分析 (Principal component analysis; PCA) 由蜂膠樣品的 TPC、TFC 和FRAP抗氧化能力數據分析顯示有明顯的三個主成分集群，與平原地區的主成分析相比，丘陵和沿海地區主成分集群更接近。 基於這項研究，酚類化合物被發現是識別蜂膠地理來源的重要化合物。UPLC 分析清楚地揭示了所有蜂膠樣品中相似類型的酚類化合物。三個地區的綠蜂膠樣品都含有相同的五個異戊烯類黃酮蜂膠素 (prenylflavanones propolin) C、D、F、G 和 H，以及微量的白楊素 (Chrysin) 和高良薑素 (Galangin) 二種黃酮類化合物。 關鍵字: 鐵降低抗氧化能力、類黃酮、福林-西奧卡特比色法、地理來源、酚類化合物、蜂膠、超高效液相層析儀
並列摘要
Propolis, a sticky resinous mixture also called as bee glue, is collected from tree buds, flowers, leaves, and other botanical sources by honeybees. After being collected, propolis is mixed with the secretions of bees, such as saliva and wax. Propolis contains resins, waxes, polysaccharides, polyphenols, and secondary metabolites such as flavonoids and propolins. Honey bees collect propolis from various plants depending on geography, season, and weather. The composition of propolis may also vary depending on the types and location of plants. Therefore, this study investigated their total phenolic content (TPC), total flavonoid content (TFC) and antioxidant capacity of propolis from three geographical regions in Taiwan. The green propolis samples were collected from Hemei, Changhua at the altitudes of 15 m above mean seal level (m amsl) in coastal area, Jiaxian, Kaohsiung (450 m amsl) in hilly area, and Neipu, Pingtung (80 m amsl) in plain area from early November 2021 to early February 2022. Collected propolis samples were extracted using distilled water and 55, 65, 75, 85 and 95% ethanol at different extraction durations (30, 60, 90, 120, 150 and 180 min) by ultrasound-assisted extraction. Among the ethanol concentrations and extraction durations tests, the extract from 85% ethanol for 90 min had the highest yield. For the analysis of chemical composition of propolis, Ultra Performance Liquid Chromatography (UPLC) at UV wavelength of 280 nm was used. To determine the TPC and TFC of propolis, Folin-Ciocalteu method (FCM) and Aluminum chloride colorimetric assay (ACCA) were used. Based on the FCM analysis, the TPC of propolis from the hill area was significantly higher than that of propolis from the coastal or plain areas. Similar trends were found while ACCA was used to determine the TFC from propolis samples. Based on an analysis of ferric reducing ability of plasma (FRAP), propolis' antioxidant capacity was positively related to TPC and TFC levels. Propolis sample from hill area (Kaohsiung) had the highest antioxidant capacity among the evaluated samples followed by coastal area (Changhua) and plain area (Pingtung). The principal component analysis (PCA) results showed that there remained clearly three clusters based on propolis' TPC, TFC and (FRAP) antioxidant capacity. In comparison to the plain area (Pingtung) cluster, the hill area (Kaohsiung) and coastal area (Changhua) clusters were closer to each other. Based on this study, phenols were found to be important compounds for identifying the geographical origin of propolis. UPLC analysis clearly revealed similar types of phenolic compounds in all propolis samples. Additionally, all three green propolis samples contain the same five main compounds as prenylflavanones propolin C, D, F, G and H, as well as trace amounts of Chrysin and Galangin. Keywords: Ferric Reducing Ability of Plasma, flavonoids, Folin-Ciocalteu method, geographic origin, phenolic compounds, propolis, Ultra Performance Liquid Chromatography