檸檬精油對於小鼠抗憂鬱之效果

Abstract

中文摘要 憂鬱症已被世界衛生組織（World Health Organization, WHO）列為廿一世紀三大疾病之一，也是三大疾病中唯一的精神疾病，並指出憂鬱症為全球性疾病負擔排名第四名。若是以失能調整人年（disability adjusted life years, DALY）估算，預估於2020年時，造成人類失能的第二大主因，將成為除了缺血性心臟病以外的高負擔疾病，將更是與癌症和後天免疫不全症候群並列為二十一世紀人類的三大殺手，到了2030年憂鬱症更成為高負擔疾病之第一位。研究指出芳香精油、檸檬精油（lemon essential oil, LEO）或其成分經由吸入或食入後，對於人類或哺乳動物具有許多的生理活性，例如抗菌功能、抗氧化功能、清除自由基功能與情緒或情感方面功能，例如鎮靜功能、抗焦慮功能與抗失智等功能，目前學者研究檸檬精油之抗憂鬱功能，僅經由嗅覺吸入而未由食入之研究。根據 Steru 等人指出，小鼠尾部懸吊試驗（the tail suspension test, TST）使用簡單、觀察容易、客觀，和「行為上的絕望」表現出一致性，並且對於藥物的大範圍劑量都敏感，是一個適合的抗憂鬱動物模式。本研究以小鼠尾部懸吊實驗動物模式進行，評估檸檬精油在管餵小鼠模式上抗憂鬱的功效。本研究以氣相層析質譜儀（Gas chromatography-mass spectrometry, GC-MS）進行成分鑑定、滯留時間與標準品之資料庫質譜比對，分別鑑定檸檬精油之主要成分為limonene（67.7%）、β-pinene（13.0%）、γ-terpinene（10.1%）、α-pinene（2.2%）、myrcene（1.9%）、ρ-cymene（1.7%）與β-myrcene （1.0%）。抗憂鬱研究之第一部分以小鼠尾部懸吊試驗拉丁方設計找出檸檬精油抗憂鬱使用之最適劑量，結果發現檸檬精油400 mg/kg bw劑量組有顯著抗憂鬱功效，與檸檬精油100、200、800與1600 mg/kg bw劑量組呈現「U型曲線」關係，且發現檸檬精油沒有累積的影響，且沒有上次尾吊實驗之記憶，並未有學習之效果。第二部份研究中，將小鼠分為控制組、檸檬精油200、400、800 mg/kg bw劑量組與正對照組（fluoxetine 18 mg/kg bw），於開放空間試驗中，小鼠管餵檸檬精油後在穿越次數、移動距離、移動時間、移動速度與排泄物的數量上，分別於全部、周圍區與中央區均沒有顯著差異，表示檸檬精油並不會造成小鼠的自主活動行為與腸胃道的影響。小鼠管餵檸檬精油後，血清中葡萄糖、總膽固醇、尿素氮與總膽紅素；肝臟、腎臟與脾臟病理切片觀察均不會造成影響。小鼠尾部懸吊試驗中發現，檸檬精油400 mg/kg bw劑量組較控制組產生極顯著性差異，檸檬精油200、400、800 mg/kg bw劑量組間，呈現「U型曲線」關係；不活動時間之差異性在後3分鐘時產生極顯著差異。小鼠腦部單胺類物質之檢測發現，檸檬精油400 mg/kg bw劑量組與控制組相比較，於前額葉皮質區，正腎上腺素之表現有顯著差異；於紋狀體區與海馬迴區，多巴胺與血清素的濃度均顯著高於控制組；多巴胺與血清素之代謝速率均較控制組顯著降低；但於皮質區與小腦區，均無顯著差異。因此，小鼠抗憂鬱機制可能經由正腎上腺素、多巴胺與血清素共同影響之結果。

Abstract Accumulating studies have indicated that either inhalation or digestion of essential oils (e.g., lemon essential oil (LEO)), impart various bioactivities on humans and mammalian species, such as antibacterials, antioxidants, and radical scavengers. These oils also display some potential effects on the control or modulation of emotions. Despite the potential effects on mood control, LEO was usually administered via inhalation in most antidepressant studies and fewer attempts were made to administrate LEO orally. It is of great interest to expand the use of LEO in foods. The chemical composition of the LEO was analyzed by GC-MS and the volatile compounds were identified by mass spectra, retention index, and comparison with known standards. The major identified compounds are limonene (67.7%), β-pinene (13.0%), γ-terpinene (10.1%), α-pinene (2.2%), myrcene (1.9%), ρ-cymene (1.7%), and β-myrcene (1.0%). Taking advantage of tail suspension test (TST) as a mouse model of depression, the aims of this study are to (1) investigate the antidepressant effect of LEO and its effective dose, (2) to examine its physiological and histopathological impact, and (3) to explore the biochemical alterations in the brains of LEO gavaged mice. Six experiments were conducted in this study. A Latin square design was first applied to establish a “U-shaped” dose-response curve and to reveal an effective dose of LEO in mice using TST. Their spontaneous locomotor activities were further examined in the open field and our result indicated that different concentrations of LEO had no effect on their spontaneous locomotion. By contrast to Fluoxetine (18 mg/kg bw, o.p.) as a positive control, the effective dose (i.e., 400 mg/kg bw) of LEO was further confirmed in another batch of naïve mice. Our physiological examination also revealed that these dosages of 200, 400, and 800 mg/kg bw LEO we used in this study did not affect their body weights, glucose, the total cholesterol, the blood urea nitrogen and the total bilirubin of serum. By using HPLC to evaluate biochemical alterations in the brain of LEO gavaged mice, significant alterations of norepinephrine, dopamine and serotonin monoamine were revealed in the prefrontal cortex, striatum, and hippocampus, which might, at least in part, account for the antidepressant effect of LEO in these mice.